Z. Liu1, M. M. Regueiro1, L. Zhang1, Y. Li1, S. Joel2, G. S. O’Connor2, S. Deo2, S. Daunert2, O. C. Velazquez1 1University Of Miami,Department Of Surgery, School Of Medicine,Miami, FL, USA 2University Of Miami,Department Of Biochemistry & Molecular Biology, School Of Medicine,Miami, FL, USA
Introduction: Atherosclerosis is an inflammatory disease. Intercellular adhesion molecule-1 (ICAM-1) is induced on the surface of inflamed endothelial cells (iEC) in atherosclerosis. Integrin LFA-1 associates with ICAM-1, and the I-domain of LFA-1 (idLFA-1) mediates binding to ICAM-1. idLFA-1/ICAM-1 pair is thus an attractive target for mediating homing of therapeutic cells to atherosclerotic lesions where iEC are presented. In this study, we tested a systemic targeted cell delivery method by coating stem cell surface with novel nanocarrier composed of idLFA-1-dendrimer complex. The nanocarriers guide the coated stem cells homing to atherosclerotic lesion via molecular recognition and association with ICAM-1 highly expressed on iEC in aorta of ApoE-/- mice. The efficiency of novel nanocarriers in mediating interaction of nanocarrier-coated cells to iEC in vitro and home to inflamed aorta in vivo was investigated.
Methods: Expression of ICAM-1 in iEC at aortic atherosclerotic lesion was examined by immunofluorescence (IF). Binding activity of recombinant human idLFA-1 to ICAM-1 was validated by testing association of Cy5-conjugated idLFA-1 with human aorta endothelial cells (HAEC) which pre-engineered to express high ICAM-1 (ICAM-1hi). Nanocarriers were created by conjugation of idLFA-1 or BSA (control) with acetylated generation 5 (Ac-G5)-dendrimers. DsRed+ human endothelial progenitor cells (EPC) were coated with idLFA-1-nanocarriers and BSA-nanocarriers, respectively. The efficiency of idLFA-1-nanocarriers in mediating interaction of DsRed+EPC to ICAM-1hi vs ICAM-1lo HAEC was tested using in vitro cell-cell binding assay. For in vivo testing, 1 x 106 luciferase (Luc)+ murine bone marrow-derived mesenchymal stem cells (MSC) coated with idLFA-1-nanocarriers or murine albumin-nanocarriers were infused twice via tail vein (i.v.) into ApoE-/- mice fed with high fat diet (HFD) (n=5/group). Aortas were harvested next day post the 2nd cell infusion. MSC homed to aorta were examined by IF using anti-Luc antibody and quantified based on fluorescence intensity in aorta sections.
Results: ICAM-1 is elevated in iEC at aortic atherosclerotic lesions of ApoE-/- mice. In vitro, Cy5-conjugated idLFA-1 preferentially binds to “inflamed” HAEC compared to control HAEC (p<0.01). idLFA-1-nanacarriers mediate increased interaction of DsRed+EPC to ICAM-1hi HAEC compared to ICAM-1lo HAEC (>2 fold, p<0.01). In vivo, idLFA-1-nanacarriers can successfully deliver more MSC to aorta in HFD fed ApoE-/- mice than control nanocarriers (>5 fold, p=0.03).
Conclusion: We demonstrated higher efficiency of idLFA-1-nanocarriers in delivering MSC to inflamed aorta in ApoE-/- mice. This work establishes a novel method for systemic cell delivery to inflamed aorta in animal model and paves way towards developing cell therapies for treatment of atherosclerosis and maybe other inflammatory diseases.