Induction of intercellular interaction and cell fusion by cell-penetrating peptide-conjugated lipids

Cell fusion is widely used in biomedical applications. However, conventional cell fusion methods, including polyethylene glycol (PEG)-based methods, often exhibit low efficiency. In this study, we investigated an alternative strategy to induce cell fusion with higher efficiency using transactivator of transcription peptide-conjugated PEG-lipids (Tat-PEG-lipids) with lauroyl chains (C12). We demonstrated the homogeneous cell fusion of human acute lymphoblastic leukemia T (CCRF-CEM) cells and heterogeneous cell fusion of CCRF-CEM cells with murine myeloma P3U1 cells, and human adipose-derived stem cells (hADSCs) with P3U1 cells. We first evaluated the cell surface modification of Tat-PEG-lipid (C12) by confocal laser scanning microscopy and flow cytometry, studying its concentration-dependent incorporation into the cell membrane and the corresponding changes in the surface charge. Kinetic analysis revealed that the half-life of Tat-PEG-lipid (C12) on the cell membrane was approximately 9 h. In addition, the critical surface density of Tat-PEG-lipid (C12) for inducing cell fusion was approximately 5.1 × 10⁴ molecules per cell. Reaction temperature was an important factor; cell fusion occurred at 37 °C but not at 4 °C, indicating that membrane fluidity was required for lipid exchange between adjacent attached cells. We found that Tat-PEG-lipid-mediated cell fusion can occur in various solutions, including phosphate-buffered saline, serum-free media, and serum-containing media, indicating that serum components did not disturb the cell fusion. Also, Tat-PEG-lipid (C12) could facilitate membrane fusion between different cells (CCRF-CEM × CCRF-CEM; 64%, CCRF-CEM × P3U1; 57%), although heterogeneous cell fusion was less efficient than homogeneous cell fusion.