Based on previous research on non-infectious uveitis, we made progress in the following areas:? ? 1)In collaboration with NCI, we established a murine-humman B cell intraocular lymphoma model and showed that it could be used for therapeutic testing and for furter understanding molecular mechanisms of Primary Intraocular Lymphoma (PIOL). We showed that an immunotoxin that is specific for B cells (HA22) can eradicate lymphoma cells in this model, proposing that this immunotoxin can be used clinically for treating PIOL patients. This report has been accepted by Cancer Research and selected as Featured Article. Continued work is required to furhter translate this novel finding into clinical trials. ? 2) Understanding the molecular mechanisms of Daclizumab in treating autoimmune originated uveitis. Daclizumab is known to block the alpha-chain (CD25) of the IL-2 receptor which is required for T cells to sense signals from T cell growth factor, IL-2. One of the arguments that was thought that might be against the application of Daclizumab was that IL-2 signaling was critical for activation induced cell death (AICD). Blockage of AICD could result in decreased apoptosis and increased lymphoproliferation. But clinical data proved that Daclizumab indeed ameliorated autoimmune uveitis and was an excellent alternative therapy. One of the hypotheses would be that in vivo blockage of CD25 in humans differ from inactivation of CD25 gene in mice. Multiple mechanisms could be contributing to Daclizumab effects besides its role in blocking IL-2 receptor. We continuousely monitored a group of patients who underwent Daclizumab therapy. We found that a sub-group of NK cells, called CD56bright, were induced in vivo after Daclizumab infusion. We showed that these cells secreted large amount of IL-10 and could be regulatory NK cells. We proposed that these induced CD56bright NK cells under Daclizumab therapy may be beneficial to the clinical outcome of uveitis patients. These findings have been published in the J. Immunology. Further studies have been designed to understand the molecular mechanisms of the induced CD56bright NK cells, e.g., are they beneficial to the patients? how do they work in terms of in the process of inflammation? ? 3) Further understanding of the triggering events for non-infectious uveitis. Despite intensive studies on animal models and using in vitro culturing system, the molecular mechanisms of non-infectious uveitis in humans are still an open question. we continue our interest in the search for antigens associated with uveitic conditions. Tropomyocin has been recently reported to cause uveitis in rats. We tested the uveitogenic tropomycin peptides. The recall responses (proliferation, cytokine profiles) of immune cells from Behcets disease patients and from normal donors to tropomycin peptides, as well as to established human retinal S-antigen showed that more than 25% of uvietis patients showed T cell response to tropomyocin. We have also demonstrated anti-tropomyocin antibodies in the serum of uvieits patients. We have proposed that tropomyocin is one self-antigen for a sub-population of Bechets patients. These findings have been published in Clin. Exp. Immunology.? 4) Intensive efforts have also been made to understand the implication of GITRL expression in autoimmune uveitis patients as well as in other autoimmune diseases by studying the effects of the interaction between GITR and GITR ligand in vitro. We showed that the GITRL expression on retinal cells are induced by inflammatory cytokines. Overexpression of GITRL reversed physiological immune privilege of ocular RPE cells. We proposed that the expression of GITRL in ocular tissues is a natural way of balance between ocular immune privilege and inflammation responses. These studies resulted in 2 publications in Eur. J. immunology and IOVS and one manuscript in preparation. ? 5) In collaboration with Dr. Egwuagu and colleagues in LI/NEI, we also published our recent observation that SOCS5, a critical immunosuppressive protein in the cellular signaling events may be used as a clinical marker for active uveitis and further used as prognostic marker for the clinical therapy. These results have been published in J. Autoimmunity and may have implications for linking cellular signaling events with clinical activities of uveitis patients. ? ? ? Future emphasis:? A) Clinically, in additon to autoimmune uveitis, we will focus on understanding the potential roles of immune mechanisms in the molecular pathogeneisis of Age-related Macular Degeneration (AMD) and Diabetic retinopathy. Recent findings of the role of macrophages (especially the type 1 and type 2 macrophages) in AMD further support our intention to pursue in this direction. We have designed several flowcytometry protocols for screening AMD and diabetic patients. Analysis on flow cytometry phenotyping data has been ongoing. We have also adopted mouse laser induced CNV model. Preliminary data from this model have helped us understand kinetic and dynamic changes of infiltration of immune cells during laser induced CNV. Further efforst are focused on understinding the molecular mechanisms. ? B) Mechanistic effects of daclizumab (humanized anti-CD25, anti-Tac, HAT) treatment on uveitis patients: Daclizumab, or humanized anti-CD25, or anti-Tac, or HAT, has been successful in treating non-infectious intermediate and posterior uveitis. Understanding the mechanisms of this therapy at the molecular level can facilitate and improve clinical treatment of autoimmune uveitis. We believe that these findings will have implications for other autoimmune disorders. Indeed, we have discovered a unqiue sub-population of cells from innate immune system that is induced by Daclizumab and may be beneficial to patients. Continued efforts on understanding the molecular mechanisms of those cells are our current focus. ? C) Streamline molecular and cellular methodology and maximize windows that help reveal potential mechanisms for the diseases: The advantage of studying patient samples is its direct clinical relevance and potential impact on clinical diagnosis and therapy. But the challenges of clinical studies are the limitation of sample sources and quantity, inconsistency of patients compliance, often difficulty to repeat the results, as well as influences of constantly changing environments that may alter the outcome of studies. One of the ways to minimize those factors is to streamline the methodologies that are used to examine patients specimen so that multiple measurements will be performed simultaneously for a single specimen and mutually supportive data will be generated . We have made progress instandardizing methodology for analysis. We wish to continue to focus on this aspect of studies. ? D) Continuouslly collaborating with intramural as well as extramural scietists and industries. Our previous experience in collaborating with intramural scientists, e.g., LI and other labs in NEI, with NCI as well as with industries, e.g., Neurotech, Inc proved to be critical for our productivity. We wish to continue on this track and expand our collaborations with all who share interst with us in our ultimate goals, e.g., to further understand the molecular mechanisms and seek for efficient therapies for ocular inflammatory diseases.
