Experimental study on inhibiting graft rejection following high-risk cornea transplatation by resolvinE1 in mice

Abstract

Background The conventional drugs for preventing and treating graft rejection have the risks of inducing adverse responses.Researches showed that resolvinE1 (RvE1) can regulate Th1 cell-mediated immunoreaction.However, whether RvE1 has an inhibit effect on high-risk corneal graft is unclear now. Objective This study was to investigate the effects of RvE1 on immune rejection in high-risk corneal grafting mouse models. Methods SPF BALB/c mice were used as recipients, C57BL/6 mice were as donors.Ninety BALB/c mice were divided into corneal allograft group, corneal allograft+ RvE1 group and corneal autograft group according to random number table.High-risk corneal graft models were established by corneal suturing for 14 days and followed by penetrating keratoplasty in recipients.Allograft keratoplasty was performed on the right eyes in the mice of corneal allograft group and corneal allograft+ RvE1 group, and self-corneal graft rotated 180°was transplanted on the right eyes in the mice of autograft group.Normal saline solution of 10 μl was subconjunctivally injected after surgery once per day for 7 days in the corneal allograft group and corneal autograft group, and 10 μl RvE1 (1 μg) was used in the same way in the corneal allograft+ RvE1 group.The recipient eyes were examined for potential rejection signals with slit lamp microscope and calculated the mean survival time and rejection index (RI). The histopathology was examined 21 days after modeling by hemotoxylin and eosin staining.The expressions of CD4 and interferon-γ (IFN-γ) in the corneas were detected by immunohistochemistry.Th1 cell (CD3+ CD8a-IFN-γ+ ) percentage in draining lymph nodes were measured by flow cytometry.The mRNA expression levels of interleukin-2 (IL-2), tumor mecrosis factor-α (TNF-α), IFN-γ and T-bet were detected by real-time fluorescence quantitative PCR. Results The mean survival time of grafts was (28.5±1.7) days in the corneal allograft group, and that in the corneal allograft+ RvE1 group was (14.0±1.6) days, showing a significant difference between them (t=4.14, P<0.001), while the survival rate was 100% at 50 days after modeling in the corneal autograft group.Corneal edema and inflammatory cell infiltration were slight in the corneal allograft+ RvE1 group and corneal autograft group compared with corneal allograft group.CD4 was positively expressed in corneal tissue, and IFN-γ was expressed in corneal epithelium.The CD4+ and IFN-γ+ cell number was decreased in the corneal allograft+ RvE1 group and corneal autograft group compared with corneal allograft group under the fluorescence microscope.The percentages of Th1 cells in lymph cells of corneal allograft + RvE1 group and corneal autograft group were (1.07±0.25)%, (0.85±0.12)%, respectively, which were significnatly lower than (1.56±0.20)% in the corneal allograft group (both at P<0.05). The expressions of IL-2, TNF-α, IFN-γ and T-bet mRNA in the corneal tissue in the corneal allograft group were higher than those in the corneal allograft+ RvE1 group and corneal autograft group (all at P<0.05). Conclusions RvE1 inhibits graft rejection in high-risk allograft mouse models probably by down-regulating the Th1 cell percentage in lymph cells and the expression of inflammation-related cytokines in corneal grafts. Key words: Cornea/immunology; Corneal transplantation/immunology; Graft rejection; Eicosapentaenoic acid/analogs & derivatives; ResolvinE1; T helper cell 1; Cytokines; Disease models, animal; Mice, inbred BALB/c; Mice, inbred C57BL/6