Breast cancer cells (SKBR-3) did not contain any detectable CTF or AdipoR1 but did contain IDE and TACE (See on-line supplement for cell and tissue staining procedure at www.​extras.​springer.​com/​2015/​978-3-319-21926-4). As CTF is absent in the cell but the target downstream protease IDE are present, SKBR-3 makes a good model for measuring the impact of CTF on intra-cellular insulin. Treatment of SKBR-3 with CTF increases the amount of intercellular insulin (See Fig. 5.1). The number of insulin loaded cells double when exposed to 10 ug/mL of CTF. The insulin loaded cells clearly showed intercellular fluorescence from the labeled insulin. The insulin loading cell model allows screening of compound to block CTF inhibition of IDE.

Cells treated with CTF maintain viability (proliferation) with no observable cell death (apoptosis) (See Fig. 5.1). Cardiomyocytes exposed to CTF also did not modulate phosphorylation of ERK and MAPK-p38 or slow proliferation (See Table 5.1). No observable cell death was noted. As insulin signaling of the PI3K/AKT/mTOR pathway increases proliferation and decreases apoptosis, insulin loading of cells would be expected to have reduce insulin receptor response and proliferation. However no impact to proliferation observed. It appears that CTF does not inhibit PI3K/AKT/mTOR intercellular signaling. This could be due to the excess of insulin used in cell culture media. An insulin straved model was not done. This proliferation effect contrasts the apoptotic impact of Adpn, which did suppress phosphorylation of ERK in cardiomyocytes (See Table 5.1) and restored intra-cellular Ca²⁺. Restored intra-cellular Ca² would inhibit calcium dependent PI3K pathway and desensitize insulin response.

Normal and diabetic rats were injected with CTF as part of a glucose tolerance challenge (See Tables 5.2–5.4 and Fig. 5.2). Normal and diabetic rats showed different behaviors. In normal rat showed substantial plasma insulin increases compared to control (See top two graphs of Fig. 5.2). Glucose was not reduced and insulin levels in plasma doubled in normal rats. This behavior matched an insulin resistance behavior. Diabetic rats which already had with impair insulin production (<10 % plasma insulin levels of normals) did not exhibit an increase in plasma insulin upon CTF exposure (See bottom two graphs of Fig. 5.2). It was further shown that diabetic rats post treatment had CTF loading in liver, pancreas, brain and lymph nodes where as normal rats did not (See Chap. 4).