In Vivo Transfection of Rat Salivary Glands With Fluorescently Tagged Aquaporin-5 Channel DNA.

Abstract

Background The acinar cells of salivary glands are responsible for most saliva production and are, unfortunately, h­­ighly radiosensitive. As such, dry mouth or xerostomia is an adverse effect experienced by half of head and neck cancer patients treated with radiation. We evaluate a novel method of gene transfection of aquaporin channels to rat salivary glands.Materials and methodsA green fluorescent protein (GFP)-tagged human Aquaporin-5 (AQP5) cDNA sequence cloned into a pCMV6-AC-GFP vector was complexed with lipofectamine 2000. One submandibular gland of the anesthetized rats was injected with the complexed cDNA and lipid solution under ultrasound guidance, while the opposite gland was injected with the vehicle control. The animals were sacrificed between 24 to 48 hours post-injection. The salivary glands were removed and evaluated via fluorescence imaging. Western blot assays were also performed to determine AQP5 cDNA expression.Results In the experiments, the submandibular glands were identified and injected under ultrasound guidance. Four control glands and eight experimental glands were evaluated. The cDNA was expressed successfully and variably within the experimental glands, noting greater intensity along the cell surface consistent with appropriate trafficking of the AQP5 channel. Western blot analysis demonstrated variable expression in the experimental sample with no expression in the control sample. Several glands across the groups showed mild to moderate interstitial edema or inflammation.Conclusion In this study, we demonstrate an alternative in vivo transfection method via lipofection and demonstrate the successful expression of the AQP5 channel in rat salivary gland tissue.