Transcriptome analyses of benign and malignant prostate epithelial cells in formalin-fixed paraffin-embedded whole-mounted radical prostatectomy specimens

Furusato, Bungo; Shaheduzzaman, Syed; Petrovics, György; Dobi, Albert; Seifert, Martin; Ravindranath, Lakshmi; Nau, Marion M.; Werner, Thomas; Vahey, Maryanne; McLeod, D. G.; Srivastava, Shiv; Sesterhenn, Isabell A.

doi:10.1038/sj.pcan.4501007

Cited by 29 publications

(19 citation statements)

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“…KLK2 and PSA are androgen regulated serine proteases expressed in prostate epithelial cells and upregulated in prostate cancer [63]. Two ets related transcription factors observed in this study, ets-related gene ( ERG ) and Sam pointed domain ets transcription factor (SPEDF ) [16] are known to be upregulated in prostate tumor epithelial cells [64,17,18]. The importance of the ERG gene is supported by its frequent involvement in complex rearrangements with a host of other gene fusion partners.…”

Section: Discussionmentioning

confidence: 98%

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Analysis of gene expression in prostate cancer epithelial and interstitial stromal cells using laser capture microdissection

et al. 2010

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BackgroundThe prostate gland represents a multifaceted system in which prostate epithelia and stroma have distinct physiological roles. To understand the interaction between stroma and glandular epithelia, it is essential to delineate the gene expression profiles of these two tissue types in prostate cancer. Most studies have compared tumor and normal samples by performing global expression analysis using a mixture of cell populations. This report presents the first study of prostate tumor tissue that examines patterns of differential expression between specific cell types using laser capture microdissection (LCM).MethodsLCM was used to isolate distinct cell-type populations and identify their gene expression differences using oligonucleotide microarrays. Ten differentially expressed genes were then analyzed in paired tumor and non-neoplastic prostate tissues by quantitative real-time PCR. Expression patterns of the transcription factors, WT1 and EGR1, were further compared in established prostate cell lines. WT1 protein expression was also examined in prostate tissue microarrays using immunohistochemistry.ResultsThe two-step method of laser capture and microarray analysis identified nearly 500 genes whose expression levels were significantly different in prostate epithelial versus stromal tissues. Several genes expressed in epithelial cells (WT1, GATA2, and FGFR-3) were more highly expressed in neoplastic than in non-neoplastic tissues; conversely several genes expressed in stromal cells (CCL5, CXCL13, IGF-1, FGF-2, and IGFBP3) were more highly expressed in non-neoplastic than in neoplastic tissues. Notably, EGR1 was also differentially expressed between epithelial and stromal tissues. Expression of WT1 and EGR1 in cell lines was consistent with these patterns of differential expression. Importantly, WT1 protein expression was demonstrated in tumor tissues and was absent in normal and benign tissues.ConclusionsThe prostate represents a complex mix of cell types and there is a need to analyze distinct cell populations to better understand their potential interactions. In the present study, LCM and microarray analysis were used to identify novel gene expression patterns in prostate cell populations, including identification of WT1 expression in epithelial cells. The relevance of WT1 expression in prostate cancer was confirmed by analysis of tumor tissue and cell lines, suggesting a potential role for WT1 in prostate tumorigenesis.

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Section: Discussionmentioning

confidence: 98%

“…normal epithelia, we and others observed genes elevated in prostate cancer epithelial cells including kallikrein proteins 2 (KLK2) , and 3 (KLK3 , or PSA) [16]. KLK2 and PSA are androgen regulated serine proteases expressed in prostate epithelial cells and upregulated in prostate cancer [63].…”

Section: Discussionmentioning

confidence: 99%

Analysis of gene expression in prostate cancer epithelial and interstitial stromal cells using laser capture microdissection

et al. 2010

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“…LCM is a well-established technique in prostate tissue [7,15,16]. However, to our knowledge, it has not been described in FFPE diagnostic needle biopsies.…”

Section: Discussionmentioning

confidence: 99%

“…Laser-capture microdissection (LCM) allows enriched cell populations to be accurately isolated from heterogeneous tissue, hence facilitating analysis of different components from a single tissue sample [6,7]. Real-time PCR (QPCR) uses small amplicon sizes which are more suited to the short templates derived from degraded RNA as compared with standard PCR.…”

Section: Introductionmentioning

confidence: 99%

Application of transcript profiling in formalin‐fixed paraffin‐embedded diagnostic prostate cancer needle biopsies

Rogerson¹,

Darby²,

Jabbar³

et al. 2008

BJU International

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OBJECTIVETo investigate the feasibility of transcript profiling in diagnostic formalin‐fixed and paraffin‐embedded (FFPE) biopsies for prostate cancer.MATERIALS AND METHODSLaser‐capture microdissection (LCM) was used to microdissect glandular epithelium as well as stromal tissue in archival prostate needle biopsies. Optimized RNA extraction, reverse transcription and real‐time PCR (QPCR) protocols were used to detect transcript expression. RNA degradation effects were assessed using hydrolysed cell line RNA and matched xenograft FFPE and frozen tumours.RESULTSLCM and RNA extraction was achieved in all biopsies from a pilot cohort of five patients. cDNA produced was successfully used to detect expression of glyceraldehyde‐3‐phosphate dehydrogenase, RPL13, prostate‐specific antigen, vimentin, inhibitor of differentiation/DNA binding 1 (Id‐1) and polycomb group protein enhancer of zeste homolog 2 (EZH2) transcripts. In the cell line and xenograft models, we investigated the effect of RNA degradation on transcript quantification by QPCR. In both models normalization of transcript quantity with a housekeeping gene resulted in restored expression in all degraded samples to within a 50% difference of control samples. Using an extended cohort of 29 biopsies, we tested application in detecting differences in EZH2 and Id‐1 expression between malignant and benign epithelium. The results confirmed that our technique was capable of quantifying significant differences in expression between malignant and benign epithelium consistent with the reported trends.CONCLUSIONThis study reports the use of standard FFPE needle biopsies for transcript profiling and supports the concept of molecular prognostic studies in tissue acquired at diagnosis in prostate cancer.

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“…Recent studies have shown that similar techniques have been successfully performed on breast, gastric, colonic, and prostatic epithelium suggesting that this methodology is a powerful means for screening of new biomarkers (23,52-54). As pathologists our primary goal was to identify candidates for the generation of immunohistochemical reagents as immunohistochemistry is available in all pathology departments and all pathologists are quite comfortable with the technique.…”

Section: Discussionmentioning

confidence: 99%

Expression Analysis of Barrett's Esophagus–Associated High-Grade Dysplasia in Laser Capture Microdissected Archival Tissue

Sabo¹,

Meitner²,

Tavares³

et al. 2008

Clinical Cancer Research

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Purpose: Identifying genes differentially expressed in nondysplastic BE (NDBE) from those expressed in high-grade dysplasia (HGD) should be of value in improving our understanding of this transition and may yield new diagnostic and/or prognostic markers.The aim of this study was to determine the differential transcriptome of HGD compared with NDBE through gene microarray analysis of epithelial cells microdissected from archival tissue specimens. Experimental Design: Laser capture microdissection was used to isolate epithelial cells from adjacent inflammatory and stromal cells. Epithelial mRNA was extracted from areas of NDBE and HGD in matched biopsies from 11 patients. mRNA was reverse transcribed and applied on Affymetrix cDNA microarray chips customized for formalin-exposed tissue. For a subset of these genes, differential gene expression was confirmed by real-time PCR and immunohistochemistry. Results: There were 131 genes overexpressed by at least 2.5-fold in HGD versus NDBE and 16 genes that were underexpressed by at least 2.5-fold. Among the overexpressed genes are several previously shown to be increased in the neoplastic progression of BE, as well as novel genes such as lipocalin-2, S100A9, matrix metallopeptidase 12, secernin 1, and topoisomerase IIa. Genes decreased in dysplastic epithelium include MUC5AC, trefoil factor 1 (TFF1), meprin A, and CD13. Real-time PCR validated the changes in expression in 24 of 28 selected genes. Immunohistochemistry confirmed increased protein expression for topoisomerase IIa, S100A9, and lipocalin-2 and decreased expression of TFF1 across the spectrum of BE-associated dysplasia from NDBE through adenocarcinoma. Conclusions: This is the first study to identify epithelial genes differentially expressed in HGD versus NDBE in matched patient samples.The genes identified include several previously implicated in the pathogenesis of BE-associated dysplasia and new candidates for further investigation.Barrett's esophagus (BE) is a premalignant condition in which the normal squamous epithelium of the distal esophagus undergoes metaplasia to a specialized intestinal epithelium. The presence of BE increases the risk of developing esophageal adenocarcinoma (EAC); approximately 1% of patients diagnosed with BE will progress from metaplasia to dysplasia and finally to BE-related EAC (1, 2). EACs are thought to develop from BE by way of a stepwise transition through low-grade dysplasia (LGD) to high-grade dysplasia (HGD). Approximately half of all patients with HGD will progress to EAC (3). Several genetic abnormalities have been implicated in the transition from BE through HGD to EAC, including microsatellite instability (4), promoter hypermethylation (5 -8), and altered expression of a large set of genes (reviewed in 9). The identification of HGD by light microscopy is still considered the gold standard by which patient follow-up and treatment is coordinated. However, the morphologic distinction between LGD and HGD can be challenging (10, 11). It would be beneficial to ide...

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Transcriptome analyses of benign and malignant prostate epithelial cells in formalin-fixed paraffin-embedded whole-mounted radical prostatectomy specimens

Cited by 29 publications

References 13 publications

Analysis of gene expression in prostate cancer epithelial and interstitial stromal cells using laser capture microdissection

Analysis of gene expression in prostate cancer epithelial and interstitial stromal cells using laser capture microdissection

Application of transcript profiling in formalin‐fixed paraffin‐embedded diagnostic prostate cancer needle biopsies

Expression Analysis of Barrett's Esophagus–Associated High-Grade Dysplasia in Laser Capture Microdissected Archival Tissue

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