Micronuclei formation in liver fibrosis samples from patients infected by hepatitis C virus.

Genetic research on fibrosis outset and its progression in chronic hepatitis (CH) by hepatitis C virus (HCV) are limited. The lack of cytogenetic data led us to investigate the presence of micronuclei (MNi), as a sign of genomic damage. Hepatocytes of hepatic parenchyma from 62 cases diagnosed with CH associated with HCV and displaying different degrees of fibrosis (F1-F4) were analyzed. These data were compared to 15 cases without fibrosis (F0). Twelve healthy liver parenchyma samples were included as control. All samples were obtained from paraffin-embedded archival material. Micronucleated hepatocytes (MN-Heps) were analyzed through Feulgen/Fast-green staining. Results showed that the rates of MN-Heps in the F4 group were statistically significant (p < 0.05) and higher than those in the control group. Like results were also obtained on comparing F4 with F0, F1, F2 and F3 cases. Conversely, differences were not significant (p > 0.05) on comparing F0, F1, F2, F3, one against the other, as well as individual versus control. Although chromosomal losses in CH were detected, it was shown that liver parenchyma with fibrosis in the initial stages (F1-F3) cannot be considered cytogenetically abnormal.

Micronuclei formation in liver fibrosis samples from patients infected by hepatitis C virus

Genetic research on fibrosis outset and its progression in chronic hepatitis (CH) by hepatitis C virus (HCV) are limited. The lack of cytogenetic data led us to investigate the presence of micronuclei (MNi), as a sign of genomic damage. Hepatocytes of hepatic parenchyma from 62 cases diagnosed with CH associated with HCV and displaying different degrees of fibrosis (F1-F4) were analyzed. These data were compared to 15 cases without fibrosis (F0). Twelve healthy liver parenchyma samples were included as control. All samples were obtained from paraffin-embedded archival material. Micronucleated hepatocytes (MN-Heps) were analyzed through Feulgen/Fastgreen staining. Results showed that the rates of MN-Heps in the F4 group were statistically significant (p < 0.05) and higher than those in the control group. Like results were also obtained on comparing F4 with F0, F1, F2 and F3 cases. Conversely, differences were not significant (p > 0.05) on comparing F0, F1, F2, F3, one against the other, as well as individual versus control. Although chromosomal losses in CH were detected, it was shown that liver parenchyma with fibrosis in the initial stages (F1-F3) cannot be considered cytogenetically abnormal.

Successful implant of long-term cryopreserved parathyroid glands after total parathyroidectomy.

BACKGROUND: Parathyroid cryopreservation is essential in some cases of parathyroid surgery. The fate of autografted tissue after long-term cryopreservation is not fully discussed in the literature. METHODS: The successful experience with the use of parathyroid tissues preserved for 21 months and 30 months is reported. RESULTS: Both patients were women with renal hyperparathyroidism who underwent total parathyroidectomy without autotransplantation. Patient 1 was a 40-year-old woman. At 21 months of follow-up, her parathyroid hormone (PTH) level was undetectable, and despite oral calcium supplements, she was hypocalcemic. Forty-five cryopreserved fragments were thawed and implanted in her forearm. Calcium levels improved, and PTH steadily increased in both arms. PTH levels at 18 months after the autograft were 37.0 pg/mL in the contralateral arm and 1150.0 pg/mL in the implant arm. Patient 2 was a 44-year-old woman. After 30 months, her PTH was undetectable, and she underwent cryopreserved tissue implantation. CONCLUSION: These cases show that parathyroid tissue may remain viable even after long-term storage.

Detection of micronuclei formation and nuclear anomalies in regenerative nodules of human cirrhotic livers and relationship to hepatocellular carcinoma.

Human cirrhosis is considered an important factor in hepatocarcinogenesis. The lack of substantial genetics and cytogenetics data in human cirrhosis led us to investigate spontaneous micronuclei formation, as an indicator of chromosomal damage. The analysis was performed in hepatocytes of regenerative, macroregenerative, and tumoral nodules from 30 cases of cirrhosis (paraffin-embedded archival material), retrospectively selected: cryptogenic, hepatitis C virus, and hepatitis C virus associated with hepatocellular carcinoma (HCC). Thirteen control liver samples of healthy organ donors were included. Micronucleated hepatocytes were analyzed with Feulgen-fast-green dyeing techniques. The spontaneous frequency of micronucleated hepatocytes in both regenerative and macroregenerative nodules of all cirrhotic patients was significantly higher than for the normal control group. There was no significant difference in frequency of micronucleated hepatocytes in regenerative nodules compared with macroregenerative nodules for all cases analyzed, whereas a significantly higher frequency of micronucleated hepatocytes was detected in tumoral nodules, compared with cirrhotic regenerative nodules and normal parenchyma. A higher frequency of the nuclear anomalies termed broken-eggs was observed in hepatitis C virus-related samples. Chromatinic losses and genotoxicity already existed in the cirrhotic regenerative nodules, which might predispose to development of HCC.

Architectural remodelling in lungs of rabbits induced by type V collagen immunization: a preliminary morphologic model to study diffuse connective tissue diseases.

The pathogenesis of diffuse connective tissue diseases (DCTD) is still unknown and has been extensively studied regarding its autoimmunity aspects related to extracellular matrix (ECM) remodelling, with an emphasis on the collagens at the inflammatory site. The present paper describes the pulmonary architectural and repair/remodelling responses to injury after immunization of rabbits with human type V collagen. The animal model consisted of rabbits immunized with collagen mixed with Freund's adjuvant and sacrificed 7, 15, 30, 75, and 120 days after the first of four doses of antigen. Pulmonary architecture remodelling response was evaluated by histology, morphometry, and the immunofluorescence method, according to compartments of reference (parenchyma and interstitium) and injury: 1 inflammation (polymorphonuclear and mononuclear cells); 2-repair (fibrosis) and 3-ECM remodelling (collagen system). The results showed an intense inflammatory involvement of the pulmonary vascular and bronchiolar parenchyma, characterized by increased wall thickness in small arteries, infiltrations by pseudoeosinophils, and mononuclear cells. Progressive remodelling of the pulmonary ECM was characterized by collagen deposition in the septal and bronchovascular interstitium, especially in rabbits sacrifices at 75 and 120 days. The ECM remodelling process was not reproduced when rabbits were inoculated with collagen types I and III. We conclude that the model reproduces morphologic changes similar to those observed in many DCTD, encouraging realization of other experiments to gain a better understanding of the pathogenesis of these diseases.

[Experimental tumors of the central nervous system: standardisation of a model in rats using the 9L glioma cells]. / Tumores experimentais do sistema nervoso central: padronização de modelo em roedores utilizando a linhagem 9L.

A number of experimental models have been established during the last decades in order to study tumor biology and the effects of treatment or manipulation of the microenvironment of malignant glial tumors. Even though those models have been well characterised and are, to a certain extent, easily reproducible, there are limitations as to their use and to the interpretation of the results. The aim of this study is to standardize a model of a malignant glial tumor and detect possible events able to modify its development. 9L cells were inoculated intracerebrally in 48 Sprague-Dawley rats; from these, 25 animals were also implanted with a device containing electrodes for the registration of the electroencephalogramm. Animals were daily evaluated by neurologic examination. Twenty four animals developed tumor - 10 animals died either in the immediate pos-operatory period or during evolution; 14 animals did not develop tumor. Macroscopically the tumor was well demarcated from the adjacent brain; by light microscopy the tumor exhibited malignant characteristics as well as extensive infiltration of the brain parenchyma. Diagnosis was that of a malignant astrocytoma. The use of the stereotaxic frame and care to infuse a small volume of liquid containing cells during a period of 120 seconds were the most important procedures to obtain sucess in the model. Additional care should be taken in counting cells in the Neubauer camera and in maintaining cells in constant agitation before injecting the tumor-containing solution. The model here developed was efficient besides being of low cost and of relatively easy execution.

Tumores experimentais do sistema nervoso central: padronização de modelo em roedores utilizando a linhagem 9L / Experimental tumors of the central nervous system: standardisation of a model in rats using the 9L glioma cells

Grande variedade de modelos experimentais foram estabelecidos em neuro-oncologia durante as últimas décadas, a fim de estudar a biologia tumoral e a eficiciência de novas drogas e novos tratamentos em gliomas malignos humanos. Embora estes modelos estejam bem caracterizados e sejam facilmente reproduzíveis e aplicáveis, háá limitações quanto ao seu uso e a resposta obtida, principalmente quando utilizados para monitorização de tratamentos. A proposta deste estudoé padronizar modelo de tumor cerebral glial experimental e detectar variáveis que possam influenciar o seu desenvolvimento. Células 9L foram inoculadas na substância branca do cérebro em 48 ratos machos Sprague-Dawley; destes, 25 animais receberam eletrodos corticais para realização de eletroencefalografia. Os animais eram avaliados diariamente por observação neurológica. Vinte e quatro ratos desenvolveram tumor - 10 animais morreram ou no pós-operatório imediato ou durante a evolução; 14 animais não desenvolveram tumor. Macroscopicamente, o tumor parecia bem delimitado; características de malignidade e extensa infiltração foram observadas a microscopia de luz; o diagnóstico foi o de astrocitoma maligno. A utilização de técnica estereotóxica e a infusão de pequeno volume de suspensão celular por um tempo longo de infusão foram considerados importantes para o bom desenvolvimento do tumor. Procedimentos que merecem atenção são: contagem precisa das células na camara de Neubauer, agitação constante da suspensão de células a serem inoculadas e fixação precisa da profundidade de inoculação. O modelo experimental desenvolvido no presente trabalho mostrou ser de execução relativamente fácil, com custo razoável e reprodutível