Ricerc@Sapienza

Female strobilus inflorescences (hops or cones) from Humulus lupulus L., commonly named hop cones or hops, are widely used in the brewing industry as preservative and flavouring additives, as well as in traditional medicine as remedies for mild sleeping disorders and as bitter stomachic. Polyphenolic compounds, mainly prenylflavonoids (viz. xanthohumol), catechins and procyanidins represent some of the characteristic phytoconstituents. Hop constituents, mainly xanthohumol and its metabolites, seems to be responsible for a variety of interesting biological properties, including antioxidant, antinflammatory, chemopreventive, antimicrobial and antiviral. On the other hand, also proanthocyanidins and catechins have been described to possess antiviral properties, particularly against influenza A, which represents a major public health problem, with high rates of morbidity and mortality. Although different strategies have been approached to prevent the disease and/or manage its complications, the development of mutant strains increase the need for further and more effective therapeutic options. Experimental evidences suggested that herbal medicines, likely due to their polyphenolic composition, can represent alternative or integrative strategies for influenza. In this context, the present study was aimed at evaluating the ability of a hydroalcoholic extract from female inflorescences of H. lupulus (provided by EPO S.r.l.), standardized to contain 0.4 % of flavonoids, to interfere with the infection development of influenza virus. To perform the experiments, Madin-Darby Canine Kidney (MDCK) epithelial cells, permissive to influenza virus replication, and the human influenza virus A/Puerto Rico/8/34 H1N1 (A/PR8) virus were used. Cytotoxicity of the treatment was evaluated by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-2H-tetrazoliumbromide (MTT) assay, while the hemagglutination inhibition assay was performed to measure the viral titre. The virus protein expression was evaluated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and western blotting analysis. The effect of the sample on the intracellular levels of GSH were also measured, being the virus infection associated to oxidative stress. In order to highlight the possible antioxidant mechanisms of the sample, the radical scavenging power and the inhibition of lipoperoxidation were assayed [6]. The polyphenolic composition of the H. lupulus extract was evaluated by chromatographic and colorimetric methods [6]. In the range of the concentrations tested (10-140 ?g/ml), the H. lupulus extract exhibited no cytotoxicity on MDCK; in contrast, it induced a statistically significant and concentration dependent inhibition of the A/PR8 H1N1 virus replication, reaching about 80% inhibition at concentration of 140 ?g/ml. A significant inhibition of viral proteins, mainly the late ones, was highlighted after treatment with the hop extract, along with an increase level of GSH into infected cells, so suggesting that a modulation of the intracellular redox state can be involved in the antiviral activity found. The sample also able to scavenge different radicals and to interfere with lipoperoxidation, thus allowing hypothesize possible protective effects on the infected cell. The phytochemical analysis of the extract highlighted the presence of different polyphenolic compounds, among which flavonoids, flavanols and tannins: the flavonoid content agreed with that declared by the supplier. In conclusion, the antiviral and antioxidant properties here found, highlight the need for further in vivo studies on the tested H. lupulus extract, in order to hypothesize its possible application as anti-influenza dietary supplement.