Hydroalkoholischer Extrakt aus Sauerampferkraut (Rumex acetosa L.) zur Prophylaxe von Mundschleimhauterkrankungen und Periodontitis: Hemmung der bakteriellen Adhäsion und zentraler Virulenzfaktoren von Porphyromonas gingivalis

Zusammenfassung

Sauerampferkraut aus den oberirdischen Teilen von Rumex acetosa L. wird traditionell gegen entzündliche Erkrankungen der Mundhöhle verwendet. Die vorliegende Studie untersuchte den Einfluss eines Proanthocyanidin-angereicherten hydroalkoholischen Extraktes (RA1) auf die Adhäsion von Porphyromonas gingivalis, eines Bakteriums, welches ein zentraler Mitverursacher der chronischen und aggressiven Parodontitis darstellt. Ein weiteres Ziel der Untersuchungen war die Identifizierung von bioaktiven Leitstrukturen mit antiadhäsiver Aktivität und die Benennung der für die antiadhäsiven Effekte verantwortlichen molekularen Mechanismen. Der vollständig quantifizierte Extrakt RA1 (5-15 µg/ml) reduzierte die bakterielle Adhäsion von P. gingivalis an humanen KB-Mundschleimhautzellen in vitro dosisabhängig bis zu 90 %. Dies wurde sowohl in einem durchflusszytometrischen Test als auch mittels Konfokaler Laser Scanning Mikroskopie belegt. Die In-vitro-Datensätze konnten auch in einem In-situ-Modell an Buccalschleimhaut der Maus bestätigt werden. Die Kombination phytochemischer Methoden mit funktionellen Adhäsionsassays identifizierte für die aus RA1 isolierten Naturstoffen 1 bis 15 Epicatechin-3-O-gallat-(4β,8)-epicatechin-3’-O-gallat (syn. Procyanidin B2-di-gallat, Verbindung 8) als stark antiadhäsives Proanthocyanidin, welches als funktionelle Leitsubstanz des Extraktes angesehen wird. Struktur-Wirkungsbeziehungen zeigten, dass eine mindestens einfache Galloylierung von Flavan-3-olen für eine antiadhäsive Wirkung notwendig ist. Die Aktivität der Verbindungen wird durch Trihydroxylierung im B-Ring und durch Oligomerisierung weiter gesteigert.

Als molekularer Angriffspunkt für diese Wirkstoffe wurde der bakterielle Virulenzfaktor Arginin-Gingipain identifiziert, ein zentraler Virulenzfaktor von P. gingivalis, der neben der bakteriellen Adhäsion des Pathogens an der Wirtszelle auch proteolytischen Abbau von Wirtsproteinen, Modulation der immunologischen Wirtsabwehr und Reifung weiterer bakterieller Virulenzfaktoren induziert.

Aus diesen Untersuchungen zeigt sich ein ausreichendes Potenzial von Sauerampferextrakt zur weiteren präklinischen und klinischen Entwicklung zu zytoprotektiven Mundhygieneprodukten.

Abstract

Hydroalcoholic extracts from Rumex acetosa L. for prophylaxis of periodontitis: Inhibition of the bacterial adhesion and virulence factors of Porphyromonas gingivalis

The aerial parts of Rumex acetosa L. have been used in traditional European medicine for inflammatory diseases of the mouth epithelial tissue. The following study aimed to investigate the influence of a proanthocyanidin-enriched extract from R. acetosa against the adhesion of Porphyromonas gingivalis, a pathogen strongly involved in chronic and aggressive periodontitis. A further goal was to define the bioactive lead structures responsible for a potential antiadhesive activity and to characterize the underlying molecular mechanisms of the antiadhesive effects.

An extract of R. acetosa (RA1) with a defined mixture of flavan-3-ols, oligomeric proanthocyanidins and flavonoids, was used. Its impact on P. gingivalis adhesion to KB cells was studied by flow cytometry, confocal laser scanning microscopy and in situ adhesion assay using murine buccal tissue. RA1 and its compounds 1 to 15 were further investigated for additional effects on gingipain activity, hemagglutination and gene expression by RT-PCR.

RA1 (5 to 15 µg/mL) reduced P. gingivalis adhesion in a dose-dependent manner to about 90 %. Galloylated proanthocyanidins were confirmed to be responsible for this antiadhesive effect with epicatechin-3-O-gallate-(4β,8)-epicatechin-3’-O-gallate (syn. procyanidin B2-di-gallate) being the lead compound. Ungalloylated flavan-3-ols and oligomeric proanthocyanidins were inactive. RA1 and the galloylated proanthocyanidins strongly interact with the bacterial virulence factor Arg-gingipain, while the corresponding Lys-gingipain was hardly influenced. RA1 inhibited also hemagglutination.

In conclusion, the proanthocyanidin-enriched extract RA1 and its main active constituent procyanidin B2-di-gallate protect cells from P. gingivalis infection by inhibiting bacterial adhesion to the host cell. RA1 and procyanidin B2-di-gallate appear to be promising candidates for future cytoprotective preparations for oral mouth care products.

Schlüsselwörter

Rumex acetosa L. - Sauerampferkraut - Adhäsion - Zytoprotektion - Parodontitis - Porphyromonas gingivalis - Proanthocyanidine

Keywords

Rumex acetosa L - Adhesion - Cytoprotection - Periodontitis - Porphyromonas gingivalis - Proanthocyanidins

¹ Universität Münster, Institut für Pharmazeutische Biologie und Phytochemie

² Heinrich-Heine-Universität, Sektion für Parodontologie, Düsseldorf

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