PTEN及Bcl-2蛋白在膀胱移行细胞癌中的表达及其意义

目的：探讨PTEN及Bcl-2蛋白在膀胱移行细胞癌的表达规律及其临床意义。方法：应用免疫组织化学链亲和素-生物素-霉复合物（SP）方法检测43例膀胱移行细胞癌和7例正常黏膜组织中PTEN及Bcl-2蛋白的表达．分析二者的表达与膀胱癌病理参数的关系。结果：（1）G1、G2、G3肿瘤PTEN表达阳性率分别为85．7％、80．4％、73．3％，浸润性肿瘤和表浅性肿瘤表达阳性率分别为70．4％和93．8％，说明PTEN表达阳性率与肿瘤病理分级、临床分期有关。（2）G。、G2、G3肿瘤Bcl-2表达阳性率分别为14．2％、57．14％、66．67％，浸润性肿瘤和表浅性肿瘤表达阳性率分别为59．3％和43．8％，说明Bcl-2表达阳性率与肿瘤病理分级、临床分期有关。（3）随着肿瘤恶性程度的增高和临床分期进展，PTEN蛋白阳性率呈下降，Bcl-2表达呈增高趋势，二者表达呈负相关关系。结论：PTEN的抑癌作用可能与Bcl-2有关．二者的异常表达在膀胱移行细胞癌的发生、发展过程中起重要作用。二者同时检测有助于判断预后。     

磺柳酸法在Array360全自动特定蛋白分析仪尿液测定中的应用

目的探讨磺柳酸法在Array360全自动特定蛋白分析仪尿液测定中的应用价值。方法用磺柳酸法对离心后的尿液样本中的蛋白质进行半定量测定,并用Array360全自动特定蛋白分析仪测定α1微球蛋白α1-MG、白蛋白(ALB)、转铁蛋白(TRF)、免疫球蛋白G(IgG)在各个稀释度的测定范围,结合尿液蛋白质半定量结果和各个项目在不同稀释度的测定范围设定初始的标本稀释度。结果蛋白质半定量结果不同的尿液标本用Ar-ray360进行测定时,各个项目有其合适的初始标本稀释度。结论磺柳酸法在Array360全自动特定蛋白分析仪尿液的测定中有重要的应用价值,可大幅度节约试剂成本,缩短测定时间。     

骨髓干细胞参与小鼠急性肾小管坏死修复的实验研究

【摘要】 目的 观察在生理和病理情况下,骨髓来源的干细胞(BMSC)能否分化成肾小管上皮细胞。 方法 绿色荧光蛋白(GFP)标记的C57BL/6转基因小鼠提供骨髓细胞。同种无荧光标记的C57BL/6小鼠100只分为正常对照组、全身照射组、缺血再灌注组、骨髓移植组、骨髓移植+缺血再灌注组。受体鼠的骨髓重建经血液常规检查和流式细胞仪检测确认,并采用荧光组织化学、免疫组织化学等方法观察绿色荧光标记的BMSC在受体鼠肾脏的分布及数量。 结果 全身致死剂量γ射线照射未造成小鼠肾脏组织结构和生理功能的明显改变。骨髓移植后第56、84天的受体鼠肾小管中有少量GFP阳性细胞的存在[(78.75±5.99)%、(79.58±4.60)%],激光共聚焦显微镜进一步证实这些细胞位于肾小管,并表达肾小管上皮细胞特异性的功能蛋白megalin。 结论 在生理和病理情况下,骨髓干细胞均可以向肾小管上皮细胞转分化,参与肾小管上皮细胞的更新,并且在急性肾小管坏死的病理条件下,骨髓干细胞的肾向转化率与肾脏受损程度有关。     

Clinical safety of enamel matrix derivative (EMDOGAIN®) in the treatment of periodontal defects

Abstract The aim of the present clinical trial was to test tolerability during 2 treatments with EMDOGAIN® in a large number of patients. An open, controlled study design in 10 Swedish specialist clinics was chosen, with a test group of 107 patients treated with EMDOGAIN® in connection with periodontal surgery at 2 surgical test sites per patient. The procedures were performed 2 to 6 weeks apart on one-rooted teeth with at least 4 mm deep intraosseous lesions. A control group of 33 patients underwent flap surgery without EMDOGAIN® at I comparable site. In total 214 test and 33 control surgeries were performed. Serum samples were obtained from test patients for analysis of total and specific antibody levels. 10 of the patients had samples taken before and after the first surgery. 56 other samples were taken after one treatment with EMDOGAIN®, and 63 after 2 treatments. None of the samples, not even from allergy-prone patients after 2 treatments, indicated deviations from established baseline ranges. This indicates that the immunogenic potential of EMDOGAIN® is extremely low when applied in conjunction with periodontal surgery. Comparison between the test and control groups demonstrated the same type and frequency of post-surgical experiences, i.e., reactions caused by the surgical procedure itself. Clinical probing and radiographic evaluation was performed at baseline and 8 months postsurgery. About half of the patients (44 test and 21 control) were also evaluated after 3 years. There was a significant difference between the test and control results at 8 months post surgery. and this difference had increased further at the 3 year follow-up. The 2.5–3 mm increase in attachment and bone level after treatment with EMDOGAIN® was of the same magnitude as seen in the studies with split-mouth design aiming for lest of effectiveness of EMDOGAIN®.     

胃粘膜相关淋巴组织型淋巴瘤MAPK和Stat3磷酸化及cyclinD1蛋白表达的意义

目的:研究胃粘膜相关淋巴组织型(MALT)淋巴瘤MAPK和Stat3磷酸化与cyclinD1蛋白表达及其意义。方法:利用免疫组织化学方法检测了45例胃MALT淋巴瘤MAPK和Stat3磷酸化及cyclinD1蛋白的表达。结果:在胃MALT淋巴瘤中p-MAPK、p-Stat3及cyclinD1蛋白的阳性率分别为73.3%(33/45)、64.4%(29/45)和68.9%(31/45);低度恶性组p-MAPK和cyclinD1蛋白的阳性表达强度明显高于高度恶性组(P<0.01),而p-Stat3表达强度无明显差异(P>0.05);在低度和高度恶性胃MALT淋巴瘤中p-MAPK和cyclinD1蛋白的阳性信号强度均呈明显的正相关(r=0.6572和0.6823,P<0.01),而p-Stat3与cyclinD1蛋白表达未见明显相关性(r=0.1927,P>0.05)。结论:提示MAPK磷酸化在胃MALT淋巴瘤中发生及演进过程中起重要作用,但Stat3的磷酸化可能与该肿瘤的恶性演进关系不明显;在胃MALT淋巴瘤的发生与发展中,p-MAPK可诱导cyclinD1过度表达,从而促使该肿瘤细胞维持高增殖状态。     

Progress toward re‐engineering non‐ribosomal peptide synthetase proteins: a potential new source of pharmacological agents

Many important pharmaceutical agents, including vancomycin, bleomycin, cyclosporin, and several antibiotics, are produced by non‐ribosomal peptide synthetase (NRPS) enzymes in microorganisms. The NRPS pathway produces an extensive library of products using multienzyme complexes acting in an assembly‐line fashion. Engineering an NRPS system to produce an even greater variety of products, some of which may also have beneficial therapeutic value, would be an enormous advantage. Several approaches have been successful in generating novel NRPS products: mutational biosynthesis during which nonnatural substrates are fed to an organism; domain and module swapping between different species to generate hybrid enzymes; and rational site‐directed mutagenesis, based either on phylogeny or computational prediction, intended to switch substrate specificity and produce altered products. This review will highlight the progress in these areas and describe research in the future that will extend the capacity for re‐engineering NRPS systems. Drug Dev. Res. 66:9–18, 2006. © 2006 Wiley‐Liss, Inc.     

Detergent fractionation with subsequent subtractive suppression hybridization as a tool for identifying genes coding for plasma membrane proteins

Abstract:  The identification of tumor-specific proteins located at the plasma membrane is hampered by numerous methodological pitfalls many of which are associated with the post-translational modification of such proteins. Here, we present a new combination of detergent fractionation of cells and of subtractive suppression hybridization (SSH) to gain overexpressed genes coding for membrane-associated or secreted proteins. Fractionation of subcellular components by digitonin allowed sequestering mRNA of the rough Endoplasmatic reticulum and thereby increasing the percentage of sequences coding for membrane-bound proteins. Fractionated mRNAs from the cutaneous T-cell lymphoma (CTCL) cell line HuT78 and from normal peripheral blood monocytes were used for SSH leading to the enrichment of sequences overexpressed in the tumor cells. We identified some 21 overexpressed genes, among them are GPR137B, FAM62A, NOMO1, HSP90, SLIT1, IBP2, CLIF, IRAK and ARC. mRNA expression was tested for selected genes in CTCL cell lines, skin specimens and peripheral blood samples from CTCL patients and healthy donors. Several of the detected sequences are clearly related to cancer, but have not yet been associated with CTCL. qPCR confirmed an enrichment of these mRNAs in the rough endoplasmic reticulum fraction. RT-PCR confirmed the expression of these genes in skin specimens and peripheral blood of CTCL patients. Western blotting verified protein expression of HSP90 and IBP2 in HuT78. GPR137B could be detected by immunohistology in HuT78 and in keratinocytes of dysplastic epidermis, but also in sweat glands of healthy skin. In summary, we developed a new technique, which allows identifying overexpressed genes coding preferentially for membrane-associated proteins.     

自制稀土永磁夹板对骨折愈合作用的观察

目的观察稀土永磁夹板对家兔骨折愈合质量及骨形态发生蛋白（BMP-2）、血管内皮细胞生长因子（VEGF）表达情况的影响,从分子生物学水平探讨磁场促进骨折愈合的作用机制。方法健康成年家兔50只,建立双前肢桡骨中段骨折模型。左侧用稀土永磁夹板固定,作为实验侧;右侧用内镶铅片小夹板固定,作为对照侧。分别于术后1、2、3、4、6周摄双前肢X线片后取材,进行HE及BMP-2、VEGF免疫组化染色并在光镜下观察。结果X线片图像定量分析结果比较,实验侧愈合明显快于对照侧。两侧差异有显著性（P〈0.05）。实验侧在1、2、3、4周VEGF的表达均比对照侧强,两侧差异有显著性（P〈0.05）。实验侧在2、3、4周BMP-2的表达均比对照侧强,两侧差异有显著性（P〈0.01）。结论静磁场在骨折愈合过程中可加强BMP-2、VEGF的表达。自制稀土永磁夹板对实验性骨折愈合有明显的促进作用。     

透明质酸对BMP-2转染的羊BMSCs增殖、分化的影响

目的观察透明质酸（HA）对携带人骨形态发生蛋白-2的腺病毒（Adv-BMP-2）转染的羊骨髓基质干细胞（BMSCs）体外增殖、分化的影响。方法将羊骨髓体外分离、扩增BMSCs,分成5组：转染Adv-BMP-2的BMSCs＋HA组（Ⅰ组）,转染Adv-BMP-2的BMSCs组（2组）,BMSCs＋HA组（3组）,BMSCs组（4组）,转染携带β半乳糖苷酶的腺病毒（Adv-β-gal）的BMSCs组（5组）。采用细胞计数、绘制细胞生长曲线和流式细胞分析观测细胞增殖;采用碱性磷酸酶（ALP）检测以及RT-PCR检测Ⅰ型胶原（Col-Ⅰ）、骨连接素（ON）和骨涎蛋白（BSP）的mRNA表达。结果①细胞增殖：3 d后各组细胞增殖无显著差异,1周后第1组和第3组的细胞增殖明显高于其他组。②ALP活性：3 d后第3组ALP活性明显低于第4、5组,而第1、2组则显著高于第4、5组;7 d后前3组ALP活性较后两组均显著增加,而第1、2组ALP活性增加更为显著。③Col-Ⅰ、ON和BSP的mRNA表达：3 d和7 d后前3组较后两组均有不同程度的增多。结论HA与BMSCs体外复合培养后可促进BMSCs的增殖、增加ALP的活性以及Col-Ⅰ、ON和BSP基因的表达。