Effect and mechanism of Prunella vulgaris L. extract on alleviating lipopolysaccharide-induced acute mastitis in protecting the blood-milk barrier and reducing inflammation.

ETHNOPHARMACOLOGICAL RELEVANCE: According to ancient literature, Prunella vulgaris L. (P vulgaris) alleviates mastitis and has been used in China for many years; however, there are no relevant reports that confirm this or the mechanism of its efficacy. AIM OF THE STUDY: To explore the anti-acute mastitis effect and potential mechanism of P vulgaris extract. MATERIALS AND METHODS: First, the active ingredients and targets of P vulgaris against mastitis were predicted using network pharmacology. Next, the relevant active ingredients were enriched using macroporous resins and verified using UV and UPLC-Q-TOF-MS/MS. Lastly, a mouse model of acute mastitis was established by injecting lipopolysaccharides into the mammary gland and administering P vulgaris extract by oral gavage. The pathological changes in mammary tissue were observed by HE staining. Serum and tissue inflammatory factors were measured by ELISA method. MPO activity in mammary tissue was measured using colorimetry and MPO expression was detected by immunohistochemistry. The expression of tight junction proteins (ZO-1, claudin-3, and occludin) in mammary tissue was detected by immunofluorescence and Western blot. iNOS and COX-2 in mammary tissue were detected by Western blot. MAPK pathway and NF-κB pathway related proteins were also detected by Western blot. RESULTS: Network pharmacology predicted that phenolic acids and flavonoids in P vulgaris had anti-mastitis effects. The contents of total flavonoids and total phenolic acids in P vulgaris extract were 64.5% and 29.4%, respectively. UPLC-Q-TOF-MS/MS confirmed that P vulgaris extract contained phenolic acids and flavonoids. The results of animal experiments showed that P vulgaris extract reduced lipopolysaccharide-induced inflammatory edema, inflammatory cell infiltration, and interstitial congestion of mammary tissue. It also reduced the levels of serum and tissue inflammatory factors TNF-α, IL-6, and IL-1ß, and inhibited the activation of MPO. Furthermore, it downregulated the expression of MAPK and NF-κB pathway-related proteins. The expressions of ZO-1, occludin, and claudin-3 in mammary gland tissues were upregulated. CONCLUSIONS: P vulgaris extract can maintain the integrity of mammary connective tissue and reduce its inflammatory response to prevent acute mastitis. Its mechanism probably involves regulating NF-κB and MAPK pathways.

Integrative transcriptomic, proteomic and metabolomic analysis reveals the dynamic regulation of secondary metabolism upon development of Prunella vulgaris L.

Prunella vulgaris L. (P. vulgaris, Labiatae) is a perennial medicinal and edible plant widely used in China, Korea, Japan and Europe. The reddish brown spica of P. vulgaris (Prunellae Spica), which is collected in summer, has been commonly used in traditional medicine and food industry, while it is also used with whole grass in Europe and Taiwan. To clarify the regulatory pathways and mechanism of quality formation in P. vulgaris, targeted metabolomic, transcriptomic, and proteomic analyses of Prunellae Spica samples from five consecutive developmental stages were carried out. The results showed that terpenoids were mainly synthesized in the maturity stage of Prunellae Spica, with the key enzymes and coding genes in downstream pathways being mainly expressed during ripening, while related enzymes in the upstream pathway showed the opposite pattern. Flavonoids mainly accumulated before ripening, with highly expressed pathway enzymes and coding genes. The accumulation of phenylpropanoids was relatively active throughout the development process. Rosmarinic acid (RA) and its synthetic intermediate products mainly accumulated via more active pathway enzymes and coding genes before ripening. The regulatory factors and metabolites related to RA synthesis were mainly enriched in phenylpropanoid biosynthesis, plant hormone signal transduction, plant pathogen interaction, oxidative phosphorylation, and endoplasmic reticulum protein processing pathways.