Sérgio Olavo Petenusci

Bone quality associated with daily intake of coffee: a biochemical, radiographic and histometric study

Caffeine induces loss of calcium and influences the normal development of bone. This study investigated the effects of coffee on bone metabolism in rats by biochemical measurement of calcium, bone densitometry and histometry. Male rats, born of female treated daily with coffee and with coffee intake since born, were anesthetized, subjected to extraction of the upper right incisor, and sacrificed 7, 21 and 42 days after surgery. Blood and urine samples were taken, and their maxilla radiographed and processed to obtain 5-µm-thick semi-serial sections stained with hematoxylin and eosin. The volume and bone quality were estimated using an image-analysis software. The results showed significantly greater amount of calcium in the plasma (9.40 ± 1.73 versus 9.80 ± 2.05 mg%) and urine (1.00 ± 0.50 versus 1.25 ± 0.70 mg/24 h) and significantly less amount in bone (90.0 ± 1.94 versus 86.0 ± 2.12 mg/mg bone), reduced bone mineral density (1.05 ± 0.11 versus 0.65 ± 0.15 mmAL), and lower amount of bone (76.19 ± 1.6 versus 53.41 ± 2.1 %) (ANOVA; p≤0.01) in animals treated with coffee sacrificed after 42 days. It may be concluded that coffee/caffeine intake caused serious adverse effects on calcium metabolism in rats, including increased levels of calcium in the urine and plasma, decreased bone mineral density and lower volume of bone, thus delaying the bone repair process.

Streptozotocin-induced diabetes: significant changes in the kinetic properties of the soluble form of rat bone alkaline phosphatase.

A soluble form of an alkaline phosphatase, obtained from the osseous plate of streptozotocin-induced diabetic rats, was purified 90-fold with a yield of 26%. The calculated Mr of the purified enzyme was 80,000 by denaturing polyacrylamide gel electrophoresis and 160,000 by gel filtration on Sephacryl S-300, suggesting a dimeric structure for its native form. In the absence of metal ions, the p-nitrophenylphosphatase activity of the purified enzyme was 4223.1 U/mg. Magnesium or calcium ion concentrations up to 2 mM increased the specific activity of the enzyme to 9896.5 and 10,796.2 U/mg, respectively. The enzyme was stimulated to a lesser extent by MnCl2 (5390.1 U/mg) and CoCl2 (5088.2 U/mg). The purified soluble alkaline phosphatase showed a broad substrate specificity, and among the less hydrolyzed substrates were pyrophosphate (1517.6 U/mg) and bis-p-nitrophenylphosphate (499.6 U/mg). The enzyme was relatively stable at 45 degrees for periods as long as 180 min, but was denatured rapidly above 50 degrees, following first order kinetics with T1/2 values ranging from 3.5 to 57.7 min. The results reported herein suggested that the soluble form of alkaline phosphatase from streptozotocin-induced diabetic rats had its kinetic properties altered, apparently as a consequence of changes in metal-binding properties.

HEPATOTOXICIDADE DE PLANTAS MEDICINAIS. XIV. AÇÃO DA INFUSÃO DE ARTEMISIA VULGARIS L. NO RATO

O objeto deste trabalho foi estudar a hepatotoxicidade da Artemisia vulgaris L., administrada em doses elevadas e doses baixas sob a forma de infusao. Foram usados ratos albinos Wistar machos, pesando 120 g em media, que receberam no bebedouro infusao de artemisia (1,39 g/125 ml ou 250 mg/125 ml de agua), durante 7 dias. Pouco antes do sacrificio pelo eter, fragmentos de figado foram colhidos e fixados em formol a 10% durante 24 h. Apos inclusao em parafina, as pecas foram cortadas com 6 mm de espessura e coradas pela hematoxilina e eosina. A estrutura hepatica foi analisada ao microscopio de luz e avaliada cariometrica e estereologicamente. Ao exame histopatologico, os cortes de figado mostraram areas de hepatocitos volumosos, com citoplasma abundante, granuloso e vacuolado. A veia centrolobular estava dilatada e o espaco-porta desorganizado com vasos dilatados e congestos, alem de colangiocitos menores com nucleos de cromatina condensada na periferia. Os ratos que receberam doses menores mostraram poucas alteracoes. Cariometricamente, os hepatocitos apresentaram nucleos semelhantes aos do controle, porem mais alongados; os colangiocitos mostraram nucleos semelhantes aos do controle. Estereologicamente, os hepatocitos dos ratos tratados com artemisia eram mais volumosos, com citoplasma abundante, e em menor numero por mm3 nos grupos tratados. Em suma, o fitoterapico em dose excessiva provocou no figado uma lesao celular reversivel (alteracao hidropica); em dose menor, houve lesao desprezivel e perfeitamente reversivel.

Hepatotoxicidade de plantas medicinais. LIII. Ação da infusão de Tabebuia avellanedae lor. ex griseb. no rato

A infusao da casca (10 g de planta em 1 litro de agua) de Tabebuia avellanedae Lor. ex Griseb. mostrou hepatotoxicidade significante no rato, quando administrada na agua do bebedouro durante 20 dias. Foram observadas alteracoes traduzidas por vasos e sinusoides dilatados e congestos, os hepatocitos eram mais volumosos com citoplasma granuloso grosseiro e vacuolado, com nucleos de menor volume. O espaco-porta estava desorganizado e foram observados focos de inflamacao cronica. Cariometricamente, os nucleos dos hepatocitos eram menos volumosos e mais arredondados. Estereologicamente, foi possivel observar um aumento dos volumes celular e citoplasmatico e menor quantidade de celulas por mm3.

Hepatotoxicidade em ratos tratados com dipirona (1-fenil-2,3-dimetil-5-pirazolona-4-metilamino-metanossulfonato de sódio)

A dipirona e um analgesico, antitermico e espasmolitico. O objetivo do presente trabalho foi estudar a hepatotoxicidade desta droga. Para tal, foram utilizados 16 ratos Wistar femeas, pesando 200 g em media. Durante 90 dias as ratas receberam via intraperitoneal 16 mg de dipirona. Apos a eutanasia pelo pentobarbital sodico a 3%, foram colhidos fragmentos de figado, os quais foram fixados em formol a 10% durante 24 horas. Apos desidratacao, diafanizacao e inclusao em parafina, as pecas foram seccionadas com 6 μm de espessura e os cortes, corados com hematoxilina e eosina. A estrutura hepatica foi examinada ao microscopio de luz e avaliada cariometrica e estereologicamente. Ao exame histologico, o figado dos animais tratados com dipirona mostrou areas com hepatocitos volumosos, granulares e vacuolizados, provavelmente repletos de glicogenio, alem de espaco-porta desorganizado. Cariometricamente, os nucleos dos hepatocitos e colangiocitos eram maiores sem modificarem a forma. Estereologicamente, os hepatocitos eram muito semelhantes aos dos animais de controle. E possivel concluir que a dipirona provocou no rato o aparecimento de algumas areas com hepatocitos volumosos, granulares e vacuolados, provavelmente repletos de glicogenio, e desorganizacao do espaco-porta. Em suma, a droga causou pequenas alteracoes hepaticas, provavelmente reversiveis.

Streptozotocin-induced diabetes influences the activity of ecto-nucleoside triphosphate diphosphohydrolase 1 of rat osseous plate membranes

We report the kinetic characterization of an ecto-nucleosidetriphosphate diphosphohydrolase 1 from rat osseous plate membranes in streptozotocin-induced diabetic rats, which arises during ectopic mineralization twenty days after a subcutaneous implantation of demineralized bone matrix, Insulin deficiency decreased the ecto-nucleoside triphosphate diphosphohydrolase activity from 1293.1 ± 39.8 (control rats) to 556.0 ± 8.2 nmol Pi/(min mg). Two families of ATP hydrolyzing sites showed cooperative effects with specific activities of 256.2 ± 7.7 nmol Pi/(min mg) and 299.8 ± 8.9 nmol Pi/(min mg), and studies on the stimulation of the enzyme by magnesium and calcium ions showed that the decrease in enzyme activity results from changes in the affinity of the enzyme for these ions. To our knowledge this is the first study associating the effects of type I diabetes with an ecto-nucleoside triphosphate diphosphohydrolase activity from rat osseous plate membranes. (Mol Cell Biochem 267: 99–106, 2004)