Ganciclovir Prophylaxis Modifies Long-Term Murine Cytomegalovirus Induced Renal Allograft Injury

Abstract

Alabama at Birmingham, Genetics, Birmingham, United States Background: Human cytomegalovirus (CMV) infection is associated with adverse outcomes in renal transplantation, but the pathogenesis remains cryptic. Ganciclovir prophylaxis is associated with improved survival in some but not all clinical studies. In a murine CMV (MCMV) donor positive/recipient negative allogeneic renal transplant model, MCMV exacerbated innate infiltrates and early histologic injury at 2 weeks post-transplant, and these parameters improved with ganciclovir prophylaxis. The effect of ganciclovir prophylaxis on late allograft histology has not been examined. Methods: Murine CMV (MCMV) infected Balb/c donor kidneys were transplanted orthotopically into C57Bl/6 recipients with cyclosporine immunosuppression (CMV group). Control transplants were performed using uninfected Balb/c donors (control group). Ganciclovir (GCV) prophylaxis was administered for 14 days post-transplant to one cohort of MCMV infected transplants (GCV group). Animals were sacrificed at day 42 post-transplant. Histology was scored according to an established grading scale (grade 0-3 for 9 criteria, maximum score 27) by a veterinary pathologist blinded to sample identity. Immune infiltrates were examined by flow cytometry in allograft kidneys, livers, and spleens. Results: At day 42, CMV transplants had higher aggregate damage scores (average 14.5) compared to control uninfected transplants (average 10.25). Allografts with GCV prophylaxis had lower damage scores (average 9.5) and appeared histologically more similar to uninfected grafts than to CMV infected grafts. Analysis of leukocyte infiltrates by flow cytometry revealed greater frequencies of CD45+ cells in CMV infected compared to uninfected grafts. Of the CD45+ population, CMV infected and uninfected grafts had similar frequencies of CD4+, CD8+, and CD19+ lymphocytes, but CMV infected grafts contained higher frequencies of CD49b+ and Gr-1+ cells compared to uninfected grafts. GCV prophylaxis reduced the frequency of Gr-1+ infiltrates but not CD49b+ infiltrates, and also contained a lower frequency of CD11b+/CD204+ cells. Grafts with GCV prophylaxis contained greater frequencies of CD4+ cells compared to both CMV and control grafts. Analysis of infiltrates in livers and spleens from allograft recipients showed no differences in frequencies of any of these leukocyte subsets between control, CMV, and GCV treated recipients. Conclusion: CMV infection exacerbates late renal allograft damage by histologic criteria and is associated with ongoing infiltration of innate leukocyte subsets including NK cells and myeloid cells. Allografts with GCV prophylaxis had less severe allograft injury and appeared histologically similar to uninfected grafts, even though prophylaxis had been discontinued for 28 days prior to terminal sacrifice. GCV prophylaxis was associated with reduction of myeloid and macrophage infiltrates but not NK infiltrates, and additionally was associated with increased CD4+ infiltrates. These results suggest that ganciclovir prophylaxis may modulate long-term CMV effects upon allograft injury, which may possibly be related to differences in recruitment of innate and CD4+ leukocytes even after cessation of prophylaxis. Intragraft responses differed from leukocyte populations in the liver and spleen, suggesting that systemic responses may not necessarily serve as surrogates for the intragraft inflammatory response.