Differential expression of estrogen-related receptors beta and gamma (ERRbeta and ERRgamma) and their clinical significance in human prostate cancer

Abstract

Estrogen-related receptor (ERR) is a nuclear receptor that modulates the estrogen-signaling pathway. Here, we investigated the expression of both ERRbeta and ERRgamma in human prostate tissues. Using original rabbit polyclonal anti-ERRbeta and anti-ERRgamma antibodies, the expression of ERRbeta and ERRgamma was evaluated by immunohistochemical analysis of cancerous lesions (n = 107) and benign foci (n = 92), obtained by radical prostatectomy. Stained slides were evaluated for the proportion of immunoreactive cells and their staining intensity. Total immunoreactivity scores (IR scores; range, 0-8) were calculated as the sum of the proportion and intensity scores. The relationship between the clinicopathological characteristics of the patients and the expression of the three ERRs (ERRalpha, ERR beta, and ERR gamma) was evaluated. IR scores for ERRbeta and ERRgamma were significantly lower in cancerous lesions than that in benign foci (P < 0.0001, for both). Clinicopathological analyses revealed that the patients with low ERRgamma IR scores (<or=4) tended to show poor cancer-specific survival (P = 0.07). Then, we used data from our previous study (Fujimura T., Int J Cancer, 2007; 120: 2325-30). Patients with a high IR score for ERRalpha and a low score for ERRgamma showed significantly poorer cancer-specific survival than those with a low IR score for ERRalpha and a high score for ERRgamma (P = 0.0003). We demonstrated the differential expression of ERRbeta and ERRgamma in prostate tissue. The combined evaluation of the expression of ERRalpha and ERRgamma could be a significant prognostic factor for prostate cancer.

Figure 1

Western blot analysis showing the estrogen‐related receptor (ERR)‐β and ERRγ proteins in 293T cells transfected with pcDNA3‐FLAG‐ERRβ (F‐ERRβ) and pcDNA3‐FLAG‐ERRγ (F‐ERRγ). All the cell extracts were subjected to immunoblotting with the generated anti‐ERRβ (a), anti‐ERRγ (b), and anti‐FLAG antibodies. Anti‐ERRβ and ERRγ antibodies were detected bands, which corresponded to the molecular weight of ERRβ (56 kD) and ERRγ (51 kD), respectively (arrows).

Figure 2

Immunohistochemical staining for estrogen‐related receptor (ERR)‐β (a–d) and ERRγ (e–h) in human renal and prostatic tissues. Positive staining for ERRβ and ERRγ was observed in the nuclei of renal glomerular and epithelial cells, respectively (a,e). Immunoreactivities (IRs) for anti‐ERRβ and ERRγ antibodies were abundant in benign prostatic epithelium (IR score, 7; immunointensity, strong) (b,f). Decreased ERRβ and ERRγ IRs were observed in low‐grade prostate cancer (PCa) (Gleason score [GS], 6) (IR score, 5; immunointensity, moderate) (c,g), and weak expression of ERRβ and ERRγ was also observed in high‐grade PCa (GS 9) (IR score, 4; immunointensity, moderate) (d,h). Original magnification, ×400; Scale bar = 50 μm.

Figure 3

Immunoassaying of estrogen‐related receptor (ERR)‐β (a) and ERRγ (b) in the human prostate. We evaluated 107 cancerous and 92 benign foci. ERRβ and ERRγ immunoreactivities (IRs) were positive in 84 of 92 (91%) and 89 of 92 cases (95%) of benign epithelium, respectively, and in 71 of 106 (67%) and 85 of 106 (80%) cancer cases, respectively. Over 80% and 92% cases of benign foci showed IR scores of ≥5 for ERRβ and ERRγ, respectively, whereas IR scores of ≥5 were obtained in the case of 35% and 55% of patients with cancerous lesions, respectively.

Figure 4

Cancer‐specific survival in patients with prostate cancer according to the immunoreactivity (IR) of estrogen‐related receptor (ERR)‐β and ERRγ (n = 107). No significant difference was observed in ERRβ IR (a), whereas patients with low ERRγ IR tended to show poor cancer‐specific survival (b). The patients with high ERRα IR and low ERRγ IRs (IR score, ≤4) showed significantly poorer cancer‐specific survival than patients with low ERRα IR and high ERRγ IRs and the other patients (P = 0.0003 and 0.07, respectively) (c).