In Situ Phosphorylation of Akt and ERK1/2 in Rat Mammary Gland, Colon, and Liver Following Treatment with Human Insulin and IGF-1

Abstract

High doses of insulin and the insulin analog AspB10 have been reported to increase mammary tumor incidence in female rats likely via receptormediated mechanisms, possibly involving enhanced IGF-1 receptor activation. However, insulin and IGF-1 receptor functionality and intracellular signaling in the rat mammary gland in vivo is essentially unexplored. The authors investigated the effect of a single subcutaneous dose of 600 nmol/ kg human insulin or IGF-1 on Akt and ERK1/2 phosphorylation in rat liver, colon, and mammary gland. Rat tissues were examined by Western blotting and immunohistochemistry by phosphorylation-specific antibodies. Insulin as well as IGF-1 caused Akt phosphorylation in mammary epithelial cells, with myoepithelial and basal epithelial cells being most sensitive. IGF-1 caused stronger Akt phosphorylation than insulin in mammary gland epithelial cells. Phosphorylation of ERK1/2 was not influenced by insulin or IGF-1. Rather, in liver and mammary gland P-ERK1/2 appeared to correlate with estrous cycling, supporting that ERK1/2 has important physiological roles in these two organs. In short, these findings supported that the rat mammary gland epithelium expresses functional insulin and IGF-1 receptors and that phosphorylation of Akt as well as ERK1/2 may be of value in understanding the effects of exogenous insulin in the rat mammary gland and colon.

Surface‐expressed insulin receptors as well as IGF‐I receptors both contribute to the mitogenic effects of human insulin and its analogues

“…Such a mechanism is plausible, because Akt is a recognized oncogene, and gain-of-function Akt mutations are involved in many human cancers (Altomare and Testa, 2005). Interestingly, insulin has recently been shown to cause rapid Akt phosphorylation in the rat mammary gland in vivo (Hvid et al, 2011).…”

“…These cells represent tissue where carcinogenic and growth-promoting effects of insulin have been observed in vivo in rats and mice (Tran et al, 2006;Hvid et al, 2011), as well as the liver, which represents target tissue for the classic metabolic effects of insulin. Therefore, cells such as these are considered relevant for inclusion in cell panels for preclinical safety assessment of new insulin analogues.…”

Metabolic effect and receptor signalling profile of a non-metabolisable insulin glargine analogue

“…injection of a 4 nmol/kg dose (Ikeda et al ., 2009). In addition, the s.c. injection of a supraphysiological dose (600 nmol/kg) of IGF-1 increased Akt phosphorylation in liver, colon and mammary gland of Sprague–Dawley rats (Hvid et al ., 2011). Akt and ERK phosphorylation also occurred in mouse mammary gland tissue only after a large bolus tail vein injection of IGF-1 (Lee et al ., 2003).…”

Differences in metabolic and mitogenic signalling of insulin glargine and insulin aspart B10 in rats

“…The insulin and IGF receptors trigger a complex range of intracellular signals for metabolism, cell growth and proliferation [16]. Their relative abundance affects intracellular signalling and has important consequences for tissue-specific responses to insulin, IGFs and insulin analogues [17, 18]. In addition, it has been demonstrated that overexpression of IR and IGF1R in human breast carcinomas allows insulin and IGF-1 hybrid receptors to form.…”

“…Previous studies have been performed in mice [39, 40]. The one rat study that has been reported to date used suprapharmacological doses and only investigated downstream signalling [17]. …”