Fig 2: Effects of erianin on the RhoA/ROCK1 pathway in vitro. (A) The mRNA levels of RHOA, ROCK1, and MYPT1 in HRVECs exposed to normoxia, CoCl2, or erianin at various concentrations. (B) (B–I) Protein levels of GTP-RhoA, ROCK1, and p-MYPT1 in HRVECs exposed to normoxia, CoCl2, or erianin at various concentrations. (B–II) Quantitative analysis of GTP-RhoA, ROCK1, and p-MYPT1; the results are presented as percentages of the normoxia group. * Significant difference between samples (paired t-test, P values < 0.05); # Comparison with the collagen group of CoCl2-incubated cells without erianin treatment (paired t-test, P values < 0.05); † Comparison with noncollagen group of normoxia cells without erianin treatment (independent sample t-test, P values < 0.05).

Fig 3: Western blot validation of specific proteins identified in the proteomic analysis.Western blot analysis was used to validate some of the targets identified on proteomic profiling. The protein expression of CDC2 was 2-fold higher in the AD cells compared to the AI tumorspheres, and PARP1, SOX2 and HDGFR were expressed > 1.5-fold higher in the AI tumorspheres compared to the AD cells. In addition, the expression levels of MIA3, RhoA, PDGFRβ, VEGFR and EGFR was also significantly (P < 0.05) higher in the AI tumorspheres, as determined with Western blot analysis. * indicates a P value < 0.05. Results represent the mean ± SEM of multiple experiments.

Fig 4: Migrasomes promote adipose tissue regeneration. (A) Schematic illustration of injection of poorly vascularized Ischemic adipose tissue with migrasomes (+ Mig group) or PBS (PBS group). N = 7 in each groups. (B) Western blot analysis of CXCL12 protein expression in adipose tissue from the + Mig and PBS groups at 7 days after surgery. (C) Semiquantitative analysis of CXCL12 protein expression. *p < 0.05 compared with PBS group. (D) Macroscopic image of adipose tissue from the + Mig and PBS groups at 30 days after surgery. Scale bar, 50 mm. (E) HE staining of adipose tissue from the + Mig and PBS groups at 30 days after surgery. Scale bar, 200 μm. (F) Immunofluorescence staining of CD34 and perilipin in adipose tissue from the + Mig and PBS groups at 30 days after surgery. Scale bar, 20 μm. (G) Quantitative analysis of infiltrated CD34+ ASCs per field in adipose tissue from the + Mig and PBS groups over time. (H) Quantitative analysis of the perilipin+ area per field in adipose tissue from the + Mig and PBS groups over time. *p < 0.05 compared with PBS group. (I) Western blot analysis of CXCR4, RhoA, and PPARγ protein expression in adipose tissue from the + Mig and PBS groups 7 days after surgery. (J-L) Semiquantitative analysis of CXCR4, RhoA, and PPARγ protein expression. *p < 0.05 compared with PBS group. The data are mean ± SEM. Statistical differences in (C) were tested by nonparametric Mann-Whitney test. (G), (H) and (J) to (L) were analyzed by One-way ANOVA

Fig 5: Activation dynamics of the GTPases RhoA and Rac1 that control cell adhesion and spreading on Fn were altered in 129 TG2−/− relative to TG2+/+ MEFs. (A,C) Representative western blots and (B,D) quantitation of activated GTP-bound RhoA (GTP-RhoA) relative to total RhoA (B) or activated GTP-bound Rac1 (GTP-Rac1) relative to total Rac1 (D) in lysates from Fn-adherent 129 TG2+/+ MEFs, TG2−/− MEFs, TG2−/− MEFs transfected with TG2 cDNA (TG2t) or TG2−/− MEFs with recombinant wild-type TG2 added to the Fn matrix (+TG2, 20 μg/cm2) at 10, 30, 60 or 90 min post-seeding, with GTPγS-treated TG2+/+ lysates as a positive control (+ve) (n = 3 experiments). *, p < 0.05; **, p < 0.01, ***, p < 0.001 for individual post hoc Bonferroni test results from two-way ANOVA. #, p < 0.05; ##, p < 0.01, ###, p < 0.001 for individual post hoc Bonferroni test results two-way ANOVA comparing TG2−/− with TG2+/+ MEFs at the indicated time points.