Study on the correlation between methylation level of PGC-1α gene and fetal distress in pregnant women with gestational diabetes mellitus / 中华内分泌外科杂志

Objective:To detect the methylation level and mRNA expression level of peroxisome proliferator activated receptory-coactivator-1α (PGC-1α) gene in placental tissue of pregnant women with gestational diabetes (GDM) and to explore the relationship between them and fetal distress.Methods:A total of 174 pregnant women with GDM admitted to in our hospital from Jul. 2018 to Dec. 2019 were selected as the study objects, among which 78 pregnant women with fetal distress were selected as the fetal distress group; and 96 pregnant women with normal delivery and without fetal distress were the control group; during the same period, 82 normal pregnant women without GDM were selected as the healthy group. The methylation level of PGC-1α gene in placenta was detected by direct sequencing after DNA was treated with sodium bisulfite; the expression of PGC-1α mRNA in placenta was detected by real-time fluorescence quantitative PCR (qRT-PCR) ; the levels of triglyceride (TG) , total cholesterol (TC) , low density lipoprotein cholesterol (LDL-C) and high density lipoprotein cholesterol (HDL-C) were measured by automatic biochemical analyzer; the relationship between methylation frequency of PGC-1α gene and the expression level of PGC-1α mRNA was analyzed; and the influencing factors of fetal distress were analyzed.Results:PGC-1α gene methylation frequency and TG level were higher in the fetal distress group [ (25.42±7.31) %, (4.72±0.68) mmol/L] than in the control group [ (9.26±2.67) %, (4.31±0.64) mmol/L] and the healthy group [ (3.24±1.07) %, (4.33±0.72) mmol/L]. PGC-1α gene methylation frequency was higher in the control group than in the healthy group, and the differences were statistically significant ( P<0.05) ; PGC-1α mRNA expression level in fetal distress group (0.67±0.16) is lower than that in the control group (0.74±0.14) and healthy group (1.00±0.27) . PGC-1α mRNA expression level in control group was lower than that in healthy group, and the difference was statistically significant ( P<0.05) ; the methylation frequency of PGC-1α gene was negatively correlated with the expression level of PGC-1α mRNA in pregnant women with fetal distress ( r=-0.515, P<0.05) ; the methylation of PGC-1α gene was an independent risk factor for fetal distress ( P<0.05) , and the high expression of PGC-1α mRNA was the protective factor of fetal distress ( P<0.05) . Conclusion:DNA methylation level of PGC-1α gene in pregnant women with GDM is related to fetal distress, which may be the target of gene modification for fetal distress.

Performance evaluation of methylation detection of plasma Septin[STHZ]9[STBZ] gene and its diagnostic value in patients with colorectal cancer / 临床检验杂志

Objective@#To evaluate the value of methylation detection of plasma Septin9 gene in the diagnosis of colorectal cancer (CRC) and verify its performance. @*Methods@#The plasma samples from 32 CRC patients before colonoscopy and 10 healthy controls during October 2016 and May 2017 were collected, and the methylation levels of Septin9 gene in these samples were detected by the detection kit of plasma Septin9 gene methylation. The coincidence rate, detection limit and precision of the kit in the diagnosis of CRC were evaluated, and its diagnostic value was compared with that of carcinoembryonic antigen (CEA) and facal immunochemical tests (FIT). @*Results@#The positive and negative coincidence rates of the plasma Septin9 gene methylation kit in the detection of CRC were 100%. The reference materials assigned the detection limit were positive, and the coefficient of variation (CV) of precision was less than 5%, which met the basic performance requirements. The sensitivity, specificity, positive predictive value and negative predictive value of the kit in the diagnosis of CRC were 62.50%, 90.00%, 95.20% and 42.90%, respectively. The detection rate of CRC by the kit was 62.50%, significantly higher than those of FIT (28.13%) and CEA (28.13%) (all P＜0.05). The area under the ROC curve (AUC ROC ) of the kit in the diagnosis of CRC was 0.762, and the detection rate of stage Ⅰ CRC by the kit was 50.00%. @*Conclusion@#The performance of the plasma Septin9 gene methylation kit meets the anticipated clinical requirements, which may be used as a serological marker for the assistant diagnosis of CRC.

Detection of plasma HOXA7 methylation in the diagnosis of non-small cell lung cancer / 中国肿瘤临床

Objective: To detect the methylation levels of homeobox protein Hox A7 (HOXA7) gene promoter in the plasma of non-small cell lung cancer (NSCLC) and to study the value of HOXA7 methylation and serum levels of the tumor markers carcinoembryonic antigen (CEA), carbohydrate antigen 199 (CA199), and cytokeratin 19 fragment (Cyfra21-1) in the auxiliary diagnosis of NSCLC. Methods: Plasma samples were collected from 80 patients with NSCLC and 50 healthy controls, which were enrolled in Henan Cancer Hospital from January 2012 to December 2016. The plasma HOXA7 methylation levels were detected by real-time quantitative methylation-specific PCR, and the plasma levels of CEA, CA199, and Cyfra21-1 were detected by electrochemical luminescence. The ROC curve was constructed to analyze the clinical value of each index in the differential diagnosis of NSCLC, and the relationship between HOXA7 methylation, clinical parameters, and tumor markers was analyzed. Results: The serum levels of HOXA7 methylation in NSCLC patients were significantly higher (χ2=36.972, P<0.000 1) than the healthy control group. The sensitivity and specificity of HOXA7 methylation in the diagnosis of NSCLC were 68.8% (55/80) and 86.0% (43/50), respectively. Plasma HOXA7 methylation was not related to gender, age, smoking history, or pathological type (all P>0.05), but was related to TNM stage (P<0.05). The plasma HOXA7 methylation level of stageⅣpatients (81.8%, 18/22) was the highest. Cyfra21-1 had the highest value in the diagnosis of NSCLC among tumor markers with a sensitivity of 70.00% and a specificity of 90.00%. The combination of HOXA7 methylation with all three tumor markers had the highest efficiency (AUC=0.893, sensitivity 73.75%, specificity 94%) in the diagnosis of NSCLC. The correlation coefficient between HOXA7 methylation and Cyfra21-1 was the highest (r=0.564, P<0.05). Conclusions: HOXA7 gene methylation is related to the degree of metastasis of NSCLC and proves as an efficient diagnostic marker for NSCLC when combined with tumor marker detection.

Methylation in the promoter regions of WT1, NKX6-1 and DBC1 genes in cervical cancer tissues of Uygur women in Xinjiang

Abstract This study aimed to explore: 1) DNA methylation in the promoter regions of Wilms tumor gene 1 (WT1), NK6 transcription factor related locus 1 gene (NKX6-1) and Deleted in bladder cancer 1 (DBC1) gene in cervical cancer tissues of Uygur women in Xinjiang, and 2) the correlation of gene methylation with the infection of HPV16/18 viruses. We detected HPV16/18 infection in 43 normal cervical tissues, 30 cervical intraepithelial neoplasia lesions (CIN) and 48 cervical cancer tissues with polymerase chain reaction (PCR) method. Methylation in the promoter regions of the WT1, NKX6-1 and DBC1 genes in the above-mentioned tissues was measured by methylation-specific PCR (MSP) and cloning sequencing. The expression level of these three genes was measured by real-time PCR (qPCR) in 10 methylation-positive cervical cancer tissues and 10 methylation-negative normal cervical tissues. We found that the infection of HPV16 in normal cervical tissues, CIN and cervical cancer tissues was 14.0, 36.7 and 66.7%, respectively. The infection of HPV18 was 0, 6.7 and 10.4%, respectively. The methylation rates of WT1, NKX6-1 and DBC1 genes were 7.0, 11.6 and 23.3% in normal cervical tissues, 36.7, 46.7 and 30.0% in CIN tissues, and 89.6, 77.1 and 85.4% in cervical cancer tissues. Furthermore, WT1, NKX6-1 and DBC1 genes were hypermethylated in the high-grade squamous intraepithelial lesion (CIN2, CIN3) and in the cervical cancer tissues with infection of HPV16/18 (both P< 0.05). The expression of WT1, NKX6-1 and DBC1 was significantly lower in the methylation-positive cervical cancer tissues than in methylation-negative normal cervical tissues. Our findings indicated that methylation in the promoter regions of WT1, NKX6-1 and DBC1 is correlated with cervical cancer tumorigenesis in Uygur women. The infection of HPV16/18 might be correlated with methylation in these genes. Gene inactivation caused by methylation might be related to the incidence and development of cervical cancer.

Research progress of imprinted gene H19 methylation in the field of assisted reproduction / 医学研究生学报

The imprinted gene H19, one of the maternally imprinted gene which had been found long time ago, is highly expressed in early embryo. It mainly concentrates in endoderm and mesoderm which plays an important role in regulating the development of embryos and in control of off spring behavior. More and more studies have shown that the alterations of methylation status in CpG islands of H19 gene imprinted control region will lead to the decline of male sperm quality and female oocyte quality, then affecting human fertility. Imprinting gene H19 is also related to the development of the embryo. This article reviews the progress of methylation of the imprinted gene H19 in assisted reproductive technology.

Detection and the Significance of SOCS-1 Gene Methylation Status in Peripheral Blood of Systemic Lupus Erythematosus Patients / 现代检验医学杂志

Objective To investigate the relationship between the systemic lupus erythematosus (SLE)and the SOCS-1 gene methylation status of the peripheral blood DNA,to provide the basis for diagnosis and treatment of systemic lupus erythema-tosus.Methods Blood samples of SLE patients (27 cases)and healthy group (19 cases)in January 2015 to April were col-lected and the DNA were extracted.Using polymerase chain reaction combining DNA agarose gel electrophoresis to detect the SOCS-1 gene methylation status.Results In patients with systemic lupus erythematosus SOCS-1 gene complete methyl-ation accounted for 44% (12/27),incomplete methylation accounted for 56% (15/27).In healthy group SOCS-1 gene com-plete methylation accounted for 74% (14/19)and incomplete methylation accounted for 26% (5/19).The rate of complete methylation of SOCS-1 gene of SLE patients was lower than that of healthy group (χ2=3.88,P=0.049).Conclusion SLE patients may have lower SOCS-1 gene methylation status in the peripheral blood DNA,which is worth for further study.

Correlation between stress and BDNF gene promoter CpG methylation in patients with major depressive disorder / 中华行为医学与脑科学杂志

Objective To investigate the effects of stress on the methylation of brain derived neurotrophic factor gene in the patients with major depressive disorder (MDD),and to investigate the relationship between BDNF gene methylation and MDD.Methods 47 cases of MDD were divided into MDD stress group (n=24) and MDD non stress group(n=23) while 27 health subjects were collected as normal control group.The methylation status of CpG island in the promoter region of BDNF gene in peripheral blood was detected by the method of heavy hydrogen and hydrogen sulfate.SPSS17.0 statistical software package was used to analyze the data.The differences of 19 CpG methylation rates and the overall level of methylation rates of three groups were analyzed.Results The CpG overall methylation rates (median,interquartile range) of MDD stress group,MDD non stress group and normal control group was 189.150 (7.575),188.500 (400)and 480.200(770) respectively,and the difference was statistically significant (P＜0.01).There was no significant difference in the overall methylation rate of CpG in the MDD stress group compared with MDD non stress group (P＞0.05).The CpG methylation rates (mean± SD or median,interquartile range) of three groups were detected as follows:CpG-1:2.600(0.275),2.700 (0.400),6.500(0.800);CpG-2:3.350(0.650),3.300(0.800),14.600(1.500);CpG-3:1.596±0.363,1.543±0.400,4.581 ±0.437;CpG-4:1.779±0.516,1.522±0.329,4.033 ±0.529;CpG-5:0.900 (0.575),0.800 (0.600),5.700 (1.500);CpG-6:6.258 ± 0.805,6.213 ±0.944,14.589±0.819;CpG-7:10.667±0.894,10.283± 1.006,15.000±0.763;CpG-8:16.421 ±0.697,16.330±0.775,24.796±0.547;CpG-9:4.713±0.565,4.891 ±0.554,28.826±0.679;CpG-10:10.254±0.902,10.378±0.777,11.381±0.538;CpG-11:24.125±2.301,24.170±2.613,37.474± 1.579;CpG-12:5.442±0.641,5.596±1.117,12.141 ±0.940;CpG-13:4.150(1.150),4.200(1.000),61.700(4.800);CpG-14:5.500±0.544,5.717±0.568,6.378±0.397;CpG-15:3.700 (0.700),4.100(1.000),63.300(2.500);CpG-16:8.200 (1.775),8.100(1.500),75.200(3.300);CpG-17:3.250(0.550),3.300(0.800),34.600(5.000);CpG-18:1.988±0.279,1.939±0.259,2.330±0.207;CpG-19:35.338±2.421,35.187±2.259,65.941 ±2.692.16 CpG methylation rates of 19 CpG were higher in MDD stress group and MDD non stress group.Compared with the normal control group,the difference was statistically significant (P＜0.01).There was no significant difference in CpG methylation rate between MDD stress group and MDD non stress group (P＞0.05).Conclusion The overall methylation rate of CpG in BDNF gene promoter region is closely related with MDD,which may affect the incidence of MDD.There was no correlation between CpG methylation in BDNF gene promoter region and MDD,and stressful life events may not be the direct cause of CpG methylation in BDNF gene promoter region in patients with MDD.

Analysis ofDBC1 gene promoter methylation in cervical cancer tissues of Uyghur women in Xinjiang / 中国癌症杂志

Background and purpose:In recent years, epigenetics research has become a new direction of cancer research. A large number of results have shown that the abnormal changes of epigenetic modifications have close connection with cancer. Genome-wide epigenetic modifications have become new markers for cancer. This study aimed to investigate the methylation of the promoter ofDBC1 gene in cervical cancer tissues of Uyghur women in Xinjiang, to explore the correlation between the gene methylation and the infection of HPV, and to evaluate whether it can be used as a tool with high sensitivity and specificity for cervical cancer screening.Methods:This study detected the infection of HPV16, 18 in 43 normal cervical tissues, 35 cervical intraepithelial neoplasia tissues and 54 cervical cancer tissues using the polymerase chain reaction (PCR) method. The methylation of the promoter ofDBC1 gene in above-mentioned tissues was detected by the methylation-specific PCR method. The expression ofDBC1 at mRNA level was measured by real-time fluorescent quantitative polymerase chain reaction (RTFQ-PCR) in 10 methylation-negative normal cervical tissues and 10 methylation-positive cervical cancer tissues.Results:In normal cervical tissues, CIN tissues and cervical cancer tissues, the infection ratios of HPV16 were 18.6%, 34.3% and 68.5%, respectively; the infection ratios of HPV18 were 2.3%, 8.6% and 16.7%, respectively; and the methylation ratios ofDBC1 gene were 23.3%, 40.0%, 87.0%, respectively. In 79 high-grade squamous intraepithelial lesions (CINⅡ and Ⅲ) and cervical cancer tissues, 50 of 79 were infected with HPV16/18, while 29 of 79 were negative. The methylation ratio ofDBC1 gene was 88.0% in HPV16/18 infection positive group while the methylation ratio was 55.2% in negative group (P<0.05). The expression ofDBC1 gene at mRNA level in 10 methy- lation-positive cervical cancer tissues was significantly lower than that in the 10 methylation-negative normal cervical tissues (P<0.05).Conclusion:The methylation ofDBC1 gene may become a molecular marker to detect cervical cancer of Uyghur women in Xinjiang.DBC1 gene methylation combined with HPV16/18 infection test can be used to aid diagnosis of cervical cancer.

Abnormal expression and its regulation mechanism of Cav-1 gene in gastric cancer / 临床与实验病理学杂志

Purpose To investigate the expression of Cav-1 in gastric cancer ( GC) cell lines and GC samples, and to analyze the pos-sible effect of gene methylation in expression of Cav-1. Methods Methylation specific PCR ( MSP) method was applied to examine the CpG methylation of the Cav-1 promoter in GC cell lines (AGS, MKN45, BGC-823) and 104 samples of GC and corresponding ad-jacent tissues. RT-PCR method was applied to examine the mRNA expression in GC cell lines. IHC were applied to examine the pro-tein expression of Cav-1 in the GC tissues. Results The expression level of Cav-1 mRNA was obviously increased after treated with 5-aza-2’-deoxycytidine (5-Aza-Dc, a demethylation agent) in AGS cell line. We detected the positive expression of Cav-1 gene in MKN45 and BGC-823 cell lines before and after treated with 5-Aza-Dc. The level of Cav-1 mRNA expression was no any change in AGS, MKN45 and BGC-823 cell lines treated with trichostatin A (TSA). MSP results showed that it can be amplified methylated bands in AGS cell line, and the methylated bands disappeared after treated with 5-Aza-Dc. MKN45 and BGC-823 cell lines were no any methylated bands amplified before and after treatment. The methylation frequency of Cav-1 gene was 29. 8% (31/104), which was significantly higher than that in adjacent tissues (P=0. 000). Furthermore, Cav-1 gene hypermethylation status was correlated with lymph node metastasis and family history of upper gastrointestinal cancers ( UGIC) , but not with pathological grade and clinical stage (P>0. 05). The positive frequency of Cav-1 expression was 51. 9%(54/104) in GC, which was significantly lower than that in adja-cent tissues (P=0. 000). The expression of Cav-1 was correlated with the frequency of gene methylation in GC tissues (P=0. 000). Conclusion The expression of Cav-1 was reduced in GC tissues and the gene hyermethylation may be one of the mechanisms causing Cav-1 gene silencing.

Radioprotective effects of FUZHENGJIEDU on rats exposed to radon / 中华放射医学与防护杂志

Objective To investigate the protective effects of FUZHENGJIEDU,a kind of Chinese herb on rats exposed to radon and it's daughter.Methods Twelve male SD rats were exposed with concentration of 40 000 Bq/m3 radon and the exposed dose was 120 work level month (120 WLM),and then all the rats were randomly divided into 2 groups.One group was 120 WLM group (positive control,n=6 ),the other group was FUZHENGJIEDU treatment group (120 WLM exposed rats were given FUZHENGJIEDU 5.0 g every day,n =6),meanwhile 6 rats which lived in the normal environment ( the concentration of radon and it's daughter was lower than 50 Bq/m3 ) as control group.The levels of tumor markers induding carcinoembryonic antigen ( CEA),neuron-specific enolase (NSE),CYFRA21-1 and p53 antibody were tested by ELISA.The rate of p16 gene methylation was measured by methylation-specific PCR (MSP).Results The levels of CEA and NSE in 120 WLM group were (396.62 ± 148.74) and (9.09 ±0.90) μg/L,respectively,significantly higher than those of control group (t =2.583,2.463,P ＜ 0.05).The levels of CEA and NSE in treatment group were (70.89 ±44.71) and (4.31 ±1.37) μg/L,respectively,remarkably lower than those of 120 WLM group (t =2.921,2.526,P ＜0.05 ).The concentrations of CYFRA21-1 and p53 antibody were not significantly different among the three groups (t =1.713,1.963,P ＞ 0.05 ).The rate of p16 gene methylation in BALF cells in 120 WLM group was 16.67％,but in control group and treatment group did not arise p16 gene methylation in BALF cells.Conclusions FUZHENGJIDU would decrease the levels of CEA and NSE and the rate of p16 gene methylation,and could exert it's protective effect on rats exposed to high concentration of radon.

The value of CT scan and tumor markers in sputum for early diagnosis of peripheral lung cancer / 重庆医科大学学报

Objective:To investigate the diagnostic value of telomerase activity and p16 gene methylation from exfoliated cells of sputum in 55 cases of solitary nodules and of being suspected of early peripheral lung cancer(T1N0M0).Methods:The sputum specimens from 34 cases of cancer nodes and 21 cases of benign lung lesion were detected for telomerase activity and p16 gene methylation by methylation analysis.Results:The qualitative diagnosis accuracy of CT scan was 61.8%(34/55) for peripheral lung cancer.Telomerase activity was positive in 29 cases:Sensitivity was 79.4%;Specificity was 90.5%;and accuracy was 83.6%.p16 gene methylation was in 11 cases:Sensitivity was 32.4%;specificity was 100%,and veracity was 58.2%.The sensitivity was increased to 86.1% by the combination of telomerase activity and p16 gene methylatiion.Conclusion:The results suggest that combining CT scan with telomerase activity and p16 gene methylation detection in sputum for patients with lung cancer may enhance the diagnostic value of conventional cytology.It is a promising approach to early diagnosis of lung cancer and massive screening in terms of its rapidity,economy and simplicity.

5-Aza-cytidine inhibit proliferation of human promyelocytic leukemia cells (HL60) by inducing apoptosis / 重庆医科大学学报

Objevtive:This study was aimed to investigate the effect of 5-aza-cytidine on the proliferation,cell cycles distribution profile and apoptosis of human promyelocytic cell line HL60.Methods: The human promyelocytic cell line HL-60 was cultured in RPMI 1640 medium.3-(4,5-Dimethylthiazol-2-y)l-2,5-diphenylte trazolium(MTT)reduction by cells was used to assess drug-induced cell growth inhibition.Cells were treated with 0.25,0.5,1,2.5?mol/L of freshly prepared 5-aza-cytidine for 48 hours before they were harvested for analyzing distribution of cell cycle phases and apoptosis by flow cytometer.Results:5-aza-cytidine inhibited HL60 cell proliferation in a time-and concentration-dependent manner.Apoptosis was induced as detected by flow cytometry.Determination of the cell cycle distribution by flow cytometry revealed an accumulation of cells in G1 phase.Conclusion:DNA methylation inhibitor 5-aza-cytidine has effects on proliferation of human promyelocytic cell line HL60.Possible mechanism of this inhibition is induction of apoptosis.

Relationship Between Methylation of Tumor Suppressor Gene and Gastric Cancer / 中国普外基础与临床杂志

Objective To investigate the relationship between methylation of tumor suppressor gene and gastric cancer. Methods The literatures in recent years about the concept of methylation, its biological significance and the relationship between DNA methylation/demethylation and gastric cancer were reviewed. The effects of methylation of different tumor suppressor genes on gastric cancer were also analyzed. Results The effect of aberrant methylation on the development and the progression of gastric cancer was still unclear but it was supposed that the inactivation of genes related with cell cycle regulation, mitotic checkpoint, apoptosis, DNA mismatch repair, metastasis suppression and so on might be attributable to the aberrant methylation in gastric cancer. Conclusion Aberrant methylation of tumor suppressor genes plays an important role in the development and progression of gastric cancer. The status of methylation of tumor suppressor genes may be used as a useful molecule marker for diagnosis, assessing metastasis and evaluating prognosis, and demethylation could possibly be a new therapy for gastric cancer.

Effect of 5-Aza-2′-deoxycytidine on experimental lung metastasis of breast cance / 中国普通外科杂志

Objective To investigate the effect of 5-Aza-CdR on experimental lung metastasis of breast cancer and the possible mechanisms.Methods MDA-MB-231 cells were divided into control group and 5-Aza-CdR-treated group.The mRNA expressions and promotor methylation status of BRMS1 and CXCR4 of the MDA-MB-231 cells were evaluated by SqRT-PCR and MSP respectively.Then,the MDA-MB-231 cells of control group and 5-Aza-CdR-treated group were injected into BALB/c nude mice through lateral tail veins,respectively.five weeks later,the mRNA abundance of the target gene HPRT and internal control gene GAPDH of the lung tissues from the mice were evaluated by FqRT-PCR.Results 5-Aza-CdR upgraded the BRMS1mRNA expression significantly than that in control group(0 versus 0.39?0.001,P0.05) and the status of unmethylated CXCR4 CpG island 1 in promotor were not changed significantly.The Ct values of HPRT and GAPDH in control and 5-Aza-CdR-treated groups were 24.75?1.55,16.19?0.69 versus 27.61?1.67,17.48?0.96 respectively,2-??Ct=0.34.The mRNA abundance of the HPRT was lower and there were fewer metastases in the lungs of 5-Aza-CdR-treated group compared with control group.Conclusions 5-Aza-CdR can activate tumor metastasis suppressor gene BRMS1 by demethylation mechanism,and thus,decreased the ability of breast cancer MDA-MB-231 cells for experimental metastasis to lungs.