Fig 1: Effects of SPJs on expression levels of genes involved in TGF-β1/Smad signaling in the renal tissue of aging rats. The mRNA expression levels of TGF-β1 (A), Smad2(B), Smad3(C), and Smad4(D). (E) Representative results of Western blot analysis with antibodies against TGF-β1, Smad2/3, and Smad4, β-actin was used as a control; and (F) The quantitative relative protein expression levels of MMP2, MMP9, TIMP1 and TIMP2 based on Western blot. All the experiments were performed in triplicate. Data in (A-D) and (F) were expressed as mean ± SD, ∗p < 0.05, n = 6.
Fig 2: RASSF10 impaired cell adhesion through MMP2 via FAK signaling. (a) Eight genes modulated by RASSF10 in HCC migration or invasion were found by cDNA adhesion array, which were further confirmed by qRT-PCR. White bars indicate the result of cDNA adhesion array, and black bars represent qRT-PCR data (the value of 2−ΔΔCT) in QGY 7703 transfected with pcDNA3.1/RASSF10 or empty vector (pcDNA3.1). (b) Western blot confirmed the association between RASSF10 and MMP2 or TIMP2 by over-expression or knock-down assay. (c) Regulatory effect of RASSF10 on FAK and MAPKs. Over-expression of RASSF10 has suppressed the accumulation of total or phosphorylation FAK and p38 MAPK; consistently, down-regulation of RASSF10 by SiRNA/RASSF10 in Huh 7 cells induced the activity of FAK, as well as p38 MAPK. The expression of ERK1/2 or JNK1/2 was independent on the level of RASSF10. (d) Regulatory effect of FAK on MMP2. Depletion of FAK suppressed the expression of MMP2. (e) Rescued assay for MMP2 in stable cell lines (QGY7703/RASSF10) was evidenced by western blot. (f) The change of cell invasion property was re-evaluated following MMP2 rescued assay. More invaded cells were observed with the restored expression of MMP2 in QGY7703/RASSF10. Invaded cells were stained with cell stain solution, counted by microscope in five random high power fields. Data are mean±s.d. The asterisk indicates statistical significance (*P<0.05, **P<0.01).
Fig 3: ColXV induced ECM remodeling dependent on MMPs. All mice were pretreated with either Ad-GFP or Ad-Col15a1. (A) Relative mRNA expressions of Col1a1, Col4a1, Col6a1 and Fibronectin in mice inguinal white adipose tissue (iWAT) with or without GM6001 (n = 6). (B–D) Relative protein expressions of Fibronectin, Col I, Col VI, MMP-2, MMP-9, TIMP-1, TIMP-2 and Cleaved Caspase-3 in mice iWAT with gradients concentration of GM6001 (n = 6). (E-F) Relative protein expressions of Col I, Col VI, MMP-2, TIMP-1 and TIMP-2 in the Control group, APMA group and co-treatment of Ad-Col15a1 and APMA group in mice iWAT (n = 6), each group has two technical repeats (E). (G–J) Representative images of Col I and MMP-9 in adipocytes by immunofluorescent staining in different groups (n = 4). (K) Representative images of collagen bundles microstructure on adipocytes surface by scanning electron microscope. Scale bar, 30 μm (top) and 10 μm (bottom) (n = 4). Arrowheads indicate disrupted collagen fibers and exposed adipocytes surface (middle). Asterisks indicate adipocytes. Values are presented as mean ± SEM. *p < 0.05, ** p < 0.01, ns, not significant.
Fig 4: Overexpression of miR-195 inhibits the expression of metastasis-associated gene and the activation of the PI3K/AKT signaling pathway in Hec1A cells. (A-C) Relative mRNA levels of TIMP-2, MMP-2 and MMP-9 were detected by RT-qPCR. (D and E) Relative protein levels were measured by western blot analysis. (F-H) The protein and phosphorylation levels of PI3K and AKT were detected by western blot analysis. Each value represents the mean ± SEM (n=3). GAPDH was detected as an internal control. aaP<0.01 vs. Control group; bbP<0.01 vs. Mock group. TIMP-2, tissue inhibitor of metalloproteinase 2; MMP, matrix metalloproteinase.
Fig 5: Expression of MMP-9, TIMP-2, E-cadherin and vimentin in the control, NC, miR-490-5p mimics, miR-490-5p mimics + MAP3K9 and MAP3K9 groups of BICR 18 and FaDu cells. (A) Relative mRNA expressions of MMP-9, TIMP-2, E-cadherin and vimentin in each group of BICR 18 cells. (B) Relative mRNA expressions of MMP-9, TIMP-2, E-cadherin and vimentin in each group of FaDu cells. (C) The original outcomes of western blot of the expressions of MMP-9, TIMP-2, E-cadherin and vimentin in each group of BICR 18 cells. (D) Relative protein levels of MMP-9, TIMP-2, E-cadherin and vimentin (of β-actin) in each group of BICR 18 cells (E) The original outcomes of western blotting of the expression of MMP-9, TIMP-2, E-cadherin and vimentin in each group of FaDu cells. (F) Relative protein levels of MMP-9, TIMP-2, E-cadherin and vimentin (of β-actin) in each group of FaDu cells. Bars indicates the mean ± standard deviation. *P<0.05 vs. NC groups; #P<0.05 vs. miR-490-5p mimics groups. miR, microRNA; NC, negative control; MAPK, mitogen activated protein kinase; E, epithelial; TIMP-2, tissue inhibitor of metalloproteinase; MMP, matrix metalloproteinase.
Supplier Page from Abcam for Anti-TIMP2 antibody