No abstract

Normal hepatocytes do not express endogenous uncoupling protein 2 (UCP2) in adult liver, although Kupffer cells do, and it is strikingly induced in hepatocytes in steatotic liver and obese conditions. However, the direct link of UCP2 with the pathogenic development of liver diseases and liver injury remains elusive. Here we report that targeted expression of UCP2 to mouse liver increases susceptibility to acute liver injury induced by lipopolysaccharide (LPS) and galactosamine (GalN). UCP2 appears to enhance proton leak, leading to mild uncoupling in a guanosine diphosphate-repressible manner. Indeed, mitochondria from the genetically manipulated mouse liver have increased state 4 respiration, lower respiratory control ratio, and reduced adenosine triphosphate (ATP) levels, which altered mitochondrial physiology. To address the underlying mechanism of how UCP2 and the reduced energy coupling efficiency enhance cell death in mouse liver, we show that the reduced ATP levels lead to activation of 5AMP-activated protein kinase (AMPK) and its downstream effector, c-Jun N-terminal kinase; thus, the increased sensitivity toward LPS/GalN-induces apoptosis. Importantly, we show that inhibition of UCP2 activity by its pharmacological inhibitor genipin prevents LPS/GalN-induced ATP reduction, AMPK activation, and apoptosis. Also, inhibition of ATP production by oligomycin promotes LPS/GalN-induced cell death both in vivo and in vitro. Conclusion: Our results clearly show that targeted expression of UCP2 in liver may result in compromised mitochondrial physiology that contributes to enhanced cell death and suggests a potential role of UCP2 in the development of liver diseases. (HEPATOLOGY 2009;50:1204-1216

Despite remarkable progress in the development of both viral and non-viral gene delivery vectors for cystic fibrosis therapy, low efficiency of gene transfer to the airway epithelium is a major obstacle to clinical application. Here we develop formulations that enhance cellular absorption of adenoviral vectors. We selected excipients from a panel of pharmaceutically acceptable com-pounds known to enhance drug absorption. Transduction efficiency of the virus in the presence of each ingredient was assessed in vitro and in vivo. Mannitol and chitosan substantially enhanced transduction efficiency in vitro and augmented expression in vivo by 4 and 8 log units, respectively. The most successful formulation (a blend of sucrose, mannitol, and Pluronic F68) transduced 100% of an A549 cell population in vitro and produced areas of intense gene expression in both large and small airways in vivo with minimal toxicity. Dose response studies also indicate that when placed in this formulation, the viral dose can be lowered by 1/2 log while maintaining superior levels of transgene expression. This formulation also enhanced the physical stability of the virus. No significant loss in titer was detected from a lyophilized formulation after storage at 25 degrees C for 30 days.