Real-time imaging of beta-lactoglobulin-targeted luciferase activity in the mammary glands of transgenic mice.
This study was aimed at establishing a new platform for real-time monitoring of milk-protein gene expression in the mammary glands. A transgenic reporter composed of the beta-lactoglobulin (BLG)/luciferase hybrid gene was targeted to the mammary glands of pregnant and lactating mice and luciferase activity was imaged in vivo with a low-light imaging system. The mammary glands of a 17-day pregnant mouse occupied an area comparable to that of a 6-day lactating mouse. Nevertheless, the intensity of the luciferase signal was much weaker and confined to regions in the inguinal and thoracic glands. A few small and defined locations of higher expression were also detected, indicating diversity in the initiation of this transgenic milk protein expression. In the lactating mice, high inter- and intra-heterogeneity among regions in a particular gland and among glands was demonstrated, and confirmed by ex vivo analysis of luciferase activity in mammary biopsies. The lack of correlation between luciferase activities and levels of beta-casein accumulation in these biopsies resulted, most probably, from the longer half-life of the native milk protein, compared to the activity of the transgenic marker in the tissue. Unilateral sealing of mammary glands for 4 hr resulted in complete abrogation of luciferase activity, establishing the BLG/luciferase transgene as a reliable tool to follow short-term stimuli. Dispersed mammary epithelial cells preserved luciferase activity in culture, and thus could be used for following mammary gland development after re-implantation. The bioluminescence-based methodology presented here eliminates averaging of heterogeneity in gene expression among glands, and misinterpretations resulting from sampling biopsies taken from inactive regions. Imaging luciferase expression in the mammary glands may enable an accurate monitoring of milk-protein gene expression during cyclic periods of development and apoptosis in a limited number of animals, and could be applied for reporting the consequences of selected drugs on milk-protein gene expression.
Barash
Itamar
I
Institute of Animal Science, Agricultural Research Organization, The Volcani Center, Bet Dagan, Israel. barashi@agri.huji.ac.il
Reichenstein
Moshe
M
D000818:Animals; D015153:Blotting, Western; D002364:Caseins / Q000378:metabolism; D003199:Computer Systems; D003952:Diagnostic Imaging / Q000379:methods*; D005260:Female; D005822:Genetic Vectors / Q000235:genetics; D007774:Lactation / Q000235:genetics / Q000378:metabolism; D007782:Lactoglobulins / Q000235:genetics*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism*; D008163:Luminescent Measurements; D008321:Mammary Glands, Animal / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D011247:Pregnancy; D011401:Promoter Regions, Genetic / Q000235:genetics
D002364:Caseins; D007782:Lactoglobulins; D008156:Luciferases