Hypothetical protein Cpn0423 triggers NOD2 activation and contributes to Chlamydia pneumoniae-mediated inflammation.

BACKGROUND: Chlamydia pneumoniae (C. pneumoniae) is pathogenic to humans, by causing pulmonary inflammation or bronchitis in both adolescents and young adults. However, the molecular signals linking C. pneumoniae components to inflammation remain elusive. This study was to investigate the effect of Chlamydia-specific Cpn0423 of C. pneumoniae on C. pneumoniae-mediated inflammation. RESULTS: Cpn0423 was detected outside of C. pneumoniae inclusions, which induced production of several cytokines including macrophage inflammatory protein-2 (MIP-2) and interleukins (ILs). Production of the Cpn0423-induced cytokines was markedly reduced in cells pretreated with NOD2-siRNA, but not with negative control oligonucleotides. Mice treated with Cpn0423 through intranasal administration exhibited pulmonary inflammation as evidenced by infiltration of inflammatory cells, increased inflammatory scores in the lung histology, recruitment of neutrophils and increased cytokines levels in the BALF. CONCLUSION: Cpn0423 could be sensed by NOD2, which was identified as an essential element in a pathway contributing to the development of C. pneumoniae -mediated inflammation.

Mast cells play an important role in chlamydia pneumoniae lung infection by facilitating immune cell recruitment into the airway.

Mast cells are known as central players in allergy and anaphylaxis, and they play a pivotal role in host defense against certain pathogens. Chlamydia pneumoniae is an important human pathogen, but it is unclear what role mast cells play during C. pneumoniae infection. We infected C57BL/6 (wild-type [WT]) and mast cell-deficient mice (Kit(W-sh/W-sh) [Wsh]) with C. pneumoniae. Wsh mice showed improved survival compared with WT mice, with fewer cells in Wsh bronchoalveolar lavage fluid (BALF), despite similar levels of cytokines and chemokines. We also found a more rapid clearance of bacteria from the lungs of Wsh mice compared with WT mice. Cromolyn, a mast cell stabilizer, reduced BALF cells and bacterial burden similar to the levels seen in Wsh mice; conversely, Compound 48/80, a mast cell degranulator, increased the number of BALF cells and bacterial burden. Histology showed that WT lungs had diffuse inflammation, whereas Wsh mice had patchy accumulations of neutrophils and perivascular accumulations of lymphocytes. Infected Wsh mice had reduced amounts of matrix metalloprotease-9 in BALF and were resistant to epithelial integral membrane protein degradation, suggesting that barrier integrity remains intact in Wsh mice. Mast cell reconstitution in Wsh mice led to enhanced bacterial growth and normal epithelial integral membrane protein degradation, highlighting the specific role of mast cells in this model. These data suggest that mast cells play a detrimental role during C. pneumoniae infection by facilitating immune cell infiltration into the airspace and providing a more favorable replicative environment for C. pneumoniae.

The sst1 resistance locus regulates evasion of type I interferon signaling by Chlamydia pneumoniae as a disease tolerance mechanism.

The sst1, "supersusceptibility to tuberculosis," locus has previously been shown to be a genetic determinant of host resistance to infection with the intracellular pathogen, Mycobacterium tuberculosis. Chlamydia pneumoniae is an obligate intracellular bacterium associated with community acquired pneumonia, and chronic infection with C. pneumoniae has been linked to asthma and atherosclerosis. C. pneumoniae is a highly adapted pathogen that can productively infect macrophages and inhibit host cell apoptosis. Here we examined the role of sst1 in regulating the host response to infection with C. pneumoniae. Although mice carrying the sst1 susceptible (sst1(S) ) locus were not impaired in their ability to clear the acute infection, they were dramatically less tolerant of the induced immune response, displaying higher clinical scores, more severe lung inflammation, exaggerated macrophage and neutrophil influx, and the development of fibrosis compared to wild type mice. This correlated with increased activated caspase-3 in the lungs of infected sst1(S) mice. Infection of sst1(S) macrophages with C. pneumoniae resulted in a shift in the secreted cytokine profile towards enhanced production of interferon-ß and interleukin-10, and induced apoptotic cell death, which was dependent on secretion of interferon-ß. Intriguingly macrophages from the sst1(S) mice failed to support normal chlamydial growth, resulting in arrested development and failure of the organism to complete its infectious cycle. We conclude that the sst1 locus regulates a shared macrophage-mediated innate defense mechanism against diverse intracellular bacterial pathogens. Its susceptibility allele leads to upregulation of type I interferon pathway, which, in the context of C. pneumoniae, results in decreased tolerance, but not resistance, to the infection. Further dissection of the relationship between type I interferons and host tolerance during infection with intracellular pathogens may provide identification of biomarkers and novel therapeutic targets.

Dual-color bioluminescent assay using infected HepG2 cells sheds new light on Chlamydia pneumoniae and human cytomegalovirus effects on human cholesterol 7α-hydroxylase (CYP7A1) transcription.

Chlamydia pneumoniae and human cytomegalovirus (HCMV) are intracellular pathogens able to infect hepatocytes, causing an increase in serum triglycerides and cholesterol levels due to the production of inflammatory cytokines. We investigated whether these pathogens could interfere with cholesterol metabolism by affecting activity of hepatic cholesterol 7α-hydroxylase (CYP7A1) promoter. CYP7A1 is the rate-limiting enzyme responsible for conversion of cholesterol to bile acids, which represents the main route of cholesterol catabolism. A straightforward dual-reporter bioluminescent assay was developed to simultaneously monitor CYP7A1 transcriptional regulation and cell viability in infected human hepatoblastoma HepG2 cells. C. pneumoniae and HCMV infection significantly decreased CYP7A1 promoter activity in a dose-dependent manner, with maximal inhibitions of 33±10% and 32±4%, respectively, at a multiplicity of infection of 1. To support in vitro experiments, serum cholesterol, high-density lipoprotein (HDL) cholesterol, triglycerides and glucose levels were also measured in Balb/c mice infected with C. pneumoniae. Serum cholesterol and triglycerides also increased in infected mice compared with controls. Although further investigation is required, this work presents the first experimental evidence that C. pneumoniae and HCMV inhibit CYP7A1 gene transcription in the cultured human hepatoblastoma cell line.

Chlamydia pneumoniae induces a pro-inflammatory phenotype in murine vascular smooth muscle cells independently of elevating reactive oxygen species.

  NADPH oxidases (Nox) are reactive oxygen species (ROS)-generating enzymes that play important physiological roles in host defence and redox signalling. However, Nox activity is upregulated in the vascular wall during atherosclerosis and contributes to plaque formation by promoting oxidative stress and inflammation.   The bacterium Chlamydia pneumoniae has been detected in vascular smooth muscle cells (VSMC) of human atheroma. We hypothesized that C. pneumoniae infection of VSMC causes Nox activation, which initially limits infection but ultimately causes oxidative stress, activation of pro-inflammatory pathways and an atherogenic phenotype.   Chlamydia pneumoniae infection of mouse cultured VSMC significantly increased ROS production by twofold but did not upregulate mRNA expression of Nox1 or Nox4. Chlamydia pneumoniae did increase Nox2 mRNA levels significantly by threefold, but this did not translate to elevated Nox2 protein expression.   The Nox inhibitor gp91ds-tat had no effect on C. pneumoniae-induced ROS production. In contrast, apocynin significantly reduced ROS levels by 75% in C. pneumoniae-infected VSMC, an effect most likely attributable to its direct anti-oxidant action.   Although apocynin had no effect on C. pneumoniae-induced expression of inflammatory markers, bacteria recovered from apocynin-treated VSMC displayed a higher degree of infectivity in HEp-2 cells.   In conclusion, C. pneumoniae infection increases ROS production in VSMC independently of Nox activity. Although elevated ROS production appears to serve a protective role by limiting the spread of infection, we speculate that this response will be detrimental over the long term by causing oxidative stress and a smouldering inflammatory response by maintaining C. pneumoniae persistence within the cell.

Deficiency of XIAP leads to sensitization for Chlamydophila pneumoniae pulmonary infection and dysregulation of innate immune response in mice.

Obligate intracellular Chlamydophila pneumoniae induce apoptosis resistance in host cells to escape eradication by immune effector cells. Apoptosis resistance depends on the increased expression and stabilization of cellular inhibitor of apoptosis proteins (cIAPs) and X-linked inhibitor of apoptosis protein (XIAP). Here we investigated the role of XIAP in experimental pulmonary infection of mice with C. pneumoniae. XIAP knock out (KO) mice were sensitized for C. pneumoniae infection compared with wild type mice. XIAP was involved in lipopolysaccharide (LPS)-induced production of nitric oxide (NO) and endotoxin shock. Hyper-secretion of tumor necrosis factor-alpha and lower NO in LPS-treated KO mouse macrophages revealed its regulatory role in inflammatory responses. Unexpectedly, activating stimuli like LPS, tumor necrosis factor-alpha, or interferon-gamma very efficiently induced apoptotic cell death in KO macrophages but not in wild type macrophages. Cell survival transcription factor nuclear factor kappaB (NF-kappaB) p65 levels were reduced in lungs and pulmonary macrophages of infected KO mice. Furthermore, a reduced CD8 T cell population and their increased sensitivity for concanavalin A and chlamydial HSP60 stimulation revealed a defect in CD8 T cells in XIAP KO mice. These data demonstrated a role of XIAP for the integrity of both innate and cellular immune responses during C. pneumoniae infection.

Chlamydophila pneumoniae downregulates MHC-class II expression by two cell type-specific mechanisms.

Chlamydophila pneumoniae was shown to prevent IFN gamma-inducible upregulation of MHC-class II molecules by secreting chlamydial protease-like activity factor (CPAF) into the cytosol of those host cells which support the complete bacterial replication cycle. CPAF acts by degrading upstream stimulatory factor 1 (USF-1). However, in cells like bone marrow-derived macrophages (BMM), which restrict chlamydial replication, we show that CPAF expression is barely detectable and the expression of USF-1 is induced upon infection with C. pneumoniae. Nevertheless, the infection still reduced base line and prevented IFN gamma-inducible MHC-class II expression. Similar results were obtained with heat-inactivated C. pneumoniae. In contrast, reduction of MHC-class II molecules was not observed in MyD88-deficient BMM. Reduction of IFN gamma-inducible MHC-class II expression by C. pneumoniae in BMM was mediated in part by the MAP-kinase p38. Infection of murine embryonic fibroblasts (MEF) with C. pneumoniae, which allow chlamydial replication, induced the expression of CPAF and decreased USF-1 and MHC-class II expression. Treatment of these cells with heat-inactivated C. pneumoniae reduced USF-1 and MHC-class II expression to a much lower extent. In summary, C. pneumoniae downregulates MHC-class II expression by two cell type-specific mechanisms which are either CPAF-independent and MyD88-dependent like in BMM or CPAF-dependent like in MEFs.

Analysis of Chlamydia pneumoniae infection in mononuclear cells by reverse transcription-PCR targeted to chlamydial gene transcripts.

Chlamydia pneumoniae (C. pneumoniae) is an important etiological agent of respiratory infections including pneumonia. C. pneumoniae DNA can be detected in peripheral blood mononuclear cells indicating that monocytes can assist the spread of infection to other anatomical sites. Persistent infection established at these sites could promote inflammation and enhance pathology. Thus, the mononuclear cells are in a strategic position in the development of persistent infection. To investigate the intracellular replication and fate of C. pneumoniae in mononuclear cells, we have established an in vitro model in the human Mono Mac 6 cell line. In the present study, we analyzed the transcription of 11 C. pneumoniae genes in Mono Mac 6 cells during infection by real-time RT-PCR. Our results suggest that the transcriptional profile of the studied genes in monocytes is different from that seen in epithelial cells. Furthermore, our study shows that genes related to secretion are transcribed, and secreted bacterial proteins are also translated during infection of monocytes, creating novel opportunities for the management of chlamydial infection of monocytes.

[The phylogenetic position of Chlamydia strains isolated from monkeys and humans with Chlamydial pathology in the family Chiamydiaceae. Genotypic and phenotypic properties of this pathogen].

Based on the results of the comparative analysis concerning relatedness and evolutional difference of the 16S - 23S nucleotide sequences of the middle ribosomal cluster and 23S rRNA I domain, and based on identification of phylogenetic position for Chlamydophila pneumoniae and Chlamydia trichomatis strains released from monkeys, relatedness of the above stated isolates with similar strains released from humans and with strains having nucleotide sequences presented in the GenBank electronic database has been detected for the first time ever. Position of these isolates in the Chlamydiaceae family phylogenetic tree has been identified. The evolutional position of the investigated original Chlamydia and Chlamydophila strains close to analogous strains from the GenBank electronic database has been demonstrated. Differences in the 16S - 23S nucleotide sequence of the middle ribosomal cluster and 23S rRNA I domain of plasmid and non-plasmid Chlamydia trachomatis strains released from humans and monkeys relative to different genotype groups (group B- B, Ba, D, Da, E, L1, L2, L2a; intermediate group - F, G, Ga) have been revealed for the first time ever. Abnormality in incA chromosomal gene expression resulting in Chlamydia life and development cycle disorder and decrease of Chlamydia virulence can be related to probable changes in the nucleotide sequence of the gene under consideration.

[Development of pulmonary chlamydia infection in inbred mice lines differentiated by genetically determinated sensitivity to tuberculosis infection].

Mice of I/St strain develop severe lung inflammation and die shortly following infection with virulent mycobacteria. The susceptibility does not depend on the Nramp1 gene, as I/St mice carry its resistant allele, but is controlled by little interacting QTL mapped to chromosomes 3, 9, 17. To find out whether the tuberculosis-susceptible I/St mice are susceptible to other intracellular bacteria taxonomically distant pathogen of Chlamydia pneumoniae was studied. Comparison of I/St and TB-resistant A/Sn mice (both Nramp1r) demonstrated that the former were more susceptible to chlamydia, displaying a significantly shortened survival time following challenge (I/St, 9.2 +/- 1.2 days; A/Sn, 22.0 +/- 0 days (p < 0.001)). To estimate the degree of chlamydial multiplication in the lungs, we suggested a quantitative real-time polymerase chain reaction (PCR)-based method which allows enumeration of the parasite's genome equivalents in infected tissue from 1 to 16 days after challenge. The interstrain difference of chlamydia burden in lungs was observed only after 24 hours after infection. Multiplication of chlamydia in the lungs was controlled efficiently after day 4 of infection. The numbers of genome equivalents dropped slightly by day 8 both in I/St and A/Sn mice. Lung pathology develops more rapidly in I/St compared to A/Sn mice following infection with chlamydia despite their similar ability to control bacterial multiplication. Lung tissue of susceptible I/St mice was markedly infiltrated with macrophages (p < 0.01), which differed significantly from the lungs of resistant A/Sn mice. In agreement with higher macrophage content in the lungs, significantly more macrophage-derived proinflammatory cytokines TNF-? and IL-6 were detected in lung tissue homogenates obtained from I/St mice (p < 0.05). Because the prominent difference in survival time did not correlate with permanent difference in bacterial multiplication, we suggested that both infections trigger fatal pathological processes whose dynamics depends strongly upon the host genetics.

The 1B vaccine strain of Chlamydia abortus produces placental pathology indistinguishable from a wild type infection.

Chlamydia abortus is one of the most commonly diagnosed causes of infectious abortion in small ruminants worldwide. Control of the disease (Enzootic Abortion of Ewes or EAE) is achieved using the commercial live, attenuated C. abortus 1B vaccine strain, which can be distinguished from virulent wild-type (wt) strains by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. Published studies applying this typing method and whole-genome sequence analyses to cases of EAE in vaccinated and non-vaccinated animals have provided strong evidence that the 1B strain is not attenuated and can infect the placenta causing disease in some ewes. Therefore, the objective of this study was to characterise the lesions found in the placentas of ewes vaccinated with the 1B strain and to compare these to those resulting from a wt infection. A C. abortus-free flock of multiparous adult ewes was vaccinated twice, over three breeding seasons, each before mating, with the commercial C. abortus 1B vaccine strain (Cevac® Chlamydia, Ceva Animal Health Ltd.). In the second lambing season following vaccination, placentas (n = 117) were collected at parturition and analysed by C. abortus-specific real-time quantitative PCR (qPCR). Two placentas, from a single ewe, which gave birth to live twin lambs, were found to be positive by qPCR and viable organisms were recovered and identified as vaccine type (vt) by PCR-RFLP, with no evidence of any wt strain being present. All cotyledons from the vt-infected placentas were analysed by histopathology and immunohistochemistry and compared to those from wt-infected placentas. Both vt-infected placentas showed lesions typical of those found in a wt infection in terms of their severity, distribution, and associated intensity of antigen labelling. These results conclusively demonstrate that the 1B strain can infect the placenta, producing typical EAE placental lesions that are indistinguishable from those found in wt infected animals.

Relevance of the genetic polymorphism of NOD1 in Chlamydia pneumoniae seropositive stroke patients.

BACKGROUND AND PURPOSE: Chronic infections with certain pathogens, such as Chlamydia pneumoniae, and genetic parameters that influence inflammatory reactions have been suggested to contribute to ischaemic stroke. NOD1 is a potent cytosolic receptor for C. pneumoniae. The aim of this study was to investigate the genetic polymorphism of NOD1 from the aspect of the development of stroke. MATERIALS AND METHODS: A total of 280 patients with ischaemic stroke were enrolled in the study; 150 healthy blood donors served as controls. The G796A (E266K) NOD1 polymorphism was determined by restriction fragment length polymorphism. Chlamydia pneumoniae seropositivity was tested by ELISA. RESULTS: There was a significant difference in NOD1 G796A genotype distribution between the controls and the stroke patients with C. pneumoniae seropositivity. The AA homozygote and GA heterozygote mutant variants were detected in 16% (25 of 152) and in 50% (77 of 152) of the C. pneumoniae-positive stroke patients, as compared with 8% (6 of 84), and 28% (24 of 84), respectively, in the C. pneumoniae-positive healthy controls. (OR = 2.559; 95% CI = 1.105-6.517, P = 0.04 and OR = 2.567; 95% CI = 1.451-4.540 P < 0.001, respectively). The stroke patients with the large vessel pathology exhibited the highest frequency of the mutant allele A (51%). In contrast, amongst the C. pneumoniae-negative subjects, no difference in genotype frequency was observed between the stroke patients and the controls. CONCLUSION: Polymorphism in NOD1 G796A alone did not prove to be a risk factor for stroke in general, but in association with C. pneumoniae infection it appeared to be accompanied by an increased risk of the development of stroke.

[Effector proteins of Clamidia].

The review summarizes the recent published data on molecular mechanisms of Chlamidiae - host cell interaction, first of all on chlamydial effector proteins. Such proteins as well as III transport system proteins that transfer many effector proteins into host cytoplasm are attractive targets for drug therapy of chlamydial infections. The majority of the data concerns two species, Chlamydia trachomatis and Chlamydophila pneumoniae. C. trachomatis protein TARP, which is presynthesized in elementary bodies, plays an essential role in the initial stages of the infection. Patogen proteins participating in the next stage, that is the intracellular inclusion traffic to the centrosome, are CT229 of C. trachomatis and Cpn0585 of C. pneumoniae, which interact with cellular Rab GTPases. In C. trachomatis, IncA protein plays a key role in chlamydial inclusions fusion, CT847 modulates life cycle of the host cell, LDA3 is essential in acquisition of nutrients. CPAF protease and inclusion membrane proteins IncG and CADD participate in suppression of apoptosis of infected cells. The proteases CPAF and CT441, as well as deubiquitinating ChlaDub1 protein, contribute to avoiding the immune response.

Temporal delay of peak T-cell immunity determines Chlamydia pneumoniae pulmonary disease in mice.

Severe chlamydial disease typically occurs after previous infections and results from a hypersensitivity response that is also required for chlamydial elimination. Here, we quantitatively dissected the immune and disease responses to repeated Chlamydia pneumoniae lung infection by multivariate modeling with four dichotomous effects: mouse strain (A/J or C57BL/6), dietary protein content (14% protein and 0.3% L-cysteine-0.9% L-arginine, or 24% protein and 0.5% L-cysteine-2.0% L-arginine), dietary antioxidant content (90 IU alpha-tocopherol/kg body weight versus 450 IU alpha-tocopherol/kg and 0.1% g L-ascorbate), and time course (3 or 10 days postinfection). Following intranasal C. pneumoniae challenge, C57BL/6 mice on a low-protein/low-antioxidant diet, but not C57BL/6 mice on other diets or A/J mice, exhibited profoundly suppressed early lung inflammatory and pan-T-cell (CD3delta(+)) and helper T-cell (CD45) responses on day 3 but later strongly exacerbated disease on day 10. Contrast analyses characterized severe C. pneumoniae disease as being a delayed-type hypersensitivity (DTH) response with increased lung macrophage and Th1 cell marker transcripts, increased Th1:Th2 ratios, and Th1 cytokine-driven inflammation. Results from functional analyses by DTH, enzyme-linked immunospot, and immunohistofluorescence assays were consistent with the results obtained by transcript analysis. Thus, chlamydial disease after secondary infection is a temporal dysregulation of the T-cell response characterized by a profoundly delayed T-helper cell response that results in a failure to eliminate the pathogen and provokes later pathological Th1 inflammation. This delayed T-cell response is under host genetic control and nutritional influence. The mechanism that temporally and quantitatively regulates the host T-cell population is the critical determinant in chlamydial pathogenesis.

The association of C (-260)-->T polymorphism in CD14 promoter and Chlamydia pneumoniae infection in ischemic stroke patients.

The C (-260) --> T polymorphism has been reported to regulate CD14 gene expression. It has also been implicated in atherosclerotic diseases, and in addition, it could be a genetic factor responsible for interindividual differences in the susceptibility to Chlamydia pneumoniae infection. This case-control study is aimed at evaluating the association between CD14 promoter polymorphisms, frequency of persistent C pneumoniae infection, and anti-chlamydial heat shock protein 60 (cHsp60) induction in stroke patients. Persistent C pneumoniae infection was observed in 43.3% of control subjects and 53.3% of patients (P = .005). The odds ratio of persistent infection was 3.25 for the CC genotype in stroke patients (P = .018). However, the serologic positive responses to cHsp60 in people with the TT genotype were significantly lower in stroke patients (9.3% vs 38.6%; P < .001). Our findings suggest that persistent C pneumoniae infection is independently associated with stroke in CC genotypes and the lowering of cHsp60 antibody levels in TT genotypes.

Chlamydia pneumoniae infection and hyperlipidaemia-induced expression of P50 and c-Fos in the heart of C57BL6J mice.

OBJECTIVE: The objective of this study was to investigate infection by Chlamydia pneumoniae (C. pneumoniae) and the combined effects of an atherogenic diet on the expression of AP-1 or the subunit of AP-1 (c-Fos) and NF-kappaB or the subunit of NF-kappaB (P50) in myocardial cells of C57BL/6J mice. METHODS AND RESULTS: Thirty-six 8-week-old female C57BL/6J mice were divided into three groups: A, B and C (twelve mice in each group). Mice in group A were fed an atherogenic diet (consisting of 15% fat and 2.5% cholesterol). Those in group B and C (as blank control) were fed a regular diet. The mice in group B were infected with C. pneumoniae. Fourteen weeks later, the expression of P50 (subunit of NF-kappaB) and c-Fos (subunit of AP-1) in the heart was determined by indirect immunofluorescence in the myocardial cells. Binding activity of NF-kappaB and AP-1 in the heart was determined by electrophoretic mobility shift assay (EMSA). The expression of NF-kappaB and AP-1 in the myocardial cells was upregulated in group A and B, compared with group C (P < 0.05). There was no significant difference between group A and B. CONCLUSION: The inflammatory process was already initiated in the myocardial cells in C57BL/6J mice at the early stage of C. pneumoniae infection and hyperlipidaemia. The expression of NF-kappaB and AP-1 was upregulated in mice infected by C. pneumoniae or fed an atherogenic diet. C. pneumoniae or hyperlipidaemia may involve the pathogenesis and progression of ischaemic cardiomyopathy.

Cholesterol uptake in the mouse aorta increases during Chlamydia pneumoniae infection.

Chlamydia pneumoniae has been suggested as a stimulator of the atherosclerotic process. Mice fed a normal diet were infected intranasally with C. pneumoniae and given one intraperitoneal injection of 14C-cholesterol tracer per day for 12 days. Bacteria were demonstrated in the aorta in the early phase of infection and in lungs and liver throughout the study period of 20 days. 14C-cholesterol was not affected in the heart but increased in the blood, liver and aorta on day 4 when the infection was clinically most severe. Furthermore, on day 20 14C-cholesterol tended to be increased in the aorta. Accordingly, copper- and zinc levels and expressions of the infection biomarkers Cxcl2 and Ifng increased in the liver on day 4 with a tendency of increased of copper, zinc and Ifng on day 20. In mice where bacteria could be cultivated from the lungs, expressions of cholesterol transporters Abca1 and Idol were both increased in the liver on day 4. The increased levels of 14C-cholesterol in blood and aorta together with increased Abca1 and Idol in the liver during C. pneumoniae infection in mice fed a normal diet suggest that this pathogen may have a role in the initiation of the atherosclerotic process.

Chlamydophila (Chlamydia) pneumoniae infection of human astrocytes and microglia in culture displays an active, rather than a persistent, phenotype.

BACKGROUND: The intracellular pathogen Chlamydia pneumoniae can cause persistent infections during which its morphologic, molecular, and pathogenic characteristics differ importantly from those of active infection. This bacterium was identified within astrocytes and microglia in the brain of late-onset Alzheimer disease patients. We investigated whether infection of these two host cell types displays an active or persistent growth phenotype. METHODS: The human astrocytoma and microglioma cell lines U-87 MG and CHME-5 (respectively) and the human epithelial cell line HEp-2 were infected by the standard method with C pneumoniae strain AR-39. Cultures were harvested at 24, 48, and 72 hours postinfection and subjected to analysis of inclusion morphology. DNA and RNA were prepared from portions of each infected culture sample and analyzed for relative chromosome accumulation and presence or absence of several specific bacterial mRNAs. RESULTS: Astrocytes and microglial cells infected in vitro with C pneumoniae displayed inclusions that were indistinguishable from those characteristic of active infection of the standard HEp-2 host cell line. Real time polymerase chain reaction (PCR) showed that the relative accumulation of chlamydial chromosome over time during infection of these two cell lines also was virtually identical to that in actively infected HEp-2 cells. Reverse transcriptase PCR (RT-PCR) analyses showed that mRNA from ftsK, pyk, and other chlamydial genes whose expression is abrogated during persistent infection were easily identifiable in infected CHME-5 and U-87 MG cells. CONCLUSIONS: In cultured human astrocytes and microglia, C pneumoniae displays an active, not a persistent, growth phenotype. This indicates normal passage through the developmental cycle with its probable concomitant destruction by lysis of some portion of host cells at the termination of that cycle.

Chlamydia pneumoniae infection does not induce or modify atherosclerosis in mice.

BACKGROUND: Seroepidemiological studies have linked Chlamydia pneumoniae (CP) to coronary heart disease, and recent experimental studies suggest that it may accelerate or even induce atherosclerosis. We therefore evaluated the effect of CP infection on atherosclerosis in atherosclerosis-prone apolipoprotein E-knockout (apoE-KO) and wild-type C57BL/6J mice. METHODS AND RESULTS: Six- to 8-week-old female mice were infected intranasally with live CP and then fed a standard chow diet for 22 weeks. A subgroup of mice was reinfected 18 weeks after primary infection. Polymerase chain reaction analysis of lung tissue confirmed successful infection with CP, and ELISA assays demonstrated development of a humoral immune response. Despite this, no statistically significant differences in aortic atherosclerotic lesions were found between CP-infected and control apoE-KO mice. Furthermore, CP infection did not induce atherosclerosis in C57BL/6J mice. CONCLUSIONS: CP does not induce atherosclerosis in wild-type mice and does not accelerate atherosclerosis in chow-fed apoE-KO mice. Further studies will be necessary to clarify the explanation for the seroepidemiological association between CP and coronary heart disease in humans.

Invariant Natural Killer T Cells Promote T Cell Immunity by Modulating the Function of Lung Dendritic Cells during Chlamydia pneumoniae Infection.

In this study, we examined the effect of invariant natural killer T (iNKT) cells on the function of lung dendritic cells (LDCs) in eliciting protective immunity against Chlamydia pneumoniae (Cpn) lung infection. We employed a combination of approaches including the use of iNKT cell-deficient, Jα18-knockout (KO) mice and LDC adoptive transfer. We found that iNKT cells significantly altered the number, phenotype and cytokine profile of LDCs following infection. Furthermore, coculture of T cells with LDCs from Cpn-infected wild-type (WT) and KO mice induced type-1 and type-2 responses, respectively. More importantly, upon adoptive transfer, LDCs from Cpn-infected WT mice (WT-LDCs) conferred protective immunity, whereas LDCs from KO mice (KO-LDCs) increased the severity of disease after challenge infection. Further cytokine analyses of the lung tissues and lung-draining lymph node cells showed that KO-LDC-recipient mice exhibited a type-2 cytokine production pattern, while WT-LDC recipients exhibited a type-1 cytokine profile. Taken together, our results provide in vivo evidence that iNKT cells play a critical role in modulating LDC function to generate protective T-cell immunity, particularly in a clinically relevant intracellular bacterial infection.