Immunohistochemical study of proteins linked to apoptosis in rat fetal kidney cells following prepregnancy adriamycin administration in the mother

Adriamycin is an antibiotic of the anthracycline group. In a previous study, we showed that administration of a single dose of adriamycin (i.p. injection, 5mg/kg body weight) 4 weeks before pregnancy in female Wistar rats induced histological changes in the fetal renal cells typical of apoptosis and also over-expression of heat shock proteins (HSP70). Using a similar experimental model, we have now examined renal cells in fetuses (gestation day 20) to investigate the pathways of the transduction signal of apoptosis in these cells that is induced by prepregnancy maternal administration of adriamycin. Immunolocalization of several proteins - p53, Bax, Apaf-1 and caspase 9 - which take part in the mitochondrial pathway of apoptosis and caspase 12, which takes part in the endoplasmic reticulum pathway of apoptosis, was determined. The results showed that adriamycin administered to the mother rat before pregnancy subsequently induced changes in fetal kidneys involving both the mitochondrial pathway of apoptosis, with increased labeling of the proteins p53, Bax, Apaf-1 and caspase 9, and the endoplasmic reticulum pathway of apoptosis, with increased labeling of caspase 12. Immunolabeling of these proteins was quantified using an image analysis program.     

真武汤对阿霉素所致大鼠肾损伤的治疗作用

 目的：研究真武汤对阿霉素肾病模型（adriamycin nephropathy,AN）大鼠足细胞裂孔隔膜蛋白分子（podocin和nephrin）表达的影响,探讨其防治阿霉素肾病大鼠蛋白尿的机制。方法：采用常规生化、病理方法（包括HE染色、Masson染色及电镜）观察真武汤对AN模型所致纤维化大鼠肾功能、肾组织形态学变化及羟脯氨酸（Hyp）含量的改善作用;采用蛋白免疫印迹等方法探索真武汤对足细胞标志蛋白podocin和nephrin信号分子表达的影响。结果：模型组大鼠尿蛋白（TP）、血尿素氮(BUN)和血清肌酐（SCr）显著增加,肌酐清除率（CCr）显著下降（P<0.05）;Hyp显著增加（P<0.05）;肾组织podocin和nephrin蛋白表达水平显著降低（P<0.05）;肾小管萎缩,基底膜增厚;足突扁平、融合、消失。肾小球集中现象明显;部分肾小管扩张,肾小管上皮细胞变性,蛋白管型明显;肾间质纤维组织增生和较多炎症细胞浸润。与模型组相比,各治疗组TP、BUN和SCr均有一定程度的下降,CCr显著提高;Hyp明显下降（P<0.05）;真武汤组肾组织podocin和nephrin蛋白表达水平显著提高（P<0.05）;肾组织病变程度轻于模型组。结论：真武汤能减少阿霉素肾病大鼠肾组织羟脯氨酸含量,改善肾功能及减轻病理损伤。 真武汤降低模型大鼠蛋白尿的作用可能与其维持足细胞podocin和nephrin的表达有关。     

Study of the activation mechanism of adriamycin on rat mast cells

Adriamycin (ADR) induces nonimmunological and noncytotoxic histamine release from peritoneal and pleural rat mast cells. This secretion is unaffected by the pretreatment with pertussis toxin, cholera toxin and benzalkonium chloride. Histamine release induced by compound 48/80, was markedly inhibited by pertussis toxin, cholera toxin, benzalkonium chloride and neuraminidase. The ADR dose-response curve is significantly shifted to the right when cells were preincubated with the unspecific phosphodiesterase inhibitor IBMX. The activation of protein kinase C (PKC) with the phorbol esther TPA increases the response to ADR, while PKC inhibition with trifluoperazine decreases histamine release. The pretreatment of mast cells with okadaic acid did not modify the response to ADR. these results suggests that ADR elicits histamine release with a mechanism notably different from compound 48/80.     

Comparison of two decalcification agents using microwave technology: a histochemical assessment of the rat cochlea

Despite the prevalence of hearing loss, there are no FDA-approved chemical entities available for either otoprotection or treatment of hearing impairment. This lack of drug-based treatments has made the assessment of the middle and inner ear a rare, challenging exercise in the pharmaceutical industry. Recently, advances in the use of various decalcifying agents, as well as heating techniques utilizing microwave technology have proven to be effective at shortening processing times, minimizing tissue damage and potentially lowering the cost of histopathological evaluation and diagnosis. This study tested decalcification agents in order to determine the most efficient decalcification procedure for rat outer, middle, and inner ear structures while preserving tissue morphology and eliminating introduction of artifacts to best facilitate histopathological analysis. Presented here are the results of rat ear histological preparations using two decalcifying agents at increasing microwave times with heat controlled at the physiological temperature of 37 °C. In this analysis, an acid-based combination decalcification-fixative solution was found to significantly speed up the decalcification process when compared with the calcium chelator EDTA.     

葛根素抗培养脑细胞阿霉素损伤作用

目的: 观察葛根素抗培养脑细胞阿霉素损伤作用。方法: 建立培养脑细胞阿霉素损伤模型, 观察葛根素对线粒体脱氧酶活性及细胞超微结构的影响。结果: 葛根素可剂量依赖性提高阿霉损伤脑细胞内线粒体脱氢酶活性及减轻细胞超微结构的损伤。结论: 葛根素对阿霉素损伤培养脑细胞具有保护作用。     

Semi-automatic counting of connexin 32s immunolocalized in cultured fetal rat hepatocytes using image processing

Connexin 32s (Cx32s) were immunolocalized in fetal rat hepatocytes and their distribution was determined qualitatively. We used image analysis using a quantitative index (QI) of Cx32 (QI Cx32) defined as the area of Cx32s/number of cells in cultured fetal rat hepatocytes. Hepatocytes from livers of fetal rats were separated by collagenase digestion and low centrifugation on gestational day 17. Cells were cultured for 3 days in dexamethasone (DEX)-supplemented medium (Dex0). The medium was replaced with fresh medium and cells were continuously cultured for 3 days with DEX or epidermal growth factor supplemented medium (Dex3 or EGF3). After culture termination, cells were fixed and stained using the fluorescein-labeled antibody method for Cx32s and diaminophenylindole staining for nuclei. Thirty pairs of histological images for Cx32s and nuclei, 180 images in total, were obtained from each condition. The QI Cx32 significantly increased from 284.1 ± 102.0 (mean and SD, n = 26) of Dex0 to 428.9 ± 101.0 of Dex3 (n = 28) (P < 0.05, Kruskal-Wallis test, then Steel-Dwass test). The increase of QI Cx32 was compatible with the morphological observations. The image analysis processing time after preparation for 180 images was reduced from 8 h needed for manual operations to 1 min using ImageJ software with our macro routine.     

白蛋白过载加重阿霉素肾病小鼠肾损害及辛伐他汀的肾脏保护作用

目的:探讨白蛋白过载是否可以通过脂质肾毒性加重阿霉素肾病小鼠肾脏损害及辛伐他汀的肾脏保护作用。方法:雄性BALB/c小鼠随机分为对照组、阿霉素肾病组(ADR组)、白蛋白过载阿霉素肾病组(ADR+BSA组)和白蛋白过载阿霉素肾病辛伐他汀治疗组(ADR+BSA+SIMV组),所有小鼠行左肾切除术,第2周末构建阿霉素肾病模型,第6周末开始构建白蛋白过载模型。0、2、6、10、14周末检测24 h尿蛋白;14周末收集标本,检测血清生化指标;光镜和电镜观察肾脏组织病理;酶比色法和油红O染色法检测肾脏组织脂质水平;real-time PCR法检测肾组织IL-1β、TGF-β1和低密度脂蛋白受体(LDLr)的mRNA表达;免疫组化法检测IL-1β和IL-17的蛋白水平;ELISA检测肾组织匀浆上清IL-17的分泌。结果:与对照组相比,ADR组肾组织IL-1β、TGF-β1、IL-17和LDLr的表达增高(P0.05),肾组织内胆固醇含量升高(P0.05)。ADR+BSA组较ADR组尿蛋白及血清肌酐水平增加,IL-1β、IL-17及LDLr表达均显著增加(P0.05),肾组织脂滴沉积及肾小球硬化加重。而ADR+BSA+SIMV治疗组上述炎症因子表达下调,肾组织内胆固醇含量减少,肾小球硬化程度降低。结论:阿霉素肾病小鼠肾组织内炎症介质可能通过上调LDLr导致肾脏局部脂质沉积,加重肾脏损害;白蛋白过载可进一步加重这一进程;而辛伐他汀可通过降低炎症介质及LDLr的表达减少脂质沉积,对肾脏起保护作用。     

Morphological effects of a large single dose of cycloheximide on the intestinal epithelium of the rat

Adult male rats received 15 mg/kg cycloheximide and the subsequent morphological effects at three and six hours after injection were evaluated using histometry, light and electron microscopy, histological demonstration of terminal web and acid phosphatase, and radioautography with tritiated thymidine. Rapid atrophy of the villi took place, progressing from the villus tip by premature exfoliation of epithelial cells. The crypts also diminished by random exfoliation of many crypt cells and by partial or complete disintegration. Mitosis and epithelial cell migration were absent. By six hours, the area occupied by the villi and the crypts per unit length of histological section was decreased by about 70–90% in most of the small intestine but only by about 40–60% in the duodenum and the terminal ileum. In the upper half of the villi, the epithelium was strongly positive for acid phosphatase and contained large numbers of round bodies resembling primary lysosomes. In the lower half, the microvillous border and terminal web were found to be disrupted. Animals receiving only 5 mg/kg cycloheximide also showed the atrophy of villi and crypts, and the round bodies resembling lysosomes. Evidence from several sources has indicated that protein synthesis in normal villus epithelial cells subsides toward the villus tip and becomes minimal at exfoliation. At exfoliation, proteins responsible for epithelial cohesion probably fail because they are no longer replenished. Cycloheximide appears to accelerate this process.     

Introduction to a special issue on kidney development and disease

Enriching our understanding of the anatomy of the kidneys, in development, health, and disease, has been the primary focus of Professor John Bertram's distinguished research career to date. Among other notable achievements, his landmark analyses of nephron number in over 400 human kidneys (the Monash Series), and his refinement of stereological techniques for renal structural analyses, have proven him an international leader in renal anatomy research. In this Special Issue, we (some of John's collaborators, colleagues, and former students) celebrate John's career with a series of 20 review and original research articles relevant to his expertise: (a) renal anatomy, physiology, and pathology, (b) kidney development, podocyte biology, and applications of renal stem cells, (c) renal developmental programming, and (d) contemporary methodologies in renal research; his accomplishments as a Head (Chair) of an Anatomy Department are also illustrated. We hope that this collection will serve as both an important resource, and a source of inspiration, to renal anatomy researchers and educators alike.     

产前超声诊断胎儿肾脏异常与妊娠结局的临床分析

目的探讨产前超声诊断胎儿肾脏异常与妊娠结局的关系。方法对2004年3月∽2013年7月产前超声筛查提示胎儿肾脏异常,并且在复旦大学附属妇产科医院产前诊断中心进行多科会诊的305例孕妇进行产前及新生儿随访观察。结果305例孕妇中有37例失访;2例妊娠中期流产;3例妊娠晚期胎死宫内;64例引产终止妊娠,其中30例为优生引产;199例分娩并随访至新生儿期,其中新生儿死亡4例、需手术治疗20例,余175例转归良好。结论产前B超提示胎儿肾脏异常不是优生引产的指征,排除致死性多发畸形和胎儿染色体疾病后,仅约14.2%的胎儿预后不良。     

Effects of exogenous ACTH on the rat fetal adrenal cortex: a histochemical and electron-microscopical observation.

ACTH was administered subcutaneously to rat fetus directly at the late stage of fetal development and acute reaction on the fetal adrenal cortex was observed histochemically and electron microscopically. By administration of ACTH the adrenal cortex became remarkably hyperemic and there were swollen cells in all layers, particularly in the middle and inner layers (corresponding to the zona fasciculata and reticularis in adult rat). Marked reduction of lipid, enlarged mitochondria with increased vesicular cristae and increased smooth-surfaced endoplasmic reticulum (SER) were characteristic. The alterations of mitochondria preceded the change of SER, and thereafter mitochondria showed rapid degeneration. The outer layer (corresponding to the zona glomerulosa in adult rat) also showed similar changes by ACTH to those of the other two layers. These results indicated that the fetal adrenal cortex of rats was exogenous ACTH-reactive and its reaction which was different from that of adult cortical cells, seemed to be specifically related to the development and differentiation of the cells.     

Histological study of the regrowth of a human melanoma xenograft exposed to single dose irradiation

The aim of this investigation was to find the location and the histological characteristics of cells that were the source of tumour regrowth after single dose irradiation. A human melanoma xenograft was irradiated with a single dose of 25.0 Gy which gives local tumour control in nearly 50% of treated animals. Serial histological sections were made from tumours removed during the first two weeks after irradiation. During the first week the perivascular organization of tumour parenchyma disappeared and the central part of the tumour became necrotic. The occurrence of vascular stasis, thrombosis and endothelial cell changes indicated that radiation injury to the vascular system was involved in the disappearance of the tumour cords. Tumour cells that remained histologically intact were located in subcapsular areas. The incidence of normal mitotic figures increased, and the fraction of abnormal mitoses and the incidence of micronuclei decreased 5 to 7 days before macroscopical regrowth was usually detected. It is concluded that cells which are the source of tumour regrowth were located in the subcapsular areas at the time of irradiation. Radiation injury to the tumour vascular system was an important factor in the necrotization of the tumour centre after treatment.     

Effect of the antioxidant phenosan on physicochemical properies of rat kidney cell membranes during carcinogenesis induced by nitrosodimethylamine

Laboratory of Biochemistry, Research Institute of Urology, Ministry of Health of the RSFSR, Moscow. Department of Pathological Physiology, Orenburg Medical Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR N. A. Lopatkin.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 106, No. 11, pp. 557–559, November, 1988.