Fig 2: Phosphorylation of Protein kinase B (PKB) associates with CDC-induced HSC activation. Murine hepatic stellate cells (mHSCs) were stimulated with CDC in the presence or absence of Phosphatidyl-inositol-3-kinase (PI3K) pan-inhibitors. (a) Phosphorylation of PKB (pPKB) was detected after stimulation with CDC (50–250 µM, control 0.01% dimethyl sulfoxide) by Western blotting (n = 6). Results are shown as mean ± SD (* p <0.05 vs. control, Fisher’s LSD). (b) HSCs were stimulated with CDC (100 µM) for 4 h on day 7 in the absence or presence of PI3K inhibitors LY294002 (LY; 5 µM) or Wortmannin (WT; 0.1 µM), respectively. Phosphorylation of PKB was confirmed by Western blotting (n = 8–11). (c) Deposition of collagen was determined after 14 days of stimulation with CDC (100 µM) in the absence or presence of LY294002 (5 µM) and Wortmannin (0.1 µM), respectively, by Sirius Red staining (n = 3–5). Controls were treated with diluent (0.1% dimethyl sulfoxide). Results are shown as mean ± SD (* p < 0.05 vs. control, Student’s t-test).

Fig 3: PIK3CD and miR-433-5p were selected as the downstream molecules related to HOTAIRM1. a GSEA enrichment analysis revealed that apoptotic pathway was closely related to PCOS. b 9 genes including HOTAIRM1 were obtained from the intersection of apoptosis-related genes and differentially expressed genes in PCOS. c Data from GSE34526 showed that PIK3CD was highly expressed in PCOS samples, p < 0.05. d A positive association between HOTAIRM1 and PIK3CD expression was identified, p = 0.0433. eThe miRNAs corresponding to PIK3CD or HOTAIRM1 were predicted by Targetscan or LncBase, respectively. Intersecting 31 common miRNAs with differentially expressed miRNAs in PCOS to obtain miR-433-5p. f Data from GSE86241 displayed that miR-433-5p was under expressed in PCOS patients, p < 0.05

Fig 4: HOTAIRM1 inhibited the growth and promoted the apoptosis of KGN cells by modulation of miR-433-5p and PIK3CD. a Data from CCK-8 assay showed that the inhibitory effect of HOTAIRM1 on KGN cells growth was suppressed by miR-433-5p mimic or PIK3CD depletion. b Analysis from CCK-8 revealed that depletion of miR-433-5p suppressed the promoting effect of si-HOTAIRM1 on KGN cells viability, upregulation of PIK3CD has a similar result. c Flow cytometry assay displayed that the promoting effect of HOTAIRM1 on KGN cells apoptosis was inhibited by miR-433-5p mimic or PIK3CD depletion. d Analysis from flow cytometry showed that knockdown of miR-433-5p attenuated the inhibitory effect of si-HOTAIRM1 on KGN cells apoptosis, while the inhibitory effect of si-HOTAIRM1 on KGN cells apoptosis was suppressed by PIK3CD upregulation. **p < 0.01 vs. control, ##p < 0.01 vs. HOTAIRM1 or si-HOTAIRM1, &p < 0.05, &&p < 0.01 vs. mimic or inhibitor

Fig 5: Genetic inhibition of PI3K p110β with siRNA reduces viability of SETD2 deficient ccRCC-derived cells(A) Bar graph showing relative cell viability as a percentage of control (si-CTL) of SETD2 (+/+) 786-0 and SETD2 (KO) 786-0 and (–/–) A498 cells in response to treatment with siRNA specific to p110α, p110β, and p110δ (singlets and doublets). *P < 0.05; **P < 0.005; ***P < 0.001; ****P < 0.0001; ns, no statistical significance observed. Standard deviations were calculated and represented for all conditions. (B) Western blot analysis of indicated proteins showing decrease in expression of p110α, p110β, and p110δ in individual or combinatorial treatments (specificity) and variations in phosphorylation levels of known downstream target pAKT-S473 (efficacy). Actin is a loading control.