Objective To explore the regulation mechanism of autophagy-related protein, microtubule-associated protein 1 light chain 3 (LC3), via c-Jun in methotrexate resistant human choriocarcinoma JEG-3 cell lines. Methods Human choriocarcinoma JEG-3 cell lines, and methotrexate resistant choriocarcinoma JEG-3 (JEG-3/MTXR) cell lines were used in our present study. Phosphorylation c-Jun (p-c-Jun) was evaluated after exposure to 0.02 ng/ml methotrexate for 72 hours in both cells by western blot. c-Jun gene was knockdown by small interference RNA (siRNA) in JEG-3/MTXR cells, and LC3 was evaluated by western blot and reverse transcription-PCR. The binding of LC3 promoter with c-Jun protein was detected via chromatin immunoprecipitation assay (ChIP) with or without 0.02 ng/ml methotrexate exposure. Results The results showed that p-c-Jun was up-regulated after methotrexate treatment for 72 hours (1.99±0.20, versus 0.20±0.06 at 0 hour;P<0.05) by western blot analysis in JEG-3/MTXR cell lines. Further investigation demonstrated that c-Jun-siRNA could inhibit the up-regulation of LC3 formation and after methotrexate exposure (LC3 mRNA:1.24±0.17 versus 3.03±0.43;LC3 protein:0.52±0.07 verus 1.20± 0.15; all P<0.05). The binding of LC3 promoter by c-Jun protein was up-regulated after methotrexate treatment by the method of ChIP in methotrexate resistant JEG-3/MTXR cells [(2.95 ± 0.35) times]. Conclusion Autophagy-related gene LC3 expression regulated by c-Jun protein may be involved in the effect mechanism of the development of methotrexate resistance in choriocarcinoma JEG-3 cells.

Objective To investigate the age distribution and types of human papillomavirus (HPV) cervical infection in patients in Wuhan region, so as to provide scientific basis for the prevention and treatment of HPV infection and cervical cancer.Methods 9 915 exfoliated cervical cell specimens from patients in Wuhan Medical and Health Center for Women and Children between January 2015 and March 2016 were performed HPV genotyping (21 subtypes), clinical data of 1 732 HPV positive cases were statistically analyzed.Results HPV-positive rates in ≤25, 26~, 36~, 46~, and ≥56 age groups were 22.15%, 15.90%, 17.04%, 19.97%, and 17.57%,respectively (χ2=36.587,P<0.01),HPV-positive rates in ≤25 and 46~ age groups were both higher than other age groups.There were significant differences in single infection and multiple infection rates among different age groups respectively(χ2=14.39, 36.51,respectively, both P<0.05),single infection rate was highest in 46~ and ≤25 age groups (15.41% and 15.24% respectively);multiple infection rates was highest in ≤25 and ≥56 age groups (6.90% and 5.86% respectively).The percentage of single infection and multiple infection were 75.58% and 24.42% respectively, the major single infection type was HPV high-risk subtype (84.34%),the major multiple infection types were high-risk and high-risk compound subtype as well as high-risk and low-risk compound subtype, accounting for 60.52% and 38.77% respectively.The main single infection types were HPV high-risk subtypes 33, 68, 31 and 16, as well as and low-risk subtype 11, the ratios of single infection to multiple infection were 3.13, 2.03, 1.71, 1.67 and 2.00 respectively.Conclusion Cervical infection rates in women in Wuhan region is high in ≤25 and 46~ age groups, there are differences in the distribution of different HPV subtypes of single infection and multiple infection.

Background Posterior capsular opacification(PCO) is common complication after extrecapsular extract of cataract.Matrix metalloproteinases-3 (MMP-3) can degrade all the extracellular matrix except polyose.The gene therapy of PCO upon MMP-3 is the researching hot topic.Fibronectin ( FN ) is a degrade gelatin,so its expression can reflect the effect of MMP-3 on LECs indirectly. Objective The aim of this study was to construct MMP-3 eukaryotic recombination plasmid and transfect to lens epithelium cells(LECs) for the observation of MMP3 expression,and to explore the feasibility of gene therapy for after cataract. Methods Six fresh lenses were obtained from pigs.LECs were cultured using explant method.The eukaryotic expression vector pEGFP-N1-MMP-3 was reconstructed with MMP-3 and pEGFP-N1 plasmids.The accuracy of MMP-3 gene fragment was confirmed by double enzyme digestion and DNA sequencing analysis.After transfecting pEGFP-N1-MMP-3 into LECs of pig,the expression of MMP-3 protein in the cells was indirectly observed by green fluorescent protein.The expression of FN in LECs was detected using Western blot. Results The result of double enzyme digestion was consistent with the base number of pEGFP-N1 plasmids and target fragment.By enlacing the result of DNA sequencing analysis with software,the resemblance of the DNA sequence of MMP-3 from recombination plasmid pEGFP-N1-MMP-3 and that of homo MMP-3 was 99.6％,indicating that the target fragment was inserted to pEGFP-N1 plasmids successfully.Green fluorescence for GFP was seen in the LECs in pEGFP-N1-MMP-3 transfected group,but absent response for GFP was in empty vector group.Western blot revealed that the relative expression level of FN in LECs was 0.666±0.008 in pEGFP-N1-MMP-3 trasfected group and 0.326 ±0.071 in empty vector group,with a significant difference between these two groups(P=0.000). Conclusions Eukaryotic recombination plasmid pEGFP-N1-MMP-3 is successfully constructed,and MMP-3 can be expressed in LECs after transfected.These results lay a foundation for the further research of MMP-3 gene therapy for PCO.

With inlet temperature, specific gravity, feeding speed as independent variables, the comprehensive evaluating indexes of content of schisandrin and arctiin as dependent variable, the experimental data were fitted to a second order polynomial equation. Based on establishing the mathematical relationship between the comprehensive evaluating indexes and respective variables, Box-Benhnken central composite test and response surface analysis method was employed to optimize the spray drying technology of Biqiu granules ethanol extract. The optimal drying parameter was as follows: the inlet temperature was 175 degrees C, the specific gravity was 1.10, feeding speed was 32 r x min(-1). Under these conditions, the comprehensive evaluating indexes of spraying dry processes was 92.68, which was close to the model prediction. The spraying dry technology of Biqiu granules ethanol extract optimized by response surface methodology was accurate and feasible, which provided theoretical experiment basis for the industrialization production.
Chemistry, Pharmaceutical ; methods ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Ethanol

Child ; Female ; Humans ; Menisci, Tibial ; abnormalities ; surgery

First with the qualified rate of granules as the evaluation index, significant influencing factors were firstly screened by Plackett-Burman design. Then, with the qualified rate and moisture content as the evaluation indexes, significant factors that affect one-step pelletization technology were further optimized by Box-Behnken design; experimental data were imitated by multiple regression and second-order polynomial equation; and response surface method was used for predictive analysis of optimal technology. The best conditions were as follows: inlet air temperature of 85 degrees C, sample introduction speed of 33 r x min(-1), density of concrete 1. 10. One-step pelletization technology of Biqiu granules by Plackett-Burman design and Box-Behnken response surface methodology was stable and feasible with good predictability, which provided reliable basis for the industrialized production of Biqiu granules.
Chemistry, Pharmaceutical ; methods ; Drugs, Chinese Herbal ; chemistry ; Temperature

Objective To investigate the relatonahip between TNF-α and ventricular arrhythmias after acute myocardial infarction(AMI)and its mechanism.Method Both the clinical and animal experiments were done.(1)Clinical experiment:Eighty patients with AMI were included in Union Hospital,Tongji Medical College,Huazhong University of Science and Techology,from May 2005 to November 2006 according to the WHO diagnostic criteria.Co-infection of diseases such as severe upper respiratory infection,lung infection,high fever,cancer,et al were excluded.The relationship between the levels of TNF-α and arrhythmias were observed at different times after AMI.A straight line correlation,analysis Was done.(2)Animal experiment:Different concentrations of TNF-αwere added to isohted rat hearts for observing the arrhythrnia effects.The effect of TNF-α on intracellular Ca2+ concentration was detected by laser confocal technique.All data were analyzed by SNK-q test using SPSS 13.0 sofeware prograrn.Results(1)The plasma levels of TNF-α were significantly associated with the Lown class of PVC after AMI and they were higher in AMI of anterior wall[(46.41±10.34)pg/ml]than other positions [(28.25±6.35)pg/ml,P<0.05].2)The frequency of ventricular arrhythmias was interrelated with the concentralions of TNF-α.Using etanercept beforehand,TNF-α induced a slight increase of intracellular Ca2+ intensity (P<0.05).Conclusions There was a relationship between TNF-αlevels and ventricular arrhythmisa in patients with AMI.Animal experiments confirmed the isolated heart perfusion with TNF-α induced ventricular arrhytrnias.Expression of TNF-α after AMI was related with the occurrence of ventricular arrhythrnias.The effect might be associated with the increased inuaeellular Ca2+ intensity caused by TNF-α.

OBJECTIVETo investigate the carcinogenic and mutagenic mechanism of bitumen fume.

METHODSThe experimental mice were forced to inhale the bitumen fume at different exposure level (55 mg/m(3), 165 mg/m(3)) and in different time (30 days, 60 days). The pathological changes of the lung tissue in mice were observed with H.E staining. The content of the DNA in the lung tissue of mice and the cell circles were determined with flow cytometry.

RESULTSThe lesion of the lung tissue in mice comprised the atypical hyperplasia of different levels and the carcinoma in situ with the increase of the containment time and dosage; the cycle index was changed: the number of the G 1 phase cells was decreased, the S phase was retarded, the cells entering the G 2/S phase were decreased, the cell proliferation index (P I) was increased and the heteroploid DNA index (DI) was increased (P < 0.05). The cell index in the 55 mg/m(3) group and the 165 mg/m(3) group was higher than that in the control group when the containment time was same. The heteroploid DNA index (DI) in the 55 mg/m(3) group was significantly higher than that in the 165 mg/m(3) group (P < 0.05 or P < 0.01). When the containment dosage was same, the DI in the 60 days treatment group (1.16 +/- 1.51 x 10(-2), 1.20 +/- 2.3 x 10(-2)) was all significantly higher than those in the 30 days treatment group (1.14 +/- 8.8 x 10(-2), 1.16 +/- 1.47 x 10(-2)) (P < 0.05).

CONCLUSIONThe precancerous lesion in the lung tissue of the mice induced by the bitumen fume may be related with the changes of the cell cycle.

Animals ; Carcinogens ; toxicity ; Cell Cycle ; drug effects ; DNA ; analysis ; Dose-Response Relationship, Drug ; Female ; Flow Cytometry ; Hydrocarbons ; toxicity ; Lung ; drug effects ; pathology ; Male ; Mice ; Mice, Inbred Strains ; Precancerous Conditions ; chemically induced ; pathology

Objective To provide applied anatomy basis for upper fibular bone flap pedieled with cir- cumflex fibular neck artery.Methods The origin,course,branches,distribution and anastomosis of cir- cumflex fibular neck artery were observed on 54 sides of adult cadaver lower limb speeimens perfused with red latex.Results The circumflex fibular neck artery derived from popliteal artery (3.3?1.1)cm beneath the tip level of fibular head,with its length of (1.4?1.0) cm and diameter of (1.4?0.5) mm.The ascending periosteal branches of circumflex fibular neck artery anastomosed consistently with the descending periosteal branches of inferior lateral genicular artery.Conclusion The transplanting operative design of upper fibular bone flap pedicled with circumflex fibular neck artery was feasible.

Objective To determine risk factors for the miss rate of colorectal adenomas during colonoscopy.Methods A total of 981 patients,diagnosed as having at least one polyp in colonoscopy,received a second colonoscopy in 6 months from November 2012 to March 2016.All polyps were removed in the second colonoscopy.Bio-information of patients such as sex,age,surveillance interval and features of polyps such as number,size,shape,location,pathology,withdrawal time,bowel preparation was retrospectively analyzed.Factors associated with the miss rates in these patients were analyzed with Chisquare and was also analyzed with Logistic regression model for multiple factors.Results A total of 981 patients were selected according to the inclusion and exclusion criteria,including 604 males and 377 females.Miss rates of males and females were 38.9％ (235/604) and 27.9％ (105/377) (P＜0.01)respectively.Age ranged from 25 to 87 years with mean age being 61.0±9.7 years.Miss rates of senior patients ＜65 and ≥65 years were 31.5％(195/619) and 40.1％(145/362) respectively (P＜0.01).A total of 1 728 adenomas were found in first colonoscopy.A total of 2 267 adenomas were found in the second colonoscopy.The adenoma miss rate was 23.8％ (539/2 267).The miss rate of adenoma whose size ≤ 5 mm was 42.5％ (311/732);and that of larger size of 6 to 9 mm was 17.8％ (194/1 090);that of even larger size,i.e.,≥10 mm,was 7.6％(34/445)(P＜0.01).Miss rates of Is,Isp,Ip,LST and Ⅱ adenomas in shape were 28.4％(489/1 720),9.3％(24/235),6.6％ (12/182),9.0％(6/67) 20.5％ (8/39) respectively (P＞0.05).Location with highest adenoma miss rate were descending colon,ascending colon and transverse colon,27.8％(64/230),25.5％(120/470),25.5％ (161/632) respectively.Miss rates of high and low risk adenoma were 44.8％ (277/618) and 17.4％ (63/363) (P＜0.01).The highest adenoma miss rate of all the pathology type was tubular adenoma.The adenoma miss rate was 26.9％ (449/1 671) (P＜0.01).Miss rates of good and poor bowel preparation were 30.2％ (271/897) and 82.1％ (69/84) (P＜0.01).Miss rates of adequate and inadequate withdrawal time were 24.3％ (174/717) and 62.9％ (166/264) (P＜0.01).Conclusion Male,old-age,diameter ≤ 5 mm,poor bowel preparation and inadequate withdrawal time,high risk adenoma are the risk factors for missed adenoma.But the shape and location of adenoma are not the risk factors.