Inflammatory pain-related miR-34a mediates mitochondrial homeostasis via XIST and attenuates cell apoptosis
The goal of this study is to elucidate the mechanism by which inflammatory pain regulated mitochondrial function and apoptosis of macrophages via down-regulation of miR-34a and up-regulation of X-inactive specific transcript(XIST).For this purpose,the expressions of XIST were measured in miR-34a over-expressed or LPS-treated J774A.1(female mouse cell line)and the SH-SY5Y(female human cell line)cells;And the in vivo correlation between the expressions of miR-34a and XIST was also analyzed.MitoTracker and TMRE(tetramethylrhodamine ethyl ester)staining were used to analyze the mitochondrial volume and membrane potential in J774A.1 cell line.The mitochondrial stress assay(Seahorse)was conducted to test the mitochondrial function in J774A.1 cells after over-expressing miR-34a and/or stimulated with LPS.TUNEL assay was used to assess cell line apoptosis.CFA(complete Freund's adjuvant)-induced acute inflammatory pain model was established to detect the pain reaction threshold and the expression of XIST and miR-34a in acute and chronic phases and healthy controls,respec-tively.Real-time PCR was used to detect the expression of XIST in the blood of complex regional pain syndrome(CRPS)pa-tients and healthy controls.The results showed that there was no significant difference in XIST expression between the control group and the CRPS group in total human samples(t=1.528,P=0.131).And there was no statistical difference between the two groups(t=0.945,P=0.353)when only male patients were analyzed.However,in female patients,the XIST expression of the CRPS group was significantly higher than that of the control group(t=2.764,P=0.008).Over-expression of miR-34a reduced XIST expression in J774A.1 and SH-SY5Y cells.The datum from patients showed that the expression of miR-34a and XIST exhibited a negative correlation(r2=0.681,P＜0.001).LPS treatment inhibited the expression of miR-34a and up-regulated XIST expression in J774A.1 cell line.In mouse pain model,XIST expression was up-regulated(t=1.810,P＜0.001)and miR-34a expression was down-regulated(t=2.220,P＜0.001)at 4 h time point,and the difference disappeared at D14.LPS up-regulated mitochondrial volume and membrane potential,while over-expression of miR-34a led to down-regulation of both.And the effects of the two treatments compensated for each other.LPS stimulation significantly decreased cellular oxygen consumption(t=4.323,P=0.014),ATP production(t=4.323,P=0.014)and the maximum oxygen consumption(t=1.885,P=0.008),while miR-34a over expression had the opposite effect(t=4.245,P=0.004).LPS reduced cell apoptosis whereas miR-34a over-expression promoted it.XIST over-expression can reverse the cell apoptosis mediated by miR-34.In conclusion,inflammation may promote the expression of XIST by down-regulating miR-34a and the former suppresses cell apoptosis by promoting the maintenance of mitochondrial function.
X-inactive specific transcriptmicro ribonucleic acid 34apainenergy metabolismapoptosis
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