Fatemeh Karami
1,2 
, Behrouz Asgari Abibeiglou
1, Saghar Pahlavanneshan
3, Ali Farrokhi
1, Amin Tamadon
4, Mohsen Basiri
1, Keynoosh Khalooghi
1, Majid Fallahi
1, Yaser Tahamtani
1,5* 1 Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
2 Faculty of Sciences and Advanced Technologies in Biology, University of Science and Culture, Tehran, Iran
3 Medical Nanotechnology and Tissue Engineering Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
4 Persian Gulf Marine Biotechnology Research Center, Persian Gulf Biomedical Sciences Research Institute, Bushehr University of Medical Sciences, Bushehr, Iran
5 Reproductive Epidemiology Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran
Abstract
Introduction: Measurement of pancreatic beta cell mass in animal models is a common assay in diabetes researches. Novel whole-organ clearance methods in conjunction with transgenic mouse models hold tremendous promise to improve beta cell mass measurement methods. Here, we proposed a refined method to estimate the beta cell mass using a new transgenic Tg(Pdx1-GFP) mouse model and a recently developed free-of-acrylamide clearing tissue (FACT) protocol.
Methods: First, we generated and evaluated a Tg(Pdx1-GFP) transgenic mouse model. Using the FACT protocol in our model, we could quantify the beta cell mass and alloxan-induced beta cell destruction in whole pancreas specimens.
Results: Compiled fluorescent images of pancreas resulted in enhanced beta cell mass characterization in FACT-cleared sections (2928869±120215 AU) compared to No-FACT cleared sections (1292372±325632 AU). Additionally, the total number of detected islets with this method was significantly higher than the other clearance methods (155.7 and 109, respectively). Using this method, we showed green fluorescent protein (GFP) expression confined to beta cells in Tg(Pdx1-GFP) transgenic. This enhanced GFP expression enabled us to accurately measure beta cell loss in a beta cell destruction model. The results suggest that our proposed method can be used as a simple, and rapid assay for beta cell mass measurement in islet biology and diabetes studies.
Conclusion: The Tg(Pdx1-GFP) transgenic mouse in conjunction with the FACT protocol can enhance large-scale screening studies in the field of diabetes.