Reduction of dietary magnesium by only 50% in the rat disrupts bone and mineral metabolism.

INTRODUCTION: The objective of this study was to determine the effect of a moderate reduction of dietary magnesium [50% of nutrient requirement (50% NR)] on bone and mineral metabolism in the rat, and to explore possible mechanisms for the resultant reduced bone mass. METHODS: Female rats were 6 weeks of age at the start of study. Serum magnesium (Mg), calcium (Ca), parathyroid hormone (PTH), 1,25(OH)(2)-vitamin D, alkaline phosphatase, osteocalcin, and pyridinoline were measured during the study at 3- and 6-month time points in control (dietary Mg of 100% NR) and Mg-deficient animals (dietary Mg at 50% NR). Femurs and tibias were also collected for mineral content analyses, micro-computerized tomography, histomorphometry, and immunohistochemical localization of substance P, TNFalpha, and IL-1beta at 3 and 6 months. RESULTS: Although no significant change in serum Mg was observed, Mg deficiency developed, as assessed by the reduction in bone Mg content at the 3- and 6-month time points (0.69+/-0.05 and 0.62+/-0.04% ash, respectively, in the Mg depletion group compared to 0.74+/-0.04 and 0.67+/-0.04% ash, respectively, in the control group; p=0.0009). Hypercalcemia did not develop. Although serum Ca level remained in the normal range, it fell significantly with Mg depletion at 3 and 6 months (10.4+/-0.3 and 9.6+/-0.3 mg/dl, respectively, compared to 10.5+/-0.4 and 10.1+/-0.6 mg/dl, respectively, in the control group; p=0.0076). The fall in serum Ca in the Mg-depleted animals was associated with a fall in serum PTH concentration between 3 and 6 months (603+/-286 and 505+/-302 pg/ml, respectively, although it was still higher than the control). The serum 1,25(OH)(2)-vitamin D level was significantly lower in the Mg depletion group at 6 months (10.6+/-7.1 pg/ml) than in the control (23.5+/- 12.7 pg/ml) (p<0.01 by the t-test). In Mg-deficient animals, no difference was noted in markers of bone turnover. Trabecular bone mineral content gain was less over time in the distal femur with Mg deficiency at 3 and 6 months (0.028+/-0.005 and 0.038+/-0.007 g, respectively, compared to 0.027+/-0.004 and 0.048+/-0.006 g, respectively, in the control group; p<0.005). Histomorphometry at these time points demonstrated decreased trabecular bone volume (15.76+/-1.93 and 14.19+/-1.85%, respectively, compared to 19.24+/-3.10 and 17.30+/-2.59%, respectively, in the control group; p=0.001). Osteoclast number was also significantly increased with Mg depletion (9.07+/-1.21 and 13.84+/-2.06, respectively, compared to 7.02+/-1.89 and 10.47+/-1.33, respectively, in the control group; p=0.0003). Relative to the control, immunohistochemical staining intensity of the neurotransmitter substance P and of the cytokines TNFalpha and IL-1beta was increased in cells of the bone microenvironment in the Mg depletion group, suggesting that inflammatory cytokines may contribute to bone loss. CONCLUSION: These data demonstrate that Mg intake of 50% NR in the rat causes a reduced bone mineral content and reduced volume of the distal femur. These changes may be related to altered PTH and 1,25(OH)(2)-vitamin D formation or action as well as to an increase release of substance P and the inflammatory cytokines TNFalpha and IL-1beta.

Alterations in growth plate and articular cartilage morphology are associated with reduced SOX9 localization in the magnesium-deficient rat.

Insufficient dietary magnesium (Mg) intake has been associated with low bone mass in humans,and recent basic science studies have indicated that this bone loss may be secondary to increased release of substance P and TNFc Much less is known about the effects of low Mg intake on cartilage. We have evaluated growth plate and articular cartilage in rats following a 6 month dietary Mg restriction. Histomorphometry demonstrated significantly decreased distal femur articular cartilage chondrocyte density and decreased tibial growth plate width in experimental animals compared to controls. Growth plates of Mg-restricted animals showed reduced chondrocyte column formation. Extracellular matrix of both articular cartilage and growth plates in experimental animals contained reduced amounts of proteoglycans. Immunolocalization of Sox9 was decreased in both articular and growth plate cartilage in experimental animals compared to controls, suggesting that reduced Mg intake causes cartilage changes that may be secondary to reduced levels of the SOX9 transcription factor.

Magnesium deficiency: effect on bone mineral density in the mouse appendicular skeleton.

BACKGROUND: Dietary magnesium (Mg) deficiency in the mouse perturbs bone and mineral homeostasis. The objective of the present study was to evaluate bone mineral density of the femur in control and Mg-deficient mice. METHODS: BALB/c mice aged 28 days at study initiation were maintained on a normal or Mg deficient (0.0002% Mg) diet, and at time points 0, 2, 4 or 6 weeks bones were harvested for bone mineral density analysis. Peripheral quantitative computed tomography (pQCT) was used to assess the trabecular metaphyseal compartment and the cortical midshaft. RESULTS: Although mean total bone density of the femoral midshaft in Mg deficient mice did not differ significantly from controls throughout the study, the trabecular bone compartment showed significantly decreased mineral content after 4 (p < 0.001) and 6 weeks (p < 0.001) of Mg depletion. CONCLUSIONS: This study demonstrates the profound effect of Mg depletion on the trabecular compartment of bone, which, with its greater surface area and turnover, was more responsive to Mg depletion than cortical bone in the appendicular skeleton of the mouse.

Magnesium deficiency: effect on bone and mineral metabolism in the mouse.

Insufficient dietary magnesium (Mg) intake has been associated in humans with low bone mass. Mg deficiency in the rat has suggested bone loss is due to increased bone resorption and/or inadequate bone formation during remodeling. The purpose of this study was to assess the effect of a low Mg diet on bone and mineral metabolism in the young and mature BALB/c mouse and explore the hypothesis that inflammatory cytokines may contribute to Mg deficiency-induced osteoporosis. Using an artificial diet, we induced targeted Mg depletion (0.002% Mg) with all other nutrients maintained at the normal level. In all Mg-depleted mice, hypomagnesemia developed and skeletal Mg content fell significantly. The serum Ca in Mg-deficient mice was higher than in control mice; however, serum PTH levels were not significantly different. Osteoprotegerin (OPG) in dosages that inhibit osteoclastic bone resorption did not prevent hypercalcemia in Mg-deficient animals. No significant difference in serum Ca was observed between groups when dietary Ca was reduced by 50%, suggesting that a compensatory increase in intestinal absorption might account for the hypercalcemia. Growth plate width decreased 33% in young Mg-deficient animals and chondrocyte columns decreased in number and length, suggesting that Mg deficiency reduced bone growth. Trabecular bone volume in the metaphysis of the tibia in these animals was decreased and osteoclast number was increased by 135%. Osteoblast number was significantly reduced. Immunohistochemistry revealed that substance P increased 230% and 200% in megakaryocytes and lymphocytes, respectively, after 1 day of Mg depletion. IL-1 increased by 140% in osteoclasts by day 3 and TNF alpha increased in osteoclasts by 120% and 500% in megakaryocytes on day 12. This study demonstrates a profound effect of Mg depletion on bone characterized by impaired bone growth, decreased osteoblast number, increased osteoclast number in young animals, and loss of trabecular bone with stimulation of cytokine activity in bone.

Cytokines associated with the pathophysiology of aggressive fibromatosis.

The rare benign extra-abdominal desmoid tumor is characterized by aggressive invasion of normal tissue. Treatment is complicated by its recurrence, invasiveness, and persistence. The etiology is unknown and the pathophysiology is obscure. Because of exuberant fibroblastic proliferation with collagenous tissue being the primary tissue component, this desmoid tumor has been compared with keloids arising from excessive scar formation in healing wounds. Numerous cytokines are associated with signaling for growth and maintenance of mesenchymal cells. Altered expression of these proteins is associated with many pathologic conditions. It has been proposed that the enhanced expression of platelet-derived growth factor and its receptor characterize desmoid tumors. We tested the hypothesis that the exuberant fibrosis of desmoid tumors may have resulted from the initiation of the cascade of molecular events producing increased expression of cytokines. We used immunohistochemical analysis of cytokines in desmoid tumors compared with keloids and skin to localize the expression of cytokines. The results showed localized increased expression of the cytokines epidermal growth factor, transforming growth factor-beta, tumor necrosis factor-alpha, vascular endothelial growth factor, interleukin-1beta, and interleukin-6 in the endothelial cells of blood vessels in the tumors. Production of tumor necrosis factor-alpha and interleukin-1beta in tumor tissue was increased, but we did not find increased expression of platelet-derived growth factor. We concluded that the increased expression of cytokines associated with angiogenesis usually found in wound healing and invasive tumors may contribute to the pathophysiology of the desmoid tumor.

Cytokines expressed in multinucleated cells: Paget's disease and giant cell tumors versus normal bone.

Human osteoclasts are well characterized multinucleated cells whose function is the directed resorption of normal bone (NB). Osteoclastic bone destruction accompanies lytic solid tumors and myeloma as well as Paget's disease (PD) of bone and giant cell tumors of bone (GCTB). The mechanism of this stimulation of osteoclastic bone resorption is unknown. This study was designed to detect cytokines present in the multinucleated cells of PD and GCTB in order to determine whether cytokine abnormalities exist to account for bone lysis. Nine cytokines, representing the functions of bone resorption, angiogenesis, tumor necrosis, bone cell proliferation, and osteoblast-osteoclast coupling, were examined by immunohistochemistry using tissue samples from 15 NB, 17 PD, and 19 GCTB patients. Standard nonparametric statistical analysis showed a significant increase (P < 0.01 to 0.05) in immunostaining between osteoclasts of PD and NB for interleukin-6 (Il-6), tumor necrosis factor beta (TNFbeta), epidermal growth factor (EGF), platelet derived growth factor (PDGF), and basic fibroblast growth factor (bFGF). There was a statistically significant decrease in immunostaining of giant cells of GCTB as compared with NB for transforming growth factor beta (TGFbeta), but no other differences from normal osteoclasts. The increase in staining of PD osteoclasts over the giant cells of GCTB was significant (P < 0.01) for Il-6, TNFbeta, PDGF, bFGF and insulin growth factor-1 (IGF-1), and (P < 0. 05) for Il-1 and EGF. It was concluded that marked cytokine differences exist in vivo between osteoclasts of NB and PD lesions consistent with stimulated resorption. Alternatively, "osteoclastoma" cells in the center of the tumor did not overexpress the cytokines associated with bone lysis, suggesting some other mechanism for stimulated resorption.

Multinucleated cells formed in vitro from Paget's bone marrow express viral antigens.

Paget's disease of bone is characterized by large numbers of osteoclasts that have viral-like nuclear and/or cytoplasmic inclusions. Pagetic osteoclasts express respiratory syncytial viral (RSV) and measles viral (MV) nucleocapsid antigens. The data suggest a possible viral etiology for Paget's disease. However, studies to characterize further the putative viral inclusions in Paget's osteoclasts have been severely hampered by the extreme difficulty in isolating large numbers of osteoclasts from pagetic bone. The recent demonstration that osteoclast-like multinucleated cells (MNC), that had certain characteristics of pagetic osteoclasts formed in marrow cultures from Paget's patients, may permit studies to describe this virus further. Therefore, we have cultured marrow samples from involved and uninvolved bones from Paget's patients and from normal subjects to determine if the MNC formed in these cultures express viral antigens. RSV and/or MV antigens were expressed in the mononuclear cells and/or the MNC formed in 12 of 12 marrow cultures from active lesions of patients with Paget's disease, with 40-50% of the cells expressing viral antigens. In contrast, less than 5% of cells isolated from cultures from normal subjects expressed RSV and/or MV. These results suggest that MNC formed in long-term marrow cultures from patients with Paget's disease frequently express paramyxoviral antigens and are very similar to pagetic osteoclasts. Thus, these marrow cultures may be useful for further characterizing the virus in Paget's disease.

[The absence of characteristics lesions in the microcirculation of non-insulin-dependent diabetic patients]. / Ausencia de lesiones características en la microcirculación de pacientes diabéticos no insulino-dependientes.

Diabetic cardiomyopathy apparently has an important role in the increased cardiovascular morbi-mortality of diabetic patients and its cause is likely to be secondary to small vessel disease. We undertook the present study to compare small and large vessel disease in hearts of patients who died with coronary disease, and determine how diabetes and/or hypertension correlates with these findings. The paraffin blocks of 52 hearts were used in this study. Cases were selected if they died from coronary artery disease and excluded if they had a previous angioplasty, revascularization surgery, congenital, rheumatic or other causes of heart disease. They were divided in two groups; diabetics and non-diabetics and each group was subdivided in hypertensives an non hypertensives. They were matched by age and sex. DM duration was 11 +/- 6 years and known hypertension of 10 +/- 4 years with no significant differences between both groups. The results were recorded without knowledge of patients clinical findings. Atherosclerotic heart disease was more advanced in DM patients, with an increased prevalence of three vessels disease, and more extensive myocardial infarctions. Diabetic subjects had increased (non significant) basal membrane thickening of the capillaries. We could not find differences in parenchymal hypertrophy, interstitial edema, proliferative endothelial lesions and luminal width in middle and large size vessels. Hypertensive patients had increased perivascular fibrosis (NS). Our results suggest that advanced atherosclerotic heart disease is more common in diabetic patients and diabetic cardiomyopathy, if present, seems not to related to a particular structural microvascular disease.