Epithelial-mesenchymal transition (EMT) is critical for carcinoma invasiveness and metastasis. To investigate the role of membrane-type-2 matrix metalloproteinase (MT2-MMP) in EMT, we generated lentiviral constructs of wild-type (WT) and an inactive Glu260Ala (E260A) mutant MT2-MMP and derived stably transfected HCT116 and A549 cell lines. WT-transfected cells appeared mesenchymal-like, whereas cells transfected with the E260A mutant were epithelial-like, as were cells treated with an MMP inhibitor (GM6001). Expression of E-cadherin, β-catenin, and zonula occludens-1 was lower in cells transfected with WT MT2-MMP compared to vector controls, cells treated with GM6001, or cells transfected with the E260A mutant. An 80-kD N-terminal fragment of E-cadherin was immunoprecipitated in conditioned medium from WT MT2-MMP cells, but not in the medium from vector controls, cells treated with GM6001, or E260A mutant cells. When endogenous expression of MT2-MMP in A2780 human ovarian cancer cells was inhibited using GM6001 or MT2-MMP-specific siRNA, levels of the 80-kD E-cadherin fragment in conditioned medium were decreased. Chick embryo chorioallantoic membrane invasion assays demonstrated that cells transfected with WT MT2-MMP were more invasive than cells transfected with control vector, treated with GM6001, or transfected with the E260A mutant. These results suggest that MT2-MMP degrades adherens and tight junction proteins and results in EMT, making it a potential mediator of EMT in carcinomas.
These results suggested that TAP variants were likely to play a substantial role in different outcomes of persistent HBV infection in the studied population.
Increasing attention has been focused on the malignant tumor microenvironment, which plays important roles in tumor occurrence, progression and metastasis. Fibroblasts are recruited by platelet-derived growth factor (PDGFs) and invade the tumor microenvironment. In the PDGF family, PDGF-B has been reported to play an important role in the recruitment and invasion programs. However, whether PDGF-D plays a role in these programs remains unclear. We generated a recombinant plasmid expressing human PDGF-D and transfected the plasmid to dermal fibroblasts to examine the effects on cell invasive activities in 3D type I collagen gels. PDGF-D plasmid transfection enhanced fibroblast invasive activities both in invasive cell numbers and invasion depth in 3D collagen gels. These effects were blocked by Snail-specific siRNA transfection. PDGF-D transfection significantly induced Snail expression at both mRNA and protein levels. PDGF-D further upregulated MT1-MMP mRNA and protein expressions and this was inhibited when Snail was knocked down by siRNA. Both Snail and MT1-MMP expressions in fibroblasts and cellular invasive activities in 3D collagen induced by PDGF-D were inhibited by LY294002, SP600125, and U1026, the inhibitors of PI3K, JNK, and ERK1/2 signaling pathways, respectively. However, no effects were observed in response to the P38MAPK signaling pathway inhibitor SB203580. These effects of PDGF-D were confirmed by using the culture supernatants of the transfectants. Taken together, these data demonstrate that PDGF-D plays important roles in the recruitment and invasion programs of fibroblasts via the activation of PI3K, JNK and ERK1/2 signaling pathways, and upregulation of Snail and downstream effecter MT1-MMP. These findings indicate that PDGF-D is an important player in the tumor microenvironment for fibroblast recruitment.
The in vitro culture of pancreatic islets reduces their immunogenicity and prolongs their availability for transplantation. Both simulated microgravity (sMG) and a polyglycolic acid scaffold (PGA) are believed to confer advantages to cell culture. Here, we evaluated the effects of sMG combined with a PGA on the viability, insulin-producing activity and morphological alterations of pancreatic islets. Under PGA-sMG conditions, the purity of the islets was ≥85%, and the islets had a higher survival rate and an increased ability to secrete insulin compared with islets cultured alone in the static, sMG, or PGA conditions. In addition, morphological analysis under scanning electron microscopy (SEM) revealed that the PGA-sMG treatment preserved the integral structure of the islets and facilitated islet adhesion to the scaffolds. These results suggest that PGA-sMG coculture has the potential to improve the viability and function of islets in vitro and provides a promising method for islet transplantation.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.