Fig 2: High expression of meCAF markers in the stroma predicts poor prognosis in patients with PDAC.a Higher expression of four meCAF marker genes (PLA2G2A, CRABP2, SERPINE2, and MFAP5) predicting a worse overall survival (OS) in patients with PDAC (TCGA PAAD database, n = 178). b Representative H&E and IHC stainings of meCAF markers (PLA2G2A and CRABP2) indicated high expression of these markers in loose-type PDAC (n = 94). High expression of meCAF markers (PLA2G2A + CRABP2) in the stroma predicted a poor OS in patients with PDAC (n = 76). c Representative IHC staining of myCAF marker (POSTN) indicated its high expression in dense stroma of PDAC (n = 94). The expression level of POSTN in stroma showed no correlation with OS of PDAC patients (n = 76). d–f Representative IHC staining of an iCAF marker (APOD) and CAF subcluster 1 and 2 markers (RGS5 and MYH11). The expression of these markers show no correlation with the overall survival in patients with PDACs. Scale bar, 100 µm. Censored samples are indicated by “+”. Kaplan–Meier curve was used for survival analysis. All statistical analyses were performed with the Log-rank test.

Fig 3: HPSMCs identification. (A) Cell morphology of freshly isolated and cultured HPSMCs at passages 0 and 3. Scale bar, 50 µm. (B) Immunofluorescence staining of a-SMA, SM22a, Desmin, Calponin and SMMHC (all green), and cell nucleus (blue) in HPSMCs at passage 3. Scale bar, 50 µm. (C) Flow cytometric analysis revealed that HPSMCs were positive for CD29, PDGFR-ß and CD90, whilst negative for CD31, CD34, CD45, CD14 and KDR. HPSMC, human prostate smooth muscle cells; SMMHC, smooth muscle myosin heavy chain; PDGFR-ß, platelet-derived growth factor receptor ß; KDR, kinase insert domain receptor; SM22a, transgelin-1.

Fig 4: High stretch induces SMC alignment but downregulates most markers of differentiated SMCs. (A) Representative images of vascular SMCs after 8 h on stretched/unstretched silicone chambers or rigid culture plates. (B) ACTA2/smooth muscle actin as well as LMOD1 were upregulated in cells grown on tissue culture plates as compared to silicon membranes. However, MYH11 showed the opposite trend. Representative images show 24 h. (C) MYOCD, TAGLN and CNN1 transcript levels were elevated after 24 h in SMCs grown on rigid plates, while PDLIM7 mRNA expression was most highly upregulated after 24 h under non-stretch conditions. Images show (A) 4×/20× magnification and (B) 63×. Insets show corresponding isotype negative control. Plots show mean with SEM. Statistical difference assessed by two-way ANOVA. Expression levels calculated according to ??CT and normalized to unstretched conditions at 8 h.

Fig 5: Mechanism diagram. Overexpression of DNMT3B in GC inhibited MYH11 expression by promoting methylation of the MYH11 promoter, thereby attenuating the repressive effect of MYH11 on TNFRSF14 transcriptional activity and promoting GC progression