Сytometric evaluation of immune cell compartments in patients after penetrating keratoplasty

The corneal graft rejection, especially in patients from the “high risk” group, is a complex immunological process that remains an urgent problem in ophthalmology. The cornea, despite its immunological privilege, loses its protective barriers under certain conditions (vascularization, inflammation), which leads to the development of immune reaction against the transplant. The key mechanisms of rejection include activation of T lymphocytes (CD4⁺ and CD8⁺), as well as an imbalance between effector and regulatory immune cells. Modern methods of immunological monitoring, including analysis of T cell populations, make it possible to detect signs of rejection in a timely manner and adjust therapy to improve long-term transplant results. The aim of our study was to conduct a comparative cytometric analysis of the population and subpopulation composition of blood lymphocytes in patients after end-to-end (penetrating) keratoplasty. The study involved 46 patients who underwent penetrating keratoplasty, being divided into two subgroups: those with transplant rejection (n = 25) and those with successful engraftment (n = 21), as well as a control group of 21 healthy volunteers. The immunological analysis included venous blood sampling and quantitative determination of lymphocyte subpopulations by flow cytometry using a Navios cytofluorimeter (Beckman Coulter, USA) and markers of CD45⁺ and CD46⁺ cell populations. Key populations of lymphocytes, including T helper cells, cytotoxic T lymphocytes, NK cells, B lymphocytes, and activated T cells, were studied to identify differences in the immune status of patients. Comparative cytometric analysis of the population and subpopulation composition of peripheral blood lymphocytes in “high-risk” patients with graft rejection showed a number of significant changes manifesting as an increased total number of T lymphocytes, T helper cells, T cytotoxic, T lymphocytes with markers of early and late activation, thus suggesting the key role of T cell-mediated immunity in development of late cellular rejection reactions. The data obtained suggest involvement of systemic cellular mechanisms into rejection of the native corneal allograft, presuming a need for a more detailed study of the T cell response of immune system in this condition. Cytometric assessment of subpopulations of immunocytes makes it possible to identify early signs of immune activation associated with rejection and opens up new opportunities for the development of personalized and effective treatment strategies aimed at improving the transplant outcomes.