Modulation of Human Beta-Defensin 2 Expression by Pathogenic Neisseria meningitidis and Commensal Lactobacilli

Antimicrobial peptides (AMPs) play an important role in the defense against pathogens by targeting and killing invading microbes. Some pathogenic bacteria have been shown to negatively regulate AMP expression, while several commensals may induce AMP expression. The expression of certain AMPs, such as human beta-defensin 2 (hBD2), can be induced via nuclear factor NF-κB, which in turn is negatively controlled by tumor necrosis factor alpha-induced protein 3 (TNFAIP3, or A20). In this work, we examined the expression of hBD1 and hBD2 during coincubation of pharyngeal epithelial cells with pathogenic Neisseria meningitidis and commensal lactobacilli. The Lactobacillus strains induced hBD2 expression in human pharyngeal cells, while the pathogen N. meningitidis did not. In coincubation experiments, meningococci were able to dampen the AMP expression induced by lactobacilli. We found that N. meningitidis induced the NF-κB inhibitor A20. Further, RNA silencing of A20 resulted in increased hBD2 expression after meningococcal infection. Since it is known that induction of A20 reduces NF-κB activity and thus hBD2 levels, meningococcal-mediated A20 induction could be a way for the pathogen to dampen AMP expression. Finally, treatment of N. meningitidis and lactobacilli with synthetic hBD2 reduced N. meningitidis viability more efficiently than L. reuteri, explaining why maintaining low AMP levels is important for the survival of the pathogen.

Histone deacetylase inhibition enhances antimicrobial peptide but not inflammatory cytokine expression upon bacterial challenge

Antimicrobial peptides (AMP) are defense effectors of the innate immunity playing a crucial role in the intestinal homeostasis with commensals and protection against pathogens. Herein we aimed to investigate AMP gene regulation by deciphering specific characteristics allowing their enhanced expression among innate immune genes, particularly those encoding proinflammatory mediators. Our emphasis was on epigenetic regulation of the gene encoding the AMP β-defensin 2 (HBD2), taken as a model of possibly specific induction, upon challenge with a commensal bacterium, compared with the proinflammatory cytokine IL-8. Using an in vitro model of colonic epithelial cells challenged with Escherichia coli K12, we showed that inhibition of histone deacetylases (HDAC) by trichostatin A dramatically enhanced induction of HBD2 expression, without affecting expression of IL-8. This mechanism was supported by an increased phosphorylation of histone H3 on serine S10, preferentially at the HBD2 promoter. This process occurred through activation of the IκB kinase complex, which also led to activation of NF-κB. Moreover, we demonstrated that NF-κB was modified by acetylation upon HDAC inhibition, partly by the histone acetyltransferase p300, and that both NF-κB and p300 supported enhanced induction of HBD2 expression. Furthermore, we identified additional genes belonging to antimicrobial defense and epithelial restitution pathways that showed a similar pattern of epigenetic control. Finally, we confirmed our finding in human colonic primary cells using an ex vivo organoid model. This work opens the way to use epigenetic pharmacology to achieve induction of epithelial antimicrobial defenses, while limiting the deleterious risk of an inflammatory response.

Human β-Defensins Suppress Human Immunodeficiency Virus Infection: Potential Role in Mucosal Protection

ABSTRACT β-Defensins are small (3 to 5 kDa in size) secreted antimicrobial and antiviral proteins that are components of innate immunity. β-Defensins are secreted by epithelial cells, and they are expressed at high levels in several mucosae, including the mouth, where the concentration of these proteins can reach 100 μg/ml. Because of these properties, we wondered whether they could be part of the defenses that lower oral transmission of human immunodeficiency virus (HIV) compared to other mucosal sites. Our data show that select β-defensins, especially human β-defensin 2 (hBD2) and hBD3, inhibit R5 and X4 HIV infection in a dose-dependent manner at doses that are compatible with or below those measured in the oral cavity. We observed that β-defensin treatment inhibited accumulation of early products of reverse transcription, as detected by PCR. We could not, however, detect any reproducible inhibition of env-mediated fusion, and we did not observe any modulation of HIV coreceptors following treatment with hBD1 and hBD2, in both resting and phytohemagglutinin-activated cells. Our data instead suggest that, besides a direct inactivation of HIV virions, hBD2 inhibits HIV replication in the intracellular environment. Therefore, we speculate that β-defensins mediate a novel antiretroviral mechanism that contributes to prevention of oral HIV transmission in the oral cavity. Immunohistochemical data on hBD2 expression in oral mucosal tissue shows that hBD2 is constitutively expressed, forming a barrier layer across the epithelium in healthy subjects, while in HIV-positive subjects levels of hBD2 expression are dramatically diminished. This may predispose HIV-positive subjects to increased incidence of oral complications associated with HIV infection.