Development of factors to convert frequency to rate for -cell replication and apoptosis quantified by time-lapse video microscopy and immunohistochemistry

Saisho Y, Manesso E, Gurlo T, Huang CJ, Toffolo GM, Cobelli C, Butler PC. Development of factors to convert frequency to rate for -cell replication and apoptosis quantified by time-lapse video microscopy and immunohistochemistry. Am J Physiol Endocrinol Metab 296: E89 –E96, 2009. First published October 21, 2008; doi:10.1152/ajpendo.90697.2008.—An obstacle to development of methods to quantify -cell turnover from pancreas tissue is the lack of conversion factors for the frequency of -cell replication or apoptosis detected by immunohistochemistry to rates of replication or apoptosis. We addressed this obstacle in islets from 1-mo-old rats by quantifying the relationship between the rate of -cell replication observed directly by time-lapse video microscopy (TLVM) and the frequency of -cell replication in the same islets detected by immunohistochemistry using antibodies against Ki67 and insulin in the same islets fixed immediately after TLVM. Similarly, we quantified the rate of -cell apoptosis by TLVM and then the frequency of apoptosis in the same islets using TdT-mediated dUTP nick-end labeling and insulin. Conversion factors were developed by regression analysis. The conversion factor from Ki67 labeling frequency (%) to actual replication rate (%events/h) is 0.025 0.003 h . The conversion factor from TdT-mediated dUTP nick-end labeling frequency (%) to actual apoptosis rate (%events/h) is 0.41 0.05 h . These conversion factors will permit development of models to evaluate -cell turnover in fixed pancreas tissue.