Sequential administration of alendronate and strontium ranelate: histomorphometry and bone biomechanics in ovariectomized animals.

Bisphosphonates are the first choice therapy for the pharmaco logical treatment of osteoporosis. Following reports of cases of bisphosphonaterelated osteonecrosis of the jaw and atypical femur fracture, the safety of longterm use of bisphosphonates has been evaluated, resulting in the proposal of strontium as an alternative drug. No experimental study using a sequential administration design has been reported to date. Hence, the aim of this study was to evaluate the effect on bone tissue of ovariectomized rats of administration of alendronate followed by strontium ranelate. Fortyeight female Wistar rats were ovariectomized on day 1 of the experiment. Beginning on day 30, they were administered 0.3 mg/kg/week of alendronate (ALN) or vehicle (VEH) for 8 weeks. Two groups (ALN and corresponding control) were euthanized at this time, and the remaining animals were divided into 4 groups and given 290 mg/kg/day of strontium ranelate (SR) in their drinking water (TW) or only water for 4 months. Experimental groups were: ALN+SR, ALN+TW, VEH+SR, VEH+TW, ALN and VEH. The tibiae and hemimandibles were resected for histomorphometric evaluation, and the right femur was used to perform biomechanical studies. ANOVA and Bonferroni test were applied. Diaphyseal stiffness, maximum elastic load and fracture load increased in animals that received alendronate, regardless of whether or not they received subsequent SR treatment. Fracture load also increased in VEH+ SR versus control (VEH+TW). Subchondral and interradicular bone volumes were significantly higher in animals that received ALN than in those that received vehicle. No difference was observed in cortical area or thickness of the tibia among treatments. The results obtained with the model presented here, evaluating tibial and mandibular interradicular bone, showed that the combination of ALN and SR and administration of ALN alone are equally effective in preventing bone loss associated with ovariectomyinduced estrogen depletion.

Si bien la primera opción terapéutica para el tratamiento farmacológico de la osteoporosis son los bisfosfonatos (BPs), luego de los primeros reportes en 2003 de los casos de osteone crosis de mandíbula asociada al uso de dichas drogas y las fracturas atípicas de fémur, se ha evaluado su seguridad a largo plazo. Además, en aquellos pacientes que no responden al tratamiento con BPs y mantienen elevado el riesgo de fractura, es necesario suspender su administración y alternar con otras drogas. Una de las que se ha utilizado en la clínica luego del tratamiento con BPs es el ranelato de estroncio (SR). Existen varios trabajos clínicos que reportan los efectos de la administra ción secuencial de ambas drogas, aunque estudios experi men tales con un diseño secuencial aun no se han reportado. Por ello el objetivo de este trabajo ha sido evaluar el efecto de la administración secuencial de alendronato, seguido de ranelato de estroncio sobre el tejido óseo de ratas ovariectomizadas. Se utilizaron 48 ratas Wistar hembras de dos meses de edad divididas en 6 grupos de 8 animales cada uno. El día 1 de experiencia todas fueron ovariectomizadas. El día 30 se comenzó con la administración de alendronato (ALN) en una dosis de 0.3 mg/kg/semana o vehículo (VEH) durante 8 semanas. Luego de este período se sacrificaron dos grupos (uno que recibió ALN y su correspondiente control (sólo vehículo). Los cuatro grupos restantes continuaron con ranelato de estroncio (SR) en el agua de bebida durante 4 meses en una dosis de 290 mg/kg/día o sólo agua corriente( TW) Luego de ese período fueron eutanasiados. Así, los grupos experimentales conformados fueron: ALN+SR, ALN+TW, VEH+SR, VEH+TW, ALN y VEH. Para los estudios histomorfométricos se extrajeron ambas tibias y hemimandíbulas; para el estudio biomecánico se utilizó el fémur derecho. Los resultados fueron analizados mediante el test de ANOVA y el test de Bonferroni. Incrementaron significativamente la rigidez diafisaria, la carga elástica límite y la carga de fractura aquellos grupos que recibieron alendronato versus aquellos que no lo recibieron, independientemente del tratamiento posterior con SR. La carga de fractura además fue mayor en el grupo VEH+SR versus el control (VEH+TW). En cuanto al volumen óseo subcondral e interradicular evaluado histomorfométricamente fue significativamente mayor en aquellos animales que recibieron ALN versus aquellos que recibieron vehículo. No se detectaron diferencias entre aquellos grupos que recibieron SR y sus controles. El área y espesor cortical de la tibia no mostraron diferencias entre grupos. Los resultados obtenidos en el modelo estudiado tanto a nivel del volumen óseo subcondral y cortical de la tibia como a nivel del hueso interradicular del maxilar inferior, mostraron que la combinación de ALN con SR y la administración aislada de ALN son igualmente efectivas para prevenir la pérdida ósea causada por la depleción estrogénica de la ovariectomía.

Sequential administration of alendronate and strontium ranelate: histomorphometry and bone biomechanics in ovariectomized animals

Bisphosphonates are the first choice therapy for the pharmaco logical treatment of osteoporosis. Following reports of cases of bisphosphonaterelated osteonecrosis of the jaw and atypical femur fracture, the safety of longterm use of bisphosphonates has been evaluated, resulting in the proposal of strontium as an alternative drug. No experimental study using a sequential administration design has been reported to date. Hence, the aim of this study was to evaluate the effect on bone tissue of ovariectomized rats of administration of alendronate followed by strontium ranelate. Fortyeight female Wistar rats were ovariectomized on day 1 of the experiment. Beginning on day 30, they were administered 0.3 mg/kg/week of alendronate (ALN) or vehicle (VEH) for 8 weeks. Two groups (ALN and corresponding control) were euthanized at this time, and the remaining animals were divided into 4 groups and given 290 mg/kg/day of strontium ranelate (SR) in their drinking water (TW) or only water for 4 months. Experimental groups were: ALN+SR, ALN+TW, VEH+SR, VEH+TW, ALN and VEH. The tibiae and hemimandibles were resected for histomorphometric evaluation, and the right femur was used to perform biomechanical studies. ANOVA and Bonferroni test were applied. Diaphyseal stiffness, maximum elastic load and fracture load increased in animals that received alendronate, regardless of whether or not they received subsequent SR treatment. Fracture load also increased in VEH+ SR versus control (VEH+TW). Subchondral and interradicular bone volumes were significantly higher in animals that received ALN than in those that received vehicle. No difference was observed in cortical area or thickness of the tibia among treatments. The results obtained with the model presented here, evaluating tibial and mandibular interradicular bone, showed that the combination of ALN and SR and administration of ALN alone are equally effective in preventing bone loss associated with ovariectomyinduced estrogen depletion.

Si bien la primera opción terapéutica para el tratamiento farmacológico de la osteoporosis son los bisfosfonatos (BPs), luego de los primeros reportes en 2003 de los casos de osteone crosis de mandíbula asociada al uso de dichas drogas y las fracturas atípicas de fémur, se ha evaluado su seguridad a largo plazo. Además, en aquellos pacientes que no responden al tratamiento con BPs y mantienen elevado el riesgo de fractura, es necesario suspender su administración y alternar con otras drogas. Una de las que se ha utilizado en la clínica luego del tratamiento con BPs es el ranelato de estroncio (SR). Existen varios trabajos clínicos que reportan los efectos de la administra ción secuencial de ambas drogas, aunque estudios experi men tales con un diseño secuencial aun no se han reportado. Por ello el objetivo de este trabajo ha sido evaluar el efecto de la administración secuencial de alendronato, seguido de ranelato de estroncio sobre el tejido óseo de ratas ovariectomizadas. Se utilizaron 48 ratas Wistar hembras de dos meses de edad divididas en 6 grupos de 8 animales cada uno. El día 1 de experiencia todas fueron ovariectomizadas. El día 30 se comenzó con la administración de alendronato (ALN) en una dosis de 0.3 mg/kg/semana o vehículo (VEH) durante 8 semanas. Luego de este período se sacrificaron dos grupos (uno que recibió ALN y su correspondiente control (sólo vehículo). Los cuatro grupos restantes continuaron con ranelato de estroncio (SR) en el agua de bebida durante 4 meses en una dosis de 290 mg/kg/día o sólo agua corriente( TW) Luego de ese período fueron eutanasiados. Así, los grupos experimentales conformados fueron: ALN+SR, ALN+TW, VEH+SR, VEH+TW, ALN y VEH. Para los estudios histomorfométricos se extrajeron ambas tibias y hemimandíbulas; para el estudio biomecánico se utilizó el fémur derecho. Los resultados fueron analizados mediante el test de ANOVA y el test de Bonferroni. Incrementaron significativamente la rigidez diafisaria, la carga elástica límite y la carga de fractura aquellos grupos que recibieron alendronato versus aquellos que no lo recibieron, independientemente del tratamiento posterior con SR. La carga de fractura además fue mayor en el grupo VEH+SR versus el control (VEH+TW). En cuanto al volumen óseo subcondral e interradicular evaluado histomorfométricamente fue significativamente mayor en aquellos animales que recibieron ALN versus aquellos que recibieron vehículo. No se detectaron diferencias entre aquellos grupos que recibieron SR y sus controles. El área y espesor cortical de la tibia no mostraron diferencias entre grupos. Los resultados obtenidos en el modelo estudiado tanto a nivel del volumen óseo subcondral y cortical de la tibia como a nivel del hueso interradicular del maxilar inferior, mostraron que la combinación de ALN con SR y la administración aislada de ALN son igualmente efectivas para prevenir la pérdida ósea causada por la depleción estrogénica de la ovariectomía.

Effect of alendronate on the mandible and long bones: an experimental study in vivo.

BACKGROUND: Bisphosphonates are anticatabolic agents that inhibit bone resorption and are widely used to treat osteoporosis and bone metastases in adults. They are also used in young patients with diseases like osteogenesis imperfecta or juvenile osteoporosis. Bone modeling/remodeling is elevated in growing subjects, and inhibition of osteoclastic activity has been shown to interfere with growth. Thus, our objective was to evaluate the effect of alendronate (ALN) on growing animals. METHODS: Healthy male Wistar rats, aged 1 mo, received ALN or vehicle for 8 wk. Serum levels (calcemia, phosphatemia, and total alkaline phosphatase) were determined. Morphometric (rat: femur and tibia weight and length and hemimandible growth) and histomorphometric parameters (thickness of tibial epiphyseal cartilage and each cartilage zone, interradicular bone volume in the first lower molar, trabeculae volume, percentage of bone and cartilage, and osteoclast number in mandibular condyles) were assessed. RESULTS: ALN caused a significant decrease in femur and tibia length, tibial cartilage thickness, and longitudinal growth of hemimandibles. It increased interradicular bone volume and mandibular condyle trabeculae volume, increasing the percentage of cartilage and osteoclast number. CONCLUSION: These findings indicate that administration of ALN to growing animals alters the endochondral ossification process, and thus alters growth.

Effect of strontium ranelate on bone remodeling.

Osteoporosis is a disease in which the microarchitecture of bone tissue deteriorates, with consequent loss of bone mass. Strontium ranelate (SrR) is currently used for treatment of the condition. SrR may have a dual effect: anabolic (stimulating pre-osteoblast replication) and anti-catabolic (reducing osteoclastic activity). However, its mechanism of action has not yet been completely elucidated. The aim of this study is to evaluate the effect of SrR on bone remodeling in healthy Wistar rats. Two-month old female Wistar rats were administered SrR (2 g/L) in drinking water for 30 weeks. Oriented histological sections were prepared from lower jaw and tibia and stained with H&E, and the following histomorphometric parameters were evaluated: a) in interradicular bone: bone volume, and percentages of bone-formation, quiescent and bone-resorption surfaces; and b) in tibia: bone volume, total thickness of growth cartilage, thickness of hypertrophic cartilage zone and number of megakaryocytes. No significant difference was found in the parameters between the control animals and those treated with SrR. The results would therefore show that SrR does not alter the bone parameters studied in this experimental design.

Effect of strontium ranelate on bone remodeling

Osteoporosis is a disease in which the microarchitecture of bone tissue deteriorates, with consequent loss of bone mass. Strontium ranelate (SrR) is currently used for treatment of the condition. SrR may have a dual effect: anabolic (stimulating pre-osteoblast replication) and anti-catabolic (reducing osteoclastic activity). However, its mechanism of action has not yet been completely elucidated. The aim of this study is to evaluate the effect of SrR on bone remodeling in healthy Wistar rats. Two-month old female Wistar rats were administered SrR (2 g/L) in drinking water for 30 weeks. Oriented histological sections were prepared from lower jaw and tibia and stained with H&E, and the following histomorphometric parameters were evaluated: a) in interradicular bone: bone volume, and percentages of bone-formation, quiescent and bone-resorption surfaces; and b) in tibia: bone volume, total thickness of growth cartilage, thickness of hypertrophic cartilage zone and number of megakaryocytes. No significant difference was found in the parameters between the control animals and those treated with SrR. The results would therefore show that SrR does not alter the bone parameters studied in this experimental design.

La osteoporosis es una enfermedad caracterizada por el deterioro de la microarquitectura del tejido oseo y la consecuente perdida de masa osea. El ranelato de estroncio (RSr) es actualmente utilizado para su tratamiento ya que poseeria un efecto dual: anabolico (estimulando la replicacion de preosteoblastos) y anticatabolico (disminuyendo la actividad osteoclastica). Sin embargo, su mecanismo de accion aun no ha sido completamente dilucidado. El objetivo del presente trabajo es evaluar el efecto del RSr sobre la remodelacion osea en ratas Wistar sanas. Se utilizaron ratas Wistar hembras de dos meses de edad a las cuales se les administro RSr (2 gr/L) en el agua de bebida durante 30 semanas. Se realizaron cortes histologicos orientados de maxilar inferior y tibia coloreados con H&E y se evaluaron los siguientes parametros histomorfometricos: a) En hueso interradicular: volumen oseo, porcentaje de superficies en formacion, reposo y reabsorcion osea. b) En tibia: volumen oseo, espesor total del cartilago de crecimiento, espesor de la zona de cartilago hipertrofiado y numero de megacariocitos. No se observaron diferencias significativas en los parametros evaluados entre los animales control y los tratados con RSr. Por lo tanto, los resultados obtenidos indicarian que el RSr no altera los parametros oseos estudiados en el presente diseño experimental.

Evaluation of vascular endothelial growth factor (VEGF) in interradicular bone marrow in olpadronate treated animals.

UNLABELLED: Vascular endothelial growth factor (VEGF) is a protein that increases vascular permeability and induces the proliferation, migration and survival of endothelial cells. Bisphosphonates (BPs) are antiresorptive drugs that are widely used in the treatment of bone metabolism diseases and bone metastases. Since 2003, cases of bisphosphonate-related osteonecrosis of the jaw (BRONJ) have been reported. Few papers explain the mechanisms that induce BRONJ; some authors mention alterations in bone remodelling and a certain antiangiogenic effect of BPs. The aim of this study is to evaluate the expression of VEGF in bone marrow cells and the number of blood vessels and area occupied by them in animals treated with the BP sodium olpadronate (OPD). We used 16 Wistar rats, 60 days old, divided into two groups, experimental (OPD) and control. The OPD group received 0.3 mg/kg/week intraperitoneal OPD for 5 weeks. The control group received an equivalent intraperitoneal volume of physiological saline solution. After euthanasia, hemimandibles were processed and mesio-distal histological sections of the first molar were prepared. Sections were stained with hematoxylin-eosin (HE), immunohistochemical detection of VEGF was performed (sc-7269) and the following histomorphometric parameters were evaluated: In HE-stained sections--number of blood vessels per sq. mm. and percentage (%) of area occupied by blood vessels in relation to total area evaluated; in sections with immunohistochemical detection of VEGF--number of VEGF+ bone marrow cells per sq. mm. Data underwent statistical analysis. Number of blood vessels/mm2 was significantly lower in the OPD group (OPD: 92 +/- 16; CONTROL: 140 +/- 31; p < 0.05) and % vascular area/total area evaluated showed no significant difference (OPD: 15.6 +/- 6.1; CONTROL: 10.2 +/- 4.2). Number of VEGF+ cells/mm2 was lower in the OPD group than in the control group, with statistically significant differences (OPD: 7804.8 +/- 597; CONTROL: 13187.6 +/- 894; p < .001). The results of this study suggest that monosodium olpadronate has an antiangiogenic effect. Further studies are needed to reveal its potential as an antitumor agent and its connection with the onset of BRONJ.