Increased expression of the interleukin-11 receptor and evidence of STAT3 activation in prostate carcinoma

Abstract

Previous investigations have shown that interleukin-6, a member of the JAK-STAT activating family of cytokines, plays an important role in prostate carcinoma. Here we demonstrate the co-expression of another member of this cytokine family, interleukin-11 (IL-11), and components of its receptor (interleukin-11 receptor; IL-11R), ie, IL-11Ralpha (involved in ligand recognition), and gp130 (involved in signal transduction) in cultured normal and malignant prostate-derived epithelial cell lines. In the DU-145 prostate carcinoma cell line, rhIL-11 stimulates a transient and dose-dependent increase in the tyrosine 705-phosphorylated, active form of STAT3 (STAT3 P-Tyr705), involved in the downstream signaling of IL-11R and other members of the gp130-dependent receptors. The ability of IL-11 to activate STAT3 in prostate-derived cells may be mechanistically important, given recent data suggesting that constitutively activated STAT3 may be associated with the malignant phenotype. In 51 human primary tissues derived from normal prostate, benign prostatic hyperplasia, and prostate carcinomas, IL-11Ralpha and gp130 were commonly expressed, with a statistically significant elevation in the expression of IL-11Ralpha in prostate carcinoma. Also, the tyrosine-phosphorylated, activated form of STAT3 was observed more prominently in the nuclei of cells residing in malignant glands compared to those in nonmalignant samples. Thus, the IL-11 receptor system is up-regulated in prostate carcinoma, and may be one part of a cytokine network that maintains STAT3 in its activated form in these tissues.

Figure 1.

Evidence for the expression of IL-11 and IL-11Rα in normal and malignant prostate epithelial cell lines. A: Expression of the mRNA for IL-11 (left) and IL-11Rα (right) in the normal prostate epithelial cell line, PrEC (lane 2) and the prostate carcinoma cell lines DU-145, PC-3, and LNCaP (lanes 3–5, respectively) as determined by RT-PCR. Experimental procedures are described in Materials and Methods. Identity of the PCR bands were confirmed by Southern blotting using antisense oligonucleotide probes specific to internal sequences within the expected PCR products (see Material and Methods). B: Secretion of IL-11 by normal and malignant prostate epithelial cell lines as detected by enzyme-linked immunosorbent assay. Experimental procedures are described in Materials and Methods. In the case of each cell line, “control” refers to the respective culture medium incubated in the absence of cells (in the case of PrEC, PrEC growth medium, DU-145, minimal essential medium with 10% fetal bovine serum; LNCaP, RPMI 1640 with 10% fetal bovine serum). The IL-11 levels in all control samples were less than the detection limits of the assay (<8 pg/ml), as indicated by the asterisk. Values represent the mean ±SD for three independent determinations. C: Immunohistochemical staining of PrECs (top left) and LNCaP cells (bottom left) for IL-11Rα. Experimental methodology is described in Material and Methods. Right: The immunostaining was performed on these cell types using antibodies to IL-11Rα that had been preincubated with a fivefold excess by weight of a IL-11Rα-specific peptide to which the antibody had been raised (blocking peptide). Original magnification, ×600. D: Activation of phosphorylation of STAT3 at Tyr705 by rhIL-11 in DU-145 prostate carcinoma cells, as determined by immunoblotting. Top: Time course in minutes (“) of STAT3 phosphorylation in the absence (left) or presence (right) of 250 ng/ml rhIL-11. Pos Cont, positive control for STAT3 P-Tyr705, SK-N-MC cells treated with ciliary neurotrophic factor. Bottom: Dose response relationship. Cell treatments and immunoblotting methodology are described in Materials and Methods.

Figure 2.

Immunohistochemical staining of normal human prostate and primary prostate carcinoma for IL-11Rα. Experimental procedures are described in Materials and Methods. A: Normal prostate tissue section showing weak epithelial cell staining, but high expression of IL-11Rα in stromal cells. B: Expression of IL-11Rα in high-grade prostatic intraepithelial neoplasia (PIN, arrows). C: High level of expression of IL-11Rα in prostate carcinoma with perineural invasion (arrow). D: High level of expression of IL-11Rα in prostate carcinoma. Original magnification, ×200.

Figure 3.

Immunohistochemical staining of prostate carcinoma (A) and normal prostate tissue (B) for the tyrosine-phosphorylated, activated form of STAT3 (STAT3 P-Tyr705). Note the strong nuclear staining in prostate carcinoma (A) as compared to the weak, more sporadic staining in the normal epithelial cells (B). Experimental methodology is described in Materials and Methods. Original magnification, ×400.