RESUMO
Acquired haemophilia is a rare bleeding disorder usually caused by the spontaneous formation of inhibitory antibodies to coagulation FVIII. The disease occurs most commonly in the elderly, and although acquired haemophilia may be associated with a variety of underlying conditions, up to 50% of reported cases are idiopathic. Treatment options have traditionally involved human FVIII or FIX replacement therapy (if the inhibitor titre allows), porcine FVIII or the use of activated pro-thrombin complex concentrates. Recombinant activated coagulation FVII (rFVIIa) was available on an emergency and compassionate use basis from 1988 to 1999 at sites in Europe and North America. It has been registered in Europe for use in treating acquired haemophilia since 1996 and has recently been licensed for this indication in the United States. By directly activating FX on the surface of activated platelets at the site of injury (thereby bypassing FVIII and FIX), rFVIIa can circumvent the actions of inhibitory antibodies present in acquired haemophilia patients. This paper provides an overview of experiences with rFVIIa for the treatment of acquired haemophilia from the NovoSeven compassionate and emergency use programmes (1989-1999), the Hemophilia and Thrombosis Research Society Registry, and independent published reports from January 1999 to September 2005. rFVIIa has been reported to provide safe and effective haemostasis as a first line therapy in patients of all ages for a variety of surgical and non-surgical bleeding situations.
Assuntos
Fator VII/uso terapêutico , Hemofilia A/tratamento farmacológico , Hemorragia/prevenção & controle , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Relação Dose-Resposta a Droga , Fator VIIa , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas Recombinantes/uso terapêutico , Resultado do TratamentoRESUMO
BACKGROUND: Neuroblastoma is a childhood tumor that often displays unusual biological behavior. The tumor may present with widespread metastases that are unresponsive to aggressive treatment. At other times, both the metastases and the primary tumor may spontaneously regress without treatment. Apoptosis, or programmed cell death, is thought to play a role in the dichotomous behavior of neuroblastoma. We hypothesize that neuroblastoma cells will interact with host tissues to release mediators that affect apoptosis. MATERIALS AND METHODS: Human neuroblastoma cells and human Chang hepatocytes are grown in a noncontact, coculture system. After incubation for 4 days, the medium from the coculture system is collected. Neuroblastoma cells and Chang hepatocytes are then plated separately with the conditioned medium and their own standard growth medium as controls. After 4 days, these cells are harvested and cytospins made for immunostaining. Tumor necrosis factor alpha (TNF-alpha), Fas ligand, and Bcl-2, are measured with immunohistochemistry. Apoptosis is detected with the TUNEL method. Immunostaining data are interpreted with computer image analysis and reported as stain index. TUNEL data are reported as percentage apoptotic cells. All data are reported as means +/- SEM. Statistical analysis is performed and P < 0.05 considered significant. RESULTS: Chang hepatocytes grown in the coculture conditioned media have an increase in TNF-alpha and Fas ligand. The neuroblastoma cells have a significant decrease in Fas ligand. There is a significant increase in the number of apoptotic hepatocytes when they are cultured in the conditioned media. In contrast, the neuroblastoma cells grown in the coculture conditioned media show no increase in apoptosis. Finally, Bcl-2 is significantly increased in the neuroblastoma cells cultured in the conditioned media. CONCLUSIONS: Neuroblastoma cells grown in coculture conditioned media show increased expression of Bcl-2 and decreased Fas ligand levels. These changes should diminish apoptosis activity in the tumor cells. In contrast, the conditioned media induce elevated levels of proapoptotic mediators in the Chang hepatocytes. A tumor's ability to successfully metastasize may be dependent on mediators generated in the tumor-host interaction, and may not be just an independent characteristic of the tumor itself.
Assuntos
Apoptose , Técnicas de Cultura de Células/métodos , Meios de Cultivo Condicionados/farmacologia , Fígado/citologia , Neuroblastoma , Comunicação Celular/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Proteína Ligante Fas , Humanos , Citometria por Imagem , Marcação In Situ das Extremidades Cortadas , Glicoproteínas de Membrana/análise , Proteínas Proto-Oncogênicas c-bcl-2/análise , Software , Células Tumorais Cultivadas/química , Células Tumorais Cultivadas/citologia , Fator de Necrose Tumoral alfa/análise , Receptor fas/análiseRESUMO
BACKGROUND: We have previously demonstrated an increase in hepatocyte apoptosis when they are cocultured with neuroblastoma cells. Death receptors in the tumor necrosis factor (TNF) family such as TNFR1 and Fas have been identified as regulators of apoptosis and may be responsible for the altered regulation of apoptosis seen in our coculture model. To evaluate the effects of released factors and remove the potential alterations induced by direct contact, a noncontact coculture system was used to study the interaction between hepatocytes and neuroblastoma cells. METHODS: Human Chang hepatocytes (HC) were plated onto Falcon cell culture inserts with 0.45-micrometer pores in the permeable membrane. Human neuroblastoma cells (NB-IMR-32) were seeded into wells of the Falcon companion plate. After 24 h, inserts containing HC were placed into wells containing NB cells and incubated for 4 days. This provided a coculture environment without actual cellular contact. Immunohistochemical staining for TNFalpha, Fas, and Fas ligand (Fas-L) was performed. Apoptosis was detected via the TUNEL method. Images were analyzed with ImagePro-Plus. Statistical analyses were done with significance determined at P < 0.05. RESULTS: Chang hepatocytes demonstrated a significant increase in the levels of TNF, Fas, and Fas-L when cocultured with neuroblastoma cells (P < 0.005). In addition, the cocultured hepatocytes had a 20-fold increase in the apoptotic rate (P < 0.001). Neuroblastoma cells had no demonstrable level of Fas or TNF when grown alone and in cocultures. Neuroblastoma cells that were grown alone had an elevated level of Fas-L, but this level diminished by 44% when cocultured with hepatocytes (P < 0.001). CONCLUSION: An upregulated TNF/Fas receptor-ligand system may be responsible for increased apoptosis in hepatocytes when cocultured with neuroblastoma. This upregulation may be due to release of neuroblastoma-derived Fas ligand into the media. Tumors may alter the regulation of apoptosis in surrounding tissues via the death receptors.