Detection of mutations in Mycobacterium tuberculosis genome determining resistance to fluoroquinolones by hybridization on biological microchips

We developed a method of identification of Mycobacterium tuberculosis with simultaneous evaluation of the sensitivity to fluoroquinolones on a biological microchip array. The method of multiplex two-staged PCR followed by hybridization of a biochip makes it possible to detect 8 mutant variants of gyrA gene occurring in fluoroquinolone-resistant strains (∼85% all resistant forms) within 1 day. Using this method we analyzed 107 cultures isolated from patients with tuberculosis and 78 sputum samples. Mutations in gyrA gene were detected in 48 (92%) resistant strains. Natural S95T polymorphism in gyrA gene was detected in all resistant and in 76% sensitive strains. The sensitivity and specificity of the proposed method calculated on the basis of the analysis of sputum samples (n=78) were 94 and 100%, respectively. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 145, No. 1, pp. 115–120, January, 2008     

Effect of cultivation temperature on blast transformation of lymphocytes from people of different ages

The effect of the cultivation temperature on blast transformation of lymphocytes from persons aged 20–35 and 90–102 years induced by phytohemagglutinin was studied. Cultivation at 39°C was found to increase, but at 41°C to reduce sharply the index of blast transformation compared with the control (37°C). The effect of cultivation temperature on the blast-transformation process has certain features which depend on the donors' age.Institute of Gerontology, Academy of Medical Sciences of the USSR, Kiev. (Presented by Academician of the Academy of Medical Sciences of the USSR D. F. Chebotarev.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 83, No. 4, pp. 470–472, April, 1977.     

Adenovirus-mediated intratumoral lymphotactin gene transfer potentiates the antibody-targeted superantigen therapy of cancer

Bacterial superantigens are extremely potent activators of murine and human T lymphocytes. To engineer superantigens for cancer immunotherapy, staphylococcal enterotoxin A (SEA) was genetically fused to the Fab region of the human colon carcinoma-reactive monoclonal antibody (mAb) C215. Fusion protein C215Fab-SEA can trigger cytotoxic T cells against C215 antigen positive tumor cells and induce tumor-suppressive cytokines. However, the antitumor effect of C215Fab-SEA is often not satisfactory because of T cell deletion after activation and failure to induce potent CTL activity after repeated administration. Lymphotactin (Lptn) is a potent chemoattractant for T cells and NK cells. To improve the therapeutic efficacy of fusion protein C215Fab-SEA we investigated in this study the antitumor responses elicited by combination of C215Fab-SEA and adenovirus-mediated intratumoral Lptn gene transfer in the preestablished C215 antigen expressing B16 melanoma murine model. More significant inhibition of tumor growth and prolonged survival time were observed in tumor-bearing mice that received combined therapy of C215Fab-SEA and Ad-Lptn than those of mice treated with C215Fab-SEA or Ad-Lptn alone. The highest CTL activity of tumor-bearing mice was induced after combined therapy. Intratumoral coadministration of C215Fab-SEA and Ad-Lptn augmented splenic NK activity of tumor-bearing mice most markedly. Our data demonstrate that the in vivo antitumor effect of C215Fab-SEA immunotherapy is potentiated significantly by combination with intratumoral Lptn gene transfer through more efficient induction of specific and nonspecific antitumor immune responses.     

Ⅰ型胶原膜、胞外基质提取物膜体外构建人工真皮能力比较

比较Ⅰ型胶原与胞外基质(extracellularmatrix,EM)提取物冻干后形成的膜(EM膜)在体外构建人工真皮的能力,以选择一种更有利于人工真皮形成的支架材料。方法分别使用小牛真皮部分提取的Ⅰ型胶原蛋白和胞外基质提取物冻干后制备基质网架,植入皮肤成纤维细胞,形成人工真皮。用扫描电镜观察I型胶原膜和EM膜的表面形态结构;选取不同的时间点,对种植于两种支架材料上的细胞进行计数,并利用间接ELISA方法检测人工真皮复合物中的细胞Ⅰ型胶原分泌情况,通过统计学的分析手段,对细胞的生长情况做出判断;用组织学方法观察人工真皮的形成情况。结果扫描电镜观察结果,两种膜在外观形态上无明显差异;细胞在EM膜上的增殖速度较Ⅰ型胶原膜快;ELISA分析显示,EM-细胞复合物中的成纤维细胞能够分泌更多的Ⅰ型胶原;与Ⅰ型胶原膜相比,EM在培养液中的降解更为缓慢,种植后的成纤维细胞在EM膜上生长旺盛,细胞层次明显多于前者,形成了较为明显的真皮样组织。结论EM膜适于成纤维细胞的生长,体外降解速率慢,是一种较Ⅰ型胶原膜更为理想的真皮支架材料。     

Metabolism of lymphocytes in mice with Ehrlich ascites carcinoma

The pattern of metabolic processes in lymphocytes of mice with Ehrlich ascites carcinoma depended on the stage of tumor growth.Translated from Byulleten Eksperimentalnoi Biologii i Meditsiny, Vol. 138, No. 11, pp. 563–566, November, 2004     

支气管哮喘病人焦虑与生命质量的相关性研究

目的　评估支气管哮喘病人焦虑与生命质量的水平 ;探讨支气管哮喘病人焦虑与生命质量的关系 .方法应用状态 -特质焦虑问卷 (STAIFORMY)和世界卫生组织生命质量量表简表 (WHOQOL -BREF) ,分别对支气管哮喘门诊病人和健康对照人群的焦虑和生命质量水平进行问卷调查 .结果　哮喘组状态焦虑为 45 1± 9 7,特质焦虑为46 5± 9 7,生命质量为 87 3± 13 2 ;健康组的状态焦虑为 3 7 7± 7 4,特质焦虑为 3 8 4± 8 1,生命质量为 95 4± 11 7.哮喘组状态焦虑、特质焦虑与生命质量均负相关 ,相关系数分别为 - 0 69和 - 0 79.结论　①支气管哮喘病人的状态焦虑及特质焦虑水平均高于健康人群 ,生命质量水平低于健康人群 ;②支气管喘病人的焦虑与生命质量负相关 .因此 ,在护理实践中应对哮喘病人进行焦虑的心理护理 ,提高其生命质量     

下胫腓联合分离内固定术式的生物力学研究

目的　探讨下胫腓联合分离采用新型内固定术式———下胫腓钩板固定器 (Hook -platefixation ,HPF)的生物力学特性 ,与螺钉、钢板和带钩固定器三种内固定方式比较 ,为临床提供科学依据。方法　采用新鲜成人尸体足标本 6具 ,运用生物力学实验应力分析方法和压敏片技术 ,测量不同内固定术式的远端胫腓骨强度、应变、负重面积、接触应力和足弓承载能力及踝关节稳定性的变化。结果　新型下胫腓钩板固定器内固定术式无论在胫腓骨强度和刚度、负重面积、接触压力、足弓的变形、移位强度和刚度、承载能力以及踝关节的稳定性方面均优于其它三种内固定术式 ,具有显著性差异 (P <0 .0 1)。结论　实验结果表明 ,采用下胫腓钩板固定器 (HPF) ,既有利于提高生物力学性能 ,又有利于改善踝关节的稳定性 ,与传统手术方式相比可减少一次手术 ,避免了断钉等并发症 ,且能有效地提高足部承载能力 ,是下胫腓联合分离内固定的一种优良术式     

Affinity fractionation of lymphocytes using a monolithic cryogel

A new type of continuous, supermacroporous, monolithic, cryogel affinity adsorbent was developed, allowing specific fractionation and separation of human peripheral blood lymphocytes in a chromatographic format. The affinity adsorbent was used to design a novel cell separation strategy, which was based on the interaction of protein A from Staphylococcus aureus with cells bearing IgG antibodies on the surface. After treating lymphocytes with goat anti-human IgG(H+L), the IgG-positive B-lymphocytes were efficiently separated from T-lymphocytes. Protein A covalently coupled to epoxy activated dimethylacrylamide (DMAA) cryogel matrix specifically bound IgG-bearing B-lymphocytes through the Fc region, while non-bound T-lymphocytes passed through the column. More than 90% of the B-lymphocytes were retained in the column while the cells in the breakthrough fraction were enriched in T-lymphocytes (81%). The viability of the T-lymphocytes isolated was greater than 90%. The bound lymphocytes released by human or dog IgG recovered 60-70% of the B-cells without significantly impairing the cell viability. The technique can be applied in general to cell separation systems where IgG antibodies against specific cell surface markers are available.     

一种低成本、低本底的免疫细胞化学反应方法

为了探讨一种适合培养细胞的低本底、低成本免疫细胞化学方法 ,本研究将低浓度的 Triton X-10 0加入抗体稀释液和洗液中 ,观察了其对 ABC反应时的抗体和 ABC的使用浓度和反应效果的影响。结果证明 ,在抗体稀释液和洗液中加入 0 .1%Triton X-10 0可以增强细胞对抗体的通透性 ,在不减弱阳性信号的状况下使 -抗的使用浓度明显降低 ,使生物素化二抗和 ABC的使用浓度达到 1/ 10 0 0～ 1/ 40 0 0 ,并大大减弱染色本底。本研究结果提示 ,在这一稀释度明显低于目前试剂盒建议的使用浓度。在充分显示阳性信号的前提下 ,使抗体的使用浓度和量大幅度减少 ,从而实现了降低本底和实验成本的目的