Decrease In Bone Calcium Content Research Articles

Beta-Cryptoxanthin stimulates bone formation and inhibits bone resorption in tissue culture in vitro.

The effect of beta-cryptoxanthin, which is greatly present in fruits, has not been clarified so far on bone metabolism. The effect of beta-cryptoxanthin on bone formation and bone resorption was investigated in tissue culture in vitro. Rat femoral-diaphyseal (cortical bone) and -metaphyseal (trabecular bone) tissues were cultured for 48 h in Dulbecco's modified Eagle's medium (high glucose, 4.5%) supplemented with antibiotics and bovine serum albumin. The experimental cultures contained 10(-8)-10(-5) M beta-cryptoxanthin. The presence of beta-cryptoxanthin (10(-6) or 10(-5) M) caused a significant increase in calcium content, alkaline phosphatase activity and deoxyribonucleic acid (DNA) content in the diaphyseal and metaphyseal tissues. These increases were completely prevented in the presence of cycloheximide (10(-6) M), an inhibitor of protein synthesis. beta-Carotene (10(-6) or 10(-5) M) or xantine (10(-6) or 10(-5) M) had no effect on the diaphyseal and metaphyseal calcium contents. The bone-resorbing factors parathyroid hormone (1-34) (PTH; 10(-7) M) or prostaglandin E2 (PGE2; 10(-5) M) caused a significant decrease in calcium content in the diaphyseal and metaphyseal tissues. The decrease in bone calcium content induced by PTH or PGE2 was completely inhibited by beta-cryptoxanthin (10(-8)-10(-6) M). In addition, beta-cryptoxanthin (10(-8)-10(-6) M) completely inhibited the PTH (10(-7) M)- or PGE, (10(-5) M)-induced increase in medium glucose consumption and lactic acid production by diaphyseal and metaphyseal tissues. The inhibitory effect of beta-cryptoxanthin (10(-7) M) on PTH (10(-7) M)- or PGE2 (10(-5) M)-stimulated decrease in the diaphyseal calcium content was significantly prevented in the presence of 10(-3) M vanadate, an inhibitor of protein tyrosine phosphatase. Vanadate (10(-3) M) did not have a significant effect on calcium content and lactic acid production in control bone tissues. The present study demonstrates that beta-cryptoxanthin has a direct stimulatory effect on bone formation and an inhibitory effect on bone resorption in tissue culture in vitro.

Inhibitory Effect of Marine Alga Sargassum horneri Extract on Bone Resorption in Tissue Culture in Vitro.

The effect of marine alga Sargassum horneri extracts on bone resorption in vitro was investigated. Femoral-diaphyseal and -metaphyseal tissues obtained from normal rats were cultured for 48 hr in Dulbecco's modified Eagle's medium (high glucose, 4.5%) supplemented with antibiotics and bovine serum albumin. The experimental cultures contained water-solubilized extracts (10, 25, and 50 μg/ml of medium) of S. horneri. The bone-resorbing factors parathyroid hormone (1-34) (PTH; 10-7 M) and prostaglandin E2 (PGE2; 10-5 M) caused a significant decrease in the diaphyseal and metaphyseal calcium content. The decrease in bone calcium content induced by PTH or PGE2 was completely inhibited by water-solubilized extracts (10, 25, and 50 μg/ml) of S. horneri. In addition, these extracts (25 and 50 μg/ml) completely prevented the PTH (10-7 M)- or PGE2 (10-5 M)-induced increase in medium glucose consumption and lactic acid production by bone tissues. Moreover, S. horneri extracts (10-50 μg/ml) blocked PTH (10-7 M)- increased acid phosphatase activity in the diaphyseal and metaphyseal tissues. These findings indicate that water-solubilized extracts of S. horneri have a direct inhibitory effect on bone resorption in tissue culture in vitro.

Inhibitory effect of menaquinone-7 (vitamin K2) on osteoclast-like cell formation and osteoclastic bone resorption in rat bone tissues in vitro.

The effect of menaquinone-7 (MK-7; vitamin K2) on osteoclast-like cell formation and osteoclastic bone resorption in rat femoral tissues in vitro was investigated. The bone marrow cells were cultured for 7 days in a a-minimal essential medium (alpha-MEM) containing a well-known bone resorbing agent [parathyroid hormone (1-34) (PTH) or prostaglandin E2 (PGE2)] with an effective concentration. Osteoclast-like cells were estimated by staining for tartrate-resistant acid phosphatase (TRACP), a marker enzyme of osteoclasts. The presence of PTH (10(-8) M) or PGE2 (10(-6) M) induced a remarkable increase in osteoclast-like multinucleated cells. These increases were significantly inhibited by MK-7 (10(-8) - 10(-5) M). MK-7 (10(-7) and 10(-6) M) significantly inhibited phorbol 12-myristate 13-acetate-induced osteoclast-like cell formation, whereas MK-7 did not inhibit dibutyryl cyclic adenosine monophosphate (DcAMP) (10(-5) M)-induced osteoclast-like cell formation. These results suggest that the inhibitory action of MK-7 is partly involved in protein kinase C signaling. The bone cells isolated from rat femoral tissues were cultured for 48 h in an alpha-MEM containing either vehicle or MK-7 (10(-8) - 10(-5) M). The presence of MK-7 (10(-6) and 10(-5) M) caused a significant decrease in the number of mature osteoclasts. Such a decrease was also seen in the presence of calcitonin (10(-10) - 10(-8) M), DcAMP (10(-6) and 10(-5) M), or calcium chloride (10(-4) and 10(-3) M). The effect of MK-7 (10(-6) M) in decreasing the number of osteoclasts was not further enhanced in the presence of calcitonin (10(-8) M), DcAMP (10(-5) M), or calcium chloride (10(-3) M), and was completely abolished by the presence of dibucaine (10(-6) M) or staurosporine (10(-7) M), which are inhibitors of Ca2+-dependent protein kinases. These results suggested that MK-7 has a suppressive effect on osteoclasts. Moreover, the femoral-metaphyseal tissues obtained from rats were cultured for 48 h in Dulbecco's modified Eagle's medium containing either vehicle, PTH (10(-7) M), orPGE2 (10(-5) M) in the absence or presence of MK-7 (10(-7) - 10(-5) M). The presence of PTH or PGE2 induced a significant decrease in bone calcium content. These decreases were significantly blocked by MK-7 (10(-7) - 10(-5) M). This study demonstrates that MK-7 has an inhibitory effect on osteoclastic bone resorption in vitro.

Calcitonin and parathyroid hormone: an evening seminar of the 17th Annual Meeting of the Japanese Society for Bone and Mineral Research, July 29, 1999.

and neurodegenerative diseases as a consequence of the paradoxical calcium overload due to age-related calcium deficiency. The main biologic action of calcitonin has resulted from its successful use in disease states characterized by increased bone resorption. Calcitonin is approved by the Food and Drug Administration (FDA) for the treatment of osteoporosis. The main biologic effect of calcitonin is to inhibit osteoclastic bone resorption. This effect is mediated by a calcitonin receptor, which is strongly expressed by osteoclasts. Thyroid C cells, which are neural crest derivatives, are the primary source of calcitonin in mammals. However, other tissue sources of calcitonin have been described, notably the pituitary cells and widely distributed neuroendocrine cells. Since the concentration of calcitonin at its several sites of biosynthesis may be sufficiently high, calcitonin may be expected to have paracrine effects at these sites. The demonstration of calcitonin and its receptors at intracranial sites may qualify calcitonin as a neurotransmitter. Most investigators find that women have lower calcitonin levels than men. The mechanism for this difference may be accounted for in part by a stimulating effect of gonadal steroids on calcitonin secretion. Analgesia is a commonly reported effect of calcitonin treatment in osteoporosis and bone malignant metastasis. In experimental studies using animals, calcitonin has acute antinociceptive effects when administered into the central nervous system. However, the effects of centrally administered calcitonin do not necessarily explain the clinical effects of intramuscular calcium injections. In this symposium, Dr. Yoshimura further focuses on the modulation of pain sensation at the level of the spinal cord in cases where there is an estrogen deficiency. By using a rat model, he initially observed the inducement of hyperalgesia after ovariectomy and the antinociceptive effect after intramuscular injections of elcatonin. He confirmed the involvement of the Parathyroid hormone and calcitonin are the principal regulators of bone and calcium metabolism in humans and most likely all terrestrial vertebrates. In bone, parathyroid hormone stimulates the resorption of bone, releasing calcium and phosphate. In the kidney, this hormone stimulates the reabsorption of calcium and inhibits the reabsorption of phosphate. Parathyroid hormone stimulates the activity of renal 1-αhydroxylase, thereby enhancing synthesis of the active form of vitamin D, which in turn increases intestinal calcium and phosphate. As a result of these parathyroid hormone-dependent actions, blood calcium concentration increases and phosphate concentration decreases. The extracellular calcium concentration is the most important regulator of parathyroid hormone secretion, and it is maintained at a constant level despite variation in the amount of daily calcium intake. In chronic calcium-deficient conditions, however, the homeostatic regulation of extracellular calcium level by parathyroid hormone seems to threaten the skeleton and nonskeletal soft tissues as well. Increased parathyroid hormone secretion readily leads to osteoporosis, which is a decrease in bone calcium content. On the other hand, a variety of cells in the extraskeletal soft tissues are exposed to the constant calcium overflow from bone so their intracellular calcium levels tend to increase. As a consequence, serious disturbance of cell function occurs, giving rise to various diseases. While systemic calcium deficiency is certainly expected to cause skeletal calcium deficiency, the associated increase of calcium within cells of nonskeletal tissue may appear to be paradoxical. In this symposium, Dr. Fujita focuses on this paradoxical calcium overload of the nonskeletal tissues and relates it to calcium deficiency. He attempts to explain the cause of various soft tissue disorders including cardiovascular diseases

Inhibitory effect of genistein on bone resorption in tissue culture

The effect of genistein on bone resorption in vitro was investigated. Femoral-metaphyseal tissues obtained from elderly female rats were cultured for 48 hr in Dulbecco's modified Eagle's medium (high glucose, 4.5%) supplemented with antibiotics and bovine serum albumin. The experimental cultures contained 10 −7 to 10 −3 M genistein. The bone-resorbing factors parathyroid hormone (1–34) (PTH; 10 −7 M), prostaglandin E 2 (PGE 2; 10 −5 M), and lipopolysaccharide (10 μg/mL) caused a significant decrease in bone calcium content. The decrease in bone calcium content induced by bone-resorbing factors was inhibited completely by genistein (10 −7 to 10 −5 M). In addition, this isoflavonoid (10 −5 M) completely inhibited the PTH (10 −7 M)- or PGE 2 (10 −5 M)-induced increase in medium glucose consumption and lactic acid production by bone tissues. Moreover, genistein (10 −5 M) blocked both PTH (10 −7 M)-increased acid phosphatase and -decreased alkaline phosphatase activities of bone tissues. The inhibitory effect of genistein (10 −5 M) on PTH (10 −7 M)-stimulated bone resorption was clearly prevented by the presence of 10 −6 M tamoxifen, an anti-estrogen reagent. Genistein (10 −5 M) did not further enhance the inhibitory effect of estrogen (10 −9 M) on PTH-stimulated bone resorption. These findings indicate that genistein has a direct inhibitory effect on bone resorption in tissue culture in vitro.

Preventive effect of beta-alanyl-L-histidinato zinc on bone metabolism in rats fed on low-calcium and vitamin D-deficient diets.

The effect of beta-alanyl-L-histidinato zinc (AHZ) on bone metabolism in the femoral diaphysis of rats fed on low-calcium and vitamin D-deficient diets was investigated. Rats were orally administered AHZ (10, 30, and 100 mg/kg per day) for 14 days and were killed on the 15th day. Feeding with low-calcium and vitamin D-deficient diets caused a significant decrease in serum 25-hydroxy-vitamin D3, calcium, and inorganic phosphorus concentrations. These decreases were not prevented by AHZ administration. Meanwhile, the femoral-diaphyseal calcium and phosphorus contents were significantly reduced by feeding with the deficient diets. Decrease in bone calcium content was significantly prevented by the doses of 30 and 100 mg AHZ/kg. Furthermore, the dose of 100 mg AHZ/kg produced a significant increase in bone deoxyribonucleic acid (DNA) content and alkaline phosphatase activity in rats fed on the deficient diets. Bone zinc content in the deficient rats was significantly increased by the doses of AHZ (30 and 100 mg/kg). The present results suggest that oral administration of AHZ has a preventive effect in the development of deteriorating bone metabolism in rats fed on low-calcium and vitamin D-deficient diets.

Inhibitory effect of beta-alanyl-L-histidinato zinc on bone resorption in tissue culture.

The inhibitory effect of beta-alanyl-L-histidinato zinc (AHZ) on bone resorption in tissue culture was investigated. Calvaria were removed from weanling rats (3-week-old male) and cultured for periods up to 48 h in Dulbecco's modified Eagle medium (high glucose, 4.5%) supplemented with antibiotics and bovine serum albumin. The experimental cultures contained 10(-7) to 10(-4) mol/l AHZ. The bone-resorbing factors, parathyroid hormone (1-34) (PTH; 10(-7) mol/l), prostaglandin E2 (10(-5) mol/l), interleukin-1 alpha (IL1 alpha; 50 U/ml), and lipopolysaccharide (10 micrograms/ml), caused a significant decrease in bone calcium content. The decreases in bone calcium content induced by bone-resorbing factors were completely inhibited by the coexistence of AHZ (10(-6) to 10(-4) mol/l). Also, AHZ (10(-5) mol/l) completely inhibited the PTH (10(-7) mol/l) or IL1 alpha (50 U/ml)-induced increase in medium glucose consumption and lactic acid production by bone tissue. Furthermore, AHZ (10(-5) mol/l) fairly blocked both PTH (10(-7) mol/l)-increased acid phosphatase and decreased alkaline phosphatase activities of bone tissue. The inhibitory effect of AHZ (10(-5) mol/l) on PTH (10(-7) mol/l)-stimulated bone resorption was clearly prevented by the presence of 10(-4) mol/l dipicolinate, a chelator of zinc. However, zinc sulfate (10(-7) to 10(-4) mol/l) did not inhibit the PTH (10(-7) mol/l)-stimulated bone resorption in tissue culture. These findings indicate that AHZ had a direct inhibitory effect on bone resorption in vitro, and the AHZ effect was found in the chemical form of zinc-chelated dipeptide.

Hypercalciuria Due to Combined Growth Hormone and Calcitriol Therapy in Uremia: Effects of Growth Hormone on Mineral Homeostasis in 75% Nephrectomized Weanling Rats

The administration of growth hormone (GH) in conjunction with calcitriol in uremia may increase urinary calcium and decrease renal phosphate excretion, which could have an adverse effect on the kidney in chronic renal insufficiency. The effect of 40 d of ovine GH, calcitriol, and the combination of GH and calcitriol on mineral excretion was studied in rapidly growing uremic rats. Uremia was produced by 75% nephrectomy, and the animals were fed a diet containing 8% protein with equal quantities of calcium (0.6%) and phosphate (0.6%). The uremic rats treated with ovine GH were significantly longer and heavier than the uremic control rats and the uremic rats treated with calcitriol alone. However, the combination of calcitriol and GH abolished the beneficial effect of GH on growth and increased urinary calcium excretion 4-fold over uremic controls whether expressed as calcium excretion per 100 g body weight, urine calcium to creatinine ratio, or as fractional calcium excretion. Calcitriol therapy alone also significantly increased calcium excretion, but not to the extent that the combination therapy did. This increased urinary calcium excretion in the GH plus calcitriol group was not associated with an increase in calcium and sodium intake, plasma ionized calcium, or urinary sodium excretion. The calcium content of the femurs from all uremic rat groups was significantly lower than that of the sham control rats; however, there was also no further decrease in bone calcium content in the GH plus calcitriol group compared with uremic controls. This indicated that bone was not the source of this excess urinary calcium.(ABSTRACT TRUNCATED AT 250 WORDS)

Bony Demineralization Following Urinary Intestinal Diversion

The effect of urinary intestinal diversion on bone mineral metabolism was investigated in 78 rats. The animals were divided into sham-operated controls and diverted animals. The diverted animals were given either no supplement, sodium bicarbonate or ascorbic acid for an eight month period. Dual photon densitometry and bone mineral content were determined. Urinary intestinal diversion resulted in a minimal systemic acidosis and little alteration in baseline renal function but a significant decrease in bone calcium content. Oral bicarbonate and ascorbic acid administration prevented the demineralization.

Calcium metabolism in relation to chronic ethanol ingestion and estrogen deficiency in rat.

The plasma calcium concentration, urinary calcium excretion, total bone calcium content and the rate of calcium uptake and release by tibias were measured in 6 groups of rats which were kept under various treatment protocols for 8 wk. Ovariectomy led to significant hypocalcemia and a decrease in bone calcium content; the former being reversed towards normal by estrogen treatment. Chronic ethanol ingestion reduced the bone calcium content without apparent effect on plasma calcium in intact rat and further reduced (P less than 0.05) the bone calcium content in ovariectomized rats. The calcium uptake by the bone measured by 45Ca injected ip at zero hour was the same among the 6 groups. When 45Ca was injected ip 17 hours earlier, the disappearance of plasma 45Ca was found to be slower in ovariectomized and alcohol treated groups, indicating a higher rate of calcium movement from bone to plasma. Moreover, there was also an increase in the rate of calcium release by tibias after ovariectomy and chronic alcohol ingestion; the release was slightly greater in the condition of ovariectomy plus alcohol. The present investigation thus, provides evidence that ethanol is likely to have a direct stimulatory effect on bone resorption as well as an indirect effect through disrupting ovarian function during chronic administration.