Analysis of DNA Ploidy, Cell Cycle and Ki67 Antigen in Nasopharyngeal Carcinoma by Flow Cytometry

Summary: The expression of DNA ploidy, the cell cycle and Ki67 antigen in nasopharyngeal carcinoma (NPC) were studied and their relationship with the clinical biological behaviors and prognosis of NPC was evaluated. Biopsied specimens of NPC were made into cell suspension. By using cytometric double labeling Ki67 and DNA method, the expression of DNA ploidy, the cell cycle and Ki67 antigen were analyzed. The patients were followed-up for about 3 years and the relationship between the above-mentioned parameters and the clinical biological behavior and prognosis of NPC were evaluated. Of the 62 cases of NPC, the DNA aneuploid accounted for 29.03 %. The S phase cells accounted for 0 to 54 % in the cell cycle and the positive expression of Ki67 ranged from 0 to 52 %. There were 40 cases of LPI (64.5 %) including 15 negative cases and 22 cases of HPI (35 5 %) respectively. The DNA anueploid content was positively related to the S phase cells. The patients having a low expression of Ki67 or DNA aneuploid in tumor cells were not sensitive to chemotherapy, liable to metastasis to distant organs and had a poor prognosis, while Ki67 showed no correlation with DNA ploidy and the cell cycle. It was suggested that DNA ploidy and Ki67 could be used as an independent and objective marker to evaluate the radiosensitivity and prognosis of NPC.

A COMPARATIVE STUDY OF DNA FLOW CYTOMETRY AND MICROSPECTROPHOTOMETRY IN GASTRIC CARCINOMAS

Flow cytometric (FCM) and microspectrophotometric (MSP) measurements of cell nuclear DNA content were made in 53 fresh gastric carcinoma specimens and in 30 gastric mucosal specimens with chronic gastritis. DNA aneuploidy was found in 32/53 (60%) of gastric carcinomas, and appeared more frequently in wellor moderately differentiated tubular adenocarcinomas (90%) than in undifferentiated and mucousa cell carcinomas (23.6%) (P【0.001). No aneuploidy was found in chronic gastritis samples, but their proliferative cell fractions were higher than in normal control gastric mucosa samples (P【0.01). A comparison was made between FCM and MSP analyses of DNA content in 29 cases of gastric carcinoma, a high correlation rate (r=0.90) was found. The advantages and limitations of both methods are discussed, they may be used in combination for more precise cytochemical analysis.

Qualitative and Quantitative Studies of Polygene Proteins Expression in Esophageal Precancerous Lesions and Esophageal Carcinoma

Objective： To examine the expressions of MDM2, P53 and P27 proteins in chronic esophagitis, para-cancer mucosa and esophageal carcinoma. Methods： Immunohistochemistry was used to detect the expressions of MDM2, P53 and P27 proteins in forty-seven patients suffering from chronic esophagitis and eighty-five cases of esophageal carcinoma and corresponding para-cancer mucosa. Flow cytometry（（FCM） was applied to detect the quantities of these proteins expressed in fresh tissues of 48 cases of esophageal cancer and their para-cancer tissues and 24 cases of relative normal mucosa at the surface of cutting edge. Results： Immunohistochemistry results showed that the expressions of the three studied proteins were very similar in the epithelia of chronic esophagitis and para-cancer mucosa （P〉0.05）. Both the qualitative and quantitative studies displayed that the P53 protein had no expression and its accumulations would appear only in the early stages of esophagus canceration while the MDM2 and P27 proteins had different degrees of expressions in cases of normal esophageal mucosa. MDM2 protein markedly increased in the advanced stages of esophageal canceration. A quantitative study showed that the expression of P27 protein had a linearity of decreasing tendency （F=9.132, P=0.002） in the course of esophageal canceration. Conclusion： Chronic esophagitis may be a precancerous lesion. Owing to the changes of the P53 and P27 proteins, we can also conclude that these occur in the early stages of esophagus oncogenesis, however the changes of MDM2 expression may occur in the advanced stage of esophageal canceration.

INTRATUMORAL DNA PLOID HETEROGENEITY IN ESOPHAGEAL SQUAMOUS CELL CARCINOMA

In order to investigate the intratumoral DNA ploid heterogeneity (PH) in esophageal squamous cell carcinoma and its clinical-pathological significance, nuclear DNA ploidy of 80 cases of squamous cell carcinoma of the esophagus were determined with multiple samples removed from the same tumor, using a flow cytometry (FCM) technique. 240 samples for flow cytimetric DNA analysis were taken from 3 different parts if each tumor of 80 cases of specimens. DNA measurement was were present in a tumor or the variation in DI value among 3 peaks in each tumor was greater than 10%. Further more at last, comparison or clinical-pathological characteristics was performed between PHTs and N-PHT which has a similar ploid pattern in 3 sampling spots of each tumor (non PHT). DNA indices ranged from 0.77～1.74, and the incidence of DNA AN was 88. 8% (71/80) in this series. Of 80 cases, 38 cases (47. 5%, 38/80) showed intratumoral heterogeneity in DNA ploidy. The heterogeneity in DNA ploidy was related to the extent of wall penetration by the tumor,the incidence of lymph node metastasis and the patients's prognosis, not to histological grades and size of the tumor. There is PH phenomena in esophageal squmous cell carcinoma, and DNA PH may be a more exact indicator in reflecting the biological chatacteristics of the tumor and patient's prognosis.

胸水DNA倍体联合细胞角蛋白-19片段检测对良恶性肺部疾病的鉴别诊断意义

目的探讨胸水DNA倍体联合细胞角蛋白-19片段(CYFRA-21-1)检测对良恶性肺部疾病的鉴别诊断意义。方法回顾性分析2021年5月1日至2023年12月1日浙江省荣军医院收治确诊为非小细胞肺癌伴胸水的患者78例作为病例组,同时纳入同期入院存在胸水的肺部良性疾病患者50例作为对照组。对所有患者的胸水及外周血标本进行DNA倍体及CYFRA-21-1阳性率的检测,并统计比较外周血及胸水标本中DNA倍体、CYFRA-21-1单一检测与联合检测的效能。结果病例组胸水、外周血的DNA倍体和CYFRA-21-1阳性率均明显高于对照组,差异均有统计学意义(均P<0.05);病例组胸水DNA倍体和CYFRA-21-1的阳性率均明显高于外周血,差异均有统计学意义(均P<0.05);胸水DNA倍体联合CYFRA-21-1鉴别良恶性肺部疾病的AUC为0.980,大于单一指标胸水DNA倍体的AUC 0.964和胸水CYFRA-21-1的AUC 0.900,差异均有统计学意义(均P<0.05);胸水DNA倍体联合CYFRA-21-1检测鉴别良恶性肺部疾病的灵敏度为1.00,特异度为0.96,准确度为0.98。结论肺癌患者胸水出现DNA异常增殖及CYFRA-21-1阳性率增高且远早于外周血标本,胸水DNA倍体及胸水CYFRA-21-1联合检测有助于提高良恶性肺部疾病的鉴别诊断效能,值得予以重视应用。

Establishment of cell clones with different metastatic potential from the metastatic hepatocellular carcinoma cell line MHCC97

AIM: To establish clone cells with different metastatic potential for the study of metastasis-related mechanisms. METHODS: Cloning procedure was performed on parental hepatocellular carcinoma (HCC) cell line MHCC97, and biological characteristics of the target clones selected by in vivo screening were studied. RESULTS: Two clones with high (MHCC97-H) and low (MHCC97-L) metastatic potential were isolated from the parent cell line. Compared with MHCC97-L, MHCC97-H had smaller cell size (average cell diameter 43 microm vs 50 microm) and faster in vitro and in vivo growth rate (tumor cell doubling time was 34.2h vs 60.0h). The main ranges of chromosomes were 55-58 in MHCC97-H and 57-62 in MHCC97-L. Boyden chamber in vitro invasion assay demonstrated that the number of penetrating cells through the artificial basement membrane was (37.5 +/- 11.0) cells/field for MHCC97-H vs (17.7 +/- 6.3)/field for MHCC97-L. The proportions of cells in G0-G1 phase, S phase, and G2-M phase for MHCC97-H/MHCC97-L were 0.56/0.65, 0.28/0.25 and 0.16/0.10, respectively, as measured by flow cytometry. The serum AFP levels in nude mice 5wk after orthotopic implantation of tumor tissue were (246 +/- 66) microg.L(-1) for MHCC97-H and (91 +/- 66) microg.L(-1) for MHCC97-L. The pulmonary metastatic rate was 100% (10/10) vs 40% (4/10). CONCLUSION: Two clones of the same genetic background but with different biological behaviors were established, which could be valuable models for investigation on HCC metastasis.

鼻咽癌DNA倍体、增殖指数对预后的预测意义(英文)

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份,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例石腊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%verse62%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%verse61%verse21%;77%verse62%verse33%,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%verse58%verse54%,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。

胃癌及癌前病变DNA含量与细胞增殖活性检测的临床意义

目的:探讨胃癌及癌前病变DNA检测对胃癌早期诊断的临床意义。方法:应用流式细胞术(FCM)检测了240例胃癌、癌前病变及对照组的胃粘膜细胞DNA含量(DI)、倍体和细胞增殖活性(PI)。结果:DI在重度肠上皮化生、Ⅲ级不典型增生及胃癌显著增加;重度肠上皮化生20%出现异倍体,Ⅲ级不典型增生及胃癌中分别有52.17%与71.42%出现异倍体;慢性萎缩性胃炎、重度肠上皮化生、不典型增生及胃癌PI显著增高。81例癌前病变者随访2年,15例病理诊断证实癌变,其中11例为DNA异倍体,4例为DNA二倍体,PI值也相应较高。结论:DNA检测对胃癌早期诊断是很有价值的临床生物学指标,有助于胃癌的早期诊断。

流式细胞计测定食管癌细胞 DNA 含量和临床及病理特性的关系

本文用流式细胞检测了36例食管癌切除标本的DNA含量,结果发现,DNA倍体水平差异较大,范围为1.68C～8.16C,大多数食管癌为非整倍体DNA含量,其中少数病人在同一标本中同时发现两种不同的非整倍体。而且,食管癌细胞的DNA含量和肿瘤病变长度、局部淋巴结转移、食管外侵犯以及病理分级有密切关系;以二倍体肿瘤预后较好,且病理分化较高。而非整倍体肿瘤则相反。

食管癌前细胞DNA、端粒酶含量及多基因表达产物的定量检测

为探讨食管癌高发区人群食管上皮癌变过程中的早期分子改变及早期癌变机理 .应用流式细胞术和免疫荧光技术及碘化丙啶DNA荧光染色方法 ,对食管上皮癌前细胞的DNA含量、端粒酶含量和多个基因p5 3、p16、cyclinD1蛋白质表达进行了定量检测 .检测结果发现 ,DNA含量在癌变形成时明显增加 ,异倍体率为 87 9% ;p5 3蛋白积聚发生在癌变早期 ,在癌细胞组的阳性率为 10 0 % (5 5 ) ;抑癌基因 p16在癌变早期有明显缺失 ;癌基因cyclinD1及端粒酶阳性率在癌细胞组都为 10 0 % (分别为 6 6 ,7 7) .研究结果表明 :在癌变早期 ,DNA含量及异倍体率增加 ,癌基因cyclinD1表达增高 ,抑癌基因 p16缺失及 p5 3蛋白积聚 ,端粒酶含量也明显增高 。

瘤体内DNA倍体异质性与食管癌组织学分级的关系

采用多点取材的方法，应用流式细胞术对８０例食管癌瘤体内ＤＮＡ倍体异质性进行研究。结果显示，倍体异质性肿瘤的检出率为４７．５％（３８／８０）；倍体异质性与组织学分级无关（Ｐ＞０．０５），而与患者的预后有关（Ｐ＜０．０５）。本研究提示，深入研究肿瘤ＤＮＡ倍体异质性将对判断患者预后，协助组织学分级具有重要的意义。

食管鳞癌组织中肺耐药蛋白表达和DNA含量的定量分析

目的:探讨食管鳞癌组织中肺耐药相关蛋白(lung resistance-related protein,LRP)的表达和DNA含量检测的临床意义。方法:应用流式细胞术(flow cytometry,FCM)定量分析52例原发食管鳞癌组织和相应癌旁组织中LRP蛋白表达及DNA含量的变化状况。结果:食管鳞癌组织LRP表达的相对荧光强度(RFI)的中位数(M)为1·39,而相应癌旁组织为0·75;差异有统计学意义,P<0·05。癌组织DNA指数(DI)、SPF和PI显著高于相应癌旁组织,P<0·05。LRP蛋白表达在不同的性别、病理分级、临床分期和有无淋巴结转移间的表达均差异无统计学意义,P>0·05。DNA倍体和SPF、PI均与性别、病理分级和临床分期无明显关系,P>0·05;但与有无淋巴结转移有关,P<0·05。非整倍体肿瘤患者淋巴结转移率(63·9%,23/36)高于二倍体肿瘤患者淋巴结转移率(18·8%,3/16),P<0·05。有淋巴结转移患者SPF和PI显著高于无淋巴结转移患者,P<0·05。非整倍体肿瘤LRP蛋白表达水平(Median=1·42)略高于二倍体肿瘤(Median=1·35),但两者差异无统计学意义,P>0·05。结论:LRP蛋白表达和DNA含量在食管鳞癌发生中起重要作用。DNA含量与淋巴结转移有关,可作为预测食管鳞癌预后的指标。

食管鳞癌DNA和RNA含量测定的临床意义

目的:为探讨食管鳞癌DNA异倍体和RNA含量的变化及临床意义。方法:应用流式细胞仪对140例食管鳞癌石蜡标本进行研究。结果:食管鳞癌DNA异倍体检出率为90％(126/140),RNA含量为21.85±6.90,DNA异倍体和RNA含量与病期、恶性程度和淋巴结转移有关。结论:提示DNA和RNA含量测定有助于判断食管鳞癌患者的病期、恶性程度和淋巴结转移情况,为术后有针对性的制定治疗方案提供依据。

食管癌DNA倍体及增殖活性与病理特征的关系

目的探讨食管癌组织DNA含量(DI)、合成期细胞比例(SPF)、增殖指数(PI)与临床病理特征间的关系。方法应用流式细胞仪测定98例新鲜食管癌组织细胞的DI、SPF、PI值,研究其和食管癌临床病理的关系。结果(1)异倍体率、DI、SPF、PI等指标在以患者性别、肿瘤大体类型、分化程度、TNM分期等病理特征为分组对象的各组间没有发现差异有统计学意义;(2)食管组织淋巴结阳性组的SPF明显大于淋巴结阴性组(P=0.03)。结论(1)食管组织一旦癌变后其细胞DNA的改变和临床分期等病理指标没有明显关系;(2)食管癌淋巴结转移和食管癌组织合成期细胞比例(SPF)有关。

食管癌DNA含量和细胞周期分析的临床意义

目的 :研究食管癌和癌旁组织DNA含量、细胞周期分布与临床分期及组织病理学分级的关系和意义。方法 :应用流式细胞术 (FCM)测定 4 0例食管癌、癌旁组织、1 0例正常食管粘膜细胞的DNA指数 (DI)、S期细胞百分率 (SPF)、异倍体率。结果 :癌灶中心组织DI、SPF、异倍体率明显高于癌旁组织 ;DI、SPF、异倍体率与年龄、性别无关 ,而随临床分期、组织病理学分级增加而逐渐增加 ;有淋巴结转移的癌组织DI、SPF、异倍体率明显高于无淋巴结转移者。结论 :DNA含量、异倍体率与癌中心灶密切相关 ,SPF可作为组织病理学分级、临床分期的一个参数。DI、SPF、异倍体率与患者的临床信息结合 ,将有助于食管癌患者的诊断。

食管鳞癌中hTERT表达、DNA含量与细胞增殖周期的关系

目的探讨食管鳞癌中hTERT表达与细胞DNA含量、细胞增殖周期的关系。方法采用免疫组化检测hTERT表达,流式细胞术检测hTERT和DNA含量及增殖指数。结果(1)癌组hTERT阳性率和荧光指数(FI)显著高于癌旁组(P<0.01);(2)癌组DNA指数(DI)和异倍体率均高于癌旁组(P<0.05),癌组增殖指数(PI)显著高于癌旁组(P<0.01)。结论hTERT过量表达、DNA含量增加和增殖指数升高与食管鳞癌的发生、发展密切相关。

周围型肺癌的CT征象与癌细胞DNA含量的关系

目的 :探讨周围型肺癌DNA含量及其与CT征象的关系。材料和方法 :用流式细胞分析术检测 5 0例有术前CT检查并经手术病理证实的周围型肺癌石蜡包埋标本的细胞核DNA含量。结果 :CT显示分叶征、毛刺征、棘突征 ,有纵隔、肺门淋巴结转移的有无 ,其DNA异倍体检出率均有统计学差异。结论 :周围型肺癌出现分叶征、毛刺征、棘突征且有纵隔、肺门淋巴结转移的CT征象与癌细胞核的DNA含量有显著关系。

胃癌和癌旁组织DNA含量、细胞增殖活性的定量分析与临床意义

目的探讨胃癌及癌旁组织DNA含量、异倍体、细胞增殖活性与胃癌手术范围及胃癌临床病理因素的关系。方法采用流式细胞术(FCM)对40例胃癌及相应癌旁组织中细胞DNA含量(DI值)、细胞增殖活性(PI值)进行定量检测,并根据DNA含量分析异倍体。结果胃癌组织中DI值为1.35±0.10,PI值为18.25±7.32,异倍体为82.5%,癌旁组织3cm、5cm、7cm、最远端断端,其DI值分别为1.20±0.08、1.05±0.05、1.01±0.03、0.96±0.01,其PI值分别为16.03±6.27、9.98±3.20、9.54±2.23、6.27±2.17,其异倍体率为70.0%、22.5%、17.5%、15.0%。异倍体率、细胞增殖活性与胃癌组织分化程度、浸润深度、淋巴结转移、手术断端癌残留、TNM分期密切相关。结论胃癌、癌旁组织3㎝组DNA含量、细胞增殖活性及异倍体率,高于癌旁组织5cm、7cm、最远端断端组,可作为胃癌手术切除范围的理论依据。异倍体率、细胞增殖活性可作为评价胃癌恶性程度、判断预后的重要指标。

肝细胞癌RNA、DNA定量测定及临床意义

应用流式细胞分析技术，对64例肝细胞癌（HCC）RNA、DNA含量进行定量分析，并对病人进行1～5年半的随访，结果表明：RNA指标和DNA指标对PHC的临床病理诊断符合率分别为93．7％和75．0％，RNA指标更敏感，双指标联合应用与临床病理诊断符合率为96.8％；DNA倍体与年龄、HCV感染密切相关（P＜0．05），与性别、AFP水平、HBsAg及瘤体大小无关；DI随组织学分级的增高而增加，RNA和DNA含量与预后密切相关。提示RNA、DNA含量可作为PHC诊断及判断预后的理想指标，RNA和DNA双指标同时应用优于任一单项指标。

结肠直肠癌细胞DNA及CD44含量与肿瘤形成和转移的关系

目的 :探讨结肠直肠癌细胞DNA含量及CD4 4的定量检测与癌形成和转移的关系。方法 :采用流式细胞仪 (flowcytometry ,FCM)对手术切除的结肠直肠癌组织及癌旁周围正常组织细胞进行DNA含量及倍体测定、CD4 4定量检测。结果 :DNA含量在结直肠癌组明显增高 ,二倍体细胞明显减少 ,而异倍体明显增多 ,在结直肠癌组异倍体率为 6 6 6 % ;同时CD4 4在结直肠癌组明显高于癌旁周围正常组织组 (P <0 0 1) ,CD4 4在有淋巴结转移组和无淋巴结转移组中有极显著差异 (P <0 0 1)。结论 :癌细胞DNA含量及异倍体率增加 ,CD4 4的表达与结肠直肠癌形成、转移有关。