• Animal cells and systems
• /
• v.5 no.3
• /
• pp.205-209
• /
• 2001

Mesenchymal cells from the leg buds of stage 24-chick embryos differentiated into chondrocytes when plated at high density. Treatment of high density micromass culture of chick mesenchymal cells with cytochalasin D(CD, 2 $\mu$M for 24 h) resulted in inhibition of chondrogenesis. CD treatment was found to affect the expression of the contractile protein $\alpha$-smooth muscle actin ($\alpha$-SM actin). In control cultures, $\alpha$-SM actin uniformly expressed from culture day 2, but the CD-treated cells induced expression of $\alpha$-SM actin from the first day of culture followed by a continuous increase. Expression of pan-cadherin (P-cadherin) decreased as chondrogenesis proceeded in the control culture, whereas the CD-treated cells showed sustained expression. These results propose a close connection of chondrogenic differentiation with expression of $\alpha$-SM actin and P-cadherin.

• Korean Journal of Veterinary Service
• /
• v.31 no.4
• /
• pp.449-456
• /
• 2008

The differentiation of mouse embryonic stem(ES) cell into smooth muscle cells(SMC) may play a major role in cardiovascular development and under pathophysiological conditions. Therefore, in the present study, we have examined the differentiation of ES cells and its related gene expression. SMC differentiation was indicated by cellular morphology and time-dependent induction of dibutyryl adenosine 3,5-cyclic monophosphate(DBcAMP)and retinoic acid(RA) on smooth muscle ${\alpha}$-actin($SM{\alpha}A$), smooth muscle myosin heavy chain(SMMHC) gene expression. The control was undifferentiated ES cells(protein expressions represent 50-60kDaOct-4). The results of this study show that morphology of embryoid body and confirmation of $SM{\alpha}A$ expression by immunocytochemistry. Moreover, SMMHC and desmin expression was significantly increased by time dependent manner(5, 7, 15 days), in contrast to $SM{\alpha}A$ expression was slightly decreased on 15days. In conclusion, DBcAMP and RA stimulate mouse ES cells differentiation into SMC and enhanced $SM{\alpha}A$, SMMHC and desmin expression.

• Journal of Life Science
• /
• v.17 no.3 s.83
• /
• pp.420-426
• /
• 2007

In the present investigation, we studied the modulating effects of (-)-epigallocatechin-3-gallate(EGCG) on the differentiation of mouse C2C12 myoblasts. We found that the strong inhibitory effect of EGCG on DNA methyltransferase-mediated DNA methylation induced transdifferentiation of C2C12 myoblasts into smooth muscle cells demonstrated by both morphological changes and immunofluorescent staining. C2C12 myoblasts treated with EGCG for 4 days expressed smooth muscle ${\alpha}-actin$ protein. Real-time PCR data revealed that smooth muscle ${\alpha}-actin$ mRNA was induced by EGCG treated C2C12 myoblasts in a concentration-dependent manner. Smooth muscle ${\alpha}-actin$ mRNA concentration increased 330% and 490% after 2 and 3 days of 50 ${\mu}M$ of EGCG treatment. The expression of another smooth muscle marker, transgelin, mRNA was also increased up to 9-fold by 4 days of EGCG treatment compared with control in a concentration-dependent manner. These results suggested that C2C12 enables to transdifferentiate into smooth muscle when gene expression patterns are changed by the inhibition of DNA methylation induced by EGCG. In conclusion, transdifferentiation of C2C12 myoblasts into smooth muscle is resulted from the modulating effects of EGCG on DNA methylation which subsequently results in changing the expression pattern of several genes playing a critical role in the differentiation of C2C12 myoblasts.

• BMB Reports
• /
• v.33 no.6
• /
• pp.531-536
• /
• 2000

Tropomyosin (TM) is an important actin binding protein involved in regulation of muscle contraction. Unacetylated striated tropomyosin failed to bind to actin whereas unacetylated smooth tropomyosin bound well to actin. It has been demonstrated that high actin affinity of unacetylated ${\alpha}-tropomyosin$ was ascribed to the carboxyl terminal amino acid residues. In order to define the role of the carboxyl terminal residues of tropomyosin molecule on actin binding, two mutant tropomyosins were constructed. TM11 is identical to the striated tropomyosin except that the carboxyl terminal last three amino acids was replaced with $^{282}NNM^{284}$ whereas in TM14 $^{276}HA^{277}$ was substituted with smooth specific $^{276}QT^{277}$. TM11 and TM14 were overproduced in Escherichia coli and analyzed for actin affinity. The apparent binding constants (Kapp) of unacetylated tropomyosins were $2.2{\times}10^6M^{-1}$ for sm9, $1.03{\times}10^6M^{-1}$ for TM14, $0.19{\times}10^6M^{-1}$ for TM11, $>0.1{\times}10^6M^{-1}$ for striated, respectively. This result indicated that higher actin affinity of the unacetylated smooth tropomyosin was primarily attributed to the presence of QT residues in the smooth sequence. In case of the Ala-Ser (AS) dipeptide extension of the amino terminus of tropomyosin, Kapp were $21.1{\times}10^6M^{-1}$ for AS-sm9, $8.0{\times}10^6M^{-1}$ for AS-11, $4.7{\times}10^6M^{-1}$ for AS-14, $3.8{\times}10^6M^{-1}$ for AS-striated. AS-TM11 showed considerably higher actin affinity than AS-TM14, implying that interaction of Ala-Ser of the amino terminus with the carboxyl terminal residues. Since Kapp of AS-TM11 was significantly lower than that of AS-sm9, the presence of QT might be required for restoration of high actin affinity of the smooth ${\alpha}-tropomyosin$. These results suggested that the carboxyl terminal amino acid residues Glutamine275-Threonine276 are important for actin affinity of the recombinant smooth ${\alpha}-tropomyosin$, particularly of unacetylated smooth ${\alpha}-tropomyosin$.

• Journal of Korean Biological Nursing Science
• /
• v.8 no.2
• /
• pp.41-59
• /
• 2006

Chronic liver diseases and hepatic cancer have been reported as 10% of cause of death in Koreans. Regardless of various causes, chronic liver disease accompanies commonly hepatic fibrosis. But still the mechanism of hepatic fibrosis remains poorly understood. Using the dimethylnitrosamine(DMN)-induced hepatic fibrosis rat model, We performed to evaluate the possible therapeutic effect of RIP(extracts of Phellodendron amurense and Patrinia scabiosaefolia) and to investigate the changes in referential connective tissue proteins($TGF-{\beta}_1$, ${\alpha}$-smooth muscle actin, and vimentin) as a marker of fibrogenesis. For these purposes, liver tissues were stained with H & E, and Azan staining for estimation of developing fibrosis. In the DMN-treated rat liver tissue, fibrosis were developed forming incomplete septal fibrosis. Whereas, in the RIP-treated rat liver tissues, the fibrosis were decreased recovering to normal morphology. The expressions of $TGF-{\beta}_1$, ${\alpha}$-smooth muscle actin($\alpha-SMA$), and vimetin were increased in the DMN-treated rat liver tissues, but decreased in the various areas of RIP-treated rat liver tissues. According to these results, RIP could be a possible therapeutic agent to reduce hepatic fibrosis, and the $TGF-{\beta}_1$, ${\alpha}$-SMA, and vimentin could be possible indicative markers of hepatic fibrosis development and recovery.

• Journal of Life Science
• /
• v.17 no.2 s.82
• /
• pp.204-210
• /
• 2007

Previous reports indicated that the carboxyl terminal residues, glutamine276-threonine277 in particular, were important for actin affinity of the unacetylated smooth ${\alpha}-tropomyosin$. To determine the role of the glutamine and threonine residues in C-terminal region in actin binding, we constructed mutant striated muscle ${\alpha}-tropomyosin$ (TMs), in which these two residues were individually substituted. These mutant tropomyosins, designated TM18 (HT) and TM19 (QA), were overexpressed in E. coli as an either unacetylated form or Ala-Ser. (AS) dipeptide fusion form, and were analyzed F-actin affinity by cosedimentation. Unacetylated TM19 (QA) bound to actin approximately three times stronger than TM18 (HT) and much stronger than ST (HA). AS/TM19 (QA) showed four times stronger, in actin affinity than AS/ST (HA) while AS/TM14 (QT) bound to actin stronger to some extent than AS/TM18 (HT). These results suggested that the presence of Gln residue at 276 be primarily attributed to higher actin affinity of smooth ${\alpha}-tropomyosin$.

• Journal of Life Science
• /
• v.17 no.7 s.87
• /
• pp.976-982
• /
• 2007

The purpose of this study was to determine the modulating effects of histone deacetylase inhibitor, trichostatin A, on the differentiation of mouse C2C12 myoblasts. We demonstrated that trichostatin A induced morphological changes of C2C12 myoblasts into smooth muscles and significantly increased the gene expression of smooth muscle markers including smooth muscle ${\alpha}-actin$ and transgelin. These results were due to the change in the expression level of cell cycle regulators in trichostatin A-treated C2C12 cells. Real-time PCR data revealed that cyclin dependent kinase inhibitor, p21, mRNA expression was significantly increased in trichostatin A-treated C2C12 cells. However, trichostaDn A rapidly decreased cyclin Dl mRNA expression necessary for cell cycle progression in 24hr after treatment. In conclusion, the strong inhibitory effects of trichostatin A on histone deacetylation induced transdifferentiation of C2C12 myoblasts into smooth muscle cells and these results are partly due to the changes in the expression of cell cycle regulators such as p21 and cyclin D1.

• Molecules and Cells
• /
• v.24 no.3
• /
• pp.441-444
• /
• 2007

Despite the importance of cell fate decisions regulated by epigenetic programming, no experimental model has been available to study transdifferentiation from myoblasts to smooth muscle cells. In the present study, we show that myoblast cells can be induced to transdifferentiate into smooth muscle cells by modulating their epigenetic programming. The DNA methylation inhibitor, zubularine, induced the morphological transformation of C2C12 myoblasts into smooth muscle cells accompanied by de novo synthesis of smooth muscle markers such as smooth muscle ${\alpha}$-actin and transgelin. Furthermore, an increase of p21 and decrease of cyclinD1 mRNA were observed following zebularine treatment, pointing to inhibition of cell cycle progression. This system may provide a useful model for studying the early stages of smooth muscle cell differentiation.

• Journal of Life Science
• /
• v.23 no.1
• /
• pp.137-142
• /
• 2013

Human adipose-derived mesenchymal stem cells (hMSCs) are highly useful for vascular regeneration of injured or inflamed tissue. Lipopolysaccharide (LPS) is a potent activator of macrophages and stimulates macrophages to release inflammatory cytokines. In the present study, we explored the role of LPS-activated macrophages in the differentiation of hMSCs to smooth muscle cells (SMCs). We demonstrated that conditioned medium from LPS-induced macrophages (LPS CM) stimulates differentiation of hMSCs to SMCs, as evidenced by increased expression of smooth muscle-specific markers, including alpha-smooth muscle actin (${\alpha}$-SMA), smooth muscle-myosin heavy chain, and calponin. LPS induced the secretion of $PGF2{\alpha}$ from macrophages, and $PGF2{\alpha}$ treatment stimulated expression levels of SMC-specific markers in hMSCs. Furthermore, small interfering RNA-mediated silencing of the $PGF2{\alpha}$ receptor inhibited LPS CM-stimulated ${\alpha}$-SMA expression. These results suggest that LPS-activated macrophages promote differentiation of hMSCs to SMCs through a $PGF2{\alpha}$-dependent mechanism.

• Korean Journal of Veterinary Service
• /
• v.35 no.2
• /
• pp.153-157
• /
• 2012

A 7-year-old, castrated male dachshund dog with tumor of the spleen portion was referred to the Kangwon National University. The tumor removed surgically and tumor size was 6~7 cm. Histopathologically, this neoplasm was the presence of perivascular whorls of fusiform cells. The cells also arranged in interlacing bundles or storiform patterns. And this neoplasm consisted of spindle cells that often formed distinct whorls around a central capillary. Immunohistochemical analysis revealed multi-focally immunoreactivity for ${\alpha}$-smooth muscle actin, whereas not immunoreactive for desmin, S-100, von Willebrand factor. On the base of the histological and immunohistochemical results, this neoplasm was diagnosed as a canine hemangiopericytoma.