Objective To induce the site-directed mutation of human cardiac troponin I (cTnI) gene, express the mutant in E. coli, and to study the effects of the mutation on the prokaryotic expression of cTnI. Methods The cDNA encoding cTnI was cloned with RT-PCR from the total RNA extracted from human myocardium tissues. A pair of primers was designed and, after the mutations were induced at the second and the fourth codons, inserted into prokaryotic vector pET-28c (+) and transform the recombinant to BL21 (DE3) bacteria. After purified with Ni-NTA resin, the histidine-tagged fusion protein expressed by IPTG-induced was identified by Western blotting and the expression yield of cTnI protein was investigated. Results The expression of the recombinant carrying processed cTnI cDNA was stronger than that in control group. Conclusion cDNA encoding cTnI was successfully cloned. The recombinant with mutations can be more efficient expressed in E. coli. The cTnI protein can be purified to near homogeneity.

Objective To study the interference effects of tetramethylpyrazine (TMP) on calcinuerin (CaN), c-fos and the nuclear antigen of proliferating cells in the proliferation of vascular smooth muscle cells (VSMCs) treated by angiotensinⅡ(Ang Ⅱ). Methods A cell proliferating model of VSMCs induced by Ang Ⅱ was established; the effects of TMP on CaN was detected by enzyme reaction phosphorus measurement; the effects of TMP on c-fos gene and PCNA expression were observed by immunocytochemical staining and image analysis technique (A value). Results The rats’ aortic smooth muscle cells were cultured in vitro successfully. CaN activities, cell proliferation activity and the expression levels of c-fos and PCNA increased significantly in VSMCs proliferation induced by Ang Ⅱ (P

Objective To induce the the 17th condon antonymous mutagenesis of salivary histatin 5(HRP5) cDNA,express the mutant and hrp5 in Pichia pastoris,and to study the effects of mutation on expression.Methods According to the Pichia pastoris' codon bias,two pairs of primers(H1 and H2,H3 and H4) were designed.H1 and H2,H3 and H4 have complementary 3' end,and the EcoR I site was added to the 5' end of H1 and H3,Sal I site to H2,H4.The cDNA of hrp5 and hrp5' was generated with PCR by H1 and H2,H3 and H4,respectively.The secrete vector pPICZ?-A,hrp5 and hrp5' were digested with EcoR I +Sal I,linkede by T4 DNA ligase and transformed to E.coli TOP10 comptetent cell,positive colonies were screened on LB plates with Zeocin.The recombinant plasmids pPICZ?-A-hrp5 and pPICZ?-A-hrp5' identified by digestion and DNA sequencing were amplified largely,linearized by Sac I and transformed to GS115 comptetent cell by electroporation,positive colonies were screened on YEPD plates with Zeocin,the recombinant GS115 were confirmed by PCR,cultured and induced expression by methanol.The amount and anticandidal activity of the expressed products was compared with synthetic HRP5.Results Both hrp5 and hrp5' were integrated into the genome of GS115 and expressed successfully,the anticandidal activity of the recombinant HRP5 and HRP5' was identical with synthetic HRP5,the amount of expressed HRP5 and HRP5' was 4?mol/L and 5?mol/L,respectively.Conclusion Both the recombinant HRP5 and HRP5' showed better anticandidal activity.The amount of expressed prodcts increased 25% by substituting Asn for Lys17 without changing anticandidal activity.

0.05);between YMDD mutation and YMDD negative,there was statistical difference(P0.05),between YMDD mutation and YMDD negative, there was significant difference(t=12.76,P

AIM and METHODS: To study the protective effects of liposomes containing L-Arg,Se and taurine on intestinal ischemia-reperfusion injury in rats. Wistar rats were divided randomly into sham operated group,ischemia-reperfusion(I/R) group,pretreatment with liposomes group and treatment with liposomes at reperfusion group. In the experiments, superior mesenteric artery was clipped for 60 min, and then unclipped. 2 hours of reperfusion later, MDA content, T-SOD and Ca 2+ -Mg 2+ -ATPase activities in intestinal tissues were detected respectively, ultrastructure and bcl-2 expression in intestinal mucosa tissue were observed. RESULTS: MDA content in liposomes-treated group was less than I/R group ( P 0.05). CONCLUSION: Liposomes containing L-arginine, Se and taurine can protect intestine against ischemia-reperfusion injury in rats,which may be related to inhibiting lipid peroxidation, stabilizing internal circumstances and inducing bcl-2 protein expression.

AIM: To study the role of calcineurin (CaN)-dependent signaling pathway in proliferation of vascular smooth muscle cells (VSMCs) by observing the effect of cyclosporin A (CsA) on proliferation of neuropeptide Y (NPY)-induced rat VSMCs. METHODS: Upon the model of cultured rat VSMCs, the study consisted of three groups: NPYgroup,CsA+NPY group and control group. CaN activity was determinated by enzyme reaction phosphorus measurement. The methods of biochemistry (MTT) and quantitative immunocytochemistry were applied to investigate the proliferation of VSMC and the expression of proliferation cell nuclear antigen (PCNA) in cultured rat VSMCs. (RESULTS:) (Compared) with the control group, the VSMC's CaN activity, proliferation activity and expression of PCNA (by photo densitometry A_(PCNA)) were obviously increased in NPY group (P

AIM: To investigate the protective effect of ischemic-preconditioning under the mild hypothermia against small intestine ischemia-reperfusion injury in rats and its mechanism. METHODS: Thirty-two rats were randomized into 4 groups (8 rats in each group): sham operated group (Sham), ischemia-reperfusion (I/R) group, ischemic-preconditioning (IP) group, mild hypothermia ischemic-preconditioning (MHIP) group. The wet/dry ratio, Ca 2+ -Mg 2+ -ATPase activity in intestine tissue, the malondialdehyde (MDA) content, activities of lactate dehydrogenase (LDH), superoxide dismutase (SOD) and total antioxdase (TAX) in blood were determined. Ultrastructure, Bcl-2 and Bax expression in intestinal mucosa tissue were also observed. RESULTS: After I/R, the intestinal tissue wet/dry ratio, the content of MDA, LDH activity, the optic density of Bcl-2 and Bax proteins were significantly higher in I/R group than those in sham group (P

OBJECTIVE:To probe into an inventory method of high efficiency and quality for PIVAS.METHODS:Data processing and calculation were carried out using Excel based on the information system of PIVAS.RESULTS & CONCLUSIONS:Digital inventory greatly improves the efficiency and quality of inventory.Under the condition of same workload and same number of staff,working time of digital inventory is 60% less than that of traditional inventory.And consistent rate between material and account is increased by 5%.

To study the expression and distribution of neuropeptide Y (NPY) and long leptin receptor (OB-Rb) in the gastrointestinal tract of giant panda, samples of three animals were collected from the key laboratory for reproduction and conservation genetics of endangered wildlife of Sichuan province, China conservation and research center for the giant panda. Paraffin sections of giant panda gastrointestinal tissue samples were observed using hematoxylin-eosin staining (HE) and strept actividin-biotin complex immunohistochemical staining (IHC). The results show that the intestinal histology of three pandas was normal and no pathological changes, and there were rich single-cell and multi-cell mucous glands, long intestinal villi and thick muscularis mucosa and muscle layer. Positive cells expressing NPY and OB-Rb were widely detected in the gastrointestinal tract by IHC methods. NPY positive nerve fibers and neuronal cell were widely distributed in submucosal plexus and myenteric plexus, especially in the former. They were arranged beaded or point-like shape. NPY positive cells were observed in the shape of ellipse and polygon and mainly located in the mucous layer and intestinal glands. OB-Rb positive cells were mainly distributed in the mucous layer and the laminae propria, especially the latter. These results confirmed that NPY and OB-Rb are widely distributed in the gut of the giant panda, which provide strong reference for the research between growth and development, digestion and absorption, and immune function.
Animals ; China ; Intestines ; metabolism ; Neuropeptide Y ; genetics ; metabolism ; Receptors, Leptin ; genetics ; metabolism ; Ursidae ; genetics ; metabolism

OBJECTIVE To explore the effect of bisphenol A (BPA) on the differentiation potential of embryonic stem cells, and provide an experimental basis for evaluation of safety of BPA. METHODS Mouse embryonic fibroblasts (MEFs) and embryonic stem cells (ESCs) were treated with BPA 0.1, 1, 10, 100 and 1000 μmol.L-1 for 8 d respectively. The viability of MEFs and ESCs was measured by CCK-8 and lC50 was calculated. The mRNA expression of α-myosin heavy chain in ESCs was tested by RT-PCR to determine lD50 . The embryonic body cultured by suspension method was treated with BPA 0.001, 0.01, 0.1 and 1 μmol.L-1 for 10 d respectively. The changes of marked genes in each blastoderm were detected by RT-PCR. RESULTS lC50 of BPA to mouse ESCs was 5.22×10-4 mol.L-1 , and to MEFs was 6. 25 × 10-4 mol.L-1 . lD50 of BPA to mouse ESCs differentiating to cardiomyocytes was 7.0×10-7 mol.L-1 . BPA 0.001 and 0.01 μmol.L-1 upregulated the expression of the marked genes of mesoderm, fetal liver kinase-1 and globin transcription factor 1. CONCLUSION BPA is a strong embry-otoxic compound. BPA of low concentration can promote the differentiation of mouse ESCs to mesoderm.