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VOL. 76 (1), 23-44, 2010 BETA-CELL HYPERPLASIA INDUCED BY HEPATIC INSULIN...

2.11. Proliferation studies in isolated pancreatic islets
2.11. and cultured cells

DNA synthesis in isolated islets was estimated by determining
BrdU incorporation by using the Cell Proliferation ELISA kit (Roche
Diagnostics GmbH, Germany). In cultured beta cells the DNA syn-
thesis was estimated by [3H]-thymidine incorporation as previously
described (20).

2.12. Statistical analysis

All values are expressed as mean +/– SEM. Statistical analyses
were carried out using a two-tailed Student’s unpaired t test, and the
null hypothesis was rejected at the 0.05 level.

3. RESULTS

3.1. Progressive liver-specific IR deletion without liver
3.1. dysfunction

The iLIRKO mice were generated as described in Materials
and Methods. These mice showed variable IR deletion and were
grouped into three groups (50, 25 and 0% of the normal receptor
complement). Ablation of IR was tissue-specific, none of other tissues
studied were affected (Figure 1).

A critical issue in constitutive LIRKO was the appearance of some
liver dysfunction. However, hematoxylin-eosin staining in liver
sections revealed no dysplastic or hyperplastic nodules in 6 month-
and 12 month-old iLIRKO mice as compared with their respective
controls. In addition, Masson staining showed no increase in collagen
infiltration or fibrosis in iLIRKO mice (Figure 2, upper panels).
Regarding enzymes of hepatic glucose metabolism, we found two
kind of evidence as compared with constitutive LIRKO mice. Thus,
late IR deletion in iLIRKO mice resulted in reduced glucokinase
(GK) protein expression and a dramatic loss of glycogen liver
content, as shown by PAS-staining of liver sections, as previously

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