laser-capture-microdissection-human-prostatic-epithelium-for-rna

Laser-capture Microdissection of Human Prostatic Epithelium for RNA Analysis

The prostate gland contains a heterogeneous milieu of stromal, epithelial, neuroendocrine and immune cell types.  Healthy prostate is comprised of ...

handling-and-assessment-of-human-primary-prostate-organoid-culture

Handling and Assessment of Human Primary Prostate Organoid Culture

This paper describes a detailed protocol for three-dimensional (3D) culturing, handling, and evaluation of human primary prostate organoids. The process ...

University of Illinois at Chicago

Vitamin D deficiency increases the risk of lethal prostate adenocarcinomas (PCa) and the majority of older men are deficient. Although PCa arises from the epithelium, the surrounding stroma has hormonal regulatory control over the epithelium and contributes to carcinogenesis. Herein, we describe regulation of microRNAs (miRs) by the active hormone dihydroxyvitamin D (1,25(OH)D) in human prostate stroma. 1,25(OH)D binds the vitamin D receptor (VDR) transcription factor to regulate gene expression, including miRs, which have emerged as potent regulators of protein expression. 1,25(OH)D-regulated miRs were identified by profiling in primary human prostatic stromal cells (PrS) and three miRs, miR-126-3p, miR 154-5p and miR-21-5p were subsequently validated in laser-capture micro-dissected prostate stromal tissue from a vitamin D3 clinical trial (N=45). Regulation of these miRs by 1,25(OH)D was VDR-dependent. Network analysis of known and putative mRNA targets of these miRs was enriched with cancer and inflammation pathways, consistent with known roles of stroma and of vitamin D in carcinogenesis. Expression of the miR processing ribonuclease, DICER1, positively correlated with vitamin D metabolite levels in the clinical trial specimens. High epithelial/stromal ratios of DICER1 were significantly associated biochemical recurrence (OR 3.1, p=0.03) in a tissue microarray of 170 matched PCa patients. In summary, these results underscore the role of the prostate stroma in regulating responses to the hormone 1,25(OH)D and identified miRs and DICER1 as being regulated in human prostate stroma. Regulation of stromal DICER1 by 1,25(OH)D may also have clinical relevance in protection against aggressive PCa.

microRNAs and DICER1 are regulated by 1,25-dihydroxyvitamin D in prostate stroma.

 African American (AA) men are disproportionately affected by both prostate cancer (PCa) and vitamin D deficiency compared with European American (EA) men. Vitamin D deficiency is linked to increased PCa aggressiveness and mortality. Therefore, it has been hypothesized that vitamin D deficiency may contribute to the PCa disparity between AA and EA men.  We studied a cross sectional group of 60 PCa patients (AA,  = 31; EA,  = 29) who underwent radical prostatectomy. Vitamin D metabolites 25-hydroxyvitamin D (25(OH)D) and 1,25-dihydroxyvitamin D (1,25(OH)D) were measured in the serum and tissue by uHPLC-MS-MS. Tissue was laser capture microdissected, and gene expression was quantified by microarray. DNA isolated from whole blood was genotyped for West African ancestry markers and vitamin D-related SNPs.  Serum concentrations of 25(OH)D were lower in AAs, but concentrations of 1,25(OH)D in the prostate tissue were higher compared with EAs. Expression of the vitamin D receptor was higher in prostate tissue from AAs. Expression of the extracellular receptor of vitamin D binding protein, , was positively associated with West African ancestry and inversely associated with tissue 25(OH)D concentrations in AAs.  The relationships between vitamin D binding protein  and vitamin D metabolites suggest that the prohormone is actively transported into the prostate, followed by intraprostatic conversion to the active hormone, rather than passive diffusion. These findings support the presence of a compensatory response in prostate tissue to vitamin D deficiency in AAs and reveal a previously unknown complexity involving tissue distribution of vitamin D metabolites.  Department of Defense Prostate Cancer Research Program Idea Award for Disparities Research PC121923 (LN and RK) and the NIH 1R01MD007105 (RK).

Prostatic compensation of the vitamin D axis in African American men.

The miR-183 cluster, which is comprised of paralogous miRs-183, -96 and -182, is overexpressed in many cancers, including prostate adenocarcinoma (PCa). Prior studies showed that overexpression of individual pre-miRs-182, -96 and -183 in prostate cells decreased zinc import, which is a characteristic feature of PCa tumours. Zinc is concentrated in healthy prostate 10-fold higher than any other tissue, and an >80% decrease in zinc is observed in PCa specimens. Here, we studied the effect of overexpression of the entire 4.8 kb miR-183 family cluster, including the intergenic region which contains highly conserved genomic regions, in prostate cells. This resulted in overexpression of mature miR-183 family miRs at levels that mimic cancer-related changes. Overexpression of the miR-183 cluster reduced zinc transporter and intracellular zinc levels in benign prostate cells, PCa xenografts and fresh prostate epithelial organoids. Microarray analysis of miR-183 family cluster overexpression in prostate cells showed an enrichment for cancer-related pathways including adhesion, migration and wound healing. An active secondary transcription start site was identified within the intergenic region of the miR-183 cluster, which may regulate expression of miR-182. Taken together, this study shows that physiologically relevant expression of the miR-183 family regulates zinc levels and carcinogenic pathways in prostate cells.

ArticleTotal : 2	Year
Publication title Cited by 15	2015
Publication title Cited by 23	2019

Article	Year
microRNAs and DICER1 are regulated by 1,25-dihydroxyvitamin D in prostate stroma. The Journal of steroid biochemistry and molecular biology\| PubMed ID: 28089917	2017
Prostatic compensation of the vitamin D axis in African American men. JCI insight\| PubMed ID: 28138564	2017
The miR-183 family cluster alters zinc homeostasis in benign prostate cells, organoids and prostate cancer xenografts. Scientific reports\| PubMed ID: 28794468	2017

Zachary Richards

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