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• W1979478895 abstract "ObjectiveToll-like receptors (TLR) mitigate innate immune responses to infection. Gram positive (GP), Mycoplasma and Ureaplasma spp. primarily engage TLR2, while TLR4 recognizes Gram negative (GN, i.e. E. Coli) bacterial ligands. We propose that the cross-talk between TLRs generates differential IAI responses. Our aim was to evaluate the intensity of IAI elicited by single vs. mixed spp. in vivo and in vitro.Study DesignWe studied amniotic fluid (AF) from 134 women (GA: 26±3 wks) with PTB. AF cultures tested positive for: 1) single GN (n=30); 2) single GP (n=31); 3) Mycoplasma or Ureaplasma (n=34); 4) mixed TLR2 (n=16) or 5) mixed TLR2 & TLR4-engaging microbes. (n=23). In vivo, IAI level was assessed by IL6 ELISA. An amniochorion explant system (term CS, n=4) was used to test the effect of prototype ligands in vitro. We generated dose response curves at 6 and 18h after challenge with ligands for TLR4 [LPS], TLR2 [Lipoteichoic acid (LTA), Pam3Cys (PAM), Mycoplasma FSL] alone or combined [LPS+PAM, LPS+FSL]. TNFα, IL6, IL8 were measured in explant media and normalized to total protein. Tissue viability was confirmed by LDH release.ResultsIn vivo, IAI with single GN had highest AF IL6 independent of GA (p<0.001). There was no difference in IL6 among single TLR2-engaging spp. In contrast, mixed GP induced higher IL6 vs. single TLR2-engaging spp. (p=0.016). IAI with mixed GN & GP had higher IL6 vs. single and mixed GP (p<0.05). In vitro, LPS induced higher responses in all cytokines at all times and doses while FSL responses were lowest (p<0.001). Of all TLR2 ligands, LTA had the highest response (p<0.05). LPS+FSL produced a 3-fold increase in all cytokines over FSL alone (p=0.03). LPS+PAM induced a 20-fold increase over PAM alone (p=0.02) and a transient 4-fold increase over LPS alone at 6h (p<0.05).ConclusionsBoth in vivo and in vitro, single TLR4-engaging bacteria generate highest inflammatory responses. Mixed infections lead to different IAI levels vs. single species alone. ObjectiveToll-like receptors (TLR) mitigate innate immune responses to infection. Gram positive (GP), Mycoplasma and Ureaplasma spp. primarily engage TLR2, while TLR4 recognizes Gram negative (GN, i.e. E. Coli) bacterial ligands. We propose that the cross-talk between TLRs generates differential IAI responses. Our aim was to evaluate the intensity of IAI elicited by single vs. mixed spp. in vivo and in vitro. Toll-like receptors (TLR) mitigate innate immune responses to infection. Gram positive (GP), Mycoplasma and Ureaplasma spp. primarily engage TLR2, while TLR4 recognizes Gram negative (GN, i.e. E. Coli) bacterial ligands. We propose that the cross-talk between TLRs generates differential IAI responses. Our aim was to evaluate the intensity of IAI elicited by single vs. mixed spp. in vivo and in vitro. Study DesignWe studied amniotic fluid (AF) from 134 women (GA: 26±3 wks) with PTB. AF cultures tested positive for: 1) single GN (n=30); 2) single GP (n=31); 3) Mycoplasma or Ureaplasma (n=34); 4) mixed TLR2 (n=16) or 5) mixed TLR2 & TLR4-engaging microbes. (n=23). In vivo, IAI level was assessed by IL6 ELISA. An amniochorion explant system (term CS, n=4) was used to test the effect of prototype ligands in vitro. We generated dose response curves at 6 and 18h after challenge with ligands for TLR4 [LPS], TLR2 [Lipoteichoic acid (LTA), Pam3Cys (PAM), Mycoplasma FSL] alone or combined [LPS+PAM, LPS+FSL]. TNFα, IL6, IL8 were measured in explant media and normalized to total protein. Tissue viability was confirmed by LDH release. We studied amniotic fluid (AF) from 134 women (GA: 26±3 wks) with PTB. AF cultures tested positive for: 1) single GN (n=30); 2) single GP (n=31); 3) Mycoplasma or Ureaplasma (n=34); 4) mixed TLR2 (n=16) or 5) mixed TLR2 & TLR4-engaging microbes. (n=23). In vivo, IAI level was assessed by IL6 ELISA. An amniochorion explant system (term CS, n=4) was used to test the effect of prototype ligands in vitro. We generated dose response curves at 6 and 18h after challenge with ligands for TLR4 [LPS], TLR2 [Lipoteichoic acid (LTA), Pam3Cys (PAM), Mycoplasma FSL] alone or combined [LPS+PAM, LPS+FSL]. TNFα, IL6, IL8 were measured in explant media and normalized to total protein. Tissue viability was confirmed by LDH release. ResultsIn vivo, IAI with single GN had highest AF IL6 independent of GA (p<0.001). There was no difference in IL6 among single TLR2-engaging spp. In contrast, mixed GP induced higher IL6 vs. single TLR2-engaging spp. (p=0.016). IAI with mixed GN & GP had higher IL6 vs. single and mixed GP (p<0.05). In vitro, LPS induced higher responses in all cytokines at all times and doses while FSL responses were lowest (p<0.001). Of all TLR2 ligands, LTA had the highest response (p<0.05). LPS+FSL produced a 3-fold increase in all cytokines over FSL alone (p=0.03). LPS+PAM induced a 20-fold increase over PAM alone (p=0.02) and a transient 4-fold increase over LPS alone at 6h (p<0.05). In vivo, IAI with single GN had highest AF IL6 independent of GA (p<0.001). There was no difference in IL6 among single TLR2-engaging spp. In contrast, mixed GP induced higher IL6 vs. single TLR2-engaging spp. (p=0.016). IAI with mixed GN & GP had higher IL6 vs. single and mixed GP (p<0.05). In vitro, LPS induced higher responses in all cytokines at all times and doses while FSL responses were lowest (p<0.001). Of all TLR2 ligands, LTA had the highest response (p<0.05). LPS+FSL produced a 3-fold increase in all cytokines over FSL alone (p=0.03). LPS+PAM induced a 20-fold increase over PAM alone (p=0.02) and a transient 4-fold increase over LPS alone at 6h (p<0.05). ConclusionsBoth in vivo and in vitro, single TLR4-engaging bacteria generate highest inflammatory responses. Mixed infections lead to different IAI levels vs. single species alone. Both in vivo and in vitro, single TLR4-engaging bacteria generate highest inflammatory responses. Mixed infections lead to different IAI levels vs. single species alone." @default.
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• W1979478895 title "76: Intra-amniotic inflammation (IAI) in response to infection with single species versus mixed bacterial flora in women with preterm birth (PTB)" @default.
• W1979478895 doi "https://doi.org/10.1016/j.ajog.2010.10.090" @default.
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