To study the effect of physiological conditions on spidroins, we analyzed NTR1SR2CT module secondary structure, aggregation and silk-formation influenced by different salts (in different concentration intervals). According to the full-length Araneus ventricosus MiSp sequence, NTR1SR2CT module was constructed and expressed in Escherichia coli BL21 (DE3), and the recombinant proteins were purified by denaturation method mediated by 8 mol/L urea. Random coil and helix are the main secondary structures of NTR1SR2CT and could be induced into beta-sheet by drying natively and lyophilization, where methanol can be used as a promoter. Furthermore, potassium and phosphate cations can cause significant NTR1SR2CT protein aggregation and silk-formation. The results could be a basis for the determination of silk-formation mechanism, and also useful for industrialized generation of high performance spider silk-like fibers.
Animals ; Fibroins ; chemistry ; Protein Structure, Secondary ; Salts ; chemistry ; Spiders

Objective To investigate the effects of esmolol on ATPase activity in myocardial mitochondria after resuscitation in rats.Methods Sixty-six male Wistar rats were randomly allocated to three groups:sham group,AD group,AD+Es group.A cardiac arrest model was reproduced by asphyxiation in rats,and then CPR was performed in these animals after 10min asphyxia.The ATPase activity of myocardial mitochondria was determined at 30th,120th and 180th minute after resuscitation.ECG,MAP,HR and temperature were measured continuously.Results Heart rates were significantly higher after adrenaline when compared with sham group.Myocardial mitochondria ATPase activity was decreased significantly in both AD and AD+Es group compared with that of sham group(P

Objective To investigate the effects of connective tissue growth factor (CTGF) on the chronic peripheral nerve compression injury and explore the function of CTGF in peripheral nerve compression injury and repair. Methods From July 2010 to September 2010, fifty aduh male SD rats were randomly divided into group A and B: group A (sham-operated group): only exposed the sciatic nerve; group B (compression group): undergone sciatic nerve entrapment operation on the right hind leg according to the method which Mackinnon adopted when he established the model of chronic sciatic nerve compression.Electron microscopy,immunohistochemistry,RT-PCR and Western-blot were performed to observe the morphological changes of the compressed nerve tissue and to determine the level of CTGF,collagen- Ⅰ,Ⅲ (COL- Ⅰ,Ⅲ),2,4,6,8,10 weeks after the surgery,respectively. Results After sciatic nerve compression,the collagen in nerve increased ; The expression of CTGF and COL- Ⅰ, Ⅲ in sciatic nerve of compressed group increased, which was statistically different compared with the sham-operation group (P ＜ 0.05); In the meanwhile,the contents of CTGF and COL- Ⅰ,Ⅲ were positively correlated in a certain period. Conclusion Peripheral nerve fibrosis can be caused by chronic nerve compression.The expression of COL- Ⅰ,Ⅲ in sciatic nerve increased and CTGF get involved in the pathophysiological process, which suggests that CTGF plays an important role in the process of neural injury and fibrosis.

ObjectiveTo investigate the effect of transforming growth factor-β1 (TGF-β1) antibody on peripheral neural fibrosis after chronic entrapment.MethodsA total of 75 rats were randomly divided into three groups, ie, Group A (sham operation, only the sciatic nerve exposed), Group B (compression only, treated with sciatic nerve entrapment) and Group C (compression plus antibody injection).Electron microscopy, immunohistoehemistry, RT-PCR and Western blot were performed to observe the morphological changes of the compressed nerve tissue and to determine the level of TGF-β1 , collagen Ⅰ and collagen Ⅲ at 2, 4, 6, 8, 10 weeks after sciatic nerve compression, respectively.Results The levels of TGF-β1 , types Ⅰ and Ⅲ collagen protein were increased significantly in the Group B compared with that in the Group A, when the expression of TGF-β1 was increased in the early phase of the compression, reached the peak at the 4th week, and then decreased slowly.The expressions of collagenⅠand collagen Ⅲ were increased after compression, reached a peak at the 6th week and then maintained a relatively high level.The number of the fibrous tissues was decreased significantly and the content of types Ⅰ and Ⅲ collagen protein declined in the Group C, with statistical difference compared with the Group B (P ＜ 0.05).ConclusionsPeripheral nerve fibrosis can be caused by chronic nerve compression.TGF-β1 plays an important role in effectively inhibiting the collagen synthesis and ameliorating the nervous fibrosis of the protein following peripheral nervous entrapment.

Objective To investigate the status of Chlamydia trachomatis(CT) infection in pregnant women in Wuxi ,and to ex‐plore the relationship between CT and abortion .Methods A total of 3 199 cases of pregnant women visiting this hospital from Jan‐uary 2013 to December 2014 were enrolled in this study and divided into the missed abortion group(2 633 cases) ,incomplete abor‐tion group(265 cases) ,the inevitable abortion group(137 cases) ,threatened abortion group(104 cases) and control group(60 cases , all women undergoing induced abortion) .The CT infection was detected by using the fluorescent quantitative polymerase chain reac‐tion(FQ‐PCR) .Results The positive rate of CT in missed abortion group ,incomplete abortion group ,the inevitable abortion group and threatened abortion group were 5 .62% ,4 .53% ,5 .11% and 4 .81% ,respectively ,all were higher than that in the control group (3 .33% ) ,without statistically significant differences (P>0 .05) .The positive rate of CT in women in 16- <21 years old age group and 21- <26 years old age group was higher than that in other age groups ,which was 7 .06% and 6 .20% respectively .However , there were no statistically significant differences in positive rate of CT between these age groups (P>0 .05) .Conclusion The posi‐tive rate in women undergoing spontaneous abortion is higher than that in women undergoing induced abortion ,and young women has a higher infection rate of CT .

OBJECTIVE:To provide reference for the sustainable development of pharmaceutical enterprises in Guizhou prov-ince. METHODS:A questionnaire was conducted for 55 pharmaceutical enterprises in Guizhou province,the basic situation of en-terprises,familiarity of related directors to Good Manufacture Practice of Drugs(GMP)and relevant policy,the current situation of implementing the new version GMP were investigated and statistically analyzed,problems were found,and corresponding coun-termeasures were put forward. RESULTS & CONCLUSIONS:Totally 55 questionnaires were sent out,49 valid ones were received with effectively recovery of 85.1%. The results showed 43 enterprises(87.8%)had passed the GMP authentication;only 13 enter-prises(26.5%)directors were very familiar with the new version GMP. In terms of personnel management,the head of production management and quality management and the authorized person of quality and personnel had not yet met related requirements of the new version GMP fully;in terms of equipment and production management,production area transformation(clean areas,lounges, warehouses,water use) and air purification system design in most enterprises met related requirements of the new version GMP, 23 enterprises (46.9%) still can not conduct fully inspection to products and materials;in terms of document management,there were still some enterprises not meeting the new version GMP standards fully,enterprises'documents(health area layout,air purifi-cation layout,management procedures,operating procedures,etc.) of production site were imperfect. According to the investiga-tion,the main existing problems included lack of funds in implementing the new version GMP reform,not enough understanding or familiarity with the new version GMP,relevant personnel management not reaching the designated position,equipment and pro-duction management needing to be strengthened,document management systemic being poor,risk management being not sound, etc. It is suggested that government should give all forms of capital policy and strengthen the training of the new version GMP;en-terprises should attach great importance to the relevant personnel management,strengthen the equipment and production manage-ment,set up perfect document management system and a sound system of risk management.

Objective To observe the effects of butylphthalide on the expression of autophagy-related protein and mRNA in l-meth-yl-4-phenyl-pyridiniumion (MPP+)-induced SH-SY5Y cells, and to explore the protective effect and possible mechanism of butylphthalide to the cell model of Parkinson's disease. Methods The SH-SY5Y cells were divided into control group (A), MPP+group (B), rapamycin pre-treated+MPP+group (C) and Butylphthalide pretreated+MPP+group (D). The relative viability of SH-SY5Y cells induced by MPP+was measured with MTT assay, the morphology of SH-SY5Y cells was observed. The expression of microtubule associated protein 1 light chain 3 (LC3)-II/I and Beclin 1 protein was detected by Western blotting. And the expression of LC3-II/I and Beclin 1 mRNA were assayed by re-al-time quantitative reverse transcription-PCR (RT-PCR). Results The viability rates of cells were significantly lower in group B than in group A (t=20.270, P<0.001), and were significantly higher in groups C and D than in group B (t>8.770, P<0.001), however, there was no significantly difference between groups C and D (t=2.270, P=0.064). The expression of LC3-II/I and Beclin 1 was higher in group B than in group A (t>6.647, P<0.01), and was higher in groups C and D than in group B (t>3.630, P<0.01), however, there was no significantly differ-ence between groups C and D (t<2.238, P≥0.05). Conclusion Butylphthalide could prevent the injury of SH-SY5Y cells induced by MPP+, which may affect Parkinson's disease by inducing autophagy.

ObjectiveTo detect specific antigens of neural cells in amniotic epithelial cells(AECs) in rats. MethodsAECs were dissociated and purified from the amnion of pregnancy 12—14 d rats. The expression of specific markers of neural stem cells (Nestin, Musashi) and differentiated cells (MAP-2,NSE,GFAP) and ChAT, NT-3 in the AECs were detected by immunocytochemistry. ResultsThe cultured AECs displayed positive immunoreactivity to MAP-2, NSE, GFAP, Nestin and Musashi. In addition, the cells also demonstrated immunoreactivity to ChAT and NT-3. ConclusionAECs are similar with neural cells and it may be useful as a sustained source to improve outcome of neural stem cells transplantation.

ObjectiveTo investigate an effective method to isolate neural stem cells(NSCs).MethodsNSCs were dissociated by digestion with trypsin, EDTA and different doses of Dispase,and serum-free culture techniques and immunohistochemistry techniques were used to verifying the dissociated.ResultsA lot of single neural stem cells were obtained by using Dispase to digest neurosphere, and the cells could keep its structure and morphology.ConclusionIt is an ideal method by using Dispase to digest neurosphere for isolating NSCs.