Adipose Tissue Expansion is Dependent on a Piezo2-Vegfr3 Signaling Axis that Regulates Lymphangiogenesis

Obesity and impaired lymphangiogenesis are closely interrelated. One of the consequences of obesity is decreased density of lymphatic vessels, whereas not fully functional lymphatic vessels lead to the accumulation of lipids primary in adipose tissue. Adipose tissue is rich in both blood and lymphatic vessels, making it constantly under mechanical perturbation by blood and lymph flow. In this project we sought to understand how mechanical signals may mediate dynamic changes in adipose expansion. For this purpose, we have focused on Piezo channels which are mechanosensitive currents activated by shear stress, fluid flow and membrane tension, resulting in an intracellular influx of calcium. Our data of single cell RNA-seq analysis performed on adipose and mesenteric endothelium indicated that different Piezo channels have their own specialized localization within lymphatic endothelium. Piezo1 was a marker of lymphatic collecting duct endothelium, but Piezo2 was a marker of lymphatic capillary endothelium. In this project we initially focused on Piezo2 due to the high density of lymphatic capillaries in adipose tissue. Generated obesogenic Piezo2 fl/fl /Prox1-Cre ERT2+ mice were found to have significantly increased insulin tolerance, significantly increased weight and epigonadal fat pads. Importantly, there was no change in the water mass of the animals. Because decreased lymphangiogenesis correlates with adipose expansion, we hypothesize that Piezo2 may regulate lymphangiogenesis. In obesogenic Piezo2 fl/fl /Prox1-Cre ERT2+ mice, adipose lymphatics lost expression of Piezo2 and Flt4 (gene encoding main regulator of lymphangiogenesis – Vegfr3). This was also observed in humans, where higher BMI correlated with decreased expression of PIEZO2 and FLT4 in lymphatic capillaries of adipose tissue. To test our hypothesis, we used PIEZO2 siRNA on human dermal lymphatic endothelial cells (HDLECs) and demonstrated a significant reduction in FLT4 mRNA, with a regression analysis of PIEZO2 and FLT4 expression of r 2 =0.722. What is more, we haven’t observed a reduction of PIEZO2 mRNA after FLT4 siRNA knock-down in HDLECs. Treatment of lipids highly present in obesogenic diet led to decreased expression of PIEZO2 and FLT4 with unchanged expression level of PIEZO1, and decreased proliferation of HDLECs. Additionally, by Ki67 staining we shown that loss of PIEZO2 significantly decreased proliferation of HDLECs. Considering the Piezo2-dependent regulation of Flt4 expression, we proposed a potential model for their interaction with proximity ligation assay. Piezo2 dependent calcium pool in the cell's cytoplasm interacts with Calmodulin, which forms a ternary protein complex with Klf2 and the transcription factor Prox1 – regulator of Vegfr3 maintenance. To determine physiological implications of Piezo2 from lymphatic endothelial cells on lymphatic vessels density in adipose tissue light sheet analysis was performed. In conclusion, we propose a novel function of Piezo2 as a regulator of Flt4 expression that affects lymphangiogenesis in adipose to regulate expansion. Project is founded by public sources: NIH HL137112; NIH HL171997 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.