• Title/Summary/Keyword: QC-PCR

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A role of carboxy-terminal region of Toxoplasma gondii-heat shock protein 70 in enhancement of T. gondii infection in mice

  • Mun, Hye-Seong;Norose, Kazumi;Aosai, Fumie;Chen, Mei;Yano, Akihiko
    • Parasites, Hosts and Diseases
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    • v.38 no.2
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    • pp.107-110
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    • 2000
  • We investigated the role of recombinant Toxoplasma gondii heat shock protein (rT.g.HSP) 70-full length, rT.g. HSP70-NH2-terminal region, or rT.g. HSP70-carboxy-terminal region in prophylactic immunity in C57BL/6 mice perorally infected with Fukaya cysts of T. gondii. At 3, 4, 5, and 6 weeks after infection, the number of T gondii in the brain tissue of each mouse was measured by quantitative competitive-polymerase chain reaction (QC-PCR) targeting the surface antigen (SAG) 1 gene. Immunization with rT.g.HSP70-full length or rT.g.HSP70-carboxy-terminal region increased the number of T. gondii in the brain tissue after T. gondii infection, whereas immunization with rT.g.HSP70-NHa-terminal region did not. These results suggest that T.g. HSP70-carboxy-terminal region as well as T.g.HSP70-full length may induce deleterious effects on the protective immunity of mice infected with a cyst-forming T. gondii strain, Fukaya.

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Endometrium from Women with Endometriosis Expresses Increased Levels of Pleiotrophin (PTN) and Midkine (MK) mRNA Compared to Normal Endometrium (자궁내막증 환자와 정상 여성의 자궁내막에서 Pleiotrophin (PTN)과 Midkine (MK) mRNA 발현 차이에 관한 연구)

  • 정혜원;허성은;문혜성
    • Development and Reproduction
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    • v.4 no.1
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    • pp.101-108
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    • 2000
  • Objectives: The pleiotrophin (PTN) and midkine (MK) are secreted heparin-binding neurokines that share 50% sequence homology. PTN and MK are expressed in the range of primary human tumors. The association of PTN and MK with carcinogenesis, enhancement of plasminogen activator activity and angiogenic factor are reported. Patients with endometriosis are characterized by the ability of the endometrium to implant; angiogenic and growth factors may play a significant role in the pathogenesis of endometriosis. To test the hypothesis that higher expression of PTN and MK in endometrium from women with endometriosis might be increase angiogenesis and growth ectopic endometriosis implants, we investigated PTN and MK expression by quantitative and competitive polymerase chain reaction (QC-PCR) in endometrium from women with and without endometriosis throughout the menstrual cycle. Design: MK and PTN mRNA expression in endomeoium from women with endometriosis and control patients without endometriosis were determined by QC-PCR throughout the menstrual cycle. Methods: Endometrial tissue was obtained from 25 patients with severe endometriosis and 30 patients without endometriosis undergoing hysterectomy or endometrial biopsy. Stage of endometrial cycle and a diagnosis of endometriosis were confirmed histologically. Total RNA was extracted and reverse transcribed into c-DNA. QC-PCR was performed to evaluate PTN and MK mRNA expression. Results were analysed by Post Hoc test. Results: MK and PTN were expressed throughout the menstrual cycle in both groups. MK expression was higher in follicular phase than luteal phase in endometrium from normal women. endometrium from endometriosis patients showed increased expression of PTN and MK compared to endometrium from normal women in the luteal phase (p<0.05). Conclusion: Our results suggest that uterine endometrium from women with endometriosis expresses higher levels of MK and PTN than endometrium from normal women during luteal phase. Increased MK and PTN expression may be related to the initiation of ectopic endometrial implants and their subsequent peritoneal invasion.

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Endometrium from Women with Endometriosis Expresses Decreased Levels of Plasminogen Activator Inhibitor-1 and Tissue Inhibitor of Metalloproteinase-3 Compared to Normal Endometrium (자궁내막증 환자와 정상 여성의 자궁내막에서 TIMP-3와 PAI-1 mRNA 발현 차이에 관한 연구)

  • 정혜원
    • Development and Reproduction
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    • v.3 no.1
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    • pp.29-38
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    • 1999
  • The pathogenesis of endometriosis is unknown, but retrograde menstruation is widely accepted as an etiology. Refluxed endometrium from endometriosis patients is more prone to implant and invade peritoneum possibly through the action of extracellular proteolysis. This proteolytic action may involve plasminogen activators and the collagenase system. Plasminogen activators (PAs) and matrix metalloproteinases (MMPs) play a critical role in the breakdown of extracellular matrix components and basement membrane in the processes of implantation and tumor invasion. PAs are inhibited by plasminogen activator inhibitor (PAI) and MMPs activity is inhibited by tissue inhibitor of metalloproteinase (TIMP). To test the hypothesis that lower expression of PAI-1 and TIMP-3 in endometrium from women with endometriosis, we investigated their PAI-1 and TIMP-3 expression by quantitative competitive RT PCR in endometrium from women with and without endometriosis. Endometrial tissues were obtained from 14 patients with severe endometriosis and 14 patients without endometriosis. Total RNA was extracted and reverse transcribed into cDNA, and quantitative competitive PCR (QC PCR) was performed to evaluate PAI-1 and TIMP-3 mRNA expression. Endometrium from patients with endometriosis showed decreased expression of PAI-1 and TIMP-3 mRNA compared to endometrium from control in luteal phase (p<0.05). Our results suggest that endometrium from women with endometriosis expresses lower levels of PAI-1 and TIMP-3 than endometrium from normal women. Endometrium from endometriosis patients may be more invasive and prone to peritoneal implantation than control because of higher PA and MMP enzymatic activity. Thus, increased proteolytic activity may be one of the reasons for the invasive properties of the endometrium resulting in the development of endometriosis.

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mRNA Expression of Thrombospondin-1 and -2 in Severe Endometriosis Patients in Korean Women (중증 자궁내막증 환자의 내막에서 thrombospondin-1과 -2의 mRNA의 발현에 관한 연구)

  • Hur, Sung Eun;Lee, Ji Young;Moon, Hye-Sung;Chung, Hye Won
    • Clinical and Experimental Reproductive Medicine
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    • v.32 no.3
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    • pp.253-259
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    • 2005
  • Objective: We investigated the expression of TSP-1 and -2 in eutopic endometrium of advanced endometriosis and control patients. Methods: The 33 endometriosis patients and 32 controls were enrolled. Endometrial samples were obtained from 65 premenopausal women aged 29-44 years, undergoing laparoscopic surgery or hysterectomy for non-malignant lesions. Sufficient samples were collected from 33 patients with endometriosis stage III and IV and 32 controls without endometriosis confirmed by laparoscopic surgery. The mRNA expression from eutopic endometrium for TSP-1 and -2 were analyzed by RT-QC PCR. Results: The mRNAs of TSP-1 and -2 were expressed in eutopic endometrium from endometriosis and normal controls throughout the menstrual cycle. There were no significant differences in expression of TSP-1 and TSP-2 in eutopic endometrium between controls and endometriosis patients. Conclusion: Our results indicated that TSP-1 and -2 had no crucial role compared to other molecules in the regulation of angiogenesis. These findings also suggest that dysregulation of other angiogenic regulators would be concerned in pathophysiologic role in endometriosis development.

mRNA Expression Differences of uPA, uPAR in Eutopic Endometrium of Advanced Stage Endometriosis Patients (중증 자궁내막증 환자의 자궁내막과 정상인 자궁내막에서 uPA, uPAR mRNA 발현의 차이에 관한 연구)

  • Hur, Sung Eun;Lee, Ji Young;Lee, Woon Jung;Chung, Hye-Sung;Chung, Hye Won
    • Clinical and Experimental Reproductive Medicine
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    • v.33 no.4
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    • pp.229-236
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    • 2006
  • Objective: We investigated the expression of uPA and uPAR in eutopic endometrium of advanced stage endometriosis and control patients. Methods: The 33 endometriosis patients and 32 controls were enrolled. Endometrial samples were obtained from 65 premenopausal women aged 29-44 years, undergoing laparoscopic surgery or hysterectomy for non-malignant lesions. Sufficient samples were collected from 33 patients with endometriosis stage Ill and IV and 32 controls without endometriosis confirmed by laparoscopic surgery. The mRNA expression of uPA and uPAR from eutopic endometrium were analyzed by RT-QC PCR. Results: The mRNAs of uPA and uPAR were expressed in eutopic endometrium from endometriosis and normal controls throughout the menstrual cycle. Uterine endometrium from women with endometriosis expresses significantly (p<0.05) higher levels of u-PA mRNA than endometrium from normal women without endometriosis in the proliferative phase. There were no significant differences in expression of uPAR in eutopic endometrium between controls and endometriosis patients. Conclusion: These results suggest that eutopic endometrium from endometriosis patients may be more invasive and prone to peritoneal implantation because of greater u-PA mRNA expression than endometrium from women without endometriosis. Thus, increased proteolytic activity may be one etiology for the invasive properties of the endometrium resulting in the development of endometriosis.

Expression of angiogenin, TGF-${\beta}$, VEGF, APEX and TNF-${\alpha}$ in oral squamous cell carcinoma

  • Lee, Ho-Sun;Kim, Kyoung-Won;Kim, Wun-Jae
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.32 no.1
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    • pp.8-18
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    • 2006
  • Purpose: The purpose of this study was to verify that the expressions of angiogenin, transforming growth factor-beta(TGF-${\beta}$), vascular endothelial growth factor(VEGF), human apurinic/apyrimidinic endonuclease(APEX) and tumor necrosis factor-alpha(TNF-${\alpha}$) were associated with the tumorigenesis of the oral squamous cell carcinoma(OSCC). Materials and Methods: Fifty-one samples of OSCC and fifteen normal oral mucosae were obtained to analyze the expression levels of above five factors. mRNA expressions were quantified by the quantitative competitive PCR(QC-PCR) method. After 2% agarose gel electrophoresis stained with ethidium bromide, the concentration of mRNA was calculated by a digital image analysis system. The expression levels of angiogenin, TGF-${\beta}$, VEGF, APEX and TNF-${\alpha}$ were compared by unpaired Student's t-tests between cancer and normal tissues. We analyzed statistically to find the cut-off values that would be useful as diagnostic markers, and the linear regression analysis between every two factors of these five factors by SAS system. Results: All of these five factors (angiogenin: P<0.0037, TGF-${\beta}$: P<0.0001, VEGF: P<0.0102, APEX: P<0.0023, TNF-${\alpha}$: P<0.0074) were significantly correlated with OSCC. In the analysis to find the cut-off values for the diagnosis, we could not find any value that had a reasonable sensitivity and specificity. In the linear regression analysis, there were correlations between angiogenin and TNF-${\alpha}$, TGF-${\beta}$ and VEGF, TGF-${\beta}$ and APEX, TGF-${\beta}$ and TNF-${\alpha}$, VEGF and APEX, VEGF and TNF-${\alpha}$, APEX and TNF-${\alpha}$. Conclusion: Our results suggest that not only angiogenin, TGF-${\beta}$, VEGF, APEX and TNF-${\alpha}$ are significantly associated with the tumorigenesis, but also the close relationship between these factors might enhance the tumorigenesis of OSCC. We can not find clinical availability for diagnosis.