Astrocyte-derived GDNF is a potent inhibitor of microglial activation

• Autor(a) principal: Rocha, Sandra Catarina Moreira
• Data de Publicação: 2010
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10400.6/3940
• topic: Doença de Parkinson; Microglia; Astrócitos; Neuroinflamação; Domínio/Área Científica::Engenharia e Tecnologia::Engenharia Química

Parkinson’s disease is characterized by the selective loss of dopaminergic neurons in the substantia nigra pars compacts. The aetiology of this disease is not completely clarified; however several hypotheses have been advanced regarding the loss of dopaminergic neurons. Among them, neuroinflammation has been increasingly recognized as a major factor in the pathogenesis of Parkinson’s disease, and increasing evidence suggests that microglial cells are a predominant source of inflammation contributing for the dopaminergic neurodegeneration. Astrocytes play vital roles in the maintenance of the normal brain function and diverse studies suggest that they could act as physiological regulators preventing excessive inflammatory microglial responses. However, little is known regarding how astrocytes may modulate the microglial activation. Due to the relevance of astrocytes-microglia interactions in the regulation of brain inflammation, it is important to identify the mediators involved in this process, which could act as natural anti-inflammatory agents in the brain. In this way, the major goal of the present work was to evaluate of the effect of soluble mediators release by astrocytes on microglial activation induced by the inflammatory agent Zymosan A, as well as to identify the nature of these mediators. For the determination of the effect of these molecules in the microglial activity, ventral midbrain microglial primary cultures were previously exposed to astrocytes conditioned media (or culture medium – control), and then treated with 5 μg/mL ofZymosan. Studies previously made indicated that this concentration of Zymosan provokes an accented increase of the microglia phagocytic activity and increased ROS generation, showing no citotoxici effect to the cells. However, the pre-incubation of the microglial cells with astrocytes conditioned media was capable to prevent the characteristic increase of the phagocytic activity and ROS production induced by Zymosan A, which levels remained at control levels. To evaluate the nature of the soluble mediators release by astrocytes able to prevent the microglial activation, specific antibodies recognizing some neurotrophics factors known by its neuroprotectors properties of substantia nigra. were used to block their action on the astrocytes conditioned media. Those antibodies were: anti-GDNF, anti-CDNF and anti condicioned media, we observed that the glial cells line-derived neurotrophic factor (GDNF) seems to be a soluble mediator capable to completely prevent the microglial activation induced by Zymosan A, whereas, the remaining mediators do not exerted an effect on microglial activation. To confirm this fact, specific knockdown of GDNF was achived in astrocytes cell cultures and the resultant condicined medium from this cultures, when applied to microglia cultures before Zymosan A, were not capable to prevent its activation. Finally, to clarify if this effect was an isolated of GDNF or if other molecules were laso involved the levels of GDNF on the astrocytes conditioned media were quntidied by ELISA assay. Based on the obtained values, three concentrations of GDNF (100 pg/mL, 200 pg/mL and 400 pg/mL), diluted in culture medium, were tested to verify its capability to prevent microglia activation induced by Zymosan A. The results have shown that the three concentrations of GDNF were capable to suppress microglial activation induced by Zymosan A. However, the achieved protection was not in a dose dependent manner, as initially expected. Taken together, the results obtained in this work demonstrate that GDNF a neurotrophic factor expressed by astrócitos, has the capacity to modulate the microglial inflammatory response. In this way, GDNF could be use to develop a potencial therapy to prevent neuroinflammation, and in this way contributing to the reduction of the development of Parkinson’s Disease pathogenesis.