In vitro studies on bone resorption in neonatal mouse calvariae using a modified dissection technique giving four samples of bone from each calvaria

Bone resorption, in vitro, is often measured as the release of prelabelled45Ca from neonatal mouse calvarial bones, or from fetal rat long bones. In this report we describe a technique to measure the breakdown of bone-matrix, in vitro. We also describe a new way to dissect neonatal mouse calvarial bones, in order to obtain large amounts of bone samples. Twelve bone fragments were dissected out from each mouse calvaria and were thereafter cultured in CMRL 1066 culture medium in serum-free conditions in 0.5 cm2 multiwell culture dishes. Matrix degradation after treatment with parathyroid hormone was assessed by measuring the amount of carboxyterminal telopeptide of type I collagen (ICTP) by RIA. The data on matrix degradation was compared to the release of prelabelled45Ca from neonatal mouse calvarial bones. We found that the dose-responses for parathyroid hormone-induced release of prelabelled45Ca and ICTP were identical. In conclusion: RIA-analysis of the ICTP-release is an easy and accurate method to measure degradation of bone-matrix, in vitro. Furthermore, the new dissection technique, described in this report, makes it easy to obtain large amounts of bone samples and thus to perform extensive experiments, e.g. dose-responses for agents that enhance bone resorption.

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Differential effects of glucocorticoids on bone resorption in neonatal mouse calvariae stimulated by peptide and steroid-like hormones

Journal of Endocrinology ◽

10.1677/joe.0.1550513 ◽

1997 ◽

Vol 155 (3) ◽

pp. 513-521 ◽

Cited By ~ 22

Author(s):

HH Conaway ◽

D Grigorie ◽

UH Lerner

Keyword(s):

Bone Resorption ◽

Vitamin D3 ◽

Neonatal Mouse ◽

Receptor Interaction ◽

Differential Effects ◽

All Trans Retinoic Acid ◽

45Ca Release ◽

Stimulation Of

Differential effects on in vitro bone resorption were observed when the glucocorticoids, hydrocortisone and dexamethasone, were added to neonatal mouse calvariae treated with either parathyroid hormone (PTH), 1,25(OH)2-vitamin D3, all trans-retinoic acid (t-RA), or prostaglandin E2 (PGE2). Bone resorption was assessed by analyzing either the release of 45Ca from [45Ca]CaCl2 prelabeled calvarial bones or the release of 3H from [3H]proline prelabeled calvariae. At PGE2 concentrations of 3 x 10(-8) and 3 x 10(-7) mol/l, co-treatment with either 10(-6) mol/l dexamethasone or 10(-6) mol/l hydrocortisone caused additive 45Ca release from neonatal mouse calvariae. In contrast, synergistic release from mouse calvarial bones of both 45Ca and 3H was found after either 10(-6) mol/l hydrocortisone or 10(-6) mol/l dexamethasone was combined with 3 x 10(-11) mol/l PTH treatment for 120 h. Dose-response studies indicated that the synergistic stimulation of 45Ca release from neonatal mouse calvariae by glucocorticoids and PTH could be elicited at glucocorticoid concentrations of 10(-8) to 10(-6) mol/l and at PTH concentrations of 10(-11) to 10(-9) mol/l. Progesterone and RU 38486 (a derivative of 19-nortestosterone with antiglucocorticoid activity) blocked the synergism noted with glucocorticoid and PTH co-treatment, suggesting that interaction between the steroids and PTH was dependent on glucocorticoid receptor interaction. Addition of either 10(-6) mol/l hydrocortisone or 10(-6) mol/l dexamethasone to neonatal mouse calvariae treated with 1,25(OH)2-vitamin D3 (10(-11) and 10(-10) mol/l) also resulted in synergistic stimulation of 45Ca release. In contrast to these observations, the stimulatory effect of t-RA (10(-8) mol/l) on 45Ca release from calvarial bones was abolished in the presence of 10(-6) mol/l dexamethasone. These results suggest that an important role of glucocorticoids may be to synergistically potentiate bone resorption stimulated by PTH and 1,25(OH)2-vitamin D3, but indicate an opposing interaction between the glucocorticoids and bone resorptive retinoids.

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Haptoglobin-stimulated bone resorption in neonatal mouse calvarial bones in vitro

Arthritis & Rheumatism ◽

10.1002/art.1780350517 ◽

1992 ◽

Vol 35 (5) ◽

pp. 587-591 ◽

Cited By ~ 10

Author(s):

Ulf H. Lerner ◽

Nils Fröhlander

Keyword(s):

Bone Resorption ◽

Neonatal Mouse

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The Biology of Corrosion and Wear Debris from Orthopedic Implants

MRS Proceedings ◽

10.1557/proc-550-321 ◽

1998 ◽

Vol 550 ◽

Author(s):

Joshua J. Jacobs ◽

Tibor T. Glant

Keyword(s):

Bone Resorption ◽

Bone Loss ◽

Wear Debris ◽

Degradation Products ◽

Orthopedic Implants ◽

Corrosion And Wear ◽

In Vitro Studies ◽

Type I ◽

Particulate Wear

AbstractBone loss (osteolysis) following total joint arthroplasty has been a subject of increasing concern in the orthopedic research community. Depending on the distribution and severity, bone loss can lead to aseptic loosening, periprosthetic fracture and formidable reconstructive problems at revision surgery. Bone loss is believed to be primarily a response to particulate wear and corrosion debris derived from the prosthetic materials. Phagocytosed particulates activate macrophages and osteoblasts (and perhaps fibroblasts) to produce factors which stimulate osteoclastic bone resorption and reduce osteoblastic bone formation. To investigate the responses of these cells to particulate corrosion and wear debris, in vitro studies have been performed by measuring factors at both the molecular and cellular levels that may trigger, maintain and/or regulate particulate biomaterial-induced pathologic bone resorption. The biological effect of a particulate species depends upon their size, concentration (number) and composition, in the order listed. Particulate wear debris of phagocytosable size (less that 10 micrometers) activate macrophages, fibroblasts and osteoblasts more effectively that those of larger sizes. As a response to phagocytosed particulates, i) macrophages produce a number of cytokines (interleukins such as II-1, II-6, TNF-alpha) and prostaglandins, which may act either in an autocrine fashion or further stimulate cells present in the periprosthetic tissue; ii) fibroblasts secrete active forms of metalloproteinases; and iii) osteoblasts have diminished collagen type I synthesis. Taken together, particulate corrosion and wear debris provoke a series of biological responses which generate an active microenvironment around prosthetic components. Strategies to modify the host response to particulate degradation products have emerged from these in vitro studies. These strategies may provide pharmacotherapeutic solutions to this important clinical problem.

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