Notch2 negatively regulates myofibroblastic differentiation of myoblasts

Abstract

Myofibroblasts are one of the key cellular components involved in fibrosis of skeletal muscle as well as in other tissues. Transforming growth factor-beta1 (TGF-beta1) stimulates differentiation of mesenchymal cells into myofibroblasts, but little is known about the regulatory mechanisms of myofibroblastic differentiation. Since Notch2 was shown to be downregulated in TGF-beta1-induced non-muscle fibrogenic tissue, we investigated whether Notch2 also has a distinctive role in myofibroblastic differentiation of myogenic cells induced by TGF-beta1. TGF-beta1 treatment of C2C12 myoblasts led to expression of myofibroblastic marker alpha-smooth muscle actin (alpha-SMA) and collagen I with concomitant downregulation of Notch2 expression. Overexpression of active Notch2 inhibited TGF-beta1-induced expression of alpha-SMA and collagen I. Interestingly, transient knockdown of Notch2 by siRNA in C2C12 myoblasts and primary cultured muscle-derived progenitor cells resulted in differentiation into myofibroblastic cells expressing alpha-SMA and collagen I without TGF-beta1 treatment. Furthermore, we found Notch3 was counter-regulated by Notch2 in C2C12 cells. These findings suggest that Notch2 is inhibiting differentiation of myoblasts into myofibroblasts with downregulation of Notch3 expression.

Notch1 Signaling in FIZZ1 Induction of Myofibroblast Differentiation

“…41 Additionally overexpression of active Notch2 is shown to inhibit TGF-␤-induced ␣-SMA expression, whereas transient knockdown of Notch2 by siRNA in C2C12 myoblasts results in differentiation into myofibroblastic cells without TGF-␤ treatment. 40 In contrast, Notch3 is required for TGF-␤-induced myofibroblastic differentiation, and counterregulated by Notch2 in these cells, indicating a countervailing regulatory system dependent on which Notch receptor is being activated. A similar mechanism is suggested by a more recent study showing that whereas Notch signaling via CSL up-regulates ␣-SMA gene transcription, signaling via HRT1 or HRT2 suppresses transcription.…”

“…35,36 Several studies also show that Notch signaling regulates ␣-SMA expression differentially via CSL or HRT in multiple cell types, including smooth muscle cells and mesenchymal fibroblasts, respectively. [37][38][39][40][41] Because Notch1 is shown to mediate activation of Th2 differentiation, 11 which is a predominant response in pulmonary fibrosis, the objective of this study was to clarify the activation of Jagged1/ Notch1 signaling in response to FIZZ1 and analyze its role in myofibroblast differentiation in pulmonary fibrosis. These results for the first time showed the importance of Notch1 signaling in FIZZ1-induced myofibroblast differentiation and bleomycin (BLM)-induced pulmonary fibrosis.…”

Transforming growth factor beta-1 expression in macrophages of human chronic periapical diseases

“…TGF-b1 appears to play an important role in inducing fibroblastic differentiation of periodontal ligament (PDL) stem/progenitor cells and in maintaining the PDL apparatus under physiological conditions (Fujii et al, 2010). It also regulates the expression of collagen type I and promotes tissue healing (Okubo et al, 2003;Fujita et al, 2004;Ono et al, 2007). In addition, the gradual increase in connective tissue growth factor (CTGF) and TGF-b1 levels in hepatic tissue is associated with liver fibrosis.…”

Presenilin-1 acts via Id1 to regulate the function of muscle satellite cells in a γ-secretase-independent manner

“…Immunocytochemistry was performed as previously described (Ono et al, 2007). Primary antibodies were used in PBS as follows: goat anti-PS1 (recognises Nterminal), anti-Id1, anti-Ki67, anti-MyHC, anti-Pax7, anti-Myog, mouse anti-GFP, rabbit anti-GFP, anti-V5 or anti-MyoD antibody at 4°C overnight.…”

“…The transfection of siRNA (Stealth siRNA; Invitrogen) into C2C12 myoblasts and primary muscle progenitors cells was performed using Lipofectamine 2000 reagent (Invitrogen) as previously described (Ono et al, 2007). Transfection of siRNA into single myofibres was carried out 20-24 hours after isolation.…”