p65-Dependent production of interleukin-1b by osteolytic prostate cancer cells causes an induction of chemokine expression in osteoblasts Jochen Schulze a,1 , Kristoffer Weber b,1 , Anke Baranowsky a,1 , Thomas Streichert c , Tobias Lange d , Alexander Simon Spiro a , Joachim Albers a , Sebastian Seitz a , Josef Zustin e , Michael Amling a , Boris Fehse b , Thorsten Schinke a,⇑ a Department of Osteology and Biomechanics, University Medical Center Hamburg-Eppendorf, Hamburg 20246, Germany b Research Department Cell and Gene Therapy, Clinic for Stem Cell Transplantation, University Medical Center Hamburg-Eppendorf, Hamburg 20246, Germany c Department of Clinical Chemistry, University Medical Center Hamburg-Eppendorf, Hamburg 20246, Germany d Institute of Anatomy and Experimental Morphology, University Medical Center Hamburg-Eppendorf, Hamburg 20246, Germany e Institute of Pathology, University Medical Center Hamburg-Eppendorf, Hamburg 20246, Germany article info Article history: Received 3 June 2011 Received in revised form 9 November 2011 Accepted 11 November 2011 Keywords: Prostate cancer p65 IL-1B Osteoblast abstract Skeletal metastases are a frequent complication of prostate, breast and lung cancer, and the interactions of tumor cells with bone-forming osteoblasts and bone-resorbing osteoclasts have been suggested to play critical roles in disease progression. We have previously shown that treatment of primary murine oste- oblasts with conditioned medium of the human osteolytic prostate cancer cell line PC-3 results in a rapid induction of chemokine expression, thereby providing further evidence for a molecular crosstalk between bone and tumor cells. The aim of our current study was to identify PC-3-derived molecules mediating this effect. Using Affymetrix Gene Chip hybridization followed by qRT-PCR we were able to confirm that the expression of chemokine-encoding genes is markedly induced in human primary osteoblasts following incubation with PC-3-conditioned medium. Since this induction was significantly affected upon alter- ation of p65-levels in PC-3 cells, we performed a second genome-wide expression analysis to identify p65-regulated cytokines, which were then tested for their ability to induce chemokine expression. Here we observed that interleukin-1b (IL-1B) did not only increase the expression of chemokines in osteo- blasts, but also the phosphorylation of p65 and thereby its own expression. Since immunohistochemistry on bone biopsy sections from prostate cancer metastases demonstrated IL-1B expression in both, tumor cells and osteoblasts, our data suggest that IL-1B is one of the relevant cytokines involved in the skeletal complications of cancer metastases. Ó 2011 Elsevier Ireland Ltd. All rights reserved. 1. Introduction Prostate cancer is one of the most common forms of cancer in men and one of the leading causes of cancer-related death in the USA [1]. Bone metastases are a frequent complication of the dis- ease, occurring in nearly 70% of the patients with advanced pros- tate cancer, and they are also found in other cancer forms, especially breast and lung cancer [2]. Depending on the skeletal complications caused by these metastases, they can be classified as osteoblastic (causing increased bone formation) and/or osteo- lytic (causing excessive bone resorption) [3]. Since both activities, bone formation and bone resorption, can only be performed by specific cell types, namely osteoblasts and osteoclasts, it has been hypothesized that tumor cells produce molecules with the ability to regulate bone cell differentiation and/or activity [4]. Moreover, since bone cell-specific production, but also resorption of the bone matrix, can result in a local increase of certain growth factors, it has been postulated that bone and tumor cells can stimulate each other [5]. This so-called vicious cycle of skeletal metastases has been a matter of intensive research over the past years, since iden- tification of the involved factors will certainly reveal new thera- peutic strategies for the respective patients. One way to study the molecular crosstalk between bone and tu- mor cells is the use of cell lines derived from patients with skeletal metastases. The cell lines MDA-PCa-2b and PC-3, which were both initiated from bone metastases of prostatic adenocarcinomas, are especially valuable in this regard, since they have a different influ- ence on bone turnover, when injected into immunodeficient mice. In fact, while MDA-PCa-2b cells trigger increased bone formation reminiscent of osteoblastic metastases, PC-3 cells do not act osteo- anabolic, but promote the development of osteolytic lesions caused by excessive bone resorption [6–8]. Since it was likely that soluble 0304-3835/$ - see front matter Ó 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.canlet.2011.11.016 ⇑ Corresponding author. Tel.: +49 40 7410 58057; fax: +49 40 7410 58010. E-mail address: schinke@uke.uni-hamburg.de (T. Schinke). 1 These authors contributed equally to this work. Cancer Letters 317 (2012) 106–113 Contents lists available at SciVerse ScienceDirect Cancer Letters journal homepage: www.elsevier.com/locate/canlet