Neurite outgrowth is impaired on HSP70-positive astrocytes through a mechanism that requires NF-kappaB activation.

In the adult central nervous system (CNS), prominent reactive astrocytosis is seen in acute traumatic brain injury, neurodegenerative diseases and a variety of viral infections. Reactive astrocytes synthesize a number of factors that could play different roles in neuronal regeneration. In this study, the effects of thermal stress were evaluated on nuclear factor-kappaB (NF-kappaB) activation and proinflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) secretion in primary astrocytic cultures. The ability of HSP70-positive astrocytes to support or inhibit neurite outgrowth was investigated in neuron-astrocyte cocultures. Cultured astrocytes from cerebral cortex of rats were exposed to transient hyperthermia (42 degrees C/30 min) and incubated at 37 degrees C for different periods of recovery. During HSP70 accumulation, astrocytes extended large and thick processes associated to rearrangement of glial fibrillary acidic protein (GFAP) filaments and an increase in protein synthesis and GFAP, suggesting an astrogliosis event. A delay of NF-kappaB activation appeared closely related to TNF-alpha secretion by HSP70-positive astrocytes. These cells demonstrated a functional shift from neurite growth-promoting to non-permissive substrate. We also found that gliotoxin, a specific NF-kappaB inhibitor, partially abrogated the inhibitory ability of reactive astrocytes. These findings may suggest a involvement of NF-kappaB and TNF-alpha in modulating the failure of HSP70-positive astrocytes to provide functional support to neuritic outgrowth.

The transcription factor NFAT3 mediates neuronal survival.

Neuronal apoptosis is critical for normal development of the mammalian nervous system and also contributes to the pathogenesis of ischemic and degenerative diseases of the brain. Apoptosis of neurons is tightly regulated by extrinsic signals including growth factors and neuronal activity, but the intracellular mechanisms by which these signals promote neuronal survival are incompletely understood. We report that the transcription factor NFAT3 plays a critical role in mediating survival of granule neurons of the developing cerebellum. NFAT3 accumulated in the nucleus of primary granule neurons under survival conditions of serum growth factors and neuronal activity that was elicited by depolarization with high K(+). In contrast, deprivation of serum and K(+), which leads to neuronal apoptosis, triggered NFAT3 nuclear export. Treatment of granule neurons with Li(+), an inhibitor of the NFAT export kinase GSK3, prevented the nuclear export of NFAT3 and increased granule cell survival even under pro-apoptotic conditions. Thus, the nuclear localization of NFAT3 correlated tightly with granule neuron survival. Consistent with a pro-survival function for NFAT3, genetic knockdown of NFAT3 by RNA interference in primary granule neurons led to increased apoptosis even in neurons cultured under survival conditions. Conversely, expression of a constitutively active form of NFAT protected neurons against apoptosis induced by serum withdrawal and low K(+). Taken together, these results reveal an essential function for NFAT3-mediated transcription in neuronal survival that may play important roles in the developing and mature brain.

The T-cell anergy induced by Leishmania amazonensis antigens is related with defective antigen presentation and apoptosis.

Leishmania amazonensis is the main agent of diffuse cutaneous leishmaniasis, a disease associated with anergic immune responses. In this study we show that the crude antigen of Leishmania amazonensis (LaAg) but not L. braziliensis promastigotes (LbAg) contains substances that suppress mitogenic and spontaneous proliferative responses of T cells. The suppressive substances in LaAg are thermoresistant (100 degrees C/1h) and partially dependent on protease activity. T cell anergy was not due to a decreased production of growth factors as it was not reverted by addition of exogenous IL-2, IL-4, IFN-gamma or IL-12. LaAg did not inhibit anti-CD3-induced T cell activation, suggesting that anergy was due to a defect in antigen presentation. It was also not due to cell necrosis, but was accompanied by expressive DNA fragmentation in lymph node cells, indicative of apoptosis. Although pre-incubation of macrophages with LaAg prevented their capacity to present antigens, this effect was not due to apoptosis of the former. These results suggest that the T cell anergy found in diffuse leishmaniasis may be the result of parasite antigen-driven apoptosis of those cells following defective antigen presentation.

The T-cell anergy induced by Leishmania amazonensis antigens is related with defective antigen presentation and apoptosis

A Leishmania amazonensis é o principal agente etiológico da leishmaniose cutânea difusa, uma doença associada a respostas imunes anérgicas. Neste estudo nós mostramos que o extrato bruto de promastigotas de Leishmania amazonensis (LaAg), mas não de L. braziliensis (LbAg), contém substâncias que suprimem respostas proliferativas, espontâneas e mitogênicas, de células T. As substâncias supressoras no LaAg são termo-resistentes (100§C/1h) e parcialmente dependentes da atividade de proteases. A anergia de células T não foi devida à diminuição na produção de fatores de crescimento, uma vez que não foi revertida pela adição de: IL-2, IL-4, IFN-g ou IL-12. O LaAg não inibiu a ativação de células T induzida por anti-CD3, sugerindo que a anergia é devida a um defeito na apresentação de antígenos. A anergia não foi devida à necrose celular, mas foi acompanhada de uma expressiva fragmentação de DNA nas células de linfonodos, indicativo de apoptose. Apesar da pré-incubação de macrófagos com LaAg ter inibido sua capacidade de apresentação de antigenos, este efeito não foi devido à apoptose dos primeiros. Estes resultados sugerem que a anergia de células T encontrada na leishmaniose difusa deve ser devida à apoptose dessas células que se segue à apresentação defeituosa de antígenos pelo antígeno do parasito.