caa a22 4500 477126154 CHVBK 20180405111650.0 cr unu---uuuuu 170330e19970901xx s 000 0 eng 10.1023/A:1012163025455 doi (NATIONALLICENCE)springer-10.1023/A:1012163025455 The Absence of Accessible Vitronectin Receptors in Differentiated Tissue Hinders Adenoviral-Mediated Gene Transfer to the Intestinal Epithelium In Vitro [Elektronische Daten] [Elke Walter, Maria Croyle, Blake Roessler, Gordon Amidon] Purpose. Adenoviral (Ad) vectors have been used as efficient tools for gene therapy in various tissues, whereas in some differentiated epithelium transduction efficiency is almost abolished. Methods. Caco-2 cell monolayers were chosen as an in vitro model for the differentiated intestinal epithelium. Fluorescence-labeled adenoviral particles were used for binding studies to cell surfaces. Internalization receptors for adenoviral uptake were decteted by a fluorescence-labeled vitronectin antibody. Gene expression was studied by using the β-galactosidase reporter gene. All experiments were done on undifferentiated and differentiated Caco-2 cells. Furthermore, adenoviral particles were allowed to bind to differentiated Caco-2 monolayers followed by a trypsinization step that disintegrates the monolayers and result in a cell suspension. Gene expression was tested after reseeding the cells into dishes. Results. The results from adenoviral binding studies, vitronectin immunofluorescence detection and gene expression are in good agreement and indicate that virion binding as well as the expression of internalization receptors almost disappear in fully differentiated cells. Nonetheless, adenoviral binding to differentiated monolayers seems to be sufficient to cause up to 53% gene expression, but only if internalization of the vector can be induced by disintegrating the monolayers and releasing free vitronectin receptors. Conclusions. These findings indicate that gene transfer to the intestinal epithelium utilizing adenoviral vectors is poor and ineffective, because of the lack of sufficient internalization receptors. If these receptors can be exposed in differentiated epithelium, transduction can be made more efficicient. Alternatively, a viral vector must be developed whose uptake mechanism is independent of integrin receptor expression like the enteral virus Ad40, or Ad5 could be conjugated to ligands that trigger viral internalization by receptor-mediated endocytosis. Plenum Publishing Corporation, 1997 adenoviral vector nationallicence Caco-2 nationallicence gene transfer nationallicence integrin nationallicence intestinal epithelium nationallicence vitronectin receptor nationallicence Walter Elke College of Pharmacy, The University of Michigan, 48109-1065, Ann Arbor, Michigan aut Croyle Maria College of Pharmacy, The University of Michigan, 48109-1065, Ann Arbor, Michigan aut Roessler Blake Department of Internal Medicine, Division of Rheumatology, University of Michigan Medical Center, 48109, Ann Arbor, Michigan aut Amidon Gordon College of Pharmacy, The University of Michigan, 48109-1065, Ann Arbor, Michigan aut Pharmaceutical Research Kluwer Academic Publishers-Plenum Publishers 14/9(1997-09-01), 1216-1222 0724-8741 14:9<1216 1997 14 11095 https://doi.org/10.1023/A:1012163025455 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1023/A:1012163025455 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Walter Elke College of Pharmacy, The University of Michigan, 48109-1065, Ann Arbor, Michigan aut NATIONALLICENCE 700 1- Croyle Maria College of Pharmacy, The University of Michigan, 48109-1065, Ann Arbor, Michigan aut NATIONALLICENCE 700 1- Roessler Blake Department of Internal Medicine, Division of Rheumatology, University of Michigan Medical Center, 48109, Ann Arbor, Michigan aut NATIONALLICENCE 700 1- Amidon Gordon College of Pharmacy, The University of Michigan, 48109-1065, Ann Arbor, Michigan aut NATIONALLICENCE 773 0- Pharmaceutical Research Kluwer Academic Publishers-Plenum Publishers 14/9(1997-09-01), 1216-1222 0724-8741 14:9<1216 1997 14 11095 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer