Venous thromboembolism ( VTE) remains the third most common cardiovascular disease with a vague pathogenesis.Conventional biomarkers exhibit poor performance in the diagnosis, surveillance and prognosis of VTE.MicroRNAs ( miRNAs ) are a class of evolutionarily conserved small non-coding RNAs that are involved in the regulation of gene expression and protein translation An array of experimental studies has shown the importance of miRNAs for disease initiation/progression.Circulating miRNAs are found in plasma, serum and other body fluids in an apparently stable form.Recent evidence revealed that circulating miRNAs, a novel family of regulatory molecules, emerge as a promising class of biomarkers in many cardiovascular diseases, malignancies as well as VTE.This review describes current understanding of miRNA biogenesis and the origins and types of circulating miRNAs and gives an outline of recent work on circulating miRNAs as well as its challenges and perspectives of the clinical utility of circulating miRNA in VTE.

AIM To study the action of danshengsuan (Dan) on expression of CD11b, P selectin, ICAM 1, VCAM 1 and E slectin in vitro. METHODS Adheshion molecule expression on the cell surface was measured by flow cytometry. RESULTS Treatment of neutrophils with N fomyl met leu phe(fMLP), resulted in a marked increase of CD11b expression. Dan inhibited the effects of fMLP in a concentration dependent manner. Dan did not inhibit the increase of P selectin expression of thrombin actived human blood platelets. Treatment of cultured human umbilical vein endothelial cells (HUVEC) with TNF ? resulted in an increase of ICAM 1, VCAM 1 and E slectin. Dan inhibited the increase of VCAM 1 and E slectin expression, not ICAM 1. CONCLUSION Dan inhibited expression of adhesion molecules CD11b, ICAM 1, VCAM 1 in human neutrophils and HUVEC.

Objective:To express the recombinant single chain Fv(scFv) in E.coli and reduce immunogenicity and molecular weight of a monoclonal antibody specific for human platelet.Methods:The variable regions of the heavy and light chains of platelet specific antibody SZ 2 were amplified by reverse transcription and polymerase chain reaction.VH and VL gene segments were cloned into pUC Tm and joined together with a (gly 4ser) 3 linker.The resulting scFv was expressed in PET expression system.The expressed recombinant protein was characterized by its size on SDS PAGE,by Western blot,by flow cytometry and its functions.Results:The VH and VL genes were homologous with the published gene sequences of mouse antibody variable region.The recombinant scFv was expressed mostly in the form of inclusion bodies,and the yield was up to 25% of the total cell proteins.Functional studies showed that SZ 2 scFv could bind to platelet and could suppress platelet aggregation induced by ristocatin and thrombin.Conclusion:A recombinant SZ 2 scFv specific against platelet was developed and characterized.

AIM: To clone and express the metalloproteinase domain of human von Willebrand factor-cleaving protease (vWF-cp). METHODS: The metalloproteinase domain of human vWF-cp, amplified from the plasmid containing the vWF-cp cDNA gene by using polymerase chain reaction, was cloned into pUC18, and its accuracy was verified by sequencing. Then the domain was inserted into the multiclone site of pET28a(+) and included a 6?His Tag at its amino terminal. After induced by IPTG, the recombinant protein was purified by using a Ni-NTA column and confirmed by Western blot. RESULTS: Comparison of the nucleotide sequence of our cloned domain with the GenBank sequence revealed no difference. High-level expression of the recombinant protein was yielded after 5-hour induction, which amounted to 28% of total bacteria protein in inclusion body. Western blot demonstrated that it possessed high specificity. CONCLUSION: The metalloproteinase domain of vWF-cp was high efficiently expressed in Escherichia coli. This might contribute to the further study of the relationship between its structure and function. [

AIM: To obtain a McAb that can inhibit the function of matrix metalloproteinase-2 (MMP-2), we expressed the fibronectin-like domain of MMP-2 (MFD) in vitro and prepared a McAb against MMP-2. METHODS: The purified MFD protein was used to immunize BALB/C mouse three times. Then the spleen of mouse was taken out and hybridized with hybridoma cells SP2/0. The positive cell clones were screened with ELISA method. The subtype and tissue specificity of the McAb were identified and its effect on endothelial cell migration and tube-formation was analyzed. RESULTS: After the spleen cells of the mouse and hybridoma cells SP2/0 were hybridized, a piece of cells that continuously secreted McAb against MMP-2 was obtained and named SZ-117. The titers of this McAb in culture supernatants and ascites were 2?10~-3 and 2?10~-5 , respectively. The heavy chain of the McAb belongs to IgG1 subclass. The McAb identified native MMP-2. MMP-2 existed in the stromal tissue of stomach, cholecystis, spleen, ovarian, prostate, salping and lymph node. It inhibited the invasion behavior of endothelial cells Eahy926 and pancreatic carcinoma cells 1990 and inhibited the tube-formation of Eahy926 cells. CONCLUSION: A useful tool for testing MMP-2 is obtained and it will be helpful to look for a kind of new anti-tumor material.

AIM: To investigate the therapeutic action of secreted endostatin (ES) on breast cancer cells. METHODS: Retroviral-mediated endostatin gene was transferred to breast cancer cell line MDA-MB-231. The ES biological properties and function were evaluated by polymerase chain reaction (PCR), MTT and a murine xenograft model. RESULTS: After retroviral transduction, endostatin genetically modified breast tumor cells were confirmed by PCR, and the integration and durative expression of endostatin gene was successfully committed. Compared with controls, endostatin secreted by genetically modified cells markedly inhibited endothelial cell proliferation (P0.05). The results of the transplanted subcutaneous tumor model in nude mice suggested that the subcutaneous growth of MDA-MB-231 was significantly inhibited by the expression of endostatin gene (P

AIM: To study the clinical significance of determining serum anti-endothelial cell antibodies (AECA) and thrombopoietin (TPO) in the differential diagnosis of idiopathic thrombocytopenic purpura (ITP) and systemic lupus erythematosus (SLE). METHODS: Serum AECA and TPO in 76 patients with ITP, 41 patients with SLE and 50 normal individuals were detected by ELISA method. RESULTS: Serum AECA level in SLE and ITP was much higher than that in normal group (P0.05), but serum TPO level in SLE was much higher than that in ITP and normal groups (P

AIM: To investigate the clinical significance in determination of the P-selectin levels in subjects with prethrombotic state or thrombosis by flow cytometry (FCM). METHODS: The P-selectin expression on platelet membrane in 42 patients with diabetes mellitus, 33 with hyperlipidemia, 23 with cerebral infarction and 20 healthy individuals, were analyzed using fluorescently-labeled SZ-51 by direct FCM comparing with indirect FCM and enzyme-linked immunosorbent assay (ELISA). RESULTS: The level of P-selectin on platelet membrane is higher in DM (23.92%?15.83%), in hyperlipidemia (18.34%?9.46%) and in cerebral infarction (19.32%?10.38%) than normal subjects (3.38%?1.11%) (P

OBJECTIVETo investigate the therapeutic effect of retroviral endostatin gene transfer on the human colon cancer cell line, LoVo.

METHODSA retroviral vector pLESSN expressing secretable endostatin was constructed and packaged with a titer of 8.2 x 10(5) CFU/ml. A LoVo cell line was subjected to retrovirus-mediated endostatin gene transfer. The proviral integration of endostatin was analyzed with PCR. The function of endostatin was tested by MTT assay in vitro and a mouse xenograft model in vivo.

RESULTSAfter transfection and superinfection, amphotropic retrovirus was collected, and transduction with amphotropic retroviruses resulted in endostatin proviral integration. The endostatin secreted by transduced LoVo cells markedly inhibited cell growth up to 67% (P<0.001), compared with the control cells. The gene expression of endostatin in LoVo colon tumor cells significantly inhibited tumor growth in vivo. There was an 86% reduction in tumor size in the endostatin-transduced group, accompanied by a reduction in vessels, compared with the control group (P<0.01).

CONCLUSIONRetroviruses can allow functional expression of the endostatin gene in human colon tumors, showing promise for an antitumor strategy using antiangiogenesis.

Cell Division ; Cell Line, Tumor ; Colonic Neoplasms ; pathology ; therapy ; Endostatins ; genetics ; Gene Transfer Techniques ; Genetic Vectors ; Humans ; Retroviridae

Aim To study the effects of ferulic acid (FA) on E-selectin expression in human umbilical vein endothelial cells (HUVECs) activated by lipopolysaccharide and leukocyte-endothelial cell adhesion. Methods The effects of FA on E-selectin and E-selectin mRNA expression were determined by flow cytometry and reverse transcription polymerase chain reaction. The effect of FA on HL60-HUVEC adhesion was evaluated with the method of staining the cells by Rose Bengal. Results The expression of tively). Conclusion FA can inhibit the expression of E-selectin and E-selectin mRNA and HL60-HUVEC adhesion. This may contribute to its protective effect against ischemia-reperfusion injury.