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Introduction: Generating new lymphatic vessels has been postulated as an innovative therapeutic strategy for various disease phenotypes, including neurodegenerative diseases, metabolic syndrome, cardiovascular disease, and lymphedema. Yet, compared to the blood vascular system, protocols to differentiate human induced pluripotent stem cells (hiPSCs) into lymphatic endothelial cells (LECs) are still lacking. Methods: Transcription factors, ETS2 and ETV2 are key regulators of embryonic vascular development, including lymphatic specification. While ETV2 has been shown to efficiently generate blood endothelial cells, little is known about ETS2 and its role in lymphatic differentiation. Here, we describe a method for rapid and efficient generation of LECs using transcription factors, ETS2 and ETV2. Results: This approach reproducibly differentiates four diverse hiPSCs into LECs with exceedingly high efficiency. Timely activation of ETS2 was critical, to enable its interaction with Prox1, a master lymphatic regulator. Differentiated LECs express key lymphatic markers, VEGFR3, LYVE-1, and Podoplanin, in comparable levels to mature LECs. The differentiated LECs are able to assemble into stable lymphatic vascular networks in vitro, and secrete key lymphangiocrine, reelin. Conclusion: Overall, our protocol has broad applications for basic study of lymphatic biology, as well as toward various approaches in lymphatic regeneration and personalized medicine. 

Surface coating with dopamine conjugated hyaluronic acid (HA–DP) can interact with lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1) to preserve the phenotypes and functionality of lymphatic endothelial cells (LECs). 

Abstract Granular hydrogels show great promise in biomedical applications by mimicking the extracellular matrix and fostering a supportive microenvironment for tissue regeneration. This study investigates how tuning granular hydrogel properties influences lymphatic tube formation. Microgels were fabricated using norbornene‐modified hyaluronic acid (NorHA) via pipetting or vortexing for 90 s (V90s) and 180 s (V180s), then assembled into granular hydrogels under loose and tight packing conditions. These conditions produced gels with varied pore morphologies and bulk rheological properties. Lymphatic capillary formation occurred only in tightly packed gels, where mechanical properties converged, highlighting the importance of gel morphology over stiffness. V180s samples showed earlier vessel formation as seen in lymphatic gene and protein expression, while pipetted gels exhibited greater capillary connectivity, forming larger vessel clusters and fewer small satellite structures. The pipetting gels also supported lower‐curvature, more linear capillary networks that bridged multiple droplets, likely due to reduced entrapment in large voids compared to vortexed gels. These findings suggest that in bulk granular gels, lymphatic tube formation is governed not by mechanical stiffness but by pore size and gel topology (periodicity). Understanding and optimizing these morphological parameters can inform future strategies in lymphatic tissue engineering and regenerative medicine.