Fig 2: LINC00961 knockdown attenuates TGF-β-induced EndMT. (A) Immunofluorescence staining was used to identify CD31+/α-SMA+ cells. α-SMA (red), CD31 (green); scale bars: 50 µm. (B) Reverse transcription-quantitative PCR was used to evaluate EndMT-related mRNA. (C) Western blotting was used to evaluate EndMT-related proteins. (a) Control, (b) TGF-β, (c) TGF-β + sh-LINC00961, (d) TGF-β + sh-Scramble, (e) TGF-β + LV-LINC00961 and (f) TGF-β + LV-Control. Each experiment was repeated at least three times, ns, P>0.05; *P<0.05 and #P<0.05. n=5 per group. EndMT, endothelial-mesenchymal transition; α-SMA, α-smooth muscle actin; sh-, short hairpin; LV, lentivirus; VE-Cad, VE cadherin; FSP-1, fibroblast specific protein 1; ns, not significant.

Fig 3: Schematic figure highlighting the connections among identified proteins and their roles. LINC00961 knockdown attenuates endothelial injuries and endothelial-mesenchymal transition in vitro by suppressing the PTEN-PI3K-AKT pathway.

Fig 4: LINC00961 knockdown attenuates injury and EndMT in HCMECs by activating the PTEN-PI3K-AKT signalling pathway. (A) Immunofluorescence staining was used to identify CD31 + /α-SMA+ cells. α-SMA (red), CD31 (green); scale bars: 50 µm. (B) Rate of cell apoptosis was evaluated using flow cytometry. (C) Reverse transcription-quantitative PCR was used to determine the expression of PTEN, PI3K, AKT and mTOR mRNA. (D) Western blotting was used to determine the protein expression of PTEN, p-PI3K, PI3K, p-AKT, AKT, p-mTOR and mTOR. (a) TGF-β + sh-LINC00961, (b) TGF-β, (c) TGF-β + LV-LINC00961 and (d) TGF-β + sh-LINC00961 + VOT. Each experiment was repeated at least three times. ns, P>0.05; *P<0.05, #P<0.05 and &P<0.05. n=5 per group. EndMT, endothelial-mesenchymal transition; HCMECs, human cardiac microvascular endothelial cells; PTEN, phosphatase and tensin homolog deleted on chromosome 10; α-SMA, α-smooth muscle actin; p-, phosphorylated; sh-, short hairpin; LV, lentivirus; VOT, VO-OHpic trihydrate; ns, not significant.