Association of carotid plaque Lp-PLA(2) with macrophages and Chlamydia pneumoniae infection among patients at risk for stroke.

BACKGROUND: We previously showed that the burden of Chlamydia pneumoniae in carotid plaques was significantly associated with plaque interleukin (IL)-6, and serum IL-6 and C-reactive protein (CRP), suggesting that infected plaques contribute to systemic inflammatory markers in patients with stroke risk. Since lipoprotein-associated phospholipase A2 (Lp-PLA(2)) mediates inflammation in atherosclerosis, we hypothesized that serum Lp-PLA(2) mass and activity levels and plaque Lp-PLA(2) may be influenced by plaque C. pneumoniae infection. METHODOLOGY/PRINCIPAL FINDINGS: Forty-two patients underwent elective carotid endarterectomy. Tissue obtained at surgery was stained by immunohistochemistry for Lp-PLA(2) grade, macrophages, IL-6, C. pneumoniae and CD4+ and CD8+ cells. Serum Lp-PLA(2) activity and mass were measured using the colorimetric activity method (CAM) and ELISA, respectively. Serum homocysteine levels were measured by HPLC. Eleven (26.2%) patients were symptomatic with transient ischemic attacks. There was no correlation between patient risk factors (smoking, coronary artery disease, elevated cholesterol, diabetes, obesity, hypertension and family history of genetic disorders) for atherosclerosis and serum levels or plaque grade for Lp-PLA(2). Plaque Lp-PLA(2) correlated with serum homocysteine levels (p = 0.013), plaque macrophages (p<0.01), and plaque C. pneumoniae (p<0.001), which predominantly infected macrophages, co-localizing with Lp-PLA(2). CONCLUSIONS: The significant association of plaque Lp-PLA(2) with plaque macrophages and C. pneumoniae suggests an interactive role in accelerating inflammation in atherosclerosis. A possible mechanism for C. pneumoniae in the atherogenic process may involve infection of macrophages that induce Lp-PLA(2) production leading to upregulation of inflammatory mediators in plaque tissue. Additional in vitro and in vivo research will be needed to advance our understanding of specific C. pneumoniae and Lp-PLA(2) interactions in atherosclerosis.

Identification of novel single nucleotide polymorphisms in inflammatory genes as risk factors associated with trachomatous trichiasis.

BACKGROUND: Trachoma is the leading preventable cause of global blindness. A balanced Th1/Th2/Th3 immune response is critical for resolving Chlamydia trachomatis infection, the primary cause of trachoma. Despite control programs that include mass antibiotic treatment, reinfection and recurrence of trachoma are common after treatment cessation. Furthermore, a subset of infected individuals develop inflammation and are at greater risk for developing the severe sequela of trachoma known as trachomatous trichiasis (TT). While there are a number of environmental and behavioral risk factors for trachoma, genetic factors that influence inflammation and TT risk remain ill defined. METHODOLOGY/FINDINGS: We identified single nucleotide polymorphisms (SNP) in 36 candidate inflammatory genes and interactions among these SNPs that likely play a role in the overall risk for TT. We conducted a case control study of 538 individuals of Tharu ethnicity residing in an endemic region of Nepal. Trachoma was graded according to World Health Organization guidelines. A linear array was used to genotype 51 biallelic SNPs in the 36 genes. Analyses were performed using logic regression modeling, which controls for multiple comparisons. We present, to our knowledge, the first significant association of TNFA (-308GA), LTA (252A), VCAM1 (-1594TC), and IL9 (T113M) polymorphisms, synergistic SNPs and risk of TT. TT risk decreased 5 times [odds ratio = 0.2 (95% confidence interval 0.11.-0.33), p = 0.001] with the combination of TNFA (-308A), LTA (252A), VCAM1 (-1594C), SCYA 11 (23T) minor allele, and the combination of TNFA (-308A), IL9 (113M), IL1B (5'UTR-T), and VCAM1 (-1594C). However, TT risk increased 13.5 times [odds ratio = 13.5 (95% confidence interval 3.3-22), p = 0.001] with the combination of TNFA (-308G), VDR (intron G), IL4R (50V), and ICAM1 (56M) minor allele. CONCLUSIONS: Evaluating genetic risk factors for trachoma will advance our understanding of disease pathogenesis, and should be considered in the context of designing global control programs.

Role of secreted conjunctival mucosal cytokine and chemokine proteins in different stages of trachomatous disease.

BACKGROUND: Chlamydia trachomatis is responsible for trachoma, the primary cause of preventable blindness worldwide. Plans to eradicate trachoma using the World Health Organization's SAFE program (Surgery, Antibiotics, Facial Cleanliness and Environment Improvement) have resulted in recurrence of infection and disease following cessation of treatment in many endemic countries, suggesting the need for a vaccine to control infection and trachomatous disease. Vaccine development requires, in part, knowledge of the mucosal host immune responses in both healthy and trachomatous conjuctivae-an area of research that remains insufficiently studied. METHODOLOGY/PRINCIPAL FINDINGS: We characterized 25 secreted cytokines and chemokines from the conjunctival mucosa of individuals residing in a trachoma endemic region of Nepal using Luminex X100 multiplexing technology. Immunomodulating effects of concurrent C. trachomatis infection were also examined. We found that proinflammatory cytokines IL-1beta (r = 0.259, P = 0.001) and TNFalpha (r = 0.168, P<0.05) were significantly associated with trachomatous disease and concurrent C. trachomatis infection compared with age and sex matched controls from the same region who did not have trachoma. In support of these findings, anti-inflammatory cytokine IL-1 receptor antagonist (IL-1Ra) was negatively associated with chronic scarring trachoma (r = -0.249, P = 0.001). Additional cytokines (Th1, IL-12p40 [r = -0.212, P<0.01], and Th2, IL-4 and IL-13 [r = -0.165 and -0.189, respectively, P<0.05 for both]) were negatively associated with chronic scarring trachoma, suggesting a protective role. Conversely, a pathogenic role for the Th3/Tr1 cytokine IL-10 (r = 0.180, P<0.05) was evident with increased levels for all trachoma grades. New risk factors for chronic scarring trachoma included IL-6 and IL-15 (r = 0.259 and 0.292, respectively, P<0.005 for both) with increased levels for concurrent C. trachomatis infections (r = 0.206, P<0.05, and r = 0.304, P<0.005, respectively). Chemokine protein levels for CCL11 (Eotaxin), CXCL8 (IL-8), CXCL9 (MIG), and CCL2 (MCP-1) were elevated in chronic scarring trachoma compared with age and sex matched controls (P<0.05, for all). CONCLUSIONS/SIGNIFICANCE: Our quantitative detection of previously uncharacterized and partially characterized cytokines, a soluble cytokine receptor, and chemokines for each trachoma grade and associations with C. trachomatis infections provide, to date, the most comprehensive immunologic evaluation of trachoma. These findings highlight novel pathologic and protective factors involved in trachomatous disease, which will aid in designing immunomodulating therapeutics and a vaccine.

Impact of annual targeted treatment on infectious trachoma and susceptibility to reinfection.

CONTEXT: The World Health Organization developed the SAFE strategy (Surgery for trichiasis; Antibiotics for Chlamydia trachomatis infection; Facial cleanliness; and Environmental improvement) to eliminate blinding trachoma globally by the year 2020. Despite a number of studies using various intervals of treatment for different prevalence rates, there has been a lack of sufficient follow-up beyond the final treatment point to determine rates of recurrence of disease and infection and the risk factors that may contribute to each. OBJECTIVE: To evaluate the impact of 2 annual targeted azithromycin treatments on active trachoma and C trachomatis infection rates over 3 years in Vietnam. DESIGN, SETTING, AND PARTICIPANTS: Three communes were randomly selected for a longitudinal study in Vietnam from November 2000 through November 2003. Individuals (n = 3186) were graded for trachoma followed by conjunctival sampling to detect chlamydiae by commercial polymerase chain reaction. Grading and chlamydial detection were repeated every 6 months for 3 years. INTERVENTION: Azithromycin was given to children aged 5 through 15 years with active trachoma and their household members in SAFE and SA communes at baseline and 12 months; these communes were compared with the S-only control commune that did not receive azithromycin targeted treatment. MAIN OUTCOME MEASURES: Prevalence and incidence of active trachoma and C trachomatis infection in all communes at baseline, 6, 12, 18, 24, and 36 months. Subgroup analysis evaluated new infection, continuing infection, and reinfection at 6, 12, 18, 24, and 36 months and risk factors for each. RESULTS: Reinfection rates increased significantly between 12 and 36 months for SAFE (from 1.6 to 29.3 per 1000; P<.001) and SA (5.1 to 25.3 per 1000; P = .002) communes but not for the S-only commune (13.4 to 6.7 per 1000; P = .55) after 24 months. Compared with the S-only commune, mixed-effects and generalized estimating equations (GEE) logistic models showed that reinfection risk was significantly higher for SAFE (odds ratio [OR], 4.1; 95% confidence interval [CI], 1.5-9.8; P = .005) and SA (OR, 4.2; 95% CI, 1.1-17.3; P = .04) communes at 36 months. CONCLUSIONS: Increasing reinfection rates suggest that treatment may interrupt the duration of infection required for developing immunity, increasing the number of individuals susceptible to reinfection and adversely affecting disease prevalence over time. Additional research is needed to determine optimal trachoma control strategies, including evaluation of the "F" and "E" components. TRIAL REGISTRATION: www.actr.org.au Identifier: 12606000360516.

Correlating Chlamydia trachomatis infectious load with urogenital ecological success and disease pathogenesis.

The association of infectious burden of Chlamydia trachomatis with patient characteristics and clinical disease may have implications for understanding disease pathogenesis. We examined chlamydial load from 171 urine samples where load was based on copy number of organisms per copy number of eukaryotic cells derived by real-time quantitative PCR. High- (E, F, G) and low-prevalence (Ia, H, J, Ja) genotypes in the population had similar loads, suggesting a similar propensity for replicating in vivo, despite their differential ecological success. Symptomatic and asymptomatic patients also had similar chlamydial loads, indicating that virulence differences are likely not associated with variations in replication. There was a significant difference in genotypes by age for F (<31 years; P = 0.031) and for H where the mean age was lower than for the most prevalent genotype, E (P=0.013). Also, men had a significantly lower load than women when the genotype was F (P=0.042), although there was no significant difference in load between partners. Patients with recurrent chlamydial infections had a significant reduction in load with each subsequent episode regardless of genotype (P=0.007), suggesting that immune defenses do not block chlamydial entry but may impact replication. Additionally, the probability of being infected with J was 7.7-fold higher in patients with prior chlamydial infections (P=0.016), and although the loads were lower when compared with patients without prior infection, the results did not reach statistical significance. These findings suggest that chlamydial burden could be an important marker for recurrence and host immune response, which would facilitate pathogenesis research.