关键词: 急性肝衰竭, 脐带间充质干细胞, 树突状细胞, TLR4, NF-κB

Abstract: Objective To investigate the role of umbilical cord mesenchymal stem cells (hU-MSCs) in the regulation of inflammatory environment by regulating the maturation and differentiation of dendritic cells (DCs) in patients with acute hepatic failure (AHF). Methods DCs derive from five patients with AHF were co-cultured with hU-MSCs. The maturation and activation levels of DCs and the proportion of regulatory DC cells (regDCs) and the cytokine levels in the cell culture supernatants were measured with or without the administration of NS398, a selective cyclooxygenase-2 (COX-2) inhibitor, to the co-culture system. The expression of Toll like receptor 4 (TLR4), inhibitor of nuclear factor kappa-B kinase subunit alpha (IKKα) and nuclear factors-κB p65 (NF-κB-p65) were then detected by Western blot. New therapeutic targets were screened by whole transcriptomic sequencing analysis on the DCs before and after co-culture with hU-MSCs. Results After co-cultured with hU-MSCs, regDCs increased. The expression levels of surface markers MHC-II, CD80 and CD86 on immature DCs (imDCs) increased. The levels of IL-1 and IL-6 decreased in cocultured DCs compared to DCs that cultured alone [IL-1α: (754.68±22.65)pg/ml vs (76.67±11.25)pg/mL;IL-1β:(336.49±55.75)pg/mL vs (85.33±12.58)pg/mL;IL-6:(38027.85±5231.14)pg/mL vs (12000.75±6793.69)pg/mL; P<0.0500)], whereas the level of IL-10 increased significantly [(0.41±0.01)pg/mL vs (0.61±0.01)pg/mL; P<0.05]. After co-cultured with hU-MSCs, the down-regulation of NF-κB-p65 [(0.777±0.01 vs 0.55±0.02; P<0.05)] and TLR4[(1.12±0.01 vs 0.54±0.01; P<0.05)] expression and up-regulation of IKKα expression in DCs were significant [(0.12±0.01 vs 0.88±0.01; P<0.05)]. After the administration of NS398 to DCs that cocultured with hU-MSCs, the expression of NF-κB-p65 and TLR4 (0.14±0.01, 0.12±0.01) in DCs were down-regulated, and IKKα was significantly up-regulated (1.18±0.01). By transcriptomic sequencing analysis it was found that the expressions of 909 genes were up-regulated and 363 genes were down-regulated in DCs co-cultured with hU-MSCs when compared with those without. Most of the genes played important roles in inflammation. Conclusion hU-MSCs showed an immuno-suppressive effect by inducing the differentiation of imDCs derived from AHF to CD11bhigh1alowCD11clowregDCs in vitro, and played an anti-inflammatory role by suppressing the inflammatory cascade reaction through TLR4/NF-κB/COX-2 pathway.

Key words: Acute hepatic failure, hU-MSCs, DCs, NF-κB