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Linda D. Hazlett

Distinguished Professor & Chair, Department of Anatomy and Cell Biology

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Linda D. Hazlett

Distinguished Professor & Chair, Department of Anatomy and Cell Biology

Anatomy & Cell Biology and Distinguished Professor Ophthalmology & Immunology/Microbiology Research Subject: Corneal infection Focus: Host immune response to bacterial keratitis

Selected Recent Published Articles:
- 1. Berger, E.A., Vistisen K.S., Barrett, R., and Hazlett, L.D. Effects of VIP on corneal reconstitution and homeostasis following Pseudomonas aeruginosa induced keratitis. IOVS, 53:7432-9, 2012. PMID 23036997 PMCID:PMC3061511 2. Jiang, X., McClellan, S.A., Barrett, R.P., Zhang, Y., Foldenauer M., and Hazlett, L.D. The role of VIP in cornea. Invest. Ophthalmol. Vis. Sci., 53: 7560-6, 2012. PMID:23074208. PMCID:PMC3493186 3. Sun, M., Zhu, M., Nie, X., Deng, Q., Chen, K., Hazlett, L., Wu, M., and Huang, X. TREM-2 promotes host resistance against Pseudomonas aeruginosa by suppressing corneal inflammation via PI3K/Akt signaling pathway. IOVS, 17:3452-62, 2013. PMCID:PMC3658264 4. Foldenauer, M., McClellan, S., and Hazlett, L.D. mTOR regulates IL-10 and resistance to Pseudomonas aeruginosa corneal infection. J. Immunology, 190:5649-58, 2013. PMCID:PMC3660542 5. Berger, E., McClellan, S., Vistisen, K., and Hazlett, L.D. HIF-1α is essential for effective PMN bacterial killing, antimicrobial peptide production and apoptosis in Pseudomonas aeruginosa keratitis PLOS PATHOGENS 9(7): 18 July, 2013 e1003457. doi:10.1371/journal.ppat.1003457. PMCID:PMC3715414 6. Jiang, X., McClellan, S., Barrett, R., Foldenauer, M., and Hazlett, L.D. HGF signaling impacts severity of P. aeruginosa keratitis. IOVS, 55:2180-90, 2014. PMCID:PMC3985408 7. Hazlett, L. Linda Hazlett speaks to Hannah Branch, Commissioning Editor. Future Microbiol. 709-710, 2013. 8. Hazlett, Linda D., Jiang, Xiaoyu, and McClellan, Sharon A. IL-10 Function, Regulation, and in Bacterial Keratitis. Journal of Ocular Pharmacology and Therapeutics 30: 373-380, 2014. PMCID:PMC4043257 9. Li, C., McClellan, S.A., Barrett, R., Hazlett, L.D. Interleukin 17 regulates Mer tyrosine kinase-positive cells in Pseudomonas aeruginosa keratitis. IOVS 55:6886-900,2014. PMCID:PMC4214210 10. High-mobility group box 1: A novel target for treatment of Pseudomonas aeruginosa keratitis J.Immunology, in press 2015.
Summary of Research:
- Dr. Hazlett’s research is focused on the host immune response to the gram negative bacterium Pseudomonas aeruginosa (P. aeruginosa). The ubiquitous bacterium is an opportunistic pathogen noted both for its ability to cause disease in particularly susceptible or compromised individuals and its ever-increasing resistance to antibiotics. Extremely common in soil and water, it is also responsible for many serious, often fatal, human infections, including upper respiratory tract colonization in cystic fibrosis patients and in ocular infections, particularly due to contact lens usage, where blindness can be the outcome. With contact lens use at an all time high, such sight-threatening infections are becoming increasingly widespread and range from purulent conjunctivitis to iridocyclitis, keratitis and iritis, corneal ulcer, and panophthalmitis. In fact, of the 30,000,000 daily-wear contact lens users worldwide, one in every 2,500 will suffer pseudomonal infection; the odds increase by a factor of five – or one in every 500 – for those using extended-wear or overnight lenses. In the United States alone, 25,000-30,000 cases of microbial keratitis are reported annually with an estimated cost of between $15 and $30 million for treatment. Dr. Hazlett has authored 167 peer reviewed publications on this topic in high quality journals, including the Journal of Immunology, Progress in Retinal and Eye Research (2004 publication, solicited Review Article by this number one eye journal) and Investigative Ophthalmology and Visual Science. She also has contributed 22 books/chapters/review articles, one of which is a chapter contribution to a clinical textbook (Duane’s Clinical Ophthalmology and Foundations of Clinical Ophthalmology, 2006) entitled, “Pathogenesis of Pseudomonas Keratitis”. The long-term objective of her studies is to reveal basic pathophysiological mechanisms of disease in the P. aeruginosa infected cornea. Her studies directly address a major objective of the current Corneal Diseases Program (National Plan for Eye and Vision Research) which parenthetically states, to “investigate corneal infectious processes and immunological responses and to develop treatments to reduce keratitis and prevent blindness.” Her studies integrate cellular and molecular biology and immunology and continue to elucidate novel targets in the immune response that will impact clinical care of patients with P. aeruginosa keratitis
Current Research
- Dr. Hazlett studies the role of the host response to bacterial infection of the cornea as a major focus of her scientific work. As her publications suggest, she has been doing this for a considerable amount of time and with high productivity and success. She has specifically focused on two areas one, Toll-like receptors (TLR) and the other, neuropeptides, that modulate the host response to bacterial infection. Her goal is to better understand the disease process in which inflammation, if uncontrolled, is deleterious and to develop rational intervention therapies that will have clinical relevance. Her current R01 focuses on the pathogenesis and treatment potential of interventions against high-mobility group box 1 (HMGB1), a prototypic alarmin. HMGB1 is a mediator of the systemic inflammatory response syndrome, is elevated late in bacterial infection/sepsis and considered a target for disease treatment, but nothing is known about its role in bacterial keratitis. Given that it is important in innate immunity, has different functions dependent on cellular localization, and has the ability to bind to TLR and other molecules such as receptor for advanced glycation end products (RAGE), the hypothesis that she is testing is that HMGB1 has significant amplification effects on resident and infiltrating inflammatory cells and is an attractive therapeutic target in Pseudomonas (P.) aeruginosa keratitis. Dr. Hazlett also is PI of Wayne State’s NEI Core Vision grant and uses the facilities of the Core for her own studies and together with the Ophthalmology chair, Mark Juzych, M.D., continues to advance vision research at Wayne State University principally through recruitment and development of the research enterprise.

Elizabeth Berger, PhD

Assistant Professor in Anatomy & Cell Biology and Ophthalmology

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Elizabeth Berger, PhD

Assistant Professor in Anatomy & Cell Biology and Ophthalmology

Research Subject: Ocular Immunology Focus: Bacterial Keratitis

Published Articles:
- Berger EA, McClellan SA,Vistisen KS, Hazlett LD. HIF-1a is essential for effective PMN bacterial killing, antimicrobial peptide production and apoptosis in Pseudomonas aeruginosa keratitis. PLoS Pathogen. 9(7):e1003457, Epub Jul 18, 2013. PMID:23874197
  Foldenauer MEB, McClellan SA, Berger EA, Hazlett LD. Mammalian target of rapamycin regulates IL-10 and resistance to Pseudomonas aeruginosa corneal infection. J Immunol. Jun 190(11):5649-5658, 2013. PMID: 23626014
  Berger EA, Vistisen KS, Barrett RP, Hazlett LD. Effects of VIP on corneal reconstitution and homeostasis following Pseudomonas aeruginosa induced keratitis. Invest Ophthalmol Vis Sci. Nov 53(12):7432-7439, 2012. PMID: 23036997
  Jiang X, McClellan S, Barrett RP,Berger EA, Zhang Y, Hazlett LD. VIP and growth factors in the infected cornea. Invest Ophthalmol Vis Sci. Aug 52(9): 6154-6161, 2011. PMID: 21666233
  Berger EA, McClellan S, Barrett RP, Hazlett LD. Testican-1 promotes resistance against P. aeruginosa induced keratitis via modulation of MMP-2 driven wound healing and ECM restoration. Invest Ophthalmol Vis Sci. Jul 52(8):5339-5346, 2011. PMID: 21613368
  Berger EA, McClellan S, Barrett RP, Hazlett LD. VIP promotes resistance in the Pseudomonas aeruginosa infected cornea by modulating adhesion molecule expression. Invest Ophthalmol Vis Sci. 51(11):5776-5782, 2010. PMID: 20592225
  Zhou Z, Barrett R, McClellan S, Zhang Y,Szliter E, Huang X, Hazlett LD. Substance P delays apoptosis in P. aeruginosa infected cornea and exacerbates disease. Invest Ophthalmol Vis Sci. Oct; 49(10): 4458-4467, 2008. PMID: 18566468
  Szliter EA, Lighvani S, Barrett RP, Hazlett LD. VIP balances pro- and anti-inflammatory cytokines in the P.aeruginosa infected cornea and protects against corneal perforation. J Immunol. Jan; 178(2): 1105-1114, 2007. PMID:17202374
  Szliter EA, Barrett RP, Gabriel MM, Zhang Y, Hazlett LD. Pseudomonas aeruginosa -induced inflammation in the rat extended-wear contact lens model. Eye Contact Lens. Jan; 32(1): 12-18, 2006. PMID: 16415687
  Szliter EA, Morris C.A, Carney F, Gabriel, MM, Hazlett LD. Development of An Extended Wear Contact Lens Model In the Rat. CLAO J. Jul; 28(3): 119-123, 2002. PMID: 12144229
Summary of Research:
- Title: Assistant Professor in Anatomy & Cell Biology and Ophthalmology.The studies carried out by our laboratory predominately focus on disease pathogenesis and the interplay between the immune and neuroendocrine systems. We mechanistically and therapeutically investigate the events of inflammation and innate immunity using a model of ocular infectious disease induced by a common pathogen of eye infections – Pseudomonas aeruginosa. This includes analyses of: host inflammatory cells (macrophages, neutrophils, T cells – both in vivo and in vitro), extracellular matrix and adhesion molecules, cytokines/chemokines, Toll-like receptors and other related molecules using a number of molecular, cellular and immunohistochemical techniques.
Current Research:
- Ongoing NIH-funded studies examine how the neuropeptide, vasoactive intestinal peptide(VIP), influences T cells and the production of IL-17, nerve regeneration, and the interaction with endogenous lipid mediator circuits. In addition, my laboratory is defining the clinical applicability of VIP as a treatment modality in the eye. Additionally, other projects include: 1) how neutrophil transdifferentiation contributes to disease pathogenesis and 2) collaborative projects looking at miRNAs, as well as a novel gene therapy strategy for the treatment of glaucoma.

Bruce Berkowitz, Ph.D.

Diabetic retinopathy; retinal physiology

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Bruce Berkowitz, Ph.D.

Diabetic retinopathy; retinal physiology

Focus: Dr. Berkowitz’s research is focused on the development and application of develop and apply new analytical MRI methods for discovering new perspectives on retinal pathophysiology and its treatment in experimental models of diabetic retinopathy. Title: Professor in Anatomy and Cell Biology, and Ophthalmology

Published Articles:
- http://www.ncbi.nlm.nih.gov/pubmed?term=berkowitz%20b*%20%5Bau%5D%20AND%20retina&cmd=Search
Summary of Research:
- Significant advances made in the Berkowitz lab include new information concerning panretinal and vascular and avascular border oxygenation, as well as the role of calcium channels, in retinal neovascular models, the benefits of steroid treatment on retinopathy, the first identification of a diabetic retinal edema model, and the importance of photoreceptor L-type calcium channels in diabetic retinopathy.
Current Research:
- Current imaging methods do not comprehensively measure essential photoreceptor functions of light detection, regulated transmission of information, and visual pigment regeneration. Our new suite of novel biomarkers greatly expands the number of analytical imaging tools available for comprehensive and objective assessment of these critical photoreceptor functions. Our new methods are transformative because they optimally measure photoreceptor function in health, in disease, and during treatment beyond that currently available, and can be applied in future translational human studies.

Tomomi Ichinose, MD, PhD

Assistant Professor

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Tomomi Ichinose, MD, PhD

Assistant Professor

Research Subject: Mechanisms of Visual Signal Processing in the RetinaFocus: Physiological functions of bipolar cells in the mouse retina

Published Articles:
- 1. Hellmer CH, Ichinose T: Recording light-evoked postsynaptic responses in neurons in dark-adapted, mouse retinal slice preparations using patch clamp techniques. Journal of Visualized Experiments (JoVE) In Press, 2015 2. Fyk-Kolodziej B, Hellmer CH, Ichinose T: Marking cells with infrared fluorescent proteins to preserve photoresponsiveness in the retina. BioTechniques 57:245-253, 2014
  3. Ichinose T, Fyk-Kolodziej B, Cohn J: Roles of ON cone bipolar cell subtypes in temporal coding in the mouse retina. Journal of Neuroscience 34:8761-8771, 2014 4. Ichinose T, Lukasiewicz PD: The mode of retinal presynaptic inhibition switches with light intensity. Journal of Neuroscience 32:4360-4371, 2012 5. Ichinose T, Lukasiewicz PD: Light and dopamine regulate sodium channels to dynamically control retinal signaling. Journal of Neuroscience 27:4756-4764, 2007 6. Ichinose T, Lukasiewicz PD: Inner and outer retinal pathways both contribute to surround inhibition of salamander ganglion cells. Journal of Physiology 565 (Pt 2):517-35, 2005 7. Ichinose T, Shields CR, Lukasiewicz PD: Sodium channels in transient retinal bipolar cells enhance visual responses in ganglion cells. Journal of Neuroscience 25(7):1856-65, 2005 8. Ichinose T, Yu S, Wang XQ, Yu SP: Ca2+-independent, but voltage- and activity-dependent regulation of the NMDA receptor outward K+ current in mouse cortical neurons. Journal of Physiology 551(Pt 2):403-17, 2003 9. Ichinose T, Lukasiewicz PD: GABA transporters regulate inhibition in the retina by limiting GABA(C) receptor activation. Journal of Neuroscience 22(8):3285-92, 2002 10. Manzerra P, Behrens MM, Canzoniero LM, Wang XQ, Heidinger V, Ichinose T, Yu SP, Choi DW: Zinc induces a Src family kinase-mediated up-regulation of NMDA receptor activity and excitotoxicity. Proceedings of the National Academy of Sciences U S A 98(20):11055-61, 2001 11. Ichinose T, Snider WD: Differential effects of TrkC isoforms on sensory axon outgrowth. Journal of Neuroscience Research 59(3):365-71, 2000
Summary of Research:
- Our visual system perceives the world as functions of time, space, and color. The temporal visual processing pathways encode time (temporal) information of an object. These pathways are broadly classified into transient pathways and sustained pathways based on their neural responses, which are thought to encode changing objects and static visual information, respectively. The transient/sustained dichotomy is recognized in retinal second order neurons, which are bipolar cells, suggesting that temporal processing originates in bipolar cells. Recently, morphological studies have revealed more than 10 subtypes of bipolar cells in the mammalian retina, which might initiate distinct pathways encoding different features of temporal signals. However, no studies have addressed how each subtype of bipolar cell in the mammalian retina encodes temporal visual information. In response to this research gap, my goal is to understand the cellular and molecular mechanisms of temporal encoding in each subtype of bipolar cell. We use electrophysiological recording techniques and immunohistochemical method to analyze temporal visual encoding in individual bipolar cells. We found that each ON bipolar cell encode different aspects of temporal visual information in a subtype-dependent manner (Figure 1) (Ichinose et al., 2014). We are currently working on the elucidation of underlying mechanisms of temporal encodings. These studies will improve our understanding of parallel processing in the retinal neural network, which will contribute to the future design of a functional retinal prosthesis and sight restoring gene therapies. Figure 1. Six subtypes of ON bipolar cells and their responses to step-light stimuli.

Renu A. Kowluru, PhD

(Ex. Research Assistant Professor in Ophthalmology) Professor

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Renu A. Kowluru, PhD

(Ex. Research Assistant Professor in Ophthalmology) Professor

Research Subject: Molecular Mechanism of Pathogenesis of Diabetic Retinopathy Focus: Mitochondrial Dysfunction and Epigenetic Modifications

Published Articles:
- Santos JM. Kowluru RA. Metabolic memory and diabetic retinopathy: Continued impaired retinal mitochondria biogenesis in diabetic rats. Invest Ophthalmol Vis Sci 52:8791-8798, 2011 (PMCID:PMC3202722) Tewari S, Santos JM and Kowluru RA. Damaged mitochondrial DNA replication system and the development of diabetic retinopathy. Antioxid Redox Signal 17:492-504, 2012 (PMCID:PMC3365359) Santos JM, Tewari S. Kowluru RA. A compensatory mechanism protects retinal mitochondria from initial insult in diabetic retinopathy. Free Rad Biol Med 53:1729-1737, 2012 (PMCID:PMC3632051) Kowluru RA, Kowluru A, Veluthakal R, Mohammad G, Syed I, Santos JM, Mishra M. TIAM1-RAC1 signalling axis-mediated activation of NADPH oxidase-2 initiates mitochondrial damage in the development of diabetic retinopathy. Diabetologia 57:1047-1056, 2014 (PMID:24554007) Zhong Q, Kowluru RA. Epigenetic modification of Sod2 in the development of diabetic retinopathy and in the metabolic memory: Role of histone methylation. Invest Ophthalmol Vis Sci 54:244-250, 2013 (PMCID:PMC3590072) Zhong Q, Kowluru RA. Regulation of matrix metallopeptidase-9 by epigenetic modifications, and the development of diabetic retinopathy. Diabetes 62:2559-2568, 2013 (PMCID:PMC3712057) Santos JM, Kowluru RA. Impaired transport of mitochondrial transcription factor A (TFAM) and the metabolic memory phenomenon associated with the progression of diabetic retinopathy. Diabetes Metab Res Rev 29:204-213, 2013 (PMCID:PMC3596457) Kowluru RA, Santos JM, Zhong Q. Sirt1, a negative regulator of matrix metalloproteinase-9 in diabetic retinopathy. Invest Ophthalmol Vis Sci 55:5653-5660, 2014 (PMID:24894401) Mishra M, Zhong Q, Kowluru RA. Epigenetic modifications of Nrf2-mediated glutamate-cysteine ligase: implications for the development of diabetic retinopathy and the metabolic memory phenomenon associated with its continued progression. Free Radic Biol Med 75:129-139, 2014 (PMID:25016074) Mishra M, Kowluru RA. Retinal mitochondrial DNA mismatch repair in the development of diabetic retinopathy, and its continued progression after termination of hyperglycemia. Invest Ophthalmol Vis Sci 55:6960-6967, 2014 (PMID:25249609). Mishra M, Zhong Q, Kowluru RA. Epigenetic modifications of Keap1 regulate its interaction with the protective factor Nrf2 in the development of diabetic retinopathy. Invest Ophthalmol Vis Sci 55:7256-7265, 2014 (PMID:25301875)
Ongoing Research:
- The overall objective of our research is to understand the pathogenesis of diabetic retinopathy, and the research is focused on investigating the role of mitochondrial damage in the development/ progression of diabetic retinopathy. Our studies have shown that capillary cells undergo accelerated apoptosis before histopathology characteristic of diabetic retinopathy can be observed in rodent models of diabetic retinopathy. Mitochondria become dysfunctional, their DNA is damaged, biogenesis is impaired, and cytochrome c starts to leak out into the cytosol activating the apoptotic machinery. Mitochondrial superoxide scavenging enzyme is subnormal, and its promoter and enhancer are epigenetically modified. However, the damage to the mitochondria precedes overproduction of reactive oxygen species by cytosolic NADPH oxidase, and continuous production of these reactive species in a hyperglycemic milieu damages the mitochondria. Using rodent models of diabetic retinopathy, and retinal vascular cells in culture, our laboratory is investigating the signaling mechanisms responsible for mitochondrial damage, and the role of epigenetic modifications of the nuclear genes and mitochondrial DNA in the homeostasis of mitochondrial function.

Ashok Kumar, Ph.D.

Assistant Professor, Department of Ophthalmology

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Ashok Kumar, Ph.D.

Assistant Professor, Department of Ophthalmology

Department of Anatomy and Cell Biology Wayne State University School of Medicine Kresge Eye Institute

Bacterial endophthalmitis is a vision-threatening complication of penetrating eye injury and intraocular surgery, notably cataract surgery, the most common ophthalmic procedure performed in older populations in the United States. Approximately, 3 out of 1000 patients develop bacterial endophthalmitis after cataract surgery. As the aged population in the US is expected to grow dramatically, the number of cataract surgeries performed will also increase significantly, resulting in a proportional increase in the incidence of endophthalmitis. The visual properties of the retina are highly sensitive to inflammation-caused damage therefore, a rapid detection and clearance of invading pathogens is critical in minimizing retinal damage. Dr. Kumar’s lab research interests are to understand the regulation of retinal innate immunity in endophthalmitis and to identify the potential targets for intervention. Recognition of microbial infection and initiation of host defense responses is controlled by multiple mechanisms. The host innate immune system uses a series of pattern recognition receptors (PRRs) to detect the presence of pathogens, thus allowing rapid host defense responses to invading microbes. A key component of such receptors is the “Toll-like receptor” (TLR). Previous studies from our laboratory and others have revealed an important role of TLRs in mediating innate immune response at the ocular surface. Although the expression and function of TLRs in many tissues have been studied extensively, few studies are related to the retina. Retinal pigment epithelial (RPE) cells have been shown to express TLR2, TLR3, TLR4, and TLR9. However, many questions fundamental to the understanding of retinal innate immunity remain elusive. For example: what cell types in the retinal tissue are involved in recognition of invading pathogens in endophthalmitis? Does the TLR innate system exist in the retina? If yes, which TLRs are involved? How might retinal cells respond to bacteria or the TLR ligand challenge? The main objective of our research is to define the role of TLRs in bacterial endophthalmitis using mouse models of S. aureus endophthalmitis and cultured retinal cells in vitro. A better understanding of host-pathogen interaction in the retina will allow more effective therapeutic strategies to prevent or treat bacterial endophthalmitis.

Awards & Honors
- 2010: Member, Scientific Review Committee, Fight for Sight foundation. 2010: Editorial board member, World Journal of Gastrointestinal Pathophysiology (WJGP) 2008: Grant Initiative Program Award, Wayne State University, School of Medicine 2008: KEI intramural research grant award. 2007: Fight for Sight grant-in-aid research award. 2006: Midwest eye bank research grant award. 2006: Association for Research in Vision and Ophthalmology (ARVO), travel grant. 2005: Fight for Sight post-doctoral fellowship award 1998: Post Grad Institute of Medical Education & Research fellowship. 1997: Secured first position in order of merit in Master’s Program For more information about Dr. Kumar, click here.

Zhuo-Hua Pan

Professor

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Zhuo-Hua Pan

Professor

Research Subject:Rodents and non-human primates Focus: Optogenetic approaches for restoration of vision and retinal visual processing Title: Professor, Department of Ophthalmology and Anatomy/Cell Biology; Edward T. and Ellen K. Dryer Endowed Professor of Ophthalmology; Scientific Director of the Ligon Research Center of Vision, Kresge Eye Institute, Wayne State University School of Medicine.

Published Articles (since 2000):
- Pan, Z.-H., Hu, H.-J. Voltage-dependent Na+ currents in mammalian retinal cone bipolar cells. J. Neurophysiol. 84:2564-2571, 2000. Pan, Z.-H. Voltage-gated Ca2+ channels and GABA receptors revealed at presynaptic terminals of mammalian retinal bipolar cells. Visual Neurosci. 18:279-288, 2001. Pan, Z.-H., Hu, H.-J., Perring, P., Andrade, R. T-type Ca2+ channels mediate neurotransmitter release in retinal bipolar cells. Neuron 32:89-98, 2001. Hu, H.-J., Pan, Z.-H. Differential expression of K+ currents in mammalian retinal bipolar cells. Visual Neurosci. 19:163-173, 2002. Ma, Y.-P., Pan, Z.-H. Spontaneous regenerative activity in mammalian retinal bipolar cells: roles of multiple subtypes of voltage-dependent Ca2+ channels. Visual Neurosci. 20:131-139, 2003. Ma, Y.-P., Cui, J., Hu, H.-J., Pan, Z.-H. Mammalian retinal bipolar cells express inwardly rectifying K+ currents (IKir) with a different distribution than that of Ih. J. Neurophysiol. 90:3479-3489, 2003. Cui, J., Ma, Y.-P., Lipton, S.A., Pan Z.-H. Glycine receptors and glycinergic synaptic input at the axon terminals of mammalian retinal rod bipolar cells. J. Physiol. 553:895-909, 2003. Ma, Y.-P., Cui, J., Pan, Z.-H. Heterogeneous expression of voltage-dependent Na+ and K+ channels in mammalian retinal bipolar cells. Visual Neurosci. 22:119-133, 2005. Bi, A., Cui, J., Ma, Y.-P., Olshevskaya, E., Pu, M., Dizhoor, A.M., and Pan Z.-H. Ectopic expression of a microbial-type rhodopsin restores visual responses in mice with photoreceptor degeneration. Neuron 50:23-33, 2006. Cui, J. and Pan Z.-H. Two types of cone bipolar cells express voltage-gated Na+ channels in the rat retina. Visual Neurosci. 25: 635-645, 2008. Hu, C., Bi, A, and Pan Z.-H. Differential expression of three T-type calcium channels in retinal bipolar cells in rat. Visual Neurosci. 26:177-187, 2009. Zhang, Y., Ivanova, E., Bi, A., and Pan, Z.-H. Ectopic expression of multiple microbial rhodopsins restores ON and OFF light responses in the retina after photoreceptor degeneration. J. Neurosci. 29:9186-96, 2009. Lu, Q., Ivanova, E., and Pan Z.-H. Characterization of GFP-expressing retinal cone bipolar cells in a 5-HTR2a transgenic mouse line. Neurosci. 163:662-668, 2009. Ivanova, E, Pan, Z.-H. Evaluation of virus mediated long-term expression of channelrhodopsin-2 in the mouse retina. Mol. Vision 15:1680-1689, 2009. Ivanova, E., Hwang, G.S. and Pan, Z.-H. Characterization of transgenic mouse lines expressing Cre-recombinase in the retina. Neurosci. 165: 233-243, 2010. Ivanova, E., Hwang, G.-S., Pan, Z.-H., and Troilo, D. Evaluation of AAV-mediated expression of chop2-GFP in the marmoset retina. IOVS 51: 5288-5296, 2010. Ivanova, E, Roberts, R. Bissig, D., Pan, Z.-H., and Berkowitz, B.A. Retinal and optic nerve channelrhodopsin-2-mediated activity in vivo evaluated with manganese-enhanced MRI. Mol. Vision 16:1059-67, 2010. Wu, C., Ivanova, E., Cui, J., Lu Q. and Pan. Z.-H. Action potential generation at an AIS-like process in the axonless retinal AII amacrine cell. J. Neurosci. 31:14654-14659, 2011. Cui, J., Ivanova, E., Lu Q., and Pan, Z.-H. Expression of CaV3.2 T-type Ca2+ channels in a subpopulation of retinal type-3 cone bipolar cells. Neurosci. 224:63-9, 2012. Shimano, T., Fyk-Kolodziej, B., Mirza, N., Asako, M., Koichi, T., Bledsoe, S., Pan, Z.-H., Molitor, S., Holt, A.G. Assessment of the AAV- mediated expression of channelrhodopsin-2 and halorhodopson in brainstem neurons mediating auditory signaling. Brain Research, 1511:138-52, 2013. Ivanova, E., Lee, P., and Pan, Z.-H. Characterization of multiple bistratified retinal ganglion cells in a purkinje cell protein 2-cre transgenic mouse line. J. Comp. Neurol. 521:2165-80, 2013. Lu, Q., Ivanova, E., Ganjawala, H. T., and Pan, Z.-H. Cre-mediated recombination efficiency and transgene expression patterns of three retinal bipolar cell-expressing Cre transgenic mouse lines. Mol. Vision, 19:1310-1320, 2013. Wu, C., Ivanova, E., Zhang, Y., Pan, Z.-H. AAV-mediated subcellular targeting of optogenetic tools in retinal ganglion cells. PloS One, 8(6):e66332, 2013. Pan, Z.-H., Ganjawala, T.H., Lu, Q., Ivanova, E., Zhang, Z. ChR2 mutants at L132 and T159 with improved operational light sensitivity for vision restoration. PloS One, 9(6):e98924, 2014.
Summary of Research:
- The major focus of the laboratory is on the development of a novel optogenetic strategy for restoring sight to people who suffer from retinal degenerative diseases, such as retinitis pigmentosa and age-related macular degeneration. The optogenetic strategy for restoring vision utilizes genetically encoded light sensors to convert surviving retinal neurons into photosensitive cells, thus imparting light sensitivity to the retina after the death of photoreceptor cells. Our earlier proof-of-concept studies demonstrated the feasibility of this strategy by AAV-mediated expression of channelrhodopsin-2 (ChR2) in inner retinal neurons of photoreceptor-deficient mice. We also demonstrated the restoration of both ON and OFF light responses by expression of ChR2 and halorhodopsin in the retina. More recently, we also made important progress on the development of more light sensitive ChR2 as well as AAV vector constructs for selective targeting of the light sensors to specific retinal neurons and/or subcellular compartments. Our long-term goal is aimed at developing effective optogenetic therapies for treating blindness caused by retinal degenerative diseases. Another area of interest of the laboratory is the investigation of retinal visual information processing, especially the roles voltage-dependent membrane channels in retinal bipolar cell processing. This will provide new insight into our understanding of visual information processing in the retina and help to develop new treatment approaches for blinding retinal diseases. The experimental techniques used in our studies include molecular biology, AAV vectors, electrophysiological recordings, immunocytochemistry, and animal behavior.
Current Research:
- 1) Develop and optimize optogenetic light sensors for restoring vision. 2) Develop virus-mediated targeting of optogenetic light sensors to specific inner retinal cell types and/or subcellular compartments. 3) Evaluate optogenetic approaches with electrophysiologial recordings and animal behavioral experiments. 4) Investigate the roles voltage-dependent membrane channels in the information processing of retinal bipolar cells.

Shunbin Xu, M.D. and Ph.D.

Associate professor

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Shunbin Xu, M.D. and Ph.D.

Associate professor

Research Subject: microRNAs in retina and ocular diseases Focus: 1) microRNAs in retinal degenerative diseases and aging. 2) microRNAs in diabetic retinopathy – diagnosis and treatment. 3) microRNAs in bacterial keratitis. 4) microRNAs in glaucoma.

Published Articles:
- 1. Cowan C, Muraleedharan CK, O’Donnell JJ, III, Singh PK, Lum H, Kumar A, and Xu S. microRNA-146 Inhibits Thrombin-induced NF-κB Activation and Subsequent Inflammatory Responses in Human Retinal Endothelial Cells. Invest Ophthalmol Vis Sci. 2014 Jul 1;55(8):4944-51. PMID: 24985472. 2. Lumayag S, Haldin CE, Corbett NJ, Wahlin KJ, Cowan C, Turturro S, Larsen P, Kovacs B, Witmer PD, Valle D, Zack DJ, Nicholson DA, and Xu S. Inactivation of the microRNA-183/96/182 cluster results in syndromic retinal degeneration. PNAS 2013 Feb 5;110(6):E507-16. doi: 10.1073/pnas.1212655110. Epub 2013 Jan 22.. PMID: 23341629. 3. Kovacs B, Lumayag S, Cowan C, Xu S. microRNAs in early diabetic retinopathy in streptozotocin-induced diabetic rats. Invest. Ophthalmol. Vis. Sci. 2011 Jun 21; 52(7): 4402-9. Print 2011 Jun. PMID: 21498619 4. Hildebrand MS, Witmer PD, Xu S, Newton SS, Kahrizi K, Najmabadi H, Valle D, Smith RJH. miRNA Mutations are Not a Common Cause of Deafness. Am J Med Genet A. 2010 Mar;152A(3):646-52. PMID: 20186779. 5. Xu S. microRNA expression in the eyes and their significance in relation to functions. Prog Retin Eye Res. 2009 Mar;28(2):87-116. Epub 2008 Nov 28. (invited review). PMID: 19071227. 6. Xu S*, Witmer D, Kovacs B, Lumayag S and Valle D*. (2007). MicroRNA Transcriptome Of Mouse Retina And Functional Study of a Sensory Organ Specific miRNA cluster. J Biol Chem. 282(34):25053-25066. PMID: 15456885. (*: corresponding authors). 7. Perez SE, Lumayag S, Kovacs B, Mufson EJ and Xu S. (2008) β-Amyloid Deposition and Functional Impairment in the Retina of the APPswe/PS1∆E9 Transgenic Mouse Model of Alzheimer’s Disease. Invest Ophthalmol Vis Sci. 2009 Feb;50(2):793-800. PMID: 18791173. 8. Kovacs B, MacCumber MW, Xu S. (2007). Adult Retinal Stem Cell Spheres Hold Potential for Diabetic Retinopathy (DR) Treatment. Retina Today. Nov/Dec 2007: 41-42. 9. Liu H, Xu S, Wang Y, Mazerolle C, Thurig S, Coles BLK, Ren J, Taketo MM, van der Kooy D, Wallace VA. (2007). Ciliary margin transdifferentiation from neural retina is controlled by canonical Wnt signaling, Developmental Biology 308(1):54-67. PMID: 17574231. 10. Xu S, Sunderland ME, et al. (2007). The proliferation and expansion of retinal stem cells require functional Pax6. Dev Biol. 304(2):713-21. PMID: 17316600. 11. Smukler SR, Runciman SB, Xu S, van der Kooy D. (2006). Embryonic stem cells assume a primitive neural stem cell fate in the absence of extrinsic influences. J Cell Biol. 172(1):79-90. PMID: 16390999.
Summary of Research:
- The research interest of my laboratory is to study the roles of a new class of molecules – microRNAs (miRNAs), in the eye and ocular diseases. miRNAs are small, non-coding, regulatory RNAs and constitute a newly recognized level of gene expression regulation. They exist in every cells of our body, and play important roles regulating the expression of protein-coding genes, and can cause diseases when they are defective. However, the roles of these newly discovered molecules in the eye and ocular diseases are largely unknown. Our long term goal is to uncover the roles miRNAs in normal development and function of the eye and in ocular diseases, and to identify novel miRNA-based diagnostic biomarkers and therapeutic targets for early diagnosis and treatment of various ocular diseases, including diabetic retinopathy, inherited retinal degeneration, glaucoma, and age-related macular degeneration and corneal keratitis.
Current Research:
- Currently, the following major projects are ongoing in the lab: 1) microRNAs in retinal degenerative diseases and aging: Previously, we identified all miRNAs exist in adult mouse and human retina. Among these retinal miRNAs, we focus on a conserved miRNA cluster – the miR-183/96/182 cluster, which produces three miRNAs, miR-183, miR-96 and miR-182, which are highly expressed in the retina and all major sensory organs (Xu et al. 2007. JBC). We showed that inactivation of miR-183/96/182 in mice causes congenital retinal dysfunction, progressive retinal degeneration, and multiple sensory defects (Lumayag et al. PNAS. 2013). Currently, we are working on molecular mechanisms underlying retinal and other sensory diseases caused by inactivation of miR-183/96/182, and identification of disease-causing mutations in miR-183/96/182 cluster in patients with inherited and age-related retinal degenerative diseases. This research will lead to molecular diagnosis of retinal degenerative diseases, and finding new therapies for these retinal diseases. In addition, we are studying miRNAs in the retina of patients with age-related macular degeneration (AMD). This research will lead to identification of miRNAs involved in AMD and new therapeutic targets for its treatment. 2) microRNAs in diabetic retinopathy – diagnosis and treatment: Diabetic retinopathy (DR) is one of the leading causes of blindness. Previously, we identified a series of key miRNA signatures reflecting pathological changes of early DR (Kovacs et al. IOVS. 2011). Currently, we focus on miR-146 and several other miRNAs to dissect their involvement in DR and test their potential as therapeutic targets for treatment of DR. In addition, we are developing miRNA biomarkers in the vitreous, aqueous and plasma as diagnostic biomarkers for DR and other retinal diseases. 3) microRNAs in neuroimmune interaction and bacterial keratitis: Pseudomonas aeruginosa (PA) is a gram-negative opportunistic pathogen capable of inducing keratitis, affecting 25,000-300,000 contact lens users annually with a cost of treatment estimated between $15 to $30 million in the USA. Up to now, bacterial keratitis is mainly treated by topical administration of antibiotics; however, frequent emergence of antibiotic resistant bacteria poses serious challenges for its effective management. Recently, collaborating with Dr. Linda D Hazlett, we showed that inactivation of miR-183/96/182 causes decreased PA-induced corneal inflammation and keratitis in mice. Currently, we are dissecting molecular mechanisms of miR-183/96/182 in bacterial keratitis and develop miRNA-based new therapy for treatment of keratitis. 4) microRNAs in glaucoma: Previously we identified miRNAs whose expression levels changed in the retina of glaucoma mice compared to normal control mice. Currently, we are studying their roles in pathogenesis of glaucoma. This research may lead to identification of new miRNA-based therapeutic targets to develop new treatment for glaucoma.

Jena J. Steinle, PhD

Professor of Anatomy and Cell Biology and Ophthalmology

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Jena J. Steinle, PhD

Professor of Anatomy and Cell Biology and Ophthalmology

Area/Subject of Research: Retinal Vascular Biology

Research Focus: Diabetic Retinopathy, Vascular Biology of Ocular Cancers Education: Post-Doc (Vascular Biology)–Texas A&M University, Temple, TX 2003 PhD—University of Kansas Medical Center, Kansas City, KS, 2001 BS–University of Bridgeport, Bridgeport, CT, 1997

Publications: Recent of a total of 65 publications
- 1) Thakran S, Zhang Q, Morales-Tirado VM, Steinle JJ. Pioglitazone restores IGFBP-3 levels through DNA PK in retinal endothelial cells cultured in hyperglycemic conditions. Invest Ophthalmol Vis Sci. 2014 Dec 18. pii: IOVS-14-15550. PMID: 25525174 2) Jiang, Y, Zhang, Q, Steinle, JJ. Etanercept restores normal insulin signal transduction in β2-adrenergic receptor knockout mice. Journal of Neuroinflammation. PMID: 25138272 3) Zhang, Q, Steinle, JJ. 2014. IGFBP-3 inhibits TNFα production and TNFR-2 signaling to protect against retinal endothelial cell apoptosis. Microvasc. Res. 95C:76-81. PMID: 25086184 4) Jiang, Y, Thakran, S, Bheemreddy, R, Ye, EA, He, H, Walker, RJ, Steinle, JJ. 2014. Pioglitazone normalizes insulin signaling in the diabetic rat retina through reduction in tumor necrosis factor α and suppressor of cytokine signaling 3. J. Biol. Chem. 289(38):26395-405. PMID: 25086044 5) Jiang, Y, Pagadala, J, Miller, DD, Steinle, JJ. 2014. Insulin-like growth factor-1 binding protein 3 (IGFBP-3) promotes recovery from trauma-induced expression of inflammatory and apoptotic factors in the retina. Cytokine. PMID: 25082650 6) Zhang, Q, Soderland, D, Steinle, JJ. 2014. TNFα inhibits IGFBP-3 through Activation of p38α and Casein Kinase 2 in Human Retinal Endothelial Cells. PLoS One. 9(7):e103578. PMID: 25073020 7) Jiang, Y, Zhang, Q, Ye, EA, Steinle, JJ. 2014. β1-adrenergic receptor stimulation by agonist Compound 49b restores insulin signal transduction in vivo. Mol Vis. 20:872-80. PMID: 24966659 8) Jiang, Y, Zhang, Q, and Steinle, JJ. 2014. Intravitreal injection of IGFBP-3 restores normal insulin signaling in diabetic rat retina. PLoS One. 9(4):e93788. PMID: 24695399 9) Zhang, Q*, Jiang, Y, Toutounchian, J, Wilson, MW, Morales-Tirado, V, Miller, DD, Yates, CR, Steinle, JJ. 2013. Novel Quinic Acid Derivative KZ-41 Prevents Retinal Endothelial Cell Apoptosis Without Inhibiting Retinoblastoma Cell Death Through p38 Signaling. Invest. Ophthalmol. Vis. Sci. 54(9):5937-43. PMID: 23942968 10) Zhang, Q*, Soderland, C, Steinle, JJ. Regulation of retinal endothelial cell apoptosis through activation of the IGFBP-3 receptor. 2013. Apoptosis. Mar;18(3):361-8. PMID: 23291901
Summary of your research:
- We have a number of projects in the laboratory. 1) We are developing the mechanism of action of a novel beta-adrenergic receptor agonist, Compound 49b, that prevents diabetic retinopathy in a rat model. Compound 49b is anti-apoptotic and anti-inflammatory. We are working on the cellular signaling pathways involved in these actions. 2) We have a project on insulin resistance in the retina. We are working on pathways by which TNFα elicits insulin resistance in retinal cells (endothelial cells and Müller cells). We are also testing drugs (both novel and commercially available) to block these actions Current Projects: 1) R01 from NEI: Compound 49b Prevents Retinal Endothelial Cell Death Through IGFBP-3 Levels Diabetic retinopathy remains the fifth leading cause of preventable blindness worldwide. Interventions to prevent progression of diabetic retinopathy are limited to improved glycemic control (a challenging goal for all diabetic patients) and to laser photocoagulation (available only for advanced stages of retinopathy). We and others have reported that adrenergic signaling is lost in the diabetic retina, suggesting that development of novel agents to restore autonomic homeostasis is necessary. Unfortunately, currently available adrenergic agents are associated with adverse systemic or non-specific effects. These problems inspired our group to synthesize compound 49b, a novel and selective β-adrenergic receptor agonist, as a potential paradigm shift in the prevention of diabetic retinopathy. Our preliminary data suggest that compound 49b prevents the formation of degenerate capillaries, which involves degenerate capillary formation, which are the hallmark pathology noted in the diabetic retinal vasculature. In addition to preventing degenerate capillaries in vivo, compound 49b prevents the cleavage of caspase 3, a well-established marker of apoptosis, in retinal endothelial cells (REC) in vitro, suggesting that Compound 49b can decrease apoptosis. In the oxygen-induced model of retinopathy, others have associated increased levels of insulin-like growth factor binding protein-3 (IGFBP-3) with protection from REC apoptosis. Furthermore, using the streptozotocin-induced diabetic rat model, we observed that chronic insulin deficiency reduced IGFBP-3 protein levels in whole retinal lysates, but topical application of compound 49b to the eye restored retinal IGFBP-3 to its control level in these insulin-deficient rats. Thus, we hypothesize that compound 49b prevents the critical vascular damage underlying diabetic retinopathy in part by restoring IGFBP-3 levels in retinal endothelial cells. This project focuses on a deeper understanding of the mechanisms underlying this protective action. 2) R01 from NEI Mechanisms of TNFalpha-Induced Insulin Resistance in Retinal Cells The proposed study will test the novel hypothesis that in the diabetic retina, hyperglycemia stimulates production of tumor necrosis factor α (TNFα), which in turn decreases insulin receptor binding leading to decreased signal transduction. The overall effect of this signaling cascade would be to create insulin resistance, exacerbate problems caused by limited insulin production in diabetes, and thus contribute to development of diabetic retinopathy seen in both type 1 and type 2 diabetes. While our preliminary data and previous reports by others support a major role for inflammatory mediators such as TNFα in diabetic retinopathy, the pathways involved are largely unknown. Our proposed studies will focus on one likely candidate, the suppressor of cytokine signaling 3 (SOCS3) pathway (Fig.1), which is poorly understood in retina and yet represents a promising therapeutic target in future treatments for diabetic retinopathy. Our overall goal is to 1) establish the role of the SOCS3 pathway in regulating insulin signaling (through insulin receptor substrate-1; IRS-1) and apoptosis in normal and diabetic rats and 2) evaluate effects of upstream drug targets on the SOCS3 pathway and their downstream effects on insulin signaling and retinal cell apoptosis

Susmit Suvas, PhD

Associate Professor in Ophthalmology

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Susmit Suvas, PhD

Associate Professor in Ophthalmology

Research Subject: To understand the immunopathogenesis of herpes simplex virus-1 infection induced keratitis Focus: Role of peripheral nervous system in regulating corneal HSV-1 infection and inflammation

Published Articles:
- • Gaddipati S, Estrada K, Rao P, Jerome A and Suvas S (2015). IL-2/anti-IL-2 complex treatment inhibits the development but not the progression of herpetic stromal keratitis. J Immunol. 194 (1): 273-82. (PMID: 25411200) • Alazabi FA, Zohdy, MA and Suvas S (2013). Parameter estimation from experimental laboratory data of HSV-1 by using alternative regression method. Systems and Synthetic Biology. 7(14): 151-60. (PMID: 24432152) • Channappanavar R, Twardy BS and Suvas S (2012). Blocking of PDL-1 interaction enhances primary and secondary CD8 T cell response to herpes simplex virus-1 infection. PLoS One. 7(7): e39757. (PMID: 22808056) • Twardy BS, Channappanavar R, Suvas S (2011). Substance P in the corneal stroma regulates the severity of herpetic stromal keratitis lesions. Invest Ophthalmol Vis Sci. 4; 52(12): 8604-13. (PMID: 21969295) • Channappanavar R, Twardy BS, Krishna P and Suvas S (2009). Advancing age leads to predominance of inhibitory receptor expressing CD4 T cells. Mech Ageing Dev 130 (10): 709-12. (PMID: 19715717) • Susmit Suvas (2008). Advancing age and immune cell dysfunction: is it reversible or not? Expert Opinion on Biological Therapy 8 (5): 657-68. (PMID: 18407768)
Summary of Research:
- The major focus of the laboratory is on the development of a novel optogenetic strategy for restoring sight to people who suffer from retinal degenerative diseases, such as retinitis pigmentosa and age-related macular degeneration. The optogenetic strategy for restoring vision utilizes genetically encoded light sensors to convert surviving retinal neurons into photosensitive cells, thus imparting light sensitivity to the retina after the death of photoreceptor cells. Our earlier proof-of-concept studies demonstrated the feasibility of this strategy by AAV-mediated expression of channelrhodopsin-2 (ChR2) in inner retinal neurons of photoreceptor-deficient mice. We also demonstrated the restoration of both ON and OFF light responses by expression of ChR2 and halorhodopsin in the retina. More recently, we also made important progress on the development of more light sensitive ChR2 as well as AAV vector constructs for selective targeting of the light sensors to specific retinal neurons and/or subcellular compartments. Our long-term goal is aimed at developing effective optogenetic therapies for treating blindness caused by retinal degenerative diseases. Another area of interest of the laboratory is the investigation of retinal visual information processing, especially the roles voltage-dependent membrane channels in retinal bipolar cell processing. This will provide new insight into our understanding of visual information processing in the retina and help to develop new treatment approaches for blinding retinal diseases. The experimental techniques used in our studies include molecular biology, AAV vectors, electrophysiological recordings, immunocytochemistry, and animal behavior.
Current Research:
- The laboratory is currently looking at the role of neuropeptides in corneal HSV-1 infection and inflammation in mouse model.

Lalit Singh Pukhrambam, Ph.D.

Assistant Professor of Anatomy/Cell Biology and Ophthalmology

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Lalit Singh Pukhrambam, Ph.D.

Assistant Professor of Anatomy/Cell Biology and Ophthalmology

Research Subject: Diabetic retinopathy Focus: Critical role of Thioredoxin Interacting Protein (TXNIP) in mitochondrial dynamics, stress response and neurovascular degeneration in diabetic retinopathy

Published Articles:
- 1. Perrone L, Matrone C, Singh L.P. (2014) Epigenetic modifications and potential new treatment targets in diabetic retinopathy. J Ophthalmol. 2014:789120. PMID: 25165577 2. Singh L.P. (2014) The NLRP3 Inflammasome and Diabetic Cardiomyopathy: Editorial to: “Rosuvastatin alleviates diabetic cardiomyopathy by inhibiting NLRP3 inflammasome and MAPK pathways in a type 2 diabetes rat model” by Beibei Luo et al. Cardiovasc Drugs Ther. 28(1):5-6. PMID: 24220800 3. Singh L.P. (2013) Thioredoxin Interacting Protein (TXNIP) and Progression of Diabetic Retinopathy. J Clin Exp Ophthalmol, 4: 287. PMID: 24353900 4. Devi TS, Hosoya KI, Terasaki T, Singh L.P. (2013) Critical role of TXNIP in oxidative stress, DNA damage, and apoptosis of pericytes in high glucose: Implications for diabetic retinopathy. Exp. Cell Res. 319:1001–1012. PMID: 23353834 5. Singh, L.P., Devi, TS, Nantwi, KD. (2012) Theophylline Regulates Inflammatory and Neurotrophic factor Signals in Functional Recovery after C2-Hemisection in Adult Rats. Exp. Neurol., 238:79-88. PMID: 22981449 6. Devi TS, Lee I, Hüttemann M, Kumar A, Nantwi KC, Singh L.P. (2012) Txnip links innate host defense mechanisms to oxidative stress and inflammation in retinal Muller glia under chronic hyperglycemia: Implications for diabetic retinopathy. Exp. Diab Res., 2012;2012:438238. PMID: 22474421 7. Devi TS, Hosoya K, Terasaki T, and Singh LP. (2011) GSK-3-CREB axis mediates IGF-1-induced ECM/adhesion molecule expression, cell cycle progression and monolayer permeability in retinal capillary endothelial cells: Implication for Diabetic Retinopathy. Biochim Biophys Acta-Molecular basis of disease 1812(9):1080-1088. PMID: 21549192 8. Sbai O, Devi TS, Melone MA, Feron F, Khrestchatisky M, Singh L.P., and Perrone L. (2010). RAGE-TXNIP axis is required for S100B-promoted Schwann cell migration, fibronectin expression and cytokine secretion. J Cell Science 123(Pt 24):4332-4332 PMID: 21098642 9. Perrone L, Devi TS, Hosoya K, Terasaki T, and Singh LP. (2010). Inhibition of TXNIP Expression In Vivo Blocks Early Pathologies of Diabetic Retinopathy. Cell Death and Disease 1, e65; doi:10.1038/cddis.2010.42. PMID: 21364670 10. Perrone L, Devi TS, Hosoya K, Terasaki T, and Singh LP. (2009). Thioredoxin-Interacting Protein (TXNIP) induces inflammation through chromatin modification in retinal capillary endothelial cells under diabetic conditions. J Cell Physiol. 221(1):262-272. PMID: 19562690
Summary of Research:
- Our laboratory is interested in developing mechanism-based and stage-specific therapies to treat diabetic retinopathy using peptide and RNAi technologies. We use both in vivo animal models of type-1 diabetes and in vitro retinal cell cultures to understand the molecular basis of early abnormalities in neurons and vasculature, especially mediated by mitochondrial (MT) dysfunction, oxidative stress and inflammation. We investigate the critical role played by TXNIP in redox imbalance, mitochondrial protein unfolding and MT-UPR and mitophagy in neurovascular degeneration and the progression of diabetic retinopathy. We use biochemical, molecular biology, proteomics, epigenetics, and in vivo non-invasive methods to measure diabetes duration-dependent neuronal injury, microvascular dysfunction and BRB breakdown.
Current Research:
- Mitochondrial dysregulation and resultant energy imbalance is associated with various chronic diseases including neuro-degeneration, ischemia/reperfusion, and diabetic complications including diabetic retinopathy. Recently, we published that pro-oxidant thioredoxin interacting protein (TXNIP) is significantly up-regulated in diabetic retinas and under hyperglycemia in retinal cells in culture such as endothelial cells, pericytes and Muller cells (MC) and mediates cellular oxidative stress, inflammation and apoptosis. TXNIP has recently been implicated by several studies as a critical protein in the pathogenesis of diabetes and its complications. TXNIP binds to thioredoxin (Trx), a redox anit-oxidant protein, inhibiting its reactive oxygen species (ROS) scavenging and thiol reducing capacity; therefore, results in cellular oxidative/nitrosative (ROS/RNS) stress and aberrant protein s-nitrosylation. Furthermore, MC are important for retinal neuronal health and activated MC (gliosis) induces aberrant gene expression for cytokines and growth factors to maintain retinal homeostasis. However, prolonged MC activation is injurious in DR. Therefore, our overall hypothesis is that TXNIP is critical for MT dysfunction, neurovascular injury, and MC activation in the development of DR. We are investigating the role of TXNIP in (i) MT dysfunction and retrograde stress responses (MT-UPR); and (ii) MT fission/fusion and mitophagy as a part of mitochondrial quality control in diabetic retinopathy. Is there a relationship between mitophagy deregulation and NLRP3 inflammasome activation and innate immune responses in diabetic retinopathy and, if any, to what extent TXNIP plays a role in this process are some of the questions being asked? To address these objectives, we will use streptozotocin (STZ)-induced type-1 diabetic models of rat and mouse in conjunction with manipulation of TXNIP expression levels (siRNA and knock-out mice) in the retina. In vitro studies using retinal MC, endothelial cells and neuronal cell cultures will be performed to dissect the molecular mechanisms as to how TXNIP induces MT dysfunction and mitophagy, which specifically removes damaged MT. Our studies may allow us to identify potential new targets for developing gene/drug therapies to prevent or slow down the progression of DR. Currently; our research is supported by NIH/NEI grant, NIH/NEI Core grant to the Department of Anatomy and Cell Biology and Research to Prevent Blindness to the Department of Ophthalmology.

Fu-Shin X. Yu, Ph.D.

Professor of Ophthalmology, Anatomy and Cell Biology

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Fu-Shin X. Yu, Ph.D.

Professor of Ophthalmology, Anatomy and Cell Biology

Research Subject: Corneal wound healing and innate immunity

Focus: i. Cellular and molecular mechanisms underlying development of peripheral neuropathy and delayed wound healing in diabetic cornea ii. Toll-like receptor-5 ligand flagellin induced innate mucosal immune protection against microbial keratitis (infection).

Published Articles:
- • Gao N, Kumar A, and Yu FS. MMP13 as a target for suppressing corneal ulceration caused by Pseudomonas aeruginosa infection. J. Infect Diseases. In press. • Liu X, Gao N, Dong C, Zhou L, Mi QS, Standiford TJ, et al. Flagellin-induced expression of CXCL10 mediates direct fungal killing and recruitment of NK cells to the cornea in response to Candida albicans infection. Eur J Immunol. 2014;44(9):2667-79. Epub 2014/06/27. doi: 10.1002/eji.201444490. PubMed PMID: 24965580; PubMed Central PMCID: PMC4165733. • Bettahi I, Sun H, Gao N, Wang F, Mi X, Chen W, et al. Genome-wide transcriptional analysis of differentially expressed genes in diabetic, healing corneal epithelial cells: hyperglycemia-suppressed TGFbeta3 expression contributes to the delay of epithelial wound healing in diabetic corneas. Diabetes. 2014;63(2):715-27. Epub 2013/12/07. doi: 10.2337/db13-1260. PubMed PMID: 24306208; PubMed Central PMCID: PMC3900551. • Yoon GS, Dong C, Gao N, Kumar A, Standiford TJ, Yu FS. Interferon regulatory factor-1 in flagellin-induced reprogramming: potential protective role of CXCL10 in cornea innate defense against Pseudomonas aeruginosa infection. Invest Ophthalmol Vis Sci. 2013;54(12):7510-21. Epub 2013/10/17. doi: 10.1167/iovs.13-12453. PubMed PMID: 24130180; PubMed Central PMCID: PMC3832217. • Gao N, Sang Yoon G, Liu X, Mi X, Chen W, Standiford TJ, et al. Genome-wide transcriptional analysis of differentially expressed genes in flagellin-pretreated mouse corneal epithelial cells in response to Pseudomonas aeruginosa: involvement of S100A8/A9. Mucosal Immunol. 2013;6(5):993-1005. Epub 2013/01/24. doi: 10.1038/mi.2012.137. PubMed PMID: 23340821; PubMed Central PMCID: PMC3722258. • Wang F, Gao N, Yin J, Yu FS. Reduced innervation and delayed re-innervation after epithelial wounding in type 2 diabetic Goto-Kakizaki rats. Am J Pathol. 2012;181(6):2058-66. Epub 2012/10/16. doi: 10.1016/j.ajpath.2012.08.029. PubMed PMID: 23063510; PubMed Central PMCID: PMC3509759. • Gao N, Yin J, Yoon GS, Mi QS, Yu FS. Dendritic cell-epithelium interplay is a determinant factor for corneal epithelial wound repair. Am J Pathol. 2011;179(5):2243-53. Epub 2011/09/20. doi: 10.1016/j.ajpath.2011.07.050. PubMed PMID: 21924232; PubMed Central PMCID: PMC3204011. • Yin J, Huang J, Chen C, Gao N, Wang F, Yu FS. Corneal complications in streptozocin-induced type I diabetic rats. Invest Ophthalmol Vis Sci. 2011;52(9):6589-96. Epub 2011/07/01. doi: 10.1167/iovs.11-7709. PubMed PMID: 21715347; PubMed Central PMCID: PMC3176029. • Xu K, Yu FS. Impaired epithelial wound healing and EGFR signaling pathways in the corneas of diabetic rats. Invest Ophthalmol Vis Sci. 2011;52(6):3301-8. Epub 2011/02/19. doi: 10.1167/iovs.10-5670. PubMed PMID: 21330660; PubMed Central PMCID: PMC3109029. • Gao N, Kumar A, Guo H, Wu X, Wheater M, Yu FS. Topical flagellin-mediated innate defense against Candida albicans keratitis. Invest Ophthalmol Vis Sci. 2011;52(6):3074-82. Epub 2011/02/12. doi: 10.1167/iovs.10-5928. PubMed PMID: 21310913; PubMed Central PMCID: PMC3109016.
Summary of your research:
- The Yu’s laboratory investigates the molecular mechanisms underlying corneal epithelial wound healing and diabetic neurotrophic keratopathy, using corneal organ culture and animal models. Specifically, the lab studies the interactions of sensory nerve, intraepithelial dendritic cells, and epithelial cells in the cornea during wound healing and their defects in diabetic corneas which lead to delayed epithelial wound closure and impaired sensory nerve regeneration. The second project of Dr. Yu’s lab is to study ocular innate immunity and protection against infection. Dr. Yu and colleagues have shown that activation of Toll-like receptors, particularly TLR5 prior to infection resulted in profound mucosal protection against a broad spectrum of keratitis causing pathogens. The focus in Dr. Yu’s lab is to define the molecular mechanisms underlying these protective pathways.
Current Research:
- Molecular regulation of corneal wound healing Mechanisms of flagellin induced protection against infection keratitis

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Linda D. Hazlett

Distinguished Professor & Chair, Department of Anatomy and Cell Biology

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Linda D. Hazlett

Distinguished Professor & Chair, Department of Anatomy and Cell Biology

Anatomy & Cell Biology and Distinguished Professor Ophthalmology & Immunology/Microbiology Research Subject: Corneal infection Focus: Host immune response to bacterial keratitis

Selected Recent Published Articles:
- 1. Berger, E.A., Vistisen K.S., Barrett, R., and Hazlett, L.D. Effects of VIP on corneal reconstitution and homeostasis following Pseudomonas aeruginosa induced keratitis. IOVS, 53:7432-9, 2012. PMID 23036997 PMCID:PMC3061511 2. Jiang, X., McClellan, S.A., Barrett, R.P., Zhang, Y., Foldenauer M., and Hazlett, L.D. The role of VIP in cornea. Invest. Ophthalmol. Vis. Sci., 53: 7560-6, 2012. PMID:23074208. PMCID:PMC3493186 3. Sun, M., Zhu, M., Nie, X., Deng, Q., Chen, K., Hazlett, L., Wu, M., and Huang, X. TREM-2 promotes host resistance against Pseudomonas aeruginosa by suppressing corneal inflammation via PI3K/Akt signaling pathway. IOVS, 17:3452-62, 2013. PMCID:PMC3658264 4. Foldenauer, M., McClellan, S., and Hazlett, L.D. mTOR regulates IL-10 and resistance to Pseudomonas aeruginosa corneal infection. J. Immunology, 190:5649-58, 2013. PMCID:PMC3660542 5. Berger, E., McClellan, S., Vistisen, K., and Hazlett, L.D. HIF-1α is essential for effective PMN bacterial killing, antimicrobial peptide production and apoptosis in Pseudomonas aeruginosa keratitis PLOS PATHOGENS 9(7): 18 July, 2013 e1003457. doi:10.1371/journal.ppat.1003457. PMCID:PMC3715414 6. Jiang, X., McClellan, S., Barrett, R., Foldenauer, M., and Hazlett, L.D. HGF signaling impacts severity of P. aeruginosa keratitis. IOVS, 55:2180-90, 2014. PMCID:PMC3985408 7. Hazlett, L. Linda Hazlett speaks to Hannah Branch, Commissioning Editor. Future Microbiol. 709-710, 2013. 8. Hazlett, Linda D., Jiang, Xiaoyu, and McClellan, Sharon A. IL-10 Function, Regulation, and in Bacterial Keratitis. Journal of Ocular Pharmacology and Therapeutics 30: 373-380, 2014. PMCID:PMC4043257 9. Li, C., McClellan, S.A., Barrett, R., Hazlett, L.D. Interleukin 17 regulates Mer tyrosine kinase-positive cells in Pseudomonas aeruginosa keratitis. IOVS 55:6886-900,2014. PMCID:PMC4214210 10. High-mobility group box 1: A novel target for treatment of Pseudomonas aeruginosa keratitis J.Immunology, in press 2015.
Summary of Research:
- Dr. Hazlett’s research is focused on the host immune response to the gram negative bacterium Pseudomonas aeruginosa (P. aeruginosa). The ubiquitous bacterium is an opportunistic pathogen noted both for its ability to cause disease in particularly susceptible or compromised individuals and its ever-increasing resistance to antibiotics. Extremely common in soil and water, it is also responsible for many serious, often fatal, human infections, including upper respiratory tract colonization in cystic fibrosis patients and in ocular infections, particularly due to contact lens usage, where blindness can be the outcome. With contact lens use at an all time high, such sight-threatening infections are becoming increasingly widespread and range from purulent conjunctivitis to iridocyclitis, keratitis and iritis, corneal ulcer, and panophthalmitis. In fact, of the 30,000,000 daily-wear contact lens users worldwide, one in every 2,500 will suffer pseudomonal infection; the odds increase by a factor of five – or one in every 500 – for those using extended-wear or overnight lenses. In the United States alone, 25,000-30,000 cases of microbial keratitis are reported annually with an estimated cost of between $15 and $30 million for treatment. Dr. Hazlett has authored 167 peer reviewed publications on this topic in high quality journals, including the Journal of Immunology, Progress in Retinal and Eye Research (2004 publication, solicited Review Article by this number one eye journal) and Investigative Ophthalmology and Visual Science. She also has contributed 22 books/chapters/review articles, one of which is a chapter contribution to a clinical textbook (Duane’s Clinical Ophthalmology and Foundations of Clinical Ophthalmology, 2006) entitled, “Pathogenesis of Pseudomonas Keratitis”. The long-term objective of her studies is to reveal basic pathophysiological mechanisms of disease in the P. aeruginosa infected cornea. Her studies directly address a major objective of the current Corneal Diseases Program (National Plan for Eye and Vision Research) which parenthetically states, to “investigate corneal infectious processes and immunological responses and to develop treatments to reduce keratitis and prevent blindness.” Her studies integrate cellular and molecular biology and immunology and continue to elucidate novel targets in the immune response that will impact clinical care of patients with P. aeruginosa keratitis
Current Research
- Dr. Hazlett studies the role of the host response to bacterial infection of the cornea as a major focus of her scientific work. As her publications suggest, she has been doing this for a considerable amount of time and with high productivity and success. She has specifically focused on two areas one, Toll-like receptors (TLR) and the other, neuropeptides, that modulate the host response to bacterial infection. Her goal is to better understand the disease process in which inflammation, if uncontrolled, is deleterious and to develop rational intervention therapies that will have clinical relevance. Her current R01 focuses on the pathogenesis and treatment potential of interventions against high-mobility group box 1 (HMGB1), a prototypic alarmin. HMGB1 is a mediator of the systemic inflammatory response syndrome, is elevated late in bacterial infection/sepsis and considered a target for disease treatment, but nothing is known about its role in bacterial keratitis. Given that it is important in innate immunity, has different functions dependent on cellular localization, and has the ability to bind to TLR and other molecules such as receptor for advanced glycation end products (RAGE), the hypothesis that she is testing is that HMGB1 has significant amplification effects on resident and infiltrating inflammatory cells and is an attractive therapeutic target in Pseudomonas (P.) aeruginosa keratitis. Dr. Hazlett also is PI of Wayne State’s NEI Core Vision grant and uses the facilities of the Core for her own studies and together with the Ophthalmology chair, Mark Juzych, M.D., continues to advance vision research at Wayne State University principally through recruitment and development of the research enterprise.

Elizabeth Berger, PhD

Assistant Professor in Anatomy & Cell Biology and Ophthalmology

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Elizabeth Berger, PhD

Assistant Professor in Anatomy & Cell Biology and Ophthalmology

Research Subject: Ocular Immunology Focus: Bacterial Keratitis

Published Articles:
- Berger EA, McClellan SA,Vistisen KS, Hazlett LD. HIF-1a is essential for effective PMN bacterial killing, antimicrobial peptide production and apoptosis in Pseudomonas aeruginosa keratitis. PLoS Pathogen. 9(7):e1003457, Epub Jul 18, 2013. PMID:23874197
  Foldenauer MEB, McClellan SA, Berger EA, Hazlett LD. Mammalian target of rapamycin regulates IL-10 and resistance to Pseudomonas aeruginosa corneal infection. J Immunol. Jun 190(11):5649-5658, 2013. PMID: 23626014
  Berger EA, Vistisen KS, Barrett RP, Hazlett LD. Effects of VIP on corneal reconstitution and homeostasis following Pseudomonas aeruginosa induced keratitis. Invest Ophthalmol Vis Sci. Nov 53(12):7432-7439, 2012. PMID: 23036997
  Jiang X, McClellan S, Barrett RP,Berger EA, Zhang Y, Hazlett LD. VIP and growth factors in the infected cornea. Invest Ophthalmol Vis Sci. Aug 52(9): 6154-6161, 2011. PMID: 21666233
  Berger EA, McClellan S, Barrett RP, Hazlett LD. Testican-1 promotes resistance against P. aeruginosa induced keratitis via modulation of MMP-2 driven wound healing and ECM restoration. Invest Ophthalmol Vis Sci. Jul 52(8):5339-5346, 2011. PMID: 21613368
  Berger EA, McClellan S, Barrett RP, Hazlett LD. VIP promotes resistance in the Pseudomonas aeruginosa infected cornea by modulating adhesion molecule expression. Invest Ophthalmol Vis Sci. 51(11):5776-5782, 2010. PMID: 20592225
  Zhou Z, Barrett R, McClellan S, Zhang Y,Szliter E, Huang X, Hazlett LD. Substance P delays apoptosis in P. aeruginosa infected cornea and exacerbates disease. Invest Ophthalmol Vis Sci. Oct; 49(10): 4458-4467, 2008. PMID: 18566468
  Szliter EA, Lighvani S, Barrett RP, Hazlett LD. VIP balances pro- and anti-inflammatory cytokines in the P.aeruginosa infected cornea and protects against corneal perforation. J Immunol. Jan; 178(2): 1105-1114, 2007. PMID:17202374
  Szliter EA, Barrett RP, Gabriel MM, Zhang Y, Hazlett LD. Pseudomonas aeruginosa -induced inflammation in the rat extended-wear contact lens model. Eye Contact Lens. Jan; 32(1): 12-18, 2006. PMID: 16415687
  Szliter EA, Morris C.A, Carney F, Gabriel, MM, Hazlett LD. Development of An Extended Wear Contact Lens Model In the Rat. CLAO J. Jul; 28(3): 119-123, 2002. PMID: 12144229
Summary of Research:
- Title: Assistant Professor in Anatomy & Cell Biology and Ophthalmology.The studies carried out by our laboratory predominately focus on disease pathogenesis and the interplay between the immune and neuroendocrine systems. We mechanistically and therapeutically investigate the events of inflammation and innate immunity using a model of ocular infectious disease induced by a common pathogen of eye infections – Pseudomonas aeruginosa. This includes analyses of: host inflammatory cells (macrophages, neutrophils, T cells – both in vivo and in vitro), extracellular matrix and adhesion molecules, cytokines/chemokines, Toll-like receptors and other related molecules using a number of molecular, cellular and immunohistochemical techniques.
Current Research:
- Ongoing NIH-funded studies examine how the neuropeptide, vasoactive intestinal peptide(VIP), influences T cells and the production of IL-17, nerve regeneration, and the interaction with endogenous lipid mediator circuits. In addition, my laboratory is defining the clinical applicability of VIP as a treatment modality in the eye. Additionally, other projects include: 1) how neutrophil transdifferentiation contributes to disease pathogenesis and 2) collaborative projects looking at miRNAs, as well as a novel gene therapy strategy for the treatment of glaucoma.

Bruce Berkowitz, Ph.D.

Diabetic retinopathy; retinal physiology

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Bruce Berkowitz, Ph.D.

Diabetic retinopathy; retinal physiology

Published Articles:
- http://www.ncbi.nlm.nih.gov/pubmed?term=berkowitz%20b*%20%5Bau%5D%20AND%20retina&cmd=Search
Summary of Research:
- Significant advances made in the Berkowitz lab include new information concerning panretinal and vascular and avascular border oxygenation, as well as the role of calcium channels, in retinal neovascular models, the benefits of steroid treatment on retinopathy, the first identification of a diabetic retinal edema model, and the importance of photoreceptor L-type calcium channels in diabetic retinopathy.
Current Research:
- Current imaging methods do not comprehensively measure essential photoreceptor functions of light detection, regulated transmission of information, and visual pigment regeneration. Our new suite of novel biomarkers greatly expands the number of analytical imaging tools available for comprehensive and objective assessment of these critical photoreceptor functions. Our new methods are transformative because they optimally measure photoreceptor function in health, in disease, and during treatment beyond that currently available, and can be applied in future translational human studies.

Tomomi Ichinose, MD, PhD

Assistant Professor

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Tomomi Ichinose, MD, PhD

Assistant Professor

Research Subject: Mechanisms of Visual Signal Processing in the RetinaFocus: Physiological functions of bipolar cells in the mouse retina

Published Articles:
- 1. Hellmer CH, Ichinose T: Recording light-evoked postsynaptic responses in neurons in dark-adapted, mouse retinal slice preparations using patch clamp techniques. Journal of Visualized Experiments (JoVE) In Press, 2015 2. Fyk-Kolodziej B, Hellmer CH, Ichinose T: Marking cells with infrared fluorescent proteins to preserve photoresponsiveness in the retina. BioTechniques 57:245-253, 2014
  3. Ichinose T, Fyk-Kolodziej B, Cohn J: Roles of ON cone bipolar cell subtypes in temporal coding in the mouse retina. Journal of Neuroscience 34:8761-8771, 2014 4. Ichinose T, Lukasiewicz PD: The mode of retinal presynaptic inhibition switches with light intensity. Journal of Neuroscience 32:4360-4371, 2012 5. Ichinose T, Lukasiewicz PD: Light and dopamine regulate sodium channels to dynamically control retinal signaling. Journal of Neuroscience 27:4756-4764, 2007 6. Ichinose T, Lukasiewicz PD: Inner and outer retinal pathways both contribute to surround inhibition of salamander ganglion cells. Journal of Physiology 565 (Pt 2):517-35, 2005 7. Ichinose T, Shields CR, Lukasiewicz PD: Sodium channels in transient retinal bipolar cells enhance visual responses in ganglion cells. Journal of Neuroscience 25(7):1856-65, 2005 8. Ichinose T, Yu S, Wang XQ, Yu SP: Ca2+-independent, but voltage- and activity-dependent regulation of the NMDA receptor outward K+ current in mouse cortical neurons. Journal of Physiology 551(Pt 2):403-17, 2003 9. Ichinose T, Lukasiewicz PD: GABA transporters regulate inhibition in the retina by limiting GABA(C) receptor activation. Journal of Neuroscience 22(8):3285-92, 2002 10. Manzerra P, Behrens MM, Canzoniero LM, Wang XQ, Heidinger V, Ichinose T, Yu SP, Choi DW: Zinc induces a Src family kinase-mediated up-regulation of NMDA receptor activity and excitotoxicity. Proceedings of the National Academy of Sciences U S A 98(20):11055-61, 2001 11. Ichinose T, Snider WD: Differential effects of TrkC isoforms on sensory axon outgrowth. Journal of Neuroscience Research 59(3):365-71, 2000
Summary of Research:
- Our visual system perceives the world as functions of time, space, and color. The temporal visual processing pathways encode time (temporal) information of an object. These pathways are broadly classified into transient pathways and sustained pathways based on their neural responses, which are thought to encode changing objects and static visual information, respectively. The transient/sustained dichotomy is recognized in retinal second order neurons, which are bipolar cells, suggesting that temporal processing originates in bipolar cells. Recently, morphological studies have revealed more than 10 subtypes of bipolar cells in the mammalian retina, which might initiate distinct pathways encoding different features of temporal signals. However, no studies have addressed how each subtype of bipolar cell in the mammalian retina encodes temporal visual information. In response to this research gap, my goal is to understand the cellular and molecular mechanisms of temporal encoding in each subtype of bipolar cell. We use electrophysiological recording techniques and immunohistochemical method to analyze temporal visual encoding in individual bipolar cells. We found that each ON bipolar cell encode different aspects of temporal visual information in a subtype-dependent manner (Figure 1) (Ichinose et al., 2014). We are currently working on the elucidation of underlying mechanisms of temporal encodings. These studies will improve our understanding of parallel processing in the retinal neural network, which will contribute to the future design of a functional retinal prosthesis and sight restoring gene therapies. Figure 1. Six subtypes of ON bipolar cells and their responses to step-light stimuli.

Renu A. Kowluru, PhD

(Ex. Research Assistant Professor in Ophthalmology) Professor

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Renu A. Kowluru, PhD

(Ex. Research Assistant Professor in Ophthalmology) Professor

Research Subject: Molecular Mechanism of Pathogenesis of Diabetic Retinopathy Focus: Mitochondrial Dysfunction and Epigenetic Modifications

Published Articles:
- Santos JM. Kowluru RA. Metabolic memory and diabetic retinopathy: Continued impaired retinal mitochondria biogenesis in diabetic rats. Invest Ophthalmol Vis Sci 52:8791-8798, 2011 (PMCID:PMC3202722) Tewari S, Santos JM and Kowluru RA. Damaged mitochondrial DNA replication system and the development of diabetic retinopathy. Antioxid Redox Signal 17:492-504, 2012 (PMCID:PMC3365359) Santos JM, Tewari S. Kowluru RA. A compensatory mechanism protects retinal mitochondria from initial insult in diabetic retinopathy. Free Rad Biol Med 53:1729-1737, 2012 (PMCID:PMC3632051) Kowluru RA, Kowluru A, Veluthakal R, Mohammad G, Syed I, Santos JM, Mishra M. TIAM1-RAC1 signalling axis-mediated activation of NADPH oxidase-2 initiates mitochondrial damage in the development of diabetic retinopathy. Diabetologia 57:1047-1056, 2014 (PMID:24554007) Zhong Q, Kowluru RA. Epigenetic modification of Sod2 in the development of diabetic retinopathy and in the metabolic memory: Role of histone methylation. Invest Ophthalmol Vis Sci 54:244-250, 2013 (PMCID:PMC3590072) Zhong Q, Kowluru RA. Regulation of matrix metallopeptidase-9 by epigenetic modifications, and the development of diabetic retinopathy. Diabetes 62:2559-2568, 2013 (PMCID:PMC3712057) Santos JM, Kowluru RA. Impaired transport of mitochondrial transcription factor A (TFAM) and the metabolic memory phenomenon associated with the progression of diabetic retinopathy. Diabetes Metab Res Rev 29:204-213, 2013 (PMCID:PMC3596457) Kowluru RA, Santos JM, Zhong Q. Sirt1, a negative regulator of matrix metalloproteinase-9 in diabetic retinopathy. Invest Ophthalmol Vis Sci 55:5653-5660, 2014 (PMID:24894401) Mishra M, Zhong Q, Kowluru RA. Epigenetic modifications of Nrf2-mediated glutamate-cysteine ligase: implications for the development of diabetic retinopathy and the metabolic memory phenomenon associated with its continued progression. Free Radic Biol Med 75:129-139, 2014 (PMID:25016074) Mishra M, Kowluru RA. Retinal mitochondrial DNA mismatch repair in the development of diabetic retinopathy, and its continued progression after termination of hyperglycemia. Invest Ophthalmol Vis Sci 55:6960-6967, 2014 (PMID:25249609). Mishra M, Zhong Q, Kowluru RA. Epigenetic modifications of Keap1 regulate its interaction with the protective factor Nrf2 in the development of diabetic retinopathy. Invest Ophthalmol Vis Sci 55:7256-7265, 2014 (PMID:25301875)
Ongoing Research:
- The overall objective of our research is to understand the pathogenesis of diabetic retinopathy, and the research is focused on investigating the role of mitochondrial damage in the development/ progression of diabetic retinopathy. Our studies have shown that capillary cells undergo accelerated apoptosis before histopathology characteristic of diabetic retinopathy can be observed in rodent models of diabetic retinopathy. Mitochondria become dysfunctional, their DNA is damaged, biogenesis is impaired, and cytochrome c starts to leak out into the cytosol activating the apoptotic machinery. Mitochondrial superoxide scavenging enzyme is subnormal, and its promoter and enhancer are epigenetically modified. However, the damage to the mitochondria precedes overproduction of reactive oxygen species by cytosolic NADPH oxidase, and continuous production of these reactive species in a hyperglycemic milieu damages the mitochondria. Using rodent models of diabetic retinopathy, and retinal vascular cells in culture, our laboratory is investigating the signaling mechanisms responsible for mitochondrial damage, and the role of epigenetic modifications of the nuclear genes and mitochondrial DNA in the homeostasis of mitochondrial function.

Ashok Kumar, Ph.D.

Assistant Professor, Department of Ophthalmology

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Ashok Kumar, Ph.D.

Assistant Professor, Department of Ophthalmology

Department of Anatomy and Cell Biology Wayne State University School of Medicine Kresge Eye Institute

Awards & Honors
- 2010: Member, Scientific Review Committee, Fight for Sight foundation. 2010: Editorial board member, World Journal of Gastrointestinal Pathophysiology (WJGP) 2008: Grant Initiative Program Award, Wayne State University, School of Medicine 2008: KEI intramural research grant award. 2007: Fight for Sight grant-in-aid research award. 2006: Midwest eye bank research grant award. 2006: Association for Research in Vision and Ophthalmology (ARVO), travel grant. 2005: Fight for Sight post-doctoral fellowship award 1998: Post Grad Institute of Medical Education & Research fellowship. 1997: Secured first position in order of merit in Master’s Program For more information about Dr. Kumar, click here.

Zhuo-Hua Pan

Professor

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Zhuo-Hua Pan

Professor

Published Articles (since 2000):
- Pan, Z.-H., Hu, H.-J. Voltage-dependent Na+ currents in mammalian retinal cone bipolar cells. J. Neurophysiol. 84:2564-2571, 2000. Pan, Z.-H. Voltage-gated Ca2+ channels and GABA receptors revealed at presynaptic terminals of mammalian retinal bipolar cells. Visual Neurosci. 18:279-288, 2001. Pan, Z.-H., Hu, H.-J., Perring, P., Andrade, R. T-type Ca2+ channels mediate neurotransmitter release in retinal bipolar cells. Neuron 32:89-98, 2001. Hu, H.-J., Pan, Z.-H. Differential expression of K+ currents in mammalian retinal bipolar cells. Visual Neurosci. 19:163-173, 2002. Ma, Y.-P., Pan, Z.-H. Spontaneous regenerative activity in mammalian retinal bipolar cells: roles of multiple subtypes of voltage-dependent Ca2+ channels. Visual Neurosci. 20:131-139, 2003. Ma, Y.-P., Cui, J., Hu, H.-J., Pan, Z.-H. Mammalian retinal bipolar cells express inwardly rectifying K+ currents (IKir) with a different distribution than that of Ih. J. Neurophysiol. 90:3479-3489, 2003. Cui, J., Ma, Y.-P., Lipton, S.A., Pan Z.-H. Glycine receptors and glycinergic synaptic input at the axon terminals of mammalian retinal rod bipolar cells. J. Physiol. 553:895-909, 2003. Ma, Y.-P., Cui, J., Pan, Z.-H. Heterogeneous expression of voltage-dependent Na+ and K+ channels in mammalian retinal bipolar cells. Visual Neurosci. 22:119-133, 2005. Bi, A., Cui, J., Ma, Y.-P., Olshevskaya, E., Pu, M., Dizhoor, A.M., and Pan Z.-H. Ectopic expression of a microbial-type rhodopsin restores visual responses in mice with photoreceptor degeneration. Neuron 50:23-33, 2006. Cui, J. and Pan Z.-H. Two types of cone bipolar cells express voltage-gated Na+ channels in the rat retina. Visual Neurosci. 25: 635-645, 2008. Hu, C., Bi, A, and Pan Z.-H. Differential expression of three T-type calcium channels in retinal bipolar cells in rat. Visual Neurosci. 26:177-187, 2009. Zhang, Y., Ivanova, E., Bi, A., and Pan, Z.-H. Ectopic expression of multiple microbial rhodopsins restores ON and OFF light responses in the retina after photoreceptor degeneration. J. Neurosci. 29:9186-96, 2009. Lu, Q., Ivanova, E., and Pan Z.-H. Characterization of GFP-expressing retinal cone bipolar cells in a 5-HTR2a transgenic mouse line. Neurosci. 163:662-668, 2009. Ivanova, E, Pan, Z.-H. Evaluation of virus mediated long-term expression of channelrhodopsin-2 in the mouse retina. Mol. Vision 15:1680-1689, 2009. Ivanova, E., Hwang, G.S. and Pan, Z.-H. Characterization of transgenic mouse lines expressing Cre-recombinase in the retina. Neurosci. 165: 233-243, 2010. Ivanova, E., Hwang, G.-S., Pan, Z.-H., and Troilo, D. Evaluation of AAV-mediated expression of chop2-GFP in the marmoset retina. IOVS 51: 5288-5296, 2010. Ivanova, E, Roberts, R. Bissig, D., Pan, Z.-H., and Berkowitz, B.A. Retinal and optic nerve channelrhodopsin-2-mediated activity in vivo evaluated with manganese-enhanced MRI. Mol. Vision 16:1059-67, 2010. Wu, C., Ivanova, E., Cui, J., Lu Q. and Pan. Z.-H. Action potential generation at an AIS-like process in the axonless retinal AII amacrine cell. J. Neurosci. 31:14654-14659, 2011. Cui, J., Ivanova, E., Lu Q., and Pan, Z.-H. Expression of CaV3.2 T-type Ca2+ channels in a subpopulation of retinal type-3 cone bipolar cells. Neurosci. 224:63-9, 2012. Shimano, T., Fyk-Kolodziej, B., Mirza, N., Asako, M., Koichi, T., Bledsoe, S., Pan, Z.-H., Molitor, S., Holt, A.G. Assessment of the AAV- mediated expression of channelrhodopsin-2 and halorhodopson in brainstem neurons mediating auditory signaling. Brain Research, 1511:138-52, 2013. Ivanova, E., Lee, P., and Pan, Z.-H. Characterization of multiple bistratified retinal ganglion cells in a purkinje cell protein 2-cre transgenic mouse line. J. Comp. Neurol. 521:2165-80, 2013. Lu, Q., Ivanova, E., Ganjawala, H. T., and Pan, Z.-H. Cre-mediated recombination efficiency and transgene expression patterns of three retinal bipolar cell-expressing Cre transgenic mouse lines. Mol. Vision, 19:1310-1320, 2013. Wu, C., Ivanova, E., Zhang, Y., Pan, Z.-H. AAV-mediated subcellular targeting of optogenetic tools in retinal ganglion cells. PloS One, 8(6):e66332, 2013. Pan, Z.-H., Ganjawala, T.H., Lu, Q., Ivanova, E., Zhang, Z. ChR2 mutants at L132 and T159 with improved operational light sensitivity for vision restoration. PloS One, 9(6):e98924, 2014.
Summary of Research:
- The major focus of the laboratory is on the development of a novel optogenetic strategy for restoring sight to people who suffer from retinal degenerative diseases, such as retinitis pigmentosa and age-related macular degeneration. The optogenetic strategy for restoring vision utilizes genetically encoded light sensors to convert surviving retinal neurons into photosensitive cells, thus imparting light sensitivity to the retina after the death of photoreceptor cells. Our earlier proof-of-concept studies demonstrated the feasibility of this strategy by AAV-mediated expression of channelrhodopsin-2 (ChR2) in inner retinal neurons of photoreceptor-deficient mice. We also demonstrated the restoration of both ON and OFF light responses by expression of ChR2 and halorhodopsin in the retina. More recently, we also made important progress on the development of more light sensitive ChR2 as well as AAV vector constructs for selective targeting of the light sensors to specific retinal neurons and/or subcellular compartments. Our long-term goal is aimed at developing effective optogenetic therapies for treating blindness caused by retinal degenerative diseases. Another area of interest of the laboratory is the investigation of retinal visual information processing, especially the roles voltage-dependent membrane channels in retinal bipolar cell processing. This will provide new insight into our understanding of visual information processing in the retina and help to develop new treatment approaches for blinding retinal diseases. The experimental techniques used in our studies include molecular biology, AAV vectors, electrophysiological recordings, immunocytochemistry, and animal behavior.
Current Research:
- 1) Develop and optimize optogenetic light sensors for restoring vision. 2) Develop virus-mediated targeting of optogenetic light sensors to specific inner retinal cell types and/or subcellular compartments. 3) Evaluate optogenetic approaches with electrophysiologial recordings and animal behavioral experiments. 4) Investigate the roles voltage-dependent membrane channels in the information processing of retinal bipolar cells.

Shunbin Xu, M.D. and Ph.D.

Associate professor

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Shunbin Xu, M.D. and Ph.D.

Associate professor

Published Articles:
- 1. Cowan C, Muraleedharan CK, O’Donnell JJ, III, Singh PK, Lum H, Kumar A, and Xu S. microRNA-146 Inhibits Thrombin-induced NF-κB Activation and Subsequent Inflammatory Responses in Human Retinal Endothelial Cells. Invest Ophthalmol Vis Sci. 2014 Jul 1;55(8):4944-51. PMID: 24985472. 2. Lumayag S, Haldin CE, Corbett NJ, Wahlin KJ, Cowan C, Turturro S, Larsen P, Kovacs B, Witmer PD, Valle D, Zack DJ, Nicholson DA, and Xu S. Inactivation of the microRNA-183/96/182 cluster results in syndromic retinal degeneration. PNAS 2013 Feb 5;110(6):E507-16. doi: 10.1073/pnas.1212655110. Epub 2013 Jan 22.. PMID: 23341629. 3. Kovacs B, Lumayag S, Cowan C, Xu S. microRNAs in early diabetic retinopathy in streptozotocin-induced diabetic rats. Invest. Ophthalmol. Vis. Sci. 2011 Jun 21; 52(7): 4402-9. Print 2011 Jun. PMID: 21498619 4. Hildebrand MS, Witmer PD, Xu S, Newton SS, Kahrizi K, Najmabadi H, Valle D, Smith RJH. miRNA Mutations are Not a Common Cause of Deafness. Am J Med Genet A. 2010 Mar;152A(3):646-52. PMID: 20186779. 5. Xu S. microRNA expression in the eyes and their significance in relation to functions. Prog Retin Eye Res. 2009 Mar;28(2):87-116. Epub 2008 Nov 28. (invited review). PMID: 19071227. 6. Xu S*, Witmer D, Kovacs B, Lumayag S and Valle D*. (2007). MicroRNA Transcriptome Of Mouse Retina And Functional Study of a Sensory Organ Specific miRNA cluster. J Biol Chem. 282(34):25053-25066. PMID: 15456885. (*: corresponding authors). 7. Perez SE, Lumayag S, Kovacs B, Mufson EJ and Xu S. (2008) β-Amyloid Deposition and Functional Impairment in the Retina of the APPswe/PS1∆E9 Transgenic Mouse Model of Alzheimer’s Disease. Invest Ophthalmol Vis Sci. 2009 Feb;50(2):793-800. PMID: 18791173. 8. Kovacs B, MacCumber MW, Xu S. (2007). Adult Retinal Stem Cell Spheres Hold Potential for Diabetic Retinopathy (DR) Treatment. Retina Today. Nov/Dec 2007: 41-42. 9. Liu H, Xu S, Wang Y, Mazerolle C, Thurig S, Coles BLK, Ren J, Taketo MM, van der Kooy D, Wallace VA. (2007). Ciliary margin transdifferentiation from neural retina is controlled by canonical Wnt signaling, Developmental Biology 308(1):54-67. PMID: 17574231. 10. Xu S, Sunderland ME, et al. (2007). The proliferation and expansion of retinal stem cells require functional Pax6. Dev Biol. 304(2):713-21. PMID: 17316600. 11. Smukler SR, Runciman SB, Xu S, van der Kooy D. (2006). Embryonic stem cells assume a primitive neural stem cell fate in the absence of extrinsic influences. J Cell Biol. 172(1):79-90. PMID: 16390999.
Summary of Research:
- The research interest of my laboratory is to study the roles of a new class of molecules – microRNAs (miRNAs), in the eye and ocular diseases. miRNAs are small, non-coding, regulatory RNAs and constitute a newly recognized level of gene expression regulation. They exist in every cells of our body, and play important roles regulating the expression of protein-coding genes, and can cause diseases when they are defective. However, the roles of these newly discovered molecules in the eye and ocular diseases are largely unknown. Our long term goal is to uncover the roles miRNAs in normal development and function of the eye and in ocular diseases, and to identify novel miRNA-based diagnostic biomarkers and therapeutic targets for early diagnosis and treatment of various ocular diseases, including diabetic retinopathy, inherited retinal degeneration, glaucoma, and age-related macular degeneration and corneal keratitis.
Current Research:
- Currently, the following major projects are ongoing in the lab: 1) microRNAs in retinal degenerative diseases and aging: Previously, we identified all miRNAs exist in adult mouse and human retina. Among these retinal miRNAs, we focus on a conserved miRNA cluster – the miR-183/96/182 cluster, which produces three miRNAs, miR-183, miR-96 and miR-182, which are highly expressed in the retina and all major sensory organs (Xu et al. 2007. JBC). We showed that inactivation of miR-183/96/182 in mice causes congenital retinal dysfunction, progressive retinal degeneration, and multiple sensory defects (Lumayag et al. PNAS. 2013). Currently, we are working on molecular mechanisms underlying retinal and other sensory diseases caused by inactivation of miR-183/96/182, and identification of disease-causing mutations in miR-183/96/182 cluster in patients with inherited and age-related retinal degenerative diseases. This research will lead to molecular diagnosis of retinal degenerative diseases, and finding new therapies for these retinal diseases. In addition, we are studying miRNAs in the retina of patients with age-related macular degeneration (AMD). This research will lead to identification of miRNAs involved in AMD and new therapeutic targets for its treatment. 2) microRNAs in diabetic retinopathy – diagnosis and treatment: Diabetic retinopathy (DR) is one of the leading causes of blindness. Previously, we identified a series of key miRNA signatures reflecting pathological changes of early DR (Kovacs et al. IOVS. 2011). Currently, we focus on miR-146 and several other miRNAs to dissect their involvement in DR and test their potential as therapeutic targets for treatment of DR. In addition, we are developing miRNA biomarkers in the vitreous, aqueous and plasma as diagnostic biomarkers for DR and other retinal diseases. 3) microRNAs in neuroimmune interaction and bacterial keratitis: Pseudomonas aeruginosa (PA) is a gram-negative opportunistic pathogen capable of inducing keratitis, affecting 25,000-300,000 contact lens users annually with a cost of treatment estimated between $15 to $30 million in the USA. Up to now, bacterial keratitis is mainly treated by topical administration of antibiotics; however, frequent emergence of antibiotic resistant bacteria poses serious challenges for its effective management. Recently, collaborating with Dr. Linda D Hazlett, we showed that inactivation of miR-183/96/182 causes decreased PA-induced corneal inflammation and keratitis in mice. Currently, we are dissecting molecular mechanisms of miR-183/96/182 in bacterial keratitis and develop miRNA-based new therapy for treatment of keratitis. 4) microRNAs in glaucoma: Previously we identified miRNAs whose expression levels changed in the retina of glaucoma mice compared to normal control mice. Currently, we are studying their roles in pathogenesis of glaucoma. This research may lead to identification of new miRNA-based therapeutic targets to develop new treatment for glaucoma.

Jena J. Steinle, PhD

Professor of Anatomy and Cell Biology and Ophthalmology

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Jena J. Steinle, PhD

Professor of Anatomy and Cell Biology and Ophthalmology

Area/Subject of Research: Retinal Vascular Biology

Publications: Recent of a total of 65 publications
- 1) Thakran S, Zhang Q, Morales-Tirado VM, Steinle JJ. Pioglitazone restores IGFBP-3 levels through DNA PK in retinal endothelial cells cultured in hyperglycemic conditions. Invest Ophthalmol Vis Sci. 2014 Dec 18. pii: IOVS-14-15550. PMID: 25525174 2) Jiang, Y, Zhang, Q, Steinle, JJ. Etanercept restores normal insulin signal transduction in β2-adrenergic receptor knockout mice. Journal of Neuroinflammation. PMID: 25138272 3) Zhang, Q, Steinle, JJ. 2014. IGFBP-3 inhibits TNFα production and TNFR-2 signaling to protect against retinal endothelial cell apoptosis. Microvasc. Res. 95C:76-81. PMID: 25086184 4) Jiang, Y, Thakran, S, Bheemreddy, R, Ye, EA, He, H, Walker, RJ, Steinle, JJ. 2014. Pioglitazone normalizes insulin signaling in the diabetic rat retina through reduction in tumor necrosis factor α and suppressor of cytokine signaling 3. J. Biol. Chem. 289(38):26395-405. PMID: 25086044 5) Jiang, Y, Pagadala, J, Miller, DD, Steinle, JJ. 2014. Insulin-like growth factor-1 binding protein 3 (IGFBP-3) promotes recovery from trauma-induced expression of inflammatory and apoptotic factors in the retina. Cytokine. PMID: 25082650 6) Zhang, Q, Soderland, D, Steinle, JJ. 2014. TNFα inhibits IGFBP-3 through Activation of p38α and Casein Kinase 2 in Human Retinal Endothelial Cells. PLoS One. 9(7):e103578. PMID: 25073020 7) Jiang, Y, Zhang, Q, Ye, EA, Steinle, JJ. 2014. β1-adrenergic receptor stimulation by agonist Compound 49b restores insulin signal transduction in vivo. Mol Vis. 20:872-80. PMID: 24966659 8) Jiang, Y, Zhang, Q, and Steinle, JJ. 2014. Intravitreal injection of IGFBP-3 restores normal insulin signaling in diabetic rat retina. PLoS One. 9(4):e93788. PMID: 24695399 9) Zhang, Q*, Jiang, Y, Toutounchian, J, Wilson, MW, Morales-Tirado, V, Miller, DD, Yates, CR, Steinle, JJ. 2013. Novel Quinic Acid Derivative KZ-41 Prevents Retinal Endothelial Cell Apoptosis Without Inhibiting Retinoblastoma Cell Death Through p38 Signaling. Invest. Ophthalmol. Vis. Sci. 54(9):5937-43. PMID: 23942968 10) Zhang, Q*, Soderland, C, Steinle, JJ. Regulation of retinal endothelial cell apoptosis through activation of the IGFBP-3 receptor. 2013. Apoptosis. Mar;18(3):361-8. PMID: 23291901
Summary of your research:
- We have a number of projects in the laboratory. 1) We are developing the mechanism of action of a novel beta-adrenergic receptor agonist, Compound 49b, that prevents diabetic retinopathy in a rat model. Compound 49b is anti-apoptotic and anti-inflammatory. We are working on the cellular signaling pathways involved in these actions. 2) We have a project on insulin resistance in the retina. We are working on pathways by which TNFα elicits insulin resistance in retinal cells (endothelial cells and Müller cells). We are also testing drugs (both novel and commercially available) to block these actions Current Projects: 1) R01 from NEI: Compound 49b Prevents Retinal Endothelial Cell Death Through IGFBP-3 Levels Diabetic retinopathy remains the fifth leading cause of preventable blindness worldwide. Interventions to prevent progression of diabetic retinopathy are limited to improved glycemic control (a challenging goal for all diabetic patients) and to laser photocoagulation (available only for advanced stages of retinopathy). We and others have reported that adrenergic signaling is lost in the diabetic retina, suggesting that development of novel agents to restore autonomic homeostasis is necessary. Unfortunately, currently available adrenergic agents are associated with adverse systemic or non-specific effects. These problems inspired our group to synthesize compound 49b, a novel and selective β-adrenergic receptor agonist, as a potential paradigm shift in the prevention of diabetic retinopathy. Our preliminary data suggest that compound 49b prevents the formation of degenerate capillaries, which involves degenerate capillary formation, which are the hallmark pathology noted in the diabetic retinal vasculature. In addition to preventing degenerate capillaries in vivo, compound 49b prevents the cleavage of caspase 3, a well-established marker of apoptosis, in retinal endothelial cells (REC) in vitro, suggesting that Compound 49b can decrease apoptosis. In the oxygen-induced model of retinopathy, others have associated increased levels of insulin-like growth factor binding protein-3 (IGFBP-3) with protection from REC apoptosis. Furthermore, using the streptozotocin-induced diabetic rat model, we observed that chronic insulin deficiency reduced IGFBP-3 protein levels in whole retinal lysates, but topical application of compound 49b to the eye restored retinal IGFBP-3 to its control level in these insulin-deficient rats. Thus, we hypothesize that compound 49b prevents the critical vascular damage underlying diabetic retinopathy in part by restoring IGFBP-3 levels in retinal endothelial cells. This project focuses on a deeper understanding of the mechanisms underlying this protective action. 2) R01 from NEI Mechanisms of TNFalpha-Induced Insulin Resistance in Retinal Cells The proposed study will test the novel hypothesis that in the diabetic retina, hyperglycemia stimulates production of tumor necrosis factor α (TNFα), which in turn decreases insulin receptor binding leading to decreased signal transduction. The overall effect of this signaling cascade would be to create insulin resistance, exacerbate problems caused by limited insulin production in diabetes, and thus contribute to development of diabetic retinopathy seen in both type 1 and type 2 diabetes. While our preliminary data and previous reports by others support a major role for inflammatory mediators such as TNFα in diabetic retinopathy, the pathways involved are largely unknown. Our proposed studies will focus on one likely candidate, the suppressor of cytokine signaling 3 (SOCS3) pathway (Fig.1), which is poorly understood in retina and yet represents a promising therapeutic target in future treatments for diabetic retinopathy. Our overall goal is to 1) establish the role of the SOCS3 pathway in regulating insulin signaling (through insulin receptor substrate-1; IRS-1) and apoptosis in normal and diabetic rats and 2) evaluate effects of upstream drug targets on the SOCS3 pathway and their downstream effects on insulin signaling and retinal cell apoptosis

Susmit Suvas, PhD

Associate Professor in Ophthalmology

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Susmit Suvas, PhD

Associate Professor in Ophthalmology

Published Articles:
- • Gaddipati S, Estrada K, Rao P, Jerome A and Suvas S (2015). IL-2/anti-IL-2 complex treatment inhibits the development but not the progression of herpetic stromal keratitis. J Immunol. 194 (1): 273-82. (PMID: 25411200) • Alazabi FA, Zohdy, MA and Suvas S (2013). Parameter estimation from experimental laboratory data of HSV-1 by using alternative regression method. Systems and Synthetic Biology. 7(14): 151-60. (PMID: 24432152) • Channappanavar R, Twardy BS and Suvas S (2012). Blocking of PDL-1 interaction enhances primary and secondary CD8 T cell response to herpes simplex virus-1 infection. PLoS One. 7(7): e39757. (PMID: 22808056) • Twardy BS, Channappanavar R, Suvas S (2011). Substance P in the corneal stroma regulates the severity of herpetic stromal keratitis lesions. Invest Ophthalmol Vis Sci. 4; 52(12): 8604-13. (PMID: 21969295) • Channappanavar R, Twardy BS, Krishna P and Suvas S (2009). Advancing age leads to predominance of inhibitory receptor expressing CD4 T cells. Mech Ageing Dev 130 (10): 709-12. (PMID: 19715717) • Susmit Suvas (2008). Advancing age and immune cell dysfunction: is it reversible or not? Expert Opinion on Biological Therapy 8 (5): 657-68. (PMID: 18407768)
Summary of Research:
- The major focus of the laboratory is on the development of a novel optogenetic strategy for restoring sight to people who suffer from retinal degenerative diseases, such as retinitis pigmentosa and age-related macular degeneration. The optogenetic strategy for restoring vision utilizes genetically encoded light sensors to convert surviving retinal neurons into photosensitive cells, thus imparting light sensitivity to the retina after the death of photoreceptor cells. Our earlier proof-of-concept studies demonstrated the feasibility of this strategy by AAV-mediated expression of channelrhodopsin-2 (ChR2) in inner retinal neurons of photoreceptor-deficient mice. We also demonstrated the restoration of both ON and OFF light responses by expression of ChR2 and halorhodopsin in the retina. More recently, we also made important progress on the development of more light sensitive ChR2 as well as AAV vector constructs for selective targeting of the light sensors to specific retinal neurons and/or subcellular compartments. Our long-term goal is aimed at developing effective optogenetic therapies for treating blindness caused by retinal degenerative diseases. Another area of interest of the laboratory is the investigation of retinal visual information processing, especially the roles voltage-dependent membrane channels in retinal bipolar cell processing. This will provide new insight into our understanding of visual information processing in the retina and help to develop new treatment approaches for blinding retinal diseases. The experimental techniques used in our studies include molecular biology, AAV vectors, electrophysiological recordings, immunocytochemistry, and animal behavior.
Current Research:
- The laboratory is currently looking at the role of neuropeptides in corneal HSV-1 infection and inflammation in mouse model.

Lalit Singh Pukhrambam, Ph.D.

Assistant Professor of Anatomy/Cell Biology and Ophthalmology

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Lalit Singh Pukhrambam, Ph.D.

Assistant Professor of Anatomy/Cell Biology and Ophthalmology

Published Articles:
- 1. Perrone L, Matrone C, Singh L.P. (2014) Epigenetic modifications and potential new treatment targets in diabetic retinopathy. J Ophthalmol. 2014:789120. PMID: 25165577 2. Singh L.P. (2014) The NLRP3 Inflammasome and Diabetic Cardiomyopathy: Editorial to: “Rosuvastatin alleviates diabetic cardiomyopathy by inhibiting NLRP3 inflammasome and MAPK pathways in a type 2 diabetes rat model” by Beibei Luo et al. Cardiovasc Drugs Ther. 28(1):5-6. PMID: 24220800 3. Singh L.P. (2013) Thioredoxin Interacting Protein (TXNIP) and Progression of Diabetic Retinopathy. J Clin Exp Ophthalmol, 4: 287. PMID: 24353900 4. Devi TS, Hosoya KI, Terasaki T, Singh L.P. (2013) Critical role of TXNIP in oxidative stress, DNA damage, and apoptosis of pericytes in high glucose: Implications for diabetic retinopathy. Exp. Cell Res. 319:1001–1012. PMID: 23353834 5. Singh, L.P., Devi, TS, Nantwi, KD. (2012) Theophylline Regulates Inflammatory and Neurotrophic factor Signals in Functional Recovery after C2-Hemisection in Adult Rats. Exp. Neurol., 238:79-88. PMID: 22981449 6. Devi TS, Lee I, Hüttemann M, Kumar A, Nantwi KC, Singh L.P. (2012) Txnip links innate host defense mechanisms to oxidative stress and inflammation in retinal Muller glia under chronic hyperglycemia: Implications for diabetic retinopathy. Exp. Diab Res., 2012;2012:438238. PMID: 22474421 7. Devi TS, Hosoya K, Terasaki T, and Singh LP. (2011) GSK-3-CREB axis mediates IGF-1-induced ECM/adhesion molecule expression, cell cycle progression and monolayer permeability in retinal capillary endothelial cells: Implication for Diabetic Retinopathy. Biochim Biophys Acta-Molecular basis of disease 1812(9):1080-1088. PMID: 21549192 8. Sbai O, Devi TS, Melone MA, Feron F, Khrestchatisky M, Singh L.P., and Perrone L. (2010). RAGE-TXNIP axis is required for S100B-promoted Schwann cell migration, fibronectin expression and cytokine secretion. J Cell Science 123(Pt 24):4332-4332 PMID: 21098642 9. Perrone L, Devi TS, Hosoya K, Terasaki T, and Singh LP. (2010). Inhibition of TXNIP Expression In Vivo Blocks Early Pathologies of Diabetic Retinopathy. Cell Death and Disease 1, e65; doi:10.1038/cddis.2010.42. PMID: 21364670 10. Perrone L, Devi TS, Hosoya K, Terasaki T, and Singh LP. (2009). Thioredoxin-Interacting Protein (TXNIP) induces inflammation through chromatin modification in retinal capillary endothelial cells under diabetic conditions. J Cell Physiol. 221(1):262-272. PMID: 19562690
Summary of Research:
- Our laboratory is interested in developing mechanism-based and stage-specific therapies to treat diabetic retinopathy using peptide and RNAi technologies. We use both in vivo animal models of type-1 diabetes and in vitro retinal cell cultures to understand the molecular basis of early abnormalities in neurons and vasculature, especially mediated by mitochondrial (MT) dysfunction, oxidative stress and inflammation. We investigate the critical role played by TXNIP in redox imbalance, mitochondrial protein unfolding and MT-UPR and mitophagy in neurovascular degeneration and the progression of diabetic retinopathy. We use biochemical, molecular biology, proteomics, epigenetics, and in vivo non-invasive methods to measure diabetes duration-dependent neuronal injury, microvascular dysfunction and BRB breakdown.
Current Research:
- Mitochondrial dysregulation and resultant energy imbalance is associated with various chronic diseases including neuro-degeneration, ischemia/reperfusion, and diabetic complications including diabetic retinopathy. Recently, we published that pro-oxidant thioredoxin interacting protein (TXNIP) is significantly up-regulated in diabetic retinas and under hyperglycemia in retinal cells in culture such as endothelial cells, pericytes and Muller cells (MC) and mediates cellular oxidative stress, inflammation and apoptosis. TXNIP has recently been implicated by several studies as a critical protein in the pathogenesis of diabetes and its complications. TXNIP binds to thioredoxin (Trx), a redox anit-oxidant protein, inhibiting its reactive oxygen species (ROS) scavenging and thiol reducing capacity; therefore, results in cellular oxidative/nitrosative (ROS/RNS) stress and aberrant protein s-nitrosylation. Furthermore, MC are important for retinal neuronal health and activated MC (gliosis) induces aberrant gene expression for cytokines and growth factors to maintain retinal homeostasis. However, prolonged MC activation is injurious in DR. Therefore, our overall hypothesis is that TXNIP is critical for MT dysfunction, neurovascular injury, and MC activation in the development of DR. We are investigating the role of TXNIP in (i) MT dysfunction and retrograde stress responses (MT-UPR); and (ii) MT fission/fusion and mitophagy as a part of mitochondrial quality control in diabetic retinopathy. Is there a relationship between mitophagy deregulation and NLRP3 inflammasome activation and innate immune responses in diabetic retinopathy and, if any, to what extent TXNIP plays a role in this process are some of the questions being asked? To address these objectives, we will use streptozotocin (STZ)-induced type-1 diabetic models of rat and mouse in conjunction with manipulation of TXNIP expression levels (siRNA and knock-out mice) in the retina. In vitro studies using retinal MC, endothelial cells and neuronal cell cultures will be performed to dissect the molecular mechanisms as to how TXNIP induces MT dysfunction and mitophagy, which specifically removes damaged MT. Our studies may allow us to identify potential new targets for developing gene/drug therapies to prevent or slow down the progression of DR. Currently; our research is supported by NIH/NEI grant, NIH/NEI Core grant to the Department of Anatomy and Cell Biology and Research to Prevent Blindness to the Department of Ophthalmology.

Fu-Shin X. Yu, Ph.D.

Professor of Ophthalmology, Anatomy and Cell Biology

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Fu-Shin X. Yu, Ph.D.

Professor of Ophthalmology, Anatomy and Cell Biology

Research Subject: Corneal wound healing and innate immunity

Published Articles:
- • Gao N, Kumar A, and Yu FS. MMP13 as a target for suppressing corneal ulceration caused by Pseudomonas aeruginosa infection. J. Infect Diseases. In press. • Liu X, Gao N, Dong C, Zhou L, Mi QS, Standiford TJ, et al. Flagellin-induced expression of CXCL10 mediates direct fungal killing and recruitment of NK cells to the cornea in response to Candida albicans infection. Eur J Immunol. 2014;44(9):2667-79. Epub 2014/06/27. doi: 10.1002/eji.201444490. PubMed PMID: 24965580; PubMed Central PMCID: PMC4165733. • Bettahi I, Sun H, Gao N, Wang F, Mi X, Chen W, et al. Genome-wide transcriptional analysis of differentially expressed genes in diabetic, healing corneal epithelial cells: hyperglycemia-suppressed TGFbeta3 expression contributes to the delay of epithelial wound healing in diabetic corneas. Diabetes. 2014;63(2):715-27. Epub 2013/12/07. doi: 10.2337/db13-1260. PubMed PMID: 24306208; PubMed Central PMCID: PMC3900551. • Yoon GS, Dong C, Gao N, Kumar A, Standiford TJ, Yu FS. Interferon regulatory factor-1 in flagellin-induced reprogramming: potential protective role of CXCL10 in cornea innate defense against Pseudomonas aeruginosa infection. Invest Ophthalmol Vis Sci. 2013;54(12):7510-21. Epub 2013/10/17. doi: 10.1167/iovs.13-12453. PubMed PMID: 24130180; PubMed Central PMCID: PMC3832217. • Gao N, Sang Yoon G, Liu X, Mi X, Chen W, Standiford TJ, et al. Genome-wide transcriptional analysis of differentially expressed genes in flagellin-pretreated mouse corneal epithelial cells in response to Pseudomonas aeruginosa: involvement of S100A8/A9. Mucosal Immunol. 2013;6(5):993-1005. Epub 2013/01/24. doi: 10.1038/mi.2012.137. PubMed PMID: 23340821; PubMed Central PMCID: PMC3722258. • Wang F, Gao N, Yin J, Yu FS. Reduced innervation and delayed re-innervation after epithelial wounding in type 2 diabetic Goto-Kakizaki rats. Am J Pathol. 2012;181(6):2058-66. Epub 2012/10/16. doi: 10.1016/j.ajpath.2012.08.029. PubMed PMID: 23063510; PubMed Central PMCID: PMC3509759. • Gao N, Yin J, Yoon GS, Mi QS, Yu FS. Dendritic cell-epithelium interplay is a determinant factor for corneal epithelial wound repair. Am J Pathol. 2011;179(5):2243-53. Epub 2011/09/20. doi: 10.1016/j.ajpath.2011.07.050. PubMed PMID: 21924232; PubMed Central PMCID: PMC3204011. • Yin J, Huang J, Chen C, Gao N, Wang F, Yu FS. Corneal complications in streptozocin-induced type I diabetic rats. Invest Ophthalmol Vis Sci. 2011;52(9):6589-96. Epub 2011/07/01. doi: 10.1167/iovs.11-7709. PubMed PMID: 21715347; PubMed Central PMCID: PMC3176029. • Xu K, Yu FS. Impaired epithelial wound healing and EGFR signaling pathways in the corneas of diabetic rats. Invest Ophthalmol Vis Sci. 2011;52(6):3301-8. Epub 2011/02/19. doi: 10.1167/iovs.10-5670. PubMed PMID: 21330660; PubMed Central PMCID: PMC3109029. • Gao N, Kumar A, Guo H, Wu X, Wheater M, Yu FS. Topical flagellin-mediated innate defense against Candida albicans keratitis. Invest Ophthalmol Vis Sci. 2011;52(6):3074-82. Epub 2011/02/12. doi: 10.1167/iovs.10-5928. PubMed PMID: 21310913; PubMed Central PMCID: PMC3109016.
Summary of your research:
- The Yu’s laboratory investigates the molecular mechanisms underlying corneal epithelial wound healing and diabetic neurotrophic keratopathy, using corneal organ culture and animal models. Specifically, the lab studies the interactions of sensory nerve, intraepithelial dendritic cells, and epithelial cells in the cornea during wound healing and their defects in diabetic corneas which lead to delayed epithelial wound closure and impaired sensory nerve regeneration. The second project of Dr. Yu’s lab is to study ocular innate immunity and protection against infection. Dr. Yu and colleagues have shown that activation of Toll-like receptors, particularly TLR5 prior to infection resulted in profound mucosal protection against a broad spectrum of keratitis causing pathogens. The focus in Dr. Yu’s lab is to define the molecular mechanisms underlying these protective pathways.
Current Research:
- Molecular regulation of corneal wound healing Mechanisms of flagellin induced protection against infection keratitis