RESUMEN
We evaluated the embryolethality and embryotoxicity of sera from patients suffering from autoimmune diseases during remission on post-implantation rat embryos cultured on these sera and determined the association between the patients' clinical history, high blood levels of specific antibodies, medications, and oxidative stress parameters. One hundred and eighty, 10.5-day-old rat embryos were cultured in their yolk sacs in 33 sera of systemic lupus erythematosus (SLE)/antiphospholipid syndrome (APS) patients, and compared with 84 embryos cultured in rat sera and 88 embryos cultured in control human sera. The sera proved to be lethal and embryotoxic but not teratogenic resulting in smaller yolk sacs and embryos, lower protein level and lower developmental score. Significantly less embryos cultured in 'toxic' SLE/APS sera had peak 2 of low molecular weight antioxidants (LMWA) wave, implying a delayed maturation of the antioxidant defense. Lower peak 1 of LMWA correlated with a history of recurrent abortions. Embryonic levels of superoxide dismutase (SOD) and catalase (CAT) did not correlate with sera toxicity, patients' clinical history or specific antibodies. We conclude that SLE/APS patients' clinical remission did not prevent death or developmental delay accompanied by later appearance of peak 2 of LMWA in post-implantation rat embryo cultures. The normal levels of the antioxidant enzymes evaluated may indicate that sera toxicity is not related to oxidative stress.
Asunto(s)
Anticuerpos/sangre , Síndrome Antifosfolípido/sangre , Embrión de Mamíferos/inmunología , Lupus Eritematoso Sistémico/sangre , Animales , Anticuerpos/inmunología , Antioxidantes/metabolismo , Síndrome Antifosfolípido/inmunología , Catalasa/metabolismo , Femenino , Humanos , Lupus Eritematoso Sistémico/inmunología , Masculino , Estrés Oxidativo , Embarazo , Ratas , Superóxido Dismutasa/metabolismoRESUMEN
Systemic lupus erythematosus (SLE) and primary anti-phospholipid syndrome (PAPS) are autoimmune diseases causing recurrent pregnancy loss. We hypothesized that anti-phospholipid antibodies (aPL), but not anti-Ro and anti-La antibodies, might have a role through direct placental damage. We cultured human placental explants in sera from women with SLE/PAPS with different antibodies. These sera were found to reduce placental growth and increase trophoblastic apoptosis. No effect was found on estradiol or progesterone secretion, but inhibition in betahCG secretion was detected. BetahCG was reduced in women with a history of recurrent pregnancy loss or thromboembolic events, and was also the most sensitive marker when examining the effects of specific antibodies. High titers of aPL were found to cause the largest reduction in betahCG. Anti-Ro and anti-La did not induce placental damage. A strong correlation was found between the rise in the number of different antibodies in the sera and the incidence of recurrent pregnancy loss, which was also accompanied by a decline in the betahCG levels. In conclusion, aPL, but not anti-Ro or anti-La, may cause placental damage in vitro. Thus betahCG levels might constitute a predictive marker for the risk of placental damage and pregnancy loss in women with SLE/PAPS.
Asunto(s)
Anticuerpos Antinucleares/fisiología , Anticuerpos Antifosfolípidos/fisiología , Síndrome Antifosfolípido/inmunología , Gonadotropina Coriónica/metabolismo , Lupus Eritematoso Sistémico/inmunología , Placentación , Síndrome Antifosfolípido/sangre , Autoantígenos/inmunología , Femenino , Humanos , Técnicas In Vitro , Lupus Eritematoso Sistémico/sangre , Ribonucleoproteínas/inmunología , Antígeno SS-BRESUMEN
Oophorectomized (OVX) rats served for many years as a popular model for 'postmenopausal' osteoporosis in spite of the fact that the rat continues to grow during these experiments. We performed OVX in rats at 1, 3, 6 and 10 months of age and compared the histomorphometric (bone size, bone trabecular and cartilage volume in different areas) and chemical (ash, Ca, P and Mg content) parameters at 2, 6, 8 and 20 weeks post-OVX (2-20 weeks) to those of sham-operated rats. Significant differences were observed only in the animals that were OVX at a young or young-mature age, i.e. mainly at 1 and 3 months and some of the rats at 6 months of age. There were no changes in bone ash and mineral contents in the OVX animals in comparison to sham-operated rats, except 2 weeks post-OVX in the 1-month-old rats where these variables were reduced. The most significant finding was a reduction in the metaphyseal bone volume. This was observed in the rats OVX at 1, 3 and 6 months of age, but not in those OVX at 10 months. In the young OVX rats there was also an increase in epiphyseal cartilage volume. The epiphyseal and diaphyseal bone volumes were not different between the groups at any time postsurgery, explaining the lack of differences in bone ash and mineral contents. There were no significant changes in the results of the histomorphometric studies between OVX and sham-operated rats when surgery was performed at 10 months of age. Since OVX exerts significant changes only in young rapidly growing rats, this approach is an inappropriate model for postmenopausal osteoporosis, which occurs long after bone growth has ended.
Asunto(s)
Enfermedades Óseas Metabólicas/patología , Enfermedades Óseas Metabólicas/fisiopatología , Osteoporosis Posmenopáusica/patología , Osteoporosis Posmenopáusica/fisiopatología , Ovariectomía/efectos adversos , Factores de Edad , Animales , Peso Corporal , Densidad Ósea/fisiología , Femenino , Placa de Crecimiento/patología , Placa de Crecimiento/fisiopatología , Humanos , Ratas , Ratas Wistar , Tibia/patología , Tibia/fisiopatología , Factores de TiempoRESUMEN
PURPOSE: To validate our hypothesis that a bisphosphonate (BP) having a nitrogen-containing heterocyclic ring on the side chain, and with no hydroxyl on the geminal carbon would possess increased activity, and better oral bioavailability due to enhanced solubility of its calcium complexes/salts and weaker Ca chelating properties. METHODS: A novel BP, 2-(2-aminopyrimidinio)ethylidene-1, 1-bisphosphonic acid betaine (ISA-13-1) was synthesized. The physicochemical properties and permeability were studied in vitro. The effects on macrophages, bone resorption (young growing rat model), and tumor-induced osteolysis (Walker carcinosarcoma) were studied in comparison to clinically used BPs. RESULTS: The solubility of the Ca salt of ISA-13-1 was higher, and the log beta(Ca:BP) stability constant and the affinity to hydroxyapatite were lower than those of alendronate and pamidronate. ISA-13-1 exhibited effects similar to those of alendronate on bone volume, on bone osteolysis, and on macrophages, following delivery by liposomes. ISA-13-1 was shown to have 1.5-1.7 times better oral absorption than the other BPs with no deleterious effects on the tight junctions of intestinal tissue. CONCLUSIONS: The similar potency to clinically used BPs, the increased oral absorption as well as the lack of effect on tissue tight junction of ISA-13-1 warrant its further consideration as a potential drug for bone diseases.
Asunto(s)
Betaína/análogos & derivados , Enfermedades Óseas/tratamiento farmacológico , Difosfonatos/síntesis química , Difosfonatos/farmacocinética , Administración Oral , Alendronato/química , Alendronato/farmacocinética , Animales , Antineoplásicos/farmacocinética , Betaína/síntesis química , Betaína/farmacocinética , Enfermedades Óseas/metabolismo , Neoplasias Óseas/tratamiento farmacológico , Calcio/orina , Cápsulas , Carcinoma 256 de Walker/tratamiento farmacológico , Línea Celular , Diuréticos Osmóticos/farmacocinética , Durapatita/química , Inyecciones Intravenosas , Absorción Intestinal/efectos de los fármacos , Macrófagos/citología , Macrófagos/efectos de los fármacos , Masculino , Manitol/farmacocinética , Osteogénesis/efectos de los fármacos , Pamidronato , Ratas , Ratas Endogámicas , Uniones Estrechas/efectos de los fármacos , Uniones Estrechas/metabolismo , Distribución TisularRESUMEN
Bisphosphonates are clinically used mainly to reduce bone resorption. We studied the transplacental effects of two bisphosphonates on the fetal skeleton in rats. Pregnant rats were treated during days 11-20 of pregnancy with daily subcutaneous injections of 0.1 mg/kg of alendronate or a newly synthesized bisphosphonate, VS-b6. This period of pregnancy was chosen because the active development of bones from mesenchyme through cartilaginous models occurs during that time. Histological examination of midlongitudinal sections of the 21-day-old fetuses showed an increase in the amount of diaphyseal bone trabeculae with slight shortening of the diaphysis in the experimental fetuses, in comparison to controls. Computerized histomorphometric studies similarly showed an increase in the amount of diaphyseal bone trabeculae with a concomitant decrease in bone marrow volume, but no change in cartilage volume. In addition, chemical analysis of the fetal bones showed an increase in calcium content in the treated fetuses. 14C-alendronate was shown to pass through the rat placenta and accumulate in the fetuses, most probably in their bones. This is presumed because bisphosphonates are known to accumulate in bone, being stored there for long periods of time. It is important, in light of our results, to give careful consideration to the treatment of women with bisphosphonates at childbearing age, whenever this is needed.
Asunto(s)
Alendronato/efectos adversos , Huesos/efectos de los fármacos , Calcificación Fisiológica/efectos de los fármacos , Difosfonatos/efectos adversos , Animales , Huesos/embriología , Femenino , Feto/efectos de los fármacos , Embarazo , Ratas , Ratas WistarRESUMEN
PURPOSE: This work was aimed at synthesizing novel bisphosphonates (BPs) and examining them in comparison to clinically used BPs such as pamidronate and alendronate, and to tetracycline, in order to evaluate their potential as anticalcification and antiresorption agents. The correlation between the various models was examined in order to establish facile experimental models for pre-screening of potential compounds. METHODS: Nitrogen-containing heterocyclic, novel BPs such as 2-(3-methylimidazolio) ethylidene-1,1-bisphosphonic acid betaine (VS-5b), 2-(2-dimethylamino-4-pyrazinio)ethylidene-1,1-bisphosphonic acid betaine (VS-6b), and 2-(2-alpha-pyridylethylthio) ethylidene-1,1-bisphosphonic acid (ISA-225), were synthesized and evaluated in comparison to clinically used BPs, in various experimental models of resorption and calcification. RESULTS: The physicochemical properties of the novel compounds are slightly different than the BPs in clinical use: the pKa values are lower, the affinity for hydroxyapatite is lower and the solubilities of the calcium salts are higher. The anticalcification potencies of the novel compounds were high and ranked as follows: alendronate = pamidronate > VS-6b = VS-5b = ISA-225 > tetracycline. The in vivo antiresorption activity of VS-5b and VS-6b in comparison to that of the clinically employed, pamidronate, was shown to be similar and higher, respectively. CONCLUSIONS: The anticalcification activity of the novel compounds as well as that of tetracycline was lower than that of alendronate. The antiresorption activity of VS-6b was similar to that of pamidronate. A good correlation between the different models was found, enabling the facile screening of novel compounds. The activities of tetracycline and EDTA highlight the distinct behavior of BPs as "crystal poison." In addition, tetracycline was found to be a potent anticalcification agent in the ectopic calcification model.
Asunto(s)
Antiinflamatorios/uso terapéutico , Resorción Ósea/tratamiento farmacológico , Huesos/efectos de los fármacos , Calcinosis/tratamiento farmacológico , Difosfonatos/síntesis química , Difosfonatos/uso terapéutico , Inhibidores de la Síntesis de la Proteína/uso terapéutico , Tetraciclina/uso terapéutico , Alendronato/uso terapéutico , Animales , Betaína/análogos & derivados , Betaína/síntesis química , Betaína/uso terapéutico , Huesos/química , Quelantes/uso terapéutico , Ácido Edético/uso terapéutico , Miembro Posterior , Hidroxiapatitas/química , Masculino , Pamidronato , Ratas , Relación Estructura-ActividadRESUMEN
The biological effects of bisphosphonates in calcium-related disorders are attributed to the incorporation of the bisphosphonates in bone, enabling direct interaction with osteoclasts and/or osteoblasts. The high accumulation of bisphosphonates in bone, due to their high affinity to hydroxyapatite (HAP), is essential for mediating in vitro and in vivo activity. In this study we examined the activity of tetrakisphosphonates, molecules containing two P-C-P type bisphosphonate moieties connected by a carbon chain. The novel compounds were examined in a battery of in vitro and in vivo models including HAP formation and dissolution, ectopic calcification, bone resorption, tumor osteolysis, and of macrophage-like cells (anti- or pro-inflammatory properties). The inhibition of ectopic calcification was ranked as follows: geminal bisphosphonates > bisacylphosphonates > tetrakisphosphonates. Pamidronate, but not the tetrakisphosphonates, was an effective antiosteolytic agent. Neither DNTP (tetrasodium 1,9-dihydroxynonane 1,1,9,9-tetrakisphosphonate) nor the bisacylphosphonate, PiBP (pimeloylbisphosphonate) seem to possess strong macrophage suppressive or inductive effects and can be considered to be relatively inactive in terms of anti- or pro-inflammatory action. A significant anticalcification effect was caused by various phosphonates, such as the tetrakisphosphonates, but DNTP, a tetrakisphosphonate, was found toxic as it impeded somatic growth and bone development.
Asunto(s)
Resorción Ósea/inducido químicamente , Calcinosis/prevención & control , Carcinoma 256 de Walker/tratamiento farmacológico , Difosfonatos/farmacología , Osteólisis/prevención & control , Animales , Bioprótesis , Calcinosis/patología , Carcinoma 256 de Walker/patología , Línea Celular , Supervivencia Celular/efectos de los fármacos , Difosfonatos/síntesis química , Difosfonatos/toxicidad , Durapatita/química , Ácido Etidrónico/farmacología , Femenino , Macrófagos/efectos de los fármacos , Ratones , Osteólisis/patología , Pamidronato , Ratas , Ratas WistarRESUMEN
Bisphosphonates are potent inhibitors of bone resorption, and are therefore used for the treatment of various bone diseases including osteoporosis. We examined whether cyclic therapy with bisphosphonates in oophorectomized osteoporotic rats had any advantage over continuous treatment. We therefore treated intact and oophorectomized young female rats for 8 weeks with 1 and 5 mg/kg/day of pamidronate. The 8-week treatment was given continuously for 6 days/week or intermittently, i.e. 6 days of pamidronate (APD) and 3 weeks off, for 2 cycles. We found an increase in tibial wet and ash weight and in the mineral content in oophorectomized rats treated continuously or intermittently with APD in comparison to nontreated oophorectomized animals. Histomorphometric analysis showed an increase in the volume of metaphyseal cartilage and bone. No changes were found in the volume of epiphyseal or diaphyseal bone. Pamidronate had very little effect on the bone of intact rats. Pamidronate seems to be more effective in inhibiting bone resorption in bone that undergoes rapid turnover (i.e. in oophorectomized animals) when compared to bone with low turnover (intact rats). Although the results of cyclic treatment were similar to those of continuous treatment, we have to remember that cyclic therapy may be more advantageous since animals receiving cyclic therapy received only 25% of the dose of rats continuously treated.
Asunto(s)
Antineoplásicos/administración & dosificación , Difosfonatos/administración & dosificación , Ovariectomía , Tibia/efectos de los fármacos , Animales , Calcio/análisis , Femenino , Inyecciones Subcutáneas , Magnesio/análisis , Tamaño de los Órganos/efectos de los fármacos , Pamidronato , Fósforo/análisis , Ratas , Ratas Wistar , Tibia/química , Tibia/patologíaRESUMEN
High doses of X-irradiation may affect the developing human embryo and fetus, causing brain, eye, skeletal and other defects. Although the doses used in diagnostic irradiation are not considered to be high enough to cause fetal anomalies, it is unknown whether they affect the long-term development of the in utero exposed children. Using the Bayley or McCarthy developmental scales we examined 52 children born to mothers exposed to diagnostic X-irradiation of the abdomen and/or pelvis during the first trimester of pregnancy, 60 children born to mothers exposed during pregnancy (58 in the first trimester) to X-rays in areas other than the abdomen or pelvis, and 62 normal children without maternal exposure to X-rays during pregnancy. All children were aged 1-5 years at examination. Parental socioeconomic status was the same in all three groups. There were no differences in the results of the medical and neurological examination, and in the motor or congnitive scores among three groups. Embryonic or fetal exposure to X-irradiation in doses below 5 rads does not seem to have any effect on development during childhood.
Asunto(s)
Desarrollo Infantil , Embrión de Mamíferos/efectos de la radiación , Feto/efectos de la radiación , Diagnóstico Prenatal , Efectos Tardíos de la Exposición Prenatal , Radiografía/efectos adversos , Factores de Edad , Preescolar , Femenino , Humanos , Lactante , Embarazo , Dosis de RadiaciónRESUMEN
Some geminal bisphosphonates are used clinically in a number of important bone and calcium-related diseases. This work reports the anticalcification and antiresorption effects of a series of bisacylphosphonates, nongeminal compounds with varying chain lengths having oxo groups in alpha positions relative to the phosphonic functions. We compared the activity of the novel compounds to clinically used geminal bisphosphonates, and to a bisphosphonate devoid of the oxo groups. The interaction of the compounds with calcium was studied by various in vitro and in vivo models. We found that keto groups in alpha positions to the phosphonic functions render activity. The bisacylphosphonates with a shorter chain [(CH2)n, = 4, 6] were found significantly to inhibit hydroxyapatite formation and dissolution in vitro, the calcification of bioprosthetic tissue implanted subdermally in rats, and bone resorption in the intact young animal model. The various in vitro results were found to be in good correlation with the in vivo results. Structure-activity relationship studies indicate that both bisacylphosphonates and geminal bisphosphonates are active only when at least three ionizable groups are present in the molecule. The role of the keto groups is related to their contribution to chelating calcium and/or to their electron-withdrawing influence on acidity.
Asunto(s)
Resorción Ósea/tratamiento farmacológico , Calcinosis/tratamiento farmacológico , Difosfonatos/uso terapéutico , Acilación , Adsorción , Animales , Simulación por Computador , Difosfonatos/administración & dosificación , Difosfonatos/química , Difosfonatos/farmacología , Modelos Animales de Enfermedad , Femenino , Válvulas Cardíacas/efectos de los fármacos , Válvulas Cardíacas/metabolismo , Cetonas/química , Riñón/efectos de los fármacos , Riñón/metabolismo , Hígado/efectos de los fármacos , Hígado/metabolismo , Espectroscopía de Resonancia Magnética , Modelos Moleculares , Ratas , Espectrofotometría Atómica , Relación Estructura-Actividad , Tibia/efectos de los fármacos , Tibia/metabolismoRESUMEN
Various studies have shown a direct effect of beta estradiol on cartilage and bone. Such effects point to the possibility that specific receptors to estradiol exist in the growth plate cartilage as well as in bone. 3H-estradiol specific binding (EB) was therefore investigated in the supernatant of cartilage homogenates from the epiphyses and ribs of young growing mice and rats. High levels of EB were observed in the cytosol fraction of cartilage homogenates in the late fetal stage and in young rats and mice. The EB levels decreased gradually from late fetal stage up to 14 days of age in both groups of animals independent of their sex. Nuclear binding of 3H estradiol was also demonstrated by autoradiography in the chondrocytes of proliferating and hypertrophic zones. Estradiol binding was inhibited by high doses of unlabelled beta-estradiol, but not by alpha estradiol. Binding was also inhibited by tamoxifen and DES but not by testosterone. High levels of estradiol binding (EBS) were also observed in serum from young animals, but not in animals 2 months of age or older. Study of estradiol binding in cartilage and in serum of rats of the same age showed a significant difference in estradiol binding between these two systems. The difference in estradiol binding between serum and cartilage was seen in the response to inhibitors, Scatchard analysis, and temperature dependence. The results of our study imply that there are specific receptors for 17 beta estradiol in growth plate cartilage; they originate from chondrocytes, and their amount decreases with age. The effects of estradiol on endochondral bone growth seems therefore to be receptor mediated.
Asunto(s)
Envejecimiento/sangre , Envejecimiento/metabolismo , Estradiol/sangre , Estradiol/metabolismo , Placa de Crecimiento/metabolismo , Animales , Autorradiografía , Dietilestilbestrol/farmacología , Estradiol/análisis , Femenino , Placa de Crecimiento/química , Placa de Crecimiento/citología , Masculino , Ratones , Unión Proteica , Ratas , Receptores de Estradiol/análisis , Receptores de Estradiol/metabolismo , Tamoxifeno/farmacología , Factores de Tiempo , TritioRESUMEN
Blood bed and structure of the compact substance of the cat tibial diaphysis have been studied at normal conditions and after repeated effect of gravitational overloadings. Hypergravitation produced a noticeable rearrangement in the microcirculatory bed and in histoarchitectonics of the bone compact substance. The date obtained demonstrate that the changes produced depend on the number of parameters participating in the gravitational effect and on the position of the animal in the container, as well as on the time when the animal falls ill after the experiment.
Asunto(s)
Huesos/anatomía & histología , Gravitación , Tibia/irrigación sanguínea , Animales , Capilares/anatomía & histología , Gatos , Centrifugación , Femenino , Masculino , Microcirculación/anatomía & histología , Postura , Factores de TiempoRESUMEN
Effects of different stimuli upon the structure of the microcirculatory bed of the compact substance of long tubular bones were studied in 89 animals (rabbits and cats). Prolonged local pressure on the bone resulted in transformations characterized by lacunar resorption of the compact substance in the site of application of the force and endosteal bone-formation on the opposite side of the same part of the bone. Hypergravitation caused a considerable transformation of the microcirculatory bed and the histoarchitectonics of the bone compact substance. Smooth and linear perivascular resorption took place around dilated vessels in osteon canals. It resulted information of resorption cavities. In addition, processes of bone formation took place. Due to effects of hypokinesia the transformation of the compact substance is of a regressive character. The fibrous structures of the bony tissue and blood vessels lose their regular orientation and acquire chaotic direction.