• Animal cells and systems
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• 제14권4호
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• pp.275-282
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• 2010

Chunghyul-dan (CHD) is a combinatorial drug known to exert anti-inflammatory effects in endothelial cells. In this study, we employed global transcriptional profiling using cDNA microarrays to identify molecular mechanisms responsible for the anti-inflammatory activity of CHD in endothelial cells. An analysis of the microarray data revealed that transcript levels of monocyte chemotactic protein-1 (MCP-1), vascular cell-adhesion molecule-1 (VCAM-1) and activated leukocyte cell-adhesion molecule were dramatically altered in CHD-treated endothelial cells. These changes in gene expression were confirmed by RT-PCR, Western blotting and ELISA. Chronic CHD treatment also appeared to decrease MCP-1 secretion, probably as a result of decreased MCP-1 expression. In addition, we determined that chronic CHD treatment inhibited lipopolysaccharide-stimulated adhesion of THP-1 leukocytes to endothelial cells. The inhibitory effect of CHD on LPS-stimulated adhesion resulted from downregulation of VCAM-1 expression. Transmigration of THP-1 leukocytes through endothelial cells was also inhibited by chronic CHD treatment. In conclusion, CHD controls a variety of inflammatory activities by regulating MCP-1 and VCAM-1 gene expression.

• The Korean Journal of Physiology
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• 제24권2호
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• pp.363-375
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• 1990

The present investigation was performed to purify bombesin-like immunoreactivity (BBS-LI) from the skin of frogs, B. orientalis inhabiting Korea. For extraction of BBS-LI, the fresh skin of 360 g from frogs was immersed in 1,800 ml of 100% methanol and then kept at $4^{\circ}C$ for 5 days. BBS-LI was partially purified by liquid chromatography using an alkaline alumina column followed by a Sephadex G-10 column. BBS-LI was further purified by using sequential HPLC of reversed phase C18 preparation, gel permeation, SP-ion exchange and reversed phase C18 analysis. BBS-LI in fractions of each step was monitored by radioimmunoassay for which bombesin antiserum with a titer of 1 : 188,800 was raised in a guinea pig. Eventually, two different BBS-LI were successfully purified and each BBS-LI showed the following character. 1) BBS-LI was well separated into two peaks in SP-ion exchange HPLC. One (BBS-LI-K1) bound to the column while the other (BBS-LI-K2) did not. 2) BBS-LI-K1, 73.8% of total BBS-LI, was not differentiated from synthetic bombesin in reversed phase C18 analytical and gel permeation HPLC. 3) BBS-LI-K2, 26.2% of total BBS-LI, eluted later than synthetic bombesin in reversed phase C18 analytical HPLC, but it eluted with a retention time identical to that of synthetic bombesin in gel permeation HPLC. 4) The two forms of BBS-LI and synthetic bombesin identically stimulated gastrin release and pancreatic exocrine secretion including volume, protein output and amylase output in anesthetized rats. It is concluded from the above results that the skin of B. orientalis contains two different forms of BBS-LI which are very identical to bombesin immunologically and biologically. In comparison with synthetic bombesin containing 14 amino acid residues, the major form shows quite similar pattern in all HPLC used in the present study, but the minor form exhibits quite different pattern in SP-ion exchange and reversed phase C18 analytical HPLG.

• 환경생물
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• 제27권1호
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• pp.88-94
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• 2009

안토시아닌(Anthocyanin)은 플라보노이드계 화합물의 한 부류로 항산화, 항암 및 항궤양, 항당뇨, 중금속해독, 시력보호, 콜레스테를 저하 등의 다양한 생리활성을 가지는 것으로 보고되어 있다. 죽상경화과정은 염증성 사이토카인의 분비 또는 혈관손상으로 인한 백혈구의 부착과 이동을 통해 시작된다. 본 연구는 이러한 죽상경화의 초기과정에서 안토시아닌 혼합물 중 single compound인 delphinidin chloride (DC) 인간혈관 내피세포주(HUVEC, human umbilical vein endothelial cell line)에서 백혈구 부착과 관련이 있는 ICAM-1 (Intraceliular Adhesion Molecule-1)과 VCAM-1 (Vascular Adhesion Molecule-1) 발현에 미치는 영향에 대해 조사하였다. 세포독성이 없는 농도에서 TNF-$\alpha$에 의해 유도된 혈관 내피세포에 대한 단핵구의 부착정도를 측정하기 위해 monocyte-endothelial cell adhesion assay와 광학현미경을 이용한 형태학적 관찰을 한 결과 DC가 처리농도 의존적으로 부착을 억제하였다. 내피세포로부터 TNF-$\alpha$에 의해 유도된 세포부착 분자인 VCAM-1과 ICAM-1의 발현에 대한 영향을 western blot analysis 및 RT-PCR방법으로 비교 분석한 결과 VCAM-1과 ICAM-1의 단백질과 mRNA수준에서의 발현이 농도 의존적으로 감소되었다. 이러한 결과들을 종합해 볼 때 안토시아닌 중에서 DC를 실험한 결과 DE는 TNF-$\alpha$에 의해 유도된 내피세포의 ICAM-1과 VCAM-1 발현 억제효과를 확인할 수 있었다.

• Applied Biological Chemistry
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• 제40권5호
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• pp.380-387
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• 1997

Pectate lyase isoenzymes을 분비하는 Erwinia carotovorn subsp. carotovora LY34는 식물조직을 연화시키는 연부균이다. 이 균주로부터 게놈 DNA를 분리하여 Sau3Al 제한효소로 부분 절단한 다음 pBluescript $SK^+$ 벡터에 클로닝하여 pectate Iyase를 분비하는 클론을 분리하였다 분리 결과 4.2 kb크기의 DNA 단편을 가지고 있었으며 이를 다시 재클로닝하여 3.1 kb크기의 pelCI유전자를 함유하는 pLYPA100을 구하였다. 이 유전자의 DNA 염기서열을 분석한 결과 374 개의 아미노산을 구성하는 1,122 bp의 ORF를 확인하였다. 시작코돈과 종결코돈은 ATG와 TAA였으며 초기 서열 22개의 아미노산으로 구성된 전형적인 원핵세포의 signal peptide가 존재하였다. PeICI의 단백질 염기서열을 다른 단백질과 유사성을 분석한 결과 Erwinia carotovera subsp. carotovora Er 균주의 PelIII, Erwinia carotevora subsp. carotovora SCR193 균주의 PeIC 및 Erwinia caretovora subsp. atroseptica C18 균주의 Pel3과 유사하였으며 PLbc family에 속하였다. PeICI의 분자량은 40,507, pI는 7.60으로 계산되었다.

• Biomolecules & Therapeutics
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• 제25권3호
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• pp.279-287
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• 2017

The chemical property of cinnamaldehyde is unstable in vivo, although early experiments have shown its obvious therapeutic effects on viral myocarditis (VMC). To overcome this problem, we used cinnamaldehyde as a leading compound to synthesize derivatives. Five derivatives of cinnamaldehyde were synthesized: 4-methylcinnamaldehyde (1), 4-chlorocinnamaldehyde (2), 4-methoxycinnamaldehyde (3), ${\alpha}$-bromo-4-methylcinnamaldehyde (4), and ${\alpha}$-bromo-4-chlorocinnamaldehyde (5). Neonatal rat cardiomyocytes and HeLa cells infected by coxsackievirus B3 (CVB3) were used to evaluate their antiviral and cytotoxic effects. In vivo BALB/c mice were infected with CVB3 for establishing VMC models. Among the derivatives, compound 4 and 5 inhibited the CVB3 in HeLa cells with the half-maximal inhibitory concentrations values of $11.38{\pm}2.22{\mu}M$ and $2.12{\pm}0.37{\mu}M$, respectively. The 50% toxic concentrations of compound 4 and 5-treated cells were 39-fold and 87-fold higher than in the cinnamaldehyde group. Compound 4 and 5 effectively reduced the viral titers and cardiac pathological changes in a dose-dependent manner. In addition, compound 4 and 5 significantly inhibited the secretion, mRNA and protein expressions of inflammatory cytokines TNF-${\alpha}$, IL-$1{\beta}$ and IL-6 in CVB3-infected cardiomyocytes, indicating that brominated cinnamaldehyde not only improved the anti-vital activities for VMC, but also had potent anti-inflammatory effects in cardiomyocytes induced by CVB3.

• Molecules and Cells
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• 제43권1호
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• pp.66-75
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• 2020

Saturated fatty acids contribute to β-cell dysfunction in the onset of type 2 diabetes mellitus. Cellular responses to lipotoxicity include oxidative stress, endoplasmic reticulum (ER) stress, and blockage of autophagy. Palmitate induces ER Ca²⁺ depletion followed by notable store-operated Ca²⁺ entry. Subsequent elevation of cytosolic Ca²⁺ can activate undesirable signaling pathways culminating in cell death. Mitochondrial Ca²⁺ uniporter (MCU) is the major route for Ca²⁺ uptake into the matrix and couples metabolism with insulin secretion. However, it has been unclear whether mitochondrial Ca²⁺ uptake plays a protective role or contributes to lipotoxicity. Here, we observed palmitate upregulated MCU protein expression in a mouse clonal β-cell, MIN6, under normal glucose, but not high glucose medium. Palmitate elevated baseline cytosolic Ca²⁺ concentration ([Ca²⁺]_i) and reduced depolarization-triggered Ca²⁺ influx likely due to the inactivation of voltage-gated Ca²⁺ channels (VGCCs). Targeted reduction of MCU expression using RNA interference abolished mitochondrial superoxide production but exacerbated palmitate-induced [Ca²⁺]_i overload. Consequently, MCU knockdown aggravated blockage of autophagic degradation. In contrast, co-treatment with verapamil, a VGCC inhibitor, prevented palmitate-induced basal [Ca²⁺]_i elevation and defective [Ca²⁺]_i transients. Extracellular Ca²⁺ chelation as well as VGCC inhibitors effectively rescued autophagy defects and cytotoxicity. These observations suggest enhanced mitochondrial Ca²⁺ uptake via MCU upregulation is a mechanism by which pancreatic β-cells are able to alleviate cytosolic Ca²⁺ overload and its detrimental consequences.

• 한국식품과학회지
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• 제34권4호
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• pp.694-699
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• 2002

본 연구에서는 분만 후 5일 이내에 분비되는 젖소 초유의 whey 및 whey 분획이 Th1 cell의 증식에 어떤 영향을 미치는지, 그리고 Th1 cell 증식에 직접적으로 관여하는 whey 분획이 macrophage의 $TNF-{\alpha}$ 분비에 미치는 영향을 조사하였다. 초유 whey를 ultrafiltration으로 분자량별로 분획한 결과, 단백질 성분의 회수율은 Fr. I, II, III가 각각 72%, 17.7%, 10.2%였으며, 당 성분의 회수율은 Fr. I, II, III가 각각 22.1%, 7.4%, 70.1%였다. Fr. II를 재분획한 Fr. P의 단백질 회수율과 Fr. O의 당 회수율은 각각 86.9%, 88.8%였다. 각각의 whey 분획의 농도 대비 Th1 cell 증식 효과를 검증한 결과, 1 mg/mL 농도에서 Fr. II가 Th1 cell을 가장 많이 자극시켰으며, 세포증식율은 67.1%였으나, Fr. II의 단백질 및 올리고당 분획의 세포증식효과는 없는 것으로 나타났다. Whey의 각각의 분획을 이용하여 $TNF-{\alpha}$ 분비 능력을 조사한 결과, Fr. O가 양성대조구로 사용한 LPS보다 약 80% 이상의 $TNF-{\alpha}$분비 유도 능력이 있는 것으로 나타났다.

• Journal of Nutrition and Health
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• 제38권10호
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• pp.817-826
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• 2005

Recently it has been reported that vitamin A and retinol binding proteins (RBPs) in blood and urine were changed in the condition of diabetes mellitus or hyperlipidemia. Fruits and vegetables are recommended to consume for the people suffered from these chronic degenerative diseases. The main components of fruits and vegetables are dietary fibers, for example cellulose and pectin, of which function to affect the absorption and excretion of dietary fat and fat-soluble substances. This study was conducted to investigate the effect of dietary fibers on RBPs mRNA expression in liver, small intestine and serum of rat fed high fat diet during 4 weeks. Sprague-Dawley rats, weighing 121g on average, were divided into four groups; (Control; $17\%$ fat & cellulose supplement diet, HF0: $25\%$ fat & fiber free diet, B:.Uc: $25\%$ fat & cellulose supplement diet and HF0: $25\%$ fat & pectin supplement diet) . The rats fed high fat diet groups (HF0, HFC, HFP) tended to consume the food less than the control group, but FER of HF0 groups was significantly higher than the control (p < 0.05) . The weight of adrenal gland in high fat diet groups (HF0, HFC, HFP) was significantly less than the control. Total lipid in feces daily excreted and in liver did not show any significant differences among the groups. Total cholesterol in HFP group was significantly different from that of HFC group. Serum total cholesterol and triglyceride in other group tended to lower than other groups and HDL cholesterol higher. Consequently, AI (atherogenic index) was the lowest in HFP group. Vit A contents in feces daily excreted tended to lower in high fat diet groups (HF0, HFP) compared to the control group. That content in adrenal gland was the lowest in HF0 group, but not in liver. In HFP group were down-regulated cRBPI mRNA in liver and cRBPII mRNA in small intestine and up-regulated RBP and transthyretin expression in serum compared to the other groups. In conclusion, dietary fibers, especially pectin, in high fat diet might down-regulate the expression of CRBP I, CRBP II mRNA in liver and small intestine, but increase the secretion of RBP into serum and therefore inhance the bioavailability of Vit A through the body. (Korean J Nutrition 38(10): 817$\sim$826,2005)

• Experimental and Molecular Medicine
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• 제50권12호
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• pp.1.1-1.14
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• 2018

DNA methylation is a regulatory mechanism in epigenetics that is frequently altered during human carcinogenesis. To detect critical methylation events associated with gastric cancer (GC), we compared three DNA methylomes from gastric mucosa (GM), intestinal metaplasia (IM), and gastric tumor (GT) cells that were microscopically dissected from an intestinal-type early gastric cancer (EGC) using methylated DNA binding domain sequencing (MBD-seq) and reduced representation bisulfite sequencing (RRBS) analysis. In this study, we focused on differentially methylated promoters (DMPs) that could be directly associated with gene expression. We detected 2,761 and 677 DMPs between the GT and GM by MBD-seq and RRBS, respectively, and for a total of 3,035 DMPs. Then, 514 (17%) of all DMPs were detected in the IM genome, which is a precancer of GC, supporting that some DMPs might represent an early event in gastric carcinogenesis. A pathway analysis of all DMPs demonstrated that 59 G protein-coupled receptor (GPCR) genes linked to the hypermethylated DMPs were significantly enriched in a neuroactive ligand-receptor interaction pathway. Furthermore, among the 59 GPCRs, six GI hormone receptor genes (NPY1R, PPYR1, PTGDR, PTGER2, PTGER3, and SSTR2) that play an inhibitory role in the secretion of gastrin or gastric acid were selected and validated as potential biomarkers for the diagnosis or prognosis of GC patients in two cohorts. These data suggest that the loss of function of gastrointestinal (GI) hormone receptors by promoter methylation may lead to gastric carcinogenesis because gastrin and gastric acid have been known to play a role in cell differentiation and carcinogenesis in the GI tract.

• Experimental and Molecular Medicine
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• 제50권12호
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• pp.2.1-2.12
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• 2018

Glucagon-like peptide-1 (GLP-1) has a broad spectrum of biological activity by regulating metabolic processes via both the direct activation of the class B family of G protein-coupled receptors and indirect nonreceptor-mediated pathways. GLP-1 receptor (GLP-1R) agonists have significant therapeutic effects on non-alcoholic fatty liver disease (NAFLD) and steatohepatitis (NASH) in animal models. However, clinical studies indicated that GLP-1 treatment had little effect on hepatic steatosis in some NAFLD patients, suggesting that GLP-1 resistance may occur in these patients. It is well-known that the gut metabolite sodium butyrate (NaB) could promote GLP-1 secretion from intestinal L cells. However, it is unclear whether NaB improves hepatic GLP-1 responsiveness in NAFLD. In the current study, we showed that the serum GLP-1 levels of NAFLD patients were similar to those of normal controls, but hepatic GLP-1R expression was significantly downregulated in NAFLD patients. Similarly, in the NAFLD mouse model, mice fed with a high-fat diet showed reduced hepatic GLP-1R expression, which was reversed by NaB treatment and accompanied by markedly alleviated liver steatosis. In addition, NaB treatment also upregulated the hepatic p-AMPK/p-ACC and insulin receptor/insulin receptor substrate-1 expression levels. Furthermore, NaB-enhanced GLP-1R expression in HepG2 cells by inhibiting histone deacetylase-2 independent of GPR43/GPR109a. These results indicate that NaB is able to prevent the progression of NAFL to NASH via promoting hepatic GLP-1R expression. NaB is a GLP-1 sensitizer and represents a potential therapeutic adjuvant to prevent NAFL progression to NASH.