Research Paper Volume 15, Issue 2 pp 441—458

Figure 1. INK ATTAC mice as a model to remove p16Ink4a⁺ cells. (A) The INK ATTAC transgene is driven by the p16^Ink4a promoter and encodes a Caspase 8 moiety that upon B/B homodimerizer administration leads to specific apoptosis of this cell subpopulation and is compared to animals treated with vehicle (referred to as controls). FLAG tag and eGFP expression can be tracked to measure effectiveness of deletion and are a surrogate marker of senescence load. Diagram modified from [21]. (B) qPCR from islets of animals treated with B/B homodimerizer and compared to control, untreated animals. A significant decrease of the transgene transcripts Casp8 and eGFP and p16^Ink4a was observed after treatment with B/B. (C) Representative confocal picture of islets from B/B treated and untreated animals showing a significant decrease of FLAG staining. (D) Quantification of FLAG intensity using image analysis software (Image J) showed a significant decrease in senescence load in islets from animals treated with B/B homodimerizer; n_control = 5 animals, 290 islets analyzed; n_treated = 8 animals, 518 islets analyzed mean+/−SEM, ^*p = 0.0004 by unpaired t-test. (E) Increased number of apoptotic nuclei per field in pancreas from 8-month-old INK-ATTAC mice; n_control = 5 animals, 201 images analyzed; n_treated = 6 animals, 223 images analyzed; mean+/−SEM, ^*p = 0.02 by unpaired t-test. (F) Heatmap of islet scRNASeq expression of Cdkn2a (encodes p16^Ink4) in different islet cell types reveals enriched expression of Cdkn2a in β-cells. No significant changes of mean of (G) blood glucose levels (mg/dL), (H) body weight (g), (I) pancreatic weight (mg) of 8/9-month-old INK-ATTAC mice, n = 6 per group; and (J) Beta cell mass (mg) of 6-month-old INK-ATTAC mice, n_control = 7 and n_treated = 8.