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Objective:To investigate the effect of the adoptive transfer of CD4+CD25+Foxp3+regulatory T cells ( iTregs) induced by 5-aza-2′-deoxycytidine (5AzaD) on pregnant outcome of the abortion-prone mice.Methods:Sixty cases of female CBA/J × male DBA/2J abortion-prone matings were taken as study group,the CD4+T cells from spleen of twenty female CBA/J mice were separated by magnetic activated cell sorting (MACS),5AzaD was applied to the conversion of CD4+CD25-T cells to iTregs,the expression of Foxp3 in Tregs was characterized by flow cytometry analysis before and after epigenetic modification.The purified iTregs were injected into abortion-prone mice on day 1 or 4 of pregnancy,respectively,which were used as therapy groups,and then the embryo resorption rate was counted on day 14 of pregnancy.Results:After the treatment of 5AzaD,the percentage of iTregs in CD4+T cells was (41.50±8.03)%.The embryonic absorption rates of the two therapy groups were 10.47%(on day 1 of pregnancy) and 21.69%(on day 4 of pregnancy) ,respectively ( P<0.05 ) .Conclusion: Epigenetic modication of CD4+CD25-T cells may solve the problem of nTregs deficiency,particularly adoptive therapy of 5AzaD-induced iTregs at early stage of pregnancy can maintain normal pregnancy.
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<p><b>OBJECTIVE</b>To investigate the expression of placenta-derived RASSF1A gene in maternal plasma during first and second trimesters, and to explore its value for the prediction of pre-eclampsia.</p><p><b>METHODS</b>For 325 pregnant women of the first trimester, free DNA of plasma samples was extracted at 7-12, 13-18, and 19-24 gestational weeks, respectively. Methylation-sensitive restriction enzyme digestion followed by fluorescence quantitative PCR (MSRE+ PCR) was employed for analyzing the concentrations of hypermethylated RASSF1A gene. Blood pressure, proteinuria and clinical feature were monitored at the same time. Those who had subsequently developed pre-eclampsia were selected as the pre-eclamptic group, 30 normal pregnant women were selected as the control group. Hypermethylated RASSF1A gene in maternal plasma was retrospectively analyzed. The relationship between clinical classification, type of pre-eclampsia and concentrations of the gene were further analyzed.</p><p><b>RESULTS</b>Twenty-six out of the 325 pregnant women developed pre-eclampsia as their only complication. At 13-18 gestational weeks, the mean concentrations of fetus-specific RASSF1A sequences were 141.62 copies/mL in maternal plasma of pre-eclamptic pregnancies, which was significantly greater than that of the controls (98.90 copies/mL). Fetus-derived RASSF1A levels were 2.03 fold higher in pre-eclamptic subjects than controls at 19-24 gestational weeks. There was a significant difference in the level of hypermethylated RASSF1A gene between the mild and severe pre-clamptic subjects at 13-24 gestational weeks (P< 0.05). The concentrations of the sequences were significantly higher in early-onset severe pre-eclampsia than late-onset severe pre-eclampsia at 19-24 gestational weeks (P< 0.05).</p><p><b>CONCLUSION</b>Altered expression of hypermethylated RASSF1A gene may be detected in maternal plasma during second trimester, which has important significance for early prediction of pre-eclampsia.</p>
Subject(s)
Female , Humans , Pregnancy , Gestational Age , Placenta , Metabolism , Pre-Eclampsia , Blood , Diagnosis , Genetics , Metabolism , Predictive Value of Tests , Pregnancy Trimester, Second , Prenatal Diagnosis , Methods , Tumor Suppressor Proteins , Blood , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To investigate the level of hypermethylated ras association domain family 1A (RASSF1A) gene in maternal plasma of pre-eclampsia and its clinical value.</p><p><b>METHODS</b>Sixty pre-eclampsia women including 30 mild and 30 severe cases were selected, 60 women with normal pregnancy were studied as control. Free DNA from plasma samples was extracted, fluorescence quantitative polymerase chain reaction (FQ-PCR) was used to detect the concentrations of RASSF1A gene before and after methylation-sensitive restriction digestion. Meanwhile, beta-actin gene was detected as a control to confirm complete enzyme digestion.</p><p><b>RESULTS</b>The median concentration of hypermethylated RASSF1A gene was 3.31-fold higher in samples from pre-eclamptic pregnancies than that in controls. There was significant difference between the mild and severe pre-eclamptic subjects (P<0.05), with the median concentrations of 1659 copies/mL and 2036.50 copies/mL, respectively.</p><p><b>CONCLUSION</b>Hypermethylated RASSF1A gene in pre-eclampsia plasma was significantly increased and the concentrations were related to the severity of pre-eclampsia.</p>
Subject(s)
Adult , Female , Humans , Pregnancy , Young Adult , DNA , Blood , Metabolism , DNA Methylation , Pre-Eclampsia , Genetics , Metabolism , Pathology , Tumor Suppressor Proteins , Genetics , MetabolismABSTRACT
Objective To study the effects of flavone compound α-NF on CYP1B1 gene in transplanted nude mice tumor with human paclitaxel resistance ovarian cancer cell. Methods Using 24 nude models of orthotopic transplantation of human ovarian cancer, they were divided randomly into four groups: control group, α-NF group, paclitaxel group and combined group. RT-PCR and Western blotting method were used to detect CYP1B1 mRNA and protein expression. Results There was no significant difference in CYP1B1 mRNA expression between α-NF group, combined group and control group(P >0.05). Compared with control group, CYP1B1 protein expression was distinctly lower in α-NF and combined group (P<0.01). Conclusion α-NF has no effect on CYPIB1 gene in mRNA level, whereas α-NF down-regnlate, CYP1B1 expression of transplanted A2780TS nude mice tumor in protein level, thus α-NF has the effect on reversing paclitaxel resistance of A2780TS cell.
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Objective To investigate the relationship between expression of CDK1,TFF2 and HPV16/18 infection in uterine cervical carcinomas.Methods Formalin-fixed,paraffin-embedded samples including 50 cases of squamous cell carcinomas,20 cases of cervical intraepithelial neoplasia CINⅡ-Ⅲand 20 cases of CINI and 20 cases of normal cervical tissues were studied.Using Polymer DetectionSystem For Immuno-Histological Staining(PV-9000)as immunohisto-chemical detecting method.the expression of TFF2 and CDK1 in these samples were examined.The infections of HPV type 16,18 DNA were determined by PCR.Results There were significant differences for CDK1 between carcinomas,CIN and normal cervices,respectively(all P＜0.05).Expression of CDKl were correlated with tumor grades(P＜0.05).The positive expression of TFF2 in carcinomas was significantly lower than that in normal cervical tissue and CIN.There was significant differences amongthem(P＜0.05).Expression of TFF2 were correlatedwith age(P＜0.05).Thepositive rate of HPV16/18 was significant difference between cervical carcinomas,CIN and normal tissues(P＜0.05),and it was not associated with age,tumor grades,clinical stages and lymph node metastasis(all P＞0.05).The expression of CDK1 was not associated with TFF2 and HPV16/18(all P＞0.05).The expression of HPV16/18 was associated with TFF2(rs=-0.500,P＜0.05).Conclusions The results suggest that in tissues of CIN and cervical carcinomas in TFF2 and CDK1 expression changes and HPVl6/18 infection interact with each other,which can effect the progression of CIN and carcinogenesis of cervical carcinomas.
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Objective To investigate the relationship between expression of survivin and CD44v6 and HPV16/18 infection in uterine cervical carcinogenesis.Methods Using Streptavidin-Peroxidase(S-P) immunohistochemical technique,the authors examined the expression of survivin and CD44v6 in samples.The infection of HPV type 16,18 DNA was determined by PCR.Results There were significant differences for survivin and CD44v6 between carcinomas,CIN and normal cervices(P
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Objective:To investigate the expression of minichromosome maintenance proteins 4(MCM4)and cell division cycle 6(CDC6)in uterine cervical carcinomas and its relationship with human papilloma virus(HPV)16/18 infection.Methods:The expression of MCM4 and CDC6 was examined in 50 squamous cell carcinoma specimens,20 cervical intraepithelial neoplasia(CIN)Ⅱ-Ⅲ specimens,20 CINⅠ specimens,and 20 normal cervical tissues by immunohistochemical method.The infections of HPV type 16,18 DNA were determined by PCR.Results:(1)The expression of MCM4 and CDC6 in uterine cervical carcinoma tissues was significantly higher than that in the CIN specimens and normal cervical tissues(Both P0.05).(2)The positive rates of(HPV)16/18 were significant different between cervical carcinomas,CIN and normal tissues(P0.05).(3)MCM4 expression were positively correlated with the expression of CDC6 in uterine cervical carcimonas(r=0.390,P
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Objective:To explore the expression of Toll-like receptor(4TLR4),nuclear factor -kappaB p65(NF-?Bp65) and tumor necrosis factor-alpha(TNF-?)in placenta of preterm birth of infected mouse model,as well as intervention of N-Acetylcysteine(NAC) in order to develop a novel strategy for prevention from premature birth.Methods:Expression of mRNA and protein of TLR4,NF-?Bp65 and TNF-? in the mouse placenta of premature birth was analyzed by using RT-PCR and immunohistochemistry.LPS injection was taken to develop the infectious mouse mode,and the incidence of premature birth was observed.The regimen of NAC intervention was performed and its modulation for placenta expression of TLR4,NF-?Bp65 and TNF-? was investigated.Results:It was shown that TLR4,NF-?Bp65 and TNF-? were expressed at a low level in control groups and in LPS model groups,their expressions were significantly increased when compared with the control groups(P