Pericyte contact alters endothelial cell metabolism by promoting exchange of lactate through SLC16A3

Abstract Intimate crosstalk between endothelial cells and pericytes is fundamental for vascular development and stability, but the metabolic dimension of this interaction remains poorly defined. Using a filter-based co-culture system that mimics the shared basement membrane of the microcirculation, we performed an integrated multi-omics analysis to investigate how direct contact reprograms both cell types. We found that initial contact does not induce immediate quiescence but rather triggers a transient, low-level activation of an endothelial-to-mesenchymal transition (EndMT)-like state in endothelial cells, characterized by specific upregulation of genes involved in extracellular matrix production (e.g., collagen isoforms) and PDGFR signaling, without a full loss of endothelial identity. Concomitantly, pericytes shifted endothelial cell metabolism, increasing glycolysis and elevating intracellular pyruvate and lactate. Proteomic analysis of the contact interface revealed enrichment of solute carriers, most notably the lactate transporter SLC16A3. Functional studies demonstrated that pericytes act as a glycolytic partner, actively shuttling lactate to endothelial cells via SLC16A3. This lactate did not fuel the TCA cycle but instead served as a signaling metabolite, driving widespread alterations in the endothelial acetylome and lactylome, particularly affecting proteins involved in glycolytic metabolism. In vitro, exogenous lactate potentiated cytokine-induced EndMT and upregulated lactylation-associated genes. The physiological relevance of this lactate shuttle was confirmed in vivo, where endothelial-specific deletion of Slc16a3 in mice impaired postnatal retinal angiogenesis, leading to reduced vessel density and diminished endothelial-pericyte overlap without affecting endothelial cell proliferation. Our findings establish that vessel maturation is orchestrated by a metabolically gated phase of plasticity initiated upon first contact, wherein a targeted EndMT-like program and a pericyte-driven lactate signaling axis are integrated to coordinate vascular network assembly. Competing Interest Statement The authors have declared no competing interest.