Human Osteoclasts and Osteoclast-Like Cells Synthesize and Release High Basal and Inflammatory Stimulated Levels of the Potent Chemokine Interleukin-8¹

Abstract

Chemokines, including interleukin-8 (IL-8), function as key mediators in diverse inflammatory disorders via promoting the recruitment, proliferation, and activation of vascular and immune cells. IL-8 levels are elevated in inflammatory diseases, such as rheumatoid arthritis, osteoarthritis, osteomyelitis, and periodontal disease, that also exhibit progressive bone loss. Therefore, it is possible that IL-8 contributes to the osteopenia associated with these pathological conditions. Although macrophages, neutrophils, and endothelial cells are considered the primary sources of inflammation-induced IL-8 increases, we report here for the first time that human bone marrowderived osteoclast-like cells (hOCL) as well as authentic bone-resorbing human osteoclasts (hOC) isolated from osteoporotic femoral heads express messenger RNA (mRNA) for IL-8 and secrete high levels of IL-8 during culture. Basal IL-8 release by cultured hOC or hOCL was orders of magnitude greater than the release of the proinflammatory cytokines IL-1␤, IL-6, and tumor necrosis factor-␣. At a cellular level, in situ hybridization analysis revealed that IL-8 mRNA was expressed in resorbing hOC of rheumatoid arthritic pannus and was substantially greater than that expressed in hOC of noninflammatory giant cell tumor of bone tissue. Therefore, the potential inflammationmediated induction of IL-8 was directly assessed using cultured hOCL. IL-8 release was stimulated by proinflammatory signals (IL-1␣, tumor necrosis factor-␣, lipopolysaccharide, or phorbol 12-myristate 13-acetate), unaffected by various other osteotropic modulators (transforming growth factor-␤1 and -␤3, IL-6, 17␤-estradiol, or calcitonin) and was decreased by interferon-␥, vitamin D 3 , and the antiinflammatory glucocorticoid dexamethasone. Changes in IL-8 secretion were paralleled by corresponding changes in IL-8 mRNA steady state levels. We conclude that hOC and hOCL synthesize and secrete high constitutive and inflammation-stimulated levels of the chemokine IL-8. Consequently, hOC-derived IL-8 could act as an important regulatory signal for bone, vascular, and immune cell recruitment and activation during normal and pathological bone remodeling. (Endocrinology 139: [4353][4354][4355][4356][4357][4358][4359][4360][4361][4362][4363] 1998)

Autocrine signaling is a key regulatory element during osteoclastogenesis

“…In addition, an increase in IL-8 levels was demonstrated at several diseases associated with osteoclast activation, including rheumatoid and psoriatic arthritis (Cho et al, 2007; Fitzgerald and Chandran, 2012), and several cancers exhibiting high propensity to metastasize to bone, such as breast (Bendre et al, 2005; Bendre et al, 2003), lung (Hsu et al, 2010), renal (Perut et al, 2009) and oral squamous cell carcinoma (Hwang et al, 2012). While it was shown more than 15 years ago that human osteoclasts produce unexpectedly high levels of IL-8 (Rothe et al, 1998), the functional significance of this phenomenon remained unknown. One of the potential explanations of the discrepancy between the demonstrated involvement of IL-8 in human diseases and lack of mechanistic insights into its action is the lack of IL-8 equivalent in rodents, which are most widely used to model human diseases.…”

Receptor Activator of NF-κB and Osteoprotegerin Expression by Human Microvascular Endothelial Cells, Regulation by Inflammatory Cytokines, and Role in Human Osteoclastogenesis

“…Standard curves were run with each assay, and control medium was analyzed for background levels of each cytokine (which were insignificant), and modulators were tested in at least 3 separate culture wells per trial for 2-8 independent HMVEC cultures. Results were normalized for cell protein using the bicinchoninic acid protein assay (Pierce) and bovine serum albumin (BSA) as a standard (33), and data were expressed as the mean Ϯ S.E. ng/ml of cytokine released per mg of cell protein during 24 h of culture.…”

“…Human PBMC were prepared from heparinized blood obtained from the American Red Cross (St. Louis, MO). Mononuclear cells were isolated by Ficoll-Paque (26,32,33), resuspended in ␣-MEM plus 10% FBS and 1% antibiotic/antimycotic, and added (1.6 ϫ 10 6 PBMC/well) to the 24-well dish containing unactivated or cytokine-pre-activated HMVEC. Some wells also received 100 ng/ml recombinant human OPG:Fc fusion peptide (Alexis Corporation, San Diego, CA).…”

The expression of monocyte chemoattractant protein-1 and other chemokines by osteoblasts

“…Bar, 50 um. This production is enhanced by IL-1beta or TNF-alpha stimulation (9,10). IL-8 production has been found in Pagetic osteoblast-like cells and osteosarcoma cells (11).…”