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.     

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.     

Biological characteristics of long passaging rat mesenchymal stem cells

AIM:To investigate multi-potential of rat bone marrow mesenchymal stem cells (rBMMSC) and mutation inclination, the rBMMSC were long passaged in vitro. METHODS:Cellular cycles of different passages were assayed by FACSan flow cytometry and karyotypes of passage 6, passage 25 and passage 45 were compared by G-binding analysis. RESULTS:The early passages and long-term passages all showed strong proliferation; passage 6, passage 25 and passage 45 all showed normal karyotype. CONCLUSION:Long-term culture and passage of rBMMSC still remains strong proliferation. With this capability, the mutation inclination is not enhanced.     

Effects of TGF-β1 and TNF-α antisense PS-ODNS on ex vivo expansion of hematopoietic stem/progenitor cells(HSPC)

AIM:To study the effect of TGF-β₁ and TNF-α antisense PS-ODNS on ex vivo expansion of hematopoietic stem/progenitor cells (HSPC). METHODS:CD₃₄⁺cells were purified from fresh umbilical cord blood by immunomagnetic beads, and mononuclear cells were purified from bone marrow by Ficoll-hypaque. The effects of TGF-β₁ and /or TNF-α antisense PS-ODNS on ex vivo expansion of CD₃₄⁺ cells、CFU-GEMM、CFU-GM、CFU-E and BFU-E were detected by using liquid and semi-solid culture systems.RESULTS:TGF-β₁ antisense PS-ODNS cooperated with cytokines increased the number of CD₃₄⁺ cells, CFU-GEMM, CFU-GM, CFU-E and BFU-E, which was as 4, 2.6, 2.7, 1.8, 2.1 times as that of the control (the cytokines combination), respectively. TNF-α antisense PS-ODNS cooperated with cytokines respectively increased the number of CD₃₄⁺ cells, CFU-GEMM, CFU-GM, CFU-E and BFU-E by 4, 2.9, 2.6, 1.7, 1.8 times as that of the control. The above two antisense PS-ODNS cooperated with cytokines could respectively increased the number of CD₃₄⁺ cells, CFU-GEMM, CFU-GM, CFU-E and BFU-E by 5.3, 2.1, 2.7, 1.9, 1.8 times as that of the control.CONCLUSION:Inhibition of endogenous TGF-β₁ and TNF-α by antisense PS-ODNS will be one of the effective methods to expand HSPC ex vivo.     

Effect of cilazapril on pulmonary vascular endothelial dysfunction in hypoxic rats

AIM:To study the effect of cilazapril on pulmonary vascular endothelial dysfunction in hypoxic rats. METHODS:The structure and function of endothelium in hypoxic rats were studied by biochemical analysis, radioimmunoassay, transmission electron microscope and correlated with hemodynamic. RESULTS:1) The change and damage of ultrastructure in endothelial cell (EC) were obsevered in hypoxic rats. 2) The contents of plasma nitric oxide (NO) and superoxide dismutase (SOD) activity in blood as well as endothelial nitric oxide synthase (eNOS) activity in the lung tissue were significantly lower in the hypoxic rat than those in contral animals. The concentrations of plasma endothelin-1(ET-1) and angiotensin converting enzyme(ACE) as well as malondialdehyde(MDA) were significantly higher in the hypoxic rat than these in contral animals. The relaxing and contracting factors had a significant positive/negative correlation with mean pulmonary artery pressure (mPAP). 3) Cilazapril significantly decreased the level of ET-1 and ACE and significantly increased the level of NO and activity of eNOS and SOD. At the same time, cilazapril extenuated hypoxia-induced injuries of EC. CONCLUSION:The results indicate that damaging structure and dysfunction of EC existes in hypoxic rats. The cilazapril effectively preventes and treates the chronic hypoxic PH by relieving the injury and improving secretion in EC.     

菟丝子致死薇甘菊

薇甘菊Mikania micrantha是入侵的害草，已经由珠江三角洲扩散到粤东、粤西沿海地区，以及部分山区。在薇甘菊的天敌调查中看到菟丝子寄生于薇甘菊，严重时可以把薇甘菊致死。我们在本所的实验园中栽种薇甘菊，并让其被菟丝子Cusuta chinensis Lam.寄生，结果证实菟丝子在两个月左右，完全可以抑制薇甘菊的生长，最终把薇甘菊致死。这可能是以草治草的一种新途径，但是菟丝子也是我国农作物的一种重要昌，如何利用菟丝子控制薇甘菊的为害，而又不使菟丝子对薇甘菊的伴生植物和鞭它农作物造成为害，是下一步要解决的问题，菟丝子致死薇甘菊的机理更需要深入研究。     

组氨醇脱氢酶抑制剂的研究动态

组氨醇脱氢酶(HDH)是组氨酸生物合成过程中最后两步反应的催化酶。对组氨醇脱氢酶的结构、催化机理、组氨醇脱氢酶抑制剂等方面的研究动态进行了较为详细的综述。     

苏云金芽孢杆菌cry基因在大肠杆菌中表达产物和生物活性

研究了几种Bt cry基因于大肠杆菌(Escherichia coli)中表达产物在pH 10.0的50mmol/L碳酸钠和20mmol/L乙醇胺溶解液中的溶解性 ,发现同样的Cry蛋白在碳酸钠中的溶解度大于乙醇胺。通过胰蛋白酶消化 ,明确Cry1Ca7、Cry1Ia8酶解产物为 38kD多肽 ;Cry1Ie1、Cry1Cb2、Cry2Ab4酶解产物为 41kD多肽 ;Cry1Ac酶解产物为60kD多肽。采用FPLC层析方法对 6种原毒素及其酶解后得到的毒素多肽进行了分离纯化 ,比较了原毒素和毒素的杀虫活性的差异。其结果表明 ,Cry1Ac的原毒素和毒素对棉铃虫初孵幼虫的校正死亡率均为 100% ,Cry2Ab4的原毒素的毒力高于其酶解毒素。     

十种常用农药与球孢白僵菌的生物学相容性

球孢白僵菌孢子粉与10种常用农药相容性的测定结果显示,随着孢子浓度上升,所试农药对孢子的抑制作用均有不同程度的增强。在1/10田间常规使用浓度下,百菌清和代森锰锌均能抑制或杀死孢子(萌发率<1％)。除阿维菌素外,所有杀虫剂均与白僵菌孢子相容,在常规使用浓度的10倍稀释液中孢子萌发率达90％以上。吡虫啉、蚜虱灵、灭多威和氟虫腈与孢子的相容性最好,其中吡虫啉和蚜虱灵对孢子萌发率的影响不明显随药剂浓度的变化而变化,即使在田间常规使用浓度下孢子萌发率也在95％以上,而阿维菌素与白僵菌的相容性极差。因此,应用白僵菌制剂防治害虫,选择生物学相容性好的农药以低剂量与白僵菌制剂混用,既可使菌剂增效,又可大幅度降低化学药剂用量。