Abstract: BACKGROUND: There is currently no effective treatment for kidney transplant failure caused by antibody-mediated rejection. Studies have shown that the expression of plasma miR-4286 in kidney transplant patients is significantly up-regulated, and miR-4286 can down-regulate the expression of transforming growth factor β to activate an immune response.
OBJECTIVE: To investigate the effect of miR-4286 silencing on up-regulating follicular regulatory T cells in the treatment of antibody-mediated rejection after kidney transplantation and its mechanism.
METHODS: Patients who underwent kidney transplantation at the Affiliated Hospital of Guizhou Medical University from December 2018 to December 2020 were enrolled and assigned into two groups (n=20 per group): antibody-mediated rejection group and immune tolerance group. Hematoxylin-eosin staining and C4d immunohistochemistry were performed for biopsy of kidney tissues. The mRNA expression of miR-4286 in peripheral blood lymphocytes and the proportions of follicular helper and follicular regulatory T cells were detected. The lymphocytes of patients suffering rejection reaction were divided into a blank vector group, a miR-4286 overexpression group, a miR-4286 silence group, and a transforming growth factor β blocking group in which the cells were simultaneously transfected with miR-4286-inhibited lentiviral vector and transforming growth factor β neutralizing antibody). After 48 hours, we detected the proportions of B cells, plasma cells, and follicular regulatory T cells, as well as the levels of transforming growth factor β, interferon γ, interleukin 6, and interleukin 21 in the cell supernatant.
RESULTS AND CONCLUSION: Hematoxylin-eosin staining results showed that the number of inflammatory cells infiltrated in the transplanted kidney tissue was higher in the antibody-mediated rejection group than the immune tolerance group (P < 0.05). Immunohistochemistry results showed that the antibody-mediated rejection group had a large number of C4d positive cells in the kidney tissue, while there were no C4d-positive cells in the immune tolerance group. The mRNA expression of miR-4286 and the proportion of follicular helper T cells in the antibody-mediated rejection group were higher than those in the immune tolerance group (P < 0.05). Spearman correlation analysis indicated that the expression of miR-4286 was positively correlated with the proportion of follicular helper T cells, while negatively correlated with the proportion of follicular regulatory T cells. Compared with the blank vector group, the level of transforming growth factor β was decreased in the miR-4286 overexpression group (P < 0.05), and the levels of interferon gamma, interleukin 6, and interleukin 21 were increased in the miR-4286 overexpression group (P < 0.05). Compared with the blank vector group, in the miR-4286 silence group, the level of transforming growth factor β was increased (P < 0.05), while the levels of interferon γ, interleukin 6, and interleukin 21 were decreased (P < 0.05). Compared with the miR-4286 silence group, the level of transforming growth factor beta was decreased in the transforming growth factor β blocking group (P < 0.05), and the levels of interferon γ, interleukin 6, and interleukin 21 was increased (P < 0.05). Compared with the blank vector group, the proportion of B cells and plasma cells was increased in the miR-4286 overexpression group (P < 0.05), and the proportion of follicular regulatory T cells was decreased (P < 0.05). Compared with the blank vector group, the proportion of B cells and plasma cells was decreased in the miR-4286 silence group (P < 0.05), and the proportion of follicular regulatory T cells was increased (P < 0.05). Compared with the miR-4286 silence group, the proportion of B cells and plasma cells was increased in the transforming growth factor β blocking group (P < 0.05), and the proportion of follicular regulatory T cells was decreased (P < 0.05). All these findings indicate that miR-4286 is highly expressed in peripheral blood lymphocytes of antibody-mediated rejection patients after kidney transplantation, and silencing of miR-4286 can up-regulate the proportion of follicular regulatory T cells in lymphocytes and inhibit humoral immunity through transforming growth factor β pathway.  

Key words: kidney transplantation, antibody-mediated rejection, miR-4286, lymphocyte, follicular regulatory T cell, plasma cell, transforming growth factor β