Fetal intestinal fibroblasts respond to insulin-like growth factor (IGF)-II better than adult intestinal fibroblasts

Abstract

Background: We compared IGF responses of fetal and adult intestinal fibroblasts to identify a developmental difference in the IGF-axis. Intestinal fibroblasts were isolated from maternal and fetal jejunum. Media was conditioned at confluence and one week afterwards. The proliferative response at confluence to 5 nM IGF-I or -II was compared.

Results: There were no significant differences in IGFBP expression at confluence. Post-confluence, fetal fibroblasts had no significant changes in IGFBP-2 and IGFBP-3 expression. Post-confluent maternal fibroblasts had increased IGFBP-3 levels that were significant compared to the fetal fibroblasts. IGF-I increased in post-confluent fetal fibroblasts, while in maternal fibroblasts it decreased (p < 0.001). IGF-II secretion decreased significantly in post-confluent maternal fibroblasts (p < 0.05). Maternal fibroblasts proliferated more with IGF-I than IGF-II (p < 0.001). Fetal fibroblasts responded to IGF-II slightly better than IGF-I and significantly greater than maternal cells (p < 0.001).

Conclusion: Fetal intestinal fibroblasts respond to IGF-II with greater proliferation and do not have the increased IGFBPs seen post-confluence in adult intestinal fibroblasts.

Figure 1

Concentrations of IGF-I and IGF-II in the conditioned media. The maternal and fetal intestinal fibroblasts produced primarily IGF-II. In this representative experiment the mean and SEM data represents the pooled data from triplicate samples from the three cell lines. The fetal fibroblasts produced greater concentrations of IGF-I and IGF-II than maternal fibroblasts but did not achieve statistical significance (8.4 ± 1.7 vs. 5.1 ± 0.03 ng/ml IGF-I, p = 0.53 and 40.3 ± 3.0 vs. 29.99 ± 1.3 ng/ml IGF-II, p = 0.08). The one week post-confluence fetal fibroblasts had increased IGF-I secretion (13.9 ± 0.4), while that of the maternal fibroblasts decreased (2.7 ± 0.3,* p < 0.001). The IGF-II secretion was decreased slightly one week post-confluence in fetal fibroblasts (29.9 ± 2.3) and significantly in maternal fibroblasts (to 9.5 ± 0.8, ^p < 0.05).

Figure 2

Ligand blot comparison of IGFBP expression. Three IGFBP bands were seen consistently with relative molecular weights that have been characterized for ovine IGFBPs-3 (42–50 kDa doublet), -2 (33 kDa) and -4 (24 kDa) [7,11,12]. The fibroblasts produced primarily IGFBP-2 in both maternal and fetal cell lines. The IGFBP levels did not significantly change one week post-confluence in fetal cells compared to maternal cells; which had a general increase in IGFBP expression.

Figure 3

The fibroblast response to IGF-I and IGF-II. The MTT assay demonstrated that all populations of fibroblasts responded to IGF treatment with statistically significant (p < 0.001) greater cell numbers. In this representative experiment the mean and SEM data represents the pooled data from duplicate samples from the three cell lines. Maternal fibroblasts responded with greater cell numbers for IGF-I than IGF-II compared to controls (88% vs 55% increase,* p < 0.001). Fetal fibroblasts responded to IGF-II better than IGF-I, although not significantly, and at the same rate as maternal fibroblasts responded to IGF-I (88% vs 79%, p = 0.34). The amount of increase in the fetal cells in response to IGF-II was significantly greater that the amount of increase seen in the maternal cells (^p < 0.001). Error bars are standard error of the mean calculated from the original absorbance data and divided by the mean absorbance of the controls to create a percent value.