Elsevier

Cytokine

Volume 47, Issue 3, September 2009, Pages 185-193
Cytokine

Eviprostat suppresses proinflammatory gene expression in the prostate of rats with partial bladder-outlet obstruction: A genome-wide DNA microarray analysis

https://doi.org/10.1016/j.cyto.2009.06.004Get rights and content

Abstract

Prostatic inflammation plays a role in the progression of benign prostatic hyperplasia (BPH). Eviprostat is an antioxidant, antiinflammatory phytotherapeutic agent widely used to treat lower urinary tract symptoms in BPH. Because Eviprostat is a mixture of compounds from multiple natural sources, however, its mechanism of action has been difficult to investigate. Here, we describe the use of oligonucleotide microarrays to investigate changes in gene expression in the prostate of rats with surgically induced partial bladder-outlet obstruction and the effect of Eviprostat on those changes. Several dozen proinflammatory genes were activated in obstructed rats, including cytokine, arachidonic acid cascade enzyme, Toll-like receptor (TLR), and transcription factor genes, and their expression was suppressed by Eviprostat. Pathway analysis revealed that several proinflammatory pathways were activated, including cytokine and TLR signaling pathways. The differential expression of selected genes was verified by real-time reverse-transcriptase polymerase chain reaction. Our findings suggest that prostate inflammation in our rat model of partial bladder-outlet obstruction is related to the increased expression of nuclear factor κB (NF-κB) and the induction of proinflammatory cytokines, and that Eviprostat suppresses their expression at the transcriptional level. The prostate inflammation seen in BPH and the clinical benefits of Eviprostat may be similarly explained.

Introduction

Benign prostatic hyperplasia (BPH) is a common proliferative disease in older men, with 40% having the disease by age 50 and over 80% by age 80 [1]. BPH leads to obstruction of the bladder outlet, which is associated with marked alterations in bladder structure and function characterized by lower urinary tract symptoms such as urinary frequency and urgency [2]. BPH is frequently accompanied by prostatic inflammation [3]. For example, in one study, the incidence of inflammation in surgically resected hyperplastic prostate tissue amounted to 98.1% [4]. It has recently been recognized that prostatic inflammation in BPH patients plays a role in the induction and progression of this disease [5].

Medicinal options for the treatment of BPH and lower urinary tract symptoms include α1-adrenergic receptor antagonists, which relax the muscles within the prostate and bladder neck; 5α-reductase inhibitors, which inhibit the conversion of testosterone to dihydrotestosterone, thus reducing cellular proliferation in the prostate; and plant extracts, or phytotherapeutic agents [6]. Eviprostat, which has been prescribed for more than 40 years in Japan and Germany, is the most widely used phytotherapeutic agent for the treatment of BPH. It is a mixture of plant extracts derived from the umbellate wintergreen (Chimaphila umbellata), aspen (Populus tremula), small pasque flower (Pulsatilla pratensis), and field horsetail (Equisetum arvense), combined with germ oil of wheat (Triticum aestivum). The individual components of Eviprostat are known to have diuretic, antiseptic and antiinflammatory effects [7]. The components of Eviprostat also show approximately equal suppression of superoxide anion and hydroxyl radical levels in cell-free systems and human neutrophils, and in rats a single oral dose of Eviprostat prevents carrageenan-induced paw edema in a rat model of inflammation [8]. Eviprostat therefore has both antioxidant and antiinflammatory activity. However, because it is a complex mixture of compounds from multiple natural sources, the molecular mechanism of its beneficial effects has been difficult to investigate. Recently, though, microarray technology has been successfully applied to the investigation of the molecular mechanisms of action of herbal drugs [9], [10], [11], and it should be similarly applicable to the investigation of the mechanisms of phytotherapeutic agents such as Eviprostat.

Animal models of bladder-outlet obstruction are usually used to investigate the effects of obstruction on bladder morphology and pharmacology [12]. For example, repeated administration of Eviprostat inhibits spontaneous bladder contractions during urine storage and restores urinary frequency in rats with partial bladder-outlet obstruction [13]. However, bladder-outlet obstruction is reported to induce not only bladder inflammation but also prostatic inflammation in the rat [14]. Here, we used a rat model of surgically induced partial bladder-outlet obstruction to investigate the effect of oral Eviprostat on gene expression in the inflamed rat prostate. Genome-wide DNA microarray analysis and quantitative real-time reverse-transcriptase polymerase chain reaction (RT-PCR) revealed that proinflammatory signaling pathways regulated by nuclear factor κB (NF-κB) were activated in the inflamed prostate and that Eviprostat repressed these pathways at the transcriptional level.

Section snippets

Animals

Male 10-week-old Sprague–Dawley (Slc:SD) rats (Japan SLC, Hamamatsu, Shizuoka, Japan) were housed two per cage in a room maintained at 20–26 °C and a relative humidity of 35–75% with an alternating 12-h light/dark cycle (the lights came on automatically at 8:00 a.m.). Food and water were freely available. All experimental procedures were approved by the Experimental Animal Research Committee of Nippon Shinyaku Co., Ltd.

Preparation of Eviprostat suspension

Eviprostat tablets produced by Nippon Shinyaku Co., Ltd., for prescription in

Histological analysis

In contrast to normal findings in sham-operated animals (Fig. 1A and B), rats that had undergone surgical partial bladder-outlet obstruction followed by treatment with vehicle showed leukocyte inflammatory infiltration, interstitial fibrosis, and hemorrhage of the prostate and urethra (Fig. 1C and E). Most of the infiltrating leukocytes were macrophages, as determined by their morphology and immunohistochemical properties (Supplementary Fig. S1). Twice-daily treatment with Eviprostat prevented

Discussion

Like Eviprostat, traditional Chinese herbal medicines are mixtures of a variety of known and unknown compounds whose molecular mechanisms of action have been difficult to investigate. Recently, however, several groups have applied microarray technology to studies on the molecular mechanism of herbal drugs [9]. For example, cDNA microarrays containing 3000 human genes derived from a leukocyte cDNA library have been used to investigate root extracts of the vine Tripterygium hypoglaucum, a Chinese

Acknowledgments

We thank Mr. K. Hamada and Mr. T. Nakanishi, Nippon Shinyaku Co., Ltd., for useful suggestions during the course of the work and Mr. K. Hamada, Mr. T. Nakanishi, and Dr. G.E. Smyth, Nippon Shinyaku Co., Ltd., for critical reading of the manuscript.

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