将以下文字翻译为英文并润色。目的：原发性胆汁性胆管炎Primary Biliary Cholangitis PBC患者的肝脏和外周血中自然杀伤细胞Natural Killer cells NK的数量明显增加可能参与其发病活化 T 细胞核因子 C1Nuclear factor of activated T cell cytoplasmic 1 NFATC1是影响 NK 细胞激活和发挥作用的重要基因。

Purpose: The number of natural killer cells (NK) in the liver and peripheral blood of patients with primary biliary cholangitis (PBC) is significantly increased, which may be involved in its pathogenesis. Nuclear factor of activated T cell cytoplasmic 1 (NFATC1) is an important gene that affects the activation and function of NK cells. MiRNA participates in the activation process of NK cells, and many studies have shown that miRNA regulates NK cells in other autoimmune diseases. However, the role of miRNA in the pathogenesis of PBC has not been reported. This study aims to investigate whether miRNA has an effect on the activation and protein secretion of NK cells in PBC and whether it is involved in its pathogenesis.

Methods: Human miRNA related to the NFATC1 gene was predicted through a gene prediction website, and the intersection of the results obtained from multiple websites was taken. Ten miRNAs with strong correlation to NFATC1 were selected, and the corresponding miRNA was synthesized after finding the relevant sequence. Peripheral blood samples from 168 PBC patients and 74 healthy controls (HC) were collected. NK cells were sorted using magnetic bead method, and RNA was extracted from them for qPCR analysis to quantify the ten miRNAs in NK cells to analyze whether there were differences in miRNA expression between the PBC group and the HC group. Dual-luciferase reporter gene experiments were used to verify whether miRNA can bind to NFATC1. The mimics and inhibitors of three differentially expressed miRNAs were constructed for overexpression or inhibition of NK cells. After the system was constructed and transferred into NK cells, the supernatant was collected by cell lysis for ELISA detection of perforin and granzyme secreted by NK cells. Flow cytometry was used to determine the purity of NK cell sorting and verify the effect of miRNA on CD molecules on the surface of NK cells. The experimental results were statistically analyzed, and corresponding software was used for plotting.

Results: The expression of hsa-miR-137-3p, hsa-miR-124-3p, and hsa-miR-506-3p in peripheral blood NK cells of the PBC group and the HC group showed significant statistical differences (P＜0.05). The Ct values of hsa-miR-137-3p, hsa-miR-124-3p, and hsa-miR-506-3p in NK cells of PBC patients were all lower than those of HC, and the differences were statistically significant (11.33±4.40 vs 11.85±4.40, P＜0.01; 8.61±2.22 vs 8.94±2.99, P＜0.01; and 7.18±1.89 vs 8.29±1.46, P＜0.05), indicating that the expression of these three miRNAs in NK cells of PBC patients was higher than that of HC. The dual-luciferase reporter gene experiments verified that miR-137-3p can bind to NFATC1 gene, and the fluorescence intensity of the group with miR-137-3p wild-type (WT) plasmid binding was weaker than the negative control group (P＜0.05), while hsa-miR-124-3p and hsa-miR-506-3p had no significant effect on WT. Mimics overexpression of miR-137-3p in NK cells resulted in a higher level of granzyme B than miR-137-3p inhibitor (348.59±7.47pg/ml vs 373.92±15.50pg/ml, P＜0.05). Flow cytometry showed that after overexpression of miR-137-3p, the proportion of CD3-CD16+ NK cells increased significantly compared to the miR-137-3p negative control group (73.2% vs 46.8%, P＜0.05), indicating that the binding of miR-137-3p and NFATC1 may increase the expression of CD16 on the surface of NK cells while decreasing CD56 molecules.

Conclusion: The expression levels of miR-137-3p, miR-124-3p, and miR-506-3p in NK cells are higher in PBC than in HC. MiR-137-3p can bind to the NFATC1 gene and promote the expression of CD16 on NK cells. Overexpression of miR-137-3p can promote the secretion of granzyme B in NK cells of PBC patients. MiR-124-3p and miR-506-3p have no effect on CD16 and perforin/granzyme B. In conclusion, miRNA-137-3p may play an important role in the activation of NK cells in PBC, possibly by binding to the NFATC1 gene to promote the expression of CD molecules on the surface of NK cells and secretion of granzyme B