Ovotransferrin expression and release by chicken cell lines infected with Marek's disease virus

Mammals posses both serum transferrin and lactoferrin, whose functions are taken over in birds by ovotransferrin, displaying both iron transport and antibacterial activities, Ovotransferrin also exerts antiviral activity towards Marek's disease virus, an avian member of the herpes famiJy of viruses, This virus infects Iymphoid organs and induces the transcription of ovotransferrin in infected chicken embryo fibroblasts. However, it has not yet been established whether ovotransferrin gene transcription is Iinked to the release or the protein outside the cells or whether ovotransferrin expression and release also occurs in chicken lymphoblastoid cclls in which the Marek 's disease virai genome is integrated. Our results indicate that both serum and egg-white isoforms of ovotransferrin are expressed and released in the supernatants of chicken embryo fibroblast and Iymphoblastoid cells in tbe absence of infection. ViraI infection of chicken embryo fibroblasts caused a slight increase of ovotransferrin release, whereas viraJ reinfection of lyrnphoblastoid cells caused a rernarkable ovotransferrin release in a virus coucentratìon-dependent manner. These findings suggest rhat ovotransferrin release in vivo may play a cruciaJ role in protecting the whole organism from virai infection spreading, and support the hypothesis that the antiviral activity of ovotransferrin is an important part of the innate immune response in birds, resembling the anriviral activity of lactoferrin in marnmals

Anti-invasive activity of bovine lactoferrin towards group A streptococci

Group A streptococci (GAS) are able to invade cultured epithelial and endothelial cells without evidence of intracellular replication. GAS, like other facultative intracellular bacterial pathogens, evolved such ability to enter and to survive within host cells avoiding the host defences, and bacterial intracellular survival could explain the recurrence of infections. We report here that 1 mg bovine lactoferrin (bLf)/mL significantly hindered the in vitro invasion of cultured epithelial cells by GAS isolated from patients suffering from pharyngitis and completely inhibited the invasiveness of GAS pretreated with subinhibiting concentrations of erythromycin or ampicillin. One milligram of bLf per millilitre was also able to increase the number of epithelial cells undergoing apoptosis following GAS invasion, although the number of intracellular GAS in the presence of bLf decreased by about 10-fold. The ability of bLf to decrease GAS invasion was confirmed by an in vivo trial carried out on 12 children suffering from pharyngitis and already scheduled for tonsillectomy. In tonsil specimens from children treated for 15 days before tonsillectomy with both oral erythromycin (500 mg t.i.d. (three times daily)) and bLf gargles (100 mg t.i.d.), a lower number of intracellular GAS was found in comparison with that retrieved in tonsil specimens from children treated with erythromycin alone (500 mg t.i.d.)

Proteolytic activity of bovine lactoferrin

Bovine lactoferrin catalyzes the hydrolysis of synthetic substrates (i.e., Z-aminoacyl-7-amido-4-methylcoumarin). Values of Km and kcat for the bovine lactoferrin catalyzed hydrolysis of Z-Phe-Arg-7-amido-4-methylcoumarin are 50 μM and 0.03 s−1, respectively, the optimum pH value is 7.5 at 25 ◦C. The bovine lactoferrin substrate specificity is similar to that of trypsin, while the hydrolysis rate is several orders of magnitude lower than that of trypsin. The bovine lactoferrin catalytic activity is irreversibly inhibited by the serine-protease inhibitors PMSF and Pefabloc. Moreover, both iron-saturation of the protein and LPS addition strongly inhibit the bovine lactoferrin activity. Interestingly, bovine lactoferrin undergoes partial auto-proteolytic cleavage at positions Arg415-Lys 416 and Lys440-Lys441. pKa shift calculations indicate that several Ser residues of bovine lactoferrin display the high nucleophilicity required to potentially catalyze substrate cleavage. However, a definitive identification of the active site awaits further studies