Mechanosensitive mTORC1 signaling maintains lymphatic valves.

Homeostatic maintenance and repair of lymphatic vessels are essential for health. We investigated the dynamics and the molecular mechanisms of lymphatic endothelial cell (LEC) renewal in adult mesenteric quiescent lymphatic vasculature using label-retention, lineage tracing, and cell ablation strategies. Unlike during development, adult LEC turnover and proliferation was confined to the valve regions of collecting vessels, with valve cells displaying the shortest lifespan. Proliferating valve sinus LECs were the main source for maintenance and repair of lymphatic valves. We identified mechanistic target of rapamycin complex 1 (mTORC1) as a mechanoresponsive pathway activated by fluid shear stress in LECs. Depending on the shear stress level, mTORC1 activity drives division of valve cells or dictates their mechanic resilience through increased protein synthesis. Overactivation of lymphatic mTORC1 in vivo promoted supernumerary valve formation. Our work provides insights into the molecular mechanisms of maintenance of healthy lymphatic vascular system.

Mechanosensitive ACKR4 scavenges CCR7 chemokines to facilitate T cell de-adhesion and passive transport by flow in inflamed afferent lymphatics.

T cell migration via afferent lymphatics to draining lymph nodes (dLNs) depends on expression of CCR7 in T cells and CCL21 in the lymphatic vasculature. Once T cells have entered lymphatic capillaries, they slowly migrate into contracting collecting vessels. Here, lymph flow picks up, inducing T cell detachment and rapid transport to the dLNs. We find that the atypical chemokine receptor 4 (ACKR4), which binds and internalizes CCL19 and CCL21, is induced by lymph flow in endothelial cells lining lymphatic collectors, enabling them to scavenge these chemokines. In the absence of ACKR4, migration of T cells to dLNs in TPA-induced inflammation is significantly reduced. While entry into capillaries is not impaired, T cells accumulate in the ACKR4-deficient dermal collecting vessel segments. Overall, our findings identify an ACKR4-mediated mechanism by which lymphatic collectors facilitate the detachment of lymph-borne T cells in inflammation and their transition from crawling to free-flow toward the dLNs.

Dielectric elastomer actuator for mechanical loading of 2D cell cultures.

We demonstrate the use of dielectric elastomer actuators (DEAs) for mechanical stimulation of cells in vitro. The development of living tissues is regulated by their mechanical environment through the modification of fundamental cellular functions such as proliferation, differentiation and gene expression. Mechanical cues have been linked to numerous pathological conditions, and progress in cellular mechanobiology could lead to better diagnosis and treatments of diseases such as atherosclerosis and cancers. Research in this field heavily relies on in vitro models due to the high complexity of the in vivo environment. Current in vitro models however build on bulky and often complex sets of mechanical motors or pneumatic systems. In this work we present an alternative approach based on DEAs, a class of soft actuators capable of large deformation (>100%) and fast response time (<1 ms). The key advantage of DEAs is that they can be integrated within the culture substrate, therefore providing a very compact solution. Here we present a DEA-based deformable bioreactor which can generate up to 35% uniaxial tensile strain, and is compatible with standard cell culture protocols. Our transparent device also includes a static control area, and enables real-time optical monitoring of both the stimulated and control cell populations. As a proof of concept we cycled a population of lymphatic endothelial cells (LECs) between 0% and 10% strain at a 0.1 Hz frequency for 24 h. We observe stretch-induced alignment and elongation of LECs, providing the first demonstration that DEAs can be interfaced with living cells and used to control their mechanical environment.

Endothelial Cell Responses to Biomechanical Forces in Lymphatic Vessels.

SIGNIFICANCE: Lymphatic vessels are important components of the cardiovascular and immune systems. They contribute both to the maintenance of normal homeostasis and to many pathological conditions, such as cancer and inflammation. The lymphatic vasculature is subjected to a variety of biomechanical forces, including fluid shear stress and vessel circumferential stretch. RECENT ADVANCES: This review will discuss recent advances in our understanding of biomechanical forces in lymphatic vessels and their role in mammalian lymphatic vascular development and function. CRITICAL ISSUES: We will highlight the importance of fluid shear stress generated by lymph flow in organizing the lymphatic vascular network. We will also describe how mutations in mechanosensitive genes lead to lymphatic vascular dysfunction. FUTURE DIRECTIONS: Better understanding of how biomechanical and biochemical stimuli are perceived and interpreted by lymphatic endothelial cells is important for targeting regulation of lymphatic function in health and disease. Important remaining critical issues and future directions in the field will be discussed in this review. Antioxid. Redox Signal. 25, 451-465.