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Ethnopharmacological Relevance: Extracts from Cranberry fruits (Vaccinium macrocarpon) are traditionally used against urinary tract infections, mainly due to antiadhesive activity against uropathogenic E. coli (UPEC), but the exact mode of action and compounds, responsible for the activity, are unknown., Aim of the Study: i. To investigate if cranberry extract acts only by a single component or must be assessed as a multi-active-compound preparation; ii to screen isolated cranberry-related natural products under in vitro conditions to pinpoint natural products with antiadhesive effects against UPEC, followed by in silico calculations (QSAR) to predict potential antiadhesive compounds; iii. investigations by using urine samples from cranberry treated volunteers for evaluation on the bacterial transcriptome and the mannose-binding side of FimH, iv. to investigate if besides Tamm Horsfall Protein induction in the kidney, the extract acts also directly against UPEC., Material and Methods: Antiadhesive activity of 105 compounds was determined by flow cytometric adhesion assay (UPEC UTI89 on T24 bladder cells). Urine samples from 16 volunteers treated with cranberry extract (p.o., 7 days, 900 mg/day) were used for ex vivo testing concerning influence on the bacterial transcriptome (Illumina RNA-seq) and interaction with the mannose binding domain of type-1 fimbriae., Results: i. The antiadhesive effect of cranberry extract cannot be attributed to a single compound or to a single fraction. ii. Unglycosylated flavones and flavonols with bulky substitution of the B ring contribute to the antiadhesive activity. 3'-8″-biflavones and flavolignans (not related to cranberry fruits) were identified as potent antiadhesive compounds against UPEC. iii. QSAR yielded a model with good statistical performance and sufficient internal and external predictive ability. iv. Urine samples from male cranberry-treated volunteers indicated significant interaction with the mannose binding domain of type-1 fimbriae, which correlated with the amount of Tamm-Horsfall Protein in the test samples. v Cranberry extract did not influence the UPEC transcriptome; gene expression of bacterial adhesins (P-, S-fimbrae, curli) was not influenced by the urine samples, while a slight, but non-significant upregulation of type 1 fimbriae was observed., Conclusions: B-ring substituted flavones and flavonols from cranberry contribute to the antiadhesive activity against UPEC by inhibition of the FimH-mediated interaction with the host cell bladder epithelium., Competing Interests: Declaration of competing interest We wish to confirm that there are no known conflicts of interest associated with this publication and there has been no significant financial support for this work that could have influenced its outcome., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Ethnopharmacological Relevance: Amaranthus caudatus is traditionally used to treat infections. Based on its traditional usage, we investigated the effect of A. caudatus on the bladder epithelial cells in the protection of E. coli infection., Materials and Methods: The direct antimicrobial effects of A. caudatus on uropathogenic bacteria were investigated using minimum inhibitory concentration (MIC) assay. Bladder epithelial cell lines T24 and 5637 and uropathogenic E. coli strain #12 were used to investigate the effect of A. caudatus. Bacterial adhesion and invasion into bladder cells treated with A. caudatus was analyzed. Expression of uroplakin-1a (UPK1A), β1 integrin (ITGB1), caveolin-1 (CAV1) and the antimicrobial peptides human β defensin-2 (DEFB4A) and LL-37 (CAMP) was evaluated using RT-PCR., Results: No direct antibacterial effect on E. coli or any of the tested uropathogenic strains was observed by A. caudatus. However, we demonstrated reduced mRNA expression of uroplakin-1a and caveolin-1, but not β1 integrin after treatment of uroepithelial cells, mirrored by the decreased adhesion and invasion of E. coli. A. caudatus treatment did not induce increased gene expression of the antimicrobial peptides, LL-37 and human β-defensin-2., Conclusions: Our results showed that A. caudatus has a protective role on bladder epithelial cells against uropathogenic E. coli infection by decreasing the bacterial adhesion and invasion, thereby preventing infection., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Ethnopharmacological Relevance: Clinopodium bolivianum is a South American plant with anti-inflammatory and anti-infective activities. The increasing antibiotic resistance urges for alternative therapy. Based on its use in traditional medicine, we investigated the effect of C. bolivianum on the ability to defend bladder epithelial cells from E. coli infection., Materials and Methods: The extract was analyzed by LC-MS. Bladder epithelial cell lines T24 and 5637 and uropathogenic E. coli No. 12, its isogenic mutant WE16 csgBA bscA::Cm and CFT073 were used to investigate the effect of C. bolivianum on uroepithelial infection. Bacterial adherence and invasion to cells treated with C. bolivianum were analyzed. Expression of uroplakin 1a, β1 integrin, caveolin-1, IL-8 and antimicrobial peptides in response to C. bolivianum treatment was assessed using RT-PCR. Protein expression was confirmed by Western blot analysis or ELISA. The antimicrobial effects of C. bolivianum on bacteria and fungus were investigated using minimum inhibitory concentration. Furthermore, the formation of biofilm was investigated with crystal violet assay., Results: C. bolivianum extract consisted of more than 70 different types of phytochemicals including sugars and phenolic compounds. The extract decreased the uroplakin 1a expression and E. coli adhesion and invasion of uroepithelial cells while up-regulated caveolin-1. In uninfected C. bolivianum treated cells, IL-8 was lower than in non-treated cells. In infected cells, however, no difference was observed between treated and non-treated cells. Further, C. bolivianum treatment reduced uropathogenic E. coli (UPEC) biofilms but did not inhibit bacterial growth., Conclusions: Our results show that C. bolivianum has a protective role on bladder epithelial cells against UPEC infection by decreasing the bacterial adhesion, invasion and biofilm formation., (Copyright © 2017 Elsevier Ireland Ltd. All rights reserved.)

Ethnopharmacological Relevance: Traditional usage suggests Citrus reticulata Blanco seeds have beneficial effects against infection. The purpose of this study was to investigate the effect of Citrus reticulata on the uroepithelium and to determine the mechanisms responsible for protection against urinary tract infection (UTI)., Materials and Methods: Human bladder cell lines T24 and 5637 were employed in a cell culture infection model to determine the effects of Citrus reticulata treatment on Escherichia coli adherence and invasion of the uroepithelium. β1 integrin and caveolin-1 mRNA expression was assessed using RT real-time PCR. β1 integrin protein expression was confirmed by Western Blot. The effect of Citrus reticulata on bacteria was investigated using antibacterial sensitivity, yeast agglutination and biofilm assays., Results: Citrus reticulata treatment decreased β1 integrin expression and reduced bacterial invasion while adhesion of uroepithelial cells was not affected. Caveolin-1 expression was not influenced either and Citrus reticulata did neither exhibit any direct antimicrobial effect nor interfered with type 1 fimbriae binding., Conclusions: Our results show that Citrus reticulata has a protective effect on the uroepithelium as seen by reduced bacterial invasion of uroepithelial cells. These properties suggest that seeds from Citrus reticulata may have therapeutic potential in preventing UTI., (© 2013 Elsevier Ireland Ltd. All rights reserved.)

Ethnopharmacological Relevance: Labisia pumila var. alata (LPva) is a traditional medicinal herb used by Malaysian women to treat many ailments of the genitourinary tract. Its phytoestrogenic properties suggest potential to prevent recurrent urinary tract infection (UTI) in women post menopause. The aim of this study was therefore to investigate the mechanisms of action of LPva in an in vitro model of UTI., Materials and Methods: Bladder epithelial cell lines T24 and 5637 and uropathogenic Escherichia coli (UPEC) strain CFT073 were used to model uroepithelial infection. The ability of LPva to induce programmed cell death was tested using the Annexin-V-FLUOS and TUNEL assays. Expression of caveolin-1, β1 integrin and antimicrobial peptides HBD-2 and LL-37 in response to LPva treatment and/or infection, was assessed using RT real-time PCR. Effects on protein expression were confirmed by Western blot analysis. Sensitivity and yeast agglutination assays were employed to determine if LPva had antimicrobial activities and/or interacted with type 1 fimbriae, respectively. Finally, bacterial adherence and invasion to cells treated with LPva was examined., Results: LPva induced uroepithelial apoptosis which was coupled with upregulated expression of caveolin-1 and downregulation of β1 integrin. LPva did not exhibit direct antimicrobial properties and did not influence antimicrobial peptide levels in cells. Additionally, LPva did not interact with type 1 fimbriae and did not affect adherence in comparison to non-treated control cells. However, LPva significantly reduced the number of intracellular UPEC in bladder epithelial cells., Conclusions: Our findings suggest that LPva has beneficial applications against UPEC infection due to its ability to induce programmed cell death and reduce bacterial invasion of the uroepithelium., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Ethnopharmacological Relevance: Uropathogenic Escherichia coli is the major cause for urinary tract infections (UTI). Due to emerging antimicrobial resistances treatment of UTI is becoming increasingly difficult. Therefore, alternative treatment strategies are required. We sought to investigate the molecular mechanisms of a traditionally used decoction from Vietnamese dandelion (Lactuca indica L.) mediating local protection of the bladder epithelium., Materials and Methods: To study the influence of herbal preparations on the interaction between bladder epithelial cells and uropthogenic Escherichia coli, a cell culture model of UTI was applied. In addition, the direct effect of the herbal extract on Escherichia coli was analyzed., Results: Although no direct antibacterial activity was determined, Lactuca indica decreased bacterial colonization of bladder epithelial cells remarkably. Reduced bacterial adherence was accompanied by lowered activation of focal adhesion kinase (FAK) in Lactuca indica exposed cells. Treatment with a chemical FAK inhibitor supported this mechanism of action., Conclusions: These findings indicate that, in addition to its diuretic action, Lactuca indica exhibits secondary effects directly on epithelial cells which may protect against Escherichia coli infection. These properties might be useful for the development of alternative strategies in the treatment of UTI., (Copyright © 2011. Published by Elsevier Ireland Ltd.)