- Background and aims: Production of phosphatidylinsoitol-3,4,5-trisphosphate (PIP3), induced by insulin is a major signaling step in the induction of insulin's metabolic actions in muscle and fat, and is impaired in various states of insulin resistance. Intracellular delivery of PIP3 into cells might therefore constitute an approach for bypassing such a signaling defect. However, this approach is challenged by the need to overcome permeability barriers through the plasma membrane, and to introduce the agent in well-defined spatial-temporal coordinates corresponding to endogenous PIP3 generation. Exogenous PIP3 may be introduced into cells using polycationic carriers which should mask its negative charges. However so far, these approaches only partially mimicked insulin's effects in normal, insulin sensitive cells, and displayed poor efficiency and low reproducibility. The overall aim of this work was to assess the intracellular transport of exogenous PIP3 to overcome cellular insulin resistance. Materials and Methods: Membrane-complex interactions and cellular localization were verified using biomimetic liposomal (lipid/polydiacetylene) membrane model and in L6 muscle cells in culture with fluorescent-PIP3 and live-cell microscopy. The ability of PIP3-carrier to induce insulin signaling and metabolic effects was evaluated in L6 myoblasts and in 3T3-L1 adipocytes electroporated with GFP-GLUT4myc plasmids. Results: A polymeric carrier, polyethyleneimine (PEI), was utilized at different molecular weights and branched/linear backbone to generate efficient electrostaticaly bound PIP3-carrier complexes. Fluorescent liposomes surface perturbation and ESR (Electron Spin Resonance) analyses revealed that PIP3-PEI formed complexes and penetrated the lipid bilayer. Branched PEI (25kDa) carrier was more efficient in membrane internalization than linear PEI of the same molecular weight. Maximal liposome-complex interaction occurred at 40 min. Live-cells kinetic studies revealed that the branched PEI-PIP3 complex enabled the retention of PIP3 at the cell periphery as compared to PIP3 alone. PIP3 delivered with PEI was found biologically available for binding to its intracellular ligand (GRP-PH) and for generating signaling effects, resulting in increased PKB phosphorylation, higher than that observed with neomycin as a carrier. Moreover, PIP3-carrier exposure increased Glut4 translocation and externalization in 3T3-L1 adipocytes. Conclusions: We demonstrated a polymeric system for exogenous PIP3 delivery that can induce insulin signaling and metabolic effect in muscle and fat cells based on biomimetic liposomes and cellular systems. Inducing insulin actions by intracellular PIP3 delivery in insulin resistant cells will shed light on the potential therapeutic use of this technology.