Studies of the Inflammatory/Immunogenic Mechanisms of Neurodegeneration in Glaucoma
A highlight of Dr. Tezel’s early career was the exciting finding of immune system involvement in glaucoma. Tezel Lab has pointed out for the first time and explored further the immunogenic aspects of glaucomatous neurodegeneration. Through the analyses of T-lymphocytes and autoantibodies isolated from patients’ blood and the studies of cell cultures, animal models, and postmortem human donor eyes, Tezel Lab has provided the initial evidence supporting a prominent activation and the neurodegenerative potential of adaptive immune responses in glaucoma. Over the years, it has become widely evident that the responses of resident glial cells to glaucoma-related stress and RGC damage create a critical damaging force on RGC somas, axons, and synapses in the long term. These innate immune responses can also promote an environment stimulating systemic immunity and facilitating tissue access and neurodegenerative outcomes of circulating immune cells, reactive T-lymphocytes, and autoantibodies to ocular antigens.
Although therapeutic modulation of glia-driven neuroinflammation is recognized as a logical strategy to avoid inflammatory neurotoxicity in glaucoma, effective strategies for immunomodulation remain elusive. The following research in Tezel Lab has focused on the molecular characterization of neurodegenerative inflammation. Accumulating research findings from in vitro and in vivo experimental models of glaucoma have demonstrated a prominent cytokine response in the retina and optic nerve, including the increased glial production of TNF-α, a major pro-inflammatory, pro-apoptotic, and neurodegenerative cytokine. Tezel Lab has continued investigating the molecular components and regulatory mechanisms of neuroinflammation signaling to develop immunomodulatory treatments for RGC protection.
It is a thrilling research field because the crucial roles of various glial responses in the vicious cycle of pathogenic processes put them in an important place to target for glaucoma treatment. When considering the widespread neurodegenerative outcomes of glia-driven neuroinflammation, from the retina to the brain, its modulation also offers a treatment strategy for widespread neuroprotection in glaucoma.
Studies of the Molecular Pathways of Glaucomatous Neurodegeneration and Potential Treatment Targets for Immunomodulation and Neuroprotection
Through the proteomic analysis of freshly isolated RGCs and glial cells from animal models, Dr. Tezel’s studies have been pioneering in defining the cell type-specific molecular responses in glaucoma, while newer techniques are now increasingly used to expand cell type-specific molecular information. Tezel Lab’s following research has utilized different experimental paradigms to test whether therapeutic restoration of specific molecular pathways, using pharmacological or transgenic approaches, suppresses neuroinflammation and protects RGCs against glaucomatous neurodegeneration. Besides testing therapeutic outcomes on glial responses and structural protection of RGC somas in the retina and axons in the optic nerve, these studies also examine treatment effects on RGCs’ electrophysiological function.
Degeneration of RGC axons at the optic nerve and apoptosis of RGC somas in the retina are pivotal features of neurodegeneration in glaucoma, controlled by unique molecular programs. Research outcomes of the studies characterizing molecular pathways have highlighted oxidative stress participating in glaucomatous neurodegeneration at different RGC compartments. Oxidative stress, mainly developed secondary to mitochondrial dysfunction, has been found to directly promote neuron injury and boost glial inflammatory responses through multiple mechanisms. Diverse roles of oxidative stress have been tested by antioxidant treatments that provided immunomodulation and neuroprotection in experimental glaucoma. Since mitochondrial dysfunction and glia-driven neuroinflammation are interdependent pathogenic processes, manipulating these converging routes may allow a unified treatment strategy for glaucomatous neurodegeneration. Investigating this stimulating aspect constitutes one of the current research goals in Tezel Lab.
By studying RGC and glial cell cultures and experimental animals, Tezel Lab has also demonstrated that the inflammatory death of RGCs in glaucoma, such as via TNF-α/TNFR1 signaling, involves a caspase-8-initiated apoptosis cascade. Interestingly, these studies using pharmacological or transgenic treatments to inhibit caspase-8 in glaucoma models detected cell type-specific functions of caspase-8. It appears that cFLIP functions as a molecular switch between caspase-8-mediated cell death/survival/inflammation pathways signaled through TNFRs, TLRs, and inflammasome, and the caspase-8/cFLIP interaction plays a key role in the regulation of different fates of RGCs and glia during glaucomatous neurodegeneration. In an ongoing project, Tezel Lab further explores this cell type-depending regulatory mechanism in mouse glaucoma models with cre/lox-based conditional deletion of glial cFLIP or cFLIPL to determine isoform-specific differences.
Studies of the NF-κB-Mediated Regulation of Glia-Driven Neuroinflammation
Bioinformatic analysis of the high-throughput proteomic datasets, along with the data from in vitro and in vivo experiments using specific treatments, has pointed to glial NF-κB as a common transcriptional activator of multiple inflammation pathways through TNFR or TLR signaling and inflammasome. Although NF-κB’s gene targets include pro-inflammatory cytokines involved in inflammatory responses and secondary injury processes, such as TNF-α, NF-κB also activates essential anti-apoptotic genes, increasing the survival of RGCs. Not to interfere with the neuronal survival program, a current project in Tezel Lab seeks the NF-κB-mediated regulation of neuroinflammation using a cell type-targeting approach. To test the importance of NF-κB for regulating glia-driven neuroinflammation, they analyze neuroinflammatory and neurodegenerative outcomes of experimental glaucoma in mice with or without cre/lox-based conditional deletion of specific genes (IκKβ or p65) in astroglia. Findings of this project so far have demonstrated a key role for astroglial NF-κB in experimental glaucoma, suggesting this molecule as a glial treatment target to provide neuroprotection by immunomodulation. This work paves the way for future clinical translation through glia-targeting transgenic strategies for glaucoma treatment. Targeted manipulation of specific molecules in glial cells is ideal for preventing or reversing the neurodegenerative inflammation in glaucoma while keeping the systemic immune defense and glial neurosupport functions intact, with no risk of undesired side effects on neurons.
An exciting extension of recent studies in Tezel Lab is based on the evidence supporting a potential role for NF-κB in cell-autonomous and non-cell-autonomous regulation of astroglia-microglia interactions in experimental glaucoma. An additional current project uses conditional knock-out and knock-in models to investigate the regulatory role of NF-κB in the astroglia-microglia partnership to amplify neuroinflammation in a stage-depending manner during glaucomatous neurodegeneration.
Studies of Glaucoma-Related Biomarkers and New Clinical Tools
Dr. Tezel’s clinically-oriented studies, besides postmortem studies of glaucomatous and non-glaucomatous human donors, have analyzed patients’ blood, aqueous humor, and imaging data to evaluate clinical correlations of the experimental observations in animal models. These studies, conducted in collaboration with glaucoma clinicians, have also aimed to identify glaucoma-related molecular biomarkers to help diagnose glaucoma early and monitor disease progression and treatment responses in the clinic.
Tezel Lab’s recent work through the mass spectrometry-based proteomic/immunoproteomic analysis of patients’ blood and aqueous humor samples has identified several molecules as glaucoma-related biomarker candidates to test in larger and prospective cohorts for biomarker validation. Concerning the multifactorial nature and individual variability of glaucoma, Tezel Lab anticipates and currently further searches for a panel of biomarker molecules (including specific molecules identified by mass spectrometry, oxidative stress-related biomarkers, autoantibody titers, cytokine profiles, and T-lymphocyte subset imbalances) that can collectively provide improved information for clinical predictions.
A recent imaging-based study has supported early localized alterations of the retinal inner plexiform layer in correlation with progressing visual field defects in glaucoma patients. This study providing clinical relevance of the early synaptic alterations in animal models to human glaucoma exemplifies a translational route from bench to bedside and back for improved understanding, treatment, and clinical testing of glaucoma.
Tezel Lab also collaborates with Silverman Lab in projects aiming to develop and test ultrasound-based super-resolution imaging for glaucoma diagnosis and ultrasound-activated perfluorocarbon nanodroplet applications for glaucoma treatment.