Expression Of Functional Interleukin-21Receptor On Microglial Cells

Objectives and Methods:Microglial cells are the primary immune effector cells in the brain. In response to brain injury, microglial cells become activated and undergo morphological as well as functional transformations. They are critically involved in lesions, neurodegenerative diseases, stroke, and brain tumors. Although activated microglia release multiple cytokines, BBB disruption and synthesis by invading immune cells can also contribute to increased cytokine levels in the brain.. Microglia are also targets for cytokines; released by auto-and paracrine signaling mechanisms, these small proteins regulate microglial cell growth, survival, differentiation, migration and effector functions. Several cytokines (including growth factors) and their receptors have been shown to be expressed and functional in microglia.Interleukin-21and its receptor (IL-21R) have been recently described and characterized. IL-21is a class I cytokine produced by activated T cells and has a significant homology to IL-2, IL-4and IL-15. The IL-21-induced signaling pathway involves Jakl, Jak3, STAT1, STAT3and STAT5. It has been shown that IL-21R is expressed on B, T and natural killer (NK) cells, and interacts with the common γ chain upon ligand binding.Recently, it was reported that peripheral immune cells express interleukin (IL)-21receptor (IL-21R) mRNA, while microglia do not.we examined whether microglial cells express IL-21R on their surface. Further, we examined whether IL-21modulates microglial cell activation, changes in morphology and functional characteristics such as phagocytosis and production of pro-and anti-inflammatory cytokines (TNF-α, MCP-1, IL-6, IL-10and IL-1ra). Furthermore, we analyzed the involvement of the pathway in microglial IL-21R signaling.Results1.We first analyzed the expression of IL-21R mRNA by conventional RT-PCR analysis and found that it was detectable in microglia. We then performed flow cytometric analysis to define the percentage of isolated microglia that express surface IL-21R. IL-21R expression was detectable in microglia, consistent with previous findings using RT-PCR and western-blot.2. we measured microglial cell proliferation after stimulation with or without LPS or IL-21. As shown in, microglial cell numbers increased in response to IL-21and LPS. In addition, IL-21induced proliferation of microglia in a dose-dependent manner.3. We investigated the effects of IL-21on the morphological and functional characteristics of microglial cells. Microglial cells, as the immune effector cells of the brain, play a pivotal role in neuroprotection and their behavior during temperature treatment with IL-21remains to be elucidated. IL-21did influence primary microglial cell viability during the whole of the experiment.This was in line with a significant increase in cell phagocytosis of unstimulated microglial cells directly after IL-21stimulation.4. As there is increasing evidence that pro-and anti-inflammatory cytokines and chemokines play a key role in neuroprotection and neurodegeneration, we investigated whether IL-21promotes the secretion of selected cytokines and chemokines, namely IL-1β,MCP-1, IL-10, IL-6and TNF-a. There were statistically significant increases in TNF-a, IL-1β, IL-6and MCP-1secretion of IL-21treatment. This is in line with the significantly decreased secretion of the anti-inflammatory cytokine IL-10after IL-21-stimulation in comparison to non-stimulated cells.5. Microglial cells were treated with rIL-21protein thatled to an appreciable increase in pSTAT3and pJAK3mRNA expression. We next assessed if IL-21induces activation of STAT1, another STAT protein that has been reported to be activated by IL-21in some cell types. With the same experimental conditions, we found no detectable change.Conclusions:1. Surface IL-21R expression on microglial cells2. IL-21induces microglial cells proliferation3. IL-21induces morphological changes of microglial cells4. IL-21induces secretion of pro-and anti-inflammatory cytokines and chemokines by microglial cells. 5. IL-21activates JAK3/STAT3signaling in microglial cells...