Effects of hyperprolactinemia and ovariectomy on the tibial epiphyseal growth plate and bone formation in mice

PURPOSE: To evaluate the effects of ovariectomy and the hyperprolactinemia procedure in the tibial epiphyseal growth plate of female mice.METHODS: In this study, the epiphyseal growth plate of ovariectomized (OVX) and/or rendered hyperprolactinemic female mice by 50 days of treatment with 200 μg metoclopramide (M) was evaluated morphologically, morphometrically and immuno-histochemically. Forty female and adult mice were divided into four groups according to treatment: V group - animals treated with saline solution; H group - hyperprolactinemic animals; Ovx/V group - ovariectomized animals and treated with saline solution; Ovx/H group - hyperprolactinemic and ovariectomized animals. After the treatment period, the animals were sacrificed, tibia was removed and fixed in 10% buffered formalin and decalcified in 10% formic acid. The material was immersed in paraffin and subjected to histological processing in paraffin. The sections were stained with Masson's trichrome and immunohistochemistry was carried out for the pro-apoptotic protein BCL-2. The images for the morphological and morphometric study were analyzed with the imaging program AxioVision 4.8 (Carl-Zeiss(r), Germany).RESULTS: The combination of hyperprolactinemia and the ovariectomy procedure decreased the number of resting chondrocytes 1.5-fold, the number of proliferative chondrocytes 1.8-fold; the percentage of resting cartilage 2.4-fold and the percentage of trabecular bone 2.1-fold, compared with respective control animals.CONCLUSION: The procedure of ovariectomy combined with the metoclopramide-induced hyperprolactinemia in female mice has showed marked bone degeneration due to significant decrease of cell proliferation in the epiphyseal growth plate and bone formation.

Bone changes caused by experimental Solanum malacoxylon poisoning in rabbits

The aim of this study was to describe the bone changes observed after a daily oral administration of the calcinogenic plant Solanum malacoxylon (syn. S. glaucophyllum) (Sm) during 9 days. The Sm-poisoned rabbits had an increase of bone resorption in the endosteal surface of the cortical zone and also in the surface covered by osteoblasts of the primary and secondary spongiosa of the trabecular bone compartment. Moreover, the epiphyseal growth plates in long bones appeared narrower than in the control rabbits, with reduction of the proliferative and hyperthrophic chondrocyte zones. The electron microscopic study revealed a significant decrease of proteoglycans in the hyperthrophic chondrocyte zone evidenced by a significant reduction of rutenium red positive granules in the poisoned rabbit. Altogether, these data suggest that cell differentiation may play a pivotal role in the pathogenesis of Sm-induced bone lesions.

Contribution of matrix vesicles and alkaline phosphatase to ectopic bone formation

Endochondral calcification involves the participation of matrix vesicles (MVs), but it remains unclear whether calcification ectopically induced by implants of demineralized bone matrix also proceeds via MVs. Ectopic bone formation was induced by implanting rat demineralized diaphyseal bone matrix into the dorsal subcutaneous tissue of Wistar rats and was examined histologically and biochemically. Budding of MVs from chondrocytes was observed to serve as nucleation sites for mineralization during induced ectopic osteogenesis, presenting a diameter with Gaussian distribution with a median of 306 ± 103 nm. While the role of tissue-nonspecific alkaline phosphatase (TNAP) during mineralization involves hydrolysis of inorganic pyrophosphate (PPi), it is unclear how the microenvironment of MV may affect the ability of TNAP to hydrolyze the variety of substrates present at sites of mineralization. We show that the implants contain high levels of TNAP capable of hydrolyzing p-nitrophenylphosphate (pNPP), ATP and PPi. The catalytic properties of glycosyl phosphatidylinositol-anchored, polidocanol-solubilized and phosphatidylinositol-specific phospholipase C-released TNAP were compared using pNPP, ATP and PPi as substrates. While the enzymatic efficiency (k cat/Km) remained comparable between polidocanol-solubilized and membrane-bound TNAP for all three substrates, the k cat/Km for the phosphatidylinositol-specific phospholipase C-solubilized enzyme increased approximately 108-, 56-, and 556-fold for pNPP, ATP and PPi, respectively, compared to the membrane-bound enzyme. Our data are consistent with the involvement of MVs during ectopic calcification and also suggest that the location of TNAP on the membrane of MVs may play a role in determining substrate selectivity in this micro-compartment.

Particulate bioglass in the regeneration of alveolar bone in dogs: clinical, surgical and radiographic evaluations

Bone loss, either by trauma or other diseases, generates an increasing need for substitutes of this tissue. This study evaluated Bioglass as a bone substitute in the regeneration of the alveolar bone in mandibles of dogs by clinical, surgical and radiological analysis. Twenty-eight adult dogs were randomly separated into two equal groups. In each animal, a bone defect was created on the vestibular surface of the alveolar bone between the roots of the fourth right premolar tooth. In the treated group, the defect was immediately filled with bioglass, while in the control, it remained unfilled. Clinical evaluations were performed daily for a week, as well as x-rays immediately after surgery and at 8, 14, 21, 42, 60, 90 and 120 days post-operative. Most animals in both groups showed no signs of inflammation and wound healing was similar. Radiographic examination revealed a gradual increase of radiopacity in the region of the defect in the control group. In the treated group, initial radiopacity was higher than that of adjacent bone, decreasing until 21 days after surgery. Then it gradually increased until 120 days after surgery, when the defect became undetectable. The results showed that Bioglass integrates into bone tissue, is biocompatible and reduced the period for complete bone regeneration.

Bone mineral density of rat femurs after hindlimb unloading and different physical rehabilitation programs

Bone weakening can occur due to the absence of load on the skeleton or even short periods of decreased physical activity. Therefore, musculoskeletal diseases that involve temporary immobilization by casts, inactivity or tension increases the risk of fractures. Physical activity is the most studied procedure both to prevent damage and to restore bone structure. The present study aimed at evaluating, by bone densitometry on rat femurs, the influence of hindlimb unloading and later running activity on treadmill or free movement. Sixty-four Wistar rats were used, aged 65 days with a mean corporal mass of 316.11g, randomly divided into eight experimental groups: group 1, the suspended control with seven animals under hindlimb unloading regimen for 28 days, then euthanized; groups 2 and 3, the trained suspended comprising of 7 and five animals, respectively, subjected to hindlimb unloading for 28 days, followed by treadmill exercise for 28 days (group 2) or 56 days (group 3), then euthanized; groups 4 and 5, designated free suspended, comprised of 7 animals each under hindlimb unloading regimen for 28 days followed by free activity in cages for 28 days (group 4) or 56 days (group 5), then euthanized; groups 6, 7 and 8, negative controls, each with 8 animals allowed to free activity in cages and euthanized at the ages of 93, 121 and 149 days, respectively. Bone mineral density (BMD) of the left femur was analyzed by bone densitometry. Unloading by tail-suspension decreased BMD while treadmill training and free activity in cages promoted its recovery in a similar way and over time.

Functional Assessment of Cancer Therapy Bone Marrow Transplantation: tradução e validação

OBJETIVO: Traduzir para o português e validar o questionário de qualidade de vida Functional Assessment of Cancer Therapy - Bone Marrow Transplantation (FACT-BMT) em pacientes transplantados de medula óssea. OBJETIVO: O estudo foi realizado em Ribeirão Preto, SP, em 2005. O FACT-BMT (versão 3) traduzido e a versão em português do Short Form-36 Health Survey (SF-36) foram aplicados simultaneamente em 55 pacientes consecutivos com leucemia, submetidos ao transplante e em seguimento. Dois parâmetros clínicos foram utilizados para testar a sensibilidade do questionário: tempo decorrido do transplante e presença ou não de doença do enxerto contra o hospedeiro. Foi utilizada a análise de variância (ANOVA) com o teste post hoc de Tukey. Aplicou-se o coeficiente alfa de Cronbach, padronizado para todas as questões, escore final e domínios. RESULTADOS: A média de idade dos pacientes foi 34,8±8,1 anos, com escolaridade média de 10,8±4,7 anos, sendo 78,1% do sexo feminino. A duração média de tempo pós-transplante foi de 29,8±32,19 meses. Nenhuma alteração do formato original do questionário foi observada no final do processo de tradução e adaptação cultural. A consistência interna foi alta (0,88). A correlação entre o questionário traduzido e o SF-36 variou de 0,35 a 0,57, considerada de moderada a boa para a maioria dos domínios de qualidade de vida. A avaliação das validades de construto e concorrente foi satisfatória e estatisticamente significativa. CONCLUSÕES: A versão para o português do FACT-BMT foi validada satisfatoriamente para a aplicação em pacientes brasileiros de ambos os sexos submetidos ao transplante de medula óssea.

Risk factors for osteoporotic fractures and low bone density in pre and postmenopausal women

OBJECTIVE: To estimate the prevalence and analyze risk factors associated to osteoporosis and low-trauma fracture in women. METHODS: Cross-sectional study including a total of 4,332 women older than 40 attending primary care services in the Greater São Paulo, Southeastern Brazil, between 2004 and 2007. Anthropometrical and gynecological data and information about lifestyle habits, previous fracture, medical history, food intake and physical activity were obtained through individual quantitative interviews. Low-trauma fracture was defined as that resulting from a fall from standing height or less in individuals 50 years or older. Multiple logistic regression models were designed having osteoporotic fracture and bone mineral density (BMD) as the dependent variables and all other parameters as the independent ones. The significance level was set at p<0.05. RESULTS: The prevalence of osteoporosis and osteoporotic fractures was 33% and 11.5%, respectively. The main risk factors associated with low bone mass were age (OR=1.07; 95% CI: 1.06;1.08), time since menopause (OR=2.16; 95% CI: 1.49;3.14), previous fracture (OR=2.62; 95% CI: 2.08;3.29) and current smoking (OR=1.45; 95% CI: 1.13;1.85). BMI (OR=0.88; 95% CI: 0.86;0.89), regular physical activity (OR=0.78; 95% CI: 0.65;0.94) and hormone replacement therapy (OR=0.43; 95% CI: 0.33;0.56) had a protective effect on bone mass. Risk factors significantly associated with osteoporotic fractures were age (OR=1.05; 95% CI: 1.04;1.06), time since menopause (OR=4.12; 95% CI: 1.79;9.48), familial history of hip fracture (OR=3.59; 95% CI: 2.88;4.47) and low BMD (OR=2.28; 95% CI: 1.85;2.82). CONCLUSIONS: Advanced age, menopause, low-trauma fracture and current smoking are major risk factors associated with low BMD and osteoporotic fracture. The clinical use of these parameters to identify women at higher risk for fractures might be a reasonable strategy to improve the management of osteoporosis.

Screening the mutagenic activities of coomonly used antiparasite drugs by the Simultest, a simplified Salmonella/microsome plate incorporation assay

The mutagenic activities of 16 anti-parasite drugs were screened by the Simultest in both qualitative (spot test) and quantitative (plate incorporation) assays with a Salmonella typhimurium pool composed by the indicator strains TA97, TA 98, TA100 and TA102. Four anti Chagas' disease drugs (nifurtimox, benznidazole, CL 64,855, and MK 436) and two anti-amebae drugs (metronidazole and tinidazole) gave positive results in qualitative tests and incorporation of rat liver microsomes did not alter the results. Comparative dose response curves of the mutagenic activities of CL 64,855, metronidazole and benznidazole obtained by the simultest and by individual Salmonella indicator strains demonstrated that both approachs have similar sensitivities. The results corroborate the validity of the Simultest, as a simplified, fast and economic version of the Ames test in preliminary screening of potential mutagenic drugs.

Cytomegalovirus and other herpesviruses infections in heart and bone marrow transplant recipients

From January 1988 to January 1989 all the heart transplant and bone marrow recipients at the Instituto do Coração of the Hospital das Clínicas of the University of São Paulo Medical School were studied for the incidence and morbidity associated with herpesviruses infections after transplantation. Five bone marrow and 5 heart transplant recipients were followed for a mean of 4.2 months post-transplantation. All the patients were seropositive for cytomegalovirus (CMV) before admission and 80% experienced one or more recurrences during the observation period. Of the 12 episodes of CMV infection, that were identified in this study, 83% were accompanied by clinical or laboratory abnormalities. However, there was only one case of severe disease. The overall incidence of infection for herpes simplex (HSV) was 50%. Although most of HSV reactivations were oral or genital, one case of HSV hepatitis occurred. One of the 6 episodes of HSV infections that were treated with acyclovir showed an unsatisfactory response and was successfully managed with ganciclovir. All the individuals had anti-varicella zoster virus antibodies, but none of them developed infection. The study emphasizes the importance of active diagnostic surveillance of herpesvirus infections in transplant patients. Both CMV and HSV reactivations showed high incidence and important morbidity and thus, deserve prophylactic therapy.

Aspergillosis in immunocompromised children acute myeloid leukemia and bone marrow aplasia.: Report of two cases

Two cases of Aspergillosis in immunocompromised children are reported. Both were caused by Aspergillns flavus. Early diagnosis and treatment led to the remission of the process. One patient had acute myeloid leukemia; the fungus was isolated from the blood. The other patient with bone marrow aplasia, presented an invasive aspergillosis of the paranasal sinuses with dissemination of fungal infection; the diagnosis was obtained by histology and culture of biopsied tissue from a palatal ulceration.

Studies on prevalence of Strongyloides infection in Holambra and Maceió, Brazil, by the agar plate faecal culture method

Prevalence of Strongyloides stercoralis infection in three areas of Brazil was surveyed by a recently developed faecal culture method (an agar plate culture). The Strongyloides infection was confirmed in 11.3% of 432 subjects examined. The diagnostic efficacy of the agar plate culture was as high as 93.9% compared to only 28.5% and 26.5% by the Harada-Mori filter paper culture and faecal concentration methods, when faecal samples were examined simultaneously by these three methods. Among the 49 positive samples, about 60% were confirmed to be positive only by the agar plate culture. These results indicate that the agar plate culture is a sensitive new tool for the correct diagnosis of chronic Strongyloides infection.

Scedosporium apiospermum SINUSITIS AFTER BONE MARROW TRANSPLANTATION: REPORT OF A CASE

A forty-year-old man underwent an allogeneic BMT from his HLA identical sister. GvHD prophylaxis was done with cyclosporine (CyA), methotrexate and prednisone (PDN). On day +90 extensive GvHD was noted and higher doses of immunosuppressive drugs alternating CyA with PDN were initiated. Patient's follow-up was complicated by intermittent episodes of leukopenia and monthly episodes of sinusitis or pneumonia. One year after BMT, the patient developed hoarseness and nasal voice. No etiologic agent could be identified on a biopsy sample of the vocal chord. Upon tapering the doses of immunosuppressive drugs, the patient had worsening of chronic GvHD and was reintroduced on high doses of cyclosporine alternating with prednisone on day +550. Three months later, GvHD remained out of control and the patient was started on azathioprine. On day +700, hoarseness and nasal voice recurred. Another biopsy of the left vocal chord failed to demonstrate infection. Episodes of sinusitis became more frequent and azathioprine was withheld 3 months after it was started. One month later, the patient had bloody nasal discharge and surgical drainage of maxillary sinuses was performed. Histopathology showed hyphae and cultures grew Scedosporium apiospermum. ltraconazole 800 mg/day was initiated. The patient developed progressive respiratory failure and died 15 days later.

Phialemonium curvatum infection after bone marrow transplantation

We report a case of cutaneous infection caused by Phialemonium curvatum GAMS et COOKE, 1983, after bone marrow transplantation. The genus Phialemonium was created by GAMS & MCGINNIS in 1983 including three new species: Ph. obovatum, Ph. curvatum and Ph. dimorphosporum, and represents an intermediate genus between Acremonium and Phialophora. Nowadays, the genus Phialemonium is considered to be a pheoid fungus which may cause the eventual lesions observed in pheo- and hyalohyphomycosis. Species of this genus have been described as opportunistic agents in humans and animals, mainly as a result of immunosuppression. In the present case, the patient had multiple myeloma and received an allogenic bone marrow transplant from his HLA-compatible brother. Two months after transplantation, he developed purplish and painful nodular lesions on the right ankle. Some of these lesions drained spontaneously and apparently hyaline mycelial filaments were observed, whose culture was initially identified as Acremonium sp. Subsequent studies showed that the fungus was Phialemonium curvatum. The infection was treated with amphotericin B, followed by ketoconazole. The patient was submitted to surgical debridement followed by two skin grafts to repair the bloody area. The duration of the treatment was 4 months and secondary prophylaxis with ketoconazole alone was maintained for one additional month. No recurrence was observed after discontinuation of treatment. The authors comment on the pathogenicity of the genus Phialemonium.