Sex-dependent differences in the secretome of human endothelial cells

Abstract

Background Cellular sex has rarely been considered as a biological variable in preclinical research, even when the pathogenesis of diseases with predictable sex differences is studied. In this perspective, proteomics, and “omics” approaches in general, can provide powerful tools to obtain comprehensive cellular maps, thus favoring the discovery of still unknown sex-biased physio-pathological mechanisms. Methods We performed proteomic and Gene Ontology (GO) analyses of the secretome from human serum-deprived male and female endothelial cells (ECs) followed by ELISA validation. Apoptosis was detected by FACS and Western blot techniques and efferocytosis through the ability of the macrophage cell line RAW 264.7 to engulf apoptotic ECs. PTX3 mRNA levels were measured by RT-qPCR. Results Proteomic and GO analyses of the secretome from starved human male and female ECs demonstrated a significant enrichment in proteins related to cellular responses to stress and to the regulation of apoptosis in the secretome of male ECs. Accordingly, a higher percentage of male ECs underwent apoptosis in response to serum deprivation in comparison with female ECs. Among the secreted proteins, we reliably found higher levels of PTX3 in the male EC secretome. The silencing of PTX3 suggested that male ECs were dependent on its expression to properly carry out the efferocytotic process. At variance, female EC efferocytosis seemed to be independent on PTX3 expression. Conclusions Our results demonstrated that serum-starved male and female ECs possess different secretory phenotypes that might take part in the sex-biased response to cellular stress. We identified PTX3 as a crucial player in the male-specific endothelial response to an apoptotic trigger. This novel and sex-related role for secreted proteins, and mainly for PTX3, may open the way to the discovery of still unknown sex-specific mechanisms and pharmacological targets for the prevention and treatment of endothelial dysfunction at the onset of atherosclerosis and cardiovascular disease.

Sex‐Specific Response to Combinations of Shear Stress and Substrate Stiffness by Endothelial Cells In Vitro

“…In static culture, endothelial cells have been shown to display sexual dimorphism for a range of functions including proliferation, vasoregulation, migration, and wound healing, tube formation and vascular endothelial growth factor stimulation, response to stressors of hyperoxia and serum starvation, secreted factors, and stimulation by sex hormones. [41,42,44,45,46,[48][49][50][72][73][74][75][76][77] Our main effect results for cell spreading agree with reports on the characteristics of male and female rat aortic and microvascular endothelial cells that found female endothelial cells spread more than male endothelial cells. [46] It is important to note that this has immediate implications in terms of barrier function and wound healing ability, i.e., being able to cover a given area.…”

An Optimized MRM-Based Workflow of the l-Arginine/Nitric Oxide Pathway Metabolites Revealed Disease- and Sex-Related Differences in the Cardiovascular Field

“…Sex-based differences are also known to exist in multiple circulating biomarkers associated with cardiovascular risk and, between them, those involved in endothelial dysfunction [53,54], suggesting possible diversities also in pathophysiological mechanisms contributing to CVD.…”

Multicellular 3D Models to Study Tumour-Stroma Interactions

“…Male and female GBMs are biologically distinct, and improving outcomes may require sex-specific approaches to treatment. Similarly, sexdependent differences have been described in ECs from different vascular beds [110][111][112][113][114][115]. Thus, sex as a biological variable should be taken into account to reconstitute a more physiologically relevant in vitro TME.…”