Effect Of Ufm1 On Human Umbilical Vein Endothelial Cells Inflammation

Objective:To investigate expression and effect of Ubiquitin-fold modifier 1(Ufm1)on HUVECs inflammation and further explore its possible mechanisms.Methods: Human Umbilical Vein Endothelial Cells(HUVECs) were used in this study. The expression and localization of Ufm1 in HUVECs were observed by immunofluorescence assay. Concentration of 100ng/ml LPS was used to induce HUVECs inflammation in vitro. After stimulation for 0h, 3h, 6h, 12 h, 24 h, 48 h, total RNA was retracted.And then the m RNA levels of inflammatory cytokines TNFα, IL-6, IL-1β, IL-12, MCP-1 and Ufm1 were detected by real-time PCR. Meanwhile, Ufm1 protein level was tested by Western Blotting. In addition, Ufm1 overexpression plasmid was constructed and transfected into HUVECs. After LPS stimulation for 3h, Real-Time PCR was used to detect the changes of inflammatory cytokines in each group. In order to explore the possible molecular mechanism of Ufm1 inhibiting the expression of inflammatory cytokines in HUVECs, the total protein level of NF-?B and its nuclear level were detected by WB.Results: Ufm1 was expressed in both the nucleus and cytoplasm of HUVECs. In LPS induced HUVECs inflammatory response, m RNA levels of inflammatory cytokines TNFα, IL-6, IL-1β, IL-12, MCP-1 and Ufm1 increased with time. And after stimulation for 3-6 h, their m RNA levels reached the peak. There was statistical difference(p<0.05). At the same time, the m RNA and protein level of Ufm1 increased with time, and in a time-dependent manner. After Ufm1 overexpression, m RNA levels of related inflammatory cytokines induced by LPS were significantly decreased. Compared with the control group, NF-?B p65 in the nucleus increased after LPS stimulation, but ufm1 overexpression decreased p65 translocation to nucleus.Conclusions: Ufm1 was expressed in HUVECs and localized in the nucleus and cytoplasm; In LPS induced HUVECs inflammation, Ufm1 m RNA and protein levels were unregulated; Ufm1 overexpression can significantly inhibit the expression of LPS induced inflammatory cytokines; Ufm1 may play an inhibitory role of HUVECs inflammation by TLR4-NF-?B signaling pathway.