Morphological Methods to Evaluate Peripheral Nerve Fiber Regeneration: A Comprehensive Review

Regeneration of damaged peripheral nerves remains one of the main challenges of neurosurgery and regenerative medicine, a nerve functionality is rarely restored, especially after severe injuries. Researchers are constantly looking for innovative strategies for tackling this problem, with the development of advanced tissue-engineered nerve conduits and new pharmacological and physical interventions, with the aim of improving patients’ life quality. Different evaluation methods can be used to study the effectiveness of a new treatment, including functional tests, morphological assessment of regenerated nerve fibers and biomolecular analyses of key factors necessary for good regeneration. The number and diversity of protocols and methods, as well as the availability of innovative technologies which are used to assess nerve regeneration after experimental interventions, often makes it difficult to compare results obtained in different labs. The purpose of the current review is to describe the main morphological approaches used to evaluate the degree of nerve fiber regeneration in terms of their usefulness and limitations

Biological effects of sub-lethal doses of glyphosate and AMPA on cardiac myoblasts

Introduction: Glyphosate is the active compound of different non-selective herbicides, being the most used agriculture pesticide worldwide. Glyphosate and AMPA (one of its main metabolites) are common pollutants of water, soil, and food sources such as crops. They can be detected in biological samples from both exposed workers and general population. Despite glyphosate acts as inhibitor of the shikimate pathway, present only in plants and some microorganisms, its safety in mammals is still debated. Acute glyphosate intoxications are correlated to cardiovascular/neuronal damages, but little is known about the effects of the chronic exposure.Methods: We evaluated the direct biological effects of different concentrations of pure glyphosate/AMPA on a rat-derived cell line of cardiomyoblasts (H9c2) in acute (1–2 h) or sub-chronic (24–48 h) settings. We analyzed cell viability/morphology, ROS production and mitochondrial dynamics.Results: Acute exposure to high doses (above 10 mM) of glyphosate and AMPA triggers immediate cytotoxic effects: reduction in cell viability, increased ROS production, morphological alterations and mitochondrial function. When exposed to lower glyphosate concentrations (1 μM—1 mM), H9c2 cells showed only a slight variation in cell viability and ROS production, while mitochondrial dynamic was unvaried. Moreover, the phenotype was completely restored after 48 h of treatment. Surprisingly, the sub-chronic (48 h) treatment with low concentrations (1 μM—1 mM) of AMPA led to a late cytotoxic response, reflected in a reduction in H9c2 viability.Conclusion: The comprehension of the extent of human exposure to these molecules remains pivotal to have a better critical view of the available data

Role of inflammatory cytokines in peripheral nerve injury

Inflammatory events occurring in the distal part of an injured peripheral nerve have, nowadays, a great resonance. Investigating the timing of action of the several cytokines in the important stages of Wallerian degeneration helps to understand the regenerative process and design pharmacologic intervention that promotes and expedites recovery. The complex and synergistic action of inflammatory cytokines finally promotes axonal regeneration. Cytokines can be divided into pro- and anti-inflammatory cytokines that upregulate and downregulate, respectively, the production of inflammatory mediators. While pro-inflammatory cytokines are expressed in the first phase of Wallerian degeneration and promote the recruitment of macrophages, anti-inflammatory cytokines are expressed after this recruitment and downregulate the production of all cytokines, thus determining the end of the process. In this review, we describe the major inflammatory cytokines involved in Wallerian degeneration and the early phases of nerve regeneration. In particular, we focus on interleukin-1, interleukin-2, interleukin-6, tumor necrosis factor-β, interleukin-10 and transforming growth factor-β

Generation of new neurons in dorsal root Ganglia in adult rats after peripheral nerve crush injury.

The evidence of neurons generated ex novo in sensory ganglia of adult animals is still debated. In the present study, we investigated, using high resolution light microscopy and stereological analysis, the changes in the number of neurons in dorsal root ganglia after 30 days from a crush lesion of the rat brachial plexus terminal branches. Results showed, as expected, a relevant hypertrophy of dorsal root ganglion neurons. In addition, we reported, for the first time in the literature, that neuronal hypertrophy was accompanied by massive neuronal hyperplasia leading to a 42% increase of the number of primary sensory neurons. Moreover, ultrastructural analyses on sensory neurons showed that there was not a relevant neuronal loss as a consequence of the nerve injury. The evidence of BrdU-immunopositive neurons and neural progenitors labeled with Ki67, nanog, nestin, and sox-2 confirmed the stereological evidence of posttraumatic neurogenesis in dorsal root ganglia. Analysis of morphological changes following axonal damage in addition to immunofluorescence characterization of cell phenotype suggested that the neuronal precursors which give rise to the newly generated neurons could be represented by satellite glial cells that actively proliferate after the lesion and are able to differentiate toward the neuronal lineage

APTIDÃO URBANA AOS INSTRUMENTOS DO ESTATUTO DA CIDADE: PERSPECTIVAS METODOLÓGICAS

A Lei Federal 10.257/01, conhecida como Estatuto da Cidade, cuja função principal é regulamentar os artigos 182 e 183 da Constituição Federal de 1988, instituiu a obrigatoriedade da elaboração de planos diretores para municípios com mais de 20.000 habitantes e municípios com situações específicas como os que integram as regiões metropolitanas ou aqueles de interesse turístico. Simultaneamente, o Estatuto da Cidade legitimou instrumentos criados a partir da década de 1950 em várias cidades brasileiras, quando a questão urbana passou a ser vista como um problema social a ser equacionado no âmbito municipal, direcionado e gerenciado pelos planos diretores, monitorado por novos instrumentos complementares aos pré-existentes. No entanto, a implementação desses instrumentos integrados às leis dos planos diretores municipais, passa pelo gargalo da seleção da maior aptidão de cada setor urbano, ou outra unidade territorial da cidade, para a implantação de cada instrumento ou de um conjunto de instrumentos. Nesse contexto, o presente artigo visa colaborar, discutindo o estabelecimento de indicadores que, relacionados a cada instrumento, possam identificar tais aptidões, através de análises simplificadas das características das cidades. É importante destacar que essa metodologia está voltada principalmente a cidades de porte médio, que se posicionam aquém das metrópoles, quanto a demandas de estudos mais apurados, face às características das variáveis que se sobrepõem em sua estrutura urbana. Outro aspecto importante que vale salientar é que a aplicação dos instrumentos baseados nos indicadores não garante por si só os resultados pretendidos, dependendo de outras variáveis de cunho político e da visão dos atores sociais. São alternativas a serem submetidas ao processo decisório

Effect of unacylated ghrelin on peripheral nerve regeneration

Ghrelin is a circulating peptide hormone released by enteroendocrine cells of the gastrointestinal tract as two forms, acylated and unacylated. Acylated ghrelin (AG) binds to the growth hormone secretagogue receptor 1a (GHSR1a), thus stimulating food intake, growth hormone release, and gastrointestinal motility. Conversely, unacylated GHR (UnAG), through binding to a yet unidentified receptor, protects the skeletal muscle from atrophy, stimulates muscle regeneration, and protects cardiomyocytes from ischemic damage. Recently, interest about ghrelin has raised also among neuroscientists because of its effect on the nervous system, especially the stimulation of neurogenesis in spinal cord, brain stem, and hippocampus. However, few information is still available about its effectiveness on peripheral nerve regeneration. To partially fill this gap, the aim of this study was to assess the effect of UnAG on peripheral nerve regeneration after median nerve crush injury and after nerve transection immediately repaired by means of an end-to-end suture. To this end, we exploited FVB1 Myh6/Ghrl transgenic mice in which overexpression of the ghrelin gene (Ghrl) results in selective up-regulation of circulating UnAG levels, but not of AG. Regeneration was assessed by both functional evaluation (grasping test) and morphometrical analysis of regenerated myelinated axons. Results obtained lead to conclude that UnAG could have a role in development of peripheral nerves and during more severe lesions