ABSTRACT
Objective To investigate the effects of CD226 knockout ( KO) on obese mice fed with high fat diet and to analyze the composition of immune cells in CD226KO obese mice for further elucidating the immunological mechanism of CD226 involved in high fat diet-induced obesity. Methods Both wild-type ( WT) and CD226KO mice were randomly divided into two groups, high-fat and normal diet groups, and fed for 14 weeks to establish the type 2 diabetes model. Immune cells in mouse spleen and peripheral blood were analyzed by flow cytometry. In in vitro experiments, NK92-MI cells were infected with pshRNA-CD226 lenti-virus to silence CD226 expression, and then qPCR was performed to detect the expression of Foxp3, TNF-αand IFN-γ at mRNA level. Results In the high-fat diet groups, CD226KO mice had lower blood glucose, serum insulin and HOMA-IR than WT mice, but higher HOMA-IS and HOMA-β. CD226KO could reduce compensatory hyperplasia of islet tissue, and significantly down-regulate the proportion of spleen NK cells in mice. The proportion of CD3-CD49b+CD25+Foxp3+regulatory NK cells (NKreg) increased significantly in CD226KO mice. CD226KO could significantly increase Foxp3 expression in NK92-MI cells and decrease the expression of TNF-α and IFN-γ. Conclusions CD226KO can alleviate insulin resistance, increase the number of islet β-cell and improve islet β-cell function in obese mice. The mechanism might be related to the up-regulation of Foxp3+ NKreg ratio.
ABSTRACT
Objective To investigate the protective mechanism of hypoxia-inducible factor-1α (HIF-1α) in vascular endothelial cells under hypoxia.Methods 1.After a hemorrhagic shock model was established in mice,the vascular endothelial cells were sorted in a shock group (n =3) and a sham operation group (n =3) for RNA-sequencing to analyze the main differential molecules.2.The expression of HIF-1α and glucose transporter-1 (GLUT1) was measured in human umbilical vein endothelial cells (HUVECs) and the mitochondrial membrane potentials were detected in a control group (normal culture,n =3) and a hypoxia group (hypoxia culture for 6 h,n =3).3.The HUVECs cells were transfected with HIF-1α siRNA for 48 h to interfere with HIF-1 α expression,and the control group(n =3) was transfected with control siRNA.The expression of HIF-1α was detected to determine the interference effect.The mRNA and protein expression of GLUT1 was detected in the interference and the control groups after 6 h of hypoxia culture.The mitochondrial membrane potential was detected by fluorescent probe method.Results 1.The transcriptome sequencing in the vascular endothelial cells in the shock and sham operation groups indicated 25 genes with significant differences.The HIF-1 α expression was significantly increased in the shock group (111.70 ± 15.97) than in the sham operation group (53.49 ± 3.26) (P =0.023).2.The expression of HIF-1 α and GLUT1 in the HUVECs cells was significantly increased in the hypoxia group compared with the control group (P ＜ 0.05).The mitochondrial membrane potential was significantly lower in the hypoxia group (0.781 ± 0.023) than in the control group (1.177 ± 0.062) (P ＜ 0.05),indicating aggravated injury.3.There was no significant difference in the mitochondrial membrane potential between the interference group (1.011 ± 0.076) and the control group (1.151 ± 0.031) (P ＞ 0.05).However,after hypoxia culture for 6 h,there was a significant difference in the mitochondrial membrane potential between the interference group (0.514 ± 0.018) and the control group (0.769 ± 0.044) (P ＜ 0.05),also indicating aggravated damage.Conclusions The expression of HIF-1 α may be significantly increased in hemorrhagic shock.In HUVECs under hypoxia,HIF-1 α may up-regulate the expression of GLUT1,promote glucose transport,improve mitochondrial damage and protect vascular endothelial cells.Thus,targeting HIF-1 α may contribute to the treatment of hemorrhagic shock.
ABSTRACT
OBJECTIVE@#To investigate the effect of miR-186 inhibition on the expression of hypoxia-inducible factor-1α (HIF-α) and mitochondrial function in hypoxic vascular endothelial cells.@*METHODS@#Human umbilical vein endothelial cells (HUVECs) cultured in routine or hypoxic conditions for 6 h were examined for the expression of miR-186. A miR-186 inhibitor was transfected in the HUVECs, and the cells were subsequently cultured in hypoxic condition for 6 h to observe the changes in the mitochondrial structure under an electron microscope. The changes in the mRNA and protein expressions of HIF-1α in response to miR-186 interference were tested using real-time fluorescent quantitative PCR and Western blotting.@*RESULTS@#The expression of miR-18 was mildly increased in HUVECs after hypoxic exposure for 6 h (=0.0188). Interference of miR-186 expression obviously promoted the mRNA and protein expressions of HIF-1α in HUVECs. In hypoxic conditions, miR-186 interference significantly reduced mitochondrial damage in HUVECs as observed under electron microscope (=0.0297).@*CONCLUSIONS@#Inhibition of miR-186 protects vascular endothelial cells against hypoxic injuries by promoting HIF-α expression to lessen mitochondrial damage, suggesting the possibility of targeted miR-186 interference for treatment of hemorrhagic shock.