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P2 receptors modulate cell migration and differentiation in primary cultures of neonatal rat calvarial osteoblast (OBC)
602
Abstracts
and CAT. Besides, GLN arrests apoptosis of enterocytes by blocking the activation of the extrinsic and intrinsic pathways and preserving the expression of Bcl2.
doi:10.1016/j.bone.2012.12.030
Hybrid matrix grafts to favor tissue regeneration in rabbit femur bone lesions D.P. Goya, E. Gorositoa, D.J. Colettaa, H.S. Costab, P. Mortarinoa, N. Acosta Pedemontea, J. Toledoa, D. Abransona, N. Zingonia, H.S. Mansura, A. Jamina, M.M. Pereirab, R. Battaglinoc, S. Feldmana a Lab Biologìa Osteoarticular, Ing Tisular y Terapias Emergentes, Fac Cs Médicas, Univ Nac Rosario, Argentina b Dep. Eng. Metalúrgica e de Materiais, Univ Fed Minas Gerais, Brazil c Department of Cytokine Biology, The Forsyth Institute, Boston, Massachusetts, USA At present, typical approaches employed to repair fractures and other bone lesions tend to use matrix grafts to promote tissue regeneration. These grafts act as templates, which promote cellular adhesion, growth and proliferation, osteoconduction, and even osteoinduction. The present work aimed to study the bonerepairing ability of hybrid matrixes (MHt) prepared with 70% polyvinyl alcohol and bioactive glass (30% bioactive glass, nominal composition of 58% SiO2 − 33% CaO − 9% P2O5) neutralized with NH4OH, in an experimental rabbit model. New Zealand rabbits were randomly divided into the control group (C) and two groups with bone lesions, in which one received a matrix implant (Im), while the other did not (NoI). Clinical monitoring showed no altered parameters from either the Im or the NoI groups as compared to C, for the variables of diet grades, day and night temperatures and hemograms. In the Im, radiologic and tomographic studies showed implanted areas with clean edges in femoral non-articular direction, and radio-dense images that suggest incipient integration. Minimum signs of phlogosis could be observed, whereas no signs of rejection at this imaging level could be identified. Histological analysis showed evidence of osteo-integration, with the formation of a trabecular bone within the implant. Together, these results show that implants of MHt are capable of promoting bone regeneration.
doi:10.1016/j.bone.2012.12.031
Impairment in bone resorption in a model of lead poisoning and experimental periodontitis. Histomorphometric study P.I. Mitre, H.E. Meheris, J.A. Garat Department of Histology and Pharmacology, Dental School, National University of Tucuman, Argentina Low levels of lead alter the function of bone cells. It has been described that lead causes osteopenia; in recent years the association between lead exposure and bone density is controversial since there were no observed reductions in bone density in children exposed to lead or instead increase it when exposed to high doses. Lead poisoning inhibits bone-forming activity, however, little is known about the effects of lead on resorptive activity. Then this study aimed to determine the toxic effects of lead in bone resorption and bone microarchitecture on alveolar bone. We use the alveolar bone given that this allows us to create experimental conditions that generate bone resorption and osteoclast recruitment very soon. Wistar rats aged 21 days were assigned to one of the following groups: GI fed a hard diet and a solution of water and acetic acid 0.00125% and G II fed a hard diet and a water solution of acetic acid and 0.00125% more lead acetate dissolved in sufficient quantity to achieve a concentration of 1000 ppm during 60 days. At day 57 periodontitis was induced by placing a ligature of cotton thread around the neck of the first molar in both groups. At day 60 animals were sacrificed, lower jaws were dissected and embedded in parafin. Mesiodistal sections of the first molar were performed and stained with hematoxylin and eosin. Determinations in the cervical and middle thirds of the interradicular bone were: (a) bone loss (BL), (B) bone volume (BV), (C) trabecular thickness, (d). trabecular number, and (e) trabecular spacement. Data were analyzed using Student's t test. Results: BL were significantly lower in GII compared with GI. No statistical differences were observed in the other parameters measured. Conclusions: Poisoning in drinking water alters bone resorption in periodontitis on periodontal surface of the interradicular bone crest.
doi:10.1016/j.bone.2012.12.032
Biomechanical analysis of trabecular and cortical bone of ovariectomized rats treated sequentially with monofluorophosphate and zoledronate M.L. Brance, L.R. Brun, H. Moreno, A. Rigalli Bone Biology Laboratory, School of Medicine, UNRosario, Argentina Monofluorophosphate (MFP) increases bone formation and zoledronate (Z) decreases bone resorption. It has been hypothesized that the effect of MFP would be more effective on a high remodeling state. The effect has been demonstrated by DEXA and histomorphometry. The aim of this study was to evaluate the biomechanical bone properties of ovariectomized (OVX) rats treated sequentially with MFP and Z with simultaneous modifications of bone remodeling by changing in the calcium diet content. Sprague Dawley rats of 7 weeks (n = 7/group) were OVX and fed with low calcium diet (LCaD) for 30 days. After that rats were treated with MFP and high calcium diet (HCaD) for 60 days and finally treated with Z and HCaD for another 60 days (group OVX.G3). Control groups were performed: Sham: simulated surgery; OVX: OVX rats without treatment; OVX.G1: OVX rats with MFP and Z treatment and normal calcium diet (NCaD). OVX.G2: OVX rats with only changes in the diet Ca content. On both femurs biomechanical tests were performed: three-point bending test to evaluate cortical tissue and compression test to evaluate trabecular bone. The results are expressed as mean ± EE. Student t test was performed for independent data comparing Sham vs OVX, and each treated group vs OVX. * Indicates p b 0.05. At 150 days, the bending test showed higher fracture load in OVX.G3 compared with OVX (227.30 ± 5.70* vs 206.70 ± 4.99 N). The CSMI remained stable which can be explained by improvement in the Young's modulus (7.47 ± 1.74 vs 5.43 ± 0.76 GPa). In the compression test the OVX.G3 group had a higher fracture load (42.42 ± 5.42* vs 26.78 ± 3.96 N), maximum load (71.77 ± 5.99* vs 55.71 ± 5.47 N) and stiffness (608.9 ± 108.4* vs 258.4 ± 48.3 N/mm) compared to OVX. This can be explained by the increase in trabecular bone histomorphometry previously found and because we also found increase in the Young's modulus (0.21 ± 0.04* vs 0.09 ± 0.02 GPa).
doi:10.1016/j.bone.2012.12.033
P2 receptors modulate cell migration and differentiation in primary cultures of neonatal rat calvarial osteoblast (OBC) V.B. Ayala Peña, J.A. Laiuppa, G.E. Santillán Departamento de Biología, Bioquímica y Farmacia, Universidad Nacional del Sur-CONICET, Argentina Bone cells are exposed to mechanical stimuli and injury and lysis during movements and trauma respectively. This favors ATP and UTP release to extracellular environment. Nucleotides can trigger different responses acting through plasma membrane P2 receptors (P2X ionotropic, and P2Y metabotropic). We have previously reported that ATP stimulates alkaline phosphatase activity in OBC after 7 days of treatment in osteogenic medium. In this work, we studied the participation of P2 receptors in the modulation of cell migration and differentiation. Cell migration was evaluated by “wound” assay. Differentiation into osteoblast was analyzed by detection of mineralized nodules after Alizarin red staining, and Bone Sioaloprotein (BSP) gene expression by qRT-PCR. Cells were incubated with 1–100 μM of several nucleotides (UTP, UDP, ADP, ATPγS or ADPβS) for 1–22 days. Treatment of OBC with 10 μM ADPβS seems to stimulate cell migration, whereas 100 μM ATPγS appears to show an inhibitory effect, with respect to controls. OBC treated with 100 μM ATP or UTP in osteogenic medium showed morphological changes from the 7th day of treatment, these changes correlate with an increased BSP expression. In addition, under the same conditions, we observed an increased calcium deposition around the 22nd day, with respect to controls. Our results suggest an important role for P2 receptors in bone cell migration and maturation. Extracellular nucleotides signaling would be involved in the regulation of cellular migration through P2Y1, 12 y 13 receptors (responsive to ADP). Moreover, P2Y2 and/or P2Y4 receptors (ATP and UTP responsive) would regulate genes expression involved in osteoblast maturation and function.
doi:10.1016/j.bone.2012.12.034
Parathyroid hormone related peptide (PTHrP) stimulates mitogenic pathways of MAP kinases in cells of human colon cancer M.J. Martin, N.G. Calvo, A. Russo de Boland, C.R. Gentili Dpto. de Biología, Bioquímica & Farmacia, Universidad Nacional del Sur, Bahía Blanca, Argentina
Abstracts
and CAT. Besides, GLN arrests apoptosis of enterocytes by blocking the activation of the extrinsic and intrinsic pathways and preserving the expression of Bcl2.
doi:10.1016/j.bone.2012.12.030
Hybrid matrix grafts to favor tissue regeneration in rabbit femur bone lesions D.P. Goya, E. Gorositoa, D.J. Colettaa, H.S. Costab, P. Mortarinoa, N. Acosta Pedemontea, J. Toledoa, D. Abransona, N. Zingonia, H.S. Mansura, A. Jamina, M.M. Pereirab, R. Battaglinoc, S. Feldmana a Lab Biologìa Osteoarticular, Ing Tisular y Terapias Emergentes, Fac Cs Médicas, Univ Nac Rosario, Argentina b Dep. Eng. Metalúrgica e de Materiais, Univ Fed Minas Gerais, Brazil c Department of Cytokine Biology, The Forsyth Institute, Boston, Massachusetts, USA At present, typical approaches employed to repair fractures and other bone lesions tend to use matrix grafts to promote tissue regeneration. These grafts act as templates, which promote cellular adhesion, growth and proliferation, osteoconduction, and even osteoinduction. The present work aimed to study the bonerepairing ability of hybrid matrixes (MHt) prepared with 70% polyvinyl alcohol and bioactive glass (30% bioactive glass, nominal composition of 58% SiO2 − 33% CaO − 9% P2O5) neutralized with NH4OH, in an experimental rabbit model. New Zealand rabbits were randomly divided into the control group (C) and two groups with bone lesions, in which one received a matrix implant (Im), while the other did not (NoI). Clinical monitoring showed no altered parameters from either the Im or the NoI groups as compared to C, for the variables of diet grades, day and night temperatures and hemograms. In the Im, radiologic and tomographic studies showed implanted areas with clean edges in femoral non-articular direction, and radio-dense images that suggest incipient integration. Minimum signs of phlogosis could be observed, whereas no signs of rejection at this imaging level could be identified. Histological analysis showed evidence of osteo-integration, with the formation of a trabecular bone within the implant. Together, these results show that implants of MHt are capable of promoting bone regeneration.
doi:10.1016/j.bone.2012.12.031
Impairment in bone resorption in a model of lead poisoning and experimental periodontitis. Histomorphometric study P.I. Mitre, H.E. Meheris, J.A. Garat Department of Histology and Pharmacology, Dental School, National University of Tucuman, Argentina Low levels of lead alter the function of bone cells. It has been described that lead causes osteopenia; in recent years the association between lead exposure and bone density is controversial since there were no observed reductions in bone density in children exposed to lead or instead increase it when exposed to high doses. Lead poisoning inhibits bone-forming activity, however, little is known about the effects of lead on resorptive activity. Then this study aimed to determine the toxic effects of lead in bone resorption and bone microarchitecture on alveolar bone. We use the alveolar bone given that this allows us to create experimental conditions that generate bone resorption and osteoclast recruitment very soon. Wistar rats aged 21 days were assigned to one of the following groups: GI fed a hard diet and a solution of water and acetic acid 0.00125% and G II fed a hard diet and a water solution of acetic acid and 0.00125% more lead acetate dissolved in sufficient quantity to achieve a concentration of 1000 ppm during 60 days. At day 57 periodontitis was induced by placing a ligature of cotton thread around the neck of the first molar in both groups. At day 60 animals were sacrificed, lower jaws were dissected and embedded in parafin. Mesiodistal sections of the first molar were performed and stained with hematoxylin and eosin. Determinations in the cervical and middle thirds of the interradicular bone were: (a) bone loss (BL), (B) bone volume (BV), (C) trabecular thickness, (d). trabecular number, and (e) trabecular spacement. Data were analyzed using Student's t test. Results: BL were significantly lower in GII compared with GI. No statistical differences were observed in the other parameters measured. Conclusions: Poisoning in drinking water alters bone resorption in periodontitis on periodontal surface of the interradicular bone crest.
doi:10.1016/j.bone.2012.12.032
Biomechanical analysis of trabecular and cortical bone of ovariectomized rats treated sequentially with monofluorophosphate and zoledronate M.L. Brance, L.R. Brun, H. Moreno, A. Rigalli Bone Biology Laboratory, School of Medicine, UNRosario, Argentina Monofluorophosphate (MFP) increases bone formation and zoledronate (Z) decreases bone resorption. It has been hypothesized that the effect of MFP would be more effective on a high remodeling state. The effect has been demonstrated by DEXA and histomorphometry. The aim of this study was to evaluate the biomechanical bone properties of ovariectomized (OVX) rats treated sequentially with MFP and Z with simultaneous modifications of bone remodeling by changing in the calcium diet content. Sprague Dawley rats of 7 weeks (n = 7/group) were OVX and fed with low calcium diet (LCaD) for 30 days. After that rats were treated with MFP and high calcium diet (HCaD) for 60 days and finally treated with Z and HCaD for another 60 days (group OVX.G3). Control groups were performed: Sham: simulated surgery; OVX: OVX rats without treatment; OVX.G1: OVX rats with MFP and Z treatment and normal calcium diet (NCaD). OVX.G2: OVX rats with only changes in the diet Ca content. On both femurs biomechanical tests were performed: three-point bending test to evaluate cortical tissue and compression test to evaluate trabecular bone. The results are expressed as mean ± EE. Student t test was performed for independent data comparing Sham vs OVX, and each treated group vs OVX. * Indicates p b 0.05. At 150 days, the bending test showed higher fracture load in OVX.G3 compared with OVX (227.30 ± 5.70* vs 206.70 ± 4.99 N). The CSMI remained stable which can be explained by improvement in the Young's modulus (7.47 ± 1.74 vs 5.43 ± 0.76 GPa). In the compression test the OVX.G3 group had a higher fracture load (42.42 ± 5.42* vs 26.78 ± 3.96 N), maximum load (71.77 ± 5.99* vs 55.71 ± 5.47 N) and stiffness (608.9 ± 108.4* vs 258.4 ± 48.3 N/mm) compared to OVX. This can be explained by the increase in trabecular bone histomorphometry previously found and because we also found increase in the Young's modulus (0.21 ± 0.04* vs 0.09 ± 0.02 GPa).
doi:10.1016/j.bone.2012.12.033
P2 receptors modulate cell migration and differentiation in primary cultures of neonatal rat calvarial osteoblast (OBC) V.B. Ayala Peña, J.A. Laiuppa, G.E. Santillán Departamento de Biología, Bioquímica y Farmacia, Universidad Nacional del Sur-CONICET, Argentina Bone cells are exposed to mechanical stimuli and injury and lysis during movements and trauma respectively. This favors ATP and UTP release to extracellular environment. Nucleotides can trigger different responses acting through plasma membrane P2 receptors (P2X ionotropic, and P2Y metabotropic). We have previously reported that ATP stimulates alkaline phosphatase activity in OBC after 7 days of treatment in osteogenic medium. In this work, we studied the participation of P2 receptors in the modulation of cell migration and differentiation. Cell migration was evaluated by “wound” assay. Differentiation into osteoblast was analyzed by detection of mineralized nodules after Alizarin red staining, and Bone Sioaloprotein (BSP) gene expression by qRT-PCR. Cells were incubated with 1–100 μM of several nucleotides (UTP, UDP, ADP, ATPγS or ADPβS) for 1–22 days. Treatment of OBC with 10 μM ADPβS seems to stimulate cell migration, whereas 100 μM ATPγS appears to show an inhibitory effect, with respect to controls. OBC treated with 100 μM ATP or UTP in osteogenic medium showed morphological changes from the 7th day of treatment, these changes correlate with an increased BSP expression. In addition, under the same conditions, we observed an increased calcium deposition around the 22nd day, with respect to controls. Our results suggest an important role for P2 receptors in bone cell migration and maturation. Extracellular nucleotides signaling would be involved in the regulation of cellular migration through P2Y1, 12 y 13 receptors (responsive to ADP). Moreover, P2Y2 and/or P2Y4 receptors (ATP and UTP responsive) would regulate genes expression involved in osteoblast maturation and function.
doi:10.1016/j.bone.2012.12.034
Parathyroid hormone related peptide (PTHrP) stimulates mitogenic pathways of MAP kinases in cells of human colon cancer M.J. Martin, N.G. Calvo, A. Russo de Boland, C.R. Gentili Dpto. de Biología, Bioquímica & Farmacia, Universidad Nacional del Sur, Bahía Blanca, Argentina