Regulation of Integrin α6 Recycling by Calcium-independent Phospholipase A2 (iPLA2) to Promote Microglia Chemotaxis on Laminin

Abstract

Microglia are the immune effector cells that are activated in response to pathological changes in the central nervous system. Microglial activation is accompanied by the alteration of integrin expression on the microglia surface. However, changes of integrin expression upon chemoattractant (ADP) stimulation still remain unknown. In this study, we investigated whether ADP induces the alteration of integrin species on the cell surface, leading to changes in chemotactic ability on different extracellular matrix proteins. Flow cytometry scans and on-cell Western assays showed that ADP stimulation induced a significant increase of ␣6 integrin-GFP, but not ␣5, on the surface of microglia cells. Microglia also showed a greater motility increase on laminin than fibronectin after ADP stimulation. Time lapse microscopy and integrin endocytosis assay revealed the essential role of calcium-independent phospholipase A 2 activity for the recycling of ␣6 integrin-GFP from the endosomal recycling complex to the plasma membrane. Lack of calcium-independent phospholipase A 2 activity caused a reduced rate of focal adhesion formation on laminin at the leading edge. Our results suggest that the alteration of integrin-mediated adhesion may regulate the extent of microglial infiltration into the site of damage by controlling their chemotactic ability.Microglia are the immune effector cells in the central nervous system (CNS). Under normal conditions, microglia exist as nonmigratory ramified cells. The ramified morphology of resting microglia is rapidly transformed into a motile ameboid form after pathological stimuli, driving the migration of microglia cells toward lesion sites (1-3). There is a growing body of evidence that the extracellular matrix (ECM) 2 and integrins are important for modulating microglial behavior. ECM influences microglial behavior as fibronectin (FN) and laminin (LN) often show opposite effects on microglial morphology, adhesion, and activation. FN promotes transformation of amoeboid microglia into ramified microglia, whereas LN causes the reverse transformation (4). FN also promotes increased secretion of amyloid precursor protein by microglial cells, whereas LN and collagen have the opposite effect (5). It has been demonstrated that microglia attach strongly to FN and vitronectin but only weakly to LN and astrocyte ECM and that LN exerts a dominant antiadhesive effect on microglial adhesion (6

Signaling Pathways Controlling Microglia Chemotaxis

“…It was further demonstrated that iPLA 2 activity is specifically required for the trafficking of c-Src back to the plasma membrane. Additional evidence for the important role of iPLA 2 in the regulation of microglia chemotaxis was from the study showing that the recycling of internalized α6 integrin vesicles and their delivery to focal adhesions also requires iPLA 2 activity during microglia chemotaxis ( Lee et al, 2016 ). Results from studies on chemotaxis of different types of cells clearly indicate that iPLA 2 takes a significant part in the regulation of chemotaxis.…”

“…The regulation of phosphorylation and dephosphorylation on VASP might be required for the membrane ruffle formation and growth of adhesion strength during membrane protrusion and retraction, exerting a direct impact on cell’s ability to chemotaxis ( Lee and Chung, 2009 ). It has been reported that ADP stimulation via P2Y 12 R induces the translocation of β1 and α6 integrins to the membrane ruffle regions ( Lee et al, 2016 ; Nasu-Tada et al, 2005 ). Redistribution of β1 was lost when the intracellular cAMP was increased, suggesting that PKA is a negative regulator of the β1 integrin translocation, resulting in a significant impact on microglia chemotaxis ( Nasu-Tada et al, 2005 ).…”

Regulation of Microglial Functions by Purinergic Mechanisms in the Healthy and Diseased CNS

“…It was reported that the PLC-mediated increase in intracellular Ca 2+ concentration-triggered proteinkinase B (Akt) activation was also involved in the second-messenger mechanisms of P2Y12Rs [75]. Eventually, P2Y12Rs initiate an adhesive interaction with the surrounding extracellular matrix through integrins [76,77]. Nerve injury-activated microglia engulf myelinated axons in the spinal cord dorsal horn in a P2Y12R-triggered and p38 signaling-mediated manner; this reaction is causally involved in the pathogenesis of neuropathic pain [78,79].…”

Discovery of novel L-type voltage-gated calcium channel blockers and application for the prevention of inflammation and angiogenesis

“…We further examined whether CCBs could affect the interactions of BV-2 cells and extracellular matrix, which affects cell motility as well as a changes in capacitance, which were expected due to decrease in intracellular Ca 2+ and thus overall cell polarity when compared to the culture medium media. We utilized the electrical cell-impedance sensing (ECIS) system, as Lee et al reported the effective use of ECIS to detect the BV-2 cell migration activity by reading resistance as well as capable of resolving the capacitance [37]. Due to the low sensitivity of the ECIS device, we used higher concentrations of the respective substances (10 μM) in the short-term experiment of 4 h. The ECIS results demonstrated a rapid decrease in the resistance at 4 kHz ( Fig.…”