Effect of pathological ocular environment on ocular tissue

Oxidative stress is implicated in retinal ganglion cell (RGC) death in glaucoma. All the cells have elaborate defense mechanisms against oxidative stress. Our studies demonstrate that oxidative preconditioning can protect the RGCs from cytotoxic effects of lethal doses. This protective effect is probably mediated by NF-κB.

We also studied how ultraviolet light induced oxidative stress can cause degenerative changes by inducing pro-apoptotic protein Bax in retinal cells using retinal ganglion and retinal pigment epithelial cell cultures after various exposure time points.

In another study we looked at how the proliferative conditions of the retina, that are major causes of visual impairment, can produce factors that changes the intraocular environment in a way which is conducive for proliferation of other cells types, creating a vicious cycle. Our results demonstrate that proliferating retinotypic cells produce factors, including VEGF that can significantly alter the intraocular environment, such that it favors cell proliferation. These retinotypic cells have molecular machinery to produce and respond to VEGF. Breaking this cycle may provide an opportunity to modulate the course of proliferative vitreoretinal conditions.

Hypoxia is a pathological condition associated with a variety of ocular diseases including choroidal neovascularization. Our approach is to identify other factors other than VEGF which upregulates during neovascularization process. One of our in vitro research is underway to identify the different hypoxic factors and the associated transcriptional regulators and their expression in relation to choroidal neovascularization. This research will be helpful to identify the different epigenetic modulators involved in hypoxic conditions.

Effect of proton beam radiation on ocular tissue

Neovascularization (newly sprouting endothelial cell) is a common complication associated with wet form of age-related macular degeneration. We are trying to utilize the new development of proton beam radiation, which has minimal entry and exit dose, for the treatment of neovascularization. Though some earlier clinical studies used this approach to treat neovascularization, none of the studies established a specific dosage. In our basic in vitro research using choroidal endothelial cells, we found that newly proliferating choroidal endothelial cells are sensitive between 8-12 CGE dosages of proton radiation. We also found that radiation-induced oxidative stress is a major cause of radiation induced sensitivity in choroidal endothelial cells. On the other hand, we are trying to establish the safety dosage to the retinal cells.

Further studies are underway to study the mechanistic or signaling pathways and the associated active proteins which are triggered by radiation and leading to apoptosis, necrosis and cell death. These in vitro results will provide a better insight for further in vivo experiments to proceed with this proton beam therapy from bench top to clinics.

Microbead fluoroanalyzer is more competitive to ELISA

Cytokines are involved in the pathology of wide range of diseases. Measurements of levels of cytokines are useful for understanding pathogenesis and as diagnostic and prognostic indicators in many diseases. The available gold standard technique for these markers is enzyme-linked immune sorbent assays (ELISA) which require large volume of sample. Eye is one of our systems where we can acquire small volume of sample such as aqueous humor.

In our studies, we determined that using microbead technology, which binds with protein of interest and produces fluorescent signal and measured using a fluoroanalyzer is a useful technology in comparison to ELISA at a lower volume of sample and at a lower concentration.