Camila Marques Freria

Opposing effects of Toll-like receptors 2 and 4 on synaptic stability in the spinal cord after peripheral nerve injury

BackgroundGlial cells are involved in the synaptic elimination process that follows neuronal lesions, and are also responsible for mediating the interaction between the nervous and immune systems. Neurons and glial cells express Toll-like receptors (TLRs), which may affect the plasticity of the central nervous system (CNS). Because TLRs might also have non-immune functions in spinal-cord injury (SCI), we aimed to investigate the influence of TLR2 and TLR4 on synaptic plasticity and glial reactivity after peripheral nerve axotomy.MethodsThe lumbar spinal cords of C3H/HePas wild-type (WT) mice, C3H/HeJ TLR4-mutant mice, C57BL/6J WT mice, and C57BL/6J TLR2 knockout (KO) mice were studied after unilateral sciatic nerve transection. The mice were killed via intracardiac perfusion, and the spinal cord was processed for immunohistochemistry, transmission electron microscopy (TEM), western blotting, cell culture, and reverse transcriptase PCR. Primary cultures of astrocytes from newborn mice were established to study the astrocyte response in the absence of TLR2 and the deficiency of TLR4 expression.ResultsThe results showed that TLR4 and TLR2 expression in the CNS may have opposite effects on the stability of presynaptic terminals in the spinal cord. First, TLR4 contributed to synaptic preservation of terminals in apposition to lesioned motor neurons after peripheral injury, regardless of major histocompatibility complex class I (MHC I) expression. In addition, in the presence of TLR4, there was upregulation of glial cell-derived neurotrophic factor and downregulation of interleukin-6, but no morphological differences in glial reactivity were seen. By contrast, TLR2 expression led to greater synaptic loss, correlating with increased astrogliosis and upregulation of pro-inflammatory interleukins. Moreover, the absence of TLR2 resulted in the upregulation of neurotrophic factors and MHC I expression.ConclusionTLR4 and TLR2 in the CNS may have opposite effects on the stability of presynaptic terminals in the spinal cord and in astroglial reactions, indicating possible roles for these proteins in neuronal and glial responses to injury.

Major histocompatability complex class I expression and glial reaction influence spinal motoneuron synaptic plasticity during the course of experimental autoimmune encephalomyelitis

Recent studies have shown that major histocompatibility complex class I (MHC I) expression directly influences the stability of nerve terminals. Also, the acute phase of experimental autoimmune encephalomyelitis (EAE) has shown a significant impact on inputs within the spinal cord. Therefore, the present work investigated the synaptic covering of motoneurons during the induction phase of disease and progressive remissions of EAE. EAE was induced in C57BL/6J mice, which were divided into four groups: normal, peak disease, first remission, and second remission. The animals were killed and their lumbar spinal cords processed for in situ hybridization (IH), immunohistochemistry, and transmission electron microscopy (TEM). The results indicated an increase in glial reaction during the peak disease. During this period, the TEM analysis showed a reduction in the synaptic covering of the motoneurons, corresponding to a reduction in synaptophysin immunolabeling and an increase in the MHC I expression. The IH analysis reinforced the immunolabeling results, revealing an increased expression of MHC I mRNA by motoneurons and nonneuronal cells during the peak disease and first remission. The results observed in both remission groups indicated a return of the terminals to make contact with the motoneuron surface. The ratio between excitatory and inhibitory inputs increased, indicating the potential for development of an excitotoxic process. In conclusion, the results presented here indicate that MHC I up‐regulation during the course of EAE correlates with the periods of synaptic plasticity induced by the infiltration of autoreactive immune cells and that synaptic plasticity decreases after recurrent peaks of inflammation. J. Comp. Neurol. 518:990–1007, 2010.

Glatiramer acetate positively influences spinal motoneuron survival and synaptic plasticity after ventral root avulsion.

Avulsion of ventral roots induces degeneration of most axotomized motoneurons. At present there are no effective strategies to prevent such neuronal loss and to preserve the affected spinal circuits. Interestingly, changes in the spinal cord network also occur during the course of the experimental model of multiple sclerosis (experimental autoimmune encephalomyelitis-EAE). Glatiramer acetate (GA) significantly reduces the seriousness of the symptoms during the exacerbation of EAE. However, little is known about its effects on motoneurons. In the present study, we investigated whether GA has an influence on synapse plasticity and glial reaction after ventral root avulsion (VRA). Lewis rats were subjected to the avulsion of lumbar ventral roots and treated with GA. The animals were sacrificed after 14 days of treatment and the spinal cords processed for immunohistochemistry. A correlation between the synaptic changes and glial activation was obtained by performing immunolabeling against synaptophysin, GFAP and Iba-1. GA treatment preserved synaptophysin labeling, and significantly reduced the glial reaction in the area surrounding the axotomized motoneurons. After ventral root avulsion, GA treatment was also neuroprotective. The present results indicate that the immunomodulator GA has an influence on the stability of nerve terminals in the spinal cord, which may in turn contribute to future treatment strategies after proximal lesions to spinal motoneurons.

Granulocyte colony stimulating factor neuroprotective effects on spinal motoneurons after ventral root avulsion

G‐CSF is a glycoprotein commonly used to treat neutropenia. Recent studies have shown that the G‐CSF receptor (G‐CSF‐R) is expressed by neurons in the central nervous system (CNS), and neuroprotective effects of G‐CSF have been observed. In this study, the influence of G‐CSF treatment on the glial reactivity and synaptic plasticity of spinal motoneurons in rats subjected to ventral root avulsion (VRA) was investigated. Lewis rats (7 weeks old) were subjected to unilateral VRA and divided into two groups: G‐CSF and placebo treated. The drug treated animals were injected subcutaneously with 200 μg/kg/day of G‐CSF for 5 days post lesion. The placebo group received saline buffer. After 2 weeks, both groups were sacrificed and their lumbar intumescences processed for transmission electron microscopy (TEM), motoneuron counting, and immunohistochemistry with antibodies against GFAP, Iba‐1, and synaptophysin. Furthermore, in vitro analysis was carried out, using newborn cortical derived astrocytes. The results indicated increased neuronal survival in the G‐CSF treated group coupled with synaptic preservation. TEM analyses revealed an improved preservation of the synaptic covering in treated animals. Additionally, the drug treated group showed an increase in astroglial reactivity both in vivo and in vitro. The astrocytes also presented an increased cell proliferation rate when compared with the controls after 3 days of culturing. In conclusion, the present results suggest that G‐CSF has an influence on the stability of presynaptic terminals in the spinal cord as well as on the astroglial reaction, indicating a possible neuroprotective action. Synapse, 2012.

Expressão do complexo de histocompatilidade principal de classe I (MHC I) no sistema nervoso central: plasticidade sináptica e regeneração

It has been recently demonstrated that the major histocompatibility complex of class I (MHC I) expressed in the central nervous system (CNS) does not only function as a molecule of the immune system, but also plays a role in the synaptic plasticity. The expression of MHC I influences the intensity and selectivity of elimination of synapses apposed to neurons that were subjected to lesion, besides influencing the reactivity of neighboring glial cells. MHC I expression and the degree of synaptic rearrangement and glial response after injury correlate with differences in the regenerative potential and functional recovery of isogenic mice strains. In this way, the new aspects regarding MHC I functions in the CNS may guide further studies aiming at searching the involvement of MCH I in neurologic disorders, as well as the development of new therapeutic strategies.Foi demonstrado recentemente que o complexo de histocompatibilidade principal de classe I (MHC I), expresso no sistema nervoso central (SNC), nao funciona somente como molecula com papel imunologico, mas tambem como parte de um mecanismo envolvido na plasticidade sinaptica. A expressao de MHC I interfere na intensidade e seletividade da retracao de sinapses em contato com neuronios que sofreram lesao e tambem influencia a reatividade das celulas gliais proximas a esses neuronios. A intensidade do rearranjo sinaptico e resposta glial apos lesao, ligadas a expressao de MHC I no SNC, repercute em diferencas na capacidade regenerativa e recuperacao funcional em linhagens de camundongos isogenicos. Dessa forma, os novos aspectos sobre a funcao do MHC I no SNC direcionam futuras pesquisas no sentido de buscar o envolvimento do MHC I em doencas neurologicas e tambem o desenvolvimento de novas estrategias terapeuticas.

Carbon Nanotubes: From Synthesis to Genotoxicity

Massive industrial production of carbon nanotubes (CNTs) is increasing year after year, and it is urgent to address their safety-related issues. Due to their morphological similarities with asbestos fibers, which are classical carcinogenic materials, these CNTs have been considered as hazardous manufactured products by regulatory agencies. In this context, genotoxic effects of CNTs and the mechanisms proposed in current literature are reviewed and discussed in this chapter. Relevant aspects of preparation and physicochemical characterization of CNTs in toxicological context as well as the recent perspectives involving cytotoxicity assessment are also highlighted. Finally, this chapter aims to contribute to point out to a proactive discussion towards a responsible and sustainable development of nanotechnology lined up with environmental, health, and safety (EH&S) requirements.

The immunomodulator glatiramer acetate influences spinal motoneuron plasticity during the course of multiple sclerosis in an animal model

The immunomodulador glatiramer acetate (GA) has been shown to significantly reduce the severity of symptoms during the course of multiple sclerosis and in its animal model--experimental autoimmune encephalomyelitis (EAE). Since GA may influence the response of non-neuronal cells in the spinal cord, it is possible that, to some extent, this drug affects the synaptic changes induced during the exacerbation of EAE. In the present study, we investigated whether GA has a positive influence on the loss of inputs to the motoneurons during the course of EAE in rats. Lewis rats were subjected to EAE associated with GA or placebo treatment. The animals were sacrificed after 15 days of treatment and the spinal cords processed for immunohistochemical analysis and transmission electron microscopy. A correlation between the synaptic changes and glial activation was obtained by performing labeling of synaptophysin and glial fibrillary acidic protein using immunohistochemical analysis. Ultrastructural analysis of the terminals apposed to alpha motoneurons was also performed by electron transmission microscopy. Interestingly, although the GA treatment preserved synaptophysin labeling, it did not significantly reduce the glial reaction, indicating that inflammatory activity was still present. Also, ultrastructural analysis showed that GA treatment significantly prevented retraction of both F and S type terminals compared to placebo. The present results indicate that the immunomodulator GA has an influence on the stability of nerve terminals in the spinal cord, which in turn may contribute to its neuroprotective effects during the course of multiple sclerosis.

Expression of class I major histocompatibility complex (MHC I) in the central nervous system: role in synaptic plasticity and regeneration

It has been recently demonstrated that the major histocompatibility complex of class I (MHC I) expressed in the central nervous system (CNS) does not only function as a molecule of the immune system, but also plays a role in the synaptic plasticity. The expression of MHC I influences the intensity and selectivity of elimination of synapses apposed to neurons that were subjected to lesion, besides influencing the reactivity of neighboring glial cells. MHC I expression and the degree of synaptic rearrangement and glial response after injury correlate with differences in the regenerative potential and functional recovery of isogenic mice strains. In this way, the new aspects regarding MHC I functions in the CNS may guide further studies aiming at searching the involvement of MCH I in neurologic disorders, as well as the development of new therapeutic strategies.Foi demonstrado recentemente que o complexo de histocompatibilidade principal de classe I (MHC I), expresso no sistema nervoso central (SNC), nao funciona somente como molecula com papel imunologico, mas tambem como parte de um mecanismo envolvido na plasticidade sinaptica. A expressao de MHC I interfere na intensidade e seletividade da retracao de sinapses em contato com neuronios que sofreram lesao e tambem influencia a reatividade das celulas gliais proximas a esses neuronios. A intensidade do rearranjo sinaptico e resposta glial apos lesao, ligadas a expressao de MHC I no SNC, repercute em diferencas na capacidade regenerativa e recuperacao funcional em linhagens de camundongos isogenicos. Dessa forma, os novos aspectos sobre a funcao do MHC I no SNC direcionam futuras pesquisas no sentido de buscar o envolvimento do MHC I em doencas neurologicas e tambem o desenvolvimento de novas estrategias terapeuticas.

Expresión del complejo principal de histocompatibilidad de clase I (MHC I) en el sistema nervioso central: plasticidad sináptica y regeneración

It has been recently demonstrated that the major histocompatibility complex of class I (MHC I) expressed in the central nervous system (CNS) does not only function as a molecule of the immune system, but also plays a role in the synaptic plasticity. The expression of MHC I influences the intensity and selectivity of elimination of synapses apposed to neurons that were subjected to lesion, besides influencing the reactivity of neighboring glial cells. MHC I expression and the degree of synaptic rearrangement and glial response after injury correlate with differences in the regenerative potential and functional recovery of isogenic mice strains. In this way, the new aspects regarding MHC I functions in the CNS may guide further studies aiming at searching the involvement of MCH I in neurologic disorders, as well as the development of new therapeutic strategies.Foi demonstrado recentemente que o complexo de histocompatibilidade principal de classe I (MHC I), expresso no sistema nervoso central (SNC), nao funciona somente como molecula com papel imunologico, mas tambem como parte de um mecanismo envolvido na plasticidade sinaptica. A expressao de MHC I interfere na intensidade e seletividade da retracao de sinapses em contato com neuronios que sofreram lesao e tambem influencia a reatividade das celulas gliais proximas a esses neuronios. A intensidade do rearranjo sinaptico e resposta glial apos lesao, ligadas a expressao de MHC I no SNC, repercute em diferencas na capacidade regenerativa e recuperacao funcional em linhagens de camundongos isogenicos. Dessa forma, os novos aspectos sobre a funcao do MHC I no SNC direcionam futuras pesquisas no sentido de buscar o envolvimento do MHC I em doencas neurologicas e tambem o desenvolvimento de novas estrategias terapeuticas.