The Cell Morphology Core at U. Penn's Gene Therapy Program is a fully equipped facility capable to carry out all aspects of histology, microscopy, and morphometric analyses. This includes routine histology procedures including necropsies as well as more advanced techniques such as multicolor immunofluorescence, in situ hybridization, enzyme activity staining and electron microscopy. All necessary instruments to process and analyze tissue samples are available within the Core's lab space (including microscopes for light and electron microscopy). Investigators can use the Core's microscopes and imaging software to acquire images from their samples. The Cell Morphology Core will perform all the histology- and microscopy-related work required for the three projects of this grant. This will include the following procedures: Project 1: mouse necropsies; immunostaining for OTC, OTC mutants, and MHCI-SIINFEKL;in situ tetramer staining for SIINFEKL-specific T cells. Project 2: mouse necropsies;GFP detection in liver;staining of livers for general pathology (H&E), fibrosis (trichrome/Sirius Red), steatosis (oil-red-O), apoptosis (TUNEL assay), inflammation (immunostaining for CD4/CDS T cell infiltrates);immunostaining for hOTC expression in liver;OTC enzyme activity staining. Project 3: mouse necropsies;detection of GFP in liver;immunostaining for hOTC expression in liver, H&E staining. The Cell Morphology Core has the advantage of having many years of experience with analyzing liver tissues for gene therapy studies and in particular with the analytical techniques proposed for each project of this grant. While some routine techniques are easy to perform, certain methods such as detection of human OTC in spf^^''liver by immunofluorescence or enzyme histochemistry require the adherence to established and tested protocols. Protocols have been worked out and previously used successfully for almost all planned histological methods and do not need to be newly developed. The Core performs all experiments such as immunostains with stringent controls to confirm the specificity of the stains and slides are reviewed by the Core director before being released to the investigator. The Administrative Core oversees the Cell Morphology Core's budget and reviews pricing of services to assure cost effectiveness.
Histology is an important readout for all three projects of this grant. The detection of hOTC expression in liver for Project 1, the histopathological analyses of livers for Project 2, and the evaluation of vector performance by analyzing OTC expression in liver for Project 3 are key components of this grant. The Cell Morphology Core has all the necessary experience and equipment to perform these studies with proper controls and in a timely and cost-effective way.
|Mikals, Kyle; Nam, Hyun-Joo; Van Vliet, Kim et al. (2014) The structure of AAVrh32.33, a novel gene delivery vector. J Struct Biol 186:308-17|
|Mays, Lauren E; Wang, Lili; Lin, Jianping et al. (2014) AAV8 induces tolerance in murine muscle as a result of poor APC transduction, T cell exhaustion, and minimal MHCI upregulation on target cells. Mol Ther 22:28-41|
|Bryant, Laura M; Christopher, Devin M; Giles, April R et al. (2013) Lessons learned from the clinical development and market authorization of Glybera. Hum Gene Ther Clin Dev 24:55-64|
|Zhong, Li; Malani, Nirav; Li, Mengxin et al. (2013) Recombinant adeno-associated virus integration sites in murine liver after ornithine transcarbamylase gene correction. Hum Gene Ther 24:520-5|
|Wang, L; Wang, H; Morizono, H et al. (2012) Sustained correction of OTC deficiency in spf(?ash) mice using optimized self-complementary AAV2/8 vectors. Gene Ther 19:404-10|
|Wilson, James M; Shchelochkov, Oleg A; Gallagher, Renata C et al. (2012) Hepatocellular carcinoma in a research subject with ornithine transcarbamylase deficiency. Mol Genet Metab 105:263-5|
|Wang, Lili; Morizono, Hiroki; Lin, Jianping et al. (2012) Preclinical evaluation of a clinical candidate AAV8 vector for ornithine transcarbamylase (OTC) deficiency reveals functional enzyme from each persisting vector genome. Mol Genet Metab 105:203-11|
|Wang, Lili; Bell, Peter; Lin, Jianping et al. (2011) AAV8-mediated hepatic gene transfer in infant rhesus monkeys (Macaca mulatta). Mol Ther 19:2012-20|
|Bell, Peter; Wang, Lili; Gao, Guangping et al. (2011) Inverse zonation of hepatocyte transduction with AAV vectors between mice and non-human primates. Mol Genet Metab 104:395-403|
|Wang, Lili; Calcedo, Roberto; Bell, Peter et al. (2011) Impact of pre-existing immunity on gene transfer to nonhuman primate liver with adeno-associated virus 8 vectors. Hum Gene Ther 22:1389-401|
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