2. Objectives Core C will provide resources for investigators studying samples from WT vs. null mice exposed to cigarette smoke (CS) vs. filtered air (FA). To accomplish this, we have developed five primary goals outlined below. 2.1. Goal 1: Provide Expertise in Modeling COPD in Mice: We will provide all three projects with the expertise and resources required to model COPD in mice. This includes access to individuals with expertise in CS exposure models (Dr. Owen), pulmonary inflammation and injury (Drs. Owen and Perrella), imaging systems (Dr. Owen), and histopathology (Dr. Perrella). We will standardize the murine model of CS-induced COPD for all three projects. Lung tissue from mice exposed to CS or FA will be used in functional aims in all three projects. Dr. Owen has been the Director of the Pulmonary Division Animal Models Core on the 9th floor of the Thorn Building at BWH for 5 years. This Core has been exposing mice to CS for -10 years. Dr. Owen supervises trained technicians to perform and quantify the exposures and to monitor mice being exposed to CS. By using the same highly trained and experienced personnel, we will ensure that mice are uniformly exposed to CS, exposures are tightly regulated, and mice are properly monitored throughout. 2.2. Goal 2: Development and Maintenance of Novel Lines of Mice for Study: Dr. Perrella has generated many gene-targeted lines of mice (18-23) and will oversee the development of new lines of mice. We have either already obtained mice for study (Ireb2-/- and Hhip+/- mice) or purchased ES cells transfected with gene-targeted constructs from the KOMP Repository which the HMS Transgenic Mouse Core will use to generate Hhip and Fam13a gene-targeted mice (vide infra). If unexpected problems arise that preclude study of the gene-targeted mice in our CS exposure model. Dr. Perrella is extremely well qualified to assist with troubleshooting of alternative cloning strategies. After new lines are generated. Dr. Owen will oversee the murine husbandry, expansion of colonies for study, and genotyping of all lines of mice in our barrier facility where we have ample murine housing space (>1000 cages). Dr. Owen will then coordinate transfers of the mice to the smoke exposure facility and their housing and CS vs. FA exposures thereafter (see 3.1). 2.3. Goal 3: Tissue Harvesting, Storage, Cataloging, and Distribution: Dr. Owen will supervise highly trained personnel to harvest lung tissue and blood, perform BAL, and isolate and culture alveolar macrophages and primary lung epithelial cells grown at an air-liquid interface (ALI). Core personnel will work closely with project investigators to ensure continuity among the projects for harvesting of DNA, RNA, cells, and lung tissue after CS or FA exposures. Core C will store, catalog, and distribute lung tissue and primary cells isolated from mice to all three projects to ensure efficient usage of tissues from mice exposed to CS. 2.4. Goal 4: Histopathology Service: Dr. Perrella has directed this service since 2001 and supervises an experienced histology technician (Bonna 1th). He will paraffin-embed and section tissues, prepare frozen sections of lungs, and perform routine histochemical staining and staining of lungs for markers of inflammation. This service will troubleshoot non-routine staining performed by PPG personnel as needed. 2.5. Goal 5: Imaging and Flow Cytometry: This Core component contains a bright field and epi-fluorescence microscope, image acquisition and analysis software, a six-color FACS Canto II flow cytometer, and access to Confocal and Electron Microscopy Cores and cell sorting cytometers within the Partners/HMS system. This Core component is overseen by Dr. Owen, and she will assist all of the PPG investigators using this resource. 2.6. Overall goals: By consolidating skilled staff and equipment and making them available to all projects. Core C will reduce the cost of producing high-quality data, facilitate and promote interactions between the different projects, and play a critically Important role in the successful completion of work proposed in this PPG.
Showing the most recent 10 out of 115 publications
