Response surface analysis (uniform precision of central composite design, SAS 9.1.3 software) was applied to optimize the four major factors (ratio of soybean meal to water, enzyme quantity, fermentation time and inoculation quantity) for soybean meal solid fermentation. According to the change of the hydrolyzation degree of soybean protein, the equation of polynomial regression was established between those factors and the response. The result showed that the optimum condition included as follows: ratio of soybean meal to water 1∶1.00,enzyme quantity 2.55%, fermentation time 65h and inoculation quantity 1.00%. Under the optimum level, the degree of hydrolyzation reached 13.3%, which increased 56% over pre-optimization.

Characters of one Candida intermedia yeast strain which isolated from nature can produce ethanol from xylose-fermenting been systemic studied. In conditions 28?C, 120 r/min, 72 h, it can produce 6.480 g/L ethanol from 7% xylose and 43.70% theoretical production of ethanol from 3% xylose. It can produce up to 21.225 g/L ethanol when incubation time prolong to 156 h from 8% xylose. It also can ferment 13% glucose produce 47.647 g/L ethanol and reach 76.90% of theoretical ethanol production, respectively. Compared to CK, ethanol productivity can be improved 9.91% when add 8% xylose in three times as 3%, 2% and 3%, respectively. Glucose can be first utilized in the mixture sugar medium. When the ratio of xylose vs. glucose is 3:1in mixture sugar, the productivity of ethanol can be improving 25%.

Adolescent ; Adult ; Cleft Lip ; surgery ; Female ; Humans ; Lip ; injuries ; surgery ; Male ; Middle Aged ; Postoperative Complications ; Reconstructive Surgical Procedures ; methods ; Surgical Flaps ; Young Adult

Objective To study on transcatheter closure of membranous ventricular septal defect (VSD) with pseudoaneurysm by patent ductus arteriosus(PDA) occlusion devices in children and summarize the skill and clinical experience. Methods The study included 20 membranous VSD cases in children.According to the finding of the left ventricular angiography, various kinds of the PDA occlusion devices was implanted. The mean diameter of the waist of the occluder was ( 10.4 ± 2.6) mm. Examination by transthoracic echocardiography (TTE) immediately and left ventricular angiography after the occluder was implanted 15minutes later to evaluate the efficacy. Results In the 20 patients, one of the Ⅳ type VSD patient was quitted because of the significant residual shunts(≥2 mm). Slightly residual shunts ( ＜ 2 mm) was found in one Ⅲ type VSD patient with multi-outlet. And disappeared in 1 month after the procedure, which VSD patient was confirmed by TTE. Thirteen cases were normal by EKG examination (or the same before procedure).Incomplete right bundle branch block was found in 4 cases. First degree atrioventricular block was found in 1 case and paroxysmal junctional tachycardia was found in 1 case. All of them were recovered in 1 week.Conclusions Transcatheter interventional therapy with PDA occlusion devices for membranous VSD with pseudoaneurysm is safe and effective. The key of the procedure is to select suitable occluder and suitable position to plant them according to the membranous morphologic characteristics,size and position of the pseudoaneurysm. It is a facultative method for transcatheter therapy this kind of congenital heart disease.

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 observe the expressions of C/EBPs mRNA and protein in the sebaceous gland and to study the relationship between C/EBPs and the differentiation of sebocytes. Methods RT-PCR and immunhischemistry were used to detect the expressions of C/EBPs mRNA and protein in the embryo and adult sebaceous glands. Results The lowest expression of C/EBP? mRNA in the sebaceous gland was found in embryonic period, but increased gradually during the developmental stages. The expression of C/EBP? mRNA in the sebaceous gland showed different expression patterns, i.e. it maintained at low level in all of the developmental stages. Expressions of C/EBP? and ? protein were found in the nuclei of sebocytes in embryonic period but in the basal cell layer of sebaceous glands in maturation phase. Conclusion The expression patterns of C/EBPs are different in the sebaceous gland from embryonic to adult stages, suggesting that C/EBP? and ? may play important roles in the development of human sebaceous gland.

Objective:To investigate the characteristics of magnetic resonance imaging and clinical application of multi-modality magnetic resonance imaging (MRI) for evaluating the response of neoadjuvant chemotherapy (NACT) in inflammatory breast cancer (IBC).Methods:A total of 36 IBC patients were enrolled in the study.The morphological, hemodynamic and diffusion-weighted imaging features of MRI were analyzed. Eleven patients underwent MRI examination before and after NAT. The imaging changes were analyzed and the efficacy of NACT was evaluated.Results:There were 38 identified breast carcinoma in these 36 cases, among which abnormal skin thickening and enhancement, extensive edema was found in 37 breast lesions. Enhancement of breast lesions in 25 cases was non-mass-like enhancement. Diffusion limitation was found in all lesions. The number of vessels in affected side was more than that in healthy side in MIP images. Thirty three cases had axillary lymph node enlargement.The sensitivity and specificity of MRI in evaluating residual breast tumors and vascular thrombus were high, but the evaluation of axillary lymph nodes was relatively low.Conclusions:Multi-modal MRI can be used for early and accurate diagnosis of IBC. It can also be used to predict and evaluate the effect of neoadjuvant chemotherapy.

Objective To examine the complement component 1 Q subcomponent-binding protein (C1QBP) gene expression in human resistance choriocarcinoma cell lines and its parental cell line JeG-3,and to investigate whether silence C 1QBP by small interference RNA could reverse the resistance of human resistance choriocarcinoma cell lines to its relevant chemotherapy drugs.Methods Expression of C1QBP mRNA and protein in cells were detected by real-time fluorogenic quantitative PCR and western blot,respectively.The difference of C 1QBP expression was compared between human resistance choriocarcinoma cell lines and its parental cell line JeG-3.Sub-cellular location was proved by confocal immunofluorescence microscopy.A lentiviral vector containing short hairpin RNA (shRNA) targeting C 1QBP was constructed and cotransfected with the packaging plasmid mixture into 293T cells by lipofectamine 2000.The human resistance choriocarcinoma cell lines were infected with the packaged lentivirus.Real-time fluorogenic quantitative PCR and western blot were used to validate whether the C 1QBP gene expression was silenced.The cell counting kit 8(CCK8)was used to determine the drug sensitivity.Results (1)The C1QBP mRNA expression levels among four human resistance choriocarcinoma cell lines[JeG-3/floxuridiuum (FUDR),JeG-3/methotrexate (MTX),JeG-3/etoposide (VP),JeG-3/dactinomycin (KSM)] were 2.520±0.680,1.770±0.230,1.940±0.090 and 1.740±0.350 folds compared to that in JeG-3 cells.The C1QBP protein was higher expression level in human resistance choriocarcinoma cell lines than that in JeG-3.The immunofluorescence methods and confocal analysis showed that C1QBP localized predominantly in the mitochondrial matrix.(2)The C1QBP mRNA expression in JeG-3/FUDR cells after infected with lentiviral vector were decreased by 93.1％ (P＜0.01).The protein expression of C 1QBP in JeG-3/FUDR cells after infected with lentiviral vector were almost completely suppressed.The resistance indexes of four human resistance choriocarcinoma cell lines(JeG-3/FUDR,JeG-3/MTX,JeG-3/VP,JeG-3/KSM) were respectively 86.3％,93.9％,92.8％ and 89.9％,which were decreased remarkably by knockdown the C 1QBP expression (P＜0.05).Conclusions C1QBP is overexpressed in human resistance choriocarcinoma cell lines compared with parental cell line JeG-3.Inhibition of C 1QBP by lentivirus-mediated small interference RNA could effectively reverses the resistance of human resistance choriocarcinoma cell lines to its relevant chemotherapy drugs.

ObjectiveTo observe the growth of BT-325 human glioma cells after interfering volume-regulated chloride channel ClC-2 gene.MethodsTwo expression recombinant vectors of ClC-2 gene were designed and constructed. The primary plasmid, pSUPER.puro-shRNA, and the two recombinant plasmids, pSUPER.puro-shRNA-ClC-21 and pSUPER.puro-shRNA-ClC-22, were transfected into BT-325 cells by LipofectamineTM2000 (Groups: control, PP1 and PP2, respectively). The mRNA expression of ClC-2 gene was detected with reverse transcription polymerasse chain reaction (RT-PCR), the cellular survival was determined with MTT assay, and the cell cycle was measured with flow cytometry (FCM). ResultsClC-2 mRNA expression and the growth of the cells in PP1 and PP2 groups were significantly lower than that of control group. The cell cycle progression was blocked in G1 phase (PP1 and PP2 vs control,P<0.01). ConclusionThe growth of BT-325 human glioma cells is prevented by knockdown of ClC-2 gene expression, which may be one of the novel targets to inhibit growth of human malignant glioma cells.

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.