Autologous Blood Clot Therapy for Wounds: Investigating the Chemotactic Effect on PBMCs and Fibroblasts in Diabetes

Topically applied autologous blood clot therapy (TABCT) is a low-cost regenerative modality proposed for complex wounds, yet its biological mechanism of action remains poorly defined. We investigated whether factors released from autologous blood clots promote peripheral blood mononuclear cell (PBMC) chemotaxis and fibroblast migration, and whether these effects are altered in individuals with metabolic syndrome/type 2 diabetes (MetS/DM). Twenty-two participants (controls n = 12; MetS/DM n = 10) were recruited. Autologous clots were generated ex vivo, cultured for 24 h, and the secretome analysed for total protein, PDGF-BB, P-selectin and CCL-5. PBMC basal activation (ROS production, phagocytosis, migration, Ca²⁺ flux) was assessed, followed by live-cell chemotaxis assays. Human dermal fibroblast migration was evaluated using a scratch assay. MetS/DM-derived PBMCs demonstrated heightened basal activation, including increased ROS production, migration velocity and sustained intracellular Ca²⁺ flux. Clots from MetS/DM participants released less total protein but significantly higher P-selectin levels, consistent with platelet hyperactivation. Despite containing measurable growth factors, clot-derived secretomes did not significantly enhance PBMC migration velocity or distance travelled, although directionality modestly increased. Similarly, secretome exposure failed to stimulate fibroblast-mediated wound closure (< 30% closure across groups). Pretreatment of blood with metformin, prednisone or amoxicillin had negligible effects on secretome composition or functional outcomes. These findings demonstrate that while TABCT-derived secretomes exhibit chemotactic properties, they do not substantially promote immune cell recruitment or fibroblast migration in vitro. The clinical benefits of TABCT may therefore be attributable primarily to its fibrin scaffold properties rather than bioactive factor release. Further mechanistic and translational studies are warranted.