Effects of the juice of Fructus Hippophae (JFH) on thrombocytopenia induced by cyclophosphamide in rats

AIM:To investigate the effects of the juice of fructus hippophae(JFH) on thrombocytopenia in rats.METHODS:Forty-eight wistar rats were divided into control group, model group, Yixuesheng-Jiaonang (YSJN) group and JFH group. Rats were injected intraperitoneally with cyclophosphamide (CTX, 30 mg/kg) or saline once a day for 3 days. And then, Saline, YSJN or JFH was administered (ig) once a day for 11 days. On the second, fouth, sixth and eighth day, the thrombocyte were counted, the mean platelet volume (MPV) and PDW were examined. On the eighth day, the cAMP, cGMP in platelets and platelet aggregation function were also examined.RESULTS:JFH could shorten the time of blood coagulation, increase the count of platelet, and enhance the platelet aggregation function. The experiment also showed that the JFH could decrease cGMP content in platelet.CONCLUSION:JFH could enhance blood coagulation and quality of platelet and prevent thrombocytopenia induced by cyclophosphamide in rats.     

Traumatic shock and systemic inflammation

Trauma is the No.1 cause of death in persons under the age of 45 and the No.3 cause of all deaths. Many trauma patients die of multiple organ failure (MOF), induced by systemic inflammatory response syndrome (SIRS). This paper reviews the relations of traumatic shock with systemic inflammation.     

瓢虫防卫生物碱中的氮杂大环内酯

化学防卫是昆虫防御外来侵害的方法之一。简要评述了墨西哥豆瓢虫Epilachna varivestis、南瓜瓢虫Epilachna borealis、苜蓿瓢虫Subcoccinella vigintiquatuorpunctata 3种瓢虫蛹分泌的化学防御物质——氮杂大环内酯的结构、生物活性以及生物合成的研究进展。     

The application of atomic force microscope in clinic diagnosis of erythrocytes

AIM: To investigate the application of atomic force microscope (AFM) in clinic diagnosis. METHODS: Topographic images and some parameters of large range field and microstructures of erythrocytes in the blood of normal subjects, lung cancer and myelodisplastic syndrome (MDS) patients were examined by atomic force microscope. RESULTS: Many clear topographic images of many erythrocytes, single erythrocyte, and microstructure of erythrocyte membrane surface were obtained. Many erythrocytes in lung cancer patients were found to change into echinocytes. One erythrocyte had 10-20 protuberances, most of which, with a mean width of 589. 0 nm and a length of 646. 7 nm, were on the edge of cells. The protuberances on the center of echinocytes are lodged and embedded. The erythrocytes of MDS patients were biconcave in shape. Many apertures with different diameters of tens to hundreds nanometer appeared on the surface of cell membrane. CONCLUSIONS: AFM can be widely applied in clinic pathological inspection, including quantification of cells, obtainment and comparison of many parameters (such as diameter, thickness, volume, surface, surface area/volume ratio), observation of topograph of single cell, and observation and comparison of membrane surface microstructure of cells, and so on.     

Bone marrow-derived stromal cells and cartilage tissue engineering

Articular cartilage repair is one of the most challenging issues which remains to be resolved in clinic work. Discovering of bone marrow stromal cells (BMSC) and its application in tissue engineering provide new methods for the treatment of cartilage defects. High seeding density, appropriate cytokines and three-dimensional culture play important roles in the process of inducing BMSC differentiating into chondrocytes, suitable scaffold is also essential in reconstructing cartilage in vitro by methods of tissue engineering.     

Detection,cloning and expression of bone morphogenetic protein-1 from human osteosarcoma cell lines

AIM:To manufacture recombinant protein of the highly conserved domain in human bone morphogenetic protein-1(BMP-1) using gene engineering methods as antigen for making wide spectrum antibody to BMP-1.METHODS:We analyzed the gene sequences and protein structures of BMP-1 and its related proteins, and chose a highly conserved fragment as target gene. Total RNA was prepared from human osteosarcoma cell line Saos-2, then the target gene was amplified with RT-PCR. The PCR product was cloned into prokaryotic expression vector pMAL c2 to get recombinant vector BMP-1(322-588aa)-pMAL c2. After transforming the recombinant plasmid into DH5-alpha and screening, several prositive clones were got for sequencing. Finally the transformed cells was induced with IPTG to get fusion protein.RESULTS:The BMP-1 gene fragment was successfully cloned into vector pMAL c2, and was able to express efficiently with IPTG inducement. The amount of expressed fusion protein is about 66%-72% in total volume of bacterial proteins.CONCLUSIONS:The recombinant protein contains several key domains(2 CUB domains and 1 EGF domain), which are shared by BMP-1 and its related proteins. Specific wide spectrum antibody to human BMP-1 and its related proteins may be generated with this recombinant protein antigen.     

Selection and amplification of the liver stem cell subset from rat bone marrow cells with a medium containing cholestatic serum in vitro

AIM: To explore the feasibility of direct separat and selective enlargement of the bone marrow-derived liver stem cells (BDLSC) from bone marrow cells with a culture system containing cholestatic serum in vitro. METHODS: Bone marrow cells of rats were cultured with selective media containing 2%, 5%, 7% and 10% cholestatic rat serum, respectively. The BDLSC were then induced to proliferate with the addition of hepatocyte growth factor (HGF) on the firth day. BDLSC were characterized using immunocytochemistry and RT-PCR for lineage markers, glycogen staining and urea synthetic assay for functions 2 weeks later. RESULTS: Bone marrow cells were unble to form colony in the presence of 2% cholestatic serum and apopotosis appeared gradually in 7% or 10% cholestatic serum. The BDLSC survived in the medium containing 5% cholestatic serum while the other types of cells did not. The survival cells proliferated with a high speed during the second week and then formed hepatocyte-like colony-forming units (H-CFU). Cells in the H-CFU expressed the characteristic proteins of fetal hepatocytes. Furthermore, they had glycogen storage and urea synthesis functions, two of the critical features of hepatocytes. CONCLUSION: The selective micro-environment effectively selected BDLSC from the bone marrow cell, and will be a new way to provide an abundant source of donor hepatocytes for clinical cell therapy.     

Effect of mutant β-catenin gene on the proliferation of hepatocytes

AIM:The β-catenin is a key molecule in the Wnt signal pathway, which plays a critical role in normal development and tumorigenesis. However, the mechanisms of the β-catenin on the cell growth control are still not completely defined. The aim of this study was to test the hypothesis that the mutant β-catenin may regulate the hepatocyte proliferation. METHODS: The immortalized murine hepatocyte cell line, AML12, was used for this study. A plasmid that contain mutant β-catenin S33Y was transfected into the AML12 cells and a stable cell line AML12S33Y was established. The cell growth property of this cell line and the parental cell were compared by flow cytometry analysis and direct cell count. The cells were also tested for the ability to form soft agar colonies, and the ability to form tumors in the severe immune deficient mice (SCID). RESULTS:1. The mutant β-catenin containing cell line AML12S33Y has higher proliferating index compared with the parental AML12 cells (P<0.01), suggesting that mutant β-catenin promotes cell growth. 2. The mutant β-catenin cells formed small colonies in soft agar after 4 weeks of culture, but did not generate tumor in SCID mice. CONCLUSION:The mutant β-catenin promotes liver cell growth.     

Influence of ischemia-reperfusion on apoptosis of sinoatrial node cells in rabbits in vivo

AIM:To study influence of ischemia-reperfusion(IR) on apoptosis and expression of apoptosis-related genes Fas-L, Bax and Bcl-2 of sinoatrial node(SAN) cells in rabbits in vivo. METHODS:Ninety healthy adult rabbits were divided randomly into control group, ischemia groups (I_{10 min}, I_{30 min}, I_{60 min} and I_{120 min}) and IR groups (I_{10 min}R_4h, I_{30 min}R_4h, I_{60 min}R_4hand I_{120 min}R_4h). IR injury model of SAN was established by occluding and loosening the start section of right coronary artery. The apoptosis of SAN cells was detected by TUNEL staining. The expression of Fas-L, Bax and Bcl-2 of SAN cells was detected by immunohistochemistry. RESULTS:①No obvious apoptosis of SAN cells was observed in control group, I_{10 min} and I_{30 min} groups. Apoptosis of different degrees in SAN cells were found in 68.3%(41/60) rabbits in I_{60 min}, I_{120 min} and 4 subgroups of IR. ②The highest expression of Fas-L and Bax was observed in I_{120 min} group and that of Bcl-2 was in I_{60 min} group. ③The highest expression of Fas-L and Bax was observed in I_{60 min}R_4h group. The peak level of Bcl-2 was observed in I_{30 min}R_4h group. ④The expression of Fas-L and Bax was significant higher in IR group than that in ischemic group at the same time point. CONCLUSION:Ischemia and IR induced apoptosis of SAN cells in rabbit in vivo. Fas-L、Bax、Bcl-2 may participate in the regulation of apoptosis and the injury during IR aggravates the apoptosis of SAN cells.     

Rat urotensin-II-induced main pulmonary artery contractions are mediated by MAPK

AIM: To investigate rat Urotensin-II(rat U-II)-induced vasoconstriction of rat main pulmonary arteries and the role of mitogen-activated protein kinase(MAPK). METHODS: The main pulmonary artery was dissected from the male Sprague-Dawley rats and artery ring width was 3-4 mm. Concentration-response curves were generated to rat U-II(0.03 nmol/L-30 nmol/L).Inhibitor of MAPK, PD 98059(0.1 μmol/L-10 μmol/L) were added into the medium after rat U-II(30 nmol/L)induced vasoconstriction had reached plateau to construct the relaxant concentration-response curves and their EC₅₀ and E_max. RESULTS：Rat U-II was a potent vasoconstrictor of isolated rat main pulmonary arteries [EC₅₀=7.95±0.40, E_max=(14.28±6.34)% of the response to 60 mmol/L KCl]; PD 98059 caused concentration-dependent relaxations of rat U-II precontracted arteries [EC₅₀=5.91±0.45, E_max=(81.39±13.65)%]. CONCLUSION: Rat U-II was a potent vasoconstrictor of rat main pulmonary arteries and this response was mediated through MAPK.