Impact of Human Dermal Microvascular Endothelial Cells on Primary Dermal Fibroblasts in Response to Inflammatory Stress.

Benjamin Sanchez,Géraldine Aimond,Guangrong Liu,Jérôme Lamartine,Linan Li,Dominique Sigaudo-Roussel,Joshua Dulong

doi:10.3389/fcell.2019.00044

Abstract

The aim of the present study was to evaluate the impact of the microenvironment produced by dermal microvascular endothelial cells, secondary to a pro-inflammatory challenge, on 2D culture models using dermal fibroblasts and in 3D reconstructed skin model using dermal fibroblasts and keratinocytes from healthy donors. We hypothesized that specific microvascular endothelial low grade inflammation could change fibroblasts phenotype and be involved in extracellular matrix (ECM) modification and skin alteration. Following IFNγ, TNFα, IL-1β pro-inflammatory stress on Human Dermal Endothelial Cells (HDMEC) we observed the increased release of Chemokine ligand 2 (CCL2), IL-6 and IL-8 but not VEGF-A in the conditioned medium (CM). The subsequent addition of this endothelial pro-inflammatory CM in dermal fibroblasts revealed an upregulation of IL6, IL8 and CCL2 but no NF-κB gene expression. The resulting ECM formation was impaired with a reduction of the collagen 1 network and a decrease in COL1A1 gene expression in 2D and 3D models. Collagen 1 and pro-LOX protein expression were significantly reduced confirming an impairment of the collagen network related to endothelial inflammation secretion. To conclude, this work showed that, without any immune cells, the endothelial secretion in response to a pro-inflammatory stress is able to activate the fibroblasts that will maintain the pro-inflammatory environment and exacerbate ECM degradation.

Highlights

Skin microcirculation is located deep in the dermis layer up to the reticular dermis close to the epidermis
Human Dermal Endothelial Cells were treated with low-grade inflammatory mix and we observed an upregulation of InterCellular Adhesion Molecule 1 (ICAM-1) and Vascular cell adhesion protein 1 (VCAM-1) (Figure 1A) demonstrating that the low-grade inflammatory mix activated the dermal microvascular endothelial cells, as confirmed by 2.5-fold increase in Vascular endothelial growth factor A (VEGF-A) gene expression (Figure 1B)
We observed a significant Human Dermal Endothelial Cells (HDMEC) transcriptional activation of the pro-inflammatory gene expression markers, chemokine ligand 2 (CCL2), Interleukin 6 (IL-6), and Interleukin 8 (IL-8) (Figure 1C) in response to the initial inflammatory cocktail that resulted of an increased in HDMEC protein secretion of CCL2, Il-6, and IL-8 in the CM10 compared to the control condition CMØ (Figure 1D)

Summary

Introduction

Skin microcirculation is located deep in the dermis layer up to the reticular dermis close to the epidermis. Skin microcirculation plays a significant role in regulating skin homeostasis, thermoregulation, blood pressure and inflammatory response (Charkoudian, 2003; Gutterman et al, 2016). It allows nutrients and systemic factors to reach the whole skin tissue. Vascular Microenvironment on Skin Integrity throughout the skin microvascular network from arteries to small capillaries close to the epidermis. Microvascular endothelial cells are the major components of dermal blood vessels and are clearly involved in skin inflammatory process. Endothelial cells participate to synthesize and secrete chemokine and cytokine and are implicated in recruiting immune cells in response to dysfunctions due to different factors (e.g., UV, heat, pathological and physiological processes) (Swerlick and Lawley, 1993)

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