Core B: Cell Culture Core
The Cell Culture and Physiology Core, designated as Core B, is a centralized facility that provides primary alveolar epithelial type 2 cells and secondary alveolar epithelial cell lines to each of the projects within the program project. Centralization of the cell culture facilities ensures that a continuous supply of high quality and purity alveolar epithelial cells is available to each of the projects.
The Cell Culture Core B is responsible for the isolation and maintenance of primary cultures of human, rat, and mouse alveolar epithelial type 2 cells, rat alveolar epithelial type I cells, rat and mouse lung fibroblasts, rat and mouse alveolar and peritoneal macrophages, and rat lipofibroblasts. The Core uses standard techniques previously described by our group. The core will purchase and propagate human, rat, and mouse alveolar epithelial cell lines. The purity of the cell cultures will be verified using a variety of immunological and functional assays.
The Core B also provides rat and mouse precision-cut lung slices. Lung slices are provided using a Leica VT1000 S vibrating blade microtome vibratome and are viable up to 48 hours in culture.
The cell culture facility personnel have extensive experience in the isolation of all the different cell types as well as general cell culture techniques. The consolidation of the cell culture facilities provides an economical means of isolating and culturing lung cells.
Figure 1. Representative photomicrographs of primary rat alveolar type 2 cells. Primary rat alveolar epithelial type 2 cells (AT2) were isolated by Core B and plated onto coverslips. (A) Rat AT2 cells were immunostained for keratin, an epithelial cell marker, with antibodies against keratin 8 and 18. (B) Phase-contrast image of AT2 cells depicted in Figure 1A. (C) Rat AT2 cells were immunostained for SP-C, an alveolar epithelial type 2 cell marker.
Figure 2
Figure 2. Representative photomicrographs of primary mouse alveolar type 2 cells. Primary mouse alveolar epithelial type 2 cells (AT2) were isolated from wild type and genetically modified mice by Core B and plated onto coverslips. Mouse AT2 cells were immunostained for SP-C, an alveolar epithelial type 2 cell marker.
Figure 3
Figure 3. Representative photomicrographs of primary human alveolar type 2 cells. Primary human alveolar epithelial type 2 cells (AT2) were isolated by Core B and plated onto coverslips. Cells were immunostained for keratin, an epithelial cell marker, with antibodies against keratin 8 and 18 (right panel). Cells were immunostained for SP-C, an alveolar epithelial type 2 cell marker (left panel).
Figure 4
Figure 4. Representative photomicrographs of primary rat alveolar type 1 cells. Primary rat alveolar epithelial type 1 cells (ATI) were isolated by Core B and plated onto coverslips. Cells were immunostained for T1 alpha (left panel) and RAGE (right panel), markers of alveolar epithelial type 1 cells (right panel).
Figure 5
Figure 5. Representative photomicrographs of primary mouse fibroblasts. Primary mouse fibroblasts were isolated by Core B and plated onto coverslips. Cells were immunostained for vimentin, a mesenchymal cell marker.
Figure 6
Figure 6. Representative photomicrographs of primary rat lipofibroblast. Primary rat lipofibroblasts were isolated by Core B and plated onto coverslips. Cells were immunostained for vimentin (green), a mesenchymal cell marker. Oil red O staining was used to visualize lipid droplets (red).
Figure 7
Figure 7. Representative photomicrographs of mouse lung precision slices. Precision-cut mouse lung slices were obtained by Core B and amount of live (calcein-AM; green) and dead (EthD-1; red) cells was determined.
Faculty:
Karen M. Ridge , Core Leader
Emilia Lecuona, Co-Leader
