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Objective:To identify the relationship between exposure to mobile phones and other electronic products and the ability development in children.Methods:Retrospective study.A total of 218 children aged 0.5-6.0 years presented to the Department of Health Care, Beijing Children′s Hospital, Capital Medical University form September 2019 to June 2020 for developmental examination were included.Those with nervous system, mental system diseases, endocrine system diseases and premature children were excluded.Their development was evaluated and the information about exposure to mobiles phones and other electronic products were collected. Chi- square test and Fisher′ s exact probability method were used to analyze the relationship between the exposure to mobile phones and other electronic products and the ability development in children.The influencing factors of children′s development in various fields were analyzed by the binary Logistic regression. Results:Abnormal fine movements of children aged 0.5-0.9 years were related to the latest time every night of exposure ( P<0.05). Abnormalities of adaptability, language and social self-care in children aged 1.0-2.9 years and abnormality of language in children aged 3.0-5.9 years were correlated with the number of hours of daily exposure (all P<0.05). Abnormalities of language and social self-care in children aged 1.0-2.9 years and abnormalities of adaptability, language and social self-care in children aged 3.0-5.9 years were correlated with the purposes of exposure (all P<0.05). Binary Logistic regression analysis indicated that the children′s gross motor was affected by the number of hours of daily exposure ( OR=1.868, P<0.05). The children′s fine motor movements were affected by mother′s educational level and the latest time every night of exposure ( OR=1.722, 2.355, all P<0.05). The children′s adaptability was affected by mother′s educational level, child caregivers, the number of hours of daily exposure and the latest time every night of exposure ( OR=1.711, 2.866, 1.895, 1.650, all P<0.05). The children′s speech was affected by the number of hours of daily exposure, the latest time every night of exposure, and the purposes of exposure (telephone or video phone, early education or study)( OR=2.348, 1.806, 0.328/0.350, all P<0.05). The children′s social interaction and self-care delay were affected by mother′s educational level, the number of hours of daily exposure and the purposes of exposure (telephone or video phone, early education or study)( OR=1.647, 2.678, 0.307/0.363, all P<0.05). Conclusions:The adverse effects of exposure to mobile phones and other electronic products on children should be well concerned.The exposure time of developing young children should be strictly controlled to prevent the adverse effects on the nervous system and development in children.For children who were already affected, relevant habits and behaviors should be timely corrected to avoid irreversible damages.
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Objective:To study the effects of breastfeeding within 2 weeks after birth on late-onset sepsis in very low birth weight infants (VLBWI).Methods:From July 2018 to June 2019, clinical data of VLBWI (birth weight <1 500 g) born in our hospital were retrospectively reviewed. According to the proportion of breastfeeding volume in total feeding volume within 2 weeks after birth, the infants were assigned into high-proportion breastfeeding group (breastfeeding >50%), low-proportion breastfeeding group (breastfeeding ≤50%) and formula group. The incidences of late-onset sepsis among the three groups were compared using the chi-square test or Fisher's exact probability method. Logistic regression was used to analyze the effects of breastfeeding within 2 weeks after birth on late-onset sepsis.Results:The incidences of late sepsis in high-proportion breastfeeding group, low-proportion breastfeeding group and formula group were 0.4% (1/216), 8.1% (5/62) and 8.0% (2/25), respectively ( P<0.001). Logistic regression analysis showed that compared with the high-proportion breastfeeding group, the low-proportion breastfeeding group ( OR=17.844, 95% CI 2.005~158.775) and the formula group ( OR=23.261, 95% CI 1.916~282.350) had increased risks of late-onset sepsis. Conclusions:For VLBWI, high proportion breastfeeding (breastfeeding >50%) within 2 weeks after birth may reduce the risk of late-onset sepsis.
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OBJECTIVE@#To study the influence of maternal sex chromosomal abnormalities on the prediction of fetal sex chromosome abnormalities (SCAs) by non-invasive prenatal testing (NIPT).@*METHODS@#Thirty-six pregnant women with a prediction for fetal SCAs by NIPT were verified as false positive after prenatal diagnosis using amniotic fluid samples. With informed consent, these women were subjected to chromosomal karyotyping or copy number variations (CNVs) analysis through high-throughput sequencing.@*RESULTS@#Sex chromosomal abnormalities were found in 8 women, which yielded an abnormal rate of 22.22% (8/36). Among these, 3 had sex chromosome aneuploidies (47, XXX), 4 had sex chromosome mosaicisms, and 1 carried structural chromosomal abnormalities. Reanalysis of the results of NIPT were consistent with the maternal CNVs by large. With the ratio of cffDNA (ChrX)/cffDNA was more than 2, 6 of the eight women were found to harbor sex chromosome abnormalities, and the fetal karyotype was normal. However, with a ratio of less than 2, only 2 of the 38 pregnant women had sex chromosome abnormalities, and 10 of the fetuses were confirmed as positive.@*CONCLUSION@#The presence of maternal sex chromosomal abnormalities can greatly influence the result of NIPT, which may also be an important reason for false prediction for fetal SCAs by NIPT. When NIPT indicates abnormal SCAs, it is necessary to analyze maternal sex chromosomes. The ratio of cffDNA(ChrX)/cffDNA may help to determine the source of abnormal signals.
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Objective To explore the changes in gravida's age and its influences on maternal and neonatal complications under China's two-child policy.Methods This study retrospectively analyzed the clinical data such as adverse gestational complications and fetal condition of 42 771 gravidas delivering at Changzhou Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University from July 2013 to December 2017.According to their age at delivery,they were divided into three groups:the younger maternal age group (1 140 cases,＜20 years),the advanced maternal age group (4 307 cases,≥ 35 years) and the median maternal age group (37 324 cases,≥ 20 and ＜35 years).Chi-square test was used to compare the differences among groups.Cochran-Armitage test was used for trend analysis.The risks of various complications in younger and advanced maternal age groups were analyzed by binary logistic regression analysis.Results (1) The proportion of advanced maternal age pregnancies tended to rise gradually year by year (Z=-9.909,P＜0.001).However,the figure of younger gravidas remained low and presented a downward trend (Z=10.685,P＜0.001).(2) The incidence of pregnant complications in the younger,advanced and the median maternal age groups were 52.8％ (602/1 140),72.3％ (3 116/4 307) and 56.5％ (21 091/37 324),respectively.Compared with the median maternal age group,the advanced maternal age group was at greater risks of premature delivery [9.0％ (3 343/37 324) vs 11.6％ (499/4 307),x2=124.233,P＜0.001],fetal growth restriction (FGR) [0.6％ (218/37 324) vs 1.2％ (50/4 307),x2=20.087,P＜0.001],postpartum hemorrhage [5.7％ (2 120/37 324) vs 7.8％ (336/4 307),x2=31.299,P＜0.05],hypertensive disorders in pregnancy(HDP) [4.2％ (1 561/37 324) vs 8.7％ (376/4 307),x2=180.013,P＜0.001],gestational diabetes mellitus (GDM) [7.6％ (2 845/37 324) vs 15.1％ (650/4 307),x2=280.126,P＜0.001]and placenta previa [1.7％ (621/37 324) vs 3.8％ (165/4 307),x2=97.904,P＜0.001],and the younger maternal age group was at greater risks of HDP [4.2％ (1 561/37 324) vs 5.9％ (67/1 140),x2=4.234,P=0.040],fetal distress [3.5％ (1 325/37 324) vs 5.1％ (58/1 140),x2=7.546,P=0.006],premature delivery [9.0％ (3 343/37 324) vs 15.0％ (171/1 140),22=48.668,P＜0.001] and FGR [0.6％ (218/37 324) vs 1.1％ (12/1 140),x2=4.086,P=0.043].(3) Gestational complications in the younger maternal age group were mainly related to the fetuses such as premature rupture of membranes (PROM) and premature delivery,while the advanced maternal age group had a higher incidence of maternal complications,especially GDM and HDP.(4) Most of the gravidas of advanced maternal age with HDP developed severe preeclampsia (47.9％,180/376),while mild preeclampsia was dominant in the median maternal aged HDP women (45.4％,708/1 561).(5) The advanced maternal age group had higher risk of stillbirth,premature delivery,FGR,placenta previa,GDM,HDP and postpartum hemorrhage [OR(95％CI):1.91 (1.29-2.84),1.33 (1.21-1.46),1.66 (1.21-2.28),2.56 (2.15-3.04),2.39 (2.19-2.61),2.36 (2.11-2.65),1.46 (1.31-1.62);all P＜0.05],but lower risks of fetal distress and PROM [OR(95％CI):0.79 (0.65-0.95) and 0.88 (0.81-0.96);both P＜0.05].The younger maternal age group had a higher risk of premature delivery [OR(95％CI):1.97 (1.61-2.40);P＜0.001],but significant lower risks of PROM and GDM [OR(95％CI):0.77 (0.62-0.95) and 0.05 (0.02-0.16);both P＜0.05].Conclusions Maternal age is closely related to the adverse outcomes of pregnancy.Two-child policy in China will bring about changes in maternal age and composition of pregnant complications.
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Objective To investigate the differences in the expression of vitamin D receptor (VDR) and serum vitamin D levels in subcutaneous adipose tissue between overweight/obese and normal-weight gravidas, and the relationship between these two indicators and gestational diabetes mellitus (GDM). Methods Women with full-term singleton pregnancies who underwent elective cesarean section in Changzhou Maternal and Child Health Care Hospital Affiliated to Nanjing Medical University from January 2015 to April 2017 were enrolled. Among them, there were 70 cases GDM women, including 35 normal-weight (NW-GDM group) and 35 overweight/obese women (OW-GDM group). During the same period, another 70 pregnant women with normal glucose tolerance who underwent scheduled cesarean delivery were selected as the control group, including 35 normal weight women (NW-control group) and 35 obese/overweight women (OW-control group). Fasting blood samples were collected before operation to determine the levels of different biomarkers, including vitamin D, lipid, fasting blood glucose, fasting insulin and adiponectin, and to calculate the homeostasis model assessment-insulin resistance (HOMA-IR). Two subcutaneous adipose tissue samples of the abdominal wall were taken during the operation to detect the expression and distribution of VDR protein with immunohistochemistry. Meanwhile, VDR mRNA transcription level was quantitatively analyzed using real-time fluorescence quantitative polymerase chain reaction. One-way analysis of variance, LSD, Kruskal-Wallis test, Mann-Whitney U test, Chi-square test and logistic regression analysis were used for statistical analysis. ResuLts (1) The body mass index (BMI) of the OW-control group and the OW-GDM group before pregnancy and delivery were all higher than that of the NW-control group and the NW-GDM group [BMI before pregnancy: (29.2±2.9), (29.4±3.8) vs (21.1±2.3) and (21.9±2.0) kg/m2, F=87.766; BMI before delivery: (35.2±3.4), (35.1±4.3) vs (27.9±2.8) and (28.8± 3.3) kg/m2, F=44.827; all P<0.001]. Newborn birth weight and the proportion of diabetic family history in the OW-GDM group were higher comparing to the NW- and OW- control group [(3 893±498) vs (3 501±402) and (3 625±332) g, F=4.751; 22.9%(8/35) vs 5.7%(2/35) and 5.7%(2/35), χ2=7.869; all P<0.05]. (2) In the OW-control group, the fasting insulin level and HOMA-IR were higher and the adiponectin and vitamin D concentration were lower than those in the NW-control group [13.3(12.3-14.5) vs 12.0(10.4-13.3) mmol/L, 2.7(2.4-3.0) vs 2.2(2.0-2.7), (61.8±20.4) vs (74.9±29.3) ng/ml, (21.6±7.2) vs (25.9±7.3) ng/ml; all P<0.05], and similar results were found between the OW-GDM group and the NW-GDM group [15.3(12.3-19.5) vs 12.0(10.1-15.8) mmol/L, 3.4(2.6-4.1) vs 2.6(2.1-3.2), (50.3±22.3) vs (62.1±23.2) ng/ml, (17.1±6.7) vs (20.6±7.9) ng/ml, all P<0.05]. Compared with the NW-control group, the NW-GDM group had higher fasting glucose and lower high density lipoprotein-cholesterol (HDL-C), adiponectin and vitamin D levels [4.6(4.3-5.1) vs 4.3(4.0-4.5) mmol/L, 1.7(1.6-1.9) vs 2.1(1.6~2.4) mmol/L, (62.1±23.2) vs (74.9±29.3) ng/ml, (20.6±7.9) vs (25.9±7.3) ng/ml; all P<0.05]. Compared with the OW-control group, fasting glucose, fasting insulin and HOMA-IR were higher and HDL-C, adiponectin and vitamin D levels were lower in the OW-GDM group [4.7(4.4-5.4) vs 4.5(4.2-4.7) mmol/L, 15.3(12.3-19.5) vs 13.3(12.3-14.5) mmol/L, 3.4(2.6-4.1) vs 2.7(2.4-3.0), 1.6(1.4-1.8) vs 1.9(1.7-2.2) mmol/L, (50.3±22.3) vs (61.8±20.4) ng/ml, (17.1±6.7) vs (21.6±7.2) ng/ml; all P<0.05]. (3)The overall vitamin D deficiency rate during the third trimester of the four groups was 78.6% (110/140), and the figure was 62.8% (22/35), 82.8% (29/35), 77.1% (27/35) and 91.4% (32/35) in the NW-control group, OW-control group, NW-GDM group and OW-GDM group (χ2=8.994, P=0.029), indicating a higher rate in the OW-GDM group than that in the NW-control group (χ2=8.102, P=0.004). (4) VDR was expressed in the nucleus of adipose tissue in all samples and statistic difference in protein expression was found among the four groups. VDR mRNA expression was higher in both GDM subgroups than that in the two control subgroups, and also higher in the two overweight/obese subgroups than in the corresponding normal-weight subgroups. (5)Serum vitamin D level was negatively correlated with fasting blood glucose and pre-pregnancy BMI, and positively correlated with adiponectin (P<0.05). The incidence of GDM was related to family history of diabetes, VDR mRNA, total cholesterol, HDL-C and HOMA-IR. ConcLusions GDM and overweight/obese patients had decreased serum vitamin D level and increased VDR in subcutaneous adipose tissue. These two factors are closely related to GDM.
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<p><b>OBJECTIVE</b>To explore the cause of failure of non-invasive prenatal testing (NIPT) using cell-free fetal DNA from peripheral maternal blood.</p><p><b>METHODS</b>A total of 31 832 cases of NIPT were retrospectively analyzed. The clinical data of pregnant women were analyzed and the outcome of pregnancy was followed up.</p><p><b>RESULTS</b>Among the 31 832 cases, 200 patients have failed for the first NIPT test. Second test has succeeded in 171 (85.9%) of 199 cases, while 28 cases (14.1%) still yielded no effective results. This gave rise for a total failure rate of 0.088%. Of the 28 cases, 11 (39.2%) were due to high content of total free DNA and could not be sequenced, 17 (60.7%) were found to have the fetal DNA content of less than 4%. Among the 171 cases which have obtained a valid result, NIPT showed that there were 4 patients with high risk of trisomy 21, 18 cases with high risk of 18 trisomy and 1 case with high risk of 13 trisomy. Karyotyping analysis of the amniocytic chromosomes has identified 3 cases with 47,XN,+21, 1 case with 46,XN,rob(21;21), 1 case with 47,XN,+18, while the 13 trisomy case was found to be false positive. For the 28 cases with failed NIPT retest, 14 had normal delivery, with no anomaly noticed in the neonates. Nine patients had opted for artificial abortion during middle or late pregnancy due to maternal factors (4 cases) or fetal factors (5 cases). Four patients developed complications of pregnancy. One case was in good condition upon follow-up. Four cases were lost during follow-up. Of the 11 pregnant women who had failed the NIPT test due to high content of total free DNA, 6 (54.5%) had opted for artificial abortion during midterm pregnancy, which was significantly higher than that of pregnant women with low free DNA content (17.6%).</p><p><b>CONCLUSION</b>Failure of NIPT testing should attract attention from researchers. Failure of single NIPT test should not be regarded as a high risk signal for fetal chromosomal aneuploidies. For those where the test has failed again, genetic counseling and strengthened perinatal care should be provided for the pregnant women.</p>
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Objective To investigate the influences of maternal gestational diabetes mellitus (GDM) on amino acid levels in neonates.Methods From June 2016 to May 2017,393 pregnancies diagnosed with GDM in Changzhou Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University and 3 924 normal pregnancies were enrolled in this study.Clinical data of the gravidas and their newborns were collected.Heel blood samples were collected at 72 to 96 hours after birth.Tandem mass spectrometry was performed to detect the levels of 11 amino acids including alanine,arginine,citrulline,glycine,leucine/ isoleucine/hydroxyproline,methionine,ornithine,phenylalanine,proline,tyrosine and valine in neonatal heel blood.Differences in amino acid levels between the two groups were compared by t test.Influences of GDM on neonatal amino acid levels were analyzed by multivariate linear regression.Results Compared with the healthy pregnancy group,neonates in the GDM group had higher levels of methionine [(21.01 ±6.30) vs (19.93±6.47) μmol/L,t=3.159,P=0.002] and phenylalanine [(47.19±9.19) vs (45.78±8.58) μ mol/L,t=3.076,P=0.002],but lower levels of alanine [(280.51 ±64.54) vs (290.15±68.40) μ mol/L,t=2.678,P=0.007],proline [(147.64±30.64) vs (152.36±33.57) μ mol/L,t=2.680,P=0.007],tyrosine [(85.21 ±29.50) vs (90.60± 33.32) μ mol/L,t=3.089,P=0.002] and ornithine [(101.22±28.79) vs (105.83±30.10) μmol/L,t=2.906,P=0.004].Multivariate linear regression analysis showed that GDM was responsible for the increase of methionine (β=0.69,95％CI:0.02 to 1.37,P=0.044) and phenylalanine (β=1.60,95％CI:0.69 to 2.51,P=0.001),and the decrease of tyrosine (β=-4.98,95％CI:-8.42 to-1.54,P=0.005) and ornithine (β=-3.16,95％CI:-6.30 to-0.02,P=0.048) in neonates.Conclusions GDM neonates has increased of methionine and phenylalanine levels and decreased tyrosine and omithine levels.
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OBJECTIVE To assess the performance of non-invasive prenatal testing (NIPT) based on massive parallel sequencing. METHODS A total of 10 275 maternal blood samples were collected. Fetal chromosomal aneuploides were subjected to low coverage whole genome sequencing. Patients with high risks received further prenatal diagnosis. The outcome of all patients were followed up. RESULTS High-throughput sequencing detected 72 pregnancies with fetal autosomal chromosomal aneuploidy, including 57 cases of trisomy 21, 14 cases of trisomy 18, and 1 case of trisomy 13. The positive predictive value for trisomies 21 and 18 were 98.25% and 91.67%, respectively. Comparing its performance in intermediate or high risk pregnancies, advanced maternal age pregnancies and volunteering to test pregnancies, the positive predictive value were 100%, 95%, 90% and 50%, respectively. The follow up result was only 1 case of 21 trisomy false negative with high risk. For the 56 cases of trisomy 21, the high risk group accounted for 55%, advanced maternal age accounted for 29%, the intermediate risk referred to 14%, the volunteering to test group accounted for 2%. CONCLUSION The performance of NIPT for trisomies 21, 18 and 13 was satisfactory. The method can be used for women with advanced gestational age. NIPT has offered an ideal secondary screening method for those with an intermediate or high risk, and can reduce the rate of birth defects.
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[ ABSTRACT] AIM: To investigate the differential expression of human leukocyte antigen-G ( HLA-G) isoforms and its receptors in human monocyte line THP-1 after human cytomegalovirus ( HCMV) infection for exploring the role of HLA-G in HCMV escaping the immune response of the organism .METHODS: THP-1 cells were infected with HCMV Towne strain.The expression of HLA-G isoforms at mRNA and protein levels was determined by RT-PCR and Western blot, respectively.The surface expression of HLA-G and its receptors ILT2/ILT4 and the cell viability were analyzed by flow cytometry.The levels of soluble HLA-G (sHLA-G) and IL-10 were measured by ELISA.RESULTS:After infection of the THP-1 cells with HCMV , no obvious apoptosis in the cells was observed , and the viability of the cells was high .A significant up-regulation of HLA-G1,-G3,-G4 and-G5 at mRNA expression level 1 d after infection was found , while the protein expression of HLA-G1 and HLA-G5 isoforms was mainly detected .The expression of HLA-G/ILT2/ILT4 was evi-dently up-regulated 1 d after infection .The level of sHLA-G was significantly increased 1 d after infection as compared with control group (P<0.01).The expression of IL-10 was obviously up-regulated 1 d post-infection as compared with control group.CONCLUSION:The differential expression of HLA-G isoforms and secretion of the receptors ILT 2/ILT4 and IL-10 in the THP-1 cells are induced after HCMV infection .This study provides experimental evidence for evaluating the immune mechanism of HCMV infection .
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Objective To investigate the induction and regulatory mechanism of placental trophoblast cell autophagy in women with preeclampsia (PE).Methods Twenty gravidas with severe PE who underwent cesarean section in the Department of Obstetrics and Gynecology of Changzhou Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University from August 2016 to November 2016 were enrolled in PE group.An equal number of normotensive gravidas without proteinuria who also underwent cesarean section during the same period were randomly selected as control group.Placental tissue samples were collected from all gravidas.Ultrastructure of placental trophoblast cells and changes in autophagosome formation were observed by transmission electron microscope.Expressions ofmicrotubule associated protein 1 light chain 3B (MAP1LC3B,or LC3B) and Beclin 1 in placental tissue samples were detected by quantitative real-time polymerase chain reaction (PCR) and Western blot.Activities of protein kinase B (PKB,also known as Akt)/mammalian target of rapamycin (mTOR) pathway in placental tissue samples were detected by Western blot.Two independent samples t-test or Mann-Whitney U test was used for statistical analysis.Results Sparse and disordered villi and many typical autophagosomes were observed in placental trophoblast cells from patients with severe PE.Significantly enhanced expression of LC3B at mRNA and protein levels and increased ratio of LC3-Ⅱ/LC3-Ⅰ were observed in the PE group as compared with the control group [3.37 (2.37-6.11) vs 0.62 (0.25-4.15),1.40±0.17 vs 1.00±0.13,1.57±0.25 vs 1.00±0.31,Zor t=--4.440,3.274 and 3.113,all P＜0.05].No significant difference in the expression ofBeclin 1 at mRNA or protein level in placental tissues was found between the two groups (both P＞0.05).Furthermore,Akt and mTOR phosphorylation in the PE group was significantly suppressed as compared with that in the control group (1.00±0.29 vs 0.64±0.21,1.00±0.32 vs 0.60±0.22,t=--3.672 and-2.895,both P＜0.05).However,the two groups showed no significant difference in the expression of Akt or mTOR protein (both P＞0.05).Conclusions Suppressed activity of Akt/mTOR pathway and enhanced induction of trophoblast cell autophagy are detected in placental tissues of patients with severe PE,indicating that excessive trophoblast cell autophagy,induced by decreased activity of Akt/mTOR pathway,may be the pathogenesis for PE.
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Objective: To investigate the inhibitory effect of T-2 toxin on proliferation of the human hepatocellular carcinoma HepG2 cells and its promotion effect of apoptosis.Methods:The HepG2 cells in the logarithmic phase were selected and divided into control group (without T-2 toxin)and experimental groups (given 0.25,2.50,25.00,250.00 and 2500.00 μg·L-1 T-2 toxin).After 24 h treatment,the morphology of cells was observed under inverted microscope;the inhibitory rate of proliferation of cells was determined by MTT assay;the cell cycle and apoptotic rate of cells were analyzed by flow cytometry;Hochest 33258 staining was used to observe the apoptotic morphology of cells;the activity of caspase-3 in HepG2 cells was detected.Results:After treated for 24 h,the inverted microscope observation results showed that the number of the cells in experimental groups was decreased significantly and the cells shrank and deformed.The MTT results showed that compared with control group,the inhibitory rates of proliferation of the cells in experimental groups were increased (P <0.01).The flow cytometry results showed that compared with control group, the percentage of the cells in SubG1 phase in experimental group was significantly increased,and the apoptotic rates of the cells in experimental groups were significantly increased.The Hoechest 33258 staining results showed that the chromatin condensation was observed in the cells in experimental groups,and the nuclei were dense and stained.Compared with control group,the activities of intracellular caspase-3 of the cells in experimental groups were significantly increased (P < 0.01 ). Conclusion:T-2 can inhibit the proliferation of human hepatocellular carcinoma HepG2 cells,and induce the apoptosis.
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Objective:To express the glycoprotein D of herpes simplex virus type 2 (gD2) in the insect cells,and to determine its immunogenicity.Methods:HSV-2 genome was used as the template for amplification of gD2 extracellular domain fragment gene by PCR.The PCR product was inserted into the vector Bacmind,and the constructed recombinant plasmid gD2-Bacmind was transfected into the sf9 cells to package the recombinant baculovirus.The Sf9 cells were infected by recombinant baculovirus seed derived from the forth passage(P4),the titer of P4 recombinant baculovirus was detected by a plaque assay and the expression of recombinant protein gD2 was determined by Western blotting method.The supernatant of infected cells was collected and purified by Ni-NTA affinity chromatography to obtain the target protein gD2,the purified gD2 protein was used to immunize the BALB/c mice in 0, 2, 4 weeks (gD2 group),and PBS was used as negative control(PBS group);the titers of gD2 specific IgG in serum were detected by ELISA assay.Results: The PCR analysis and sequencing results proved that gD2-Bacmind was constructed correctly.The titer of recombinant baculovirus was 2.0×109 pfu·mL-1,the purified gD2 was about 37 000 with expectation,the percentage of gD2 in total protein was 90%.The average value of Log10 of titer of gD2 specific IgG in serum detected by ELISA assay in gD2 group at the sixth week was 4.34,and there was significant difference compared with PBS group(P<0.01).Conclusion: The gD2 expressed by insect-baculovirus expression vector system has the immunogenicity and can be selected as candidate protein for HSV-2 vaccine.
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Objective:To design the small ubiquitin modification-fibroblast growth factor receptor 4 (SUMO-FGFR4) fusion gene and construct the expression vector pET22b-SUMO-FGFR4, to optimize the expression conditions. Methods:The SUMO-FGFR4 fusion gene was obtained by Overlap PCR and was connected to pET22b;the recombinant expression vector pET22b-SUMO-FGFR4 was obtained. The influence of lactose concentration, induction time,induction temperature,induction point and adding mode of lactose in the expression levels was observed,and the best induction condition was determined; then the solubility of recombinant protein was analyzed.Results:The SUMO-FGFR4 fusion protein was highly expressed,the molecular weight of the fusion protein was about 40 000 and it could bind with FGFR4 specific antibody.When the lactose concentration was 1.0 g·L-1 ,the induction time was 3 h,the induction temperature was 37℃,the value of A (600)was 0.8,the expression level was highest;but adding mode of lactose had no remarkable effect on the protein expression.The expression level of recombinant protein induced by lactose was higher than IPTG.SUMO-FGFR4 protein existed in a form of inclusion body.Conclusion:The SUMO-FGFR4 fusion protein is expressed successfully in this study while lactose is used as inducer and the best expression conditions are confirmed.
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@#An analysis of low molecule weight heparins(LMWH)patents throughout China and aroud the world is made. The suggestion, in terms of patent technical solution, is given to the R&D of LMWH, especially the enoxaparin, dalteprin and nadroparin.
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Objective To investigate the incidence of subclinical hypothyroidism (SCH) during the third trimester of pregnancy and its effects on pregnancy outcomes and neonatal hypothyroidism.Methods A total of 10 695 women in the third trimester of pregnancy (28-42 weeks of gestation) who labored from January 1,to December 31,2012 in Changzhou Maternal and Child Health Care Hospital Affiliated to Nanjing Medical University were enrolled.The levels of thyroid stimulating hormone (TSH),free thyroxine (FT4) and thyroid peroxidase antibody (TPOAb) of the mothers were quantified by electrochemical immunoassay.The time-resolved fluorescence immunoassay was used to detect neonatal thyroid hormone levels.Using t test or x2test,the incidence of adverse pregnancy outcomes was compared between SCH group and euthyroid group and between SCH women with positive (n=40) or negative TPOAb (n=176).Results The incidence of SCH was 2.02％ (216/10 695) and the positive rate of TPOAb in SCH women was 18.5％ (40/216).No neonatal thyroid dysfunction was found.According to the age matched,222 cases were randomly selected as controls from 7 757 euthyroid women.Compared with the controls,SCH women had a higher incidence of premature rupture of membranes [28.7％ (62/216) vs 14.9％ (33/222),x2=12.34],anemia [11.6％ (25/216) vs 4.1％ (9/222),x2=8.65],pregnancy-induced hypertension [9.7％ (21/216) vs 4.5％ (10/222),x2=4.53],premature labor [8.8％ (19/216) vs 3.6％ (8/222),x2=5.10] and intrahepatic cholestasis of pregnancy [8.3％(18/216) vs 2.3％ (5/222),x2=8.14] (all P＜0.05).The overall incidence of adverse pregnancy outcomes was also higher in SCH group than in the controls [69.4％ (150/216) vs 49.5％ (110/222),x2=17.96,P＜0.01].The incidence of fetal growth restriction and still birth in SCH mothers with positive TPOAb was higher than in those with negative TPOAb [7.5％ (3/40) vs 0.0％ (0/176),x2=13.32,P＜0.01; 2.5％ (1/40) vs 0.0％ (0/176),x2=4.40,P＜0.05],but there was no significant difference in the overall incidence of adverse pregnancy outcomes compared with TPOAb-negative mothers [65.0％ (26/40) vs 70.5％ (124/176),x2=0.46,P=0.50].Conclusions SCH diagnosed in the third trimester may lead to adverse pregnancy outcomes.Early screening for thyroid dysfunction is necessary.
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Objective: To observe curative effect of thoracic sympathetic block on patients with diabetes mellitus (DM) complicated cardiac chamber enlargement and heart failure (HF). Methods: A total of 76 DM patients with cardiac chamber enlargement and HF were randomly divided into routine treatment group (n=36) and epidural nerve block group (nerve block group, n=40, received thoracic epidural nerve block based on routine treatment), and four weeks were a course of treatment. Changes of left ventricular ejection fraction (LVEF), left ventricular end-diastolic dimension (LVEDd), left atrial diameter (LAD), levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), brain natriuretic peptide (BNP) and glycosylated hemoglobin (HbA1c) were observed in two groups. Results: There were no significant difference in all indexes between two groups before treatment (P>0.05). After treatment, total effective rate of cardiac function improvement in nerve block group was significantly higher than that of routine treatment group (95% vs. 66.7%, P<0.05). Compared with before treatment, there were significant decrease in LVEDd [(71.63±9.26)mm vs. (65.4±8.12)mm] and LAD[(45.62±7.10)mm vs. (37.14 ±6.28)mm] (P<0.05 both), significant increase in LVEF [(31.3±11.46)% vs. (42.12± 10.42) % , P< 0.01], and significant decrease in levels of TNF-α [(73.55±16.36)ng/L vs. (28.35±10.08) ng/L], IL-6 [(17.62±5.47) ng/L vs. (7.88±2.29) ng/L], BNP[(13.49±2.94) ng/L vs. (6.28±2.28) ng/L] and HbA1c [(7.9±2.6) % vs. (6.8±1.9) %] in nerve block group after treatment, P<0.05 or <0.001; and levels of LVEF, TNF-α, IL-6 and BNP in nerve block group after treatment were significantly improved than those of routine treatment group, P<0.05 or <0.01. Conclusion: Thoracic sympathetic block can significantly decrease levels of neuroendocrine and inflammatory factors, improve cardiac function, and elevate quality of life in patients with diabetes mellitus complicated heart failure.
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Objective To explore the relationship between the HLA-G 14 bp insertion/deletion polymorphism and the infection of Epstein-Barr virus(EBV) for children.Methods The study genotyped HLA-G 14 bp insertion/deletion polymorphism of 102 infectious mononucleosis children and 165 normal controls by PCR-PAGE,detected the plasma sHLA-G level of 51 infectious mononucleosis children and 146 normal controls by ELISA.Results A significant difference was observed for the frequencies of the HLA-G 14 bp genotype between the two groups( x2 =6.742,P=0.034 ),and a significant difference was also observed for the 14 bp allele frequencies between the two groups( x2 =6.672,P=0.01 ).The plasma sHLA-G levels in the infectious mononucleosis children were dramatically higher than that in normal controls,and a significant difference was observed between the two groups( Z=-9.472,P＜0.01 ).Among the infectious mononucleosis children,levels of sHLA-G was find a significant difference between the three genotypes of HLA-G 14 bp insertion/deletion polymorphism( H=6.09,P =0.048 ),and the level of s HLA-G with 14 bp+/+ genotype was markedly lower than that of the two other genotypes (Z=-2.376,P=0.01 8).Conclusion There was a relationship between the HLA-G 14 bp insertion/deletion polymorphism and the susceptibility to the infectious mononucleosis for children.Children who carried the 14 bp-/- genotype or deleted the 14 bp allele may have a significantly increased risk of the infection of EBV.The plasma sHLA-G might be considered as an index for auxiliary diagnosis infectious mononucleosis.
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Objective To establish a method for detection of gene mutations in β-thalassemia by high-resolution melting (HRM) and study its preliminary clinical application.Methods Two common mutations [ IVS-2-654 ( C ＞ T ) and -28 ( A ＞ G ) ]of β-thalassemia in Wenzhou city population were selected.The plasmid DNA fragments of these mutations were constructed by TA clone technology as PCR templates or genotyping controls.A method for detection of β-thalassemia gene mutations based on HRM analysis was established and its specificity,sensitivity and repeatability were methodologically evaluated.One hundred and seventeen patients with clinically suspected β-thalassemia from Second Affiliated Hospital and Yu ying Children's Hospital of Wenzhou Medical College were enrolled into this study.The genomic DNA was extracted from whole blood cells and detected by HRM method.The results were compared with the direct sequencing data.Results HRM method could detect the mutations [ IVS-2-654( C ＞ T) and -28 ( A ＞ G ) ]of β-thalassemia and the results did not show any non-specific amplified fragments.All within-run and between-run coefficients of variation for different DNA types' Tm were smaller than 0.1％.And minimum 103 copies of DNA of each assay and 10％ mutation could be determined by this method.One hundred and seventeen patients with clinically suspected β-thalassemia were detected with HRM and all the results were in accordance with direct DNA sequencing.There were 45 IVS-2-654 ( C ＞ T)heterozygous mutation and 9-28 ( A ＞ G)heterozygous mutation and none homozygous mutation.Conclusion The method of rapid identification of β-thalassemia gene mutations based on HRM analysis is successfully established,which is a convenient,rapid,specific,sensitive and accurate technique for screening gene mutations in β-thalassemia as well as a general technical platform to identify other gene mutations.
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Purpose To examine the regulatory effect of recombinant human fibroblast growth factor-21 on the expression of liver X receptor α and glucose transporter protein 1 in the type 2 diabetes mellitus rats.Methods The rat models of type 2 diabetic mellitus were divided into four groups at random, ic. rhFGF-21 every day, after eight weeks of these treatment, Inspect the fasting blood glucose (FBG), fructosamine(FA), triglyceride(TG), T-cholesterol(TC), high density lipoprotein cholesterol(HDL-C) and low density lipoprotein cholesterol(LDL-C) of these rats, then detecting the mRNA expression of LXRα and GLUT1 by RT-PCR.Results (1) rhFGF-21 can reduce blood glucose steadily to near normal levels in diabetic rats. (2) The expression of LXRα and GLUT1 level was significantly higher in the rhFGF-21 treatment group than that in the model group. (3) rhFGF-21 megadoses and middle doses decreased FA, TG, TC,and LDL-C and elevated HDL-C.Conclusion rhFGF-21 could regulate the mRNA expression of LXRα and GLUT1 in diabetes rats, increase basal level glucose transport, then reduce blood glucose, improve lipid metabolize dysfunction.
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Objective To evaluate the effect of trimetazidine(TMZ) and metoprolol combining the other regular medicine on the stable angina peetoris patients with type 2 diabetes mellitus. Methods 78 patients with type2 diabetes mellitus and the patients with no less than two months stable and revoked-angina pectoris by effort were colleeted. They were divided into two groups. In one group,20mg of TMZ was used three times a day. In the other group,25mg metoprolol was used and two times a day. The two groups all combined with the regular medicine,bamyl, caleiume-antagonist and long-efficency nitrate and rutine antilipemic, antidiabefie agents for ten weeks and analyzed the statistical difference of two groups. Meanwhile some parameter were observed including the heart rate,blood pressure, blood glucose, TG, TC, HDL-C and LDL-C during the treatment. Results In two groups, cardiac pectoris frequency and lasting-time and the used-nitroglycerin decreased significantly, while the time of next angina outcome and left ventrieular ejection fraction(EF% ) increased significantly. The therapy hasen't great differene(P>0.05) in two groups. But in TMZ group, HR, bllod pressure, blood glucose and blood fat haven't great difference(P > 0.05) before ad after thereapy. In metoprolol group the difference is significant(P < 0.05, P < 0.01) and twogroups have great difference(P < 0.05). Conclusion The results suggest that the TMZ is effect and safe on the sta-ble angina peetoris patients with type 2 diabetes mellitus.