Event Title
Downregulation of Caveolin-1 on Acinar Formation of Mammary Epithelial Cells
Location
Suwanee, GA
Start Date
15-5-2018 1:00 PM
Description
Altered expression of caveolin-1 (cav-1), the main structural protein of caveolae responsible for macromolecular transcytosis and regulation of signal transduction, has been shown to support breast tumorigenesis. We have shown a link between the loss of cav-1 and abnormal ductal architecture as well as fibronectin expression and epithelial cell growth in murine mammary glands. This study evaluates the role of the downregulation of cav-1 on the development of mammary epithelial acini, the structural/functional component of mammary glands. To determine if mammary epithelial cells form acinar structures on Matrigel, a basement membrane extract, human mammary epithelial MCF10-2A were grown atop a Matrigel matrix and cultured for varying time points. Acinar growth/structure was analyzed using immunofluorescence for the nuclear marker DAPI. Using a confocal microscope, the thickness of the acinar structures were analyzed and measured using ImageJ along with the circumference of 20x DAPI stained acinar structures. Results from these analyses showed an increase in both thickness and width of the structures. To downregulate cav-1 in MCF10-2A cells, shRNA against cav-1 was used. Eight unique trials of transfection methods and reagents were utilized to optimize transfection efficiency. Results showed 3ul lipofectamine and 20ul GFP was most efficientInitial results show healthy acinar growth on Matrigel and successful transfection of GFP; however, additional trials will be performed.
Embargo Period
8-15-2018
Downregulation of Caveolin-1 on Acinar Formation of Mammary Epithelial Cells
Suwanee, GA
Altered expression of caveolin-1 (cav-1), the main structural protein of caveolae responsible for macromolecular transcytosis and regulation of signal transduction, has been shown to support breast tumorigenesis. We have shown a link between the loss of cav-1 and abnormal ductal architecture as well as fibronectin expression and epithelial cell growth in murine mammary glands. This study evaluates the role of the downregulation of cav-1 on the development of mammary epithelial acini, the structural/functional component of mammary glands. To determine if mammary epithelial cells form acinar structures on Matrigel, a basement membrane extract, human mammary epithelial MCF10-2A were grown atop a Matrigel matrix and cultured for varying time points. Acinar growth/structure was analyzed using immunofluorescence for the nuclear marker DAPI. Using a confocal microscope, the thickness of the acinar structures were analyzed and measured using ImageJ along with the circumference of 20x DAPI stained acinar structures. Results from these analyses showed an increase in both thickness and width of the structures. To downregulate cav-1 in MCF10-2A cells, shRNA against cav-1 was used. Eight unique trials of transfection methods and reagents were utilized to optimize transfection efficiency. Results showed 3ul lipofectamine and 20ul GFP was most efficientInitial results show healthy acinar growth on Matrigel and successful transfection of GFP; however, additional trials will be performed.