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Hydrogels preserve native phenotypes of valvular fibroblasts through an elasticity-regulated PI3K/AKT pathway.

ABSTRACT: Matrix elasticity regulates proliferation, apoptosis, and differentiation of many cell types across various tissues. In particular, stiffened matrix in fibrotic lesions has been shown to promote pathogenic myofibroblast activation. To better understand the underlying pathways by which fibroblasts mechano-sense matrix elasticity, we cultured primary valvular interstitial cells (VICs) isolated from porcine aortic valves on poly(ethylene glycol)-based hydrogels with physiologically relevant and tunable elasticities. We show that soft hydrogels preserve the quiescent fibroblast phenotype of VICs much better than stiff plastic plates. We demonstrate that the PI3K/AKT pathway is significantly up-regulated when VICs are cultured on stiff gels or tissue culture polystyrene compared with freshly isolated VICs. In contrast, myofibroblasts de-activate and pAKT/AKT decreases as early as 2 h after reducing the substrate modulus. When PI3K or AKT is inhibited on stiff substrates, myofibroblast activation is blocked. When constitutively active PI3K is overexpressed, the myofibroblast phenotype is promoted even on soft substrates. These data suggest that valvular fibroblasts are sensing the changes in matrix elasticity through the PI3K/AKT pathway. This mechanism may be used by other mechano-sensitive cells in response to substrate modulus, and this pathway may be a worthwhile target for treating matrix stiffness-associated diseases. Furthermore, hydrogels can be designed to recapitulate important mechanical cues in native tissues to preserve aspects of the native phenotype of primary cells for understanding basic cellular responses to biophysical and biochemical signals, and for tissue-engineering applications.

SUBMITTER: Wang H

PROVIDER: S-EPMC3845151 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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Hydrogels preserve native phenotypes of valvular fibroblasts through an elasticity-regulated PI3K/AKT pathway.

Wang Huan H Wang Huan H Tibbitt Mark W MW Langer Stephen J SJ Leinwand Leslie A LA Anseth Kristi S KS

Proceedings of the National Academy of Sciences of the United States of America 20131111 48

Matrix elasticity regulates proliferation, apoptosis, and differentiation of many cell types across various tissues. In particular, stiffened matrix in fibrotic lesions has been shown to promote pathogenic myofibroblast activation. To better understand the underlying pathways by which fibroblasts mechano-sense matrix elasticity, we cultured primary valvular interstitial cells (VICs) isolated from porcine aortic valves on poly(ethylene glycol)-based hydrogels with physiologically relevant and tun ...[more]

PMID: 24218588

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Project description:BACKGROUND:As the most abundant epigenetic modification on mRNAs and long non-coding RNAs, N6-methyladenosine (m6A) modification extensively exists in mammalian cells. Controlled by writers (methyltransferases), readers (signal transducers), and erasers (demethylases), m6A influences mRNA structure, maturation, and stability, thus negatively regulating protein expression in a post-translational manner. Nevertheless, current understanding of m6A's roles in tumorigenesis, especially in gastric cancer (GC) remains to be unveiled. In this study, we assessed m6A's clinicopathological relevance to GC and explored the underlying mechanisms. METHODS:By referring to a proteomics-based GC cohort we previously generated and the TCGA-GC cohort, we merged expressions of canonical m6A writers (METTL3/METTL14), readers (YTHDF1/YTHDF2/YTHDF3), and erasers (ALKBH5/FTO), respectively, as W, R, and E signatures to represent m6A modification. We stratified patients according to these signatures to decipher m6A's associations with crucial mutations, prognosis, and clinical indexes. m6A's biological functions in GC were predicted by gene set enrichment analysis (GSEA) and validated by in vitro experiments. RESULTS:We discovered that W and R were potential tumor suppressive signatures, while E was a potential oncogenic signature in GC. According to W/R/E stratifications, patients with low m6A-indications were accompanied with higher mutations of specific genes (CDH1, AR, GLI3, SETBP1, RHOA, MUC6, and TP53) and also demonstrated adverse clinical outcomes. GSEA suggested that reduced m6A was correlated with oncogenic signaling and phenotypes. Through in vitro experiments, we proved that m6A suppression (represented by METTL14 knockdown) promoted GC cell proliferation and invasiveness through activating Wnt and PI3K-Akt signaling, while m6A elevation (represented by FTO knockdown) reversed these phenotypical and molecular changes. m6A may also be involved in interferon signaling and immune responses of GC. CONCLUSIONS:Our work demonstrated that low-m6A signatures predicted adverse clinicopathological features of GC, while the reduction of RNA m6A methylation activated oncogenic Wnt/PI3K-Akt signaling and promoted malignant phenotypes of GC cells.

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Hydrogels preserve native phenotypes of valvular fibroblasts through an elasticity-regulated PI3K/AKT pathway.

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Hydrogels preserve native phenotypes of valvular fibroblasts through an elasticity-regulated PI3K/AKT pathway.

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