Sb Modulates avb5 Integrin Activity treatment of gastroenteritis and IBD [18] through the modulation of host cell signaling pathways implicated in the pro-inflammatory response. Moreover, we previously demonstrated both in vitro and in mice that Sb promoted intestinal restitution [15]. In the present study, we analyzed the effect of Sb on the capacity of intestinal epithelial cells to heal a wound. According to our findings, the impact of Sb on epithelial cells can be summarized as follows: (1) SbS contains heat-sensitive compounds that decreased avb5 integrin-mediated adhesion to Vn by competing with the integrin; (2) SbS-mediated changes in cell adhesion to Vn results in a reduction of the avb5 signaling pathway; (3) This perturbation is related to Sb-induced cell migration. According to these data, we postulated that SbS contains motogenic factor(s) that improved intestinal cell restitution by down-modulating the avb5 integrin- Vn interaction. We demonstrated that SbS contains factors that blocked the cell- Vn interaction. Indeed, our adhesion assays showed that SbS competed with av integrin subunit in binding to Vn. Indeed, (1) SbS blocked cell adhesion to Vn and (2) addition of SbS onto Vn prior to cell plating impaired cell adhesion. This blockade is selective since SbS did not block the cell interaction with Fn (this paper), laminin or type I collagen [15]. Moreover this phenom- enon could be common to all intestinal epithelial cell lines since the same results were observed using HT29-D4 cells, another intestinal cell line. Vn associates with a wide variety of ligands including av integrin, IGF-I family members, and urokinase plasminogen activator receptor [36] [11,37]. The binding site(s) present in Vn for SbS factors remain to be elucidated. One might postulate that the tripeptide sequence RGD, involved in binding integrin to both Fn and Vn, does not play a role in this process since SbS did not block cell interaction with Fn. Further studies are required to explore whether sequences flanking the RGD peptide could interact with SbS , as they have been reported to be important for integrin selectivity [38]. On the other hand, other Vn domains could be involved in the interaction with compound(s) found in SbS. For example, Protein E from Hemophilus influenza Figure 8. SbS alters the functionality of adherence structures. (A) HCT-8/E11 cells were plated on either fibronectin or vitronectin for 2 h and then fixed. Tyrosine phosphorylated proteins were stained using a PY20 mAb. (B) Focal adhesion labeling was quantified by measuring the area of each focal adhesion for all cells. Data represent the mean+SD of 3 separate experiments. (C) HCT-8/E11 cells were allowed to adhere on vitronectin after SbS pretreatment. The phosphorylation of tyrosine residues in FAK (Tyr-397FAK) was determined after cell lysis at the indicated times of cell adhesion. Samples were analyzed by western blot analysis. Equal amounts of protein were analyzed and loading amounts were verified by probing the blot with anti-FAK (FAK) or anti- b-tubulin (b-tubulin) Abs. *** P,0.001. doi:10.1371/journal.pone.0045047.g008 PLOS ONE | www.plosone.org 9 September 2012 | Volume 7 | Issue 9 | e45047