Increased fluid secretion after adenoviral-mediated transfer of the aquaporin-1 cDNA to irradiated rat salivary glands

Abstract

A replication-deficient, recombinant adenovirus encoding human aquaporin-1 (hAQP1), the archetypal water channel, was constructed. This virus, AdhAQP1, directed hAQP1 expression in several epithelial cell lines in vitro. In polarized MDCK cell monolayers, hAQP1 was localized in the apical and basolateral plasma membranes. Fluid movement across monolayers infected by AdhAQP1 in response to an osmotic gradient was Ϸ4-fold that seen with uninfected monolayers or monolayers infected by a control virus. When AdhAQP1 was administered to rat submandibular glands by retrograde ductal instillation, significant hAQP1 expression was observed by Western blot analysis in crude plasma membranes and by immunohistochemical staining in both acinar and ductal cells. Three or four months after exposure to a single radiation dose (17.5 or 21 Gy, respectively), AdhAQP1 administration to rat submandibular glands led to a two-to threefold increase in salivary secretion compared with secretion from glands administered a control virus. These results suggest that hAQP1 gene transfer may have potential as an unique approach for the treatment of postradiation salivary hypofunction.

Systemic action of human growth hormone following adenovirus-mediated gene transfer to rat submandibular glands

“…The resulting plasmid was subjected to DNA sequencing to confirm the nucleotide sequences in the two joining regions, and then cotransfected with pJM17 (a gift from Dr F Graham, McMaster University, Hamilton, Ontario, Canada) into 293 cells as previously described. 25,26 The recombinant adenovirus so generated (termed AdCMVhGH) was plaque-purified.…”

Safety and efficacy of adenovirus-mediated transfer of the human aquaporin-1 cDNA to irradiated parotid glands of non-human primates

“…AdhAQP1 is a replication-deficient recombinant virus encoding hAQP1 that includes the cytomegalovirus promoter/enhancer and a simian virus 40 polyadenylation signal. 11 A functionally irrelevant virus for these experiments, AdCMVH3 (encoding the human salivary protein histatin 3), was used as control virus. 16 Recombinant adenoviruses were plaque-purified and isolated by equilibrium centrifugation on a CsCl gradient as reported previously.…”

“…10,11 Figure 5A demonstrates positive immunostaining for AQP1, primarily in capillaries and venules in control gland tissue (AdCMVH3; animal E). In animals receiving AdhAQP1, positive staining was seen in some acinar and ductal elements in addition to the vascular immunoreactivity.…”

“…However, gene transfer in vivo offers a novel way to treat conditions affecting parenchymal tissues, 8 -10 such as the secretory hypofunction that occurs secondary to radiation damage to major salivary glands. 11 Recently, work from our laboratory showed that it was possible to increase salivary fluid flow from irradiation (IR)-damaged submandibular glands in a rat model through an adenoviral-mediated transfer of human aquaporin-1 (hAQP1) cDNA to surviving epithelial cells. 11 In irradiated rats whose submandibular gland salivary flow rate was reduced ϳ65%, AQP1 cDNA transfer led to near normal levels of salivary secretion 3 days after recombinant virus administration.…”

“…11 Recently, work from our laboratory showed that it was possible to increase salivary fluid flow from irradiation (IR)-damaged submandibular glands in a rat model through an adenoviral-mediated transfer of human aquaporin-1 (hAQP1) cDNA to surviving epithelial cells. 11 In irradiated rats whose submandibular gland salivary flow rate was reduced ϳ65%, AQP1 cDNA transfer led to near normal levels of salivary secretion 3 days after recombinant virus administration. Thus, initial experiments indicated the possible value of gene transfer to manage this clinical problem.…”