Abstract
BackgroundMonocyte chemoattractant protein (CCL2/MCP-1) is a chemokine that attracts cells involved in the immune/inflammatory response. As microglia are one of the main cell types sustaining inflammation in brain, we proposed here to analyze the direct effects of MCP-1 on cultured primary microglia.MethodsPrimary microglia and neuronal cultures were obtained from neonatal and embryonic Wistar rats, respectively. Microglia were incubated with different concentrations of recombinant MCP-1 and LPS. Cell proliferation was quantified by measuring incorporation of bromodeoxyuridine (BrdU). Nitrite accumulation was measured using the Griess assay. The expression and synthesis of different proteins was measured by RT-PCR and ELISA. Cell death was quantified by measuring release of LDH into the culture medium.ResultsMCP-1 treatment (50 ng/ml, 24 h) did not induce morphological changes in microglial cultures. Protein and mRNA levels of different cytokines were measured, showing that MCP-1 was not able to induce proinflammatory cytokines (IL-1β, IL6, MIP-1α), either by itself or in combination with LPS. A similar lack of effect was observed when measuring inducible nitric oxide synthase (NOS2) expression or accumulation of nitrites in the culture media as a different indicator of microglial activation. MCP-1 was also unable to alter the expression of different trophic factors that were reduced by LPS treatment. In order to explore the possible release of other products by microglia and their potential neurotoxicity, neurons were co-cultured with microglia: no death of neurons could be detected when treated with MCP-1. However, the presence of MCP-1 induced proliferation of microglia, an effect opposite to that observed with LPS.ConclusionThese data indicate that, while causing migration and proliferation of microglia, MCP-1 does not appear to directly activate an inflammatory response in this cell type, and therefore, other factors may be necessary to cause the changes that result in the neuronal damage commonly observed in situations where MCP-1 levels are elevated.
Highlights
Monocyte chemoattractant protein (CCL2/MCP-1) is a chemokine that attracts cells involved in the immune/inflammatory response
The results presented here indicate that the changes caused by MCP-1 in microglia may not be responsible for the neuronal damage observed in certain situations where MCP-1 expression is elevated, and support the hypothesis which proposes a neuroprotective role for MCP-1
Microglia activation The induction of the enzyme NOS2 is considered to be a marker of those microglial modifications that lead to MCP-1 induces microglia proliferation A BrdU immunoassay was used to evaluate the proliferation of microglia in response to MCP-1
Summary
Monocyte chemoattractant protein (CCL2/MCP-1) is a chemokine that attracts cells involved in the immune/inflammatory response. An exaggerated inflammatory response can cause damage to cells that are necessary for the correct functioning of the tissue/organ they are part of. This is important in brain, where damage to or loss of constitutive cells can have worse consequences than in different tissues with higher regenerative capacities. The main immune cells in brain are microglia [4]; they continuously inspect their environment and react to changes that could threaten homeostasis [5]. This reaction can be detected mainly by changes in their morphology as well as by their expression of different genes or release of certain cytokines. Microglia can receive some signals that make them attack healthy neurons as is the case in neurodegenerative diseases such as Alzheimer’s disease, HIV-associated dementia or multiple sclerosis, among others [8]
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