• Title/Summary/Keyword: quantitation of mRNA by RT-PCR

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Interleukin-8 gene expression in the human colon epithelial cell line, HT-29, exposed to Entamoeba histolytica (이질아메바에 의한 인체 대장상피세포주 HT-29에서의 interleukin-8 유전자의 발현)

  • 김정목;정현채
    • Parasites, Hosts and Diseases
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    • v.33 no.4
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    • pp.357-364
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    • 1995
  • The protozoan parasite, Entcmoeba histoIWticc, is one of major causative agents of intestinal disease all over the world. In acute experimental infection, the early host response to 5. histoIHtica is characterized by an infiltration of neutrophils. However, the chemotactic signal for this response is not well known. Based on the (jading that human epithelial cells produce the potent neutrophil chemoattractant and activator, interleukin-8 (IL-8), IL-8 gene expression was examined thoroughly in human colon epithelial cells exposed to 5. histolvtica trophozoites. Cellular RNAs were extracted from HT-29 or Caco-2 human colon epithelial cells exposed to 5. histoLvtica trophozoites for 30 minutes, 1 and 3 hours. IL-8 mRNA transcripts were measured by reverse transcriptional polprnerase chain reaction (RT-PCR) using synthetic standard RNA. The number of IL-8 mRNA molecules increased from 30 minutes to 3 hours of exposure period, reaching 3.1 H 107 molecules/ug of total RNA. Expression pattern of IL-8 mRNA transcripts was parallel to the amounts of IL-8 protein measured by enzyme-linked immunosorbent assay (ELISA) . Lysates of 5. histoIVtica also induced expression of mRNA for IL-8 in colon epithelial cells. These results sugf:esc that acute inflammatory reaction by 5. histoIVticc may be initially triggered by proinflammatory cytokines such as IL-8 secreted from epithelial cells of the colon.

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Blood Glucose Lowering Activity and Mechanism of Sangbackpitang (SBPT) in db/db Mouse (db/db 마우스에서 상백피탕의 혈당강하 활성 및 기전연구)

  • 이성현;안세영;두호경;정성현
    • YAKHAK HOEJI
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    • v.43 no.6
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    • pp.818-826
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    • 1999
  • Antidiabetic activity and mechanism of Sangbackpitang (SBPT) was examined in db/db mice, which is a spontaneously hyperglycemic, hyperinsulinemic and obese animal model. SBPT and acarbose were administered orally for 4 weeks. Fasting and non-fasting serum glucose, glycated hemoglobin and triglyceride were all reduced when compared between db/db control group and SBPT treated group. At 12th week after birth, SBPT increased an insulin secretion although statistic significance was not seen. Total activities of sucrase, maltase and lactase in SBPT treated group were all decreased when compared to db/db control. On the other hand, sucrase and maltase activities in acarbose treated groups were increased. Effect of SBPT on mRNA expression of glucose transporter(GLUT-4) was also examined. Quantitation of glucose transporter was performed by RT-PCR and in vitro transcription with co-amplification of rat-action gene as an internal standard. Muscular GLUT-4 mRNA expression in SBPT treated group was increased significantly. These results may suggest that SBPT lowered blood glucose ascribing to inhibition of glycosidase-catalyzed reaction and upregulation of muscular GLUT-4 mRNA expression.

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Anti-hyperlipidemic Effects of Scutellariae Radix, Aucklandiae Radix and Bupleuri Radix (SAB) extract in FL83B cells (지방간 유도 세포모델에서 황금(黃芩), 목향(木香), 시호(柴胡) 복합 추출물의 이상지질혈증에 대한 효과)

  • Do, Hyun Ju;Kim, Kyungho;Oh, Tae Woo
    • The Korea Journal of Herbology
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    • v.35 no.5
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    • pp.23-31
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    • 2020
  • Objectives : This study was conducted to evaluate the anti-hyperlipidemia effect of Scutellariae Radix, Aucklandiae Radix and Bupleuri Radix(SAB). Methods : FL83B cells were mouse liver hepatocytes, and we used this cell line. FL83B cells were treated with 0.5 mM oleic acid(OA) for 24 h, SAB extract was treated. After OA treatment, intracellular triglyceride (TG) and free fatty acid contents were measured with AdiopoRed™ assay and Free Fatty Acid Quantitation assay kit, respectively. Further, we evaluated several lipogenesis and metabolic markers such as sterol regulatory element-binding transcription factor-1c (SREBP-1c), acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), 3-hydroxy3-methyl-glutaryl CoA reductase (HMGCR), hormone-sensitive lipase (HSL), carnitine palmitoyltransferase (CPT-1), peroxisome proliferator activated receptor alpha (PPARα), and cluster of differentiation (CD36) using RT-PCR and Western-blot analysis. Results : OA markedly increased intracellular TG and free fatty acid, which plays a key role in reducing hepatic lipid accumulation, in FL83B cells. These increases were alleviated by SAB extract. The mRNA and protein expression of Fatty acid(FA) oxidation factors (CPT-1, PPARα), lipolysis factor(HSL), FA transporter(CD36), cholesterol synthesis factors (HMGCoA) and Lipodenesis (SREBP-1c, FAS, and ACC-1) were significantly increased by treatment of SAB extract in the OA-induced fatty liver cell model. Conclusions : In summary, the treat of SAB extract showed a significant reduction of the influx of fatty acids into hepatocytes, promoted the oxidation of fatty acids, and regulated fat synthesis-related factors, thereby regulating the accumulation of TG and free fatty acids.