The Loss of Glycoprotein Nonmetastatic Melanoma Protein B (GPNMB) Worsens Endothelial Cell Permeability, Metabolism, and Survival During Infectious Challenge
During severe systemic infections, also known as sepsis, excessive cytokines and reactive oxygen species lead to endothelial dysfunction. Glycoprotein nonmetastatic melanoma protein B (GPNMB) has been implicated in regulating cellular functions, particularly within the vasculature during inflammation, but its effect on infection-mediated endothelial injury remains unclear. Data obtained from the Gene Expression Omnibus (GEO) show that GPNMB expression is systemically reduced following an infectious challenge. Therefore, to investigate the role of GPNMB during infection-mediated endothelial inflammation, we utilized human microvascular endothelial cells (HMVECs) with or without GPNMB knockdown (siGPNMB) and exposed them to heat-killed Escherichia coli (HKEC), one of the most common pathogens associated with sepsis. Silencing GPNMB altered the expression of 1,556 genes via RNA sequencing, affecting cytoskeleton function and the response to stimuli. When assessing the endothelial monolayer under basal conditions, siGPNMB cells displayed higher transendothelial electrical resistance (TEER), consistent with RNA sequencing pathway analysis, but exposure to HKEC resulted in increased barrier dysfunction compared to controls. Furthermore, compared to controls, assessments of viability, proliferation, and migration were compromised in siGPNMB cells following HKEC exposure. Exposure to HKEC decreased the oxygen consumption rate in controls and increased the extracellular acidification rate, but neither were changed in siGPNMB cells, indicating impaired metabolic adaptation and further corroborating aspects of the RNA sequencing data. Our findings demonstrate that GPNMB reduction hinders the endothelial response to infectious stimuli, resulting in decreased metabolic fluxes and a dysfunctional endothelium during infectious challenges. Overall design: Total of 6 sample were analyzed, 3 TERT-immortalized dermal human microvascular endothelilal cells (HMVECs) treated for 72 hours with scrambled (control) siRNA and 3 TERT-immortalized dermal HMVECs treated with siRNA targeting glycoprotein nonmetastatic melanoma protein B (GPNMB)