Cocaine induces production of infectious large extracellular vesicles (IEV) and regulates neuro-inflammation

Abstract: Endothelial injury is hypothesized to importantly contribute to the respiratory failure and thromboemboli seen in COVID-19. Yet, how SARS- CoV-2 may injure pulmonary endothelium has not been well elucidated. Though viral components of SARS-CoV-2 were reported to be occasionally detected in pulmonary endothelial cells of patient samples, it's still not clearly understood whether the virus actively replicates in these cells. We hypothesize that circulating mediators released from neighboring infected lung epithelial cells such as extracellular vesicles (EVs) harboring Spike-proteins or soluble Spike protein subunits may mediate inflammation and dysfunction of the vascular system. In addition, we propose that methamphetamine (Meth), one of the most commonly used illicit drugs known to be associated with an acute vascular syndrome, further exacerbates the SARS-CoV-2 induced endothelial disease by activating pro-coagulative pathways in endothelial cells. The overall objective of this proposal is to study the effects or Spike harboring EVs (Spike-EVs) or soluble Spike proteins derived from Spike expressing lung epithelial cells on elicitation of inflammation and endothelial injury and dysfunction. Further we will explore how Meth augments Spike protein-induced endothelial pathobiology. Specific points of innovation include: (i) Characterizing the EVs released from SARS-CoV-2 Spike protein expressing primary lung epithelial cells, (ii)Analyzing the effects of Spike-EVs or soluble Spike proteins on the release of pro-inflammatory cytokines in the presence or absence of Meth and studying the effects of Meth alone and Meth in combination with Spike-EVs or soluble Spike proteins on modulating the ACE2/Ang II/AT1R pathway in Human Lung Microvascular Endothelial Cells (HMVEC-L), (iii) Addressing whether Meth enhances Spike-EVs or soluble Spike proteins induced alteration of HMVEC-L monolayer integrity and permeability, (iv) Studying the effects of Meth on Spike-EVs or soluble Spike protein-induced expression of endothelial injury markers and pro-coagulative molecules, and (v) Exploring whether blocking TNF-α receptors in the cell membrane inhibits Meth and Spike-EVs or soluble Spike protein- induced endothelial injury. We set out three specific aims to: (i) Explore the effects of Spike-EVs derived from lung epithelial cells or soluble spike proteins on elicitation of inflammation in HMVEC-Ls and further study their role in inducing endothelial dysfunction and impairing monolayer integrity in the presence or absence of Meth, and (ii) Evaluate whether Spike-EVs or soluble Spike proteins augment Meth-enhanced expression of tissue factor (TF) and impairment of tissue factor pathway inhibitor, which contribute to an acute vascular syndrome. Deciphering the role of Spike-EVs and Meth in the pathogenesis of EC injury and dysfunction may provide insights into developing novel therapeutic strategies against severe COVID-19 complications in Meth using SARS-CoV-2 infected hosts.