1.Experimental study of ballon-dilatation in an establishment of restenosis model in carotid artery of rabbit
Yongqin LI ; Ming XU ; Zhaoxin YAO
Journal of Interventional Radiology 1994;0(02):-
Objective To study the mechanism of restenosis for the prevention and treatment by PTA (percutaneous transcather angioplasty) through establishing an model.Methods Under median cervical incision, to eplore right rabbit carotid artery and then clip the segments of ICA at the origination and CCA near the cardiac side of bifucation, balloon catheter with guided wire were finally inserted from proximal cardiac side of the ligated ECA to perform TA after withdrawing the clipper. Histological and morphological analysis with the controll of normal left CCA were carried out during different follow up periods. Results Histopathological and morphometric analysis indicating thrombosis was the main changes in early stage and followed by VSMC from media to intima and irregular proliferation leading to obviously intimal thicking and restenosis at 15 days after PTA.Conclusions This experimental study show low mortality, high practicability and good reproducibility of the model as an ideal presentation for study the mechanism and the prevention of restenosis.
2.Expressions and significance of tumor-associated genes in tissues adjacent to human epithelial ovarian cancer of orthotopic implantation in nude mice
Genhai ZHU ; Zhaoxin YANG ; Shengtan WANG ; Junhong CAI ; Chunying CHEN ; Maozhong YAO ; Lan HONG ; Shuying YANG ; Lili DING
Journal of Chinese Physician 2012;(11):1445-1450
Objective To investigate the feasibility in screening of normal ovarian tissues by evaluating the expressions of tumor-associated genes in tissues adjacent to human epithelial ovarian cancer of orthotopic implantation in nude mice.Methods Human epithelial ovarian cancer cell lines OVCAR3 were grown in subcutaneous tissues,and the tumor tissues were orthotopic implanted.The expressions of CK-7,CA125,P53,survivin,MMP-2,and TIMP-2 were detected by immunohistochemical staining in proximal tissues,middle tissues,distal tissues adjacent to tumor,tumor tissues,and normal ovarian tissues of nude mice.Results 35 samples ovarian tissues with normal biopsy were gained from 40 cases of human epithelial ovarian cancers of orthotopic implantation model in nude mice.The expression rate of CK-7,CA125,P53,survivin,MMP-2,and TIMP-2 were 95.0% (38/40),95.0% (38/40),75.0% (30/40),85.0% (34/40),77.5% (31/40),and 77.5% (31/40) in tumor tissues,respectively; 71.4% (25/35),68.6%(24/35),57.1% (20/35),62.9% (22/35),60.0% (21/35),and 57.1% (20/35) in proximal tissues adjacent to tumor,respectively; 34.3% (12/35),31.4% (11/35),31.4% (11/35),31.4% (11/35),34.3% (12/35),and 31.4% (11/35) in middle tissues adjacent to tumor,respectively; and 25.7% (9/35),22.9% (8/35),25.7% (9/35),25.7% (9/35),28.6%(10/35),and 31.4% (11/35) in distal tissues adjacent to tumor,re respectively.The expressions of CK-7,CA125,P53,survivin,MMP-2,and TIMP-2 were all negative in 23 samples normal ovarian tissues adjacent to tumor.The expressions of CK-7,CA125,P53,survivin,MMP-2 and TIMP-2 in proximal tissues adjacent to tumor were lower than those in tumor tissues(P<0.05),and higher than those in middle tissues and in distal tissues adjacent to tumor(P<0.01).There were no expression difference between in middle tissues and in distal tissues (P>0.05).The strong positive expressions of CK-7,CA125,and survivin were higher than those of P53,MMP-2,and TIMP-2 (P<0.01).No significant difference was found in those expressions in the normal ovarian tissues adjacent to tumor gained from orthotopic implantation model with different severity.The expressions of CK-7,CA125,P53,survivin,MMP-2,and TIMP-2 were all negative in 20 normal ovarian tissues of nude mice.Conclusions The expression of CK-7,CA125,P53,survivin,MMP-2,and TIMP-2 showed decreasing trend to non-cancer direction.Negative expressions of these tumor-associated genes can be used as standard in screening of normal ovarian tissues adjacent to tumor.Relative safe normal ovarian tissues can be obtained from the tissues adjacent to tumor.
3.Cloning and expression of organic solvent tolerant lipase gene from Staphylococcus saprophyticus M36.
Yanchong TANG ; Yaping LU ; Fengxia LÜ ; Xiaomei BIE ; Yao GUO ; Zhaoxin LU
Chinese Journal of Biotechnology 2009;25(12):1989-1995
Lipases are important biocatalysts that are widely used in food processing and bio-diesel production. However, organic solvents could inactivate some lipases during applications. Therefore, the efficient cloning and expression of the organic solvent-tolerant lipase is important to its application. In this work, we first found out an organic solvent-tolerant lipase from Staphylococcus saprophyticus M36 and amplified the 741 bp Lipase gene lip3 (GenBank Accession No. FJ979867), by PCR, which encoded a 31.6 kD polypeptide of 247 amino acid residues. But the lipase shared 83% identity with tentative lip3 gene of Staphylococcus saprophyticus (GenBank Accession No. AP008934). We connected the gene with expression vector pET-DsbA, transformed it into Escherichia coli BL21 (DE3), and obtained the recombinant pET-DsbA-lip3. With the induction by 0.4 mmol/L of isopropyl beta-D-thiogalactopyranoside at pH 8.0, OD600 1.0, 25 degrees C for 12 h, the lipase activity reached up to 25.8 U/mL. The lipase expressed was stable in the presence of methanol, n-hexane, and isooctane, n-heptane.
Amino Acid Sequence
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Base Sequence
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Cloning, Molecular
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Enzyme Stability
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Escherichia coli
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genetics
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metabolism
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Lipase
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biosynthesis
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genetics
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Molecular Sequence Data
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Organic Chemicals
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chemistry
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Recombinant Proteins
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biosynthesis
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genetics
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Solvents
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chemistry
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Staphylococcus saprophyticus
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enzymology
4.Fusion expression of fibrinolytic enzyme gene PPFE-I from endophytic Paenibacillus polymyxa in Escherichia coli and activity analysis.
Fengxia LÜ ; Zhaoxin LU ; Xiaomei BIE ; Qian LIN ; Chong ZHANG ; Lin CAO ; Yao GUO ; Yanchong TANG
Chinese Journal of Biotechnology 2010;26(8):1128-1134
With the genomic DNA of strain EJS-3 as the template, we amplified the gene of fibrinolytic enzyme from Paenibacillus polymyxa (PPFE-I) by PCR. We purified the PCR product and ligated it into pMD19-T. After DNA sequencing, we cloned the PPFE-I gene into expression vector pET-DsbA and transformed it into Escherichia coli BL21(DE3). Upon induction of IPTG, we found that the activity of recombinant fibrinolytic enzyme fused with DsbA expressed in Escherichia coli was 228 IU/mL. SDS-PAGE analysis showed that the recombinant enzyme was soluble and accounted for about 18.4% of total cell protein. Western blotting demonstrated that the recombinant protein was DsbA-PPFE-I. We purified the recombinant enzyme by Ni affinity chromatography, thrombin digestion and sephadex G-100 gel-filtration, and identified the molecular weight of purified product to be 66.3 kDa with MALDI-TOF mass spectrometry. The purified enzyme exhibited distinct fibrinolytic activity on fibrin plate.
Antifibrinolytic Agents
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pharmacology
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Cloning, Molecular
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Escherichia coli
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genetics
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metabolism
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Fibrinolytic Agents
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metabolism
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Genetic Vectors
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genetics
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Paenibacillus
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chemistry
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enzymology
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Recombinant Fusion Proteins
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biosynthesis
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genetics
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pharmacology