Inhibitory Effect of Thymoquinone on Testosterone-Induced Benign Prostatic Hyperplasia in Wistar Rats Bahaa Al-Trad, 1 * Mazhar Al-Zoubi, 2 Janti Qar, 1 Khalid Al-Batayneh, 1 Emad Hussien, 1 Riyadh Muhaidat, 1 Alaa Aljabali, 3 Hakam Alkhateeb 2 and Ghada Al Omari 1 1 Department of Biological Sciences, Yarmouk University, Irbid, Jordan 2 Department of Basic Medical Sciences, Faculty of Medicine, Yarmouk University, Irbid, Jordan 3 Faculty of Pharmacy, Yarmouk University, Irbid, Jordan This study addresses the possible protective effects of thymoquinone (TQ) against the development of experimentally-induced benign prostatic hyperplasia (BPH) in Wistar rats. Eighteen adult male rats were divided into three groups; the negative control group (n = 6) received vehicle, and two groups received subcutaneous testosterone injection (3 mg/kg). Animals receiving testosterone were randomized to untreated BPH group (n = 6) and BPH + TQ treated group (n = 6, 50 mg/kg orally for 14 days). Histological changes and the mRNA levels of transforming growth factor-β 1 (TGF-β 1 ) and vascular endothelial growth factor-A (VEGF-A) were analyzed. Additionally, dihydrotestosterone and interleukin-6 (IL-6) serum levels were determined. The presented research shows significant increases in prostate weight/body weight ratio, prostate epithelial thickness, serum IL-6 and dihydrotestosterone levels, and the prostatic expressions of TGF-β 1 and VEGF-A in the untreated BPH rats. Histological examination of the prostate tissues in the BPH rats showed an elevated level of proliferation in the stromal area and glandular epithelia with abundant intraluminal papillary folds. However, a reduction in prostate weight/body weight ratio, epithelial hyperplasia, serum IL-6 levels, and the expressions of TGF-β 1 and VEGF-A were observed in the BPH + TQ treated rats compared with the untreated BPH rats. The findings support TQ as a useful natural treatment for animal BPH model. Copyright © 2017 John Wiley & Sons, Ltd. Keywords: thymoquinone; prostatic hyperplasia; testosterone; Wistar rats. INTRODUCTION Benign prostatic hyperplasia (BPH), clinically described as a benign enlargement of the prostate and lower urinary tract symptom, is one of the major age-related urogenital disorders in men (Roehrborn, 2008). BPH is defined histologically by a progressive increase in the number of fibromuscular stromal and glandular epithelial elements of the prostate (White et al., 2013). Although the benign neoplastic process of BPH is a multifactorial disease and is not yet completely under- stood, disruption of the molecular mechanisms that regulate stromal and glandular epithelial cell prolifera- tion, differentiation, and apoptosis is proposed to be associated with the pathogenesis of BPH (Roehrborn, 2008). During this process, expression of different growth factors, such as transforming growth factor-β 1 (TGF-β 1 ), is upregulated (Kyprianou et al., 1996; Rick et al., 2011). Alterations in the prostatic vascularization have also been suggested to play an active role in the develop- ment of BPH (Sciarra et al., 2002; Walsh et al., 2002). This hypothesis is supported by studies that showed that the expression of vascular endothelial growth factor (VEGF), an angiogenic factor, is increased in both BPH and prostate cancer cells (Doll et al., 2001; Walsh et al., 2002). VEGF stimulates endothelial cell growth (i.e. neovascularization) that is a key factor required for pathogenesis and progression of BPH and prostate cancer. In a human prostate cancer cell line, tumor growth has been prevented by the suppression of angio- genesis by using anti-VEGF antibodies (Borgström et al., 1998). In addition, increasing evidence has shown that the presence of prostatic inflammation might stimulate the uncontrolled stromal and epithelial cell proliferation through different molecular pathways involving hormonal changes and release of several cytokines (such as, IL-17 and IL-6) and growth factors (De Nunzio et al., 2016). At present, there are a variety of drugs that can treat BPH. However, side effects such as erectile dysfunction, are still a major challenge (Bullock and Andriole, 2006; Traish et al., 2011). In light of this, the search for substances that effectively inhibit the development of BPH combined with proven safety and no induction of side effects is urgently needed. Herbally derived remedies may provide an alternative to new drugs for BPH treatment without adverse side effects. Thymoquinone (TQ), a natural compound from the medicinal plant Nigella sativa, has been shown to exhibit * Correspondence to: Dr Bahaa Al-Trad, Department of Biological Sci- ences, Yarmouk University, Irbid, Jordan. E-mail: bahaa.tr@yu.edu.jo Contract/grant sponsor: Deanship of Research and Graduate Studies at Yarmouk University; contract/grant number: 14/2015. PHYTOTHERAPY RESEARCH Phytother. Res. (2017) Published online in Wiley Online Library (wileyonlinelibrary.com) DOI: 10.1002/ptr.5936 Copyright © 2017 John Wiley & Sons, Ltd. Received 28 January 2017 Revised 10 August 2017 Accepted 06 September 2017