调宁蛋白在胆固醇结石形成中的作用及疏肝利胆方药的干预机制

目的:探讨疏肝利胆方药防治胆石病的作用机制。方法:将受试豚鼠随机分为四组,即正常组、模型组、疏肝利胆中药组、熊去氧胆酸西药组。后三组采用高胆固醇致石食饵诱发法建立豚鼠胆结石动物模型,其中疏肝利胆中药组、熊去氧胆酸西药组分别给予相应药物进行防治实验,连续7周后,分别对实验豚鼠胆结石生成情况、胆囊组织调宁蛋白(Cap)的蛋白表达进行观察。结果:疏肝利胆方药防治组能有效防止胆结石的生成(与模型组比较P<0.01),并且其效果优于熊去氧胆酸西药组(P<0.01)。模型组动物胆囊组织Cap蛋白表达水平较正常组显著升高(P<0.01),疏肝利胆中药组和熊去氧胆酸西药组胆囊组织Cap蛋白表达水平虽均高于正常组(分别为P<0.01,P<0.05),但与模型组比较,其胆囊组织Cap蛋白表达水平都呈明显降低(均P<0.01),且疏肝利胆中药组和熊去氧胆酸西药组两组胆囊组织Cap蛋白表达水平比较无显著性差异(P>0.05)。结论:疏肝利胆中药能显著降低胆结石的形成,其作用机制可能与下调胆囊平滑肌组织调宁蛋白的水平,提高胆囊平滑肌收缩能力有关。     

RPE-rip after intravitreal bevacizumab (Avastin) treatment for vascularised PED secondary to AMD

Background Retinal pigment epithelial (RPE)-rips in age-related macular degeneration (AMD)-associated pigment epithelial detachment (PED) occur in the natural course of the disease but also after therapy (e.g. laser photocoagulation, photodynamic therapy), possibly triggered by the specific therapy. We report here on four patients that received intravitreal bevacizumab (Avastin) for AMD-associated vascularised PED and developed RPE-rips during the follow-up. Methods The case reports of four consecutive patients that developed RPE-rips after intravitreal injection of bevacizumab at 1 mg/0.1 ml were reviewed. Results The RPE-rips occurred in all patients between 1 week and 1 month following intravitreal injection. Two of the four patients improved in vision despite the rip, but 3 months after the initial intervention, three patients suffered deterioration in visual acuity and had to be re-injected. Conclusion Improvement in visual acuity may occur following intravitreal bevacizumab despite RPE-rips, but the patients need close follow-up and eventual re-treatment in the case of deterioration. None of the authors has a propietary interest in the presented cases.     

原发性胆囊癌与胆囊结石（附84例分析）

本文分析作者收治的８４例胆囊癌的临床特点并结合文献探讨胆囊结石的关系。目前尚无证据说明胆囊结石是胆囊癌的致癌因，但它在成癌过程中很可能起着促进作用。年龄超过５０岁、女性、合并胆囊炎或接触化学致癌物质可以认为是胆囊结石者易患胆囊癌的高危因素。在目前，早期诊断胆囊癌依然十分困难，治疗预后恶劣的情况下，对于具有高危因素者如不能确认胆囊形态正常，施行预防性胆囊切除是合理的。     

视网膜色素上皮受体介导的内吞作用

目的 研究视网膜色素上皮(RPE)细胞内吞光感受器细胞外基质(IPM)内大分子可溶性物质的可能性及其机制。方法 采用荧光标记的甲醛作用后的血清白蛋白(F-FSA),作为清道夫受体配体的指示物,与组织块培养的猪RPE细胞共同孵育,用荧光显微镜和电子显微镜观察RPE细胞对F-FSA的内吞情况。结果 RPE细胞能够摄入F-FSA;这一作用随配体浓度增加而提高,随孵育时间延长而增强;相同或相似的配体之间存在竞争作用;F-FSA进入RPE细胞后,以微囊泡的形式存在于细胞中。结论 RPE细胞能够摄入携带负电荷的大分子物质,这一作用是通过受体介导的内吞作用完成的,清道夫受体可能是介导这一作用的受体;受体介导的RPE细胞的内吞作用可能是IPM内大分子物质代谢的重要机制。（中华眼科杂志,2004,40：539-544)     

视网膜下间隙移植经基因修饰的视网膜色素上皮细胞对大鼠光感受器变性及视功能的保护作用

目的 以RCS大鼠作模型,研究经基因修饰的永生化视网膜色素上皮细胞(RPE)视网膜下移植对光感受器变性的保护作用。方法 在绿色荧光蛋白基因逆转录病毒感染的基础上,利用脂质体介导节状神经生长因子(CNTF)表达质粒转移,修饰成人RPE细胞系CRL-2302。将1×105个表达绿色荧光蛋白(GFP)或GFP及CNTF的RPE细胞移植到4～5周龄RCS大鼠右眼视网膜下间隙,左眼不移植或注射。PBS作为对照。术后2、4、6、8、10和12周作荧光显微镜、光镜、电镜及电生理检查。结果 荧光显微镜观察,术后1周移植的人RPE细胞在RCS大鼠视网膜下间隙已扩散到几乎整个眼底,但随时间延长移植的细胞逐渐减少,术后6周仅残留少量移植细胞。光镜及电镜观察显示移植眼保留的光感受器数量明显较对照眼多,凋亡细胞则较对照跟少。此外,移植眼宿主RPE细胞形态较正常,并可见吞噬小体。视网膜电图(ERG)检查结果表明部分移植眼视网膜功能明显较对照眼好。结论 经过基因修饰的RPE细胞移植可延缓RCS大鼠视网膜光感受器变性,为治疗视网膜变性提供了新的途径。(中华眼科杂志,2004,40:552-556)     

A boussinesq model of natural convection in the human eye and the formation of Krukenberg's spindle

The cornea of the human eye is cooled by the surrounding air and by evaporation of the tear film. The temperature difference between the cornea and the iris (at core body temperature) causes circulation of the aqueous humor in the anterior chamber of the eye. Others have suggested that the circulation pattern governs the shape of the Krukenberg spindle, a distinctive vertical band of pigment on the posterior cornea surface in some pathologies. We modeled aqueous humor flow the human eye, treating the humor as a Boussinesq fluid and setting the corneal temperature based on infrared surface temperature measurements. The model predicts convection currents in the anterior chamber with velocities comparable to those resulting from forced flow through the gap between the iris and lens. When paths of pigment particles are calculated based on the predicted flow field, the particles circulate throughout the anterior chamber but tend to be near the vertical centerline of the eye for a greatest period of time. Further, the particles are usually in close proximity to the cornea only when they are near the vertical centerline. We conclude that the convective flow pattern of aqueous humor is consistent with a vertical pigment spindle. © 2002 Biomedical Engineering Society. PAC2002: 4266Ew, 8710+e, 8719Pp     

Cytochrome P-450 expression and lipid peroxidation in gallbladder cancer

BACKGROUND AND OBJECTIVES: Carcinoma of the gallbladder is the commonest neoplasm of the biliary tract; however, the etiology of this tumor remains unclear. METHODS: A case-control study was carried out comprising 50 patients with gallbladder cancer and 50 patients with cholelithiasis. Cytochrome P-450 estimation was carried out in the gallbladder tissue, and lipid peroxide product melonaldehyde (MDA) was measured in gallbladder bile. Statistical analysis was carried out by Student's t-test, and one-way analysis of variance (ANOVA). The Pearson correlation coefficient was calculated, and linear regression analysis was performed. RESULTS: The mean cytochrome P-450 concentration was significantly higher in patients with carcinoma of the gallbladder than in controls (t = 3.9, P < 0.001; F = 15.2, P = 0.002). Similarly, the mean concentration of melonaldehyde was significantly higher among cancer patients than in patients with gallstones (t = 2.7, P < 0.003; F = 7.37, P = 0.0078). No significant correlation was observed between tissue MDA and cytochrome P-450 levels (Pearson r = 0.1, P = 0.28). CONCLUSIONS: The results of the present study show that the rise in concentration of MDA is independent of the rise in concentration of cytochrome P-450; however, this is statistically insignificant. This could be due to biliary stasis leading to a rise in biliary concentration of melonaldeyde that is disproportionate to its production.     

复制缺陷型逆转录病毒载体介导基因工程化视网膜色素上皮细胞的制备

目的制备稳定高表达生物活性神经生长因子（nerve growth factor,NGF）工程视网膜色素上皮(retinal pigment epithelium,RPE)细胞,拟将转基因基因治疗与视网膜移植技术结合,用于视网膜变性疾病的治疗。方法首先经亚克隆和克隆构建复制缺陷性逆转录病毒质粒重组体pLXSN-NGF,然后经包装细胞pT67包装,获得含有NGF基因的复制缺陷性逆转录病毒重组体;NIH3T3细胞测定其滴度,然后转导培养RPE细胞;反转录聚合酶链反应和Western blot分析NGF基因在RPE细胞中的表达;PC12细胞测定表达产物的生物活性。结果复制缺陷性逆转录病毒滴度为1.2×10     

视网膜色素上皮细胞吞噬纤维连接蛋白时细胞内信号传导通路的研究

目的 探讨视网膜色素上皮（retinal pigment epithelium PRE)细胞对纤维连接蛋白（fibronectin,FN）的吞及相关细胞内信号传导通路的作用。方法 以FN包被的聚苯乙烯微球（微球）作为吞噬标记物, 建立人胎儿RPE细胞吞噬模型。所研究的细胞内信号传导通路包括蛋白激酶C（protein kinase C,PKC)通路、酪氨酸激酶（tyrosine kinase,TK）通路和磷酯酰肌醇3－激酶（phosphatidylinositol 3-kinase,PI3-K)通路,分别使用这3种信号通路的抑制剂预处理RPE细胞,37℃吞噬3h,流式细胞仪定量检测吞噬指数。结果 与对照组比较,RPE细胞对FN包被微球有明显吞噬作用（P＜0．05）。PKC抑制剂PMA（phorbol 12-myristate 13-acetate,100nmol/L)或Calphostin C(400nmol/L)非特异性地增加了PRE细胞对FN包被或未包被微球的吞噬指数（P＜0．01）;TK和PI3K的抑制剂Genistein(100μg/ml）与Wortmannin(5μmol/L）分别降低了FN包被微球的吞噬指数（P＜0．001）,但对对照组无影响。联合使用不同抑剂Genistein或Wortmannin可对抗PMA对FN吞噬的增强作用,但二者联合使用则具有相累加的抑制吞噬作用。结论 PRE细胞对FN的吞噬受细胞内PKC、TK和PI3－K信号传导通路的调节,此结论将为进一步研究治疗增殖性玻璃体视网膜病变的药物提供理论依据。     

绿色荧光蛋白逆转录病毒及其介导的视网膜色素上皮细胞基因转移研究

目的 制备携带绿色荧光蛋白（green fluorescent protein,GFP)基因的逆转录病毒,感染原代与建株的人视网膜色素上皮（retinal pigment epithelium,RPE)细胞,评价逆转录病毒对人RPE细胞的感染能力,为视网膜病变的基因治疗奠定基础。方法 分离编码GFP的cDNA片段并插入逆转录病毒载体pLNCX,借助脂质体将重组逆转录病毒载体pLNCX-GFP转入单嗜性及双嗜性包装细胞,G418筛选抗性克隆,分离GFP表达水平最高的克隆;收集含病毒颗粒的上清液感染NIH3T3及体外培养的人原代与建株的RPE细胞。结果 重组逆转录病毒载体pLNCX-GFP转染各种包装细胞后,可产生GFP逆转录病毒。由双嗜性包装细胞产生的GFP逆转病毒能够感染原代与建株的人RPE细胞,GFP基因可持续在RPE细胞内表达。结论 逆转录病毒能够简便、快速、稳定将目的基因转入RPE细胞,可作为眼底病变基因治疗介导目的基因转移的重要工具。