Gram-positive pathogenic bacteria induce a common early response in human monocytes

<p>Abstract</p> <p>Background</p> <p>We infected freshly isolated human peripheral monocytes with live bacteria of three clinically important gram-positive bacterial species, <it>Staphylococcus aureus</it>, <it>Streptococcus pneumoniae </it>and <it>Listeria monocytogenes </it>and studied the ensuing early transcriptional response using expression microarrays. Thus the observed response was unbiased by signals originating from other helper and effector cells of the host and was not limited to induction by solitary bacterial constituents.</p> <p>Results</p> <p>Activation of monocytes was demonstrated by the upregulation of chemokine rather than interleukin genes except for the prominent expression of interleukin 23, marking it as the early lead cytokine. This activation was accompanied by cytoskeleton rearrangement signals and a general anti-oxidative stress and anti-apoptotic reaction. Remarkably, the expression profiles also provide evidence that monocytes participate in the regulation of angiogenesis and endothelial function in response to these pathogens.</p> <p>Conclusion</p> <p>Regardless of the invasion properties and survival mechanisms of the pathogens used, we found that the early response comprised of a consistent and common response. The common response was hallmarked by the upregulation of interleukin 23, a rather unexpected finding regarding <it>Listeria </it>infection, as this cytokine has been linked primarily to the control of extracellular bacterial dissemination.</p

Necrosis is the dominant cell death pathway in uropathogenic Escherichia coli elicited epididymo-orchitis and is responsible for damage of rat testis

Male infertility is a frequent medical condition, compromising approximately one in twenty men, with infections of the reproductive tract constituting a major etiological factor. Bacterial epididymo-orchitis results in acute inflammation most often caused by ascending canalicular infections from the urethra via the continuous male excurrent ductal system. Uropathogenic Escherichia coli (UPEC) represent a relevant pathogen in urogenital tract infections. To explore how bacteria can cause damage and cell loss and thus impair fertility, an in vivo epididymo-orchitis model was employed in rats by injecting UPEC strain CFT073 into the vas deference in close proximity to the epididymis. Seven days post infection bacteria were found predominantly in the testicular interstitial space. UPEC infection resulted in severe impairment of spermatogenesis by germ cell loss, damage of testicular somatic cells, a decrease in sperm numbers and a significant increase in TUNEL (+) cells. Activation of caspase-8 (extrinsic apoptotic pathway), caspase-3/&#8722;6 (intrinsic apoptotic pathway), caspase-1 (pyroptosis pathway) and the presence of 180 bp DNA fragments, all of which serve as indicators of the classical apoptotic pathway, were not observed in infected testis. Notably, electron microscopical examination revealed degenerative features of Sertoli cells (SC) in UPEC infected testis. Furthermore, the passive release of high mobility group protein B1 (HMGB1), as an indication of necrosis, was observed in vivo in infected testis. Thus, necrosis appears to be the dominant cell death pathway in UPEC infected testis. Substantial necrotic changes seen in Sertoli cells will contribute to impaired spermatogenesis by loss of function in supporting the dependent germ cells

Protective Immunity to Listeria Monocytogenes Infection Mediated by Recombinant Listeria innocua Harboring the VGC Locus

In this study we propose a novel bacterial vaccine strategy where non-pathogenic bacteria are complemented with traits desirable for the induction of protective immunity. To illustrate the proof of principle of this novel vaccination strategy, we use the model organism of intracellular immunity Listeria. We introduced a, low copy number BAC-plasmid harbouring the virulence gene cluster (vgc) of L. monocytogenes (Lm) into the non-pathogenic L. innocua (L.inn) strain and examined for its ability to induce protective cellular immunity. The resulting strain (L.inn::vgc) was attenuated for virulence in vivo and showed a strongly reduced host detrimental inflammatory response compared to Lm. Like Lm, L.inn::vgc induced the production of Type I Interferon's and protection was mediated by Listeria-specific CD8+ T cells. Rational vaccine design whereby avirulent strains are equipped with the capabilities to induce protection but lack detrimental inflammatory effects offer great promise towards future studies using non-pathogenic bacteria as vectors for vaccination

Uropathogenic E. coli Induce Different Immune Response in Testicular and Peritoneal Macrophages: Implications for Testicular Immune Privilege

Infertility affects one in seven couples and ascending bacterial infections of the male genitourinary tract by Escherichia coli are an important cause of male factor infertility. Thus understanding mechanisms by which immunocompetent cells such as testicular macrophages (TM) respond to infection and how bacterial pathogens manipulate defense pathways is of importance. Whole genome expression profiling of TM and peritoneal macrophages (PM) infected with uropathogenic E. coli (UPEC) revealed major differences in regulated genes. However, a multitude of genes implicated in calcium signaling pathways was a common feature which indicated a role of calcium-dependent nuclear factor of activated T cells (NFAT) signaling. UPEC-dependent NFAT activation was confirmed in both cultured TM and in TM in an in vivo UPEC infectious rat orchitis model. Elevated expression of NFATC2-regulated anti-inflammatory cytokines was found in TM (IL-4, IL-13) and PM (IL-3, IL-4, IL-13). NFATC2 is activated by rapid influx of calcium, an activity delineated to the pore forming toxin alpha-hemolysin by bacterial mutant analysis. Alpha-hemolysin suppressed IL-6 and TNF-α cytokine release from PM and caused differential activation of MAP kinase and AP-1 signaling pathways in TM and PM leading to reciprocal expression of key pro-inflammatory cytokines in PM (IL-1α, IL-1β, IL-6 downregulated) and TM (IL-1β, IL-6 upregulated). In addition, unlike PM, LPS-treated TM were refractory to NFκB activation shown by the absence of degradation of IκBα and lack of pro-inflammatory cytokine secretion (IL-6, TNF-α). Taken together, these results suggest a mechanism to the conundrum by which TM initiate immune responses to bacteria, while maintaining testicular immune privilege with its ability to tolerate neo-autoantigens expressed on developing spermatogenic cells

The TIR Domain Containing Locus of Enterococcus faecalis Is Predominant among Urinary Tract Infection Isolates and Downregulates Host Inflammatory Response

Based on Toll/interleukin-1 receptor (TIR) domain structure homology, we detected a previously uncharacterized gene encoding for a TIR domain containing protein (Tcp) in the genome of Enterococcus faecalis. We assigned this gene the name tcpF (as in Tcp of E. faecalis). Screening of E. faecalis samples revealed that tcpF is more common in isolates from urinary tract infections (UTIs) than in human faecal flora. tcpF alleles showed moderate single nucleotide polymorphism (SNP) among UTI isolates. Infection of mouse RAW264.7 macrophages with a tcpF knock-out mutant led to elevated cytokine response compared to the isogenic wild type E. faecalis strain. In silico analysis predicted significant tertiary structure homology to the TIR domain of human TLR1 (TLR1-TIR). When transiently expressed in cultured eukaryotic cells, TcpF caused suppression of TLR2-dependent NF-κB activation suggesting for TcpF a role as a factor in E. faecalis that benefits colonization by modulating the host’s immune responses

Uropathogenic Escherichia coli block MyD88-dependent and activate MyD88-independent signaling pathways in rat testicular cells.

International audienceUropathogenic Escherichia coli (UPEC) is the most common etiological cause of urogenital tract infections and represents a considerable cause of immunological male infertility. We examined TLR 1-11 expression profiles in testicular cells and the functional response to infection with UPEC. All testicular cell types expressed mRNAs for at least two TLRs and, in particular, synthesis of TLR4 was induced in testicular macrophages (TM), Sertoli cells (SC), peritubular cells (PTC), and peritoneal macrophages (PM) after UPEC exposure. Even though MyD88-dependent pathways were activated as exemplified by phosphorylation of mitogen-activated protein kinases in TM, SC, PTC, and PM and by the degradation of IkappaBalpha and the nuclear translocation of NF-kappaB in PTC and PM, treatment with UPEC did not result in secretion of the proinflammatory cytokines IL-1alpha, IL-6, and TNF-alpha in any of the investigated cells. Moreover, stimulated production of these cytokines by nonpathogenic commensal E. coli or LPS in PM was completely abolished after coincubation with UPEC. Instead, in SC, PTC, TM, and PM, UPEC exposure resulted in activation of MyD88-independent signaling as documented by nuclear transfer of IFN-related factor-3 and elevated expression of type I IFNs alpha and beta, IFN-gamma-inducible protein 10, MCP-1, and RANTES. We conclude that in this in vitro model UPEC can actively suppress MyD88-dependent signaling at different levels to prevent proinflammatory cytokine secretion by testicular cells. Thus, testicular innate immune defense is shifted to an antiviral-like MyD88-independent response

Measurement of proinflammatory cytokine levels in serum.

<p>Sera was obtained from mice on days 1, 2, 3, and 4 post-infection after inoculation with 10³ cfu <i>Lm</i>, 10⁷ cfu <i>L.inn</i>, or 10⁷ cfu <i>L.inn::vgc</i>. Levels of IL-1ß, IL-6, IL-12(p70), and TNF-alpha were quantified using a multiplex cytokine assay kit. *P<0.05 (EGD-e vs. <i>L.inn</i> and <i>L.inn::vgc</i> strains).</p

Examination of spleens and DTH response after infection with <i>Lm</i> and the recombinant <i>L.inn::vgc</i> strain.

<p><b>A.</b> Morphological examination of spleens from mice inoculated i.v. with the wild type <i>Lm</i> and the recombinant <i>L.inn::vgc</i> strain. Spleens of mice infected i.v. as mentioned in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035503#pone-0035503-g001" target="_blank">Fig. 1</a> were isolated on day 3 after infection. Shown is a spleen from mice infected with the wild type <i>Lm</i>, the wild type <i>L.inn</i> and its recombinant mutant strain <i>L.inn::vgc</i>. Infiltration of monocytic cells and granulomatous lesions are only detectable in the spleens isolated from mice infected with the wild type <i>Lm</i>. <b>B.</b> Spleen sections were stained with HE and examined. Granulomas with massive leukocyte aggregates can only be detected in spleens of mice infected with <i>Lm</i>. <b>C.</b> DTH response to listerial antigen 9 days after primary infection. Mice were infected with 10³ CFU of <i>Lm</i>, 10⁷ CFU of <i>L.inn</i>, or 10⁷ CFU of <i>L.inn::vgc</i> strain. 9 days after infection, DTH was triggered through injection of soluble somatic listerial antigen. Twenty-four hours later, the specific skin response was determined. The mean value ± S.E. of five animals of a representative experiment is shown.*P<0.05 (EGD-e vs. <i>L.inn::vgc</i> strain).</p