Plasma cytokine profiles in subjects with high-functioning autism spectrum disorders

Background

Accumulating evidence suggests that dysregulation of the immune system is involved in the pathophysiology of autism spectrum disorders (ASD). The aim of the study was to explore immunological markers in peripheral plasma samples from non-medicated subjects with high-functioning ASD.

Methodology/Principal Findings

A multiplex assay for cytokines and chemokines was applied to plasma samples from male subjects with high-functioning ASD (n = 28) and matched controls (n = 28). Among a total of 48 analytes examined, the plasma concentrations of IL-1β, IL-1RA, IL-5, IL-8, IL-12(p70), IL-13, IL-17 and GRO-α were significantly higher in subjects with ASD compared with the corresponding values of matched controls after correction for multiple comparisons.

Conclusion/Significance

The results suggest that abnormal immune responses as assessed by multiplex analysis of cytokines may serve as one of the biological trait markers for ASD.     

Performance of swabs, lavage, and diluents to quantify biomarkers of female genital tract soluble mucosal mediators

Background

Measurement of immune mediators and antimicrobial activity in female genital tract secretions may provide biomarkers predictive of risk for HIV-1 acquisition and surrogate markers of microbicide safety. However, optimal methods for sample collection do not exist. This study compared collection methods.

Methods

Secretions were collected from 48 women (24 with bacterial vaginosis [BV]) using vaginal and endocervical Dacron and flocked swabs. Cervicovaginal lavage (CVL) was collected with 10 mL of Normosol-R (n = 20), saline (n = 14), or water (n = 14). The concentration of gluconate in Normosol-R CVL was determined to estimate the dilution factor. Cytokine and antimicrobial mediators were measured by Luminex or ELISA and corrected for protein content. Endogenous anti-HIV-1 and anti-E. coli activity were measured by TZM-bl assay or E. coli growth.

Results

Higher concentrations of protein were recovered by CVL, despite a 10-fold dilution of secretions, as compared to swab eluents. After protein correction, endocervical swabs recovered the highest mediator levels regardless of BV status. Endocervical and vaginal flocked swabs recovered significantly higher levels of anti-HIV-1 and anti-E. coli activity than Dacron swabs (P<0.001). BV had a significant effect on CVL mediator recovery. Normosol-R tended to recover higher levels of most mediators among women with BV, whereas saline or water tended to recover higher levels among women without BV. Saline recovered the highest levels of anti-HIV-1 activity regardless of BV status.

Conclusions

Endocervical swabs and CVL collected with saline provide the best recovery of most mediators and would be the optimal sampling method(s) for clinical trials.     

The homeostatic chemokine CCL21 predicts mortality and may play a pathogenic role in heart failure

Background

CCL19 and CCL21, acting through CCR7, are termed homeostatic chemokines. Based on their role in concerting immunological responses and their proposed involvement in tissue remodeling, we hypothesized that these chemokines could play a pathogenic role in heart failure (HF).

Methodology/Principal Findings

Our main findings were: (i) Serum levels of CCL19 and particularly CCL21 were markedly raised in patients with chronic HF (n = 150) as compared with healthy controls (n = 20). A CCL21 level above median was independently associated with all-cause mortality. (ii) In patients with HF following acute myocardial infarction (MI; n = 232), high versus low CCL21 levels 1 month post-MI were associated with cardiovascular mortality, even after adjustment for established risk factors. (iii). Explanted failing human LV tissue (n = 29) had markedly increased expression of CCL21 as compared with non-failing myocardium (n = 5). (iv) Our studies in CCR7^−/− mice showed improved survival and attenuated increase in markers of myocardial dysfunction and wall stress in post-MI HF after 1 week, accompanied by increased myocardial expression of markers of regulatory T cells. (v) Six weeks post-MI, there was an increase in markers of myocardial dysfunction and wall stress in CCR7 deficient mice.

Conclusions/Significance

High serum levels of CCL21 are independently associated with mortality in chronic and acute post-MI HF. Our findings in CCR7 deficient mice may suggest that CCL21 is not only a marker, but also a mediator of myocardial failure. However, while short term inhibition of CCR7 may be beneficial following MI, a total lack of CCR7 during long-term follow-up could be harmful.     

An Analysis of Regulatory T-Cell and Th-17 Cell Dynamics during Cytomegalovirus Replication in Solid Organ Transplant Recipients

Background

CMV-specific T-cells are crucial to control CMV-replication post-transplant. Regulatory T-cells (T-regs) are associated with a tolerant immune state and may contribute to CMV-replication. However, T-cell subsets such as T-regs and IL-17 producing T-cells (Th-17) are not well studied in this context. We explored T-regs and Th-17 frequencies during CMV-replication after transplantation.

Methods

We prospectively evaluated 30 transplant patients with CMV-viremia. We quantified CMV-specific CD4⁺ and CD8⁺ T-cells, T-regs (CD4⁺CD25⁺FoxP3⁺) and Th-17 frequencies using flow-cytometry and followed patients requiring anti-viral treatment. Two subsets were compared: anti-viral treatment requirement (n = 20) vs. spontaneous clearance of viremia (n = 10).

Results

Higher initial CMV-specific CD4⁺ T-cells and lower T-regs were observed in patients with spontaneous clearance (p = 0.043; p = 0.021 respectively). Using a ratio of CMV-specific CD4⁺ T-cells to T-regs allowed prediction of viral clearance with 80% sensitivity and 90% specificity (p = 0.001). One month after stop of treatment, the same correlation was observed in patients protected from CMV-relapse. The ratio of CMV-specific CD4⁺ T-cells to T-regs allowed prediction of relapse with 85% sensitivity and 86% specificity (p = 0.004). Th-17 responses were not correlated with virologic outcomes.

Conclusions

This study provides novel insights into T-regs and Th-17 subpopulations during CMV-replication after transplantation. These preliminary data suggest that measurement of CMV-specific CD4⁺ T-cells together with T-regs has value in predicting spontaneous clearance of viremia and relapse.     

T regulatory cells in cord blood--FOXP3 demethylation as reliable quantitative marker

Background

Regulatory T-cells (Tregs), characterized as CD4+CD25^hi T-cells expressing FOXP3, play a crucial role in controlling healthy immune development during early immune maturation. Recently, FOXP3 demethylation was suggested to be a novel marker for natural Tregs in adults. In cord blood, the role and function of Tregs and its demethylation is poorly understood. We assessed FOXP3 demethylation in cord blood in relation to previously used Treg markers such as CD4⁺CD25^hi, FOXP3 mRNA, protein expression, and suppressive Treg function.

Methodology

Cord blood mononuclear cells (CBMC) were isolated from 70 healthy neonates, stimulated for 3 days with the microbial stimulus lipid A (LpA), and allergen Dermatophagoides pteronyssinus (Derp1). Tregs (CD4+CD25^hi, intracellular, mRNA FOXP3 expression, isolated cells), DNA methylation of the FOXP3-locus and suppressive Treg function were assessed.

Principal Findings

Demethylation of FOXP3 in whole blood was specific for isolated CD4+CD25^hi Tregs. Demethylation of FOXP3 was positively correlated with unstimulated and LpA-stimulated FOXP3 mRNA-expression (p≤0.05), and CD4⁺CD25^hi T-cells (p≤0.03). Importantly, increased FOXP3 demethylation correlated with more efficient suppressive capacity of Tregs (r = 0.72, p = 0.005). Furthermore, FOXP3 demethylation was positively correlated with Th2 cytokines (IL-5, IL-13) following LpA-stimulation (p = 0.006/0.04), with Th2 and IL-17 following Derp1+LpA-stimulations (p≤0.009), but not Th1 cytokines (IFN-γ).

Conclusions

FOXP3 demethylation reliable quantifies Tregs in cord blood. FOXP3 demethylation corresponds well with the suppressive potential of Tregs. The resulting strict correlation with functionally suppressive Tregs and the relative ease of measurement render it into a valuable novel marker for large field studies assessing Tregs as qualitative marker indicative of functional activity.     

Why CCR2 and CCR5 blockade failed and why CCR1 blockade might still be effective in the treatment of rheumatoid arthritis

Background

The aim of this study was to provide more insight into the question as to why blockade of CCR1, CCR2, and CCR5 may have failed in clinical trials in rheumatoid arthritis (RA) patients, using an in vitro monocyte migration system model.

Methodology/Principal Findings

Monocytes from healthy donors (HD; n = 8) or from RA patients (for CCR2 and CCR5 antibody n = 8; for CCR1 blockade n = 13) were isolated from peripheral blood and pre-incubated with different concentrations of either anti-CCR1, anti-CCR2, or anti-CCR5 blocking antibodies (or medium or isotype controls). In addition, a small molecule CCR1 antagonist (BX471) was tested. Chemotaxis was induced by CCL2/MCP-1 (CCR2 ligand), CCL5/RANTES (CCR1 and CCR5 ligand), or by a mix of 5 RA synovial fluids (SFs), and cellular responses compared to chemotaxis in the presence of medium alone. Anti-CCR2 antibody treatment blocked CCL2/MCP-1-induced chemotaxis of both HD and RA monocytes compared to isotype control. Similarly, anti-CCR5 antibody treatment blocked CCL5/RANTES-induced chemotaxis of RA monocytes. While neither CCR2 nor CCR5 blocking antibodies were able to inhibit SF-induced monocyte chemotaxis, even when both receptors were blocked simultaneously, both anti-CCR1 antibodies and the CCR1 antagonist were able to inhibit SF-induced monocyte chemotaxis.

Conclusions/Significance

The RA synovial compartment contains several ligands for CCR1, CCR2, and CCR5 as well as other chemokines and receptors involved in monocyte recruitment to the site of inflammation. The results suggest that CCR2 and CCR5 are not critical for the migration of monocytes towards the synovial compartment in RA. In contrast, blockade of CCR1 may be effective. Conceivably, CCR1 blockade failed in clinical trials, not because CCR1 is not a good target, but because very high levels of receptor occupancy at all times may be needed to inhibit monocyte migration in vivo.     

Involvement of CCR6/CCL20/IL-17 axis in NSCLC disease progression

Objectives

Autocrine and paracrine chemokine/chemokine receptor-based interactions promote non-small-cell-lung-cancer (NSCLC) carcinogenesis. CCL20/CCR6 interactions are involved in prostatic and colonic malignancy pathogenesis. The expression and function of CCL20/CCR6 and its related Th-17 type immune response in NSCLC is not yet defined. We sought to characterize the role of the CCL20/CCR6/IL-17 axis in NSCLC tumor growth.

Methods

A specialized histopathologist blindly assessed CCL20/CCR6 expression levels in 49 tissue samples of NSCLC patients operated in our department. Results were correlated to disease progression. Colony assays, ERK signaling and chemokine production were measured to assess cancer cell responsiveness to CCL20 and IL-17 stimulation.

Results

CCL20 was highly expressed in the majority (38/49, 77.5%) of tumor samples. Only a minority of samples (8/49, 16.5%) showed high CCR6 expression. High CCR6 expression was associated with a shorter disease-free survival (P = 0.008) and conferred a disease stage-independent 4.87-fold increased risk for disease recurrence (P = 0.0076, CI 95% 1.52–15.563). Cancerous cell colony-forming capacity was increased by CCL20 stimulation; this effect was dependent in part on ERK phosphorylation and signaling. IL-17 expression was detected in NSCLC; IL-17 potentiated the production of CCL20 by cancerous cells.

Conclusion

Our findings suggest that the CCL20/CCR6 axis promotes NSCLC disease progression. CCR6 is identified as a potential new prognostic marker and the CCL20/CCR6/IL-17 axis as a potential new therapeutic target. Larger scale studies are required to consolidate these observations.     

Hepatic expression patterns of inflammatory and immune response genes associated with obesity and NASH in morbidly obese patients

Background

Obesity modulates inflammation and activation of immune pathways which can lead to liver complications. We aimed at identifying expression patterns of inflammatory and immune response genes specifically associated with obesity and NASH in the liver of morbidly obese patients.

Methodology/Principal Findings

Expression of 222 genes was evaluated by quantitative RT-PCR in the liver of morbidly obese patients with histologically normal liver (n = 6), or with severe steatosis without (n = 6) or with NASH (n = 6), and in lean controls (n = 5). Hepatic expression of 58 out of 222 inflammatory and immune response genes was upregulated in NASH patients. The most notable changes occurred in genes encoding chemokines and chemokine receptors involved in leukocyte recruitment, CD and cytokines involved in the T cell activation towards a Th1 phenotype, and immune semaphorins. This regulation seems to be specific for the liver since visceral adipose tissue expression and serum levels of MCP1, IP10, TNFα and IL6 were not modified. Importantly, 47 other genes were already upregulated in histologically normal liver (e.g. CRP, Toll-like receptor (TLR) pathway). Interestingly, serum palmitate, known to activate the TLR pathway, was increased with steatosis.

Conclusion/Significance

The liver of obese patients without histological abnormalities already displayed a low-grade inflammation and could be more responsive to activators of the TLR pathway. NASH was then characterized by a specific gene signature. These findings help to identify new potential actors of the pathogenesis of NAFLD.     

Association of differentiation state of CD4+ T cells and disease progression in HIV-1 perinatally infected children

Background

In the USA, most HIV-1 infected children are on antiretroviral drug regimens, with many individuals surviving through adolescence and into adulthood. The course of HIV-1 infection in these children is variable, and understudied.

Methodology/Principal Findings

We determined whether qualitative differences in immune cell subsets could explain a slower disease course in long term survivors with no evidence of immune suppression (LTS-NS; CD4%≥25%) compared to those with severe immune suppression (LTS-SS; CD4%≤15%). Subjects in the LTS-NS group had significantly higher frequencies of naïve (CCR7+CD45RA+) and central memory (CCR7+CD45RA−) CD4+ T cells compared to LTS-SS subjects (p = 0.0005 and <0.0001, respectively). Subjects in the rapid progressing group had significantly higher levels of CD4+ T_EMRA (CCR7−CD45RA+) cells compared to slow progressing subjects (p<0.0001).

Conclusions/Significance

Rapid disease progression in vertical infection is associated with significantly higher levels of CD4+ T_EMRA (CCR7−CD45RA+) cells.     

Monitoring of regulatory T cell frequencies and expression of CTLA-4 on T cells, before and after DC vaccination, can predict survival in GBM patients

Purpose

Dendritic cell (DC) vaccines have recently emerged as an innovative therapeutic option for glioblastoma patients. To identify novel surrogates of anti-tumor immune responsiveness, we studied the dynamic expression of activation and inhibitory markers on peripheral blood lymphocyte (PBL) subsets in glioblastoma patients treated with DC vaccination at UCLA.

Experimental Design

Pre-treatment and post-treatment PBL from 24 patients enrolled in two Phase I clinical trials of dendritic cell immunotherapy were stained and analyzed using flow cytometry. A univariate Cox proportional hazards model was utilized to investigate the association between continuous immune monitoring variables and survival. Finally, the immune monitoring variables were dichotomized and a recursive partitioning survival tree was built to obtain cut-off values predictive of survival.

Results

The change in regulatory T cell (CD3⁺CD4⁺CD25⁺CD127^low) frequency in PBL was significantly associated with survival (p = 0.0228; hazard ratio = 3.623) after DC vaccination. Furthermore, the dynamic expression of the negative co-stimulatory molecule, CTLA-4, was also significantly associated with survival on CD3⁺CD4⁺ T cells (p = 0.0191; hazard ratio = 2.840) and CD3⁺CD8⁺ T cells (p = 0.0273; hazard ratio = 2.690), while that of activation markers (CD25, CD69) was not. Finally, a recursive partitioning tree algorithm was utilized to dichotomize the post/pre fold change immune monitoring variables. The resultant cut-off values from these immune monitoring variables could effectively segregate these patients into groups with significantly different overall survival curves.

Conclusions

Our results suggest that monitoring the change in regulatory T cell frequencies and dynamic expression of the negative co-stimulatory molecules on peripheral blood T cells, before and after DC vaccination, may predict survival. The cut-off point generated from these data can be utilized in future prospective immunotherapy trials to further evaluate its predictive validity.     

Coreceptor and cytokine concentrations may not explain differences in disease progression observed in HIV-1 clade A and D infected Ugandans

Background

The use of cellular coreceptors and modulation of cytokine concentrations by HIV to establish a productive infection is well documented. However, it is unknown whether the expression of these proteins affects the course of HIV clade A and D disease, reported to have different progression rates.

Methodology/Principal Findings

We investigated whether the number of CD4⁺ T-cells expressing CCR5 or CXCR4, the density of these coreceptors and concentrations of specific immune proteins linked to HIV pathogenesis vary between individuals infected with HIV clade A or D. We undertook additional analyses stratifying participants by early (CD4>500 cells/µl) or late (CD4<200 cells/µl) disease stage. Whole blood samples were taken from 50 HIV-1 infected individuals drawn from cohorts in rural south-west Uganda. Late stage participants had less than half the number of CD4⁺/CCR5⁺ T-cells (p = 0.0113) and 5.6 times fewer CD4⁺/CXCR4⁺ cells (p<0.0001) than early stage participants. There was also a statistically significant difference in the density of CXCR4 on CD4⁺ cells between clade A and D infected early stage participants (142 [A] vs 84 [D]; p = 0.0146). Across all participants we observed significantly higher concentration of Th₁ cytokines compared to Th₂ (66.4 vs 23.8 pg/ml; p<0.0001). Plasma concentrations of IFNγ and IL-2 were 1.8 and 2.4 fold lower respectively in Late-D infected participants compared to Late-A participants. MIP-1β levels also decreased from 118.0 pg/ml to 47.1 pg/ml (p = 0.0396) as HIV disease progressed.

Conclusions/Significance

We observed specific alterations in the abundance of CD4⁺/CCR5⁺ and CD4⁺/CXCR4⁺ T-cells, and concentrations of immune proteins across different HIV clades and as infection progresses. Our results suggest that these changes are unlikely to explain the observed differences in disease progression between subtype A and D infections. However, our observations further the understanding of the natural progression of non-clade B HIV infection and how the virus adapts to exploit the host environment.     

CCR5Δ32 genotype leads to a Th2 type directed immune response in ESRD patients

Background

In patients with end stage renal disease (ESRD) we observed protection from inflammation-associated mortality in CCR5Δ32 carriers, leading to CCR5 deficiency, suggesting impact of CCR5Δ32 on inflammatory processes. Animal studies have shown that CCR5 deficiency is associated with a more pronounced Th2 type immune response, suggesting that in human CCR5Δ32 carriers the immune response may be more Th2 type directed. So, in the present study we determined the Th1-Th2 type directed immune response in ESRD patients carrying and not carrying the CCR5Δ32 genetic variant after stimulation.

Methodology/Principal Findings

We tested this hypothesis by determining the levels of IFN-γ and IL-4 and the distribution of Th1, Th2 and Th17 directed circulating CD4+ and CD8+ T cells and regulatory T cells (Tregs) after stimulation in ESRD patients with (n = 10) and without (n = 9) the CCR5Δ32 genotype. The extracellular levels of IFN-γ and IL-4 did not differ between CCR5Δ32 carriers and non carriers. However, based on their intracellular cytokine profile the percentages IL-4 secreting CD4+ and CD8+ T cells carrying the CCR5Δ32 genotype were significantly increased (p = 0.02, respectively p = 0.02) compared to non carriers, indicating a more Th2 type directed response. Based on their intracellular cytokine profile the percentages IFN-γ and IL-17 secreting T cells did not differ between carriers and non-carriers nor did the percentage Tregs, indicating that the Th1, Th17 and T regulatory response was not affected by the CCR5Δ32 genotype.

Conclusions/Significance

This first, functional human study shows a more pronounced Th2 type immune response in CCR5Δ32 carriers compared to non carriers. These differences may be involved in the previously observed protection from inflammation-associated mortality in ESRD patients carrying CCR5Δ32.     

Effects of maraviroc and efavirenz on markers of immune activation and inflammation and associations with CD4+ cell rises in HIV-infected patients

Background

Maraviroc treatment for HIV-1 infected patients results in larger CD4⁺ T cell rises than are attributable to its antiviral activity alone. We investigated whether this is due to modulation of T cell activation and inflammation.

Methods and Findings

Thirty maraviroc-treated patients from the Maraviroc versus Efavirenz Regimens as Initial Therapy (MERIT) study were randomly selected from among those who had CCR5-tropic (R5) HIV on screening and achieved undetectable HIV RNA (<50 copies/mL) by Week 48. Efavirenz-treated controls were matched for baseline characteristics to the maraviroc-treated patients selected for this substudy. Changes in immune activation and inflammation markers were examined for associations with CD4⁺ T cell changes. Maraviroc treatment tended to result in more rapid decreases in CD38 expression on CD4⁺ T cells and in plasma D-dimer concentrations than did treatment with efavirenz. The proportion of patients with high-sensitivity C-reactive protein >2 µg/mL increased from 45% to 66% in the efavirenz arm, but remained constant in the maraviroc arm (P = 0.033). Decreases in CD38 expression on CD8⁺ T cells were correlated with CD4⁺ T cell rises for maraviroc treatment (r = −0.4, P = 0.048), but not for treatment with efavirenz.

Conclusions

Maraviroc-treated patients had earlier, modest decreases in certain markers of immune activation and inflammation, although in this small study, many of the differences were not statistically significant. Levels of high-sensitivity C-reactive protein remained constant in the maraviroc arm and increased in the efavirenz arm. Decreases in immune activation correlated with increased CD4⁺ T cell gains.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00098293","term_id":"NCT00098293"}}NCT00098293     

A randomized trial to assess anti-HIV activity in female genital tract secretions and soluble mucosal immunity following application of 1% tenofovir gel

Background

Preclinical and early phase clinical microbicide studies have not consistently predicted the outcome of efficacy trials. To address this gap, candidate biomarkers of microbicide pharmacodynamics and safety were evaluated in a double-blind, placebo-controlled trial of tenofovir gel, the first microbicide to demonstrate significant protection against HIV acquisition.

Methods

30 women were randomized to apply a single daily dose of tenofovir or placebo gel for 14 consecutive days. Anti-HIV activity was measured in cervicovaginal lavage (CVL) on Days 0, 3, 7, 14 and 21 by luciferase assay as a surrogate marker of pharmacodynamics. Endogenous activity against E. coli and HSV-2 and concentrations of immune mediators were quantified in CVL as candidate biomarkers of safety. Tenofovir levels were measured in CVL and blood.

Results

A significant increase in anti-HIV activity was detected in CVL from women who applied tenofovir gel compared to their endogenous anti-HIV activity in genital tract secretions on Day 0 and compared to activity in CVL from women in the placebo group. The activity correlated significantly with CVL concentration of tenofovir (r = 0.6, p<0.001) and fit a sigmoid E_max pharmacodynamic model. Anti-HIV activity in CVL from women who applied tenofovir persisted when virus was introduced in semen, whereas endogenous anti-HIV activity decreased. Tenofovir did not trigger an inflammatory response or induce sustained loss in endogenous antimicrobial activity or immune mediators.

Conclusions

Tenofovir gel had no deleterious impact on soluble mucosal immunity. The increased anti-HIV activity in CVL, which persisted in the presence of semen and correlated with tenofovir concentration, is consistent with the efficacy observed in a recent clinical trial. These results promote quantified CVL anti-HIV activity as a surrogate of tissue pharmacodynamics and as a potential biomarker of adherence to product. This simple, feasible and inexpensive bioassay may promote the development of models more predictive of microbicide efficacy.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00594373","term_id":"NCT00594373"}}NCT00594373     

Evidence That Lipopolisaccharide May Contribute to the Cytokine Storm and Cellular Activation in Patients with Visceral Leishmaniasis

Background

Visceral leishmaniasis (VL) is characterized by parasite-specific immunosuppression besides an intense pro-inflammatory response. Lipopolisaccharide (LPS) has been implicated in the immune activation of T-cell deficient diseases such as HIV/AIDS and idiopathic lymphocytopenia. The source of LPS is gram-negative bacteria that enter the circulation because of immunological mucosal barrier breakdown. As gut parasitization also occurs in VL, it was hypothesized that LPS may be elevated in leishmaniasis, contributing to cell activation.

Methodology/Principal Findings

Flow cytometry analysis and immunoassays (ELISA and luminex micro-beads system) were used to quantify T-cells and soluble factors. Higher LPS and soluble CD14 levels were observed in active VL in comparison to healthy subjects, indicating that LPS was bioactive; there was a positive correlation between these molecules (r = 0.61;p<0.05). Interestingly, LPS was negatively correlated with CD4⁺ (r = −0.71;p<0.01) and CD8⁺ T-cells (r = −0.65;p<0.05). Moreover, higher levels of activation-associated molecules (HLA-DR, CD38, CD25) were seen on T lymphocytes, which were positively associated with LPS levels. Pro-inflammatory cytokines and macrophage migration inhibitory factor (MIF) were also augmented in VL patients. Consistent with the higher immune activation status, LPS levels were positively correlated with the inflammatory cytokines IL-6 (r = 0.63;p<0.05), IL-8 (r = 0.89;p<0.05), and MIF (r = 0.64;p<0.05). Also, higher plasma intestinal fatty acid binding protein (IFABP) levels were observed in VL patients, which correlated with LPS levels (r = 0.57;p<0.05).

Conclusions/Significance

Elevated levels of LPS in VL, in correlation with T-cell activation and elevated pro-inflammatory cytokines and MIF indicate that this bacterial product may contribute to the impairment in immune effector function. The cytokine storm and chronic immune hyperactivation status may contribute to the observed T-cell depletion. LPS probably originates from microbial translocation as suggested by IFABP levels and, along with Leishmania antigen-mediated immune suppression, may play a role in the immunopathogenesis of VL. These findings point to possible benefits of antimicrobial prophylaxis in conjunction with anti-Leishmania therapy.     

Anterior Cervical Discectomy with Arthroplasty versus Arthrodesis for Single-Level Cervical Spondylosis: A Systematic Review and Meta-Analysis

Objective

To estimate the effectiveness of anterior cervical discectomy with arthroplasty (ACDA) compared to anterior cervical discectomy with fusion (ACDF) for patient-important outcomes for single-level cervical spondylosis.

Data sources

Electronic databases (MEDLINE, EMBASE, Cochrane Register for Randomized Controlled Trials, BIOSIS and LILACS), archives of spine meetings and bibliographies of relevant articles.

Study selection

We included RCTs of ACDF versus ACDA in adult patients with single-level cervical spondylosis reporting at least one of the following outcomes: functionality, neurological success, neck pain, arm pain, quality of life, surgery for adjacent level degeneration (ALD), reoperation and dysphonia/dysphagia. We used no language restrictions. We performed title and abstract screening and full text screening independently and in duplicate.

Data synthesis

We used random-effects model to pool data using mean difference (MD) for continuous outcomes and relative risk (RR) for dichotomous outcomes. We used GRADE to evaluate the quality of evidence for each outcome.

Results

Of 2804 citations, 9 articles reporting on 9 trials (1778 participants) were eligible. ACDA is associated with a clinically significant lower incidence of neurologic failure (RR  = 0.53, 95% CI  = 0.37–0.75, p = 0.0004) and improvement in the Neck pain visual analogue scale (VAS) (MD  = 6.56, 95% CI  = 3.22–9.90, p = 0.0001; Minimal clinically important difference (MCID)  = 2.5. ACDA is associated with a statistically but not clinically significant improvement in Arm pain VAS and SF-36 physical component summary. ACDA is associated with non-statistically significant higher improvement in the Neck Disability Index Score and lower incidence of ALD requiring surgery, reoperation, and dysphagia/dysphonia.

Conclusions

There is no strong evidence to support the routine use of ACDA over ACDF in single-level cervical spondylosis. Current trials lack long-term data required to assess safety as well as surgery for ALD. We suggest that ACDA in patients with single level cervical spondylosis is an option although its benefits and indication over ACDF remain in question.     

Cytokine profiles, CTL response and T cell frequencies in the peripheral blood of acute patients and individuals recovered from hepatitis E infection

Background

Hepatitis E is a major public health problem in the developing countries. Pathogenesis of hepatitis E virus (HEV) infection is poorly understood.

Methods

This case-control study included 124 Hepatitis E patients (46 acute and 78 recovered), 9 with prior exposure to HEV and 71 anti-HEV negative healthy controls. HEV induced CTL response by Elispot, cytokines/chemokines quantitation by Milliplex assay and peripheral CD4+ & CD8+ T cell frequencies by flow cytometry were assessed.

Results

Among the patient categories, HEV specific IFN-γ responses as recorded by Elispot were comparable. Comparisons of cytokines/chemokines revealed significantly high levels of IL-1α and sIL-2Rα during acute phase. Circulating peripheral CD4/CD8+ T-cell subsets in acute and recovered individuals were comparable compared to controls, while among patient categories CD8+T cell subset was significantly higher in recovered individuals.

Conclusions

Our findings suggest that IL-1α and sIL-2Rα play a role in the pathogenesis of acute Hepatitis E infection. Lack of robust HEV ORF2-specific CTL response in the peripheral blood of HEV infected patients during the acute and recovered phases of the disease may be associated with involvement of innate immune cells/localization of the immune events at the site of infection.     

The effectiveness of online cognitive behavioral treatment in routine clinical practice

Context

Randomized controlled trails have identified online cognitive behavioral therapy as an efficacious intervention in the management of common mental health disorders.

Objective

To assess the effectiveness of online CBT for different mental disorders in routine clinical practice.

Design

An uncontrolled before-after study, with measurements at baseline, posttest, 6-week follow-up, and 1-year follow-up.

Participants & Setting

1500 adult patients (female: 67%; mean age: 40 years) with a GP referral for psychotherapy were treated at a Dutch online mental health clinic for symptoms of depression (n = 413), panic disorder (n = 139), posttraumatic stress (n = 478), or burnout (n = 470).

Interventions

Manualized, web-based, therapist-assisted CBT, of which the efficacy was previously demonstrated in a series of controlled trials. Standardized duration of treatment varied from 5 weeks (online CBT for Posttraumatic stress) to 16 weeks (online CBT for Depression).

Main Outcome Measures

Validated self-report questionnaires of specific and general psychopathology, including the Beck Depression Inventory, the Impact of Event Scale, the Panic Disorder Severity Scale-Self Report, the Oldenburg Burnout Inventory, and the Depression Anxiety Stress Scales.

Results

Treatment adherence was 71% (n = 1071). Study attrition was 21% at posttest, 33% at 6-week FU and 65% at 1-year FU. Mixed-model repeated measures regression identified large short-term reductions in all measures of primary symptoms (d = 1.9±0.2 to d = 1.2±0.2; P<.001), which sustained up to one year after treatment. At posttest, rates of reliable improvement and recovery were 71% and 52% in the completer sample (full sample: 55%/40%). Patient satisfaction was high.

Conclusions

Results suggest that online therapist-assisted CBT may be as effective in routine practice as it is in clinical trials. Although pre-treatment withdrawal and long-term outcomes require further study, results warrant continued implementation of online CBT.     

Adoptive immunotherapy in postoperative hepatocellular carcinoma: a systemic review

Purpose

The effectiveness of immunotherapy for postoperative hepatocellular carcinoma patients is still controversial. To address this issue, we did a systemic review of the literatures and analyzed the data with emphasis on the recurrence and survival.

Methods

We searched six randomized controlled trials that included adoptive immunotherapy in the postoperative management of hepatocellular carcinoma and compared with non-immunotherapy postoperation. A meta-analysis was carried out to examine one- and 3-year recurrence and survival.

Results

The overall analysis revealed significantly reduced risk of 1-year recurrence in patients receiving adoptive immunotherapy (OR = 0.35; 95% CI, 0.17 to 0.71; p = 0.003), in that the risk of 3-year recurrence with a pooled OR estimated at 0.31 (95% CI 0.16 to 0.61; p = 0.001). However, no statistically significant difference was observed for 3-year survival between groups with adoptive immunotherapy and without adjuvant treatment (OR = 0.91; 95% CI, 0.45 to 1.84; P = 0.792).

Conclusions

Adjuvant immunotherapy with cytokine induced killer cells or lymphokine activated killer cells may reduce recurrence in postoperative hepatocellular carcinoma patients, but may not improve survival.     

A pilot study of IL-2Rα blockade during lymphopenia depletes regulatory T-cells and correlates with enhanced immunity in patients with glioblastoma

Background

Preclinical studies in mice have demonstrated that the prophylactic depletion of immunosuppressive regulatory T-cells (T_Regs) through targeting the high affinity interleukin-2 (IL-2) receptor (IL-2Rα/CD25) can enhance anti-tumor immunotherapy. However, therapeutic approaches are complicated by the inadvertent inhibition of IL-2Rα expressing anti-tumor effector T-cells.

Objective

To determine if changes in the cytokine milieu during lymphopenia may engender differential signaling requirements that would enable unarmed anti-IL-2Rα monoclonal antibody (MAbs) to selectively deplete T_Regs while permitting vaccine-stimulated immune responses.

Methodology

A randomized placebo-controlled pilot study was undertaken to examine the ability of the anti-IL-2Rα MAb daclizumab, given at the time of epidermal growth factor receptor variant III (EGFRvIII) targeted peptide vaccination, to safely and selectively deplete T_Regs in patients with glioblastoma (GBM) treated with lymphodepleting temozolomide (TMZ).

Results and Conclusions

Daclizumab treatment (n = 3) was well-tolerated with no symptoms of autoimmune toxicity and resulted in a significant reduction in the frequency of circulating CD4+Foxp3+ TRegs in comparison to saline controls (n = 3)( p = 0.0464). A significant (p<0.0001) inverse correlation between the frequency of TRegs and the level of EGFRvIII specific humoral responses suggests the depletion of TRegs may be linked to increased vaccine-stimulated humoral immunity. These data suggest this approach deserves further study.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00626015","term_id":"NCT00626015"}}NCT00626015