• 동의생리병리학회지
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• 제20권2호
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• pp.383-387
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• 2006

To investigate the effect of GyeongshinhaeGihwan 1(GGT1) frequently used as an anti-obesity herbal medicine in oriental medicine on the expression of obesity-related genes, we measured the changes in mRNA levels of these genes by GGT1 in human growth hormone transgenic (hGHTg) obese female rats, and these effects by GGT1 were compared with those of reductil (RD), an anti-obesity drug approved by FDA. Rats received once daily oral administrations of autoclaved water, RD, or GGT1 for 8 weeks. At the end of study, rats were sacrificed and tissues were harvested. Total RNA from adipose tissue, liver and kidney was prepared and the mRNA levels for LPL (lipoprotein lipase), $PPAR{\gamma}$ (peroxisome proliferator activated receptor-gamma), $PPAR{\delta}$ (peroxisome proliferator activated receptor-delta), leptin, $TNF{\alpha}$ (tumor necrosis factor-alpha), and internal standard G3PDH (glyceraldehyde-3-phosphate dehydrogenase) were analyzed by RT-PCR. Compared with control group, $PPAR{\gamma}$ mRNA levels of liver and kidney were decreased in both RD and GGT1 groups, and the effects were more prominent in GGT1 group than in RD group, suggesting that GGT1 is effective in the inhibition of lipid storage by decreasing the $PPAR{\gamma}$ expression. $PPAR{\delta}$ mRNA levels of adipose tissue were increased by RD and GGT1 compared with DW, and the magnitude of increase were higher in GGT1 group than in RD group, indicating that GGT1 stimulates fatty acid oxidation and energy metabolism by activating $PPAR{\delta}$ expression. GGT1 group had higher concentrations of serum leptin, a well-known inhibitor of appetite, than control and RD groups. However, The mRNA levels of leptin, LPL, and $TNF{\alpha}$ were not changed by GGT1. These results indicate that GGT1 can prevent obesity in hGHTg obese female rats by down-regulating and up-regulating the mRNA expression of $PPAR{\gamma}$ and $PPAR{\delta}$, respectively, and that this anti-obesity effects were more pronounced in GGT1 group compared with RD group. In addition, GGT1 seems to inhibit obesity by increasing the circulating leptin levels.

• Asian Pacific Journal of Cancer Prevention
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• 제15권8호
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• pp.3451-3455
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• 2014

Background: The serum carcinoembryonic antigen (CEA) level can reflect tumor growth, recurrence and metastasis. It has been reported that epidermal growth factor receptor (EGFR) mutations in exons 19 and 21may have an important relationship with tumor cell sensitivity to EGFR-TKI therapy. In this study, we investigated the clinical value of EGFR mutations and serum CEA in patients with non-small cell lung cancer (NSCLC). Materials and Methods: The presence of mutations in EGFR exons 19 and 21 in the tissue samples of 315 patients with NSCLC was detected with real-time fluorescent PCR technology, while the serum CEA level in cases who had not yet undergone surgery, radiotherapy, chemotherapy and targeted therapy were assessed by electrochemical luminescence. Results: The mutation rates in EGFR exons 19 and 21 were 23.2% and 14.9%, respectively, with the two combined in 3.81%. Measured prior to the start of surgery, radiotherapy, chemotherapy and targeted treatment, serum CEA levels were abnormally high in 54.3% of the patients. In those with a serum CEA level <5 ng/mL, the EGFR mutation rate was 18.8%, while with 5~19 ng/mL and ${\geq}20ng/mL$, the rates were 36.4% and 62.5%. In addition, in the cohort of patients with the CEA level being 20~49 ng/mL, the EGFR mutation rate was 85.7%, while in those with the CEA level ${\geq}50ng/mL$, the EGFR mutation rate was only 20.0%, approximately the same as in cases with the CEA level<5 ng/mL. Conclusions: There is a positive correlation between serum CEA expression level and EGFR mutation status in NSCLC patients, namely the EGFR mutation-positive rate increases as the serum CEA expression level rises within a certain range (${\geq}20ng/mL$, especially 20~49 ng/mL). If patient samples are not suitable for EGFR mutation testing, or cannot be obtained at all, testing serum CEA levels might be a simple and easy screening method. Hence, for the NSCLC patients with high serum CEA level (${\geq}20ng/mL$, especially 20~49 ng/mL), it is worthy of attempting EGFR-TKI treatment, which may achieve better clinical efficacy and quality of life.

• Asian Pacific Journal of Cancer Prevention
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• 제17권4호
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• pp.2019-2025
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• 2016

Hepatocellular carcinoma (HCC) is major health problem with high mortality rates, especially in patients with hepatitis B virus (HBV) infection. Telomerase function is one of common mechanisms affecting genome stability and cancer development. Recent studies demonstrated that genetic polymorphisms of telomerase associated genes such as telomerase associated protein 1 (TEP1) rs1713449 and PIN2/TERF1-interacting telomerase inhibitor 1 (PINX1) rs1469557 may be associated with risk of HCC and other cancers. In this study, 325 patients with HCC and 539 non-HCC groups [193 healthy controls, 80 patients with HBV-related liver cirrhosis (LC) and 266 patients with HBV-related chronic hepatitis (CH)] were enrolled to explore genetic polymorphisms of both SNPs using the allelic discrimination method based on MGB probe TaqMan real time PCR. We demonstrated that all genotypes of both genes were in Hardy-Wienberg equilibrium (P>0.05). Moreover, there was no significant association between rs1713449 genotypes and HCC risk, HCC progression and overall survival (P>0.05). Interestingly, we observed positive association of rs1469557 with risk of HCC when compared with the LC group under dominant (CC versus CT+TT, OR=1.89, 95% CI= 1.06-3.40, P=0.031) and allelic (C versus T alleles, OR=1.75, 95% CI=1.04-2.94, P=0.033) models, respectively. Moreover, overall survival of HCC patients with CC genotype of rs1469557 was significantly higher than non-CC genotype (Log-rank P=0.015). These findings suggest that PINX1 rs1469557 but not TEP1 rs1469557 might play a role in HCC progression in Thai patients with LC and be used as the prognosis marker to predict overall survival in HCC patients.

• 대한한의학방제학회지
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• 제25권1호
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• pp.1-10
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• 2017

Objectives : The fruit of Prunus mume Siebold & Zucc. has been used as an alternative medicine and functional food in Korea and Japan for preventive and therapeutic purposes. However, its molecular actions and mechanism on anti-inflammatory activity have not been clearly investigated. The aim of this study was to clarify the anti-inflammatory activity of the ethanol extract of P. mume fruit (EEPM) in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells, and sought to understand the associated molecular mechanisms. Methods : Cytotoxicity was assessed by an MTT assay. The amount of nitric oxide (NO) production was determined by nitrite assay. The mRNA expression of inducible nitric oxide synthase (iNOS) was analyzed by RT-PCR. In addition, expression levels of iNOS, nuclear factor-erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) protein were detected by Western blotting. Results : Our data indicated that EEPM inhibited NO production in LPS-stimulated RAW264.7 cells in a concentration-dependent manner. At the mRNA and protein levels, EEPM suppressed LPS-induced iNOS expression. On the other hand, EEPM markedly enhanced HO-1 expression, which was associated with an induction and nuclear translocation of Nrf2. Moreover, the inhibitory effect of EEPM against LPS‑induced NO production was significantly enhanced by hemin, a HO-1 inducer; however, EEPM's effect on the production of NO was abolished by zinc protoporphyrin IX, a HO-1 inhibitor. Conclusion : The results suggest that EEPM can act as a suppressor agent on NO production through an activation of Nrf2/HO-1 signaling pathway, and may be a promising candidate for the treatment of inflammatory diseases.

• 대한한방내과학회지
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• 제29권1호
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• pp.130-148
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• 2008

Objective : This study was designed to investigate the effect of the water extract of Gamisamgibopae-tang(GMSGBPT), an oriental herbal formulation, on the growth of NCI-H460 and A549 human non-small-cell lung cancer cell lines. Methods : Cytotoxicity and cell morphology were evaluated by MTT assay and inverted microscope, respectively. Apoptosis was detected using agarose gel electrophoresis and flow cytometer. The expression levels of mRNAs and proteins of target genes were determined by RT-PCR and western blot analyses, respectively Result and Conclusion : We found that exposure of A549 cells to GMSGBPT resulted in the growth inhibition in a dose-dependent manner as measured by MTT assay, but GMSGBPTdid not affect the growth of NCI-H460 cells. The anti-proliferative effect of GMSGBPT treatment in A549 cells was associated with morphological changes, formation of apoptotic bodies and DNA fragmentation, and flow cytometry analysis confirmed that GMSGBPT treatment increased the populations of apoptotic-sub G1 phase. Growth inhibition and apoptotic cell death by GMSGBPT were connected with a up-regulation of cyclin-dependent kinase inhibitor p21 (WAF1/CIP1) mRNA and protein in a tumor suppressor p53-independent fashion. However GMSGBPT treatment did not affect other growth regulation-related genes such as early growth response-1 (Egr-1), nonsteroidal anti-inflammatory drug (NSAID)-activated gene-1 (NAG-1), inducible nitric oxide synthase (iNOS), cyclooxygenases (COXs), telomere-regulatory factors in A549 orNCI-H460 cells. Taken together, these findings partially provide novel insights into the possible molecular mechanism of the anti-cancer activity of GMSGBPT.

• Molecular & Cellular Toxicology
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• 제4권4호
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• pp.278-284
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• 2008

The X-linked inhibitor of apoptosis (XIAP) is a member of a novel family of inhibitors of apoptosis and has several BIR domains (BIR1, BIR2, and BIR3) and a carboxy-terminal RING zinc-finger. Since suppressionof apoptosis is fundamentally important for carcinogenesis and tumor growth, we investigated the expression and function of XIAP in DEN-induced carcinogenesis using rat model. Wistar rats were injected intraperitoneally with DEN at a dose of 50 mg/kg in twice a week for 12 weeks (Group II) and 16 weeks (Group III) followed by the recovery periods, respectively. The evaluation of DEN-induced carcinogenesis carried out the blood, RT-PCR, histopathological and western blot analysis. The level of blood chemistry including GOT/GPT, albumin, and total bilirubin were significantly exchanged comparing to control and Group I/Group II. The expression of albumin and collagen mRNA were significantly exchanged (P<0.05) in both groups. In addition, AFP mRNA expression decreased more after recovery periods than Group II. XIAP was expressed constitutively in normal rat liver as well as DEN-induced Groups I and Group II. In addition, XIAP expression increased more in Group I with 4 weeks recovery periods than Group I. However, XIAP expression shown to increase in Group lI, otherwise, it was decreased in Group II with 10 weeks repair periods. Taken together, these results suggest the alteration of XIAP expression could be involved in hepatocellular carcinogenesis.

• 한국수정란이식학회지
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• 제25권2호
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• pp.103-110
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• 2010

The first morphogenetic event of preimplantation development, compaction, was required efficient production of porcine embryos in vitro. Compaction of the porcine embryo, which takes place at post 4-cell stage, is dependent upon the adhesion molecule E-cadherin. The E-cadherin through ${\beta}$-catenin contributes to stable cell-cell adhesion. Rho-associated kinase (ROCK) signaling was found to support the integrity of E-cadherin based cell contacts. In this study, we traced the effects of ROCK-1 on early embryonic development and structural integrity of blastocysts in pigs. Then, in order to gain new insights into the process of compaction, we also examined whether ROCK-1 signaling is involved in the regulation of the compaction mediated by E-cadherin of cellular adhesion molecules. As a result, real-time RT-PCR analysis showed that the expression of ROCK-1 mRNA was presented throughout porcine preimplantation stages, but not expressed as consistent levels. Thus, we investigated the blastocyst formation of porcine embryos treated with LPA and Y27632. Blastocysts formation and their qualities in LPA treated group increased significantly compared to those in the Y27632-treated group (p < 0.05). Then, to determine whether ROCK-1 associates embryonic compaction, we explored the effect of activator and/or inhibitor of ROCK-1 on compaction of embryos in pigs. The rate of compacted morula in LPA treated group was increased compared to that in the Y27632-treated group (39.7 vs 12.0%). Furthermore, we investigated the localization and expression pattern of E-cadherin at 4-cell stage porcine embryos in both LPA- and Y27632-treated groups by immunocytochemical analysis and Western blot analysis. The expression of E-cadherin was increased in LPA-treated group compared to that in the Y27632-treated group. The localization of E-cadherin in LPA-treated group was enriched in part of blastomere contacts compared to that Y27632-treated group. ROCK-1 as a crucial mediator of embryo compaction may plays an important role in regulating compaction through E-cadherin of the cell adhesion during the porcine preimplantation embryo. We concluded that ROCK-1 gene may affect the developmental potential of porcine blastocysts through regulating embryonic compaction.

• 생명과학회지
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• 제16권6호
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• pp.964-971
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• 2006

기질의 침윤과 전이를 특징으로 하는 악성종양 세포는 세포외 기질이나 기저막에 의존적으로 작용한다. 세포외 기질을 분해하는 효소인 matrix metalloproteinase (MMP) 계들의 발현 및 활성증가는 대부분의 악성종양세포에서 전이와 침윤을 촉진시킨다. MMP family 가운데 특히 type IV collagenase 활성을 지닌 MMP-2와 MMP-9은 세포외기질의 중요한 구성분인 collagen, fibronectin을 분해하는 특성을 가지며 암 전이를 용이하게 하는 주요한 효소로 잘 알려져 있다. 본 연구에서는 항암후보물질인 disulfiram이 골 육종 (U20S), 신장암 (Caki-1) 및 자궁암 (Caski) 세포에서 MMP-2와 MMP-9의 효소활성 및 발현억제에 대해 조사하였다. MTT assay를 이용하여 disulfiram에 대한 암세포 viability 실험에서는 disulfiram이 암세포의 viability를 저해하였다. 또한 zymography, western blot 및 RT-PCR 등을 이용한 type IV collagenase의 활성 및 발현 실험에서 disulfiram은 type IV collagenase의 활성을 비롯하여 단백질 및 mRNA 발현을 억제시키는 것을 확인하였다. 따라서 disulfiram이 MMP-2와 MMP-9의 활성 및 발현 억제 기전을 통하여 골 육종, 신장암 및 자궁경부암 세포의 작용을 억제한다는 연구 결과는 disulfiram이 각종 악성종양의 침윤과 전이를 억제 또는 방지하기 위한 치료물질로서 임상에서 활용할 수 있는 가능성을 보여준다.

• 대한본초학회지
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• 제31권5호
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• pp.71-78
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• 2016

Objectives : Diabetic nephropathy is one of the most common chronic complications of diabetes and a leading cause of end-stage renal failure in the world. Mesangial cell proliferation is known as the major pathologic features such as glomerulosclerosis and renal fibrosis. Wiryeongtang (WRT) is a well-known traditional herbal formula as therapeutic agents for chronic edema and dysuresia of renal homeostasis. In the present study, we investigated whether WRT inhibits high glucose (HG)-induced renal dysfunction by TGF-β/Smads signal regulation in cultured mesangial cells.Methods : Inhibitory effect of WRT (10-50 ㎍/ml) on HG-stimulated mesangial cells proliferation and dysfunction were evaluated by [³H]-thymidine incorporation, Western blot, and RT-qPCR.Results : WRT significantly decreased HG-accelerated thymidine incorporation in human renal mesangial cell in a dose-dependent levels. WRT induced down-regulation of cyclins/CDKs and up-regulation of CDK inhibitor, p21waf1/cip1 and p27kip1 expression. In addition, HG enhanced expression of dysfunction biomarker such as collagen IV and CTGF, which was markedly attenuated by WRT. WRT decreased TGF-β1 and Smad-2/Smad-4 expression, whereas increased Smad-7 expression under HG. Furthermore, WRT inhibited HG-induced inflammatory factors level such as ICAM-1 and MCP-1 as well as NF-κB p65 nuclear translocation and intracellular ROS production.Conclusions : These results suggested that WRT may alleviate mesangial proliferation and inflammation possibly involved in renal fibrotic process, further diabetic nephropathy through disturbing TGF-β1/Smad signaling and NF-κB/ROS pathway. Thus, WRT might prove to be effective in the treatment of renal dysfunction leading to diabetic nephropathy.

• International Journal of Oral Biology
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• 제34권1호
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• pp.7-14
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• 2009

Nitric oxide (NO) acts as an intracellular messenger at the physiological level but can be cytotoxic at high concentrations. The cells within periodontal tissues, such as gingival and periodontal fibroblasts, contain nitric oxide syntheses and produce high concentrations of NO when exposed to bacterial lipopolysaccharides and cytokines. However, the cellular mechanisms underlying NO-induced cytotoxicity in periodontal tissues are unclear at present. In our current study, we examined the NO-induced cytotoxic mechanisms in human gingival fibroblasts (HGF). Cell viability and the levels of reactive oxygen species (ROS) were determined using a MTT assay and a fluorescent spectrometer, respectively. The morphological changes in the cells were examined by Diff-Quick staining. Expression of the Bcl-2 family and Fas was determined by RT-PCR or western blotting. The activity of caspase-3, -8 and -9 was assessed using a spectrophotometer. Sodium nitroprusside (SNP), a NO donor, decreased the cell viability of the HGF cells in a dose- and time-dependent manner. SNP enhanced the production of ROS, which was ameliorated by NAC, a free radical scavenger. ODQ, a soluble guanylate cyclase inhibitor, did not block the SNP-induced decrease in cell viability. SNP also caused apoptotic morphological changes, including cell shrinkage, chromatin condensation, and DNA fragmentation. The expression of Bax, a member of the proapoptotic Bcl-2 family, was upregulated in the SNP-treated HGF cells, whereas the expression of Bcl-2, a member of the anti-apoptotic Bcl-2 family, was downregulated. SNP augmented the release of cytochrome c from the mitochondria into the cytosol and enhanced the activity of caspase-8, -9, and -3. SNP also upregulated Fas, a component of the death receptor assembly. These results suggest that NO induces apoptosis in human gingival fibroblast via ROS and the Bcl-2 family through both mitochondrial- and death receptor-mediated pathways. Our data also indicate that the cyclic GMP pathway is not involved in NO-induced apoptosis.