Dyslipidemia-associated atherogenic oxidized lipids induce platelet hyperactivity through phospholipase Cγ2-dependent reactive oxygen species generation

Abstract

Oxidized low-density lipoprotein (oxLDL) and associated oxidized phosphocholine-headgroup phospholipids (oxPCs) activate blood platelets through ligation of the scavenger receptor CD36. Previously, we found that oxLDL stimulated phosphorylation of phospholipase Cγ2 (PLCγ2). However, the functional relevance of PLCγ2 phosphorylation in oxLDL-mediated platelet hyperactivity remained elusive. Here, we set out to explore the functional importance of PLCγ2 in oxLDL-mediated platelet activation using human and genetically modified murine platelets. The CD36-specific oxidized phospholipid (oxPC) triggered the generation of reactive oxygen species (ROS) in platelets under static and arterial flow conditions. The ROS generation in response to oxPC was sustained for up to 3 h but ablated in CD36- and PLCγ2-deficient platelets. The functional importance of ROS generation in response to atherogenic lipid stress was examined through measurement of P-selectin expression. OxPC induced P-selectin expression, but required up to 60 min incubation, consistent with the timeline for ROS generation. P-selectin expression was not observed in CD36- and PLCγ2-deficient mice. The ability of oxPC and oxLDL to stimulate P-selectin expression was prevented by incubation of platelets with the ROS scavenger N-acetyl-cysteine (NAC) and the NOX-2 inhibitor gp91ds-tat, but not with the NOX-1 inhibitor ML171. In summary, we provide evidence that prolonged exposure to oxLDL-associated oxidized phospholipids induces platelet activation via NOX-2-mediated ROS production in a CD36- and PLCγ2-dependent manner.

Rh-CSF1 Attenuates Oxidative Stress and Neuronal Apoptosis via the CSF1R/PLCG2/PKA/UCP2 Signaling Pathway in a Rat Model of Neonatal HIE

“…In the CNS, PLCG2 was mainly expressed on microglia, but was also expressed on neurons in the granulosa cell layer of the dentate gyrus [ 61 ]. PLCG2 is involved in alleviating ROS levels and is closely associated with OS [ 26 , 27 ]. Other studies found that PLCG2 regulated mTOR through the diacylglycerol/protein kinase C signaling branch, thereby affecting the proliferation and apoptosis of B cells [ 62 ].…”

“…Studies showed that PLCG2 phosphorylation induced by the CSF1R activation could increase the rapid release of Ca 2+ in monocytes and promote the differentiation of monocytes [ 25 ]. PLCG2 is expressed on microglia and neurons in the CNS and is involved in regulating OS [ 26 , 27 ]. PLCG2 activates the cyclic adenosine monophosphate- (cAMP-) response element-binding protein through activation of the Ca 2+ signaling pathway induced by protein kinase A (PKA) [ 28 , 29 ].…”

Oxidised Low-Density Lipoprotein-Induced Platelet Hyperactivity—Receptors and Signalling Mechanisms

“…Consistent with canonical signalling from GPVI, SFK couple CD36 to the activation of Syk, which leads to the activation of a RhoA-dependent inhibition of myosin light chain phosphatase that underpins spreading and shape change [ 46 ]. Downstream signalling from Syk leads to phosphorylation of phospholipase Cγ2 (PLCγ2), which appears to be critical for oxLDL-induced platelet activation, since either pharmacological inhibition or genetic deletion blunted platelet hyperactivity [ 46 , 89 , 140 ]. The role of Syk in activating the other key signalling elements, such as ERK, p38, Mkk4/JNK, and Vav1/3, is unclear; although, it is likely, given that Syk activation lies downstream of other tyrosine kinase-linked receptors in platelets, for example, GPIb-IX-V, GPVI, Clec-2, and α IIb β 3 [ 141 , 142 , 143 , 144 ].…”

Oxidative Stress and New Pathogenetic Mechanisms in Endothelial Dysfunction: Potential Diagnostic Biomarkers and Therapeutic Targets

“…Nox2 inhibition also improved hind-limb blood flow in mice, reducing ROS level and inflammation after TNF-α administration [ 274 ]. Besides, NOX-2 inhibitor gp91ds-tat, combined with the ROS scavenger N-acetyl-cysteine (NAC), has proven to be effective against platelet activation induced by prolonged exposure to oxLDL-associated oxidized phospholipids [ 275 ]. GLX351322, a selective inhibitor of NOX4, has been suggested to be effective in T2D [ 276 ], while Plumbagin, another Nox4 specific inhibitor, has been reported to counteract oxidative stress-induced endothelial dysfunction and preadipocyte apoptosis under hyperinsulinemic conditions [ 277 , 278 ].…”