Journal: Scientific reports

Article Title: Targeting Deficiencies in the TLR5 Mediated Vaginal Response to Treat Female Recurrent Urinary Tract Infection.

doi: 10.1038/s41598-017-10445-4

Figure Lengend Snippet: Figure 1. VK2 E6/E7 and RT4 DEFB4 and BD2 responses to UPEC and flagellin challenges. End point PCR panels of host defence peptide gene expression in VK2 E6/E7 and RT4 cells following 24 hr challenge with PBS (control), flagellin (250 ng/ml) or UPEC (105) (A). Full length gels are shown in Supplementary File 1 (S1A–D). DEFB4 transcript expression in VK2 E6/E7 and RT4 cells following challenges with flagellin (250 ng/ ml) and heat-killed flagellated (NCTC10418) or non-flagellated (NU14) E. coli (105) (N = 3, n = 9) (B). BD2 concentrations measured in VK2 E6/E7 and RT4 cell media following 24 hour challenges with flagellin (250 ng/ ml) and heat-killed CFT073 (105) (N = 3, n = 6) (C). BD2 peptide concentrations of primary ureteric urothelial cells challenged with E.coli flagellin (250 ng/ml) for 24 hours (N = 3, n = 6) (D). DEFB4 transcript expression, presented as relative expression, of primary vaginal epithelial cells challenged with E.coli flagellin (50 ng/ ml) for 24 hours (N = 6) (E). E.coli swarming motility on motility agar media; BD2 peptide concentrations measured in VK2 E6/E7 and RT4 cell media following 24 hour challenges with flagellin (250 ng/ml) and heat- killed bacteria (105) (N = 3, n = 6) (F). DEFB4 transcript expression of VK2 E6/E7 cells transfected with either

Article Snippet: VK2 E6/E7 cells (ATCC CRL-2616)54 were maintained without antibiotics in keratinocyte serum-free medium (GIBCO, Paisley, UK) containing 0.4 mM calcium with 0.1 ng/ml human recombinant Epidermal Growth Factor (EGF) and 0.05 mg/ml bovine pituitary extract supplements.

Techniques: Gene Expression, Control, Expressing, Bacteria, Transfection

Journal: Scientific reports

Article Title: Targeting Deficiencies in the TLR5 Mediated Vaginal Response to Treat Female Recurrent Urinary Tract Infection.

doi: 10.1038/s41598-017-10445-4

Figure Lengend Snippet: Figure 5. Effects of estrogen treatment on BD2 responses in vitro and rUTI patients. VK2 E6/E7 cell DEFB4 mRNA expression following estrogen (4 nM) seven day pretreatment and 24 h challenge with either cyclodextrin (15 nM), flagellin (50 ng/ml), or flagellin (50 ng/ml) and estrogen (4 nM) N = 3 (A). BD2 peptide concentrations measured in the VK2 E6/E7 cell media following estrogen (4 nM) seven day pretreatment and either 24 h or 48 h challenge with either cyclodextrin (15 nM), estrogen (4 nM), flagellin (50 ng/ml) or flagellin and estrogen (N = 3) (B). Vaginal douche BD2 peptide concentrations of rUTI patients before and after 6–8 weeks of treatment with antibiotic prophylaxis and/or advice (N = 12) (C), or 6 to 8 weeks treatment with vaginal estrogen (Vagifem, 10 mcg, twice weekly) (N = 9) (D). Staining of vaginal tissue from estrogen treated ( + E) and control subjects for immunoreactive BD2 (E). Vaginal douche BD2 peptide concentrations of rUTI TLR5 SNP patients before and after treatment (6 to 8 weeks) with vaginal estrogen (N = 3) or antibiotic

Article Snippet: VK2 E6/E7 cells (ATCC CRL-2616)54 were maintained without antibiotics in keratinocyte serum-free medium (GIBCO, Paisley, UK) containing 0.4 mM calcium with 0.1 ng/ml human recombinant Epidermal Growth Factor (EGF) and 0.05 mg/ml bovine pituitary extract supplements.

Techniques: In Vitro, Expressing, Staining, Control

Journal: Scientific Reports

Article Title: Impact of vaginal douching products on vaginal Lactobacillus , Escherichia coli and epithelial immune responses

doi: 10.1038/s41598-021-02426-5

Figure Lengend Snippet: Impact of douching products on vaginal epithelial cells. An immortalized vaginal epithelial cell line was cultured with a 25% solution of douching product diluted in the cell culture medium for 1 h, followed by replacement with fresh media for 23 h. Cell death ( a ) was measured using an LDH assay and IL6 ( b ), IL8 ( c ), IL1β ( d ) and IL1-RA ( e ) were measured in supernatant by ELISA. Results are presented as mean ± SD from at least two separate experiments.

Article Snippet: Vaginal epithelial cells (VK2 E6/E7, ATCC) were seeded at 40,000 cells/well in a 96-well plate, in Keratinocyte-Serum Free medium (KSF) (with BPE and EGF, but no antibiotics), and allowed to grow overnight to confluence.

Techniques: Cell Culture, Lactate Dehydrogenase Assay, Enzyme-linked Immunosorbent Assay

Journal: Scientific Reports

Article Title: Impact of vaginal douching products on vaginal Lactobacillus , Escherichia coli and epithelial immune responses

doi: 10.1038/s41598-021-02426-5

Figure Lengend Snippet: Impact of bacteria on vaginal epithelial cells. An immortalized vaginal epithelial cell line was cultured with a suspension of Lactobacillus iners, L. crispatus, L. gasseri or L. jensenii in cell culture medium (OD 0.1) for 24 h. Cell death ( a ) was measured using an LDH assay and IL6 ( b ), IL8 ( c ), IL1β ( d ) and IL1-RA ( e ) were measured in supernatant by ELISA. Results are presented as mean ± SD from at least two separate experiments.

Article Snippet: Vaginal epithelial cells (VK2 E6/E7, ATCC) were seeded at 40,000 cells/well in a 96-well plate, in Keratinocyte-Serum Free medium (KSF) (with BPE and EGF, but no antibiotics), and allowed to grow overnight to confluence.

Techniques: Bacteria, Cell Culture, Suspension, Lactate Dehydrogenase Assay, Enzyme-linked Immunosorbent Assay

Journal: Scientific Reports

Article Title: Impact of vaginal douching products on vaginal Lactobacillus , Escherichia coli and epithelial immune responses

doi: 10.1038/s41598-021-02426-5

Figure Lengend Snippet: Impact of bacteria on vaginal epithelial cell death with douching exposure. An immortalized vaginal epithelial cell line was cultured with a 25% solution of douching product diluted in the cell culture medium for 1 h, followed by replacement with a suspension of L. crispatus or L. iners (OD 0.1) in cell culture media for 23 h. Cell death was measured with an LDH assay and values compared by ANOVA. Results are presented as mean ± SD from at least two separate experiments.

Article Snippet: Vaginal epithelial cells (VK2 E6/E7, ATCC) were seeded at 40,000 cells/well in a 96-well plate, in Keratinocyte-Serum Free medium (KSF) (with BPE and EGF, but no antibiotics), and allowed to grow overnight to confluence.

Techniques: Bacteria, Cell Culture, Suspension, Lactate Dehydrogenase Assay

Journal: Scientific Reports

Article Title: Impact of vaginal douching products on vaginal Lactobacillus , Escherichia coli and epithelial immune responses

doi: 10.1038/s41598-021-02426-5

Figure Lengend Snippet: Impact of bacteria on vaginal epithelial cell death immune response to douching exposure. An immortalized vaginal epithelial cell line was cultured with a 25% solution of douching product diluted in the cell culture medium for 1 h, followed by replacement with a suspension of L. iners, L. crispatus, L. gasseri or L. jensenii (OD 0.1) in cell culture media for 23 h. Supernatant was used to measure IL6 ( a ), IL8 ( b ), IL1β ( c ) and IL1-RA ( d ) with ELISA, and values compared by ANOVA. Results are presented as mean ± SD from at least two separate experiments.

Article Snippet: Vaginal epithelial cells (VK2 E6/E7, ATCC) were seeded at 40,000 cells/well in a 96-well plate, in Keratinocyte-Serum Free medium (KSF) (with BPE and EGF, but no antibiotics), and allowed to grow overnight to confluence.

Techniques: Bacteria, Cell Culture, Suspension, Enzyme-linked Immunosorbent Assay