LncRNA H19 inhibits ER stress induced apoptosis and improves diabetic cardiomyopathy by regulating PI3K/AKT/mTOR axis

Figures

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  Figure 1. H19 improved left ventricular dysfunction. (A) Representative M-mode echocardiographic images. (B) LVEF and LVFS. Data are expressed as mean ± SEM. ∗P < 0.05. DCM vs. DCM+H19 group, n = 6/group.

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  Figure 2. H19 reduced cardiomyocyte apoptosis and cardiac fibrosis in DM. (A) TUNEL staining was used to examine the apoptosis of cardiomyocytes. (B) Masson's trichrome staining was used to investigate the involvement of H19 in cardiac fibrosis in myocardial tissue. (C) The binding site of lncRNA H19 and miR-140-5p and relative expression of lncRNA H19 and miR-140-5p levels were determined by Q-PCR in each group. (D) Binding site of mir-140-5p to PIK3CA. (E) Immunofluorescence intensity of collagen III in each group. (F) The protein levels of p-PI3K, t-PI3K, a-SMA, and collagen-I/III in each group. #P < 0.05; ##P < 0.01 vs. Sham group; *P < 0.05; **P < 0.01 vs. DCM group. between groups.

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  Figure 3. Overview of lncRNAH19 and PI3KCA expression in different DCM datasets. GSE101585, GSE26887 and GSE124405 were selected and downloaded from GEO database to identify the DEGs between DCM tissues and corresponding controls. (A and B) The heatmaps were screened out to illustrate the DEGs in DCM and corresponding controls using 124405 and GSE26887. (C) A volcano map was plotted to illustrate the DEGs using GSE101585. (D) GO enrichment analysis of abnormal regulation DEGs using GSE101585. (E) KEGG enrichment analysis of abnormal regulation DEGs using GSE101585. (F) GO biological processes analysis for the DEGs from GSE124401. (G and H) GO and KEGG enrichment analysis of abnormal regulation DEGs using GSE124405. (I) The PPI network of DEGs using GSE26887. (J) The top 10 hub genes in the PPI network were screened by Cytoscape plugin CytoHubba. (K) The Cytoscape co-expression analysis between differently expressed mRNAs and lncRNAs. (L) lncRNA-RNA interaction prediction.

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  Figure 4. Effect of H19 on ERS-associated apoptosis markers and PI3K/AKT/mTOR pathway in DM. (A) Protein bands exposed the expression of p- PI3K, AKT and mTOR, Nrf2 in each group. (B) Immunofluorescence intensity of GRP78 and p-IRE1a in each group. (C) protein levels of p-PERK, p-IRE1α, ATF6, CHOP, cleaved caspase-3, cleaved caspase-9, cleaved caspase-12, and BAX proteins in all groups. #P < 0.05; ##P < 0.01 vs. DCM group; *P < 0.05; **P < 0.01 vs. sham group, n = 6/group.

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  Figure 5. H19 repressed ERS in vitro. (A) Immunofluorescence staining exposed the expression of p-PERK and CHOP in four cell groups. (B) The statistical results of relative fluorescence intensity demonstrated the protein expression of p-PERK and CHOP in four cell groups. ##P < 0.05 Low glucose vs. High glucose. **P < 0.05 High glucose+H19 OE vs. High glucose. &&P < 0.05 High glucose vs. High glucose+ Thapsigargin, n = 3/group.

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  Figure 6. H19 attenuated ERS-induced apoptosis and activated PI3K in HL-1 cells after HG. (A) Protein bands exposed the expression of p-PI3K, Nrf2, p-PERK, p-IRE1α, GRP 78 and GRP94 proteins in four cell groups. (B) Protein bands exposed the expression of CHOP, cleaved caspase-3, cleaved caspase-9, cleaved caspase-12, and BAX proteins in four cell groups, n = 3/group.

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  Figure 7. H19 suppressed ROS, and ERS by activating PI3K-Nrf2 signals. (A) Protein bands of the p-PI3K, Nrf2, NOX-2/-4, GRP78, GRP94, p-PERK, and p-IRE1a in each group. (B) Statistical data of each group, n = 3/group.

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  Figure 8. Diagrammatic sketch for lncRNA H19’s protective roles in DCMs by activating PI3K/AKT signals and inhibiting ROS, ERS induced apoptosis.