摘要:

AbstractThe action mechanism of hPTH and hPTHrP‐(1–34) on phosphate uptake in opossum kidney (OK) cells was studied using [Nle8,18Tyr34]hPTH‐(3–34)‐NH2, a potent competitive inhibitor of adenylate cyclase‐coupled PTH receptor. We examined the effects of hPTH‐(1–34), hPTHrP‐(1–34), and hPTH‐(3–34) separately or in combination on the change in renal cyclic AMP production and phosphate uptake in OK cells. Both hPTH‐(1–34) and hPTHrP‐(1–34) stimulated intracellular cyclic AMP production to the same degree at concentrations between 10−10and 10−7M and inhibited phosphate uptake equipotently on a molar basis (27.5 ± 2.0 and 33.2 ± 1.2% inhibition at 10−7M, respectively). Both exogenous addition of (Bu)2cAMP and endogenous stimulation of cAMP by forskolin inhibited phosphate uptake in a dose‐dependent manner. Cyclic AMP production induced by either hPTH‐(1–34) or hPTHrP‐(1–34) was inhibited by both [Nle8,18Tyr34]‐hPTH‐(3–34)‐NH2and [Tyr34]‐hPTH‐(7–34)‐NH2. However, [Nle8,18Tyr34]hPTH‐(3–34)‐NH2and [Tyr34]‐hPTH‐(7–34)‐NH2inhibited hPTH‐induced cAMP production more strongly. The inhibitory action of phosphate uptake by hPTH‐(1–34) and hPTHrP‐(1–34) was prevented in the presence of a 100‐fold greater concentration of [Nle8,18Tyr34]hPTH‐(3–34)‐NH2. The antagonistic action of [Nle8,18Tyr34]hPTH‐(3–34)‐NH2on the inhibition of phosphate uptake induced by hPTH‐(1–34) and hPTHrP‐(1–34) became weaker with time (0–120 minutes), and [Nle8,18Tyr34]hPTH‐(3–34)‐NH2did not antagonize the inhibition of phosphate uptake induced by hPTHrP‐(1–34) at 120 minutes of incubation. Our results indicated that PTHrP inhibits renal phosphate transport, at least in part through an adenylate cyclase‐cAMP‐coupled PTH receptor, but the response to the competitive inhibition of this peptide‐induced cAMP production and inhibition of phosphate uptak

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051002

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY

摘要:

AbstractWe examined the effects of TPA on the high Ca2+‐stimulated accumulation of inositol phosphates in bovine parathyoid cells to determine whether protein kinase C modulates phosphoinositide turnover in a fashion similar to that observed in other cell types stimulated by more classic Ca2+mobilizing hormones. Following exposure of parathyroid cells to TPA (10−6M) for 10 or 30 minutes, there was a time‐ and dose‐dependent inhibition of the accumulation of inositol monophosphate (IP), inositol bisphosphate (IP2), and inositol trisphosphate (IP3) stimulated by 3 mM Ca2+. Half the maximal observed inhibition took place at 1–10 nM TPA, with 50–60% inhibition of high Ca2+‐stimulated accumulation of inositol phosphates at 10−6M TPA. The active phorbol ester, 4β‐phorbol didecanoate, produced similar effects; the inactive derivative, 4α‐phorbol didecanoate, was without effect. When parathyroid cells were exposed to TPA (10−6M) for varying times and were then incubated with high (3 mM) Ca2+, inhibition of inositol phosphate accumulation was observed with 10 or 30 minutes preincubation. In contrast, preincubation of cells with TPA for 3 or 18 h markedly enhanced the high (3 mM) Ca2+‐induced increase in inositol phosphates. In cells preincubated with TPA for 18 h, binding sites for [3H]phorbol dibutyrate and total protein kinase C (PKC) activity were reduced by greater than 95% and by 71%, respectively, consistent with downregulation of the enzyme. These results suggest that the high extracellular Ca2+‐stimulated increase in accumulation of inositol phosphates in parathyroid cells, which has been postulated to result from a receptorlike process, can be modulated by agonists of protein kinase C in a fashion similar to that observed with more classic Ca2+mobilizing hormones. Activators of kinase C initially inhibit the generation of inositol phosphates, presumably as a result of reduced turnover of phosphoinositides, but subsequently enhance inositol phosphate accumulation, probably because of down‐r

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051003

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY

摘要:

AbstractHuman periodontal ligament (PDL) cells were derived from healthy premolars extracted for orthodontic treatment and were utilized for in vitro experiments in passages 4–6. Human PDL cells were seeded in tissue culture tubes and incubated with interleukin‐1α (Il‐1α), IL‐1β, tumor necrosis factor‐α (TNF‐α), interferon‐γ (IFN‐γ), indomethacin, parathyroid hormone (PTH), or their combinations, for 1 h. The medium was then replaced with serum‐free BGJbmedium and incubated for 24 h without further additions. Prostaglandin E (PGE) concentrations in the conditioned media (CM) were measured by radioimmunoassay, and bone‐resorbing activity was measured using45Ca‐labeled neonatal mouse calvariae. The results of this study indicated that (1) unstimulated cultured PDL cells produced PGE, and PDL CM stimulated bone resorption; (2) cytokine‐treated (IL‐1α, IL‐1β, and TNF‐α) PDL cells had increased production of PGE and bone‐resorbing activity compared to unstimulated PDL cells; (3) indomethacin completely inhibited PGE production from unstimulated PDL cells but only partially inhibited bone‐resorbing activity, indicating that PDL cells produced nonprostaglandin bone‐resorbing factor(s); (4) IFN‐γ did not change PGE or bone‐resorbing activity production by cytokine‐stimulated PDL cells; and (5) PTH treatment of PDL cells in addition to cytokines (IL‐1α, IL‐1β, and TNF‐α) had additive effects on the production of bone‐resorbing activity and synergistic

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051004

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY

摘要:

AbstractBisphosphonates (BP) are powerful inhibitors of bone resorption. We have previously shown that 4‐amino‐1‐hydroxybutylidene‐1,1‐bisphosphonate (AHBuBP), 3‐amino‐1‐hydroxypropylidene‐1,1‐bisphosphonate (AHPrBP), and dichloromethylenebisphosphonate (Cl2MBP) inhibit the proliferation of macrophages in vitro at concentrations that do not affect the viability of nonproliferating cells. In this study we further investigated whether the antiproliferative effect of these three BP is, among the hematopoietic series, preferential to the mononuclear phagocyte lineage. BP were unable to inhibit more than 30–40% of the [3H]thymidine (3H‐TdR) incorporation into bone marrow cells stimulated to proliferate by multilineage colony‐stimulating activity containing conditioned medium (multi‐CSA). From the analysis of the colonies induced in semisolid medium by multi‐CSA and recombinant murine granulocyte‐macrophage colony stimulating factor (rmGM‐CSF), a dose‐dependent disappearance specific to the macrophage‐containing colonies emerged. In contrast, the number and composition of colonies other than macrophage and, in particular, the granulocyte colonies were not affected by these compounds, even at high concentrations (100μM) previously also shown to be toxic for nonproliferating macrophages. Since the macrophages, differently from polymorphonuclear phagocytes, are known to be highly pinocytotic, it is possible that by this means they selectively concentrate BP intracellularly, leading to toxic concentrations. We postulate tht BP may also act in vivo in addition to their effect on osteoclast activity, by a similar mechanism on osteoclast precursors and on bone resident macrophages, a source of cytokines stimulating bone resorption and leading to impaired osteoclast recruitment and activity. Furthermore, their specific effect on the mononuclear phagocyte lineage may also be relevant in other pathologic situat

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051005

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY

摘要:

AbstractNonsteroidal anti‐inflammatory drugs (NSAIDs) affect bone metabolism in vitro and in vivo. They delay but do not alter the outcome of healing processes in bone. In some bone loss models, they block bone resorption and slow the rate of loss. We studied the effect of naproxen, a potent NSAID, on cancellous bone of the proximal tibial metaphysis of 6‐month‐old adult female ovariectomized rats.Animals were ovariectomized, divided into groups, and fed standard diets differing only in naproxen content for 42 days. The rats of the groups ate 2.0, 5.5, 12.7, and 32 mg naproxen per kg body weight per day, respectively. Serum levels of naproxen were determined. Bone volume, mineralizing surface, osteoblast activity, osteoclast surface, and bone resorption rate were determined by bone histomorphometric techniques.The rats' dose‐related serum naproxen levels ranged from 4 to 28 μg/ml. Naproxen inhibited up to 70% of the bone loss occurring after ovariectomy at a serum level of 4 μg/ml. We deduced that naproxen blocked bone resorption in ovariectomized rats by slowing osteoclast activity at all doses. In contrast, naproxen slowed bone formation only at serum levels>20 μg/ml in ovariectomized rats. These findings may have clinical relevance in helping to prevent postmenopausal bone los

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051006

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY

摘要:

AbstractThe enzyme carbonic anhydrase has been suggested as a critical participant in osteoclast‐mediated bone resorption. In humoral hypercalcemia of malignancy (HHM) intense osteoclastic bone resoprtion is principally responsible for the observed hypercalcemia. We therefore undertook to examine the effect of the carbonic anhydrase inhibitor acetazolamide on the hypercalcemia induced by the H500 Leydig cell tumor in Fisher rats, a well‐described model of HHM. Acetazolamide treatment for 10 h at 10 mg/h resulted in a significant fall in serum calcium in the five drug‐treated animals (14.2 ± 0.9 to 11.5 ± 0.1 mg/dl,p<0.05). Conversely, the six animals infused with vehicle alone showed a significant rise in serum calcium (12.5 ± 0.5 to 13.8 ± 0.1 mg/dl,p<0.05). At the end of the infusion, the acetazolamide‐treated animals had a significantly lower mean serum calcium than those receiving vehicle alone (11.5 ± 0.1 versus 13.8 ± 0.1,p<0.05). There was no significant change in serum phosphorus, urine calcium, urine phosphorus, or nephrogenous cyclic AMP excretion between the two groups. Acetazaolamide and HTS 5‐(3‐hydroxybenzoyl)‐2‐thiophenesulfonamide, another carbonic anhydrase inhibitor, both significantly inhibited in vitro bone resorption induced by 5 × 10−9M36Tyr(1–36)‐PTHrP‐amide (PTHrP, parathyroid hormone‐related protein). Acetazolamide also inhibited the resorption induced by 10−8M (1–141)‐PTHrP and 2.5 × 10−9M (1–74)‐PTHrP. We conclude that acetazolamide is effective in lowering the serum calcium in animals with humoral hypercalcemia of malignancy. The data are consistent with the hypothesis that the mechanism of action for this effect is direct inhi

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051007

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY

摘要:

AbstractThe majority of in vivo competitive binding of parathyroid hormone (PTH) in the endosteal metaphysis of rat long bones was recently shown to be localized in the intertrabecular tissue to a cell that is distinct from a differentiated osteoblast. In the present report we have further characterized this cell, termed a parathyroid hormone target (PT) cell, by light and electron microscopy using radioautography and histochemical techniques. These studies demonstrate that the PT cell is a mononuclear cell with a large cell body located at times between clusters of differentiated osteoblasts, as well as in other regions of the intertrabecular tissue. Its long cytoplasmic processes extend from the bone matrix through the intertrabecular region toward vascular structures, interdigitating with various cells of the endosteum. A distinctive tubular structure originating in the Golgi system and often associated with long mitochondria and glycogen particles extends throughout the cytoplasmic processes of the PT cell. Based on its capacity to incorporate [3H]thymidine, the PT cell appears to divide rather slowly. The identification of occasional hybrid cells with ultrastructural features of both the PT cell and the differentiated osteoblast and the presence of histochemical evidence for alkaline phosphatase activity suggest that the PT cell is of the osteoblast lineage. These studies therefore morphologically define a major osseous target cell for PTH that, although of the osteoblast lineage, is not a differentiated osteoblast and provide in vivo evidence that characteristics of the “osteoblast phenotype” are not restricted to a sole osseous cell t

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051008

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY

摘要:

AbstractOsteoclasts are effector cells in bone breakdown, and the active bone resorption is confined to the ruffled border zone of these cells. An acid milieu is maintained in this zone which is probably a prerequisite for bone resorption. Tartrate‐resistant acid phosphatase (TRAP) activity has been recognized as a characteristic property of osteoclasts and in several studies proposed as a cytochemical marker of osteoclasts. We have previously isolated and characterized a tartrate‐resistant and iron‐activated acid ATPase (TrATPase) from rat bone, the enzyme being a member of the TRAP family. In the present study the ultrastructural localization of this enzyme was delineated by employing immunogold technique on low temperature‐embedded maxillar rat bone. Intensive immunolabeling was seen on the bone surfaces facing the ruffled border zone while lower amounts of marker were seen in adjacent bone areas, that is, on the bone surfaces facing the clear zone and deeper into the bone. Within the osteoclasts gold markers were observed mainly in vesicular structures interpreted as lysosomes. Immunolabeling was also observed in the recently described endocytic cells located near osteoblasts and osteoclasts. Also in these cells, the marker was confined to lysosomelike structures. The amount of label in bone facing osteoblasts was low, as was the amount within osteoblasts. Our observation of extracellular localization, in particular accumulation of TrATPase in bone matrix facing the ruffled border area of the osteoclasts, favors the view that the enzyme is exported to areas of active bone resorption, thereby indicating a potential role for the enzyme in this

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051009

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY

摘要:

AbstractThe frequency distributions of mineral apposition rate (MAR) and mineralizing surface (MS), measured separately on the intracortical, endocortical, and cancellous surfaces in 46 normal subjects and 79 patients with postmenopausal osteoporosis, indicated that MAR has a finite lower limit of 0.3 μ/day (uncorrected for section obliquity) but that MS has no finite lower limit. We conclude that in the absence of labels MAR, and indices derived from it, must be treated as missing values, but that MS and indices with MS in the numerator should be allowed to take values of zero. To avoid infinite values for indices with MS in the denominator, we propose that osteoid mineralization rate (the reciprocal of mineralization lag time) and osteoblast vigor (the reciprocal of formation period) be used instead. For surfaces with genuine single labels (SL) but no double labels, we propose that MS is calculated as SL/2 and that for MAR either the lower limit of 0.3 or the mean measured value from other surfaces be used for calculating derived indices

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051010

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY

摘要:

AbstractThe remodeling response of bone tissue to disuse in four normal adult male turkeys and four adult males metabolically altered by castration was compared by functionally isolating the left ulna of each animal via transverse epiphyseal osteotomies. The right ulna in each animal was left intact and served as a control. After 8 weeks, the animals were euthanized, the ulnae harvested, and 100 μm undecalcified cross sections of the midshaft microradiographed. Areal properties, osteon mineral apposition rates from in vivo fluorochrome labels, and the number and ratios of bone‐forming and bone‐resorbing foci were quantitated. Compared to their control ulnae, the magnitude of bone resorbed from the functionally isolated ulnae of normal versus castrated males was not significantly different (‐12.8 ± 3.7 versus −10.7 ± 3.5%, respectively). However, in the functionally isolated ulnae of normal birds, 94% of the total bone loss resulted from expansion of the corticoendosteal envelope, and 97% of the decrease in cross‐sectional areas of the ulnae in the castrated birds was due to intracortical porosity. Furthermore, there was a significant interaction between disuse and castration, increasing the total number of intracortical remodeling events (9.4 ± 0.9) when compared to disuse alone (4.7 ± 1.4,p<0.01), or to the intact ulnae of castrated (2.1 ± 0.5) and normal adult males (2.0 ± 1.1). This work emphasizes that the manner in which the bone tissue responds to local changes in its physical environment is directly dependent on the status of the organism's

ISSN:0884-0431    

DOI:10.1002/jbmr.5650051011

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1990

数据来源: WILEY