Localized Adenovirus-Mediated Gene Transfer Into Vascular Smooth Muscle in the Hamster Cheek Pouch MARY D. FRAME,* †‡ JOSEPH M. MIANO, ‡§ JAY YANG,*  AND RICHARD J. RIVERS* † *Department of Anesthesiology, † Department of Biomedical Engineering, ‡ Center for Cardiovascular Research, § Department of Medicine, and  Department of Pharmacology and Physiology, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA ABSTRACT Objective: Our purpose was to develop a method for adenovirus delivery to the hamster cheek pouch to experimentally target gene transfer in tissue used for microvascular studies. Methods: Separate constructs were tested with transgenes for lacZ or green fluorescent protein (GFP) driven by three promoters: RSV, CMV, and SM22. With university approval, adenovirus was delivered in anesthetized (pentobar- bital, 70 mg/kg) hamsters (n  28) by using either a vascular systemic injection or tissue infiltration (interstitial space behind the pouch). During 3 days, animals receiving infiltration gained the expected weight, whereas those receiving vas- cular injection lost weight; no other behavior changes were noted. Results: On day 3 postadenoviral delivery (infiltration), expression of lacZ (his- tology, -galactosidase) or GFP (fluorescence microscopy) was confirmed across the tissue (CMV and RSV promoters) and exclusively in vascular smooth muscle cells (specific SM22 promoter), without evidence of tissue inflammation. In vitro microvascular experiments verified normal responses in the cheek pouch of day 3 postadenoviral delivery animals. We tested local dilation to methacholine, adenosine, remote dilation to methacholine, adenosine, nitroprusside, and LM609 ( v  3 integrin agonist), flow-dependent dilation, and flow recruitment. Conclusions: Thus, this method enables targeted, cell-specific gene transfer to one tissue important for microvascular studies, without significant systemic ex- posure and without adverse inflammation. Microcirculation (2001) 8, 403– 413. KEY WORDS: adenovirus, non-invasive delivery, microcirculation, smooth muscle, promoter INTRODUCTION Increasingly, reports in the literature show that sys- temic adenovirus administration can evoke compli- cations, including an undesirable systemic immune response (10,16,26). Such a systemic immune re- sponse potentially interferes with the physiological mechanism under study. Nonetheless, genetic ma- nipulation is a very specific approach to determine key protein involvement in the control of cellular responses. One alternative to systemic treatment is adenovirus delivery to only the organ being studied (4,5,11,24), which has historically required surgery to directly expose the vasculature of the desired or- gan. However, surgery itself could potentially com- plicate the animal’s homeostasis. The hamster cheek pouch remains an ideal tissue for microvascular studies (17,19,25) and now also a tis- sue that can be preferentially exposed to adenovirus for targeted genetic manipulation, without surgery. Supported by American Heart Association Grant EI0040197 and National Institutes of Health Grants HL55492 (to MDF), HL62572 (to JMM), and GM57578 (to JY). For reprints of this article, contact Mary D. Frame, Ph.D., De- partment of Anesthesiology, University of Rochester School of Medicine and Dentistry, 601 Elmwood Ave., Rochester, NY 14642, USA; e-mail: molly_frame@urmc.rochester.edu Received: 1 June 2001; accepted 8 August 2001 Microcirculation (2001) 8, 403–413 © 2001 Nature Publishing Group 1073-9688/01 $17.00 www.nature.com/mn