Objective To elucidate the impact of over-expression of S100A7 on migration,invasion,proliferation, cell cycle, and epithelial-mesenchymal transition (EMT) in human cervical cancer HeLa and CaSki cells.Methods(1)Immunohistochemistry of SP was used to examine the expression of S100A7 in 40 cases of squamous cervical cancer tissues and 20 cases of normal cervical tissues.(2)The vectors of pLVX-IRES-Neo-S100A7 and pLVX-IRES-Neo were used to transfect human cervical cancer HeLa and CaSki cells, and the positive clones were screened and identified. Next, transwell migration assay, cell counting kit-8(CCK-8)assay and fluorescence activating cell sorter(FACS)were used to detect the effect of S100A7-overexpression on the migration, invasion, proliferation and cell cycle of cervical cancer cells. Furthermore, western blot was performed to observe the expression of epithelial marker(E-cadherin)and mesenchymal markers(N-cadherin,vimentin,and fibronectin)of EMT. Results(1)S100A7 expression was significantly higher in cervical squamous cancer tissues(median 91.6)than that in normal cervical tissues(median 52.1;Z=-2.948,P=0.003).(2)Stable S100A7-overexpressed cells were established using lentiviral-mediated gene delivery in HeLa and CaSki cells. S100A7 was detected by real-time quantitative reverse transcription PCR,S100A7 mRNA of S100A7-overexpressed cells were 119 ± 3 and 177 ± 16, increased significantly compared with control groups of median(P<0.01).Compared with the control cells, the number of S100A7-overexpressed HeLa and CaSki cells that passed the transwell membrane assay were increased significanatly(572 ± 51 vs 337 ± 25, P<0.01;100 ± 8 vs 41 ± 4, P<0.01).Matrigel invasion assay showed that the number of S100A7-overexpressed HeLa and CaSki cells that passed the transwell membrane were respectively 441±15 and 110±14,elevated significantly compared with control cells(156±21 and 59±7;P<0.05). However, S100A7 overexpression didn′t influence the proliferation and cell cycle progression of HeLa and CaSki cells(P>0.05). Expression of E-cadherin was dramatically decreased, while N-cadherin, vimentin, and fibronectin increased in S100A7-overexpressed cells. Conclusion S100A7 enhances the migration, invasion and EMT of HeLa cells and CaSki cells, and may be plays an important role in the development of cervical cancer.

In order to enhance gamma-aminobutyric acid production from L-glutamate efficiently, we amplified the key enzyme glutamate decarboxylase (GAD) encoding gene lpgad from the strain Lactobacillus plantarum GB 01-21 which was obtained by way of multi-mutagenesis and overexpressed it in E. coli BL21. Then we purified GAD by Ni-NTA affinity chromatography and characterized the enzyme to optimize the conditions of the whole-cell transformation. The results showed that the recombinant E. coli BL21 (pET-28a-lpgad) produced 8.53 U/mg GAD, which was increased by 3.24 fold compared with the GAD activity in L. plantarum. The optimum pH and temperature of the enzyme were pH 4.8 and 37 degrees C, respectively. At the same time, we found that Ca2+ and Mg2+ could increase the activity significantly. Based on this, we investigated gamma-aminobutyric acid transformation in 5 L fermentor under the optimum transformation conditions. Accordingly, the yield of gamma-aminobutyric acid was 204.5 g/L at 24 h when the 600 g L-glutamate was added and the mole conversion rate had reached 97.92%. The production of gamma-aminobutyric acid was improved by 42.5% compared with that under the unoptimized transformation conditions. This paved a way for the gamma-aminobutyric acid construction of the industrial applications.
Cloning, Molecular ; Escherichia coli ; enzymology ; genetics ; metabolism ; Glutamate Decarboxylase ; biosynthesis ; genetics ; Glutamic Acid ; metabolism ; Lactobacillus plantarum ; enzymology ; genetics ; Recombination, Genetic ; gamma-Aminobutyric Acid ; biosynthesis

OBJECTIVETo explore the effects on cell proliferation and invasion as well as molecular basis after suppressing EZH2 expression in endometrial carcinoma cells by using siRNAs.

METHODSRT-PCR was used to examine the expression of EZH2 in endometrial carcinoma and their paracancerous tissues. SiRNAs targeting to EZH2 were transfected to endometrial carcinoma cells, and MTT, FACS, and boyden assays were utilized to examine cell proliferation, cell cycle change, and cell invasion. Finally, the molecular mechanisms of EZH2 on cell function alteration were investigated.

RESULTSCompared with paracancerous tissues, increased expression trend of EZH2 mRNA was showed in endometrial carcinoma tissues. Further, knocking down EZH2 expression inhibited cell growth, cell cycle transition from G1 to S phase, and cell invasion ability. Molecular basis indicated that suppression of EZH2 downregulated the expression of E2F1 and MMP9 and upregulated tumor suppressor p21 expression.

CONCLUSIONEZH2 expression is increased in endometrial carcinoma tissues. Knocking down EZH2 expression suppresses the cell growth, cell cycle transition and cell invasion by downregulated E2F1 and MMP9, and upregulated tumor suppressor p21 expression.

Cell Line, Tumor ; Cell Proliferation ; E2F1 Transcription Factor ; metabolism ; Endometrial Neoplasms ; genetics ; pathology ; Enhancer of Zeste Homolog 2 Protein ; Female ; Genes, Tumor Suppressor ; Humans ; Matrix Metalloproteinase 9 ; metabolism ; Polycomb Repressive Complex 2 ; genetics ; RNA Interference ; RNA, Messenger ; genetics ; RNA, Small Interfering ; Transfection

With the optimization of surgical technologies and postoperative management regimens, the number of lung transplantation has been significantly increased, which has become an important treatment for patients with end-stage lung disease. However, due to the impact of comprehensive factors, such as bronchial ischemia and immunosuppression, the incidence of airway stenosis after lung transplantation is relatively high, which severely affects postoperative survival and quality of life of lung transplant recipients. In recent years, with the improvement of perioperative management, organ preservation and surgical technologies, the incidence of airway stenosis after lung transplantation has been declined, but it remains at a high level. Early diagnosis and timely intervention play a significant role in enhancing clinical prognosis of patients with airway stenosis. In this article, the general conditions, diagnosis, treatment and prevention of airway stenosis after lung transplantation were reviewed, aiming to provide reference for comprehensive management of airway stenosis after lung transplantation and improving clinical prognosis of lung transplant recipients.

2-Hydroxybutyric acid (2-HBA) is an important intermediate for synthesizing biodegradable materials and various medicines. Chemically synthesized racemized 2-HBA requires deracemization to obtain optically pure enantiomers for industrial application. In this study, we designed a cascade biosynthesis system in Escherichia coli BL21 by coexpressing L-threonine deaminase (TD), NAD-dependent L-lactate dehydrogenase (LDH) and formate dehydrogenase (FDH) for production of optically pure (S)-2-HBA from bulk chemical L-threonine (L-Thr). To coordinate the production rate and the consumption rate of the intermediate 2-oxobutyric acid in the multi-enzyme cascade catalytic reactions, we explored promoter engineering to regulate the expression levels of TD and FDH, and developed a recombinant strain P21285FDH-T7V7827 with a tunable system to achieve a coordinated multi-enzyme expression. The recombinant strain P21285FDH-T7V7827 was able to efficiently produce (S)-2-HBA with the highest titer of 143 g/L and a molar yield of 97% achieved within 16 hours. This titer was approximately 1.83 times than that of the highest yield reported to date, showing great potential for industrial application. Our results indicated that constructing a multi-enzyme-coordinated expression system in a single cell significantly contributed to the biosynthesis of hydroxyl acids.
Escherichia coli/genetics* ; Formate Dehydrogenases ; Hydroxybutyrates ; Threonine Dehydratase

OBJECTIVE：To establish sustainable development ability evaluation index system for ethnomedicine enterprises in Guizhou province ，and to promote the sustainable development of the ethnomedicine industry. METHODS ：The draft of evaluation index system had been made by documents collection and a meeting of focus group discussion ，the final indexes and weight had been determined by Delphi method for conducting 2 rounds expert questionnaire survey ；the index system was used to measure and compare the results with the evaluation results of the drug regulatory department and the authoritative experts of the industry. RESULTS：The draft of evaluation index system included 6 indexes in the first level indicators and 49 indexes in the second. The expert’s positive coefficients was 100％ after 2 rounds of consultation ；the authoritative coefficients on the opinions of experts for the levels of indicators were 0.86，the coordination coefficient of experts was 0.22 in the first round and 0.48 in the second. After 2 rounds of expert consultation ，the final established evaluation index system contained 6 indexes in the first level indicators and 33 indexes in the second. Fifteen ethnomedicine enterprises in Guizhou province were selected for on-site testing. The evaluation results were not much different from those of the drug regulatory department and the authoritative experts of the industry. CONCLUSIONS：Established sustainable development ability evaluation index system for ethnomedicine enterprises is scientific ， reasonable and feasible ，and can provide standardized reference for regular monitoring and evaluation.

Objective To analyze the influencing factors of survival of patients with airway stenosis requiring clinical interventions after lung transplantation. Methods Clinical data of 66 patients with airway stenosis requiring clinical interventions after lung transplantation were retrospectively analyzed. Univariate and multivariate Cox’s regression models were adopted to analyze the influencing factors of survival of all patients with airway stenosis and those with early airway stenosis. Kaplan-Meier method was used to calculate the overall survival and delineate the survival curve. Results For 66 patients with airway stenosis, the median airway stenosis-free time was 72 (52,102) d, 27% (18/66) for central airway stenosis and 73% (48/66) for distal airway stenosis. Postoperative mechanical ventilation time [hazard ratio (HR) 1.037, 95% confidence interval (CI) 1.005-1.070, P=0.024] and type of surgery (HR 0.400, 95%CI 0.177-0.903, P=0.027) were correlated with the survival of patients with airway stenosis after lung transplantation. The longer the postoperative mechanical ventilation time, the higher the risk of mortality of the recipients. The overall survival of airway stenosis recipients undergoing bilateral lung transplantation was better than that of their counterparts after single lung transplantation. Subgroup analysis showed that grade 3 primary graft dysfunction (PGD) (HR 4.577, 95%CI 1.439-14.555, P=0.010) and immunosuppressive drugs (HR 0.079, 95%CI 0.022-0.287, P<0.001) were associated with the survival of patients with early airway stenosis after lung transplantation. The overall survival of patients with early airway stenosis after lung transplantation without grade 3 PGD was better compared with that of those with grade 3 PGD. The overall survival of patients with early airway stenosis after lung transplantation treated with tacrolimus was superior to that of their counterparts treated with cyclosporine. Conclusions Long postoperative mechanical ventilation time, single lung transplantation, grade 3 PGD and use of cyclosporine may affect the survival of patients with airway stenosis after lung transplantation.