Human endothelial cells display a rapid and fluid flow dependent tensional stress increase in response to tumor necrosis factor-α

Abstract

As endothelial cells form the inner layer of blood vessels they display the first barrier to interstitial tissues, which results in a crucial role for inflammation. On the global, systemic level an important element of the complex process controlling the inflammatory response is the release of the cytokine tumor necrosis factor-α (TNF-α). While other pro-inflammatory agents like thrombin or histamine are known to induce acute but transient changes in endothelial cells which have been well studied biologically as well as mechanically, TNF-α is primarily known for its sustained effects on permeability and leukocyte recruitment. These functions are associated with transcriptional changes that take place on the timescale of hours and days. Here we show that already 4 minutes after the addition of TNF-α onto monolayers of human umbilical vein endothelial cells, a striking rise in mechanical substrate traction force and internal monolayer tension can be recorded. As expected, the traction forces act primarily at the boundary of the monolayer. While the traction forces increase monotonically during the initial cellular response, we find that the internal monolayer tension displays a rapid peak that can be abolished when applying a shear flow to the cells. The increased internal monolayer tension may provide a mechanical signal for the cells to prepare for the recruitment of leukocytes, additionally to the well studied biochemical response.

Borrelia burgdorferi modulates the physical forces and immunity signaling in endothelial cells

“…However, when we added MDP onto ECs we did not observe any change in EC force transduction but also any upregulation of NFkB target genes in ECs (data not shown). On the contrary, when we exposed ECs to TNFa, NFkB target genes were dramatically upregulated (10-to 100-fold) and immediate and sustained upregulation of EC force transduction was observed, consistent with previous studies (Stroka et al, 2012;Urbano et al, 2017;Brandt et al, 2022). This result suggests that the relationship between EC physical forces and innate immune signaling is likely not monotonic.…”

“…This remodeling results in dynamic alterations of the forces that cells transmit to their matrix and to each other, which are regulated to a large extent through the action of the actomyosin contractile apparatus (Komarova et al, 2017). Pro-inflammatory mediators and secreted cytokines can reinforce (Brandt et al, 2022) or alternatively release the tensile stresses between ECs, thus enhancing vascular permeability (Lee et al, 2011). A decrease in cell-ECM forces can also lead to attenuation in EC monolayer stresses as a result of the crosstalk between adherens junctions and focal adhesions (Komarova et al, 2017).…”

“…Our assay however is under static conditions and does not account for additional extracellular physical forces that ECs experience in vivo. For example, in the vasculature ECs are exposed to fluid shear stresses and gradients which can greatly impact EC behavior (e.g., proliferation, motility etc) and mechanotransduction (Barakat and Lieu, 2003;Brandt et al, 2022;Ostrowski et al, 2014;Tan et al, 2021). The fluid shear flow regime can also impact the way Bb adheres on ECs (Ebady et al, 2016)…”

“…The intriguing correlation between weakening in EC mechanotransduction and upregulation of NFkB target genes led us wonder whether EC force transduction would change in response to a cytokine that is known to upregulate NFkB like tumor necrosis factor alpha (TNFa) or to muramyl dipeptide (MDP), which is found in the cell wall of many bacteria and is known to upregulate NOD signaling and NFkB in certain cell types (Stroka et al, 2012;Urbano et al, 2017;Lappas, 2013;Brandt et al, 2022). We discovered that addition of TNFa onto EC monolayers led to an immediate and sustained increase of traction and monolayer stresses while the biomechanical responses of MDP-exposed ECs appeared identical to those of nonexposed ECs (Figures S7A-S7C).…”

Dystrophin is a mechanical tension modulator

“…To compare the homeostatic force generation properties at the tissue level to those as the single myoblast level, we exploit the fact that they both mechanically interact and pull on their environment, which can be quantified by the contractile moment determined by Traction Force Microscopy (TFM) [12,29,47,8] . Therefore, we performed TFM of single myoblasts on physiological 15 kPa (Young’s modulus) PAA gels that include fluorescent nanoparticles to track potential deformations (Figure 3 a).…”