In this study, we show that human milk immunoglobulin fraction (sIgA), fSC and Lf inhibit adhesion and invasion to HeLa cells by three Shigella species. The inhibitory activity seems to be similar among the three Shigella species studied. Thus, the human milk inhibitory activity could have the same target mechanism, impairing the virulence of the three species studied.
Several reports have pointed out an important role for immunoglobulins, mainly sIgA, on human milk protective effect against diarrhea [14, 17, 18]. sIgA inhibits the adherence of many pathotypes of E. coli, including EIEC which shows similar virulence aspects when compared to Shigella [18, 19]. Similarly, two glycoproteins, Lf and fSC, were shown to be involved in the inhibition of ETEC, EAEC and EPEC adherence [23, 24]. In the present work, we have shown that these two glycoproteins, in concentrations lower than those found in human milk, inhibited the adhesion and, consequently, the invasion of the three Shigella species. Furthermore, our findings indicate that sIgA and Lf bind to Shigella superficial proteins with molecular mass similar to the virulence proteins involved in the invasion mechanism , suggesting that these components could inhibit the two virulence processes independently.
Little is know about the role the fSC found in secretions, although some studies suggest that it may play a protective role against diarrhea [18, 20]. In our work show that fSC from human milk inhibited the adhesion and invasion of Shigella in vitro, indicating that this compound may be an important non-specific defense factor. Interestingly, fSC is also found in intestinal secretions where it may provide protection to mucous membranes, the initial target in most infections. Our findings further corroborated this hypothesis since anti-SC diminishes the inhibition activity of sIgA and fSC on Shigella invasion and adhesion. Furthermore, it is interesting to note that anti-SC inhibits the effect of both sIgA and fSC, therefore the bounded SC of sIgA may be involved on the inhibitory activity. On the other hand, sIgA complexed with the anti-SC may lead to esteric impairment.
Lf is considered as an important non-specific defense factor against gastrointestinal infections, causing microorganisms iron-deprivation by its iron-scavenging ability [3, 26]. In the present work, we show that Lf isolated from the human milk also affected adhesion and invasion of S. flexneri BS176 (S. flexneri M90T cured of the virulence plasmid), S. flexneri, S. dysenteriae and S. sonnei wild type strains.
Previous studies  have established the association of fucosylated oligosaccharides in human milk with adhesion inhibition of enteropathogens to Hep-2 cells. Lf, sIgA and fSC, rich in fucosylated residues, when treated with sodium metaperiodate, lose the ability to inhibit both, Shigella adhesion and invasion. This result confirms the importance of the sugar residues in natural glycoproteins. Gomez et al. showed that recombinant Lf have no effect on the adhesion of the S. flexneri M90T strain to HeLa cells , probably, due to the recombinant lactoferrin not to possess the same structure of sugar residues, loosing the capacity of inhibiting the Shigella adhesion.
Studies have suggested a role of human milk glycocompounds as cell surface homologues, inhibiting pathogen binding to host cell receptors . Newburg et al., 1992, described that glycolipid Gb3 from human milk, a natural shiga toxin receptor, could null the effect of this toxin on the host cells . Human proteins, such as Lf and fSC could play the same role, probably acting as receptor analogues of adhesins or invasines. On the other hand, the work of Gomez et al.  indicates that recombinant lactoferrin promotes a degradation of virulence factors, such as Ipa B, impairing the formation of the IpaBC complex on the surface of the host cell. Moreover, our findings suggest that sIgA and Lf bind to proteins with similar mass of Shigella virulent proteins: IpaJ and MxiJ (26 kDa), VirB (35 kDa), IpaD (37 kDa), VirA (45 kDa) or IpaA (68 kDa). These results suggest that Lf and sIgA prevent the interaction between the bacteria and host cells, impairing the invasion complex formation.