Influence of Laboratory and Clinical Factors on the Assessment of Gastric Serum Parameters

Insulin-like Growth Factor Binding Protein-4 (IGFBP-4) is a binding protein that enhances the half-life and regulates the activity of Insulin-like Growth Factor-1 (IGF-1).The downstream effects of IGF-1 are also modulated by Epidermal Growth Factor (EGF) whereby EGF prevents the reduction of IGF-1R expression by IGF-1, resulting in synergistic effects on intestinal epithelial proliferation.We have previously shown that IGFBP-4 whole body knockout (KO) mice have larger intestinal crypts compared to wild-type (WT) mice, and that IGFBP-4 dose-dependently decreases epithelial proliferation in vitro, establishing IGFBP-4 as an inhibitor of basal intestinal proliferation.However, the mechanism of IGFBP-4 action and regulation of its expression are unclear.We hypothesized that IGFBP-4 inhibits intestinal proliferation by inhibiting the IGF-1-IGF-1R axis, and that IGF-1 and EGF together stimulate the expression of IGFBP-4.To study the mechanism through which IGFBP-4 inhibits proliferation, rat intestinal epithelial IEC-6 cells were treated with IGFBP-4 or vehicle and/or IGF-1R inhibitor (NVP-AEW541).While IGFBP-4 and NVP-AEW541 both inhibited IEC-6 proliferation, IGFBP-4 with NVP-AEW541 together did not additively reduce proliferation.In order to study the regulation of IGFBP-4 expression, IEC-6 cells were treated for 6hr with IGF-1 and/or EGF.IGF-1+EGF treatment enhanced IGFBP-4 mRNA expression by 1.4-fold while this response was absent with IGF-1 or EGF alone.In addition, immunofluorescence of IGFBP-4 increased 1.9-fold with EGF and 1.8-fold with IGF-1+EGF treatment for 24hr.Similarly, by immunoblot, we observed 1.3-and 1.4-fold increases in cellular IGFBP-4 protein expression with 24hr EGF and IGF-1+EGF treatment respectively, and a 2-fold increase in IGFBP-4 secretion with IGF-1+EGF treatment.To study the regulation of IGFBP-4 expression in vivo, mucosal scrapes from inducible, intestinal epithelial IGF-1 receptor (IE-IGF-1R) KO and WT mice were collected 11d after induction of KO.Mucosal IGFBP-4 mRNA expression was not different for KO mice.Next, the IE-IGF-1R KO mice and WT mice were given an oral ErbB inhibitor (CI-1033) or vehicle for 11d.There were no differences in intestinal weight, length, or body weight across groups.However, IE-IGF-1R KO mice given CI-1033 displayed a 2-fold increase in mucosal IGFBP-4 mRNA expression compared to WT mice given CI-1033.Our studies demonstrate that IGFBP-4 exerts its inhibitory effects on proliferation by inhibiting the IGF-1-IGF-1R axis and that IGF-1+EGF together increase IGFBP-4 expression and secretion in vitro.However in vivo, IGFBP-4 expression is upregulated in response to inhibition of the ErbB and IGF-1R signaling, potentially enhancing the half-life of IGF-1 within the intestinal mesenchyme.These findings suggest a nuanced role for IGFBP-4 in regulating intestinal proliferation.