Adenoassociated virus-mediated transfer of a functional water channel intosalivary epithelial cells in vitro and in vivo

Aquaporin 1 (AQP1) is the archetypal member of a family of integral membran e proteins that function as water channels. Previously we have shown that t his protein can be expressed transiently from a recombinant adenovirus (Adh AQP1) in vitro in different epithelial cell lines, and in vivo in rat subma ndibular glands. In the present study we have constructed a recombinant ade noassociated virus (rAAV) containing the human aquaporin 1 gene (rAAVhAQP1) . rAAVhAQP1 was produced at relatively high titers. approximate to 10(11)-1 0(12) particles/ml and approximate to 10(8)-10(9) transducing units/ml. We show that the rAAVhAQP1 can transduce in vitro four epithelial cell lines o f different origins, at a level sufficient to detect the recombinant hAQP1 protein by either Western blot or confocal microscopic analysis. The recomb inant hAQP1 was correctly targeted to the plasma membranes in all cell line s. Function of the recombinant hAQP1 was measured as fluid flow, in respons e to an osmotic gradient, across a monolayer of transduced epithelial cells . The data show that even at a low level of transduction, typically approxi mate to 10% of the cells in the monolayer, transepithelial fluid movement i s enhanced about threefold above basal levels, In addition, we report that rAAVhAQP1 can transduce epithelial cells in the salivary glands and liver o f mice in vivo. These results suggest that rAAVs may be useful gene transfe r vectors to direct the production of functional transgenes in salivary epi thelial cell types.