• Asian-Australasian Journal of Animal Sciences
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• v.20 no.6
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• pp.827-831
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• 2007

Myostatin is a member of the transforming growth factor-${\beta}$(TGF-${\beta}$ super-family. It acts as a negative regulator for skeletal muscle growth. Myostatin mutations are characterized by a visible, generalized increase in muscle mass in double muscled cattle breeds. To understand the biochemistry and physiology of the Chinese Yellow bovine myostatin gene, we report here for the first time expression of the gene in Escherichia coli (E. coli). Primers of the myostatin gene of Chinese Yellow Cattle were designed on the basis of the reported bovine myostatin mRNA sequence (Gen-Bank Accession No. NM005259) and optimized for E. coli codon usage. XhoI and EcoRI restriction enzyme sites were incorporated in the primers, and then cloning vector and expression vector were constructed in a different host bacterium. The expressed protein had a molecule mass of about 16 kDa as determined by SDS-PAGE under reducing conditions. The expressed protein reacted specifically with myostatin monoclonal antibody on immunoblots. Our studies should lead to the investigation of the differences in myostatin genes of various cattle and could benefit human health and food animal agriculture.

• Fisheries and Aquatic Sciences
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• v.10 no.2
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• pp.60-67
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• 2007

Myostatin (MSTN; also known as GDF8) is a member of the transforming growth factor ${\beta}-superfamily$ of proteins. MSTN negatively regulates mammalian skeletal muscle growth and development by inhibiting myoblast proliferation. Mice and cattle possessing mutant MSTN alleles display a 'double muscling' phenotype characterized by extreme skeletal muscle hypertrophy and/or hyperplasia. We isolated the full-length cDNA of a novel MSTN gene from S. schlegeli muscle tissue and examined its expression pattern in various tissues. The full-length gene (GenBank DQ423474) consists of 1941bp with an open reading frame of 1134 bp, encoding 377 amino acids that show 62-92% amino acid similarity to other vertebrate MSTNs. The predicted protein contains a conserved proteolytic cleavage site (RXRR) and nine conserved cysteine residues at the C terminus. RT-PCR revealed that the unprocessed and prodomain myostatin mRNAs were predominantly present in muscle, with limited expression in other tissues. However, the mature myostatin mRNA was highly expressed in brain and muscle, intermediately expressed in the gills, intestine, heart, and kidney, and weakly expressed in the liver and spleen.

• BMB Reports
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• v.45 no.10
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• pp.571-576
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• 2012

Radiotherapy is considered to cause detrimental effects on bone tissue eventually increasing bone loss and fracture risk. However, there is a great controversy on the real effects of irradiation itself on osteoblasts, and the mechanisms by which irradiation affects osteoblast differentiation and mineralization are not completely understood. We explored how X-ray radiation influences differentiation and bone-specific gene expression in mouse calvarial osteoblasts. Irradiation at 2 Gy not only increased differentiation and mineralization of the cells, but also upregulated the expression of alkaline phosphatase, type I collagen, osteopontin, and osteocalcin at early stages of differentiation. However, irradiation at higher doses (>2 Gy) did not stimulate osteoblast differentiation, rather it suppressed DNA synthesis by the cells without a toxic effect. Additional experiments suggested that transforming growth factor-beta 1 and runt-transcription factor 2 play important roles in irradiation- stimulated bone differentiation by acting as upstream regulators of bone-specific markers.

• Journal of Korean Medicine Rehabilitation
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• v.29 no.4
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• pp.1-14
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• 2019

Objectives The object of this study was to assess the effect of Gyejibokryunghwan (GBH) on anti-oxidant and anti-inflammatory activities in RAW 264.7 cells and on factors associated with fracture union in tibia-fractured rats. Methods The 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radical scavenging activity was measured to assess anti-oxidant activity. The production of nitric oxide (NO), interleukin-6 (IL-6), interleukin-$1{\beta}$ ($IL-1{\beta}$) and tumor necrosis factor-${\alpha}$ ($TNF-{\alpha}$) in the RAW 264.7 cells were measured to assess anti-inflammatory activity. The production of osteocalcin, calcitonin, carboxy-terminal telepeptides of type II collagen (CTXII), transforming growth factor-${\beta}$ ($TGF-{\beta}$), bone morphogenetic protein-2 (BMP-2) in serum of tibia-fractured rats were measured to assess the effects of fracture union. X-rays were taken every two weeks from 0 to 4th week to assess fracture union effect. Results DPPH radical scavenging activity of GBH was increased according to concentration of GBH in RAW 264.7 cell. NO, prostaglandin $E_2$ ($PGE_2$), IL-6, $IL-1{\beta}$ and $TNF-{\alpha}$ were significantly decreased, indicating anti-inflammatory effect. Osteocalcin, calcitonin, $TGF-{\beta}$ were significantly increased in the experimental groups. CTXII was significantly decreased in the experimental groups. BMP-2 was not significantly changed in the experimental groups. The X-ray showed that the experimental group has better healing effects on tibia-fractured rats than control group. Conclusions From above result, GBH has an effect on anti-oxidant, anti-inflammatory activities in RAW 264.7 cells. GBH showed significant results in factors related with fracture union and radiologic examination. In conclusion, GBH can help fracture union and it well be expected to be used actively in clinics.

• Journal of Korean Neurosurgical Society
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• v.47 no.1
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• pp.30-35
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• 2010

Objective: The purpose of this study is to explain the effect and reciprocal action among tumor necrosis factor (TNF) like weak inducer of apoptosis (TWEAK), fibroblast growth factor-inducible 14 (Fn14), and transforming growth factor-$\beta1$ (TGF-$\beta1$) on degeneration of human intervertebral disc (IVD). Methods: Human intervertebral disc tissues and cells were cultured with Dulbecco's Modified Eagle's Medium/Nutrient F-12 Ham (DMEM/F-12) media in $37^{\circ}C$, 5% $CO_2$ incubator. When IVD tissues were cultured with TWEAK, Fn14 that is an antagonistic receptor for TWEAK and TGF-$\beta1$, the level of sulfated glycosaminoglycan (sGAG) was estimated by dimethyl methyleneblue (DMMB) assay and sex determining region Y (SRY)-box 9 (Sox9) and versican messenger ribonucleic acid (mRNA) levels were estimated by reverse transcriptase polymerase chain reaction (RT-PCR). Results: When human IVD tissue was cultured for nine days, the sGAG content was elevated in proportion to culture duration. The sGAG was decreased significantly by TWEAK 100 ng/mL, however, Fn14 500 ng/mL did not change the sGAG production of IVD tissue. The Fn14 increased versican and Sox9 mRNA levels decreased with TWEAK in IVD tissue TGF-$\beta1$ 20 ng/mL elevated the sGAG concentration 40% more than control. The sGAG amount decreased with TWEAK was increased with Fn14 or TGF-$\beta1$ but the result was insignificant statistically. TGF-$\beta1$ increased the Sox9 mRNA expression to 180% compared to control group in IVD tissue. Sox9 and versican mRNA levels decreased by TWEAK were increased with TGF-$\beta1$ in primary cultured IVD cells, however, Fn14 did not show increasing effect on Sox9 and versican. Conclusion: This study suggests that TWEAK would act a role in intervertebral disc degeneration through decreasing sGAG and the mRNA level of versican and Sox9.

• Tuberculosis and Respiratory Diseases
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• v.69 no.5
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• pp.337-347
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• 2010

Background: Transglutaminase-2 (TG-2) has been reported to play an important role in the process of fibrosis. However, TG-2 studies on fibroproliferation of acute lung injury (ALI) are absent. The purpose of this study was to investigate the role of TG-2 in the fibroproliferation of lipopolysaccharide (LPS)-induced ALI. Methods: The male C57BL/6 mice of 5 weeks age were divided into 3 groups; control group (n=30) in which $50{\mu}L$ of saline was given intratracheally (IT), LPS group (n=30) in which LPS 0.5 mg/kg/$50{\mu}L$ of saline was given IT, and LPS+Cyst group treated with intraperitoneal 200 mg/kg of cystamine, competitive inhibitor of TG-2, after induction of ALI by LPS. TG-2 activity and nuclear factor $(NF)-{\kappa}B$ were measured in lung tissue homogenate. Tumor necrosis factor (TNF)-${\alpha}$, interleukin (IL)-$1{\beta}$, IL-6, myeloperoxidase (MPO), and transforming growth factor (TGF)-${\beta}1$ were measured using bronchoalveolar lavage fluids. Histopathologic ALI score and Mallory's phosphotunistic acid hematoxylin (PTAH) for collagen and fibronectin deposition were performed. Results: The TG-2 activities in the LPS group were significantly higher than the control and LPS+Cyst groups (p<0.05). The TNF-${\alpha}$ and IL-$1{\beta}$ concentrations and $NF-{\kappa}B$ activity were lower in the LPS+Cyst group than the LPS group (p<0.05). The LPS+Cyst group showed lower MPO, ALI score, TGF-${\beta}1$ concentration, and Mallory's PTAH stain than the LPS group, but the differences were not significant (p>0.05). Conclusion: Inhibition of TG-2 activity in the LPS-induced ALI prevented early inflammatory parameters, but had limited effects on late ALI and fibroproliferative parameters.

• The Journal of Korean Medicine
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• v.30 no.6
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• pp.35-43
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• 2009

Objectives: Hwangryunhaedok-tang (HRHDT), a prescription composed of four herbs, has been wi dely used in Oriental Medicine for the treatment of cerebral infarction. However, the mechanisms by which the herbal formula affects on the production of pro- and anti-inflammatory cytokines in cerebral infarction patients remain unknown yet. Methods: The levels of pro- and anti-inflammatory cytokines, including tumor necrosis factor (TNF)-a, interleukin (IL)-1b, and IL-6, IL-10, and TGF-${\beta}1$ were determined in peripheral blood mononuclear cells (PBMCs) from cerebral infarction patients under our experimental conditions. Results: The secretory levels of pro- and anti-inflammatory cytokines, including tumor necrosis factor (TNF)-a, interleukin (IL)-1b, and IL-6, and IL-10 were significantly increased in phytohemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMCs) from cerebral infarction patients. However, pretreatment with HRHDT significantly inhibited the secretion of pro- and anti-inflammatory in PBMCs. Also, HRHDT induced a significant increase of transforming growth factor (TGF)-b1 in PBMCs. Conclusions: These data indicate that HRHDT may be beneficial in the suppression of inflammatory processes of cerebral infarct through suppression of TNF-$\alpha$, IL-$1{\beta}$, IL-6, and IL-10 and induction of TGF-${\beta}1$.

• The Korean Journal of Pharmacology
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• v.26 no.2
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• pp.193-207
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• 1990

Transforming growth factor ${\beta}(TGF-{\beta})$ is a multifunctional polypeptide with diverse effects on the proliferation, differentiation and other functions in many cell types. $TGF-{\beta}$ is highly abundant in bone matrix and induces divergent responses in many aspects of bone cell metabolism . Several lines of investigation indicate that matrix-associated $TGF-{\beta}$ is the products of bone cells themselves. However, exact bone cell type reponsible for the production of $TGF-{\beta}$ is still in controversy, The present study was undertaken to determine the cellular origin of matrix-associated $TGF-{\beta}$ and to assess how different bone cells respond to $TGF-{\beta}$. As a prerequisite for this, 5 bone cell populations of distinct phenotype were isolated from fetal calvaria with sequential enzyme digestion protocol and biochemical characterization. Calvarial cell populations released in early stage showed fibroblastic features whereas populations relesed later was enriched with osteoblast-like cell as judged by their acid and alkaline phosphatase activities, cAMP responsiveness to parathyroid hormone, calcitonin and prostaglandin $E_2$ and collagen synthesis rate. By polyacylamide gel and immunoblot analysis of bone and calvarial cell extracts, presence of $TGF-{\beta}$ in bone tissues and production of $TGF-{\beta}$ by bone cells were confirmed again. Subsequent analysis of calvarial cell extracts prepared as individual population revealed that all calvarial cell populations synthesize $TGF-{\beta}$. Exogenously added $TGF-{\beta}$ induced biphasic response upon bone cell proliferation under serum-free condition. In osteoblastic cell populations, it was stimulatory whereas inhibitory in fibroblastic cell populations. In contrast, collagen and noncollagen protein synthesis of all calvarial cell populations were stimulated by $TGF-{\beta}$. Enhancement of protein synthesis was found to be more general rather than specific for collagen synthesis. In addition, effects of $TGF-{\beta}$ on protein synthesis were independent to its effects on cell proliferation. In summary, production of $TGF-{\beta}$ by bone cells and differential actions on various cell populations observed in this study suggest that $TGF-{\beta}$ may play an important role in the regulation of bone metabolism by modulating the specific cellular functions in autocrine and paracrine fashion.

• Molecules and Cells
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• v.44 no.1
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• pp.38-49
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• 2021

Airway mucus secretion is an essential innate immune response for host protection. However, overproduction and hypersecretion of mucus, mainly composed of the gel-forming MUC5AC protein, are significant risk factors for patients with asthma and chronic obstructive pulmonary disease (COPD). The transforming growth factor β (TGFβ) signaling pathway negatively regulates MUC5AC expression; however, the underlying molecular mechanism is not fully understood. Here, we showed that TGFβ significantly reduces the expression of MUC5AC mRNA and its protein in NCI-H292 cells, a human mucoepidermoid carcinoma cell line. This reduced MUC5AC expression was restored by a TGFβ receptor inhibitor (SB431542), but not by the inhibition of NF-κB (BAY11-7082 or Triptolide) or PI3K (LY294002) activities. TGFβ-activated Smad3 dose-dependently bound to MUC5AC promoter. Notably, TGFβ-activated Smad3 recruited HDAC2 and facilitated nuclear translocation of HDAC2, thereby inducing the deacetylation of NF-κB at K310, which is essential for a reduction in NF-κB transcriptional activity. Both TGFβ-induced nuclear translocation of Smad3/HDAC2 and deacetylation of NF-κB at K310 were suppressed by a Smad3 inhibitor (SIS3). These results suggest that the TGFβ-activated Smad3/HDAC2 complex is an essential negative regulator for MUC5AC expression and an epigenetic regulator for NF-κB acetylation. Therefore, these results collectively suggest that modulation of the TGFβ1/Smad3/HDAC2/NF-κB pathway axis can be a promising way to improve lung function as a treatment strategy for asthma and COPD.

• Microbiology and Biotechnology Letters
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• v.49 no.4
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• pp.543-551
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• 2021

Nocardiopsis species produce bioactive compounds, such as antimicrobial and anti-cancer agents and toxins. However, no reports have described their anti-inflammatory and anti-fibrotic effects during nasal polyp (NP) formation. In this study, we investigated whether marine-derived bacterial Nocardiopsis sp. 13G027 exerts anti-inflammatory and anti-fibrotic effects on lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages and transforming growth factor (TGF)-β1-induced NP-derived fibroblasts (NPDFs). Nitric oxide (NO) and prostaglandin E₂ (PGE₂) levels were analyzed. Extract from Nocardiopsis sp. 13G027 significantly inhibited the upregulation of NO and PGE₂ in LPS-activated RAW 264.7 macrophages. The expression of mitogen-activated protein kinases (MAPKs) and protein kinase B (Akt/PKB) in LPS-induced RAW 264.7 macrophages was evaluated; smooth muscle alpha-actin (α-SMA), collagen type I (Col-1), and fibronectin also phosphorylated small mothers against decapentaplegic (SMAD) 2 and 3 in TGF-β1-stimulated NPDFs. The Nocardiopsis sp. 13G027 extract suppressed the phosphorylation of MAPKs and Akt and the DNA-binding activity of activator protein 1 (AP-1). The expression of pro-fibrotic components such as α-SMA, Col-1, fibronectin, and SMAD2/3 was inhibited in TGF-β1-exposed NPDFs. These findings suggest that Nocardiopsis sp. 13G027 has the potential to treat inflammatory disorders, such as NP formation.