Fig 2: The lncRNA MEG3 promotes trophoblast cell metastasis and invasiveness through its regulation of the miR-21/BMPR2 signaling pathway. A,B) RT-qPCR analysis of the transcription levels of miR-21-5p and MEG3 in the placenta for the normal pregnancy group and PE patients. C) Correlation analysis between the transcription levels of miR-21-5p and MEG3 in the placental tissues of PE patients; *p<0.05 relative to the normal pregnancy group. Data are presented as the means ±SEM, n = 3.

Fig 3: The effect of OIP5 knockdown on downstream genes. (A) Analysis of downstream genes of OIP5 using IPA analysis. (B) RT-qPCR was used to determine the changes in the expression of OIP5 downstream genes in Huh7 cells at 72 h following pGCSIL-shOIP5 transfection. Samples were normalized to GAPDH mRNA expression. RT-qPCR results are indicated as mean ± standard deviation of three independent experiments performed in triplicate. GAPDH protein was used as the internal control. *P<0.05 and **P<0.01 vs. pGCSIL-shCtrl-transfected cells. (C) The expression of OIP5 downstream proteins were analyzed by western blot analysis in Huh7 cells transfected with pGCSIL-shOIP5 lentivirus. #, target genes with a Z-score >3.0; RT-qPCR, reverse transcription-quantitative polymerase chain reaction; Ctrl, control; sh, short hairpin; OIP5, Opa interacting protein 5; mir-1, microRNA1; CDKN2A, cyclin dependent kinase inhibitor 2A; TWF1, twinfilin actin binding protein 1; H3F3A, H3 histone family member 3A; HNRNPU, heterogeneous nuclear ribonucleoprotein U; EEF1A1, eukaryotic translation elongation factor 1a1; RB1, retinoblastoma 1; MAD2L1, mitotic arrest deficient 2 like 1; BTG2, B-cell translation gene 2; BMPR2, bone morphogenetic protein receptor type 2; CHEK1, checkpoint kinase 1; RAC1, Rac family small GTPase 1; PDIA3, protein disulfide isomerase family A member 3; RPS6KB1, ribosomal protein S6 kinase B1; CUL4B, cullin 4B.

Fig 4: Irisin promoted the phosphorylation of ERK/STAT through binding to integrin receptor aV and up-regulates the expression of BMP2 to enhance osteogenic differentiation. (A) Cell lysates were subjected to immunoblotting analysis. Protein expression levels of integrin aV, p-Erk1/2, p-STAT3, BMPR2, p-Smad1/5/9, and Smad4 of BMSCs induced into osteoblasts for 3 days in the presence of r-irisin and SB273005. (B) ALP and ARS staining. (C) Protein expression levels of BMP2, Smad1/5/9, p-Smad1/5/9 and Smad4 after 3 days of osteogenic induction of BMSCs treated with r-irisin, and the ratio of p-Smad1/5/9 to Smad1/5/9 of the protein grayscale values. (D) Immunohistochemical staining of p-Smad1/5/9 in the femur of WT and FNDC5-/- mice (n=6, *P <0.05, ***P <0.001, ****P <0.0001).

Fig 5: Expression assay results of c.419-43delT carriers and wild-type patients. (A) Dot plot representing the fold change between carriers and wild-type patients without taking into consideration cell cycle stage. The results showed a slight increase in the variant carriers’ BMPR2 expression, and these data were not statistically significant. (B) Dot plot representing the fold change between variant carriers and wild-type patients taking into account cell cycle stage. Data showed a rise of gene expression in the G0/G1 and a slight decrease for S phase for the carriers, while in the G2/M, it showed almost identical expression between both groups. There were no statistically significant differences between stages.