scholarly journals Combination of PD98059 and TGF-β1 Efficiently Differentiates Human Urine-Derived Stem Cells into Smooth Muscle Cells

2021 ◽  
Vol 22 (19) ◽  
pp. 10532
Author(s):  
Yongha Hwang ◽  
Seon-Heui Cha ◽  
Donghee Kim ◽  
Hee-Sook Jun
Keyword(s):  
Stem Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Transforming Growth Factor ◽  
Quantitative Polymerase Chain Reaction ◽  
Muscle Cells ◽  
Immunocytochemical Staining ◽  
Sphincter Muscle ◽  
Differentiated Cells ◽  
Tgf Β1

Pluripotent adult stem cells have potential applications in cell therapy and tissue engineering. Urine-derived stem cells (UDSCs) differentiate into various cell types. Here, we attempted to differentiate human UDSCs (hUDSCs) into smooth muscle cells (SMCs) using transforming growth factor-beta 1 (TGF-β1) and/or PD98059, an extracellular signal-regulated kinase (ERK) inhibitor. Both quantitative polymerase chain reaction (qPCR) and Western blot analysis showed that the expression of messenger ribonucleic acid (mRNA) and proteins for alpha-smooth muscle actin (α-SMA), calponin (CNN1), and smooth muscle myosin heavy chain (SM-MHC), which are specific markers for SMCs, increased on day 9 after differentiation and again on day 14. The differentiated cells from human UDSCs (hUDSCs) with a combination of TGF-β1 and PD98059 showed the highest expression of SMC marker proteins. Immunocytochemical staining performed to assess the molecular expression revealed CNN and α-SMA colocalizing in the cytoplasm. The cells that differentiated from hUDSCs with a combination of TGF-β1 and PD98059 showed the strongest expression for CNN1, α-SMA, and SM-MHC. Functional testing of the differentiated cells revealed a stronger contractile capacity for the cells differentiated with a combination of PD98059 and TGF-β1 than those differentiated with a single factor. These results suggest the combination of PD98059 and TGF-β1 to be a more effective differentiation method and that differentiated SMCs could be used for restoring the functions of the sphincter muscle or bladder.

scholarly journals TGF-β1 Regulates Smooth Muscle Cells Differentiation from Adipose-Derived Stem Cells via the Wnt/β-Catenin Pathway

2021 ◽  
Author(s):  
Xuling Lv ◽  
Hao Chen ◽  
Zikai Zhang ◽  
Tian Li ◽  
Qing Wei ◽  
...  
Keyword(s):  
Stem Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Pelvic Floor ◽  
Signaling Pathway ◽  
Myogenic Differentiation ◽  
Pelvic Floor Dysfunction ◽  
Muscle Cells ◽  
Adipose Derived Stem Cells ◽  
Tgf Β1

Abstract Background: Pelvic floor dysfunction (PFD) is a spectrum of disorders including stress urinary incontinence and pelvic organ prolapse. Transforming growth factor-β1 (TGF-β1) can induce mesenchymal stem cells (MSCs) to differentiate into smooth muscle cells (SMCs). SMCs derived from adipose-derived stem cells (ADSCs) can be used to repair damaged pelvic floor smooth muscle tissues, which is of great interest for clinical applications using stem cell therapy for PFD. The Wnt/β-catenin pathway acts as a decisive factor in the fate of stem cells.Methods and Results: In this study, we used medium containing TGF-β1, TGF-β1 inhibitor LY2109761, or Wnt/β-catenin inhibitor KYA1797K, to induce ADCSs to differentiate into SMCs in vitro to explore the influence of TGF-β1 on the myogenic differentiation of ADCSs via the Wnt/β-catenin pathway. Results: 1) TGF-β1 induces ADSC-derived SMCs to hyper-express the SMC markers including SMA-α, Desmin, Calponin, and SMMHC ; 2) TGF-β1 activates the Wnt/β-catenin signaling pathway in ADSCs. After blocking TGF-β1, the Wnt/β-catenin pathway and myogenic differentiation in cells were inhibited; 3) the Wnt/β-catenin pathway is involved in the differentiation of ADSCs into SMCs. After differentiation induction, the synchronized changes in the activation of Wnt/β-catenin signaling and the expression of SMC-specific proteins showed a trend of simultaneous changes, and after the inhibition of the Wnt pathway, the adult muscle differentiation was significantly inhibited.Conclusions: We established a simpler and more efficient method for inducing ADSCs to differentiate into SMCs using TGF-β1 and demonstrated that the Wnt/β-catenin signaling pathway is activated during this process.

Transforming growth factor-β1 regulation of C-type natriuretic peptide expression in human vascular smooth muscle cells: dependence on TSC22D1

2010 ◽  
Vol 299 (6) ◽  
pp. H2018-H2027 ◽  
Author(s):  
Maria C. Mendonça ◽  
Nancy Koles ◽  
Sonia Q. Doi ◽  
Donald F. Sellitti
Keyword(s):  
Smooth Muscle ◽  
Coronary Artery ◽  
Growth Factor ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Natriuretic Peptide ◽  
Vascular Smooth Muscle Cells ◽  
Transforming Growth Factor ◽  
Muscle Cells ◽  
Tgf Β1

C-type natriuretic peptide (CNP) possesses nitric oxide-like signaling mechanisms and actions in the vasculature, including the inhibition of fibrosis and vascular remodeling through counterregulation of transforming growth factor-β (TGF-β) signaling. The leucine zipper protein transforming growth factor stimulated clone 22 domain 1 (TSC22D1), cloned via its presumed binding to a GC-rich element in the CNP promoter, was the first protein to be described as a CNP transcription factor, but the lack of supporting evidence since its discovery and its lack of a classical DNA-binding site have left in question its role in the regulation of CNP by TGF-β and other factors. To define a specific role for TSC22D1 in CNP transcription, we have examined the effects of the profibrotic growth factors TGF-β1 and PDGF-BB on CNP mRNA expression in cultured human vascular smooth muscle cells (SMC) in which TSC22D1 expression was suppressed with small interfering RNA. Results showed that TGF-β and PDGF-BB significantly increased CNP expression in all three SMC types. Twenty-four-hour TGF-β-induced elevations in CNP were strongly correlated with changes in TSC22D1 mRNA levels, and both genes exhibited their greatest response to TGF-β1 in coronary artery SMC. Furthermore, siRNA suppression of TSC22D1 expression in coronary artery and aortic SMC by ∼90% resulted in 45–65% reductions of both PDGF- and TGF-β-stimulated CNP expression, respectively. These results support a postulated role of TSC22D1 as an enhancer of CNP transcription and suggest that TGF-β-induced upregulation of CNP expression in SMC may be mediated in part by increased transcription of TSC22D1.

scholarly journals MiR-149 attenuates the proliferation and migration of TGF-β1-induced airway smooth muscle cells by targeting TRPM7 and affecting downstream MAPK signal pathway

2020 ◽  
Author(s):  
Zhengyu Zhu ◽  
Liya Zhang ◽  
Ting Jiang ◽  
Yan Qian ◽  
Yun Sun ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Airway Smooth Muscle ◽  
Transforming Growth Factor ◽  
Muscle Cells ◽  
Airway Smooth Muscle Cells ◽  
And Migration ◽  
Tgf Β1 ◽  
Proliferation And Migration

Asthma is considered as a general term for various chronic inflammatory diseases of the respiratory tract. Growing evidences have supported that microRNAs were involved in mediating cell proliferation, migration, and other cellular functions. MiR-149 has been found to take part in the development of various cancers. However, whether miR-149 participated in the proliferation and migration of transforming growth factor beta 1 (TGF-β1)-induced airway smooth muscle cells was still unknown. In this study, the expression level of miR-149 in human airway smooth muscle cells (ASMCs) was decreased after TGF-β1 treatment in vitro. Additionally, the over-expression of miR-149 obviously suppressed proliferation and migration in human ASMCs. Besides, we found that overexpression of miR-149 could inhibit the expression of transient receptor potential melastatin 7 (TRPM7) both in protein and gene levels. Furthermore, we demonstrated that miR-149 could inhibit the cell proliferation and migration in human ASMCs by targeting TRPM7 through modulating mitogen-activated protein kinases (MAPKs) signaling pathway. Taken together, we strongly supported that miR-149 might be a key inhibitor of asthma by targeting TRMP7. Therefore, our finding suggests a promising biomarker for the development of further targeted therapies for asthma.

scholarly journals Shear Stress–Induced miR-143-3p in Collateral Arteries Contributes to Outward Vessel Growth by Targeting Collagen V-α2

2020 ◽  
Vol 40 (5) ◽  
Author(s):  
Kerstin Troidl ◽  
Thomas Hammerschick ◽  
Julian Albarran-Juarez ◽  
Georg Jung ◽  
Wilma Schierling ◽  
...  
Keyword(s):  
Blood Flow ◽  
Shear Stress ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Fluid Shear Stress ◽  
Transforming Growth Factor ◽  
Quantitative Polymerase Chain Reaction ◽  
Muscle Cells ◽  
Collateral Arteries ◽  
Collateral Artery

Objective: Arteriogenesis, describing the process of collateral artery growth, is activated by fluid shear stress (FSS). Since this vascular mechanotransduction may involve microRNAs (miRNAs), we investigated the FSS-induced expression of vascular cell miRNAs and their functional impact on collateral artery growth during arteriogenesis. Approach and Results: To this end, rats underwent femoral artery ligation and arteriovenous anastomosis to increase collateral blood flow to maximize FSS and trigger collateral vessel remodeling. Five days after surgery, a miRNA expression profile was obtained from collateral tissue, and upregulation of 4 miRNAs (miR-24-3p, miR-143-3p, miR-146a-5p, and miR-195-5p) was verified by quantitative polymerase chain reaction. Knockdown of miRNAs at the same time of the surgery in an in vivo mouse ligation and recovery model demonstrated that inhibition of miR-143-3p only severely impaired blood flow recovery due to decreased arteriogenesis. In situ hybridization revealed distinct localization of miR-143-3p in the vessel wall of growing collateral arteries predominantly in smooth muscle cells. To investigate the mechanotransduction of FSS leading to the increased miR-143-3p expression, cultured endothelial cells were exposed to FSS. This provoked the expression and release of TGF-β (transforming growth factor-β), which increased the expression of miR-143-3p in smooth muscle cells in the presence of SRF (serum response factor) and myocardin. COL5A2 (collagen type V-α2)—a target gene of miR-143-3p predicted by in silico analysis—was found to be downregulated in growing collaterals. Conclusions: These results indicate that the increased miR-143-3p expression in response to FSS might contribute to the reorganization of the extracellular matrix, which is important for vascular remodeling processes, by inhibiting collagen V-α2 biosynthesis.

scholarly journals Role of complement 3 in renin generation during the differentiation of mesenchymal stem cells to smooth muscle cells

2020 ◽  
Vol 318 (5) ◽  
pp. C981-C990 ◽  
Author(s):  
Lan Chen ◽  
Noboru Fukuda ◽  
Shoichi Shimizu ◽  
Hiroki Kobayashi ◽  
Sho Tanaka ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Transcription Factor ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Nuclear Translocation ◽  
Muscle Cells ◽  
Wky Rats ◽  
Differentiated Cells ◽  
Undifferentiated State

We showed that increased expression of complement 3 (C3) induces dedifferentiation of mesenchymal cells and epithelial mesenchymal transition, which activate the local renin-angiotensin system (RAS) that contributes to cardiovascular and renal remodeling in spontaneously hypertensive rats (SHRs). In the present study, to investigate contributions of C3 to the development of the pathogenesis of hypertension, we evaluated the formation of renin-producing cells and roles of C3 in renin generation during differentiation of primary bone marrow-mesenchymal stem cells (MSCs) from C57BL/6 mice, Wistar-Kyoto (WKY) rats, and SHRs to smooth muscle cells (SMCs) with transforming growth factor-β1. The expression of renin transiently increased with increases in transcription factor liver X receptor α (LXRα), and expression of C3 and Krüppel -like factor 5 (KLF5) increased during differentiation of MSCs from C57BL/6 mice, WKY rats, and SHRs to SMCs. Exogenous C3a stimulated renin and LXRα expression accompanied by nuclear translocation of LXRα. C3a receptor antagonist SB290157 suppressed renin and LXRα expression, with inhibition of nuclear translocation of LXRα during the differentiation of mouse MSCs to SMCs. The expression of C3 and KLF5 was significantly higher in the differentiated cells from SHRs compared with the cells from WKY rats during differentiation. Renin-producing cells were formed during differentiation of MSCs to SMCs, and renin generation was observed in undifferentiated SMCs, in which transient expression of renin in the differentiated cells with lower differentiation stage was stronger from SHRs than that from WKY rats. Expression and nuclear localization of LXRα in the differentiated cells from SHRs were stronger than that from WKY rats. C3 was important in forming and maintaining this undifferentiated state of SMCs from MSCs to generate renin with increases in transcription factor LXRα and KLF5. Increases in C3 expression maintain the undifferentiated state of SMCs from MSCs to generate renin that activates RAS and contributes to the pathogenesis of hypertension in SHRs.

Down-regulation of calponin destabilizes actin cytoskeletal structure in A7r5 cells

2007 ◽  
Vol 85 (2) ◽  
pp. 225-232 ◽  
Author(s):  
Ava C. Dykes ◽  
Gary L. Wright
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Smooth Muscle Cells ◽  
Laser Scanning ◽  
Transforming Growth Factor ◽  
Muscle Cells ◽  
Stress Fibers ◽  
Resting Cells ◽  
Tgf Β1 ◽  
Actin Stress Fibers

The effects of changes in the expression levels of h1 calponin (CaP) on actin cytoskeletal organization were studied in control and phorbol-ester-treated A7r5 smooth muscle cells. Protein association and expression in control and stimulated A7r5 smooth muscle cells were evaluated by Western blotting, laser scanning confocal microscopy (LSCM), and fluorescence resonance energy transfer (FRET) microscopy in cells treated with either 2 × 10−6 mol/L TGF-β1 or 2 × 10−5 mol/L PDGF-BB to alter h1 calponin expression. Single immunostained samples showed that CaP and α-actin, localized in fibers in unstimulated control A7r5 smooth muscle cells, were translocated to podosomes following treatment with phorbol-12,13-dibutyrate (PDBu). Confocal colocalization imaging and FRET analysis both indicated substantial association of CaP with α-actin in stress fibers of control cells and in podosomes of PDBu-treated cells. PKCα, which showed evidence of only slight association with CaP in control cells, exhibited markedly increased (293%) association in PDBu-contracted cells. Platelet-derived growth factor (PDGF)-BB down-regulated CaP to non-detectable levels, whereas transforming growth factor (TGF)-β1 up-regulated (424%) the expression of CaP without affecting the levels of α-actin or PKCα. PDGF-BB resulted in a significant loss in α-actin stress fibers (–47%) and reduced podosome formation (–69%). By comparison, TGF-β1 had no effect on stress fibers in control cells but also reduced (–70%) podosome formation. The results suggest that CaP could play a major role in the stabilization of actin stress fibers in resting cells and may contribute to podosome formation in PDBu-treated cells.

Sign in / Sign up

Export Citation Format

Share Document