C.W.G.M. (Clemens) Löwik

Parathyroid hormone (PTH) and PTH-like protein (PLP) stimulate interleukin-6 production by osteogenic cells: A possible role of interleukin-6 in osteoclastogenesis

Osteogenic cells mediate PTH-stimulated osteoclastic bone resorption by a yet unidentified mechanism. We show that primairy rat osteoblast-like cells and the clonal osteogenic sarcoma cell line UMR-106 produce interleukin-6 (IL-6) and that bPTH(1-84) and synthetic hPLP(1-34) stimulate this production dose-dependently. With both peptides a close relation between IL-6 and cyclic-AMP production was found, though for PTH concentrations higher than 2.10(-8) M a clear dissociation was observed. Significant IL-6 activity was also detected in media of cultures of 17-day-old fetal mouse radii and metacarpals which was clearly stimulated by PTH. The source of IL-6 in these bone explants seems to be the osteogenic (cartilage) cells. Treatment of bone explants with IL-6 induced osteoclastic resorption which, however, depended on the bone resorption system used. This bone resorbing action of IL-6 is exerted probably through an effect on the formation of osteoclasts (osteoclastogenesis) rather than on the activation of already existing mature osteoclasts. We suggest that IL-6 produced by osteogenic cells may be a mediator in PTH-stimulated osteoclastic bone resorption.

Bisphosphonates inhibit the adhesion of breast cancer cells to bone matrices in vitro.

Bisphosphonates are used with increasing frequency in the management of skeletal complications in patients with breast cancer. In this paper, we have investigated whether bisphosphonates, besides their known beneficial effects on tumor-associated osteoclastic resorption, are capable of inhibiting breast cancer cell adhesion to bone matrix. For that we used two in vitro models for bone matrix (cortical bone slices and cryostat sections of trabecular bone from neonatal mouse tails). Four bone matrix-bound nitrogen-containing bisphosphonates (pamidronate, olpadronate, alendronate, and ibandronate) inhibited adhesion and spreading of breast cancer cells to bone dose-dependently, whereas etidronate and clodronate had little or no effect. Strikingly, the relative order of potency of the bisphosphonates in inhibiting the adhesion of cancer cells to cortical and trabecular bone corresponded to their relative antiresorptive potencies in vivo as well as their ranking in in vitro bone resorption assays with predictive value for their clinical efficacy. It appears that nitrogen-containing bisphosphonates alter selectively the adhesive properties of the extracellular bone matrix preventing the attachment of breast cancer cells to it. Besides the beneficial effects of bisphosphonates on tumor-induced osteoclastic resorption, the previously unrecognized effect presented in this paper makes these agents suitable for earlier pharmacologic intervention in patients with breast cancer at risk of developing bone metastases.

Interleukin 17 synergises with tumour necrosis factor α to induce cartilage destruction in vitro

Background: Interleukin 17 (IL17) is produced by activated T cells and has been implicated in the development of bone lesions and cartilage degradation in rheumatoid arthritis (RA). Objective: To determine whether IL17, alone or together with tumour necrosis factor α (TNFα), induces cartilage destruction in vitro. Methods: Fetal mouse metatarsals stripped of endogenous osteoclast precursors were used to study the effect of IL17 on cartilage degradation independently of osteoclastic resorption. Cartilage destruction was analysed histologically by Alcian blue staining. Results: IL17 alone, up to 100 ng/ml, had no effect on the cartilage of fetal mouse metatarsals. IL17 (≥0.1 ng/ml), however, induced severe cartilage degradation when given together with TNFα (≥1 ng/ml). The cytokine combination decreased Alcian blue staining, a marker of proteoglycans, throughout the metatarsals and induced loss of the proliferating and early hypertrophic chondrocyte zones. TNFα alone also decreased Alcian blue staining, but not as dramatically as the cytokine combination. In addition, it did not induce loss of chondrocyte zones. Treatment with inhibitors of matrix metalloproteinase (MMP) activity and nitric oxide synthesis showed that MMP activity played a part in cartilage degradation, whereas nitric oxide production did not. Conclusions: IL17, together with TNFα, induced cartilage degradation in fetal mouse metatarsals in vitro. IL17 may, therefore, participate in the development of cartilage destruction associated with RA by enhancing the effects of TNFα and may provide a potential therapeutic target.

Accelerated Atherosclerosis and Calcification in Vein Grafts: A Study in APOE*3 Leiden Transgenic Mice

Abstract— Vein grafts fail due to development of intimal hyperplasia and accelerated atherosclerosis. Many murine genetic models in which genes are overexpressed, deleted, or mutated have been introduced recently. Therefore, mouse models are very well suited to dissect the relative contribution of different genes in the development of accelerated atherosclerosis. In the present study, we evaluated whether accelerated atherosclerosis in human vein grafts could be mimicked in hypercholesterolemic APOE*3 Leiden transgenic mice. Venous bypass grafting was performed in the carotid artery in APOE*3 Leiden mice fed either a standard chow diet or a high cholesterol–rich diet for 4 weeks. At several time points (0 hour to 28 days), mice were euthanized and the morphology of the vein grafts was analyzed. In normocholesterolemic mice, vein graft thickening up to 10-fold original thickness, predominantly consisting of &agr;-smooth muscle cell actin–positive cells, was observed after 28 days. In hypercholesterolemic mice, accelerated atherosclerosis with accumulation of lipid-loaded foam cells was observed within 7 days after surgery. This accelerated atherosclerosis progressed in time and resulted in significant increase in vein graft thickening up to 50 times original thickness with foam cell–rich lesions and calcification within 28 days after surgery. The atherosclerotic lesions observed in these murine grafts show high morphological resemblance with the atherosclerotic lesions observed in human vein grafts. This accelerated, diet-dependent induction of atherosclerotic-like lesions in murine vein grafts provides a valuable tool in evaluating the mechanisms of accelerated atherosclerosis and therapeutic interventions of vein graft disease.

Ovariectomy and orchidectomy induce a transient increase in the osteoclastogenic potential of bone marrow cells in the mouse.

Withdrawal of gender steroids in both women and men is associated with an increase in bone turnover with bone resorption exceeding bone formation leading to bone loss. To further investigate this process, the osteoclastogenic potential of mouse bone marrow cells was assessed at different timepoints after ovariectomy (ovx) or orchidectomy (orx). Cocultures of osteoclast-free fetal mouse long bones together with bone marrow from ovariectomized or orchidectomized mice indicated that the withdrawal of gender steroids in female and male mice induces a transient increase in osteoclastogenesis. The osteoclastogenic potential of the bone marrow cells was increased 7 days after ovx or orx. However, osteoclastic resorption was not increased at 3 days after surgery and had normalized 30 days after either ovx or orx. These results suggest that the withdrawal of gender steroids induces a transient increase in osteoclastogenesis in mice of both genders, which is associated with the early phase of rapid bone loss.

Effect of alendronate treatment on the osteoclastogenic potential of bone marrow cells in mice.

Bisphosphonates suppress bone resorption by inhibiting the activity of mature osteoclasts as well as the formation of osteoclasts from bone marrow-derived precursor cells. It is not yet known at which level of osteoclast development this latter action is exerted and whether this is due to a systemic effect of circulating bisphosphonate or to an action at the bone surface, given the property of these compounds to concentrate specifically at active bone sites. We addressed these questions in an ex vivo study in mice. The animals were treated with the bisphosphonate alendronate for various periods, and the central compartment of the bone marrow was isolated and cultured together with osteoclast-free fetal mouse bone explants. In this way the capacity of bone marrow cells, exposed previously to alendronate in vivo, to generate osteoclasts and induce resorption in vitro was examined. Alendronate (0.25 mg/kg, subcutaneously) given to mice for periods up to 4 weeks suppressed bone resorption, as expected, but did not affect the capacity of bone marrow cells to develop into mature osteoclasts and resorb the calcified matrix of bone explants. In addition, using monoclonal antibodies specific for different macrophage-granulocyte precursor subsets, we found that alendronate treatment did not affect the pattern of antigen expression of bone marrow cells, which is in line with the lack of an effect of the bisphosphonate on the ability of bone marrow cells to induce resorption in vitro. In contrast, in control experiments, lipopolysaccharide (0.1 mg) treatment induced marked changes in the pattern of antigen expression of bone marrow cells. In conclusion, our studies demonstrate that the inhibition of bone resorption by alendronate does not involve alteration of the osteoclastogenic potential of osteoclast progenitors (precursors) from the central bone marrow compartment. Moreover, this treatment did not alter the expression of markers specific for monocyte-macrophage precursors as well as mature macrophages. These results suggest that the effects of alendronate are exerted at the bone surface at late osteoclast precursors (and mature osteoclasts).