VASCULAR ENDOTHELIAL GROWTH FACTOR IS PRESENT IN HUMAN MILK AND ITS RECEPTOR ON INTESTINAL EPITHELIAL CELLS

Abstract

33 Aims: VEGF (vascular endothelial growth factor) is a multifunctional cytokine active on blood vessel cells. Its concentration in the serum of normal adults ranges between 0 and 0.227 mg/ml. We hypothesized that breast milk, a biological fluid rich in growth factors, may contain VEGF which may bind on specific receptors on the newborn intestinal mucosa. Methods: Monomeric VEGF, containing 165 residues, (VEGF165) was examined by ELISA in the breast milk of 43 mothers, up to 5 months postpartum. 20/43 had premature (<37 weeks) and 23/43 term gestation. Breast milk VEGF concentration in the first week of lactation, were compared between mothers of infants born at term (n=13) and mothers of preterm (n=20) infants. 7/20 mothers of preterm infants had gestational age of 26-31 weeks and 13/20 of 32-36 weeks. We also compared the VEGF concentrations in the breast milk of mothers of fullterm infants during the first week of lactation (n=13) with the second (n=7). The interaction of VEGF with the intestinal epithelial cells was assessed on Caco-2 cells, a recognized model of intestinal epithelium. To determine the specific binding of VEGF, we incubated Caco-2 cells with increasing concentrations of 1251-VEGF165 in the presence of 100-fold excess of unlabeled recombinant human VEGF165. The proliferation effect of VEGF on intestinal epithelial cells was assessed in serum free media. The receptor, Flt-1, mRNA was examined in Caco-2 cells by RT-PCR. Human endothelial cells and fibroblasts were used as positive and negative controls respectively. Results: The concentration of VEGF in the breast milk was two orders of magnitude greater than in serum (mean 54.1 + SD 31.5 ng/ml). The mean VEGF of the mothers who had fullterm pregnancy (82.2 ± SD 41.3 ng/ml) was greater than those whose gestational age were 26-31 weeks (42.7 ± SD 26 ng/ml) or 32-36 weeks (37.5 ± SD27.3 ng/ml), p<0.05. VEGF decreased with lactation in fullterm infants: first week postpartum (mean 82.2 ± standard deviation 41.3 ng/ml) was greater than the second week (mean 29.3 ± SD 21.4 ng/ml), p< 0.01. Scatchard analysis of radioligand-receptor binding demonstrated the presence of specific receptors for 1251-VEGF165 on the surface of Caco-2 cells with a Kd of 2.85 to 4 nM. Flt-1 mRNA was detected in Caco-2 cells by RT-PCR and not in fibroblasts. VEGF had no effect on intestinal proliferation. Conclusion: VEGF is present in high concentrations in human milk. It binds specifically to receptors on the apical surface of intestinal epithelial cells. These receptors are likely to be of the flt-1 type because flt-1 receptor mRNA was detected by PCR. Interactions between VEGF and its receptor in intestinal epithelial cells may therefore occur in the newborn infant.