Abstract
The short‐form glucose‐dependent insulinotropic polypeptide (GIP) (1–30) is released from islet alpha cells and promotes insulin secretion in a paracrine manner in vitro. However, it is not well elucidated how GIP (1–30) is involved in glucose metabolism in vivo, since a specific assay system for GIP (1–30) has not yet been established. We first developed a sandwich enzyme‐linked immunosorbent assay (ELISA) specific for GIP (1–30) by combining a novel antibody specific to the GIP (1–30) C terminus with the common antibody against GIP N terminus. Then, we explored cross‐reactivities with incretins and glucagon‐related peptides in this ELISA. GIP (1–30) amide, but not GIP (1–42), GLP‐1, or glucagon increased absorbance in a dose‐dependent manner. We next measured plasma GIP (1–30) concentrations in nondiabetic participants (ND) during a 75‐g oral glucose tolerance test or cookie meal test (carbohydrates 75 g, lipids 28.5 g, proteins 8.5 g). Both glucose and cookie load increased GIP (1–30) concentrations in ND, but the increases were much lower than those of GIP (1–42). Furthermore, the DPP‐4 inhibitor significantly increased GIP (1–30) concentrations similarly to GIP (1–42) in ND. In conclusion, we for the first time developed an ELISA specific for GIP (1–30) and revealed its secretion in ND.
Highlights
Glucose-dependent insulinotropic polypeptide (GIP) was purified from porcine gut mucosal extracts and was originally termed “gastric inhibitory polypeptide” based on its gastric acid inhibitory activity (Brown, Mutt, & Pederson, 1970; Brown, Pederson, Jorpes, & Mutt, 1969)
GIP is established as an incretin, which consists of 42 amino acids, and is secreted from intestinal K cells upon food intake and promotes insulin secretion from pancreatic beta cells in a glucose-dependent manner (Baggio & Drucker, 2007)
We developed a sandwich enzyme-linked immunosorbent assay (ELISA) for GIP (1–30) amide, which consisted of two antibodies, 6A1A and 72A1 (Figure 1a). 6A1A is a mouse monoclonal antibody that binds to the N terminus of active forms of GIP such as GIP (1–42) and GIP (1–30) (27201, IBL). 72A1 was used as a capture antibody to C terminus of GIP (1–30) and HRP conjugated 6A1A Fab’ was used as a detection antibody
Summary
Glucose-dependent insulinotropic polypeptide (GIP) was purified from porcine gut mucosal extracts and was originally termed “gastric inhibitory polypeptide” based on its gastric acid inhibitory activity (Brown, Mutt, & Pederson, 1970; Brown, Pederson, Jorpes, & Mutt, 1969). Several initial reports showed that GIP immunoreactivity was observed in the islet alpha cells (Ahrén, Håkanson, Lundquist, & Sjölund, 1981; Alumets, Håkanson, O'Dorisio, Sjölund, & Sundler, 1978; Smith, Merchant, Johnson, Fujimoto, & Williams, 1977) Afterward, those observations were suspected due to the homology between GIP and glucagon and GIP was concluded not to be a constituent of the mammalian pancreas, after development of monoclonal antibody of the C-terminus of GIP (1–42) (Buchan, Ingman-Baker, Levy, & Brown, 1982). Fujita and colleagues reported that “short-form” GIP—GIP (1–30), which consists of 30 amino acids—is localized in the islet alpha cells and promotes insulin secretion in a paracrine manner (Fujita, Wideman, et al, 2010) They showed that GIP (1–30) retained insulinotropic activity equivalently to GIP (1–42) by using perfused mouse pancreas (Fujita, Wideman, et al, 2010). Here, we first developed a sandwich enzyme-linked immunosorbent assay (ELISA) system specific for GIP (1–30), evaluated its specificity and measured plasma GIP (1–30) concentrations in nondiabetic participants
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