DNA ploidy and S-phase fraction of neoplastic and non-neoplastic lesions of the human gallbladder

Studies on the cell kinetics of the human gallbladder are difficult because of epithelial degeneration by bile. Using the epithelial isolation technique, however, we were able to determine the degree of degeneration and to examine the cell kinetics of gallbladder lesions in freshly resected surgical specimens. Normal and neoplastic epithelial were isolated nonenzymatically from freshly resected gallbladder. The nuclear DNA content and S-phase fraction were estimated in 110 patients with gallbladder lesions by flow cytometry (FCM). Normal tissues and all lesions except carcinomas were diploid. The S-phase fraction of gallstone cases was significantly higher (1.47 ± 0.70%; mean ±SD) than normal (0.79 ± 0.39%) (P <0.0006). All gallbladder carcinomas were multiploid, and their S-phase fraction was 11.63 ± 3.65%. Cell renewal of normal gallbladder is low. In the gallstone cases, the S-phase fraction was increased, possibly correlated with carcinogenesis. © 1996 Wiley-Liss, Inc.     

Prognostic potential of flow cytometric S-phase and ploidy prospectively determined in primary breast carcinomas

In a prospective study of a consecutive breast cancer series accumulated in the period 1978–82, the S-phase fraction (SPF) and ploidy status were determined by flow cytometry performed on cell nuclei derived from samples of 580 primary tumors. Sixty percent of the tumors were non-diploid. After correction for debris the median SPF values were 7.3% overall, 12% for non-diploid tumors, and 2.9% for diploid tumors (2.6% when nodal subsets N2 and N3 and cases with metastases at presentation were excluded). The SPF values correlated both to tumor size (p=0.008) and to the number of positive axillary lymph nodes (p=0.03).At clinical follow-up in 1986, 467 unilateral breast cancer patients who had undergone radical treatment for cure could be evaluated with respect to the prognostic value of both the SPF value and ploidy status. The median duration of follow-up was then 59 months (range 2–90), and the median time-to-recurrence 24 months (range 2–69, n=137).At follow-up in 1991, 201/467 of the patients had died, the median duration of follow-up being 50 months (range 2–126) for the deceased, and 119 (range 6–148) for the survivors. In multivariate analysis (Cox's proportional hazards models), the strongest independent predictors of distant recurrence-free survival (DRFS) were the number of positive axillary lymph nodes (p<0.0001), the debris-corrected SPF value alone (p=0.003,versus p=0.05 for uncorrected value), and ploidy status combined with the corrected SPF value (p=0.0002). When age was taken into account, both the corrected SPF value and the ploidy-SPF combination were predictors of crude survival (p=0.006 and p=0.002, respectively).In univariate life-table analysis, the 5-year DRFS rate was 93% in node-negative (N0) cases with an SPF<7.3%, as compared to 80% in those with an SPF7.3% (p=0.005). Among node-positive cases, the prognostic value of the SPF was confined to those with 1–3 positive nodes, the 5-year DRFS rate being 68% in cases with an SPF<7.3%, as compared to 40% in cases with an SPF7.3% (p=0.01).Ploidy status and SPF were combined to form four groups: diploid & SPF<2.6% (DL), diploid & SPF2.6% (DH), non-diploid & SPF<12% (NDL), and non-diploid & SPF12% (NDH). Among node-negative patients, the DRFS rate fell from 95% in the DL group to 87% in the NDL group, with the DH group at an intermediate level, as compared with 74% (p=0.03) for the NDH group which accounted for the bulk of the early distant recurrences. Among patients with 1–3 positive lymph nodes, the 5-year DRFS rate was 68% in both the groups with low SPF values (DL and NDL), as compared with 45% in the DH group (p=0.03), and 37% in the NDH group (p=0.006).In this study, the flow cytometry SPF value, alone or in combination with ploidy status, yielded the most profound additional prognostic information, enabling both node-negative patients with a high probability of cure and patients at risk of early relapse to be identified. Among node-positive patients, the prognostic value of the SPF value was confined to those with 1–3 positive axillary lymph nodes (the predominant node-positive subgroup), enabling a high and a low DRFS rate subgroup to be distinguished – a useful distinction where selection for adjuvant drug treatment is concerned. As the predictive strength of the SPF value was enhanced when correction was made for debris, we would recommend that the effect of such factors as debris be minimized as far as possible when flow cytometry-derived SPF values are to be used for prognostic purposes.     

DNA ploidy and S-phase fraction in medullary carcinoma of the breast -a flow cytometric analysis using archival material

Summary In a population of 110 primary breast cancers with medullary features, registered in the Danish Breast Cancer Cooperative Group (DBCG) from 1977-82, we have determined ploidy and S-phase fraction (SF) by flow cytometry (FCM) on paraffin embedded tumour tissue. The distribution of DNA ploidy is not different from the distribution described for breast cancers in general. No difference is found between the subgroups of medullary and non-medullary cancer when using a new simplified histopathological definition of medullary carcinoma of the breast, recently proposed by us. When using the definition proposed by Ridolfiet al. in 1977, we find significantly more tumours with aneuploidy and high SF in the groups of typical medullary carcinoma (TMC) and atypical medullary carcinoma (AMC) than in the small group of non-medullary carcinoma (NMC), which seems a paradox, as patients with NMC have the worst prognosis. However, the number of patients in the NMC group is very small, and the percentage of aneuploid tumours is very low. In 84 protocolled patients we found no statistically prognostic importance of ploidy or SF, either in the whole group assessed or when stratifying for the histopathological subgroups. However, a prognostic influence of SF can be traced for the non-medullary cancers, according to the new definition, but not for the medullary cancers of the breast. The result emphasizes the impression of MC as being biologically different from other histological types of breast cancer.     

胶质瘤DNA倍性及增殖活性测定的临床意义

目的　检测胶质瘤DNA倍体特性、DI、SPF及PI值 ,探讨它们与胶质瘤级别的关系及临床意义。方法　 46例胶质瘤石蜡标本按级别分为Ⅰ级组 (15例 ) ,Ⅱ级组 (13例 ) ,Ⅲ～Ⅳ级组 (18例 ) ,10例正常脑组织作为对照组。用流式细胞技术 (FCM)检测其DNA含量、DI、SPF及PI值。结果　对照组均为二倍体 ;肿瘤组DNA异倍体率为 47 8% ;随着胶质瘤级别的增高 ,其DNA异倍体率、DI值、SPF值、PI值均增高。结论　DNA异倍体是恶性胶质瘤特征性标志 ;DI值 ,SPF值、PI值与胶质瘤的级别呈正相关     

细胞DNA倍体图像分析在宫颈癌及癌前病变筛查中的应用

目的利用显微图像分析技术对宫颈脱落细胞做DNA倍体检测、分析,以探讨该技术在宫颈癌及癌前病变筛查中的应用价值。方法对619例宫颈脱落细胞样本分别采用常规细胞学镜检和细胞DNA倍体分析法作检测,其中阳性样本患者行组织活检,并将3种检测结果进行对比分析。结果619例样本,常规细胞学镜检共检出ASC—US42例、ASC—H6例、LSIL29例、HSILl8例;采用细胞倍体分析法检出DI≥2．5时,可见DNA倍体异常细胞（1～2个）的样本102例、DNA倍体异常细胞（≥3个）的样本41例、DNA倍体异常细胞（≥25个）的样本14例;48例组织活检样本检出炎症7例、CINI20例,CINⅡ13例、CINⅢ2例、IC6例。CINⅡ～Ⅲ对应TBS系统中的HSIL计算灵敏度为81．0％;对应SIL和对应ASC＋SIL的灵敏度分别为85．7％和100％。HSIL与CINⅡ～Ⅲ比较有非常显著的统计学意义。CINⅡ～Ⅲ对应DNA倍体异常细胞（≥25个）计算灵敏度为66．7％;对应可见DNA倍体异常细胞（≥3个）的灵敏度为100％。DNA倍体异常细胞（≥25个）与CINⅡ～Ⅲ比较也有非常显著的统计学意义。结论细胞DNA倍体图像分析法在宫颈癌及癌前病变筛查中有较高的灵敏度,若联合常规宫颈细胞学检查,可明显提高宫颈癌筛查的质量控制水平。     

DNA ploidy and S-phase in recurrent astrocytomas: a retrospective study by flow cytometry of deparaffinized specimens

Summary Twenty-two patients with recurrent astrocytic tumors were treated surgically two or even three times. At the time of the first surgery 6 tumors were fibrillary astrocytomas (grade II), 9 anaplastic astrocytomas (grade III) and 7 glioblastomas (grade IV).Histopathological specimens from second surgery demonstrated in 12 cases signs of higher grades of malignancy. Flow cytometry (FCM) did not reveal any significant changes of S-phase fraction (p = 0.55). This study supports the theory that, given enough time, the histopathology of brain tumors change significantly from more benign forms to more malignant ones. The flow cytometry (FCM) could trace a weak tendency to higher S-phase fractions at the time of the second surgery. No apparent change of ploidy pattern was observed. In spite of the unequivocal histopathological changes of the recurrent astrocytomas the flow cytometry failed to indicate similar changes in terms of ploidy and S-phase fraction parameters.     

前列腺癌的细胞核DNA图像分析研究

目的探讨DNA图像分析技术在前列腺癌(PC)诊断中的应用价值。方法应用计算机图像分析技术测定对5例正常前列腺(NP)和30例PC的标本DNA倍体。结果5例NP中均为整倍体(2C和3~4C),30例PC中有23例出现非整倍体,并且随着PC分级的增高,其倍体率也增大。结论细胞核DNA含量和倍体测定是反映前列腺癌细胞增殖能力的重要生物学定量指标,能较客观反映PC的预后,为正确诊断恶性肿瘤提供依据。     

Prognostic impact of DNA ploidy pattern,S-phase fraction (SPF), and proliferating cell nuclear antigen (PCNA) in patients with primary gastric lymphoma

BACKGROUND: DNA ploidy, S-phase fraction (SPF), and proliferating cell nuclear antigen (PCNA) are considered to be significant prognostic factors in non-Hodgkin lymphomas. However, reports on their prognostic importance in gastric lymphoma patients are relatively lacking. METHODS: In the present study, we retrospectively studied the above-mentioned parameters in 29 patients with primary gastric lymphoma; 11/29 had B-low grade mucosa associated lymphoid tissue lymphoma (B-MALT), while 18/29 had diffuse large B-cell lymphoma (DLBCL), according to WHO classification. Proliferative activity was studied by staining against PCNA; in addition, the prognostic significance of DNA ploidy and SPF, as determined by flow cytometry, were investigated and compared to the results of the PCNA stainings. RESULTS: Seven out of 29 patients were found to have aneuploid tumors; DNA index values were >1 for all aneuploid lymphomas. There was no difference in DNA aneuploidy in MALT vs. DLBCL. The mean percentage of SPF was 11.4. SPF was found significantly lower in MALT vs. DLBCL (P < 0.05). The mean percentage of PCNA positive tumor cells was 52.6. PCNA protein expression was significantly lower in MALT vs. DLBCL (P < 0.0001). There was a significant positive correlation between PCNA score and SPF (P < 0.01, by Spearman analysis). DNA ploidy had no impact on survival in the present study. Both SPF and PCNA expression were important prognostic factors in the univariate analysis; however, in the multivariate analysis, the only independent prognostic factor for survival was PCNA expression. CONCLUSIONS: These findings indicate that SPF and PCNA are significant prognostic factors in patients with primary gastric lymphomas. However, in the present study, DNA ploidy had no impact on survival in patients with primary gastric lymphomas.     

DNA ploidy as a major prognostic factor in resectable ampulla of vater cancers

The archival paraffin-embedded specimens from 63 ampulla of Vater cancers after pancreaticoduodenectomy between 1965 and 1991 were analyzed by flow cytometry. Of the 63 cancers, 31 (49.2%) were diploid DNA cancers and 32 (50.8%) were aneuploid. Patients with diploid DNA cancer had a median survival time of 159.0 months, and patients with aneuploid DNA cancer had 24.0 months. This difference is statistically significant (P = 0.0257). The aneuploid group did have a poorer prognosis than the diploid group. The multivariate analysis demonstrated that DNA ploidy was an independent and very important prognostic factor, even stronger than the stage and lymph node status. There was a tendency toward higher values of S-phase fraction, proliferative index, and total aneuploid DNA fraction in the shorter survival groups, but they were of no statistical significance. These data suggest that DNA ploidy appears to be the most important and the only pre-operative predictor of prognosis in resectable ampulla of Vater cancers since endoscopic biopsy is feasible. © 1993 Wiley-Liss, Inc.     

A comparison of static cytofluorometry and flow cytometry for the estimation of ploidy and DNA replication in human breast cancer

Summary 332 primary invasive breast carcinomas were analysed by static cytofluorometry and flow cytometry. The ploidy distributions were similar, and 54% of the tumors were judged DNA aneuploid by both methods.The coefficient of variation of the G₀–G₁ peaks ranged from 2.0 to 8% with both techniques, but the mean was somewhat lower with flow cytometry — 4.1%, compared to 4.9% for the static measurements.The proportion of S-phase cells was possible to estimate from 80% of the flow histograms and 70% of the static histograms. S-phase was not estimated from the static histograms if less than 150 tumor cells were measured. With 160 tumors S-phase was measured by both methods. The range was 0 to 27% with the static measurements and 0.7 to 25% with flow cytometry. Corresponding mean values were 7.6% and 8.2%, which are similar to thymidine labeling index results with breast cancers reported in some studies. A close correlation was obtained (r = 0.927) comparing S-phase fractions estimated from aneuploid tumors with flow cytometry and static cytofluorometry if more than 200 cells were measured with the latter. The proportion of S-phase cells was significantly lower for the diploid tumors.We conclude that both techniques can be useful for the estimation of DNA ploidy and replication in human breast cancer.     

Prognostic significance of DNA ploidy in patients with stage II colorectal cancer

DNA ploidy status associated with colorectal cancer has been investigated widely in recent years. But the relationship between DNA analysis and prognosis has not been confirmed.[1(3] Some investigations showed that DNA aneuploid status was associated with lymph node involvement and poor differentiation, and thus may predict a poor clinical outcome in patients with colorectal cancer. These observations suggest that DNA ploidy abnormality may influence the development and progression of colo…     

鼻咽癌新鲜肿瘤组织DNA倍体性与预后的关系

背景与目的:因肿瘤有生物学异质性,部分肿瘤的预后和TNM分期并不符合;寻找有效的生物学预后指标作为临床分期的补充,可为今后鼻咽癌的个体化治疗提供一个新的依据.本研究探讨初治鼻咽癌患者新鲜肿瘤组织细胞的DNA倍体性与疗效、预后的关系.方法:1999年1月至2000年2月,53例初治鼻咽癌患者进入本研究,其中单纯放疗32例,另21例患者于放疗第4周接受了一个疗程的PF方案化疗.患者治疗前均活检取新鲜肿瘤组织,用流式细胞仪进行DNA倍体检测.结果:53例患者中,二倍体32例(60.4%),异倍体21例(39.6%).不同倍体组患者的年龄、性别、临床分期、N分期、化疗与否的差异无统计学意义(P=0.695、0.657、0.088、0.972和0.335).全组患者中位随访时间73个月(12～84个月).全组5年总生存率65.61%,其中二倍体组为80.92%,异倍体组为42.86%(P=0.002);5年无远处转移生存率二倍体组为84.26%,异倍体组为44.53%(P=0.003);5年无复发生存率二倍体组为92.59%,异倍体组为72.65%(P=0.118).单因素分析结果显示,临床分期是无复发生存率的影响因素,DNA倍体性、临床分期和T分期是总生存率和无远处转移生存率的影响因素.多因素分析结果显示,与总生存率相关的独立预后因素为DNA倍体性(P=0.020)和临床分期(P=0.007),与无转移生存率相关的预后因素亦为DNA倍体性(P=0.017)和临床分期(P=0.011).结论:采用流式细胞术检测新鲜组织细胞的DNA倍体性和临床分期一样可以预测鼻咽癌患者的预后;DNA异倍体患者比二倍体患者更容易出现远处转移而导致治疗失败.     

肿瘤细胞DNA干系倍体分析及其临床应用

DNA非整倍体是恶性肿瘤的特征性标志之一。测量和分析细胞核DNA含量与倍体对恶性肿瘤的病理诊断、恶性程度判定、疗效估价、预测预后具有重要价值。其主要测定方法有流式细胞术和细胞图像光度术。本文主要综述了肿瘤细胞DNA干系倍体分析及其临床应用的现状以及流式细胞术和细胞图像光度术两种检测方法的优缺点和相应的改进方法,希望对其应用于临床以提高诊断的准确性和预测肿瘤的发展起到积极的作用。     

鼻咽癌 DNA倍体、增殖指数对预后的预测意义

背景与目的：放射治疗是鼻咽癌的基本治疗方法,改变分割次数放疗及化学治疗已成为晚期鼻咽癌综合治疗方案的一部分,但是,已知的预后因素在预测鼻咽癌治疗结果时有较大的偏差,本研究采用流式细胞仪测定鼻咽癌肿瘤细胞DNA含量及增殖指数,确定其能否成为判断预后的参考指标。方法：对1994年至1995年间205例石蜡包埋存档的鼻咽癌活检组织块进行DNA倍体。细胞增殖指标包括S－期细胞分数,G2／M期细胞分数及增殖细胞期分数的测定并与相应病人的临床参数及预后进行相关分析。结果：205例检测的标本中,117例肿瘤活检组织符合DNA流式细胞仪会议指南标准,其中35例（30％）为异倍体肿瘤,82例（70％）为二倍体肿瘤,DNA倍体及各增殖指数与年龄,性别,病理类型,T／N分期及临床分期无显著性相关,异倍体肿瘤的S期细胞分数及增殖期细胞分数显著高于二倍体肿瘤,5年无瘤生存率亦存在显著差异（43％verse 632%p=0.035)。低、中、高S期细胞分数组及增殖期细胞分数组,5年无瘤生存率分别存在显著性差异（81％,verse 61% vese21%;77% verse62%verse 33%,P=0.000及P=0.048),低、中、高G2／M期细胞分数组,5年无瘤生存率无明显差异（35％verse 58% verse 54%,P=0.8617)。结论：DNA倍体,S期细胞分数及增殖细胞分数为鼻咽癌治疗的预后指标,因此,连同临床,病理参数一起,可做为筛选出预后较差的鼻咽癌个体病人的参考标准。     

DNA ploidy and p53 expression associated with tumor site and lymph node metastasis in colorectal cancer

The development of human colorectal cancer ischaracterized by a variety of genetic alterations. Geneticchanges associated with colorectal cancer have beeninvestigated intensively in recent years. p53 genemutation is one of the most common genetic alterationsin colorectal tumors and is linked to cellular proliferationand genetic instability.['l A relationship between thecarcinogenesis of colorectal cancer and p53 gene andDNA ploidy pattern also has been investigated widely.['--']p53 gene mutat…     

组织原位分析鼻咽癌细胞核DNA干系倍体及其异质性

组织原位分析鼻咽癌细胞核的DNA干系倍体及其异质性。收集42例鼻咽癌病例的归档蜡块,4μm、8μm连续组织学切片各一张。每个病例选取三个癌巢或肿瘤组织区域(共126个样本),使用TIGER细胞图像分析仪4μm组织切片测量鼻咽癌细胞核DNA的光密度等参数,8μm组织切片测量单个完整鼻咽癌细胞核的体积等参数,计算获得每个癌巢或肿瘤组织区域的以单个完整鼻咽癌细胞核的体积为单位的DNA含量(以体积积分光密度表述),以同一切片内正常鼻咽部上皮非基底细胞作为其内参照,计算其DNA干系倍体。结果显示:鼻咽癌细胞核的DNA干系倍体主要集中在1.76～3.00(DNA指数为0.88～1.50)之间,14个样本为DNA干系四倍体,仅2个样本为DNA干系亚二倍体;DNA干系倍体异质性率为66.67%。可得结论:(1)鼻咽癌DNA干系倍体主要在近二倍体与近四倍体之间;(2)鼻咽癌中存在DNA干系倍体异质性。     

Prognostic value of DNA ploidy and automated assessment of stroma fraction in prostate cancer

The combination of DNA ploidy and automatically estimated stroma fraction has been shown to correlate with recurrence and cancer death in colorectal cancer. We aimed to extend this observation and evaluate the prognostic importance of this combined marker in prostate cancer. DNA ploidy status was determined by image cytometry and the stroma fraction was estimated automatically on hematoxylin and eosin stained sections in three tumor samples from each patient to account for tumor heterogeneity. The optimal threshold for low (≤56%) and high (>56%) stroma fraction was identified in a discovery cohort (n = 253). The combined marker was validated in an independent patient cohort (n = 259) with biochemical recurrence as endpoint. The combined marker predicted biochemical recurrence independently in the validation cohort. Multivariable analysis showed that the highest risk of recurrence was observed for patients with samples that had both non-diploid ploidy status and a high stroma fraction (hazard ratio: 2.51, 95% confidence interval: 1.18–5.34). In conclusion, we suggest the combination of DNA ploidy and automatically estimated stroma fraction as a prognostic marker for the risk stratification of prostate cancer patients. It may also be a potential generic marker as concurrent results have been described in colorectal cancer.     

Study of c-erbB-2 protein and epidermal growth factor receptor expression and DNA ploidy pattern in colorectal carcinoma

Correlation of c-erbB-2 protein (n = 44), epidermal growth factor receptor (EGFR) (n = 41) expression, and DNA ploidy pattern (n = 42) with clinical outcomes of human colorectal cancers was studied. Using monoclonal antibodies against c-erbB-2 protein and EGFR, an immunohis-tochemical study of the expression of c-erbB-2 protein and EGFR in frozen tissue sections from the lesion was performed. There was no significant correlation between the expression of c-erbB-2 protein and clinicopatho-logical findings such as, tumor size, histological type, depth of invasion, lymph node metastasis, lymphatic vessel invasion, or venous invasion. However, the incidence of c-erbB-2 protein expression in Dukes D was significantly higher (9/10, 90%) than that in Dukes A to C (16/34, 47.1%). Similar tendency was also observed in the expression of EGFR. Aneuploid case was observed in 12 of observed 25 (48%) cases without lymph node metastasis, while it was observed in 16 of 17 cases (94.1%) with lymph node metastasis and there was significant association between DNA ploidy pattern and lymph node metastasis (P< 0.01) and most of the cases (17/20, 85%) were aneuploidy in Dukes C and D. The results above suggest that the expression of c-erbB-2 protein or EGFR was associated with distant metastasis, while on the other hand DNA ploidy pattern was correlated with lymph node metastasis. © 1993 Wiley-Liss, Inc.