Jichu yixue yu linchuang (Apr 2022)

Down-regulation of lncRNA-H19 expression promotes apoptosis and autophagy of human keloid fibroblasts

  • HUANG Yun, XU Xi-sheng, CHEN Kai, HE Xiu

DOI
https://doi.org/10.16352/j.issn.1001-6325.2022.04.023
Journal volume & issue
Vol. 42, no. 4
pp. 633 – 639

Abstract

Read online

Objective To investigate the effects of down-regulation of long non-coding RNA-H19 (lncRNA-H19) expression on apoptosis and autophagy of human keloid fibroblast cell line and to explore potential mechanism. Methods Human scar fibroblasts cell line (KFs) and normal human dermal fibroblasts cell line (HDFs) were collected and KFs cells were infected with lncRNA-H19F low-expressing adenovirus (si-lncRNA-H19) and empty virus vector without si-lncRNA-H19 (si-eGFP). They were divided into HDFs group, KFs group, si-lncRNA-H19+KFs group and si-eGFP+KFs group. KFs cells were co-transfected with si-lncRNA, mTOR over-expressing adenovirus (Ad-mTOR) and empty vector (Ad-eGFP-m), then divided into KFs group, si-lncRNA-H19+Ad-mTOR group,si-eGFP+Ad-mTOR group, si-lncRNA-H19+Ad-eGFP group and si-eGFP+Ad-eGFP group. RT-qPCR was used to detect the expression of lncRNA-H19; flow cytometry was used to detect the cell apoptosis ; autophagy was examined by EM microscopy with fluorescence (acridine orange,AO) staining; immuno-histochemical method was used to detect the positive expression of autophagy pathway-mammalian target of rapamycin (mTOR); Western blot was used to detect the expression of apoptosis and autophagy-related proteins. Results Compared with the HDFs group, the expression of lncRNA-H19 and mTOR activity in the KFs group increased and the number of autophagic vesicles and the expression of apoptosis and autophagy-related proteins decreased (P＜0.05); after down-regulating lncRNA-H19 in KFs cells, the activity of mTOR in KFs cells decreased and apoptosis/autophagy increased (P＜0.05). Up-regulating mTOR while down-regulating lncRNA-H19 lead to more autophagy and apoptotic in KFs cells as compared to that resulted from down-regulation of lncRNA-H19 alone (P＜0.05). Conclusions Down-regulation of lncRNA-H19 expression may promote the autophagy and apoptosis of KFs cells through inhibiting activation of mTOR pathway.

Keywords