A comparison between naturally occurring macroamylasaemia and macroamylasaemia induced by hydroxyethyl-starch

Macroamylasaemia was produced in vitro by incubation of hydroxyethylstarch with serum, and in vivo by intravenous infusion of hydroxyethylstarch. Gel filtration on Sephadex G-100 revealed distinct differences in molecular size distribution between such hydroxyethylstarch-induced macroamylase and the usual form of naturally occurring macroamylase which was observed in a few patients from our hospital. Further studies demonstrated that the gel filtration elution pattern of amylase activity in serum containing hydroxyethylstarch-induced macroamylase is significantly altered with time in vitro and in vivo, probably because of an enzymatic degradation of the hydroxyethylstarch components of the macromolecular complexes. In a healthy volunteer the serum amylase activity was elevated to a maximum of 797 u/l and the renal clearance rate of amylase was diminished to a minimum of 0.3 ml/min after infusion of 500 ml of a 6% solution of hydroxyethylstarch, as compared to 300 u/l, and 0.95 ml/min, respectively, during the pre-infusion period.     

骨唾液酸蛋白在体外培养人骨髓间充质干细胞向成骨细胞分化中的角色

目的：骨唾液酸蛋白在骨矿化形成方面扮演重要角色，实验观察其是否能诱导体外培养的人骨髓间充质干细胞向成骨细胞分化。方法：实验于2005—10／2006—12在解放军广州军区广州总医院医学实验科完成。①材料来源：选取在本院体检的健康志愿者2人，对本实验均知情同意。采用Ni-NTA亲和纯化技术，从本室构建的毕赤酵母GS115／pPICZaA-hbsp发酵上清中纯化重组人骨唾液酸蛋白。②实验方法：对健康志愿者进行髂骨穿刺抽取骨髓液，采用贴壁法培养得到骨髓间充质干细胞。设立4组：骨唾液酸蛋白组添加0．1nmol／L骨唾液酸蛋白；成骨诱导液组添加10nmol／L地塞米松、10mmol／L磷酸甘油、50mg／L抗坏血酸；联合组添加上述两组的所有试剂；空白对照组不添加任何处理因素；各组均处理细胞12d。⑧实验评估：光镜及电镜观察培养的细胞形态；以细胞计数法测定生长曲线，应用流式细胞仪分析细胞周期，采用免疫荧光细胞化学法和流式细胞分析检测干细胞标志物STRO-1的表达；生化试剂盒测定碱性磷酸酶活性；von Kossa染色法检测钙沉积。结果：①单个骨髓间充质干细胞为长梭形，经骨唾液酸蛋白处理后细胞大而扁平，原来密集的克隆分散开。②与空白对照组比较，骨唾液酸蛋白组引起细胞生长曲线右移，细胞Go／G，期比例平均增加12．09％（P〈0．01），S期比例减少65．92％（P〈0．01），STRO-1阳性细胞百分率下降26．54％（P〈0．01）。⑧与空白对照组比较，骨唾液酸蛋白组细胞碱性磷酸酶活性增加50．0％，成骨诱导液组增加59．5％，联合组增加71．43％，并且随着处理时间的延长，活性增加越显著。④空白对照组细胞vonKossa染色呈阴性，其余各组均呈阳性。其中联合组的黑色矿化结节体积最大、数目最多；骨唾液酸蛋白组的结节体积较小、数目较少；成骨诱导液组居中。结论：骨唾液酸蛋白对人骨髓间充质干细胞有促进成骨分化和矿化作用，且与成骨诱导液联用效果更佳。     

振幅型光栅法人眼调制传递函数测定的理论分析

目的:介绍测量视网膜调制传递函数的方法和光学原理,验证振幅型正弦光栅加零级空间滤波的方法测量视网膜调制传递函数使测量仪器小型化的可行性。方法:目前测量仪器中获得两相干点光源主要是使用平行板法、双道威棱镜法和光栅法3种,从理论上分析比较3种方法的优劣性,并重点分析矩形光栅、振幅型正弦光栅的各级衍射光强分布特点。结果:①平板法和双道威棱镜法可获得两相干点光源,也是在视网膜调制传递函数测量仪中应用比较成熟的技术,其光学原理简单,获得相干点光源的相干性良好,光强稳定。另外,使用双道威棱镜法时,可以用电位带动反射镜的移动来控制两点光源的间距,易于装置的智能化控制,但所设计的仪器外观尺寸较大,移动性较差,不利于测量装置的手持式和小型化发展。②矩形光栅法可以实现仪器的小型化,但由于矩形光栅的衍射图样是单缝衍射图样与多光束干涉图样相互调制的结果,即衍射光强的多光束干涉条纹干扰测量结果。③振幅型正弦光栅的透过率函数可以实现0￣1,以余弦波形式对入射光波产生振幅调制,且衍射光强只有三级谱线(0级和±1级),且±1级谱线的光强分布具有对称性,只要进行零级空间滤波,就可以得到作为视标调制度的两个点光源。结论:振幅型正弦光栅加零级空间滤波可以获得相干性良好相干点光源,并且可以实现测量仪器的小型化。     

Phosphoenolpyruvate in the rejuvenation of stored red cells in SAGM medium: optimal conditions and the indirect effect of methemoglobin formation

Red cells stored in saline-adenine-glucose-mannitol (SAGM) medium were rejuvenated by incubation with phosphoenolpyruvate (PEP) under conditions that can be achieved easily in ordinary blood banking. Regeneration of 2,3 diphosphoglycerate (2,3 DPG) and adenine nucleotides of stored red cells was dependent on the pH of the incubation medium and the incubation time. In red cells stored for 3 and 5 weeks, the optimal pH and incubation time for regeneration of 2,3 DPG and adenine nucleotides were 5.8 and 90 minutes and 6.1 and 60 minutes, respectively. During the incubation of red cells with PEP, methemoglobin was formed; it increased when the medium pH was below 6.0 and the incubation time exceeded 60 minutes. We conclude that incubation at a medium pH of 6.1 for 60 minutes is optimal for the rejuvenation of stored red cells with PEP. Under such incubation conditions, the concentrations of 2,3 DPG and adenine nucleotides in red cells stored for 5 weeks were restored to normal without methemoglobin formation.     

转染癌基因的骨髓基质干细胞体外分化特征

目的:观察转染癌基因的骨髓基质干细胞在体外分化情况,为肝细胞癌的细胞源研究提供实验依据。方法:实验于2003-05/2004-06在南方医科大学药理教研室实验室完成。①两步法获取大鼠肝细胞,梯度离心法分离大鼠骨髓基质干细胞。②单基因转染是单独将c-myc或K-ras癌基因瞬时转染大鼠骨髓基质干细胞,6孔培养板中培养,24h后荧光显微镜下观察骨髓基质干细胞转染结果。双基因转染步骤相同,只是将c-myc和K-ras癌基因同时转染大鼠骨髓基质干细胞。③c-myc癌基因转染组、K-ras癌基因转染组、双癌基因转染组常规培养,加入含体积分数为0.1胎牛血清的DMEM培养基,于37℃、体积分数为0.05的CO2孵箱培养,每24h半量更换培养液。④c-myc癌基因转染 肝细胞组、K-ras癌基因转染 肝细胞组、双癌基因转染 肝细胞组将已转染癌基因的骨髓基质干细胞,置于叠加的培养板半透膜的上方(细胞密度均为1×105个/cm2),再将肝细胞置于半透膜的下方(每孔细胞密度为3×105/cm2)进行共培养,其余步骤同常规培养。⑤通过反转录聚合酶式反应和细胞免疫组化检测骨髓基质干细胞分化情况。结果:①癌基因转染24h骨髓基质干细胞检测结果:单独转染c-myc或K-ras癌基因的细胞,其绿色荧光蛋白呈均匀一致分布;双基因转染的细胞,绿色荧光蛋白呈点片状分布。②各组骨髓基质干细胞向肿瘤细胞分化检测结果:c-myc癌基因转染组、K-ras癌基因转染组、双癌基因转染组的骨髓基质干细胞,均未向肿瘤细胞分化;c-myc癌基因转染 肝细胞组、K-ras癌基因转染 肝细胞组、双癌基因转染 肝细胞组的骨髓基质干细胞,均向肝细胞癌发展;空白对照组骨髓基质干细胞细胞均为阴性。此外,双癌基因转染 肝细胞组的骨髓基质干细胞分化增殖迅速,反转录聚合酶式反应和免疫组化检测发现,培养第7天出现甲胎蛋白表达,并迅速增加,而第7天出现的白蛋白和细胞角蛋白18表达迅速减弱,第14天消失。结论:转染癌基因的骨髓基质干细胞,在向肝细胞诱导的条件下,部分癌基因可以使干细胞分化为肝癌细胞;多癌基因转染时,更易于使干细胞分化为肝癌细胞。     

胶原-壳聚糖复合材料的制备及生物安全性检测

目的:制备胶原-壳聚糖复合细胞载体,并对该载体进行系统的生物学性能检测。方法:实验于2003-07/2006-06在天津市医药科学研究所完成。将胶原溶液和壳聚糖溶液按照一定比例(9∶1)混匀,交联后真空冷冻干燥,制成胶原-壳聚糖复合载体,对该载体进行复合材料性状及理化性能检测。并采用急性全身毒性试验、皮内刺激试验、皮肤致敏试验、溶血试验、细胞毒性试验和致突变试验进行生物学性能检测。结果:①胶原-壳聚糖复合材料具有三维立体多孔结构,适合细胞的三维生长,能为新生组织提供良好的支架。②急性全身毒性试验、皮内刺激试验均阴性,致敏率仅为6.25%,细胞毒性为0级,无诱变能力。结论:从仿生学角度出发,制备出可生物降解的胶原-壳聚糖复合材料,经过系统的生物学评价发现,胶原-壳聚糖复合材料具有良好的生物安全性,对机体无毒,不致突变,对细胞有较强的亲和作用,利于细胞生长和分化。     

Voriconazole for serious fungal infections

Voriconazole is a new second generation triazole effective against a wide spectrum of fungal pathogens. A randomised, controlled trial has shown it to be superior to amphotericin B in invasive aspergillosis, and it is a potential alternative to amphotericin B in neutropenic sepsis and to fluconazole in oesophageal candidiasis. Early clinical reports and in vitro susceptibility data suggest that it may also be a valuable antifungal against fluconazole-resistant Candida species and certain emerging fungal pathogens, which cause infections that are often refractory to conventional therapies. There is limited evidence of azole cross-resistance of clinical importance. Voriconazole is available as intravenous and oral formulations and has excellent tissue penetration and a good safety profile, the main problems being transient visual impairment and hepatotoxicity in patients with liver disease. It is metabolised by cytochrome P-450 isoenzymes causing important drug interactions but, in contrast to amphotericin B, is safe in renal failure and rarely causes infusion-related reactions. This review outlines the pharmacology of voriconazole and focuses on its clinical applications and safety profile.