OBJECTIVETo examine the incidence of congenital heart disease (CHD) in children aged 0-3 years in the rural areas of Chongqing, and to determine the suitable "screening-diagnosis-follow-up" system and screening indicators for CHD in these areas.

METHODSChildren aged 0-3 years from rural areas of the Fuling Disctrict of Chongqing were selected by cluster sampling. Using the "screening-diagnosis-evaluation system" employed at the levels of village/town, district/county, and province/city, the children were screened for seven indicators, i.e., family history of CHD, dyspnea, cyanosis, unique facial features, other congenital malformations, heart murmurs, and blood oxygen saturation (SpO<95%). Children who were positive for one or more indicators accepted echocardiography (ECG) for the diagnosis of CHD. CHD patients were evaluated for disease progression, given guided treatments, and followed-up by pediatric cardiologists.

RESULTSScreening was performed for 10 005 out of the 10 281 children enrolled in the study (97.32% response rate). Among the 175 children who were positive for the indicators, 166 underwent ECG and 60 (0.6‰) were diagnosed with CHD, including 46 cases of simple CHD (76.65%), 11 cases of combined CHD (18.33%), and 3 cases of complex CHD (5.00%). Of the 7 screening indicators, heart murmur had the largest area under the ROC curve for the diagnosis of CHD. In addition, a combination of screening indicators (heart murmur, unique facial features, and other congenital malformations) was most effective for screening out CHD. The CHD patients were given surgical or intervention treatments, and followed up for 6 to 18 months. Ten patients improved without treatment, 13 patients received interventional or surgical treatment, 1 patient died of non-cardiac reasons. The remaining 36 patients were subjected to further follow-up.

CONCLUSIONSHeart murmur alone and in combination with unique facial features and other congenital malformations are valuable tools for CHD screening in children aged 0-3 years. The "village/town-district/county-province/city" screening-diagnosis-evaluation systems are useful for the early detection, diagnosis, and treatment of CHD in infants and young children from the rural areas of Chongqing.

Child, Preschool ; Female ; Follow-Up Studies ; Heart Defects, Congenital ; diagnosis ; Heart Murmurs ; diagnosis ; Humans ; Infant ; Infant, Newborn ; Male ; Oxygen ; blood

OBJECTIVETo explore the application value of morphology assessment of sperm from fresh semen in routine in vitro fertilization (IVF).

METHODSWe analyzed the morphology of the sperm from fresh or optimized semen samples and, based on the sperm morphology of the raw semen, allocated 908 IVF cycles due to the pure tubal factor to different groups: morphologically normal sperm (MNS) ≤ 4%, > 4% - ≤ 15%, and > 15% in Trial 1 and MNS ≤ 1%, > 1% - ≤ 2%, > 2% - ≤ 3%, and > 3%-- ≤ 4% in Trial 2. We compared the rates of fertilization, cleavage, high-quality embryo, -blastocyst formation, and pregnancy among different groups.

RESULTSThe total fertilization rate was significantly lower in the MNS ≤ 4% than in the MNS > 4% - ≤ 15% and >15% groups (74.40% vs 78.61% and 80.03%, P < 0.01). Compared with the MNS ≤ 1%, > 1% - ≤ 2%, and > 2% - ≤ 3% groups, the MNS > 3% - ≤ 4% group showed remarkably increased rates of 2PN normal fertilization (77.23%, 78.97% and 78.99% vs 85.47%, P < 0.01), cleavage (95.71%, 96.01% and 97.27% vs 98.73%, P < 0.05), and blastocyst formation (53.85%, 49.01% and 49.55% vs 63.41%, P < 0.01). No statistically significant differences were observed in the rates of clinical pregnancy, implantation, early abortion, live birth, or malformation at birth among different groups (P > 0.05).

CONCLUSIONMNS ≤ 4% affected the total rate of fertilization while MNS ≤ 3% reduced the rate of normal fertilization in IVF. However, even MNS ≤ 1% did not result in fertilization disorder or failure. Therefore, teratozoospermia alone was not an indicator of ICSI and sperm mor- phology assessment had no obvious value for predicting the rates of embryo quality, clinical pregnancy, and live birth in IVF.

Female ; Fertilization in Vitro ; Humans ; Male ; Pregnancy ; Pregnancy Outcome ; Spermatozoa ; cytology

OBJECTIVETo explore the effect of Se-riched soybean peptide (SSP) on antioxidant function in rats of fatty liver caused by high-fat diet.

METHODSForty Wistar rats were divided into 4 groups randomly and fed with standard diet and water (NC), high-fat diet and water (HC), high-fat diet and SSP (0.1 g/d) (SeH), standard diet and SSP (0.1 g/d) (SeN) respectively. After 10 weeks, the rats were killed to investigate the pimelosis level in liver tissues by Sudan III staining and the expression of hepatic GRP78 by immunohistochemical analysis. We also analyzed the changes of liver function, blood lipid, the glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) activity and malondialdehyde (MDA) content in livers and serum.

RESULTSThe pimelosis level, total cholesterol (TC), triglyceride (TG), MDA contents and the expression of GRP78 in HC group were significantly higher than those in NC, SeN, SeH groups. The activities of GSH-Px and SOD in liver and serum were markedly up-regulated in SeH (P < 0.01). There was no significant difference between NC and SeN groups.

CONCLUSIONSSP can improve liver cell injury and the antioxidant functions in rats with fatty liver effectively and decrease the expression of GRP78 in liver.

Animals ; Antioxidants ; metabolism ; Diet, High-Fat ; adverse effects ; Disease Models, Animal ; Fatty Liver ; chemically induced ; metabolism ; Heat-Shock Proteins ; metabolism ; Liver ; drug effects ; metabolism ; Male ; Rats ; Rats, Wistar ; Selenium ; pharmacology ; Soybean Proteins ; pharmacology ; Soybeans ; chemistry

OBJECTIVETo study the effect of chronic intermittent hypoxia-induced inflammatory cytokines and reoxygenation on glucose transporter 4 (GLUT-4) expression in rat skeletal muscles.

METHODSTwenty-four male Sprague-Dawley rats were randomly assigned to blank control group, chronic intermittent hypoxia (CIH) group, and reoxygenation group. At the end of the experiment, fasting blood glucose (FBG), fasting blood insulin (FINS) and serum inflammatory cytokine levels were measured with glucose oxidase-peroxidase, insulin radioimmunoassay and ELISA, respectively. Homeostasis model assessment (IRI) was used to evaluate insulin resistance in the rats, and GLUT-4 protein expression in the skeletal muscles was measured with Western blotting.

RESULTSCompared with the blank control group, CIH resulted in significantly increased fasting blood glucose, blood insulin levels and insulin resistance index (IRI) (P<0.05); fasting blood glucose was significantly elevated in reoxygenation group (P<0.05). Inflammatory cytokines levels (IL-6 and TNF-α) were significantly higher in CIH group than in the blank control and reoxygenation groups (P<0.05), and were higher in reoxygenation group than in the blank control group. GLUT-4 expression in the skeletal muscles was significantly reduced after CIH (P<0.05) but increased after subsequent reoxygenation (P<0.05).

CONCLUSIONCIH can cause increased release of inflammatory cytokines to lower GLUT-4 protein expression in the skeletal muscles, which contributes to insulin resistance in adult rats.

Animals ; Blood Glucose ; Glucose Transporter Type 4 ; metabolism ; Hypoxia ; Insulin ; blood ; Insulin Resistance ; Interleukin-6 ; Male ; Muscle, Skeletal ; metabolism ; Rats ; Rats, Sprague-Dawley ; Tumor Necrosis Factor-alpha ; blood

The receptor-binding domain(RBD) protein of HCoV-NL63 is a major target in the development of diagnostic assay and vaccine, it has a pivotal role in receptor attachment, viral entry and membrane fusion. In this study, we prepared 2 purified recombinant HCoV-NL63 RBD proteins using in E. coli system and identified the proteins by Western blotting. We first optimized codon and synthesized the RL (232-684aa)coding gene, then amplified the RL or RS(476-616aa) coding gene via PCR using different primers . The RL or RS coding gene was cloned into the pM48 expression vector fused with TrxA tag. The RBD (RL and RS) of HCoV-NL63 were expressed majorly as inclusion body when expressed in E. coli BL21pLys S under different conditions. The expressed products were purified by affinity chromatography then analyzed by SDS-PAGE and Western blotting. Our results showed that the recombinant RBD proteins were maximally expressed at 37 degrees C with 0. 8mM IPTG induction for 4h. RL or RS protein with 95 % purity was obtained and reacted positively with anti-sera from mice immunized with the recombinant vaccinia virus (Tiantan strain) in which HCoV-NL63 RL or RS protein was expressed. In conclusion, the purified recombinant RBD proteins(RL and RS)derived from E. coli were first prepared in China and they might provide a basis for further exploring biological role and vaccine development of HCoV-NL63.
Animals ; Coronavirus Infections ; metabolism ; virology ; Coronavirus NL63, Human ; chemistry ; genetics ; metabolism ; Escherichia coli ; genetics ; metabolism ; Gene Expression ; Humans ; Mice ; Mice, Inbred BALB C ; Protein Engineering ; Protein Structure, Tertiary ; Receptors, Virus ; metabolism ; Viral Envelope Proteins ; chemistry ; genetics ; metabolism

OBJECTIVETo establish the method of simultaneous determination of methylcarbamate (MC) and ethylcarbamate (EC) in yellow rice wine by gas chromatography-mass spectrometry (GC/MS).

METHODSMC and EC in yellow rice wine were derived by 9-xanthydrol, and then the derivants were detected by GC/MS; and quantitatively analyzed by D5-EC isotope internal standard method.

RESULTSThe linearity of MC and EC ranged from 2.0 µg/L to 400.0 µg/L, with correlation coefficients at 0.998 and 0.999, respectively. The limits of quantitation (LOQ) and detection (LOD) were 0.67 and 2.0 µg/kg. When MC and EC were added in yellow rice wine at the range of 2.0-300.0 µg/kg, the intraday average recovery rate was 78.8%-102.3%, relative standard deviation was 3.2%-11.6%; interday average recovery rate was 75.4%-101.3%, relative standard deviation was 3.8%-13.4%. 20 samples of yellow rice wine from supermarket were detected using this method, the contents of MC were in the range of ND (no detected) to 1.2 µg/kg, the detection rate was 6% (3/20), the contents of EC in the range of 18.6 µg/kg to 432.3 µg/kg, with the average level at 135.2 µg/kg.

CONCLUSIONThe method is simple, rapid and useful for simultaneous determination of MC and EC in yellow rice wine.

Carbamates ; analysis ; Food Contamination ; analysis ; Gas Chromatography-Mass Spectrometry ; methods ; Oryza ; Urethane ; analysis ; Wine ; analysis

OBJECTIVETo understand the content status of ethyl carbamate (EC) in yellow rice wine and the changes in storage period and shelf life in Zhejiang province.

METHODSA total of 475 samples of yellow rice wine purchased randomly from supermarkets and food stores in Zhejiang province during 2008-2012, and 49 samples collected from manufacturers were measured for EC content. The sample collected from manufacturers by filter sterilization was placed at 4 °C, room temperature and 37 °C for 400 d, respectively;a bottled wine and a wine in bag were bought from market were placed for 400 d in room temperature to conduct shelf life storage test, and measure the content in every point in 2011. The EC of the samples was determinated by gas chromatography-mass spectrometry after the samples were diluted with D5-EC isotope dilution technique, and purified by alkaline diatomite solid phase extraction column.

RESULTSThe overall detection rate of EC was 99% (472/475) in yellow rice wine of Zhejiang province in 2008-2012, the median value was 70-112 µg/kg, the 90th percentile was 190-333 µg/kg, the 95th percentile was 214-393 µg/kg, and the maximum value was 430-515 µg/kg. The content of EC was increased gradually along with the increasing of storage age in commercially yellow rice wine, and the average content of EC were positively correlated with storage age(r = 0.988). The contents of EC in yellow rice wine after sterilization increased from 74 µg/kg to 86 µg/kg, 127 µg/kg and 509 µg/kg at 4 °C, room temperature and 37°C, respectively for 400 d storage, the differences had statistical significance (F = 14.73, P < 0.01). The content of EC in yellow rice wines in shelf life, which stored in room temperature with bottle and bag package, was decreased slightly with increasing storage time in the beginning, from 215 to 184 µg/kg and 196 to 158 µg/kg, respectively, and increased again with increasing storage time after 250 d, with 252 µg/kg and 210 µg/kg in bottle and bag package after 400 d, respectively, the differences had statistical significance (Z = 2.37, P < 0.05).

CONCLUSIONEC is widespread in rice wine, the content of EC was correlated with storage time and temperature.

China ; Oryza ; Urethane ; analysis ; Wine ; analysis

In this report, we developed a HBV infection model in C57BL/6 mouse line by in vivo injection of a recombinant adeno-associated virus 8 vector carrying 1. 3 copies of HBV genome (ayw subtype) (rAAV8-1. 3HBV). We firstly prepared and purified the rAAV8-1. 3HBV and then injected it into three C57BL/6 mice with the dose of 2 x 10e11vg, respectively. HBsAg and HBeAg were assayed in sera collected at different time points post injection. Ten weeks post injection, the three mice were sacrificed and blood and liver tissue were taken for assay. Copies of HBV DNA were detected by real time PCR and the way of HBV DNA replication was identified by PCR. Subsequently, detection of HBV antigen by immunohistochemistry and pathology analysis of liver tissue of mice were performed. The results suggested that expression of HBsAg and HBeAg lasted for at least 10 weeks in mice sera. Among mice injected with rAAV8-1. 3HBV, HBsAg levels were showed an 'increasing-decreasing-increasing' pattern (the lowest level at the 4th week post injection), while HBeAg levels were kept high and relatively stable. HBV DNA copies were 4.2 x 10(3), 3.6 x 10(3), 2.5 x 10(3) copies/mL in sera and 8.0 x 10(6), 5.7 x 10(6), 2.6 x 10(6) copies/g in hepatic tissues of three mice, respectively. We found that the linear 1. 3HBV DNA in the rAAV8-1. 3HBV could self form into circular HBV genome and replicate in livers of HBV transfected mice. HBsAg and HBcAg were both positive in liver tissue of mice injected with rAAV8-1. 3HBV and no obvious pathological characters were found in liver of mice injected with rAAV8-1. 3HBV. In conclusion, we successfully developed a HBV chronic infection model in C57BL/6 mouse line by in vivo transduction with the recombinant virus rAAV8-1. 3HBV, in which HBV genes could be continuously expressed and replicated over 10 weeks, and paved a way for further characterization of the human chronic hepatitis B virus infection and evaluation of vaccine and anti-HBV agents.
Animals ; Dependovirus ; genetics ; metabolism ; Disease Models, Animal ; Gene Dosage ; Genetic Vectors ; genetics ; metabolism ; Genome, Viral ; Hepatitis B virus ; genetics ; physiology ; Hepatitis B, Chronic ; virology ; Humans ; Mice ; Mice, Inbred C57BL ; Transduction, Genetic ; Virus Replication

To develop a HBV infection mouse model by hydrodynamic-based transfection and further to optimize the method of development of HBV infection mouse model. We first developed a construct which contained inverted terminal repeat elements (ITR) of adeno-associated virus (AAV) and 1. 3 copies of HBV genome (ayw subtype). The pAAV-HBV1. 3 DNA was then injected hydrodynamically into the tail veins of C57BL/6 mice in 5 seconds. The virus load in serum and liver was assayed by ELISA and Real-time PCR. The expression of virus antigen and the pathologic changes of liver were analyzed by HE and immunohistochemical staining. Meanwhile, to develop HBV transfected immunosuppressied mouse, mice were injected intraperitoneally triple with 0.2 ml dexamethason (50 mg/kg) every two days before HBV transfection. The levels of HBsAg and HBeAg were assayed by ELISA. Our data showed: (1) HBsAg and HBeAg were positive (100%) in serum and liver of experimental normal mouse at day 10 after HBV transfection, and became negative at day 30 and day 60. Meanwhile the viral load in serum and liver in experimental group was significantly higher than that in control group at day 10, 30 and 60 after HBV transfection (P < 0.01, P < 0.05, respectively). (2) HBsAg and HBeAg in serum in immunosuppressed mouse model were positive until 60 days. In conclusion, a HBV infection mouse model was developed successfully by hydrodynamic-based transfection. By suppressing the immune status of mice injected with dexamethasone, the expression of HBV antigens was extended longer than that in normal adult mice. These models pave a way for HBV research and evaluation of HBV vaccine and drug development.
Animals ; Dependovirus ; genetics ; metabolism ; Dexamethasone ; administration & dosage ; immunology ; Disease Models, Animal ; Female ; Gene Expression Regulation, Viral ; drug effects ; Genetic Vectors ; genetics ; metabolism ; Hepatitis B ; immunology ; virology ; Hepatitis B Antigens ; genetics ; metabolism ; Hepatitis B virus ; drug effects ; genetics ; physiology ; Humans ; Immunosuppressive Agents ; administration & dosage ; immunology ; Liver ; immunology ; virology ; Mice ; Mice, Inbred C57BL ; Transfection ; methods

To identify the structure of three related substances in potassium sodium dehydroandrographolide succinate (PSDS), an HPLC preparation method was used to separate the impurities. These main impurities were identified using LC-ESI/TOFMS, LC-ESI/MSn, NMR, UV and IR. One of the main impurities was a hydrolyzed and oxidized product of PSDS, which has not been reported previouely. The other two impurities were hydrolyzed products of PSDS after losing different succinic acids. The results indicate that PSDS can be easily hydrolyzed and oxidized. It should be stored at cool and dry places.
Andrographis ; chemistry ; Antiviral Agents ; chemistry ; isolation & purification ; Chromatography, High Pressure Liquid ; Diterpenes ; chemistry ; isolation & purification ; Drug Contamination ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Molecular Structure ; Plants, Medicinal ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization