Gene transfer of cyto-protective molecules in corneal endothelial cells and cultured corneas: Analysis of protective effects in vitro and in vivo Nianqiao Gong a,b,1 , Ines Ecke c,1 , Stefan Mergler a,d , Jun Yang c , Sylvia Metzner a , Sabine Schu c , Hans-Dieter Volk c , Uwe Pleyer a , Thomas Ritter c,e, * a Department of Ophthalmology, Charite ´—University Medicine Berlin, Germany b Institute of Organ Transplantation, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China c Institute of Medical Immunology, Charite ´—University Medicine Berlin, Germany d Department of Internal Medicine, Division of Hepatology and Gastroenterology, Charite ´—University Medicine Berlin, Germany e Department of Medicine, Regenerative Medicine Institute (REMEDI), National Centre for Biomedical Engineering Science (NCBES), Orbsen Building, National University of Ireland, Galway, Ireland Received 22 March 2007 Available online 2 April 2007 Abstract The loss of corneal endothelial cells plays a critical role in many corneal diseases and is a common phenomenon following cornea transplantation. In addition, the non-regenerative capacity of human corneal endothelial cells (HCEC) ultimately requires appropriate protection of corneal tissues during ex vivo storage to ensure vitality of the cells. However, only 70% of donor corneas can be used for grafting because of endothelial deficiencies. Corneal endothelial cell loss during storage is mainly induced by apoptotic cell death. This study was undertaken, for proof of principle, to investigate whether over-expression of cyto-protective molecules Bcl-x L , Bag-1, and HO-1 prevents the loss of corneal endothelial cells both in vitro and in vivo. We demonstrate that gene transfer of both Bcl-x L and HO-1 has cyto-protective effects on HCEC in vitro. However, gene transfer of a single cyto-protective molecule does not prevent its rejection upon transplantation in a MHC class I/II disparate rat model. Ó 2007 Elsevier Inc. All rights reserved. Keywords: Gene therapy; Anti-apoptotic genes; Cyto-protection; Bcl-x L ; Transplantation; Cornea; Adenovirus The loss of corneal endothelial cells plays a critical role in many corneal diseases and is a common phenomenon following penetrating keratoplasty (cornea transplanta- tion) [1–3]. In addition, the non-regenerative capacity of human corneal endothelial cells (HCEC) ultimately requires appropriate protection of corneal tissues during ex vivo storage to ensure vitality of the cells. However, despite improved storage conditions only 70% of the donor corneas can be used for grafting because of endothelial deficiencies [4]. Corneal endothelial cell loss during storage is mainly induced by apoptotic cell death [3]. Recently, it has been described that inflammatory cytokines can induce apoptosis of corneal endothelium [5]. Many intracellular proteins are involved either in the induction or prevention of apoptosis. Bcl-x L is a member of the Bcl-2-family, which is able to regulate the membrane permeability and the release of cytochrome c from mito- chondria [6]. This protein also confers resistance to a wide variety of pro-apoptotic stimuli. Bag-1, another member of the family of anti-apoptotic genes, exerts its protective effects mainly by binding and stabilizing Bcl-2 [7]. In animal models of retinitis pigmentosa, gene transfer of Bcl-2 and Bag-1 is effective in the prevention of photoreceptor cell loss 0006-291X/$ - see front matter Ó 2007 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2007.03.146 * Corresponding author. Address: Department of Medicine, Regenera- tive Medicine Institute (REMEDI), National Centre for Biomedical Engineering Science (NCBES), Orbsen Building, National University of Ireland, Galway, Ireland. Fax: +353 (0)91 495547. E-mail address: thomas.ritter@nuigalway.ie (T. Ritter). URL: www.nuigalway.ie/remedi/ (T. Ritter). 1 These authors contributed equally to the work presented in this manuscript. www.elsevier.com/locate/ybbrc Biochemical and Biophysical Research Communications 357 (2007) 302–307