Fluorescence in situ hybridization for detecting urothelial carcinoma

BACKGROUND:

Because urothelial carcinoma (UC) is associated with a significantly high risk of disease recurrence and progression, patients with UC require long‐term surveillance. Fluorescence in situ hybridization (FISH) has been shown to be more sensitive than cytology in the detection of UC. The current study evaluated the use of FISH for detecting UC.

METHODS:

A pathology database was used to identify patients who had urine cytology and FISH performed at the study institution between 2004 and 2006. Urinary specimens were analyzed using UroVysion FISH probes for abnormalities in centromeric chromosomes 3, 7, and 17 and locus‐specific 9p21. FISH results were correlated with cytologic findings and a minimal clinical follow‐up of 24 months.

RESULTS:

A total of 1006 consecutive urinary specimens from 600 patients (448 men and 152 women) who were monitored for recurrent UC (915 specimens) or evaluated for urinary symptoms (91 specimens) were identified. On FISH analysis, 669 specimens were found to be negative for UC and 272 specimens were positive for UC. Sixty‐five (6%) specimens were insufficient for FISH analysis. The sensitivity and specificity of FISH for UC were 58% and 66%, respectively, and 59% and 63%, respectively, when FISH and cytology results were combined. Factors contributing to decreased FISH sensitivity included the paucity or absence of tumor cells, low‐grade tumors, degenerated cells, method of specimen collection, type of specimen, and obscuring inflammatory cells or lubricant.

CONCLUSIONS:

UroVysion FISH appeared to have good sensitivity and specificity for detecting UC in urinary specimens. It is important to correlate the FISH results with the cytologic findings. Cancer (Cancer Cytopathol) 2010. © 2010 American Cancer Society.     

荧光原位杂交在多发性骨髓瘤中的应用

多发性骨髓瘤(MM)是一种常见的浆细胞恶性肿瘤,重要的染色体异常导致MM进展,在MM中具有独立的预后判断价值.荧光原位杂交(FISH)检测技术不需要中期分裂相,可分析大量间期细胞,具有操作相对简单、重复性好、敏感性和特异性高的优点,在中期分裂相不易获得的MM中具有更突出的优势,可检测出与预后密切相关的累及基因位点的微缺失,异常克隆检出率显著提高,为MM的诊断、分型和预后提供理论依据,指导临床个体化治疗.     

Fluorescence in situ hybridization studies on direct smears

BACKGROUND:

In the fine‐needle aspiration biopsy (FNAB) diagnosis of B‐cell non‐Hodgkin lymphomas (B‐NHL), the role of flow cytometry (FC) can be limited because of nondiagnostic findings. Fluorescence in situ hybridization (FISH) studies, similar to FC, can be helpful in establishing clonality and in subclassifying the lymphoma. The aim of the current study was to determine whether FISH studies performed on unstained direct smears improved the ability to diagnose and/or subclassify B‐NHL on FNAB.

METHODS:

A total of 181 cases of B‐NHL diagnosed by FNAB were retrieved. The cytomorphology, ancillary study results, clinical information, and available pathologic follow‐up were reviewed.

RESULTS:

Of the 181 cases, FISH studies were performed in 106 cases (59%). The indications for FISH studies were for subclassification (59 cases; 56%) and nondiagnostic or unavailable FC results (47 cases; 44%). Of the 59 cases submitted for subclassification, 23 cases (39%) were successfully subclassified. The 47 cases with nondiagnostic or unavailable FC results included cases in which FC demonstrated a surface immunoglobulin‐negative population (19 cases; 40%), had insufficient cellularity (18 cases; 38%), yielded negative results (6 cases; 13%), or had no specimen submitted (4 cases; 9%). In this group, 26 cases (55%) demonstrated an immunoglobulin heavy‐chain gene rearrangement and/or chromosomal translocation.

CONCLUSIONS:

The results of the current study illustrate that FISH studies performed on unstained direct smears play a complementary role to FC in establishing the diagnosis and/or subclassification of B‐NHL. Thus, the preparation of unstained smears at the time of FNAB can be helpful for potential FISH studies. Cancer (Cancer Cytopathol) 2009. © 2009 American Cancer Society.     

Fluorescence in situ hybridization to evaluate dysplasia in Barrett's esophagus: a pilot study

BACKGROUND: Histological disagreement is frequent in the diagnosis and grading of dysplasia in Barrett's esophagus (BE). AIMS: To identify selective markers for dysplasia in BE and to improve the differentiation between low-grade dysplasia (LGD) and high-grade dysplasia (HGD). METHODS: Eight BE esophageal mucosectomies (7 males) were analyzed by conventional histology and immunohistochemistry for p53 and Fluorescence In situ Hybridization (FISH) for chromosomes X, Y, 4, 8, 17, 18. The female mucosectomy was considered as a control for the XY probe. RESULTS: p53 confirmed multifocal dysplasia in all cases. All patients displayed increased aneusomy for chromosomes 4, 8, 17 and 18 along the sequence of cancer progression. There was also a trend for chromosome 8 to be below the FISH cutoff; 50% of cases showed aneusomy for chromosome 18 in areas with differing grades of dysplasia. Aneusomy was increased for chromosomes 4 and 17, to a similar extent in LGD and HGD. In male specimens, the presence of chromosome Y was revealed in Barrett's mucosa and LGD, but not in HGD and intramucosal carcinoma. CONCLUSIONS: FISH in BE may be useful diagnostic to confirm the diagnosis of HGD. Loss of chromosome Y might be a selective marker of HGD in male patients.     

Fluorescence in situ hybridization evaluation of minimal residual disease on stem-cell harvests

The usefulness of fluorescence in situ hybridization (FISH) analysis to detect minimal residual disease (MRD) in autologous bone marrow and peripheral blood stem-cell harvests has been tested in three patients with hematologic malignancies. Conventional cytogenetics and FISH were used to characterize the leukemic clones identifying the specific chromosomal abnormalities (monosomy 7 in a myelodysplastic patient and trisomy 8 in two acute myeloid leukemic patients). Such analysis was useful to monitor the MRD persistent after treating these patients with intensive chemotherapy. The myelodysplastic patient underwent eight peripheral blood-stem cell harvests in which FISH detected the persistence of monosomy 7 cells, precluding their use for autologous transplantation. This patient relapsed and died. In two acute myeloid leukemia patients who underwent an autologous marrow harvest, FISH did not show a significant proportion of trisomy 8 cells. Nevertheless, autologous transplantation was not performed, owing to an insufficient CD34 cell content in the harvests. One of these patients relapsed with the reappearance of trisomy 8 and died. The other patient, on the contrary, is alive in complete remission 3 years after the bone marrow harvest. The usefulness and applicability of MRD quantification in stem-cell harvests is discussed on the basis of the sensitivity of the methodology applied.     

淋巴组织增生性疾病组织中EB病毒的原位杂交检测

目的探讨ＥＢ病毒与我国各类淋巴组织增生性疾病的关系。方法以ＥＢ病毒ＬＭＰ基因为探针,对２１４例淋巴组织增生性疾病组织中ＥＢ病毒进行原位杂交检测,并用ＳＹＳＴＡＴ软件对实验结果进行分析。结果霍奇金淋巴瘤（ＨＤ）、非霍奇金淋巴瘤（ＮＨＬ）、淋巴组织良性增生（ＢＬＰ）组织中ＥＢ病毒阳性率分别为３０．０％（１５／５０）,１４．０％（１８／１２９）及２．９％（１／３５）。在ＮＨＬ中,高度恶性（ＨＮＨＬ）、中度恶性（ＭＮＨＬ）及低度恶性（ＬＮＨＬ）ＥＢ病毒阳性率分别为２８．１％（９／３２）、１０．５％（９／８４）及０％（０／９）。ＨＤ与ＨＮＨＬ间ＥＢ病毒阳性率均显著高于ＢＬＰ（Ｐ＜０．０１,Ｐ＜０．０５）,ＭＮＨＬ和ＬＮＨＬ与ＢＬＰ间ＥＢ病毒阳性率差异无显著性（Ｐ＞０．０５）。结论ＥＢ病毒与ＨＤ及ＨＮＨＬ的发生有关,而与ＭＮＨＬ及ＬＮＨＬ的发生关系不大。     

荧光原位杂交和尿细胞学检查对膀胱癌诊断意义的Meta分析

背景与目的：目前膀胱癌疗效和监测的主要方法是膀胱镜和尿细胞学检查,前者为侵人性检查,令患者感到不适;后者虽无创且特异性高．但敏感性太低,且受主观因素影响大。本研究拟对中、英文有关比较荧光原位杂交（fluorescence in situ hybridization,FISH）和尿细胞学检查诊断膀胱癌研究的结果进行系统分析,以明确FISH对膀胱癌的诊断意义。方法：采用Cochrane系统评价方法,MEDLINE（1966年1月～2008年6月）、EMBASE（1988年1月。2008年6月）、Cochrane图书馆、中国生物医学期刊文献数据库（CMCC,1979年。2008年6月）、CNKI数字图书馆（1979年1月～2008年6月）进行有关FISH和尿细胞学检查诊断膀胱癌文献的检索、质量评价和资料提取,采用MetaDiScl．4软件进行Meta分析。结果：共检索到相关研究242篇,排除230篇,符合纳入标准12篇进入Meta分析,涉及研究对象3430例。异质性检验提示无阈值效应,但存在其它原因导致的异质性。按随机效应模型进行Meta分析．FISH和尿细胞学诊断膀胱癌的准确度指标敏感度、特异度、阳性似然比、阴性似然比以及诊断优势比等汇总及95％C1分别为74％（71％-77％）VS．57％（54％-61％）、88％（86％-90％）VS．85％ （83％-87％）、6．18（3．56～10．73）VS．4．15（2．78～6．20）、0．29（0．19～0．45）VS．0．51（0．41～0．63）及24．17（9．33～62．64）VS．9．59（5．91～15．57）。FISH和尿脱落细胞学检查的敏感度随肿瘤分级、分期的升高而增高。综合受试者工作特征曲线下面积分别为0．8938、0．8247．Q^＊值分别为0．7847、0．7226。结论：FISH诊断膀胱癌的准确度较高,但对高分期的敏感度较细胞学低,目前尚不能取代传统的尿细胞学检查,但可作为膀胱癌术前诊断、术后监测和随访的指标。     

尿脱落细胞FISH检测在膀胱癌诊断中的应用

目的:研究尿脱落细胞FISH检测提高膀胱癌早期诊断的可行;方法:收集68例腺膀胱炎患者晨尿通过FISH检测GLP p16、CSP3、CSP17和CSP7染色体异常信号,并用统计学方法计算出阈值.为了验证FISH检测的优越,实验选取临床诊断膀胱癌最常用的尿脱落细胞学检测作为研究对照方法,收集100例疑似膀胱癌患者尿液标本,分别对尿脱落细胞和FISH检查诊断膀胱癌的敏感、特异、FISH检测与膀胱癌临床及病理特征的关系进行统计学分析.结果:100例疑似膀胱癌患者尿脱落细胞及FISH检测的阳率分别为56.0％ (56/100)和77.0％(77/100),敏感度分别为56.10％(46/82)和82.02％(73/89),经统计学分析两种检查方法之间的差异具有统计学意义.FISH检测的敏感和总阳率明显高于尿脱落细胞,而特异两者差异无统计学意义.FISH检测与膀胱癌的病理分级和临床分期均无相关.结论:FISH检测技术是能够成为提高膀胱癌早期诊断的一种新技术.     

应用荧光原位杂交技术进行胚胎种植前遗传学诊断的临床分析

目的：探讨应用荧光原位杂交（fluorescence in situ hybridisation,FISH）技术对染色体异常携带者进行种植前胚胎遗传学诊断（preimplantation genetic diagnosis,PGD）的临床意义。方法：根据携带者染色体异常种类,分别选择相应的亚端粒探针和着丝粒探针或性染色体探针,进行1次或者2次杂交,对7例染色体异常携带者进行了胚胎种植前遗传学诊断。结果：7例染色体异常携带者进行了7个周期的PGD,获卵131枚,活检77枚胚胎,检出卵裂球87枚,移植20枚胚胎,4例临床妊娠,其中2例已分娩健康婴儿。结论：应用荧光原位杂交技术对染色体异常携带者的胚胎进行种植前遗传学诊断是一种有效方法。     

荧光原位杂交检测宫颈脱落细胞端粒酶基因扩增临床价值的探讨

目的:检测宫颈脱落细胞中端粒酶基因扩增的检出率,探讨其在宫颈病变中诊断的价值.方法:收集2007-07-27-2009-01-09哈尔滨医科大学附属第三临床医学院妇科113例宫颈脱落细胞标本,其中非典型鳞状上皮细胞(ASC)患者27例、低度鳞状上皮内病变(LSIL)患者33例、高度鳞状上皮内病变(HSIL)患者26例、...     

Fluorescence in situ hybridization analysis of chromosome aberrations in 60 Chinese patients with multiple myeloma

Conventional cytogenetic analysis is often hampered owing to the low mitotic index of multiple myeloma (MM) cells in bone marrow samples of MM. Interphase fluorescence in situ hybridization (I-FISH) analysis combined with magnetic-activated cell sorting (MACS) has substantially enhanced the sensitivity of cytogenetic analysis. Here, we used I-FISH to explore the incidence of chromosomal abnormalities in 60 Chinese patients with newly diagnosed MM. Five different specific probes for the regions containing 13q14.3 (D13S319), 14q32 (IGHC/IGHV), 1q21, 1p12, and 17p13 were used to detect chromosomal aberrations, and LSI IGH/CCND1, LSI IGH/FGFR3, and LSI IGH/MAF probes were further applied to detect t(11;14)(q13;q32), t(4;14)(p16;q32), and t(14;16)(q32;q23) in patients with 14q32 rearrangement. Fifty of the patients (83.3%) had at least one type of abnormalities regarding the regions analyzed. Nine patients (15%) had one abnormality; 10 patients (16.7%) had two abnormalities; 31 patients (51.7%) had three or more abnormalities. The most frequent abnormality in the patients was illegitimate IgH rearrangement (70%), followed by 13q14 deletion (63.3%), 1q21 amplification (61.7%), 1p12 deletion (33.3%), and 17p13 deletion (13.3%). These aberrations are not randomly distributed, but strongly interconnected. Patients with 17p13 deletion or t(4;14)(p16;q32) had significant higher ??2-microglobulin level (P?<?0.05). However, all these abnormalities had no correlation with age, gender, disease stage, and Ig isotype; yet, it was showed that the frequencies of the individual chromosomal abnormalities were very high. Taken together, MACS in combination with I-FISH may be a promising tool to detect the molecular cytogenetic abnormalities of MM.     

Expression of albumin messenger RNA detected by in situ hybridization in preneoplastic and neoplastic lesions in rat liver

The process of chemical hepatocarcinogenesis is characterized by the appearance of preneoplastic lesions showing changes in the expression of various marker enzymes. We have analyzed the phenotype of small preneoplastic foci and expansively growing nodules in liver sections obtained from rats treated with various carcinogens. Changes within the lesions in canalicular adenosine triphosphatase, gamma-glutamyl transpeptidase, NADPH-(cytochrome P-450) reductase, cytochrome P-450 PB2, epoxide hydrolase, and glycogen content were detected by means of enzyme histochemical and immunohistochemical staining procedures. In parallel sections the expression of albumin messenger RNA was investigated by in situ hybridization using a 35S-labeled albumin specific complementary DNA probe. In general, small preneoplastic lesions showed unchanged levels of albumin messenger RNA. In contrast, the expression of albumin messenger RNA was found to be reduced to varying degrees in large hepatic nodules. An expression of alpha-fetoprotein messenger RNA could not be detected in any of the nodules. No direct correlation between the enzyme phenotype of the lesions and the degree in reduction of albumin messenger RNA could be established except that the reduction was most pronounced in nodules which had lost their ability to store glycogen. Since the synthesis and excretion of albumin is a typical function of the differentiated hepatocyte in the adult animal, the observed decrease in albumin messenger RNA expression in large hepatic nodules is in accordance with the hypothesis of a gradual dedifferentiation or retrodifferentiation of the cell population during carcinogenesis. Hyperplastic nodules produced by continuous treatment of rats with 4-dimethylaminoazobenzene showed increased rather than decreased albumin levels. The analysis of albumin messenger RNA expression might therefore be used as a tool to discriminate between nodules of differing biological nature and fate.     

Targeted cytogenetic analysis of gastric tumors by in situ hybridization with a set of chromosome-specific DNA probes

Fluorescent in situ hybridization (FISH) with biotinated chromosome-specific repetitive DNA probes was used for the cytogenetic study of ten gastric adenocarcinomas. All tumors (eight male, two female patients) were histologically moderately or poorly differentiated and nine of ten had metastasized to regional lymph nodes. The authors applied a set of satellite DNA probes, specific for chromosomes 1, 7, 17, X, and Y in order to detect numerical chromosome aberrations in freshly isolated tumor cell nuclei. Normal diploid human lymphocyte nuclei and, in a number of cases, normal gastric mucosa served as controls. Parallel with the hybridization experiments DNA flow cytometric study of acridine orange (AO)-stained tumor cells was carried out. By means of FISH the authors found seven cases to be aneuploid, the other three cases appeared diploid. This was confirmed by DNA flow cytometric analysis with AO. Furthermore, loss of the Y chromosome in a high percentage of cells was seen by FISH in six of eight tumors from male patients. In the other two male samples a possible loss was observed in a small proportion of cells (15%). In three patients from whom the authors had normal gastric mucosa the Y loss was restricted to the tumor cells. These data indicate that in situ hybridization with chromosome-specific repetitive DNA probes can serve as a cytogenetic tool for the analysis of interphase nuclei of solid human tumors, at least with respect to the detection of numerical chromosome abnormalities.     

多色荧光原位杂交技术检测结直肠癌早期染色体改变的研究

目的 分析我国结直肠癌(CRC)及结直肠腺瘤染色体畸变情况,探讨通过多色荧光原位杂交(M-FISH)技术检测染色体变化,用于CRC早期诊断及评估腺瘤恶性变倾向的可行性和有效性.方法 选择7、8、12和20号染色体探针应用M-FISH技术检测161例患者的183份结直肠病变组织,包括65份腺瘤组织、19份癌旁腺瘤组织和99份腺癌组织.分析结直肠腺癌和腺瘤与临床病理参数之间的关系.结果 7、8、12和20号染色体在腺癌组织中有较高的增益畸变率,分别为82.1%(55/67)、70.1%(47/67)、60.9%(56/92)和72.8%(67/92);在癌旁腺瘤组织亦有较高的增益畸变率,分别为76.5%(13/17)、41.2%(7/17)、50%(9/18)和72.2%(13/18);7号和20号染色体增益畸变同时出现在同一个患者的腺癌与腺瘤标本中的频率较高.结论 7、8、12和20号染色体在腺癌及腺瘤组织中均存在较高的染色体数目畸变率;其中7号与20号染色体增益畸变有可能成为CRC早期预警标志.M-FISH技术检测可能有助于CRC的早期诊断.