Clonable T lymphocytes in T cell-depleted bone marrow transplants correlate with development of graft-v-host disease

Early clinical trials using T lymphocyte-depleted human marrow for transplantation have reported that such grafts reduce, to varying degrees, both the incidence and the severity of graft-v-host disease (GVHD). However, to date, no clear estimates have been made as to what degree of T cell depletion is necessary to prevent GVHD in every case. To address this problem, we used a limiting dilution assay (LDA) to quantitate residual clonable T lymphocytes in human T cell-depleted bone marrow in 31 HLA-identical transplants for leukemia. The number of phytohemagglutinin -interleukin 2-responsive T lymphocytes determined by LDA and expressed as T cell per kilogram recipient weight was found to correlate with the subsequent development of GVHD: no patients who received less than 1 X 10(5) T cell per kilogram developed GVHD (N = 24). Of the seven patients who received 1 X 10(5) to 4.4 X 10(5) T cell per kilogram, four patients developed grade I or II skin GVHD. This study thus provides a quantitative estimate of the number of T lymphocytes necessary to initiate clinically detectable GVHD in an HLA- identical host.     

Cell kinetic analysis of intact rat colonic crypts by confocal microscopy and immunofluorescence

BACKGROUND & AIMS: Precise quantitative and spatial analysis of cell cycle-related biomarkers in colonic crypts is often vital for studies of colon carcinogenesis and cancer prevention. To overcome the limitations of histology, confocal laser microscopy of microdissected whole crypts was used to quantitate S phase and mitotic cells. METHODS: Microdissected distal colonic crypts were studied in a modified rat starvation refeeding model. S phase cells were labeled in vivo with 5- bromodeoxyuridine. Mitotic cells were labeled with MPM2 (antibody to mitosis-specific epitope) and also assessed for chromatin morphology with propidium iodide. Sequential optical crypt sections, produced by confocal microscopy, were digitally imaged. S phase labeling indices per whole crypt were also compared with those derived by conventional immunohistochemistry. RESULTS: S phase and mitotic cells were clearly discriminated without background staining. The labeled S phase cell number and fraction per whole crypt were significantly decreased with starvation and increased with refeeding. Variability in the labeling index between whole crypts analyzed by confocal microscopy was significantly smaller than between histological crypt sections. Consequently, the intervention contributed to 92.2% of the total variability of the labeling index in whole crypts but only to 59% of the variability in histological sections. CONCLUSIONS: Major limitations of histology are overcome by crypt microdissection and confocal microscopic analysis. The total crypt cell population as well as labeled M phase and S phase cells can be imaged, localized, and quantitated with improved precision. (Gastroenterology 1996 Dec;111(6):1493-500)     

A K-ras oncogene increases resistance to sulindac-induced apoptosis in rat enterocytes

BACKGROUND & AIMS: Mutations of c-K-ras occur commonly in colonic neoplasms. The aim of this study was to determine how c-K-ras mutations alter the responses to the chemopreventive agent sulindac. METHODS: The parental rat intestinal cell line IEC-18 and c-K-ras-transformed derivatives were treated with sulindac sulfide. Cell cycle distribution was determined by flow-cytometric analysis (fluorescence-activated cell sorter), apoptosis by DNA fragmentation (laddering), flow cytometry, and microscopy, and changes in gene expression by immunoblotting. RESULTS: Sulindac sulfide inhibited cell growth and induced apoptosis in a time- and dose-dependent manner more rapidly in and at lower concentrations in parental cells than ras-transformed cells. Expression of the sulindac sulfide arrested cells in G0/G1, but cells entered apoptosis throughout the cell cycle. Proapoptotic protein Bak was relatively high in untreated parental cells and increased markedly after sulindac sulfide but was low in untreated ras-transformed cells and did not increase after sulindac sulfide. Expression of other Bcl-2 family members was unchanged after sulindac sulfide. However, sulindac sulfide reduced levels of cyclin D1 protein and cyclin E- and cyclin D1- associated kinase activity. CONCLUSIONS: c-K-ras-transformed enterocytes are relatively resistant to sulindac sulfide-induced growth inhibition and apoptosis, which may result from specific reduction of bak expression. (Gastroenterology 1997 Dec;113(6):1892-900)     

中国六城市中老年人群腰椎骨关节炎患病危险因素地区调查：6128名资料分析

目的:腰椎骨关节炎所致疼痛和关节活动受限等,严重影响中老年患者的生活质量。了解中老年人群腰椎骨关节炎患病危险因素、患病率及疾病与暴露因素之间联系强度的指标,可为腰椎骨关节炎的社区预防干预提供量化性参考数据。方法:调查于2005-07/08在西安、石家庄、上海、广州、哈尔滨、成都等六城市完成。①调查对象:选择40岁及以上具有正式户口常住男女人群6218名,其中男2916名,年龄40～94岁;女3302名,年龄40～86岁;男女性别比为0.88∶1.00。纳入标准:中国汉族在本地具有正式户口的居民并对本次调查知情同意。排除标准:因各种原因外出超过1年以上居民不属本次调查对象。②调查方法:采用分层多阶段整群抽样方法对中国六城市(西安、石家庄、上海、广州、哈尔滨、成都)6218名符合调查对象人群进行腰椎骨关节炎的流行病学问卷调查(调查内容包括一般情况、现病史、既往史、体格检查、X射线片检查情况、疾病诊断),并对其中4808例有症状者进行X射线片腰椎正侧位投照,对83个变量进行多因素非条件Logistic回归分析。表示疾病与暴露因素之间联系强度的指标用比值比(OR),若OR>1,说明疾病发生危险性增加,与暴露因素呈正关联;若OR<1,说明疾病发生危险性减少,与暴露因素呈负关联。结果:纳入调查对象6218名,全部进入结果分析。①中国六城市40岁及以上男女人群腰椎骨关节炎总患病率为29.4%,各城市患病率差异有非常显著性意义(P<0.01)。②腰椎骨关节炎在大部分城市有共同的危险因素如年龄因素(OR=1.031～1.121),使用蹲位排便(OR=1.012-1.044)和共同保护性因素如饮酒(OR=0.241～0.930);而日常骑摩托(OR=3.122,西安),专职体育(OR=6.377,石家庄),骨质疏松史(OR=5.925,石家庄),神经关节病史(OR=5.528,上海),日常乘公共汽车(OR=5.784,广州),动脉粥样硬化史(OR=4.797,广州),糖尿病史(OR=4.064,广州),日常爬楼梯(坡)(OR=1.018,哈尔滨),兄弟骨关节炎史(OR=11.791,成都)等危险因素分别在不同地区出现;而常吃水果(OR=0.725,西安),喝啤酒(OR=0.507,石家庄)等保护性因素也分别在不同城市出现。结论:中国六个地区腰椎骨关节炎患病有共同的危险因素和保护性因素,同时,不同地区主要危险因素又有一定差异。