大鼠肾脏发育中肾小体数目的变化

目的探讨胚胎及出生后大鼠肾脏发育过程中的肾小体数目的变化规律。方法采用光镜连续切片技术,结合体视学分析方法,对不同发育阶段大鼠肾小体数目的变化进行体视学测量。结果肾小体的数目从胚龄18天到出生后7天大约增大7倍,出生后7天达到高峰,出生后7天至出生后40天基本不再增加。结论大鼠肾小体的数目于胚龄18天到出生后7天逐渐增多,出生后7天之后基本稳定。     

小鼠后肾集合管水通道蛋白表达及其上皮细胞超微结构研究

目的观察小鼠后肾集合管水通道蛋白(AQP)-4的表达及其上皮细胞超微结构变化。方法应用透射电镜、免疫组织化学及图像分析技术观察并检测小鼠后肾不同发育阶段集合管的超微结构及AQP-4表达。结果小鼠胚龄18 d见发育早期集合管主细胞,生后7 d～21 d,其形态结构发育基本完善。A型闰细胞于胚胎18 d出现,B型闰细胞生后21天出现。AQP-4于胚龄14 d始见表达,分布于集合管管壁上皮细胞侧基底细胞膜,随胚龄增加表达逐渐增强,生后1 d达高峰。结论集合管管壁上皮细胞于胚胎时出现,但其形态结构在生后才逐渐完善。AQP-4对小鼠胚胎时期肾水平衡调节可能起重要的作用。     

胚胎小鼠肾发育过程中nNOS的表达

目的观察神经型一氧化氮合酶(nNOS)在胚胎小鼠肾发育过程中的表达,探讨nNOS在小鼠肾脏发育早期的作用。方法选取胚龄(E)12、14、16、18 d及新生组(P0 d)小鼠,应用免疫组织化学技术观察nNOS的表达部位。应用体视学检测体密度值方法和免疫印迹技术检测各组小鼠肾组织中nNOS含量。结果免疫组化染色显示,E12 d,nNOS在输尿管芽呈现阳性表达;E14 d,nNOS在皮质生肾区、近端小管和远端小管表达。E16 d,nNOS在输尿管芽、生肾区和近端小管的表达减弱,在远端小管的表达加强。至P0 d,nNOS在输尿管芽、生肾区和近端小管的表达进一步减弱,在远端小管尤其是致密斑处呈强表达。从E12 d至P0 d,除了在成熟肾小体的肾小囊偶见nNOS的阳性表达外,在发育各期的肾小体中均未见nNOS的阳性表达。在集合管中也无表达。体视学分析结果显示,从E12 d至P0 d,各组小鼠肾组织中nNOS的体密度值逐渐增加。免疫印迹结果显示,从E12 d至P0 d,nNOS的表达量逐渐增加。结论在胚胎小鼠肾发育过程中,nNOS表达部位主要在远端小管致密斑,可能对远端小管的发育起重要的调节作用。     

缺血性急性肾损伤大鼠肾小管细胞凋亡及caspase-3表达

目的观察缺血性急性肾损伤(IAKI)大鼠肾小管细胞凋亡微细结构及caspase-3表达变化。方法应用光学和透射电镜技术、免疫组织化学和免疫印迹技术对缺血性急性肾损伤大鼠肾小管的细胞凋亡进行系统观察,并对caspase-3表达变化进行分析。结果肾缺血60 min再灌注24 h后,近髓质和髓质外带的近端小管出现了大量散在的凋亡细胞;凋亡细胞皱缩,核染色质固缩边聚,多数凋亡细胞脱落到小管腔内随尿液排除,小部分形成凋亡小体被邻近细胞所吞噬,在凋亡细胞内可见较多的自噬小体和自噬溶酶体。应用caspase-3免疫组化染色观察和免疫印迹分析结果显示,经caspase家族激活途径的细胞凋亡构成了IAKI诱导的近端小管细胞凋亡。结论大鼠肾缺血60 min再灌注24 h部分近端小管细胞发生了凋亡,推测这种细胞凋亡可能属于caspase依赖的细胞死亡;自噬作用参与了凋亡细胞处理细胞器的过程。     

肌上皮细胞的表型分化在涎腺发育及多形性腺瘤中的意义

背景与目的探讨肌上皮细胞与涎腺发生、涎腺多形性腺瘤发生的关系以及肌上皮细胞的分化状态与肿瘤生物学行为的关系。材料与方法采用组织学、免疫组化方法对不同发育阶段(胚7~8周、胚9~10周、胚11~14周、胚15~20周)的涎腺胚胎组织及涎腺多形性腺瘤中肌上皮细胞的表型分化及功能状态进行比较分析。结果在涎腺发育过程中,导管腔面及基底层细胞表达CK19,偶可表达CK14,而不表达肌上皮细胞的标记物α-SMA,为非肌上皮来源;而原始腺泡和闰管中肌上皮细胞的标记物阳性表达,为肌上皮前体细胞分化而来。在多形性腺瘤中,非管腔的肿瘤实质中,梭形细胞、上皮样细胞表达肌上皮细胞标记物CK14、P63、α-SMA,为肌上皮来源,软骨样成分和粘液样成分偶可表达肌上皮细胞标记物CK14和α-SMA,可能亦为肌上皮来源。管腔样结构、浆细胞样细胞、透明细胞不表达肌上皮细胞标记物,可能来自管腔细胞系。结论在涎腺发育过程中,腺泡和闰管来自肌上皮细胞系;导管系统来自管腔细胞系。以肌上皮分化较好的肿瘤预后较好,其中可能的原因为肌上皮细胞分化异常,失去其自身的表型特征,从而失去抑制肿瘤生长和侵袭的作用。     

诱导型一氧化氮合酶在糖尿病大鼠早期肾皮质中的定量分析

目的 观察I型糖尿病早期大鼠肾脏中诱导型一氧化氮合酶(iNOS)变化,并行定量分析.方法 成年健康SD大鼠60只,随机分成糖尿病组(DM组)和正常对照组(NC组),DM组采用一次性腹腔内注射STZ诱导制造糖尿病大鼠模型.成模后和NC组分别于第1、2和4周取材,取右肾用硝酸还原酶法和吸光度比较法测定肾组织中NO含量和NOS、iNOS的活性;左肾制作石蜡切片,行免疫组织化学染色,光镜下观察各组肾脏形态学变化及iNOS分布情况.结果 DM组大鼠肾组织中NO含量、NOS和iNOS活力较NC组均升高,具有显著性差异(P＜0.05),其中NO含量和NOS活力呈持续升高;iNOS活力却是于第2周达到高峰后随之降低.免疫组织化学染色,iNOS在DM组大鼠肾脏中,阳性部位定位于肾小球上皮细胞、系膜细胞、近端小管和远端小管上皮细胞胞浆,而且在第1、2和4周表达强度呈先高后低的变化.图像分析显示:iNOS在第2周平均光密度值显著低于第1周(P＜0.05)和第4周(P＜0.05).结论 I型DM大鼠早期肾皮质iNOS表达增加,其合成的NO在早期肾脏损害中发挥了重要作用.     

一种新的肿瘤免疫治疗载体--Exosome

Exosome是细胞的多泡体(MVB)与细胞膜融合后释放到细胞外环境中的单层膜包裹的小囊泡,它可从一种细胞到达另一种细胞,在细胞间传递多种分子.Exosome在免疫系统中具有调节作用,在实验性肿瘤免疫治疗中取得了显著效果,在正常生理状态下也具有重要功能.     

nm23参与α粒子辐射致大鼠气管上皮细胞恶性转化的原发过程

目的：克隆α粒子辐射致细胞恶性转化相关基因。方法：采用ｍＲＮＡ差异显示技术识别原代大鼠气管上皮细胞同由α粒子辐射诱发恶转的大鼠气管上皮细胞之间的差别表达基因。结果;共克隆到１５个可能同α粒子辐射致细胞恶性转化相关的差异表达基因片段,共向ＧｅｎＢａｎｋ登录７条新基因序列。其中克隆ＺＣ７６６同肿瘤转移抑制基因ｎｍ２３高度同源,经Ｎｏｒｔｈｅｒｍ杂交证实其在恶转的大鼠气管上皮细胞中高表达,序列分析提示ｎ     

甲状腺转录因子-1在肺癌细胞核中表达的定量研究

目的: 观察甲状腺转录因子-1 (TTF-1) 在正常成人肺泡Ⅱ型上皮细胞、人胚胎肺泡上皮细胞、肺癌以及淋巴结转移癌癌细胞胞核中的表达, 从量化角度探讨其在肺癌发生及其转移过程中的意义。方法: 石蜡包埋切片, 应用免疫组织化学SP法及LeicaQ500MC图像分析系统,对TTF- 1表达强度进行定量分析。结果: 胚胎肺泡上皮细胞胞核TTF 1阳性单位(PU) 值小于正常成人肺泡Ⅱ型上皮细胞胞核TTF- 1的PU值, 差异具有极显著性(P<0 .001); 不同类型肺癌癌细胞胞核TTF 1的PU值均小于胚胎肺泡上皮细胞和正常成人肺泡Ⅱ型上皮细胞胞核TTF -1的PU值, 且差异具有极显著性(P<0. 001); 肺腺癌和肺小细胞癌癌细胞胞核TTF 1的PU值均大于肺鳞癌和肺大细胞癌癌细胞胞核TTF 1的PU值, 且差异均具有极显著性(P<0 .001); 肺鳞癌癌细胞胞核TTF 1的PU值大于肺大细胞癌癌细胞胞核TTF- 1的PU值, 差异具有极显著性(P<0. 001)。肺腺癌、肺鳞癌和肺大细胞癌淋巴结转移灶中癌细胞胞核TTF 1的PU值均大于其癌原发灶癌细胞胞核TTF 1的PU值, 差异具有显著性(P<0 .001, P<0 .001,P<0. 05); 肺小细胞癌淋巴结转移灶癌细胞胞核与其原发灶癌细胞胞核TTF 1的PU值基本相同, 差异无显著性(P>0 .05)。有淋巴结转移的肺癌癌细胞胞核TTF 1的PU值大于无淋巴结转移     

鼻咽癌相关基因NAG4编码蛋白的表达模式

目的：明确人鼻咽癌相关基因NAG4编码产物的特性和活细胞内定位,了解其与癌变的关系。方法：构建了绿色荧光蛋白GFP与NAG4融合基因的真核表达载体,通过脂质体倡导分别转染非洲绿猴肾永生化细胞系COS7、人宫颈癌细胞系HeLa、人鼻咽癌细胞系HNE1以及原代培养正常鼻咽上皮PNNE,瞬时表达后荧光显微镜观察NAG4编码蛋白的活细胞内定位及其在鼻咽癌中改变。结果：NAG4基因编码产物在COS7细胞的胞浆和胞核内可见表达,而在HeLa细胞中仅在细胞浆存在;有30％的鼻咽癌细胞仅分布在胞浆,而在所有的正常鼻咽上皮细胞胞核和胞浆均有表达。结论：NAG4基因编码核转录蛋白在细胞恶变过程中可能有细胞浆到细胞核的移位障碍。     

Morphological and immunohistochemical studies of the estrogen-induced Syrian hamster renal tumor: probable cell of origin

Chronic natural or synthetic estrogen treatment of Syrian golden hamsters leads to the development of malignant renal neoplasms. In the present study, morphological and immunohistochemical studies were performed to further characterize the estrogen-induced hamster renal tumors. The neoplasms were composed of two distinct cell populations: a large-cell component that appeared highly epithelial, and a poorly differentiated small-cell component. Importantly, both cell types had epithelial characteristics, since they contained desmosomes at their cell surfaces. However, the large-cell component possessed additional epithelial features such as microvilli, intracytoplasmic lumens, and cilia. Comparative studies of renal tumors and developing renal tissue from fetal and newborn hamsters revealed remarkable histological similarities. Morphologically, the large tumor cells resembled early metanephric tubules and the small tumor cells were very similar to the blastemal cells of the developing kidney. The earliest tumor foci were found after 4.5 months of treatment. They were consistently found in the kidney interstitium in proximity to large arteries. Immunohistochemical staining for intermediate filaments in developing fetal and newborn kidneys demonstrated cytokeratin in renal tubules, desmin in blastemal cells, and vimentin in stromal cells. Estrogen-induced renal tumor cells uniquely possessed reactivity for all three intermediate filaments, clearly demonstrating their epithelial and mesenchymal characteristics. Based on their morphological resemblance to developing embryonic kidney cells and the presence of both epithelial and mesenchymal intermediate filaments, our findings provide strong evidence that the cell of origin of this malignant tumor is a precursor cell that is committed to an epithelial differentiation pathway.     

Transformation of kidney epithelial cells by a quinol thioether via inactivation of the tuberous sclerosis-2 tumor suppressor gene

Although hydroquinone (HQ) is a rodent carcinogen, because of its lack of mutagenicity in standard bacterial mutagenicity assays it is generally considered a nongenotoxic carcinogen. 2,3,5-Tris-(glutathion-S-yl)HQ (TGHQ) is a potent nephrotoxic metabolite of HQ that may play an important role in HQ-mediated nephrocarcinogenicity. TGHQ mediates cell injury by generating reactive oxygen species and covalently binding to tissue macromolecules. We determined the ability of HQ and TGHQ to induce cell transformation in primary renal epithelial cells derived from the Eker rat. Eker rats possess a germline inactivation of one allele of the tuberous sclerosis-2 (Tsc-2) tumor suppressor gene that predisposes the animals to renal cell carcinoma. Treatment of primary Eker rat renal epithelial cells with HQ (25 and 50 microM) or TGHQ (100 and 300 microM) induced 2- to 4-fold and 6- to 20-fold increases in cell transformation, respectively. Subsequently, three cell lines (The QT-RRE 1, 2, and 3) were established from TGHQ-induced transformed colonies. The QT-RRE cell lines exhibited a broad range of numerical cytogenetic alterations, loss of heterozygosity at the Tsc-2 gene locus, and loss of expression of tuberin, the protein encoded by the Tsc-2 gene. Only heterozygous (Tsc-2(EK/+)) kidney epithelial cells were susceptible to transformation by HQ and TGHQ, as wild-type cells (Tsc-2(+/+)) showed no increase in transformation frequency over background levels following chemical exposure. These data indicate that TGHQ and HQ are capable of directly transforming rat renal epithelial cells and that the Tsc-2 tumor suppressor gene is an important target of TGHQ-mediated renal epithelial cell transformation.     

Identification of a high-frequency model for renal carcinoma by the induction of renal tumors in the mouse with a single dose of streptozotocin

In order to develop a high-frequency, single-dose model of renal epithelial tumor induction in the mouse, streptozotocin (250 mg/kg) was administered i.v. to 6-week-old males and females of the CBA/H/T6J strain. In groups of 26 males and 30 females representing the effective number of survivors eligible for tumor estimates, renal cell tumors were found in 73% of males and 97% of females, including incidences of lesions diagnosed as renal carcinoma in 31 and 60%, respectively. The difference in renal tumor frequency between the sexes was not considered a real effect owing to the significantly decreased survival of the males. The neoplastic lesions ranged from small papillary or cystopapillary adenomas with a benign appearance to large, solid carcinomas with a low potential for metastasis. In the females, 22% of the larger tumors metastasized to distant sites, mainly the lungs. Intermediate lesions incorporating both papillary and solid adenomatous profiles suggested a sequential development of carcinomas along this pathway from the smaller papillary foci. The data establish the administration of a single dose of streptozotocin in the female CBA/H/T6J as a suitable high incidence model for the study of renal epithelial carcinogenesis in the mouse.     

Isolation of a morphologically-transformed epithelial cell-line from rat kidney following an in vivo dose of dimethylnitrosamine

In an in vivo-in vitro system, a rat kidney epithelial cell-line was isolated into continuous culture 48 h after a single administration intraperitoneally of dimethylnitrosamine (DMN). The dose used, 60 mg/kg, was known to induce approximately a 40% incidence of renal cortical epithelial tumors, in addition to a much higher incidence of renal mesenchymal tumors. The cell-line was established from several persisting islands of epithelial cells which were cultured in Leibowitz medium supplemented with various-growth enhancing agents and periodically, the antifibroblastic agent cis-hydroxyproline. Through succeeding subcultures, the cells retained an epithelial morphology and at confluence piled-up into 3-dimensional ridges and domes above the monolayer. The cells were characterized by continuous growth in culture, the ability to form colonies at very low seeding rates, loss of anchorage-dependence as determined by growth in semi-solid media and formation of multicellular aggregates in suspension culture, and agglutination in the presence of conconavalin A. These altered properties of growth and behavior indicate that the renal epithelial cells constitute a morphologically transformed line. As such, they may represent the same target cell population from which DMN-induced epithelial tumors arise in the rat kidney.     

Change in morphological and functional cytodifferentiation induced by seminal vesicle mesenchyme in cell suspensions of rat Dunning prostatic adenocarcinoma cells

Previous experiments have shown that seminal vesicle mesenchyme (SVM) can induce small 0.5 mm fragments of the rat Dunning tumor (DT) to undergo secretory differentiation with a concomitant reduction in tumorigenesis. In the present experiments Dunning tumor epithelial cells (DTE) were purified from DT cell suspensions by Percoll gradient centrifugation and recombined with neonatal rat SVM. The resultant tissue recombinants (SVM + DTE) were grafted under the renal capsule of male athymic mice and grown for 2 months. Under these conditions SVM induced the DTE to exhibit a highly differentiated secretory phenotype by forming ducts lined with tall columnar epithelial cells or large clear cells with pale cytoplasm. Undifferentiated epithelial cells of the parental DT were rarely observed in these tissue recombinants. The loss of tumorigenicity in SVM + DTE recombinants was associated with a striking reduction of epithelial 3H-thymidine labeling index in SVM + DTE recombinants (DT = 8.31%, SVM + DTE recombinants = 1.10%). Differences in putative secretory proteins were also observed by SDS-PAGE in SVM + DTE recombinants in comparison with DT. Testosterone metabolism was examined in epithelial cells recovered from grafts of DT vs. SVM + DTE tissue recombinants by thin layer chromatography and revealed that the major metabolite produced by DTE was androstenedione, whereas in epithelium isolated from SVM + DTE tissue recombinants the major androgen metabolite was 5α-DHT. Thus, after induction by SVM the DTE metabolized androgens in a pattern similar to the normal rat dorsal prostate. The SVM-induced changes in DTE suggest the possibility that emerging or established carcinomas might be regulated at least in part by their connective tissue microenvironment. © 1996 Wiley-Liss, Inc.     

人胎大肠和小肠上皮内淋巴细胞的比较分析

目的 :比较和分析人胎大小肠上皮内淋巴细胞 (iIEL)的发生、亚型分布和数量变化。材料和方法 :用免疫组织化学方法显示 6 2例小肠和 4 8例大肠CD3、CD4 、CD8、CD2 0 阳性淋巴细胞并进行体视学计数分析。结果 :小肠iIEL比大肠早一周出现 ,iIEL第 10周可见于小肠 ,第 11周可见于大肠 ;大小肠iIEL形态相同 ,位置相似 ,其数量均随胎龄而增加 ,但在各时期小肠iIEL均显著多于大肠 ,小肠iIEL约为大肠iIEL的 4 - 9倍。无论小肠还是大肠 ,均以T细胞为主 ,且以CD8 T细胞为主 ,CD4 T细胞很少 ;而固有层则有较多B细胞 ,且T细胞也是以CD4 T细胞为主 ,CD8 T细胞很少。结论 :大小肠iIEL有基本相同的发育规律 ,但发生时间和数量上有较大差异。     

Eruptive keratoacanthoma en plaque in an immunosuppressed patient

Immunosuppressive therapy predisposes to the development of a variety of neoplasms. A 63-year-old man developed multiple eruptive epithelial tumors 15 years after renal transplantation. The tumors included what had been diagnosed as squamous cell carcinomas, Bowen's disease, and solitary and eruptive keratoacanthomas. In addition, he developed a large plaque studded with small nodules that had the histologic features of keratoacanthoma. To our knowledge, this is the first example of acquired eruptive keratoacanthoma forming a large plaque.     

Development of resistance to the growth inhibitory effects of transforming growth factor beta 1 during the spontaneous transformation of rat liver epithelial cells.

The temporary maintenance of a rat liver epithelial cell population at confluence before passaging followed by periods of rapid proliferation resulted in the generation of spontaneous transformants after about 108 population doublings. The appearance of morphologically aberrant transformants correlated directly with an increased resistance of the population to the growth inhibitory effects of transforming growth factor beta 1 (TGF-beta 1). Clonal cell lines derived from the transformants were resistant to TGF-beta 1 dependent inhibition of DNA synthesis. These cell lines were also highly tumorigenic and aneuploid, with characteristic gross chromosomal abnormalities, and they expressed a number of phenotypic markers common to rat liver epithelial cells transformed by oncogenes or chemicals. In contrast, apparently normal looking cell lines cloned from the same population were nontumorigenic and near diploid, with few chromosomal abnormalities, and they were as sensitive to TGF-beta 1 as early passage normal rat liver epithelial cells. Morphologically normal late passage rat liver epithelial cells were sensitive to transformation by the DNA hypomethylating agent 5-aza-2-deoxycytidine, in contrast to earlier passage cells, and this transformation was accompanied by the development of resistance to the growth inhibitory effects of TGF-beta 1. These findings suggest that acquisition of resistance to the effects of growth inhibitors such as TGF-beta 1 is an important and possibly essential stage in the spontaneous transformation of rat liver epithelial cells.