Histological and radiographic evaluation of the muscle tissue of rats after implantation of bone morphogenic protein (rhBMP-2) in a scaffold of inorganic bone and after stimulation with low-power laser light.

Objective: The present study histologically and radiologically evaluates the muscle tissue of rats after implantation of bone morphogenic protein (rhBMP-2) in a natural inorganic bone mineral scaffold from a bull calf femur and irradiation with low-power light laser. Materials and Methods: The right and left hind limbs of 16 rats were shaved and an incision was made in the muscle on the face corresponding to the median portion of the tibia, into which rhBMP-2 in a scaffold of inorganic bone was implanted. Two groups of limbs were formed: control (G1) and laser irradiation (G2). G2 received diode laser light applied in the direction of the implant, at a dose of 8 J/cm2 for three minutes. On the 7th, 21st, 40th and 112th days after implantation, hind limbs of 4 animals were radiographed and their implants removed together with the surrounding tissue for study under the microscope. The histological results were graded as 0=absence, 1=slight presence, 2=representative and 3=very representative, with regard to the following events: formation of osteoid structure, acute inflammation, chronic inflammation, fibrin deposition, neovascularization, foreign-body granuloma and fibrosis. Results: There were no statistically significant differences in these events at each evaluation times, between the two groups (P>0.05; Mann-Whitney test). Nevertheless, it could be concluded that the natural inorganic bone matrix with rhBMP-2, from the femur of a bull calf, is a biocompatible combination. Conclusions: Under these conditions, the inductive capacity of rhBMP-2 for cell differentiation was inhibited. There was a slight acceleration in tissue healing in the group that received irradiation with low-power laser light.

Microwave-induced fast decalcification of rat bone for electron microscopic analysis: an ultrastructural and cytochemical study

Bone decalcification is a time-consuming process. It takes weeks and preservation of the tissue structure depends on the quality and velocity of the demineralization process. In the present study, a decalcification methodology was adapted using microwaving to accelerate the decalcification of rat bone for electron microscopic analysis. The ultrastructure of the bone decalcified by microwave energy was observed. Wistar rats were perfused with paraformaldehyde and maxillary segments were removed and fixed in glutaraldehyde. Half of specimens were decalcified by conventional treatment with immersion in Warshawsky solution at 4ºC during 45 days, and the other half of specimens were placed into the beaker with 20 mL of the Warshawsky solution in ice bath and thereafter submitted to irradiation in a domestic microwave oven (700 maximum power) during 20 s/350 W/±37ºC. In the first day, the specimens were irradiated 9 times and stored at 40ºC overnight. In the second day, the specimens were irradiated 20 times changing the solution and the ice after each bath. After decalcification, some specimens were postfixed in osmium tetroxide and others in osmium tetroxide and potassium pyroantimonate. The specimens were observed under transmission electron microscopy. The results showed an increase in the decalcification rate in the specimens activated by microwaving and a reduction of total experiment time from 45 days in the conventional method to 48 hours in the microwave-aided method.

A preservação da estrutura de ossos é dependente da qualidade e da velocidade em que ocorre o processo de desmineralização. Neste estudo foi observada a ultraestrutura de maxila de rato descalcificada utilizando microondas. Ratos Wistar sofreram perfusão com paraformaldeído e o segmento de maxila retirado e fixado em glutaraldeído. Após esta etapa algumas amostras foram descalcificadas por imersão em solução de Warshawsky durante 45 dias a 4(0)C. Outras amostras foram submetidas a irradiação por microondas (forno de microondas doméstico 700 Watts de potência), durante 20 s/350 W/ ± 37ºC. No primeiro dia foram realizadas um total de 9 irradiações e os espécimes foram deixadas posteriormente a 4ºC por 12 h na solução descalcificadora sem agitação. No segundo dia, os fragmentos foram submetidos à nova irradiação totalizando 20 banhos, trocando-se a solução e o gelo a cada banho. A seguir algumas amostras foram pós-fixadas com tetróxido de ósmio e outras com tetróxido de ósmio e piroantimonato de potássio. As amostras foram observadas em microscópio eletrônico de transmissão. Os resultados mostraram que o processo de descalcificação ativado por microondas reduziu para 48 h o período de descalcificação, o qual pelo método tradicional ocorre em 45 dias.