N. Partridge - Academia.edu (original) (raw)

Papers by N. Partridge

Journal of Biological Chemistry, 2007

The clinical findings that alendronate blunted the anabolic effect of human parathyroid hormone (... more The clinical findings that alendronate blunted the anabolic effect of human parathyroid hormone (PTH) on bone formation suggest that active resorption is involved and enhances the anabolic effect. PTH signals via its receptor on the osteoblast membrane, and osteoclasts are impacted indirectly via the products of osteoblasts. Microarray with RNA from rats injected with human PTH or vehicle showed a strong association between the stimulation of monocyte chemoattractant protein-1 (MCP-1) and the anabolic effects of PTH. PTH rapidly and dramatically stimulated MCP-1 mRNA in the femora of rats receiving daily injections of PTH or in primary osteoblastic and UMR 106-01 cells. The stimulation of MCP-1 mRNA was dose-dependent and a primary response to PTH signaling via the cAMP-dependent protein kinase pathway in vitro. Studies with the mouse monocyte cell line RAW 264.7 and mouse bone marrow proved that osteoblastic MCP-1 can potently recruit osteoclast monocyte precursors and facilitate receptor activator of NF-B ligand-induced osteoclastogenesis and, in particular, enhanced fusion. Our model suggests that PTH-induced osteoblastic expression of MCP-1 is involved in recruitment and differentiation at the stage of multinucleation of osteoclast precursors. This information provides a rationale for increased osteoclast activity in the anabolic effects of PTH in addition to receptor activator of NF-B ligand stimulation to initiate greater bone remodeling.

Metabolic Bone Disease and Related Research

Prostaglandins are potent bone resorblng agents, the most potent of those tested being prostaglan... more Prostaglandins are potent bone resorblng agents, the most potent of those tested being prostaglandin E=. The labile prostsnold, prostacylin, is also capable Of resorbing bone, but its potency is not known. The responsiveness of rat bone and bone turnout cell adenylate cyclaso has been used to assess the relative efficaclas of a wide range of stable and labile prosteglandins, their metabolltes and analogues upon bone. Prostaglandins may be Important local regulators in bone, and it will be critical, therefore, to define the precise action of prostaglandin-Iike molecules on bone cells. There is ample evidence for a role for prostaglandins in the pathogenesis of malignant hypercalcaemla in a number of animei tumour models. Evidence is also accumulating in human cancers, suggesting that prostaglandin production may be Important in the establishment of osseous metastases, and in generating excessive resorption adjacent to metastatic deposits in bone, particularly in breast and renal cor-Aical carcinomas. The nature of the rasponsible prostaglandin-Iike molecules is undetermined, and this needs to be invastigated by defining the pathways of arachidonic acid matabolism in tumours. When this has been achieved, drug therapy can be directed at the appropriate enzyme activlUas. The current application of cyclo~xygenase Inhibitors in these clinical syndromes is of tittle benefit.

Journal of Cellular Physiology, 2016

Mesenchymal stem cells (MSCs) are multipotent cells and their differentiation into the osteoblast... more Mesenchymal stem cells (MSCs) are multipotent cells and their differentiation into the osteoblastic lineage is strictly controlled by several regulators, including microRNAs (miRNAs). Runx2 is a bone transcription factor required for osteoblast differentiation. Here, we used in silico analysis to identify a number of miRNAs that putatively target Runx2 and its co-factors to mediate both positive and negative regulation of osteoblast differentiation. Among these miRNAs, miR-590-5p was selected and its expression was found to be increased during osteoblast differentiation. When mouse MSCs (mMSCs) were transiently transfected with a miR-590-5p mimic, we detected an increase in both calcium deposition and the mRNA expression of osteoblast differentiation marker genes such as alkaline phosphatase (ALP) and type I collagen genes. Smad7 was found to be among the putative target genes of miR-590-5p and its mRNA and protein expression decreased after miR-590-5p mimic transfection in human osteoblast-like cells (MG63). Our analysis indicated that Runx2 was not a putative target of miR-590-5p. However, Runx2 protein, but not mRNA expression, increased after miR-590-5p mimic transfection in MG63 cells. Runx2 protein expression was increased with knockdown of Smad7 expression by Smad7 siRNA in these cells. We further identified that the 3'-untranslated region of Smad7 was directly targeted by miR-590-5p; this was done using the luciferase reporter gene system. It is known that Smad7 inhibits osteoblast differentiation via Smurf2-mediated Runx2 degradation. Hence, based on our results, we suggest that miR-590-5p promotes osteoblast differentiation by indirectly protecting and stabilizing the Runx2 protein by targeting Smad7 gene expression. J. Cell. Physiol. 232: 371-380, 2017. © 2016 Wiley Periodicals, Inc.

Biochemical Society Transactions, 2000

FASEB journal : official publication of the Federation of American Societies for Experimental Biology, Jan 19, 2015

Bone minerals are acquired during growth and are key determinants of adult skeletal health. Durin... more Bone minerals are acquired during growth and are key determinants of adult skeletal health. During puberty, the serum levels of growth hormone (GH) and its downstream effector IGF-1 increase and play critical roles in bone acquisition. The goal of the current study was to determine how bone cells integrate signals from the GH/IGF-1 to enhance skeletal mineralization and strength during pubertal growth. Osteocytes, the most abundant bone cells, were shown to orchestrate bone modeling during growth. We used dentin matrix protein (Dmp)-1-mediated Ghr gene deletion in mice (DMP-GHRKO) to address the role of the GH/IGF axis in osteocytes. We found that DMP-GHRKO did not affect linear growth but compromised overall bone accrual. DMP-GHRKO mice exhibited reduced serum inorganic phosphate (Pi) and parathyroid hormone (PTH) levels and decreased bone formation indices and were associated with an impaired response to intermittent PTH treatment. Using an osteocyte-like cell line along with in v...

Molecular cell biology research communications : MCBRC, 2000

Matrix metalloproteinases (MMPs) are a family of secreted or transmembrane proteins that have bee... more Matrix metalloproteinases (MMPs) are a family of secreted or transmembrane proteins that have been implicated in multiple physiological and pathological processes related to extracellular matrix turnover. Recent evidence strongly suggests a role for collagenase-3 (MMP-13) in tumor metastasis and invasion. We report here that collagenase-3 is constitutively expressed in the breast cancer cell line MDA-MB231 (MDA) and outline the molecular mechanism regulating its expression. Functional analysis of the collagenase-3 promoter showed that both the activator protein-1 (AP-1) site and the runt domain (RD) binding site were required for maximal constitutive expression of collagenase-3 in MDA cells. Determination of factors binding to those sites by Northern analysis and transient transfections identified the requirement of Fra-1, c-Jun, and Cbfa1 for basal collagenase-3 promoter activity in MDA cells.

The Journal of biological chemistry, Jan 25, 1982

The pattern of cyclic AMP-dependent protein kinase isoenzyme response to acute hormonal activatio... more The pattern of cyclic AMP-dependent protein kinase isoenzyme response to acute hormonal activation has been studied in cultured cells derived from rat osteogenic sarcoma and osteoblast-rich cells grown from newborn rat calvaria. Using multiple small anion exchange columns and a batch elution technique, a rapid method of separating the isoenzymes of cyclic AMP-dependent protein kinase was developed and the acute activation by parathyroid hormone and prostaglandin E2 of each isoenzyme was studied. Activation was rapid, being detectable at 5 s, maximal at 15-30 s, and persisting for up to 6 h. Both hormones showed a dose-dependent activation of each isoenzyme in both cell types, but the patterns of response differed. Parathyroid hormone predominantly stimulated isoenzyme I in the clonal osteogenic sarcoma cells but showed equivalent activation of each isoenzyme in calvarial cells. Prostaglandin E2 also predominantly stimulated isoenzyme I in the malignant cells, whereas in the calvaria...

Methods in enzymology, 1987

The methods for establishing osteoblast-rich rat calvarial cell cultures have been described, tog... more The methods for establishing osteoblast-rich rat calvarial cell cultures have been described, together with methods for the use of clonal osteogenic sarcoma cells of osteoblast phenotype. The latter clonal lines are useful for several purposes, but all the precautions and quality control measures necessary in the study of clonal lines must be observed. Some of the techniques for studying biochemical responses to hormones in these cells have also been detailed, but clearly others are applicable, including studies of the synthesis of matrix constituents. Osteoclast-like cells have not been considered in this chapter, because osteoclast culture methods have not yet been developed to the degree of purity and reproducibility necessary for this type of biochemical approach.

The Journal of biological chemistry, Jan 25, 1990

We have isolated clones for rat collagenase from a rat osteoblastic cell cDNA library. These clon... more We have isolated clones for rat collagenase from a rat osteoblastic cell cDNA library. These clones have been sequenced and the amino acids deduced. The calculated molecular weight is 51,352 for the proenzyme and 42,229 for the active enzyme. The deduced amino acid sequence was compared to those previously reported for: 1) human collagenase, 2) rat transin 1 (stromelysin), 3) human stromelysin, and 4) rabbit collagenase. The number of amino acids conserved was 47, 47, 50, and 47%, respectively. We also compared the collagenase mRNA and protein in different rat cells (osteoblast, uterine smooth muscle, synovial fibroblast) and determined that in rat uterine cells the message is slightly larger, although collagenase protein in all three cell types was identical in size. Parathyroid hormone dramatically induces the 2.9-kilobase collagenase mRNA in the rat osteoblastic cells, UMR 106-01. Nuclear run-on studies in UMR 106-01 cells demonstrated a 4-8-fold induction in the rate of synthesi...

Prostaglandins, 1985

SEPTEMBER1985VOL.30NO. 3 527 PROSTAGLANDINS predominant effect of the growth factor was an inhibi... more SEPTEMBER1985VOL.30NO. 3 527 PROSTAGLANDINS predominant effect of the growth factor was an inhibition of both PGE and PGI production by the osteoblastic cells. The'present gesults suggest that the boneresorbing hormones do not act to cause an increase in PG by the osteoblast and that any increase in PG production by these cells may be in response to vascular agents.

Principles of Bone Biology, 2002

Osteoporosis International, 2008

Osteoarthritis and Cartilage, 2006

Objective: Matrix metalloproteinase-13 (MMP-13) is an extracellular MMP that cleaves type II coll... more Objective: Matrix metalloproteinase-13 (MMP-13) is an extracellular MMP that cleaves type II collagen, the major protein component of cartilage, with high specificity and has been implicated in the pathology of osteoarthritis. The present study aimed to characterize the binding and internalization kinetics of MMP-13 in normal rabbit chondrocytes and whether MMP-13 affected cell signalling. Methods: Rabbit chondrocytes were used in [ 125 I]-MMP-13 binding assays to investigate the MMP-13 binding kinetics and Western analysis allowed for the assessment of intracellular signalling cascades. Results: Rabbit chondrocytes were found to express the cartilage-specific genes aggrecan and type II collagen throughout their in vitro culture period. Appreciable specific cell-association of [ 125 I]-MMP-13 was detected after 10 min of exposure to the ligand and equilibrium was obtained after 2 h. Binding of [ 125 I]-MMP-13 to chondrocytes was specific and approached saturation at 75 nM. Internalization of MMP-13 was evident after 20 min, reached a maximum at 30 min and had returned to baseline by 90 min. Addition of receptor-associated protein (RAP) inhibited the internalization of MMP-13 indicating a likely role for low-density lipoprotein receptor-related protein-1 (LRP1) in this process. Interestingly the presence of MMP-13 induced phosphorylation of the extracellular signal-regulated kinase 1/2 (ERK1/2) protein showing that there is initiation of a signalling process in response to MMP-13 being bound and internalized by rabbit chondrocytes. However, this activation does not involve the MMP-13 internalization receptor LRP1. Conclusion: These studies demonstrate and characterize the MMP-13 binding and internalization system in rabbit chondrocytes and indicate that MMP-13 may regulate the phenotype of the chondrocytes through this receptor system.

Molecular Endocrinology, 1992

Collagenase is synthesized and secreted by rat osteoblastic cells in response to PTH. We have pre... more Collagenase is synthesized and secreted by rat osteoblastic cells in response to PTH. We have previously demonstrated that this effect involves a substantial increase in collagenase mRNA via transcription. Northern blots and nuclear run-on assays were performed to further investigate the induction of collagenase by PTH in the rat osteoblastic cell line UMR 106-01. Detectable amounts of collagenase mRNA were not apparent until 2 h of PTH treatment, showed the greatest abundance at 4 h, and declined to approximately 30% of maximum by 8 h. The changes in the rate of transcription of the collagenase gene in response to PTH paralleled and preceded the changes in the steady state mRNA levels. After an initial lag period of about 1 h, collagenase transcription rates increased from very low levels to a maximal response at 2 h, returning to about 50% of maximum by 10 h. The increased transcriptional rate of the collagenase gene was found to be dependent on the concentration of PTH, with a half-maximal response at approximately 7 x 10(-10) M rat PTH-(1-34) and a maximal effect with a dose of 10(-8) M. The PTH-mediated induction of collagenase transcriptional activity was completely abolished by cycloheximide, while transcription of the beta-actin gene was unaffected by the translation inhibitor. These data suggest that a protein factor(s) is required for PTH-mediated transcriptional induction of collagenase. Since PTH increases intracellular levels of several potential second messengers, agents that mimic these substances were employed to determine which signal transduction pathway is predominant in the PTH-mediated stimulation of collagenase transcription.(ABSTRACT TRUNCATED AT 250 WORDS)

Molecular Endocrinology, 1992

We thank Katja Kopp and Deborah K. Clifton for their technical assistance. 13. McGrath JP, Capon ... more We thank Katja Kopp and Deborah K. Clifton for their technical assistance. 13. McGrath JP, Capon DJ, Goeddel DV, Levinson AD 1984 Comparative biochemical properties of normal and activated human ras ~21 orotein. Nature 310:644-649

Journal of Dental Research, 2012

Bisphosphonates are therapeutic agents in the treatment of post-menopausal osteoporosis. Although... more Bisphosphonates are therapeutic agents in the treatment of post-menopausal osteoporosis. Although they have been associated with delayed healing in injured tissues, inappropriate femoral fractures, and osteonecrosis of the jaw (ONJ), the pathophysiological mechanisms involved are not clear. Our hypothesis is that alendronate, a member of the N-containing bisphosphonates, indirectly inhibits osteoblast function through the coupling of osteoclasts to osteoblasts by ephrinB-EphB interaction. We found that alendronate increased gene and protein expression of ephrinB1 and EphB1, as well as B3, in femurs of adult mice injected with alendronate (10 µg/100 g/wk) for 8 weeks. Alendronate suppressed the expression of bone sialoprotein (BSP) and osteonectin in both femurs and bone marrow osteoblastic cells of mice. After elimination of pre-osteoclasts from bone marrow cells, alendronate did not affect osteoblast differentiation, indicating the need for pre-osteoclasts for alendronate’s effects...

The Journal of Clinical Endocrinology & Metabolism, 2011

Chronic high levels of PTH may be associated with up-regulation of proteases and cytokines. Monoc... more Chronic high levels of PTH may be associated with up-regulation of proteases and cytokines. Monocyte chemoattractant protein-1 (MCP-1) is an inflammatory cytokine, produced predominantly by macrophages and endothelial cells, and is expressed in adipose tissue. More recently it has been shown that PTH administration increases MCP-1 expression in osteoblasts. Because both PTH and MCP-1 levels are higher in obesity, the goal was to determine whether the high MCP-1 occurs only in the presence of high serum PTH and is independent of adiposity and examine its relationship with bone mineral density (BMD) and turnover. In this case-control clinical design, 111 eligible women were categorized into four groups: leaner women [body mass index (BMI) 23 ± 2 kg/m(2)] with normal or higher PTH and obese (BMI 44 ± 7 kg/m(2)) with normal or higher PTH. Serum MCP-1 levels were higher (P < 0.01) in the high (PTH = 74.9 ± 27.0 pg/ml, MCP-1 = 421.5 ± 157.0 pg/ml) compared with normal PTH (PTH = 32.5 ± 10.4 pg/ml, MCP-1 = 322.5 ± 97.8 pg/ml) group, independent of BMI. C-reactive protein and adiponectin were influenced only by BMI and not PTH. MCP-1 was positively associated with osteocalcin and propeptide of type 1 collagen in the leaner (r > 0.3, P < 0.05) but not the obese women and was not associated with BMD in either group. Together these data suggest that MCP-1 is higher only in the presence of increased PTH and that adiposity alone cannot explain the higher MCP-1 levels in obesity.

Journal of Biological Chemistry, 2007

The clinical findings that alendronate blunted the anabolic effect of human parathyroid hormone (... more The clinical findings that alendronate blunted the anabolic effect of human parathyroid hormone (PTH) on bone formation suggest that active resorption is involved and enhances the anabolic effect. PTH signals via its receptor on the osteoblast membrane, and osteoclasts are impacted indirectly via the products of osteoblasts. Microarray with RNA from rats injected with human PTH or vehicle showed a strong association between the stimulation of monocyte chemoattractant protein-1 (MCP-1) and the anabolic effects of PTH. PTH rapidly and dramatically stimulated MCP-1 mRNA in the femora of rats receiving daily injections of PTH or in primary osteoblastic and UMR 106-01 cells. The stimulation of MCP-1 mRNA was dose-dependent and a primary response to PTH signaling via the cAMP-dependent protein kinase pathway in vitro. Studies with the mouse monocyte cell line RAW 264.7 and mouse bone marrow proved that osteoblastic MCP-1 can potently recruit osteoclast monocyte precursors and facilitate receptor activator of NF-B ligand-induced osteoclastogenesis and, in particular, enhanced fusion. Our model suggests that PTH-induced osteoblastic expression of MCP-1 is involved in recruitment and differentiation at the stage of multinucleation of osteoclast precursors. This information provides a rationale for increased osteoclast activity in the anabolic effects of PTH in addition to receptor activator of NF-B ligand stimulation to initiate greater bone remodeling.

Metabolic Bone Disease and Related Research

Prostaglandins are potent bone resorblng agents, the most potent of those tested being prostaglan... more Prostaglandins are potent bone resorblng agents, the most potent of those tested being prostaglandin E=. The labile prostsnold, prostacylin, is also capable Of resorbing bone, but its potency is not known. The responsiveness of rat bone and bone turnout cell adenylate cyclaso has been used to assess the relative efficaclas of a wide range of stable and labile prosteglandins, their metabolltes and analogues upon bone. Prostaglandins may be Important local regulators in bone, and it will be critical, therefore, to define the precise action of prostaglandin-Iike molecules on bone cells. There is ample evidence for a role for prostaglandins in the pathogenesis of malignant hypercalcaemla in a number of animei tumour models. Evidence is also accumulating in human cancers, suggesting that prostaglandin production may be Important in the establishment of osseous metastases, and in generating excessive resorption adjacent to metastatic deposits in bone, particularly in breast and renal cor-Aical carcinomas. The nature of the rasponsible prostaglandin-Iike molecules is undetermined, and this needs to be invastigated by defining the pathways of arachidonic acid matabolism in tumours. When this has been achieved, drug therapy can be directed at the appropriate enzyme activlUas. The current application of cyclo~xygenase Inhibitors in these clinical syndromes is of tittle benefit.

Journal of Cellular Physiology, 2016

Mesenchymal stem cells (MSCs) are multipotent cells and their differentiation into the osteoblast... more Mesenchymal stem cells (MSCs) are multipotent cells and their differentiation into the osteoblastic lineage is strictly controlled by several regulators, including microRNAs (miRNAs). Runx2 is a bone transcription factor required for osteoblast differentiation. Here, we used in silico analysis to identify a number of miRNAs that putatively target Runx2 and its co-factors to mediate both positive and negative regulation of osteoblast differentiation. Among these miRNAs, miR-590-5p was selected and its expression was found to be increased during osteoblast differentiation. When mouse MSCs (mMSCs) were transiently transfected with a miR-590-5p mimic, we detected an increase in both calcium deposition and the mRNA expression of osteoblast differentiation marker genes such as alkaline phosphatase (ALP) and type I collagen genes. Smad7 was found to be among the putative target genes of miR-590-5p and its mRNA and protein expression decreased after miR-590-5p mimic transfection in human osteoblast-like cells (MG63). Our analysis indicated that Runx2 was not a putative target of miR-590-5p. However, Runx2 protein, but not mRNA expression, increased after miR-590-5p mimic transfection in MG63 cells. Runx2 protein expression was increased with knockdown of Smad7 expression by Smad7 siRNA in these cells. We further identified that the 3'-untranslated region of Smad7 was directly targeted by miR-590-5p; this was done using the luciferase reporter gene system. It is known that Smad7 inhibits osteoblast differentiation via Smurf2-mediated Runx2 degradation. Hence, based on our results, we suggest that miR-590-5p promotes osteoblast differentiation by indirectly protecting and stabilizing the Runx2 protein by targeting Smad7 gene expression. J. Cell. Physiol. 232: 371-380, 2017. © 2016 Wiley Periodicals, Inc.

Biochemical Society Transactions, 2000

FASEB journal : official publication of the Federation of American Societies for Experimental Biology, Jan 19, 2015

Bone minerals are acquired during growth and are key determinants of adult skeletal health. Durin... more Bone minerals are acquired during growth and are key determinants of adult skeletal health. During puberty, the serum levels of growth hormone (GH) and its downstream effector IGF-1 increase and play critical roles in bone acquisition. The goal of the current study was to determine how bone cells integrate signals from the GH/IGF-1 to enhance skeletal mineralization and strength during pubertal growth. Osteocytes, the most abundant bone cells, were shown to orchestrate bone modeling during growth. We used dentin matrix protein (Dmp)-1-mediated Ghr gene deletion in mice (DMP-GHRKO) to address the role of the GH/IGF axis in osteocytes. We found that DMP-GHRKO did not affect linear growth but compromised overall bone accrual. DMP-GHRKO mice exhibited reduced serum inorganic phosphate (Pi) and parathyroid hormone (PTH) levels and decreased bone formation indices and were associated with an impaired response to intermittent PTH treatment. Using an osteocyte-like cell line along with in v...

Molecular cell biology research communications : MCBRC, 2000

Matrix metalloproteinases (MMPs) are a family of secreted or transmembrane proteins that have bee... more Matrix metalloproteinases (MMPs) are a family of secreted or transmembrane proteins that have been implicated in multiple physiological and pathological processes related to extracellular matrix turnover. Recent evidence strongly suggests a role for collagenase-3 (MMP-13) in tumor metastasis and invasion. We report here that collagenase-3 is constitutively expressed in the breast cancer cell line MDA-MB231 (MDA) and outline the molecular mechanism regulating its expression. Functional analysis of the collagenase-3 promoter showed that both the activator protein-1 (AP-1) site and the runt domain (RD) binding site were required for maximal constitutive expression of collagenase-3 in MDA cells. Determination of factors binding to those sites by Northern analysis and transient transfections identified the requirement of Fra-1, c-Jun, and Cbfa1 for basal collagenase-3 promoter activity in MDA cells.

The Journal of biological chemistry, Jan 25, 1982

The pattern of cyclic AMP-dependent protein kinase isoenzyme response to acute hormonal activatio... more The pattern of cyclic AMP-dependent protein kinase isoenzyme response to acute hormonal activation has been studied in cultured cells derived from rat osteogenic sarcoma and osteoblast-rich cells grown from newborn rat calvaria. Using multiple small anion exchange columns and a batch elution technique, a rapid method of separating the isoenzymes of cyclic AMP-dependent protein kinase was developed and the acute activation by parathyroid hormone and prostaglandin E2 of each isoenzyme was studied. Activation was rapid, being detectable at 5 s, maximal at 15-30 s, and persisting for up to 6 h. Both hormones showed a dose-dependent activation of each isoenzyme in both cell types, but the patterns of response differed. Parathyroid hormone predominantly stimulated isoenzyme I in the clonal osteogenic sarcoma cells but showed equivalent activation of each isoenzyme in calvarial cells. Prostaglandin E2 also predominantly stimulated isoenzyme I in the malignant cells, whereas in the calvaria...

Methods in enzymology, 1987

The methods for establishing osteoblast-rich rat calvarial cell cultures have been described, tog... more The methods for establishing osteoblast-rich rat calvarial cell cultures have been described, together with methods for the use of clonal osteogenic sarcoma cells of osteoblast phenotype. The latter clonal lines are useful for several purposes, but all the precautions and quality control measures necessary in the study of clonal lines must be observed. Some of the techniques for studying biochemical responses to hormones in these cells have also been detailed, but clearly others are applicable, including studies of the synthesis of matrix constituents. Osteoclast-like cells have not been considered in this chapter, because osteoclast culture methods have not yet been developed to the degree of purity and reproducibility necessary for this type of biochemical approach.

The Journal of biological chemistry, Jan 25, 1990

We have isolated clones for rat collagenase from a rat osteoblastic cell cDNA library. These clon... more We have isolated clones for rat collagenase from a rat osteoblastic cell cDNA library. These clones have been sequenced and the amino acids deduced. The calculated molecular weight is 51,352 for the proenzyme and 42,229 for the active enzyme. The deduced amino acid sequence was compared to those previously reported for: 1) human collagenase, 2) rat transin 1 (stromelysin), 3) human stromelysin, and 4) rabbit collagenase. The number of amino acids conserved was 47, 47, 50, and 47%, respectively. We also compared the collagenase mRNA and protein in different rat cells (osteoblast, uterine smooth muscle, synovial fibroblast) and determined that in rat uterine cells the message is slightly larger, although collagenase protein in all three cell types was identical in size. Parathyroid hormone dramatically induces the 2.9-kilobase collagenase mRNA in the rat osteoblastic cells, UMR 106-01. Nuclear run-on studies in UMR 106-01 cells demonstrated a 4-8-fold induction in the rate of synthesi...

Prostaglandins, 1985

SEPTEMBER1985VOL.30NO. 3 527 PROSTAGLANDINS predominant effect of the growth factor was an inhibi... more SEPTEMBER1985VOL.30NO. 3 527 PROSTAGLANDINS predominant effect of the growth factor was an inhibition of both PGE and PGI production by the osteoblastic cells. The'present gesults suggest that the boneresorbing hormones do not act to cause an increase in PG by the osteoblast and that any increase in PG production by these cells may be in response to vascular agents.

Principles of Bone Biology, 2002

Osteoporosis International, 2008

Osteoarthritis and Cartilage, 2006

Objective: Matrix metalloproteinase-13 (MMP-13) is an extracellular MMP that cleaves type II coll... more Objective: Matrix metalloproteinase-13 (MMP-13) is an extracellular MMP that cleaves type II collagen, the major protein component of cartilage, with high specificity and has been implicated in the pathology of osteoarthritis. The present study aimed to characterize the binding and internalization kinetics of MMP-13 in normal rabbit chondrocytes and whether MMP-13 affected cell signalling. Methods: Rabbit chondrocytes were used in [ 125 I]-MMP-13 binding assays to investigate the MMP-13 binding kinetics and Western analysis allowed for the assessment of intracellular signalling cascades. Results: Rabbit chondrocytes were found to express the cartilage-specific genes aggrecan and type II collagen throughout their in vitro culture period. Appreciable specific cell-association of [ 125 I]-MMP-13 was detected after 10 min of exposure to the ligand and equilibrium was obtained after 2 h. Binding of [ 125 I]-MMP-13 to chondrocytes was specific and approached saturation at 75 nM. Internalization of MMP-13 was evident after 20 min, reached a maximum at 30 min and had returned to baseline by 90 min. Addition of receptor-associated protein (RAP) inhibited the internalization of MMP-13 indicating a likely role for low-density lipoprotein receptor-related protein-1 (LRP1) in this process. Interestingly the presence of MMP-13 induced phosphorylation of the extracellular signal-regulated kinase 1/2 (ERK1/2) protein showing that there is initiation of a signalling process in response to MMP-13 being bound and internalized by rabbit chondrocytes. However, this activation does not involve the MMP-13 internalization receptor LRP1. Conclusion: These studies demonstrate and characterize the MMP-13 binding and internalization system in rabbit chondrocytes and indicate that MMP-13 may regulate the phenotype of the chondrocytes through this receptor system.

Molecular Endocrinology, 1992

Collagenase is synthesized and secreted by rat osteoblastic cells in response to PTH. We have pre... more Collagenase is synthesized and secreted by rat osteoblastic cells in response to PTH. We have previously demonstrated that this effect involves a substantial increase in collagenase mRNA via transcription. Northern blots and nuclear run-on assays were performed to further investigate the induction of collagenase by PTH in the rat osteoblastic cell line UMR 106-01. Detectable amounts of collagenase mRNA were not apparent until 2 h of PTH treatment, showed the greatest abundance at 4 h, and declined to approximately 30% of maximum by 8 h. The changes in the rate of transcription of the collagenase gene in response to PTH paralleled and preceded the changes in the steady state mRNA levels. After an initial lag period of about 1 h, collagenase transcription rates increased from very low levels to a maximal response at 2 h, returning to about 50% of maximum by 10 h. The increased transcriptional rate of the collagenase gene was found to be dependent on the concentration of PTH, with a half-maximal response at approximately 7 x 10(-10) M rat PTH-(1-34) and a maximal effect with a dose of 10(-8) M. The PTH-mediated induction of collagenase transcriptional activity was completely abolished by cycloheximide, while transcription of the beta-actin gene was unaffected by the translation inhibitor. These data suggest that a protein factor(s) is required for PTH-mediated transcriptional induction of collagenase. Since PTH increases intracellular levels of several potential second messengers, agents that mimic these substances were employed to determine which signal transduction pathway is predominant in the PTH-mediated stimulation of collagenase transcription.(ABSTRACT TRUNCATED AT 250 WORDS)

Molecular Endocrinology, 1992

We thank Katja Kopp and Deborah K. Clifton for their technical assistance. 13. McGrath JP, Capon ... more We thank Katja Kopp and Deborah K. Clifton for their technical assistance. 13. McGrath JP, Capon DJ, Goeddel DV, Levinson AD 1984 Comparative biochemical properties of normal and activated human ras ~21 orotein. Nature 310:644-649

Journal of Dental Research, 2012

Bisphosphonates are therapeutic agents in the treatment of post-menopausal osteoporosis. Although... more Bisphosphonates are therapeutic agents in the treatment of post-menopausal osteoporosis. Although they have been associated with delayed healing in injured tissues, inappropriate femoral fractures, and osteonecrosis of the jaw (ONJ), the pathophysiological mechanisms involved are not clear. Our hypothesis is that alendronate, a member of the N-containing bisphosphonates, indirectly inhibits osteoblast function through the coupling of osteoclasts to osteoblasts by ephrinB-EphB interaction. We found that alendronate increased gene and protein expression of ephrinB1 and EphB1, as well as B3, in femurs of adult mice injected with alendronate (10 µg/100 g/wk) for 8 weeks. Alendronate suppressed the expression of bone sialoprotein (BSP) and osteonectin in both femurs and bone marrow osteoblastic cells of mice. After elimination of pre-osteoclasts from bone marrow cells, alendronate did not affect osteoblast differentiation, indicating the need for pre-osteoclasts for alendronate’s effects...

The Journal of Clinical Endocrinology & Metabolism, 2011

Chronic high levels of PTH may be associated with up-regulation of proteases and cytokines. Monoc... more Chronic high levels of PTH may be associated with up-regulation of proteases and cytokines. Monocyte chemoattractant protein-1 (MCP-1) is an inflammatory cytokine, produced predominantly by macrophages and endothelial cells, and is expressed in adipose tissue. More recently it has been shown that PTH administration increases MCP-1 expression in osteoblasts. Because both PTH and MCP-1 levels are higher in obesity, the goal was to determine whether the high MCP-1 occurs only in the presence of high serum PTH and is independent of adiposity and examine its relationship with bone mineral density (BMD) and turnover. In this case-control clinical design, 111 eligible women were categorized into four groups: leaner women [body mass index (BMI) 23 ± 2 kg/m(2)] with normal or higher PTH and obese (BMI 44 ± 7 kg/m(2)) with normal or higher PTH. Serum MCP-1 levels were higher (P < 0.01) in the high (PTH = 74.9 ± 27.0 pg/ml, MCP-1 = 421.5 ± 157.0 pg/ml) compared with normal PTH (PTH = 32.5 ± 10.4 pg/ml, MCP-1 = 322.5 ± 97.8 pg/ml) group, independent of BMI. C-reactive protein and adiponectin were influenced only by BMI and not PTH. MCP-1 was positively associated with osteocalcin and propeptide of type 1 collagen in the leaner (r > 0.3, P < 0.05) but not the obese women and was not associated with BMD in either group. Together these data suggest that MCP-1 is higher only in the presence of increased PTH and that adiposity alone cannot explain the higher MCP-1 levels in obesity.