Adenoviral transduction of enterocytes and M-cells using in vitro models based on Caco-2 cells: the coxsackievirus and adenovirus receptor (CAR) mediates both apical and basolateral transduction.
Understanding virus-cell interaction is a key to the design of successful gene delivery vectors. In the present study we investigated Ad5 transduction of enterocytes and M-cells utilizing differentiated Caco-2 cells and cocultures of Caco-2 cells with lymphocytes. Transduction inhibition studies showed that CAR is the major receptor mediating apical and basolateral virus entry in differentiated Caco-2 cells. Integrins and heparan sulfate glycosaminoglycans do not appear to play a significant role. Immunofluorescence localized CAR to sites of cell-cell contact, with staining mostly observed on the cell perimeter. Staining was observed even in nonpermeabilized monolayers, suggesting apical accessibility of the receptor. Cocultures with mouse Peyer's patch lymphocytes or Raji B human lymphocytes were more susceptible to transduction than Caco-2 cells, and the effects were dose-dependent. Similar to Caco-2 cells, CAR and not integrins mediated apical transduction. In conclusion, contrary to other epithelial cell lines, both apical and basolateral transduction of absorptive enterocytes and M-cells is mediated by binding to CAR. The coculture system can be used to study the interactions between M-cells and gene delivery vectors.
Kesisoglou
Filippos
F
Department of Pharmaceutical Sciences, College of Pharmacy, The University of Michigan, Ann Arbor, Michigan 48109, USA.
Schmiedlin-Ren
Phyllissa
P
Fleisher
David
D
Zimmermann
Ellen M
EM
D000256:Adenoviridae / Q000235:genetics*; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D018938:Caco-2 Cells; D002460:Cell Line; D002478:Cells, Cultured; D018920:Coculture Techniques; D000090702:Constitutive Androstane Receptor; D020895:Enterocytes / Q000378:metabolism* / Q000648:ultrastructure / Q000821:virology*; D006801:Humans; D008214:Lymphocytes / Q000166:cytology / Q000378:metabolism; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D046529:Microscopy, Electron, Transmission; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D014161:Transduction, Genetic / Q000379:methods*
D000090702:Constitutive Androstane Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D000469:Alkaline Phosphatase