Intra-articular CD1c-expressing myeloid dendritic cells from rheumatoid arthritis patients express a unique set of T cell-attracting chemokines and spontaneously induce Th1, Th17 and Th2 cell activity

Introduction

Myeloid dendritic cells (mDCs) are potent T cell-activating antigen-presenting cells that have been suggested to play a crucial role in the regulation of immune responses in many disease states, including rheumatoid arthritis (RA). Despite this, studies that have reported on the capacity of naturally occurring circulating mDCs to regulate T cell activation in RA are still lacking. This study aimed to evaluate the phenotypic and functional properties of naturally occurring CD1c (BDCA-1)⁺ mDCs from synovial fluid (SF) compared to those from peripheral blood (PB) of RA patients.

Methods

CD1c⁺ mDC numbers and expression of costimulatory molecules were assessed by fluorescence-activated cell sorting (FACS) analysis in SF and PB from RA patients. Ex vivo secretion of 45 inflammatory mediators by mDCs from SF and PB of RA patients was determined by multiplex immunoassay. The capacity of mDCs from SF to activate autologous CD4⁺ T cells was measured.

Results

CD1c⁺ mDC numbers were significantly increased in SF versus PB of RA patients (mean 4.7% vs. 0.6%). mDCs from SF showed increased expression of antigen-presenting (human leukocyte antigen (HLA) class II, CD1c) and costimulatory molecules (CD80, CD86 and CD40). Numerous cytokines were equally abundantly produced by mDCs from both PB and SF (including IL-12, IL-23, IL-13, IL-21). SF mDCs secreted higher levels of interferon γ-inducible protein-10 (IP-10), monokine induced by interferon γ (MIG) and, thymus and activation-regulated chemokine (TARC), but lower macrophage-derived chemokine (MDC) levels compared to mDCs from PB. mDCs from SF displayed a strongly increased capacity to induce proliferation of CD4⁺ T cells associated with a strongly augmented IFNγ, IL-17, and IL-4 production.

Conclusions

This study suggests that increased numbers of CD1c⁺ mDCs in SF are involved in the inflammatory cascade intra-articularly by the secretion of specific T cell-attracting chemokines and the activation of self-reactive T cells.     

Differential response of BDCA-1+ and BDCA-3+ myeloid dendritic cells to respiratory syncytial virus infection

Background

Respiratory syncytial virus (RSV) is the leading cause of respiratory infections in children, elderly, and immunocompromised individuals. Severe infection is associated with short- and long-term morbidity including pneumonia, recurrent wheezing, and abnormal pulmonary function, and several lines of evidence indicate that impaired adaptive immune responses during infection are critical in the pathophysiology of RSV-mediated disease. Myeloid Dendritic cells (mDCs) play a pivotal role in shaping antiviral immune responses in the respiratory tract; however, few studies have examined the interactions between RSV and individual mDC subsets. In this study, we examined the effect of RSV on the functional response of primary mDC subsets (BDCA-1⁺ and BDCA-3⁺) isolated from peripheral blood.

Methods

BDCA-1⁺ and BDCA-3⁺ mDCs were isolated from the peripheral blood of healthy adults using FACS sorting. Donor-matched BDCA-1⁺ and BDCA-3⁺ mDCs were infected with RSV at a multiplicity of infection (MOI) of 5 for 40 hours. After infection, cells were analyzed for the expression of costimulatory molecules (CD86, CD80, and PD-L1), cytokine production, and the ability to stimulate allogenic CD4⁺ T cell proliferation.

Results

Both BDCA-1⁺ and BDCA-3⁺ mDCs were susceptible to infection with RSV and demonstrated enhanced expression of CD86, and the inhibitory costimulatory molecules CD80 and PD-L1. Compared to BDCA-3⁺ mDCs, RSV-infected BDCA-1⁺ mDC produced a profile of cytokines and chemokines predominantly associated with pro-inflammatory responses (IL-1β, IL-6, IL-12, MIP-1α, and TNF-α), and both BDCA-1⁺ and BDCA-3⁺ mDCs were found to produce IL-10. Compared to uninfected mDCs, RSV-infected BDCA-1⁺ and BDCA-3⁺ mDCs demonstrated a reduced capacity to stimulate T cell proliferation.

Conclusions

RSV infection induces a distinct pattern of costimulatory molecule expression and cytokine production by BDCA-1⁺ and BDCA-3⁺ mDCs, and impairs their ability to stimulate T cell proliferation.The differential expression of CD86 and pro-inflammatory cytokines by highly purified mDC subsets in response to RSV provides further evidence that BDCA-1⁺ and BDCA-3⁺ mDCs have distinct roles in coordinating the host immune response during RSV infection. Findings of differential expression of PD-L1 and IL-10 by infected mDCs, suggests possible mechanisms by which RSV is able to impair adaptive immune responses.     

Enterovirus-Infected β-Cells Induce Distinct Response Patterns in BDCA1+ and BDCA3+ Human Dendritic Cells

Enteroviruses often cause mild disease, yet are also linked to development of autoimmune diabetes. Dendritic cells (DCs) shape both innate and adaptive immune responses, including anti-viral responses. How different human DC subsets shape anti-viral responses, whether they have complementary or overlapping functions and how this relates to autoimmune responses is largely unknown. We used enterovirus-infected β-cells and freshly isolated human myeloid DC (mDC) subsets as a model for autoimmune type 1 diabetes. Our data show that both the BDCA1⁺ and BDCA3⁺ mDC subsets engulf mock- as well as virus-infected β-cells, albeit BDCA1⁺ mDCs are more efficient. Uptake of enterovirus-infected, but not mock-infected cells, activated both DC subsets as indicated by the induction of co-stimulatory molecules and secretion of type I and type III interferons. Both subsets produced similar amounts of interferon-α, yet the BDCA3⁺ DC were superior in IFN-λ production. The BDCA1⁺ mDCs more strongly upregulated PD-L1, and were superior in IL-12 and IL-10 production as compared to the BDCA3⁺ DC. Despite lack of IL-12 production by the BDCA3+ DC, both BDCA1⁺ and BDCA3⁺ DCs activated T cells in allogeneic mixed lymphocyte reaction towards a Th1-type reactivity while suppressing Th2-associated cytokines.     

Peripheral blood CD4+CD25+CD127low regulatory T cells are significantly increased by tocilizumab treatment in patients with rheumatoid arthritis: increase in regulatory T cells correlates with clinical response

IntroductionTocilizumab (TCZ), an anti-interleukin-6 receptor antibody, is clinically effective against rheumatoid arthritis (RA), and several reports have indicated how TCZ influences a number of mechanisms underlying RA pathogenesis. However, it is still unclear whether TCZ affects inflammatory cells in peripheral blood and whether any such changes are associated with clinical response. We evaluated associations between proportions of subsets of peripheral immune cells and clinical response in patients with RA treated with TCZ.MethodsThirty-nine consecutive patients with RA who started to receive TCZ as their first biologic between March 2010 and April 2012 were enrolled. The proportions of several subsets of peripheral cells with their levels of expression of differentiation markers, activation markers and costimulatory molecules were measured sequentially from baseline to week 52 by flow cytometry analysis.ResultsClinical Disease Activity Index (CDAI) remission was achieved in 53.8% of patients at week 52 of TCZ therapy. The proportions of CD4⁺CD25⁺CD127^low regulatory T cells (T_reg) and HLA-DR⁺ activated T_reg cells significantly increased with TCZ therapy (P < 0.001 and P < 0.001, respectively), whereas proportions of CD3⁺CD4⁺CXCR3⁻CCR6⁺CD161⁺ T helper 17 cells did not change over the 52 weeks. The proportions of CD20⁺CD27⁺ memory B cells, HLA-DR⁺CD14⁺ and CD69⁺CD14⁺ activated monocytes, and CD16⁺CD14⁺ monocytes significantly decreased (P < 0.001, P < 0.001, P < 0.001 and P < 0.001, respectively). Among them, only the change in T_reg cells was inversely correlated with the change in CDAI score (ρ = −0.40, P = 0.011). The most dynamic increase in T_reg cells was observed in the CDAI remission group (P < 0.001).ConclusionThis study demonstrates that TCZ affected proportions of circulating immune cells in patients with RA. The proportion of T_reg cells among CD4⁺ cells correlated well with clinical response.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0526-4) contains supplementary material, which is available to authorized users.     

Constitutively altered frequencies of circulating follicullar helper T cell counterparts and their subsets in rheumatoid arthritis

Introduction

Circulating CD4 T cells expressing CXCR5, ICOS and/or PD-1 are counterparts of follicular helper T cells (Tfh). There are three subpopulations of circulating Tfh (cTfh): CXCR5 + CXCR3 + CCR6- (Tfh-Th1), CXCR5 + CXCR3-CCR6- (Tfh-Th2) and CXCR5 + CXCR3-CCR6+ (Tfh-Th17). Our objective was to study the B cell helping capacity of cTfh subsets, and examine their frequency in Rheumatoid Arthritis (RA) patients, together with the frequency of circulating plasmablasts (CD19 + CD20-CD38^high).

Methods

Peripheral blood was drawn from RA patients with active disease (RA-a, DAS28 >2.6) (n = 17), RA in remission (RA-r, DAS28 <2.6) (n = 17) and healthy controls (HC) (n = 34). cTfh and plasmablast frequencies were determined by flow cytometry. Cocultures of sorted CD4 + CXCR5+ T cell subpopulations were established with autologous CD19 + CD27- naïve B cells of HC, and concentrations of IgG, A and M were measured in supernatants.

Results

Isolated Tfh-Th2 and Tfh-Th17 but not Tfh-Th1 cells, induced naïve B cells to secrete IgG and IgA. The frequency of CXCR5+ cells gated for CD4+ T cells was not different among HC, RA-a and RA-r. In contrast, both RA-a and RA-r patients demonstrated an increased frequency of CD4 + CXCR5 + ICOS+ T cells and augmented (%Tfh-Th2 + %Tfh-Th17)/%Tfh-Th1 ratio as compared with HC. In addition, RA-a but not RA-r patients, showed an increased frequency of circulating plasmablasts.

Conclusion

Both RA-a and RA-r patients demonstrate an increased frequency of cTfh and overrepresentation of cTfh subsets bearing a B cell helper phenotype, suggesting that altered germinal center dynamics play a role in RA pathogenesis. In contrast, only RA-a patients show an increased proportion of circulating plasmablasts.     

Regulation of Th1/Th2 and Th17/Treg by pDC/mDC imbalance in primary immune thrombocytopenia

This study investigates the regulatory effect of plasmacytoid dendritic cells (pDC)/myeloid dendritic cells (mDC) imbalance on balance of Th1/Th2 and Th17/Treg in primary immune thrombocytopenia (ITP). A total of 30 untreated ITP patients and 20 healthy controls were recruited. Compared with healthy control, the pDC proportion of ITP patients was significantly reduced (P = 0.004), while the mDC proportion was not significantly changed (P = 0.681), resulting in a decrease in the pDC/mDC ratio (P = 0.001). Additionally, compared with controls, serum levels of interleukin (IL)-6, IL-12, and IL-23 were increased in ITP patients (P < 0.001), and mRNA levels of IL-12p40, IL-12p35, and IL-23p19 were also increased (P =0.014, P = 0.043, P < 0.001). Compared with the healthy control, the proportion of Th1 and Th17 cells in ITP patients increased (P = 0.001, P = 0.031). Serum levels of interferon gamma (IFN-γ) and IL-17 in ITP patients also increased (P = 0.025, P = 0.005). Furthermore, T-bet and RORγt mRNA levels were increased in peripheral blood of ITP patients (P = 0.018, P < 0.001). Correspondingly, the proportion of Th2 and Treg cells decreased (P = 0.007, P < 0.001), along with a decrease in serum IL-4 and transforming growth factor beta (TGF-β) (P = 0.028, P = 0.042), and an increase in GATA-3 mRNA (P < 0.001). However, there was no significant difference in Foxp3 mRNA levels (P = 0.587). Pearson correlation analysis showed that the proportion of total dendritic cells (DCs) was positively correlated with IL-12 (r = 0.526, P = 0.003) and IL-23 (r = 0.501, P = 0.005) in ITP patients. Th1/Th2 ratio, IFN-γ, and IL-12 levels were negatively correlated with platelet counts (r = −0.494, P = 0.009; r = –0.415, P = 0.028; r = –0.492, P = 0.032). However, IL-23 was positively correlated with IL-17 (r = 0.489, P = 0.006) and negatively correlated with platelet count (r = –0.564, P = 0.001). The ratio of IL-6 and Th17 cells was negatively correlated with platelet count (r = –0.443, P = 0.014; r = –0.471, P = 0.011). The imbalance of pDC/mDC and the increase of IL-6, IL-12, and IL-23 lead to the increased differentiation of CD4+ T cells into Th1 and Th17 cells, which might be the important mechanisms underlying the imbalance of Th1/Th2 and Th17/Treg in ITP patients.     

Expansion of GARP-Expressing CD4+CD25−FoxP3+ T Cells and SATB1 Association with Activation and Coagulation in Immune Compromised HIV-1-Infected Individuals in South Africa

Although antiretroviral treatment lowers the burden of human immunodeficiency virus (HIV)-related disease, it does not always result in immunological recovery. This manifests as persistent chronic inflammation, immune activation or exhaustion that can promote the onset of co-morbidities. As the exact function of regulatory T (Treg) cells in HIV remains unclear, this cross-sectional study investigated three expression markers (Forkhead box protein P3 [FOXP3], glycoprotein A repetitions predominant [GARP], special AT-rich sequence binding protein 1 [SATB1]) and compared their expansion between CD4⁺CD25⁻ and CD4⁺CD25⁺⁺ T cells. Age-matched study subjects were recruited (Western Cape, South Africa) and sub-divided: HIV-negative subjects (n = 12), HIV-positive naïve treated (n = 22), HIV-positive treated based on CD4 count cells/µL (CD4 > 500 and CD4 < 500) (n = 34) and HIV-treated based on viral load (VL) copies/mL (VL < 1000 and VL > 1000) (n = 34). Markers of immune activation (CD38) and coagulation (CD142) on T cells (CD8) were assessed by flow cytometry together with FOXP3, GARP and SATB1 expression on CD4⁺CD25⁻ and CD4⁺CD25⁺⁺ T cells. Plasma levels of interleukin-10 (IL-10; anti-inflammatory marker), IL-6 (inflammatory marker) and D-dimer (coagulation marker) were assessed. This study revealed three major findings in immuno-compromised patients with virological failure (CD4 < 500; VL > 1000): (1) the expansion of the unconventional Treg cell subset (CD4⁺CD25⁻FOXP3⁺) is linked with disease progression markers; (2) increased GARP expression in the CD4⁺CD25⁻ and CD4⁺CD25⁺⁺ subsets; and (3) the identification of a strong link between CD4⁺CD25⁻SATB1⁺ cells and markers of immune activation (CD8⁺CD38⁺) and coagulation (CD8⁺CD142⁺ and D-dimer).Supplementary InformationThe online version contains supplementary material available at 10.1007/s12250-021-00386-8.     

Association of CMV-Specific T Cell-Mediated Immunity with CMV DNAemia and Development of CMV Disease in HIV-1–Infected Individuals

Background

Among HIV-1–infected individuals, cytomegalovirus (CMV) reactivation and disease occur in the setting of advanced immunosuppression. The value of a standardized assessment of CMV-specific T-cell mediated immunity by the CMV QuantiFERON assay (CMV-QFT) has not yet been thoroughly investigated in HIV-1–infected subjects.

Methods

Prospective, longitudinal study in 153 HIV-1–infected subjects with a CD4⁺ T cell count < 350/μL who simultaneously underwent CMV-QFT, CMV serology testing and CMV-DNA quantification. Factors associated with CMV-QFT were evaluated. Clinical screening for CMV manifestations was then performed every 3 months.

Results

Among the 141 CMV IgG-seropositive individuals the CMV-QFT assay yielded reactive results in 84% (118/141), negative results in 15% (21/141) and indeterminate (negative mitogen IFN-gamma response) results in 1% (2/141) of subjects. The mean actual CD4⁺ T cell count was significantly higher in CMV-QFT reactive subjects, when compared to CMV-QFT non-reactive individuals (183 ± 102 vs. 126 ± 104 cells/μL, P = 0.015). A significantly lower proportion of CMV-QFT reactive vs. non-reactive patients displayed CMV DNAemia > 100 copies/mL (23% (27/118) vs. 48% (11/23), P = 0.02). Furthermore, a statistically significant inverse association between mitogen IFN-gamma response and CMV-DNAemia > 1000 copies/mL was observed (P < 0.001). During the observational period, 5 CMV end-organ manifestations were observed. In three of the CMV cases the CMV-QFT yielded indeterminate results.

Conclusions

While CMV-QFT reactivity indicates CMV-specific immunity, indeterminate results due to negative mitogen IFN-gamma response might reflect HIV-1-induced immunodeficiency. Thus, dependency upon CD4⁺ T cell count should be considered when interpreting CMV-QFT results.     

Bone Marrow Alterations and Lower Endothelial Progenitor Cell Numbers in Critical Limb Ischemia Patients

Background

Critical limb ischemia (CLI) is characterized by lower extremity artery obstruction and a largely unexplained impaired ischemic neovascularization response. Bone marrow (BM) derived endothelial progenitor cells (EPC) contribute to neovascularization. We hypothesize that reduced levels and function of circulating progenitor cells and alterations in the BM contribute to impaired neovascularization in CLI.

Methods

Levels of primitive (CD34⁺ and CD133⁺) progenitors and CD34⁺KDR⁺ EPC were analyzed using flow cytometry in blood and BM from 101 CLI patients in the JUVENTAS-trial ({"type":"clinical-trial","attrs":{"text":"NCT00371371","term_id":"NCT00371371"}}NCT00371371) and healthy controls. Blood levels of markers for endothelial injury (sE-selectin, sICAM-1, sVCAM-1, and thrombomodulin), and progenitor cell mobilizing and inflammatory factors were assessed by conventional and multiplex ELISA. BM levels and activity of the EPC mobilizing protease MMP-9 were assessed by ELISA and zymography. Circulating angiogenic cells (CAC) were cultured and their paracrine function was assessed.

Results

Endothelial injury markers were higher in CLI (P<0.01). CLI patients had higher levels of VEGF, SDF-1α, SCF, G-CSF (P<0.05) and of IL-6, IL-8 and IP-10 (P<0.05). Circulating EPC and BM CD34⁺ cells (P<0.05), lymphocytic expression of CXCR4 and CD26 in BM (P<0.05), and BM levels and activity of MMP-9 (P<0.01) were lower in CLI. Multivariate regression analysis showed an inverse association between IL-6 and BM CD34⁺ cell levels (P = 0.007). CAC from CLI patients had reduced paracrine function (P<0.0001).

Conclusion

CLI patients have reduced levels of circulating EPC, despite profound endothelial injury and an EPC mobilizing response. Moreover, CLI patients have lower BM CD34⁺-cell levels, which were inversely associated with the inflammatory marker IL-6, and lower BM MMP-9 levels and activity. The results of this study suggest that inflammation-induced BM exhaustion and a disturbed progenitor cell mobilization response due to reduced levels and activity of MMP-9 in the BM and alterations in the SDF-1α/CXCR4 interaction contribute to the attenuated neovascularization in CLI patients.     

A Higher Frequency of Circulating IL-22+CD4+ T Cells in Chinese Patients with Newly Diagnosed Hashimoto’s Thyroiditis

Background

IL-22 and IL-17A are implicated in the pathogenesis of autoimmune diseases. However, the role of IL-22⁺ and IL-17A⁺ CD4⁺ T cells in the pathogenesis of Hashimoto’s thyroiditis (HT) is not fully understood. This study investigates serum IL-22 and IL-17A levels and determines the frequency of circulating IL-22⁺ CD4⁺ T cells in HT patients to understand their roles in the pathogenesis of HT.

Methods

The levels of serum IL-22, IL-17A and IFN-γ and the frequency of circulating IL-22⁺CD4⁺ and IL-17A⁺CD4⁺ T cells in 17 HT patients and 17 healthy controls (HC) were determined by enzyme-linked immunosorbent assay (ELISA) and flow cytometry. The levels of serum free triiodothyronine (FT4), free thyroxine (FT3), thyroid stimulating hormone (TSH), anti-thyroid peroxidase (TPO) and anti-thyroglobulin antibodies (TgAb) by chemiluminescent enzyme immunoassay and radioimmunoassay.

Results

The percentages of circulating IL-22⁺CD4⁺ and IL-17⁺CD4⁺ T cells (p<0.0001, p<0.0001) and the levels of serum IL-22, IL-17A and IFN-γ (p<0.0001, p<0.0001, p = 0.0210) in the HT patients were significantly higher than that in the HC. The percentages of IL-22⁺CD4⁺ T cells were positively correlated with Th17 cells (r = 0.8815, p<0.0001) and IL-17A⁺IL-22⁺CD4⁺ T cells (r = 0.8914, p<0.0001), but were negatively correlated with Th1 cells (r = −0.6110, p<0.0092) in the HT patients. The percentages of Th22 cells, Th17 cells and IL-17A⁺IL-22⁺CD4⁺ T cells were negatively correlated with the levels of serum TSH in the HT patients (r = −0.8402, p<0.0001; r = −0.8589, p<0.0001; r = −0.8289 p<0.0001, respectively).

Conclusions

A higher frequency of circulating IL-22⁺CD4⁺ and IL-17A⁺CD4⁺ T cells may be associated with the development of HT in Chinese patients.     

Biochemical and haematological changes in HIV subjects receiving winniecure antiretroviral drug in Nigeria

Background

Hematological and biochemical abnormalities are among the most common clinicopathological manifestations of HIV patients on ART. Consequently, the development and assessment of indigenous antiretroviral drugs with minimal abnormalities becomes a necessity. The objective of this investigation was to assess potential haematological and biochemical abnormalities that may be associated with the administration of Winniecure ART in HIV patients undergoing treatment in Nigeria. Fifty (50) confirmed HIV positive ART naïve patients aged 36 ± 10 were observed for haematological and biochemical responses for 12 weeks. Haematological responses were assessed thrice at 6 weeks interval using coulter Ac-T differential analyser and biochemical indicators (bilirubin, creatine, urea, amylase, ALT, ALP, AST, albumin) assayed spectrophotometrically.

Results

The biochemical parameters ALP (P < 0.05), ALT (P < 0.0001), AST (P < 0.001) and amylase (P < 0.05) slightly increased at the 12^th week, no significant change was observed in plasma creatinine and urea concentrations while albumin levels decreased non-significantly (P > 0.002). Haematological results showed consistent reduction of ESR, eosinophil, absolute and differential lymphocytes, granulocytes and total WBC in the test subjects throughout the assessment period. Conversely, haemoglobin, platelet and PCV increased significantly (P < 0.05). At the 12^th week thrombocytopenia (10.30%) and anaemia (76%) were reduced to 2% and 31% respectively while neutropenia (4.2 to 8%), leucopenia (26.8 to 30%) and lymphopenia (1 to 10%) increased. No cases of neutrophilia, lymphocytosis, eosinophilia and leukocytosis was observed.

Conclusion

The drug has a reduced haematological abnormalities and normal kidney function was unaffected though there were signs of possible abnormal levels of hepatic enzymes beyond 12 weeks of treatment.     

Intestinal-Specific TNFα Overexpression Induces Crohn’s-Like Ileitis in Mice

Background and Aim

Human and animal studies have clearly established tumor necrosis factor (TNF)α as an important mediator of Crohn’s disease pathogenesis. However, whether systemic or only local TNFα overproduction is required for the development of chronic intestinal inflammation and Crohn’s disease remains unclear. The aim of this study was to assess the contribution of intestinal epithelial-derived TNFα to the development of murine Crohn’s-like ileitis.

Methods

We adapted the well-established TNF^∆ARE/+ mouse model of Crohn’s disease (which systemically overexpresses TNFα) to generate a homozygous mutant strain that overexpress TNFα only within the intestinal epithelium. Intestinal-specific TNF^{i∆ARE/i∆ARE} mice were examined for histopathological signs of gut inflammation and extraintestinal manifestations of Crohn’s disease. The mucosal immune phenotype was characterized, and the contribution of specific lymphocyte populations to the pathogenesis of TNF^{i∆ARE/i∆ARE} ileitis was assessed.

Results

TNF^{i∆ARE/i∆ARE} mice had increased mucosal and systemic TNFα levels compared to wild-type controls (P<0.001), as well as severe chronic ileitis with increased neutrophil infiltration and villous distortion, but no extraintestinal manifestations (P<0.001 vs. wild-type controls). The gut mucosal lymphocytic compartment was also expanded in TNF^{i∆ARE/i∆ARE} mice (P<0.05), consisting of activated CD69⁺ and CD4⁺CD62L^- lymphocytes (P<0.05). FasL expression was significantly elevated in the mesenteric lymph nodes of TNF^{i∆ARE/i∆ARE} mice (P<0.05). Adoptive transfer of mucosal TNF^{i∆ARE/i∆ARE} lymphocytes resulted in ileitis in immunologically naïve severe combined immunodeficiency recipients (P<0.05 vs. wild-type controls), indicating an effector phenotype that was associated with increased production of both Th1 (IFNγ) and Th2 (IL-5, IL-13) cytokines.

Conclusion

Intestinal epithelial-derived TNFα is sufficient for the induction of Crohn’s-like ileitis, but not for the occurrence of extraintestinal manifestations, in TNF^{i∆ARE/i∆ARE} mice. These effects were associated with generation of effector lymphocytes within the intestinal mucosa and dysregulated apoptosis. Thus, targeted intestinal blockade of TNFα may provide an effective means to neutralize gut-derived TNFα with reduced side effects.     

Hierarchy Low CD4+/CD8+ T-Cell Counts and IFN-γ Responses in HIV-1+ Individuals Correlate with Active TB and/or M.tb Co-Infection

Objective

Detailed studies of correlation between HIV-M.tb co-infection and hierarchy declines of CD8+/CD4+ T-cell counts and IFN-γ responses have not been done. We conducted case-control studies to address this issue.

Methods

164 HIV-1-infected individuals comprised of HIV-1⁺ATB, HIV-1⁺LTB and HIV-1⁺TB^- groups were evaluated. Immune phenotyping and complete blood count (CBC) were employed to measure CD4+ and CD8+ T-cell counts; T.SPOT.TB and intracellular cytokine staining (ICS) were utilized to detect ESAT6, CFP10 or PPD-specific IFN-γ responses.

Results

There were significant differences in median CD4+ T-cell counts between HIV-1⁺ATB (164/μL), HIV-1⁺LTB (447/μL) and HIV-1⁺TB^- (329/μL) groups. Hierarchy low CD4+ T-cell counts (<200/μL, 200-500/μL, >500/μL) were correlated significantly with active TB but not M.tb co-infection. Interestingly, hierarchy low CD8+ T-cell counts were not only associated significantly with active TB but also with M.tb co-infection (P<0.001). Immunologically, HIV-1⁺ATB group showed significantly lower numbers of ESAT-6-/CFP-10-specific IFN-γ+ T cells than HIV-1⁺LTB group. Consistently, PPD-specific IFN-γ+CD4+/CD8+ T effector cells in HIV-1⁺ATB group were significantly lower than those in HIV-1⁺LTB group (P<0.001).

Conclusions

Hierarchy low CD8+ T-cell counts and effector function in HIV-1-infected individuals are correlated with both M.tb co-infection and active TB. Hierarchy low CD4+ T-cell counts and Th1 effector function in HIV-1+ individuals are associated with increased frequencies of active TB, but not M.tb co-infection.     

Allergic airway inflammation induces the migration of dendritic cells into airway sensory ganglia

Background

A neuroimmune crosstalk between dendritic cells (DCs) and airway nerves in the lung has recently been reported. However, the presence of DCs in airway sensory ganglia under normal and allergic conditions has not been explored so far. Therefore, this study aims to investigate the localisation, distribution and proliferation of DCs in airway sensory ganglia under allergic airway inflammation.

Methods

Using the house dust mite (HDM) model for allergic airway inflammation BALB/c mice were exposed to HDM extract intranasally (25 μg/50 μl) for 5 consecutive days a week over 7 weeks. With the help of the immunohistochemistry, vagal jugular-nodose ganglia complex (JNC) sections were analysed regarding their expression of DC-markers (MHC II, CD11c, CD103), the neuronal marker PGP 9.5 and the neuropeptide calcitonin gene-related peptide (CGRP) and glutamine synthetase (GS) as a marker for satellite glia cells (SGCs). To address the original source of DCs in sensory ganglia, a proliferation experiment was also carried in this study.

Results

Immune cells with characteristic DC-phenotype were found to be closely located to SGCs and vagal sensory neurons under physiological conditions. The percentage of DCs in relation to neurons was significantly increased by allergic airway inflammation in comparison to the controls (HDM 51.38 ± 2.38% vs. control 28.16 ± 2.86%, p < 0.001). The present study also demonstrated that DCs were shown to proliferate in jugular-nodose ganglia, however, the proliferation rate of DCs is not significantly changed in the two treated animal groups (proliferating DCs/ total DCs: HDM 0.89 ± 0.38%, vs. control 1.19 ± 0.54%, p = 0.68). Also, increased number of CGRP-positive neurons was found in JNC after allergic sensitisation and challenge (HDM 31.16 ± 5.41% vs. control 7.16 ± 1.53%, p < 0.001).

Conclusion

The present findings suggest that DCs may migrate from outside into the ganglia to interact with sensory neurons enhancing or protecting the allergic airway inflammation. The increase of DCs as well as CGRP-positive neurons in airway ganglia by allergic airway inflammation indicate that intraganglionic DCs and neurons expressing CGRP may contribute to the pathogenesis of bronchial asthma. To understand this neuroimmune interaction in allergic airway inflammation further functional experiments should be carried out in future studies.     

Acquired defects in CFTR-dependent β-adrenergic sweat secretion in chronic obstructive pulmonary disease

Rationale

Smoking-induced chronic obstructive pulmonary disease (COPD) is associated with acquired systemic cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction. Recently, sweat evaporimetry has been shown to efficiently measure β-adrenergic sweat rate and specifically quantify CFTR function in the secretory coil of the sweat gland.

Objectives

To evaluate the presence and severity of systemic CFTR dysfunction in smoking-related lung disease using sweat evaporimetry to determine CFTR-dependent sweat rate.

Methods

We recruited a cohort of patients consisting of healthy never smokers (N = 18), healthy smokers (12), COPD smokers (25), and COPD former smokers (12) and measured β-adrenergic sweat secretion rate with evaporative water loss, sweat chloride, and clinical data (spirometry and symptom questionnaires).

Measurements and main results

β-adrenergic sweat rate was reduced in COPD smokers (41.9 ± 3.4, P < 0.05, ± SEM) and COPD former smokers (39.0 ± 5.4, P < 0.05) compared to healthy controls (53.6 ± 3.4). Similarly, sweat chloride was significantly greater in COPD smokers (32.8 ± 3.3, P < 0.01) and COPD former smokers (37.8 ± 6.0, P < 0.01) vs. healthy controls (19.1 ± 2.5). Univariate analysis revealed a significant association between β-adrenergic sweat rate and female gender (β = 0.26), age (−0.28), FEV₁% (0.35), dyspnea (−0.3), and history of smoking (−0.27; each P < 0.05). Stepwise multivariate regression included gender (0.39) and COPD (−0.43) in the final model (R² = 0.266, P < 0.0001).

Conclusions

β-adrenergic sweat rate was significantly reduced in COPD patients, regardless of smoking status, reflecting acquired CFTR dysfunction and abnormal gland secretion in the skin that can persist despite smoking cessation. β-adrenergic sweat rate and sweat chloride are associated with COPD severity and clinical symptoms, supporting the hypothesis that CFTR decrements have a causative role in COPD pathogenesis.     

Olaparib attenuates sepsis-induced acute multiple organ injury via ERK-mediated CD14 expression

Sepsis is characterized by persistent systemic inflammation, which can cause multi-organ dysfunction. The poly polymerase-1 inhibitor olaparib possesses anti-inflammatory properties. This study aimed to assess the effects of olaparib (pre- and post-treatments) on sepsis, and to investigate whether it could suppress CD14 expression via the ERK pathway in polymicrobial sepsis and peritoneal macrophages models. Sepsis was induced by cecal ligation and puncture in C57BL/6 male mice. Fifty mice were randomly divided into five groups: The sham group was treated with vehicle or olaparib, the cecal ligation and puncture group with vehicle or with olaparib (5 mg/kg i.p.) 1 h before or 2 h after surgery. Olaparib pretreatment significantly improved the survival of septic mice (P < 0.001). Pre- and post-treatment of mice with olaparib partly alleviated cecal ligation and puncture-induced organ injury by decreasing the amounts of the pro-inflammatory mediators TNF-α and IL-6 as well as bacterial burden in the serum, peritoneal lavage fluid, and organs (P < 0.05). The protective effect of olaparib was associated with CD14 suppression via inhibition of ERK activation. Olaparib facilitated negative regulation of ERK-mediated CD14 expression, which may contribute to multi-organ injury in sepsis.     

Increased oxidative phosphorylation in lymphocytes does not atone for decreased cell numbers after burn injury

The acute systemic inflammatory response syndrome (SIRS) and multiorgan dysfunction (MOD) that occur in large burn injuries may be attributed, in part, to immunosuppressive responses such as decreased lymphocytes. However, the mitochondrial bioenergetics of lymphocytes after severe burn injury are poorly understood. The purpose of this study was to examine mitochondrial function of lymphocytes following severe burns in a swine model. Anesthetized Yorkshire swine (n = 17) sustained 40% total body surface area full-thickness contact burns. Blood was collected at pre-injury (Baseline; BL) and at 24 and 48 h after injury for complete blood cell analysis, flow cytometry, cytokine analysis, and ficoll separation of intact lymphocytes for high-resolution mitochondrial respirometry analysis. While neutrophil numbers increased, a concomitant decrease was found in lymphocytes (P < 0.001) after burn injury, which was not specific to CD4⁺ or CD8⁺ lymphocytes. No changes in immune cell population were observed from 24 h to 48 h post-injury. IL 12-23 decreased while a transient increase in IL 4 was found from BL to 24h (P < 0.05). CRP progressively increased from BL to 24h (P < 0.05) and 48h (P < 0.001) post-injury. Routine and maximal mitochondrial respiration progressively increased from BL to 24h (P < 0.05) and 48 h post-injury (P < 0.001). No changes were found in leak respiration or residual oxygen consumption. When considering the reduction in lymphocyte number, the total peripheral lymphocyte bioenergetics per volume of blood significantly decreased from BL to 24h and 48h (P < 0.05). For the first time, we were able to measure mitochondrial activity in intact lymphocyte mitochondria through high-resolution respirometry in a severely burned swine model. Our data showed that the non-specific reduction in peripheral T cells after injury was larger than the increased mitochondrial activity in those cells, which may be a compensatory mechanism for the total reduction in lymphocytes. Additional studies in the metabolic activation of T cell subpopulations may provide diagnostic or therapeutic targets after severe burn injury.     

A One Year Follow-Up Study of Natural Killer and Dendritic Cells Activities in Multiple Sclerosis Patients Receiving Glatiramer Acetate (GA)

Background

Multiple sclerosis (MS) is a chronic inflammatory, demyelinating and neurodegenerative disease. It is thought to be mediated by CD4⁺ Th1/Th17 cells. More recently, cells of the innate immune system such as dendritic cells (DCs) and natural killer (NK) cells have been in focus. Glatiramer acetate (GA) is an approved drug for treating MS patients.

Methodology/Principal Findings

In the current study we examined the activities of NK and DCs in nine relapsing remitting MS patients for up to one year after initiation of GA treatment. We observed that NK cells isolated from most of these patients have increased cytotoxic activity against K562 cells. Further analysis showed that the same NK cells lysed both autologous immature (i) and mature (m) DCs. In most patients this increased activity was correlated with increased NK cell activating cytotoxicity receptors such as NKp30, NKp44, NKp46 and NKG2D, and reduced expression of the inhibitory molecule CD158 on the surface of these NK cells. The expression of HLA-DR was increased on iDCs and mDCs in the majority of the patients, but no consistency was observed for the expression of HLA-I or HLA-E. Also, the co-stimulatory receptors CD80, CD83 or CD86 expression was down-regulated on iDCs and mDCs in most cases. Further, the expression of CCR6 was increased on mDCs at later time points of therapy (between 32–48 weeks).

Conclusions/Significance

Our results are the first showing the effects of GA treatment on NK cells in MS patients, which may impact future use of this and other drugs to treat this disease.