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Isolation and genome-wide expression and methylation characterization of CD31+ cells from normal and malignant human prostate tissue.

Luo W, Hu Q, Wang D, Deeb KK, Ma Y, Morrison CD, Liu S, Johnson CS, Trump DL - Oncotarget (2013)

Bottom Line: The data, although preliminary, indicates that there exist widespread differences in methylation and transcription between TdECs and NdECs.Interestingly, only a small proportion of perturbed gene was overlapped between American (AA) and Caucasian American (CA) patients with prostate cancer.Future studies will be required to further characterize the isolated ECs and determine the biological features that can be exploited in the prognosis and therapy of prostate cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Therapeutics, Roswell Park Cancer Institute, Buffalo, New York.

ABSTRACT
Endothelial cells (ECs) are an important component involved in the angiogenesis. Little is known about the global gene expression and epigenetic regulation in tumor endothelial cells. The identification of gene expression and epigenetic difference between human prostate tumor-derived endothelial cells (TdECs) and those in normal tissues may uncover unique biological features of TdEC and facilitate the discovery of new anti-angiogenic targets. We established a method for isolation of CD31+ endothelial cells from malignant and normal prostate tissue obtained at prostatectomy. TdECs and normal-derived ECs (NdECs) showed >90% enrichment in primary culture and demonstrated microvascular endothelial cell characteristics such as cobblestone morphology in monolayer culture, diI-acetyl-LDL uptake and capillary-tube like formation in Matrigel®. In vitro primary cultures of ECs maintained expression of endothelial markers such as CD31, von Willebrand factor, intercellular adhesion molecule, vascular endothelial growth factor receptor 1, and vascular endothelial growth factor receptor 2. We then conducted a pilot study of transcriptome and methylome analysis of TdECs and matched NdECs from patients with prostate cancer. We observed a wide spectrum of differences in gene expression and methylation patterns in endothelial cells, between malignant and normal prostate tissues. Array-based expression and methylation data were validated by qRT-PCR and bisulfite DNA pyrosequencing. Further analysis of transcriptome and methylome data revealed a number of differentially expressed genes with loci whose methylation change is accompanied by an inverse change in gene expression. Our study demonstrates the feasibility of isolation of ECs from histologically normal prostate and prostate cancer via CD31+ selection. The data, although preliminary, indicates that there exist widespread differences in methylation and transcription between TdECs and NdECs. Interestingly, only a small proportion of perturbed gene was overlapped between American (AA) and Caucasian American (CA) patients with prostate cancer. Our study indicates that identifying gene expression and/or epigenetic differences between TdECs and NdECs may provide us with new anti-angiogenic targets. Future studies will be required to further characterize the isolated ECs and determine the biological features that can be exploited in the prognosis and therapy of prostate cancer.

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The association of differentially expressed genes with differently methylated lociAmong the differentially expressed genes with differential methylation events, a portion of them have loci whose direction of methylation expression change is opposite to that of gene expression change. A) The comparison between tumor and normal in AA group. B) The comparison between tumor and normal in CA group. “up_down” indicates the genes with increased expression and hypomethylation events in tumor versus normal comparision, while “down_up” means the genes with decreased expression and hypermethylation events in tumor versus normal comparision.
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Figure 7: The association of differentially expressed genes with differently methylated lociAmong the differentially expressed genes with differential methylation events, a portion of them have loci whose direction of methylation expression change is opposite to that of gene expression change. A) The comparison between tumor and normal in AA group. B) The comparison between tumor and normal in CA group. “up_down” indicates the genes with increased expression and hypomethylation events in tumor versus normal comparision, while “down_up” means the genes with decreased expression and hypermethylation events in tumor versus normal comparision.

Mentions: To explore the interplay between expression and methylation, we examined the genes which are detected in both the expression microarray and methylation beadchip. By intercepting the differential expression results with the differential methylation results, we found 548 and 179 differentially expressed genes also have differently methylated loci in AA and CA group, respectively. A portion of them have loci whose direction of methylation expression change is opposite to that of gene expression change (Figure 7). For example, in the AA group, for the 548 differentially expressed genes with differential methylation events in the TdECs vs. NdECs comparison, 157 (29%) are up-regulated and have at least one hypomethlayed locus, while 159 (29%) are down-regulated and have at least one hypermethlayed locus. In CA group, for the 179 differentially expressed genes with differently methylated events in TdECs vs. NdECs comparison, 24 (13%) are up-regulated and have at least one hypomethlayed locus, while 71 (40%) are down-regulated and have at least one hypermethlayed locus. Gene activation might result from hypomethylation, while hypermethylation could lead to gene silencing[44]. Therefore, our data indicate the possibility that epigenetic process might partly influence the observed expression variations between TdECs and NdECs.


Isolation and genome-wide expression and methylation characterization of CD31+ cells from normal and malignant human prostate tissue.

Luo W, Hu Q, Wang D, Deeb KK, Ma Y, Morrison CD, Liu S, Johnson CS, Trump DL - Oncotarget (2013)

The association of differentially expressed genes with differently methylated lociAmong the differentially expressed genes with differential methylation events, a portion of them have loci whose direction of methylation expression change is opposite to that of gene expression change. A) The comparison between tumor and normal in AA group. B) The comparison between tumor and normal in CA group. “up_down” indicates the genes with increased expression and hypomethylation events in tumor versus normal comparision, while “down_up” means the genes with decreased expression and hypermethylation events in tumor versus normal comparision.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3824530&req=5

Figure 7: The association of differentially expressed genes with differently methylated lociAmong the differentially expressed genes with differential methylation events, a portion of them have loci whose direction of methylation expression change is opposite to that of gene expression change. A) The comparison between tumor and normal in AA group. B) The comparison between tumor and normal in CA group. “up_down” indicates the genes with increased expression and hypomethylation events in tumor versus normal comparision, while “down_up” means the genes with decreased expression and hypermethylation events in tumor versus normal comparision.
Mentions: To explore the interplay between expression and methylation, we examined the genes which are detected in both the expression microarray and methylation beadchip. By intercepting the differential expression results with the differential methylation results, we found 548 and 179 differentially expressed genes also have differently methylated loci in AA and CA group, respectively. A portion of them have loci whose direction of methylation expression change is opposite to that of gene expression change (Figure 7). For example, in the AA group, for the 548 differentially expressed genes with differential methylation events in the TdECs vs. NdECs comparison, 157 (29%) are up-regulated and have at least one hypomethlayed locus, while 159 (29%) are down-regulated and have at least one hypermethlayed locus. In CA group, for the 179 differentially expressed genes with differently methylated events in TdECs vs. NdECs comparison, 24 (13%) are up-regulated and have at least one hypomethlayed locus, while 71 (40%) are down-regulated and have at least one hypermethlayed locus. Gene activation might result from hypomethylation, while hypermethylation could lead to gene silencing[44]. Therefore, our data indicate the possibility that epigenetic process might partly influence the observed expression variations between TdECs and NdECs.

Bottom Line: The data, although preliminary, indicates that there exist widespread differences in methylation and transcription between TdECs and NdECs.Interestingly, only a small proportion of perturbed gene was overlapped between American (AA) and Caucasian American (CA) patients with prostate cancer.Future studies will be required to further characterize the isolated ECs and determine the biological features that can be exploited in the prognosis and therapy of prostate cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Therapeutics, Roswell Park Cancer Institute, Buffalo, New York.

ABSTRACT
Endothelial cells (ECs) are an important component involved in the angiogenesis. Little is known about the global gene expression and epigenetic regulation in tumor endothelial cells. The identification of gene expression and epigenetic difference between human prostate tumor-derived endothelial cells (TdECs) and those in normal tissues may uncover unique biological features of TdEC and facilitate the discovery of new anti-angiogenic targets. We established a method for isolation of CD31+ endothelial cells from malignant and normal prostate tissue obtained at prostatectomy. TdECs and normal-derived ECs (NdECs) showed >90% enrichment in primary culture and demonstrated microvascular endothelial cell characteristics such as cobblestone morphology in monolayer culture, diI-acetyl-LDL uptake and capillary-tube like formation in Matrigel®. In vitro primary cultures of ECs maintained expression of endothelial markers such as CD31, von Willebrand factor, intercellular adhesion molecule, vascular endothelial growth factor receptor 1, and vascular endothelial growth factor receptor 2. We then conducted a pilot study of transcriptome and methylome analysis of TdECs and matched NdECs from patients with prostate cancer. We observed a wide spectrum of differences in gene expression and methylation patterns in endothelial cells, between malignant and normal prostate tissues. Array-based expression and methylation data were validated by qRT-PCR and bisulfite DNA pyrosequencing. Further analysis of transcriptome and methylome data revealed a number of differentially expressed genes with loci whose methylation change is accompanied by an inverse change in gene expression. Our study demonstrates the feasibility of isolation of ECs from histologically normal prostate and prostate cancer via CD31+ selection. The data, although preliminary, indicates that there exist widespread differences in methylation and transcription between TdECs and NdECs. Interestingly, only a small proportion of perturbed gene was overlapped between American (AA) and Caucasian American (CA) patients with prostate cancer. Our study indicates that identifying gene expression and/or epigenetic differences between TdECs and NdECs may provide us with new anti-angiogenic targets. Future studies will be required to further characterize the isolated ECs and determine the biological features that can be exploited in the prognosis and therapy of prostate cancer.

Show MeSH
Related in: MedlinePlus