BACKGROUND

Asthma exacerbations contribute to significant morbidity, mortality and healthcare utilization. Furthermore, viral infections are associated with asthma exacerbations by mechanisms that are not fully understood.

OBJECTIVE

The aim of this analysis was to determine whether cytokine patterns in patients with colds could identify risks for subsequent asthma exacerbations.

METHODS

We analysed cytokine levels in nasal lavage fluid (NLF) in 59 subjects (46 with asthma) with acute upper respiratory symptoms and after symptomatic resolution. Analyte choice was based on potential relevance to asthma exacerbations: antiviral (IFN-α, IFN-β, IFN-γ, IFN-λ1, IP-10, TRAIL), cell recruiting (G-CSF, IL-1β, IL-8, MCP-1, MCP-3, TNF-α), polarizing (CXCL13, IL-10, IL-13, IL-17, TSLP), and injury remodelling (fibronectin, IL-33, MMP-9, VEGF).

RESULTS

The overall cytokine response induced during viral infections was not different between asthmatic and non-asthmatic individuals for a wide array of cytokines. However, mean levels of VEGF, TNF-α and IL-1β were 1.7-, 5.1- and 4.7-fold higher in samples from asthma subjects who exacerbated in the first 3 weeks of the cold compared with those who did not exacerbate (P = 0.006, 0.01, 0.048, respectively). Using receiver operating characteristic curve-defined thresholds, high VEGF and TNF-α levels predicted a shorter time-to-exacerbation after NLF sampling (25% exacerbation rate: 3 vs. 45 days, and 3 vs. 26 days; P = 0.03, 0.04, respectively).

CONCLUSIONS

Although they produce similar cytokine responses to viral infection as non-asthmatics, asthmatics with higher levels of VEGF and TNF-α in NLF obtained during acute cold phases predicted subsequent asthma exacerbations in this cohort of patients with mild-to-moderate disease. In the future, stratifying the risk of an asthma exacerbation by cytokine profile may aid the targeting of personalized treatment and intervention strategies.

The aim of present study was to characterize long non-coding RNA (lncRNA) AK141205 as a cellular regulator of osteogenic differentiation of mice mesenchymal stem cells (MSCs) towards osteogenic growth peptide (OGP) stimulation. Mice MSCs cells were isolated, transfected with si-AK141205, pcDNA-AK141205 or control, and stimulated with OGP. The AK141205, CXC chemokine ligand-13 (CXCL13), and osteogenic differentiation-associated parameters were determined by western blotting or quantitative RT-PCR. To determine the role of AK141205/CXCL13 in SMCs osteogenic differentiation, SMCs subjected to co-transfection of pcDNA-AK141205 and si-CXCL13 or si-AK141205 and pcDNA-CXCL13, and were submitted for osteogenic differentiation-associated parameters analyses. The results showed that stimulation of SMCs with OGP induced upregulation of both AK141205 and CXCL13, and osteogenic differentiation of MSCs. Transfection of si-AK141205 partly suppressed OGP-induced formation of calcium salt nodules, alkaline phosphatase (ALP) activity and osteogenic differentiation-associated gene expression, suggesting key regulatory role of AK141205. Analysis of CXCL13 expression in SMCs(pcDNA-AK141205) revealed that AK141205 positively promoted CXCL13 expression via acetylation of H4 histone in the promoter region. This signal transduction was demonstrated to be essential for OGP-induced osteogenic differentiation of MSCs through osteogenic differentiation analysis in simultaneously AK141205/CXCL13 controlled SMCs. In summary, we report a completely novel role of AK141205/CXCL13 as a regulator of OGP-induced osteogenic differentiation of SMCs. Our finding provides a potential therapeutic targeting of AK141205 for enhancing disease-treatment effect of SMCs.

DNA vaccination is a promising method to induce specific immune responses. However, it remains challenging to enhance DNA vaccine potency. Chemokines were used as adjuvants to improve the efficacy of DNA vaccination. Herein, we fused murine chemokine CCL20 or CXCL13 to a model antigen green fluorescent protein (GFP). The fusion DNA vaccines enhanced specific anti-GFP immune responses in mice compared with vector pEGFP-N1. Co-immunization with both of chemokine-GFP fusion constructs induced the significantly highest level of humoral immune responses. CCL20-GFP or CXCL13-GFP fusion DNA vaccine induced predominant IgG2a or IgG1 response respectively. However, co-immunization with both of these fusion DNA vaccines induced a predominant IgG2a response. Therefore, fusing chemokine CXCL13 or CCL20 to antigen provides new attractive strategy to enhance the immunogenicity of DNA vaccine and modulate immune responses.

OBJECTIVE

To evaluate gene expression by microarray analyses of inflammatory mediators in the sinus mucosa of children with and without chronic rhinosinusitis (CRS).

METHODS

Prospective molecular genetics analysis.

METHODS

Children's National Medical Center, Washington, DC.

METHODS

Eleven patients with CRS who underwent endoscopic sinus surgery and 10 control children who underwent craniofacial resection or neurosurgical procedures.

METHODS

Gene expression levels of sinus tissue from 6 patients with CRS and 6 controls and messenger RNA expression levels of upregulated inflammatory/immune response genes, as well as cytokines of interest, determined by quantitative reverse transcription-polymerase chain reaction.

RESULTS

Gene expression using the Plier algorithm yielded the most consistent grouping of samples: 96 genes were significantly upregulated more than 2-fold, and 123 genes were downregulated by at least 50% in the CRS sinus tissues compared with controls (P < .05). GeneSpring analysis demonstrated significant changes in several ontology categories in the CRS samples, including inflammatory/immune response genes. The chemokines CXCL13 and CXCL5, serum amyloid A, serpin B4, and defensin beta1 were highly upregulated >> or =5-fold). Increased expression of these genes was validated by quantitative reverse transcription-polymerase chain reaction in an independent set of tissues. Expression levels of interleukins 5, 6, and 8 were similar in both cohorts; these results were validated by reverse transcription-polymerase chain reaction.

CONCLUSIONS

Microarray analyses of sinus mucosa in children with CRS showed an increased expression of inflammatory genes involved in innate and adaptive immune systems. This technology can be successfully used to identify genes implicated in the pathogenesis of pediatric CRS.

Chemokine-directed leukocyte migration is a critical component of all innate and adaptive immune responses. The atypical chemokine receptor ACKR2 is expressed by lymphatic endothelial cells and scavenges pro-inflammatory CC chemokines to indirectly subdue leukocyte migration. This contributes to the resolution of acute inflammatory responses in vivo. ACKR2 is also universally expressed by innate-like B cells, suppressing their responsiveness to the non-ACKR2 ligand CXCL13, and controlling their distribution in vivo. The role of ACKR2 in autoimmunity remains relatively unexplored, although Ackr2 deficiency reportedly lessens the clinical symptoms of experimental autoimmune encephalomyelitis induced by immunization with encephalogenic peptide (MOG(35-55)). This was attributed to poor T-cell priming stemming from the defective departure of dendritic cells from the site of immunization. However, we report here that Ackr2-deficient mice, on two separate genetic backgrounds, are not less susceptible to autoimmunity induced by immunization, and in some cases develop enhanced clinical symptoms. Moreover, ACKR2 deficiency does not suppress T-cell priming in response to encephalogenic peptide (MOG(35-55)), and responses to protein antigen (collagen or MOG(1-125)) are characterized by elevated interleukin-17 production. Interestingly, after immunization with protein, but not peptide, antigen, Ackr2 deficiency was also associated with an increase in lymph node B cells expressing granulocyte-macrophage colony-stimulating factor (GM-CSF), a cytokine that enhances T helper type 17 (Th17) cell development and survival. Thus, Ackr2 deficiency does not suppress autoreactive T-cell priming and autoimmune pathology, but can enhance T-cell polarization toward Th17 cells and increase the abundance of GM-CSF(+) B cells in lymph nodes draining the site of immunization.

OBJECTIVE

The homeostatic chemokine, B cell attracting chemokine 1 (CXCL13), has been implicated in the pathogenesis of lymphocyte-mediated diseases. We investigated the cellular expression of this chemokine in the spectrum of ocular adnexal lymphoproliferative lesions.

METHODS

Laboratory investigation.

METHODS

CXCL13 expression in paraffin-embedded adnexal biopsy specimens was determined by indirect immunohistochemistry with antigen retrieval.

RESULTS

In 15 of 16 biopsy specimens, including reactive lymphoid hyperplasia (n = 7), atypical lymphoid hyperplasia (n = 3), and B cell lymphoma (n = 6), CXCL13 was detected. CD20-positive B-cells, as well as dendritic cells and endothelial cells, expressed the chemokine.

CONCLUSIONS

B-cells in ocular adnexal lymphoproliferative lesions demonstrate expression of CXCL13, a chemokine that may participate in tumor pathogenesis and is a potential target for novel therapies.

Skin manifestations of angioimmunoblastic T-cell lymphoma (AITL) are frequent, sometimes as first manifestations of the disease. In the absence of a specific marker for neoplastic cells, diagnosis of AITL in skin biopsies is often difficult. CD10 and CXCL13 have been recently recognized as characteristic markers of AITL, but have not been yet investigated in the skin. We analyzed 15 skin biopsies from 8 patients with AITL having skin manifestations and compared them to 14 skin biopsies from patients with various cutaneous lymphocytic infiltrates. A few CD10 lymphocytes were found in only 2 samples of the AITL group, the identification of which was hampered by the presence of a dermal CD10 cell population with dendritic features. By contrast, CXCL13 lymphoid cells were identified in most AITL cutaneous biopsies (n=12, 80%), whereas, absent in all samples from control cases. Among 12 biopsies with CXCL13 cells, cutaneous involvement by AITL was suspected in only 5 on the basis of light microscopy and classic immunophenotyping. In another case, a diagnosis of cutaneous marginal zone B-cell lymphoma had been proposed. In conclusion, this study shows that neoplastic AITL CXCL13 T cells localize in the skin and that accurate diagnosis of AITL lesions can be done in skin specimens using CXCL13 immunostaining on paraffin-embedded tissues.

Angioimmunoblastic T-cell lymphoma is immunologically defined by the expression of CD10 and the follicular helper T cell (T(FH)) markers such as CXCL13, programmed death-1 (PD-1) and inducible T-cell costimulator (ICOS). This T(FH) profile has been mainly reported by immunohistochemistry. Here, using multiparametric flow cytometry, the relevance of ICOS and PD-1 to angioimmunoblastic T-cell lymphoma diagnosis was evaluated in lymph node (n=15) as well as in peripheral blood (n=13) among a series of 28 angioimmunoblastic T-cell lymphoma cases, in addition to the CD10 expression (available in 26 lymph node and 15 peripheral blood specimens). In this series, CD10 expression was present in 23/26 (88%) lymph node and in 12/15 (80%) peripheral blood cases and ICOS in 13/15 (87%) lymph node and in 6/13 (47%) peripheral blood cases, whereas neither significant CD10 nor ICOS T cells were identified in the control group (lymph nodes with reactive hyperplasia=10, peripheral blood of healthy donors=15). PD-1 expression was less informative as observed in both angioimmunoblastic T-cell lymphoma and control cases. The multiparametric approach allowed us to confirm the frequent blood dissemination in angioimmunoblastic T-cell lymphoma and to show that circulating neoplastic T cells correspond more often to a CD10-positive subset than to an ICOS-positive subset. Consequently, if ICOS constitutes an additional feature for the diagnosis of angioimmunoblastic T-cell lymphoma, it appears less sensitive than CD10 expression for the detection of circulating neoplastic T cells.

Interferon gamma (IFN-γ) is a cytokine that plays a very important role in defining Th1 immune response in all vertebrates. In this study, recombinant IFN-γ (rIFN-γ) from the olive flounder (Paralichthys olivaceus) was produced in an Escherichia coli system using a pET expression vector. Stimulation of whole kidney leukocytes (immune-related cells) in vitro with the resulting rIFN-γ significantly induced the gene expression of interleukin-1β (IL-1β), signal transducer and activator of transcription 1 (STAT1), CXCL13-like chemokine (CXCL13), and IFN-γ. rIFN-γ also weakly induced the expression of IL-1β, tumor necrosis factor-α (TNF-α), CXCL13, and IFN-γ in olive flounder-derived HINAE (non-immune) cells. The effects of rIFN-γ against Edwardsiella tarda infection in vivo were assessed by intraperitoneally injecting a mixture of rIFN-γ (100 ng) and E. tarda (1 × 10(5) CFU/ml) into the olive flounder. The survival rate in the rIFN-γ-injected group was 60% compared to 0% in the group treated with E. tarda only, demonstrating that olive flounder IFN-γ is effective in reinforcing immune responses and preventing against edwardsiellosis.

Disease progression of neuropsychiatric symptoms in Lyme/tick-borne diseases can be better understood by greater attention to psychoimmunology. Although there are multiple contributors that provoke and weaken the immune system, infections and persistent infections are significant causes of pathological immune reactions. Immune mediated ef-fects are a significant contributor to the pathophysiological processes and disease progression. These immune effects in-clude persistent inflammation with cytokine effects and molecular mimicry and both of these mechanisms may be present at the same time in persistent infections. Sickness syndrome associated with interferon treatment and autoimmune limbic encephalopathies are models to understand inflammatory and molecular mimicry effects upon neuropsychiatric symp-toms. Progressive inflammatory reactions have been proposed as a model to explain disease progression in depression, psychosis, dementia, epilepsy, autism and other mental illnesses and pathophysiological changes have been associated with oxidative stress, excitotoxicity, changes in homocysteine metabolism and altered tryptophan catabolism. Lyme dis-ease has been associated with the proinflammatory cytokines IL-6, IL-8, IL-12, IL-18 and interferon-gamma, the chemokines CXCL12 and CXCL13 and increased levels proinflammatory lipoproteins. Borrelia burgdorferi surface gly-colipids and flagella antibodies appear to elicit anti-neuronal antibodies and anti-neuronal antibodies and Borrelia burgdorferi lipoproteins can disseminate from the periphery to inflame the brain. Autism spectrum disorders associated with Lyme/tick-borne diseases may be mediated by a combination of inflammatory and molecular mimicry mechanisms. Greater interaction is needed between infectious disease specialists, immunologists and psychiatrists to benefit from this awareness and to further understand these mechanisms.

UNASSIGNED

In patients with triple-negative breast cancer (TNBC), the extent of tumor-infiltrating lymphocytes (TILs) in the residual disease after neoadjuvant chemotherapy (NACT) is associated with better prognosis. Our objective was to develop a gene signature from pretreatment samples to predict the extent of TILs after NACT and then to test its prognostic value on survival.

UNASSIGNED

Using 99 pretreatment samples, we generated a four-gene signature associated with high post-NACT TILs. Prognostic value of the signature on distant relapse-free survival (DRFS) was first assessed on the training set (n = 99) and then on an independent validation set (n = 115).

UNASSIGNED

A four-gene signature combining the expression levels of HLF, CXCL13, SULT1E1, and GBP1 was developed in baseline samples to predict the extent of lymphocytic infiltration after NACT. In a multivariate analysis performed on the training set, this signature was associated with DRFS [hazard ratio (HR): 0.28, for a one-unit increase in the value of the four-gene signature, 95% confidence interval (CI): 0.13-0.63)]. In a multivariate analysis performed on an independent validation set, the four-gene signature was significantly associated with DRFS (HR: 0.17, 95% CI: 0.06-0.43). The four-gene signature added significant prognostic information when compared with the clinicopathologic pretreatment model (likelihood ratio test in the training set P = 0.004 and in the validation set P = 0.002).

UNASSIGNED

A four-gene signature predicts high levels of TILs after anthracycline-containing NACT and outcome in patients with TNBC and adds prognostic information to a clinicopathological model at diagnosis.

At present, diagnosis for neurosyphilis remains a major clinical challenge. Venereal Disease Research Laboratory (VDRL) titer of the cerebrospinal fluid (CSF) is suboptimally sensitive to diagnose neurosyphilis, which can be negative in neurosyphilis patients, especially in asymptomatic neurosyphilis patients. In the search for biomarkers of neurosyphilis, we investigated the chemokine profile in CSF of neurosyphilis patients and found that the concentrations of CXCL13, CXCL10 and CXCL8 were selectively elevated in neurosyphilis patients and correlated with CSF protein concentration and CSF-VDRL titer. After antibiotic treatment, the concentration of these chemokines was dramatically reduced. The area under the ROC curve (AUC) of CSF CXCL13, CXCL8,CXCL10 and the CSF/serum ratio of CXCL13, CXCL8,CXCL10 in the diagnosis of neurosyphilis were 0.940, 0.899, 0.915, 0.963, 0.846 and 0.926, respectively. The corresponding sensitivities/specificities of CSF CXCL13, CXCL8,CXCL10 and the CSF/serum ratio of CXCL13, CXCL8,CXCL10 in diagnosis of neurosyphilis were 85.4%/89.1%, 79%/90.1% and 79.6%/91.1%, 86.6%/99%, 79%/73.3% and 86%/92.1%, respectively. Our results suggest that the elevated concentrations of CXCL13, CXCL8, and CXCL10 or their increasing CSF/serum ratios may be potential biomarkers of neurosyphilis, particularly for asymptomatic neurosyphilis. Reduced concentration of these chemokines may indicate the prognosis of antibiotic therapy.

Development of lung cancer is closely related to smoking in a majority of patients. Most smokers, however, do not develop lung cancer in spite of a high mutational load accumulating in the lung tissue. Here we investigate whether a cancer-specific footprint can be revealed by investigating circulating inflammatory markers in patients with non-small cell lung cancer (NSCLC) compared with patients with chronic obstructive pulmonary disease (COPD), both cohorts characterised by similar smoking history. Serum concentrations of 57 cytokines and matrix metalloproteinases (MMPs) from 43 patients with advanced NSCLC were evaluated by multiplex immunoassays and compared with serum samples from 35 patients with COPD. Unsupervised hierarchical clustering and non-parametric analyses were performed. False discovery rate was used to adjust for multiple testing. Clustering of cytokine and MMP concentrations in the serum revealed a distinct separation of the NSCLC patients from the COPD group. Individual concentrations of thymus and activation-regulated cytokine (C-C motif chemokine ligand 17), Gro-b (C-X-C motif chemokine ligand 2 (CXCL2)), CXCL13, interleukin (IL)-1ra, IL-6, IL-8 (CXCL8), IL-16, IL-17A, macrophage migration inhibitory factor (MIF), granulocyte colony-stimulating factor, platelet-derived growth factor subunit B, MMP-2, MMP-8 and MMP-12 were significantly different in serum from NSCLC and COPD patients. Moreover, the interferon-γ/IL-10 ratio was lower in cancer patients compared with COPD patients, consistent with a cytokine milieu favouring tumour tolerance. Our results suggest that NSCLC is characterised by a distinct inflammatory signature in serum. The different cytokine profiles in NSCLC and COPD patients may represent tumour-promoting and tumour-suppressing immune responses developing in response to mucosal inflammation and mutations induced by smoking.

The treatment of drug-resistant cancer is a clinical challenge, and thus screening for novel anticancer drugs is critically important. We recently demonstrated a strong enhancement of the antitumor activity of snake (Walterinnesia aegyptia) venom (WEV) in vitro in breast carcinoma, prostate cancer, and multiple myeloma cell lines but not in normal cells when the venom was combined with silica nanoparticles (WEV+NP). In the present study, we investigated the in vivo therapeutic efficacy of WEV+NP in breast cancer- and prostate cancer-bearing experimental mouse models. Xenograft breast and prostate tumor mice models were randomized into 4 groups for each cancer model (10 mice per group) and were treated with vehicle (control), NP, WEV, or WEV+NP daily for 28 days post tumor inoculation. The tumor volumes were monitored throughout the experiment. On Day 28 post tumor inoculation, breast and prostate tumor cells were collected and either directly cultured for flow cytometry analysis or lysed for Western blot and ELISA analysis. Treatment with WEV+NP or WEV alone significantly reduced both breast and prostate tumor volumes compared to treatment with NP or vehicle alone. Compared to treatment with WEV alone, treatment of breast and prostate cancer cells with WEV+NP induced marked elevations in the levels of reactive oxygen species (ROS), hydroperoxides, and nitric oxide; robust reductions in the levels of the chemokines CXCL9, CXCL10, CXCL12, CXCL13, and CXCL16 and decreased surface expression of their cognate chemokine receptors CXCR3, CXCR4, CXCR5, and CXCR6; and subsequent reductions in the chemokine-dependent migration of both breast and prostate cancer cells. Furthermore, we found that WEV+NP strongly inhibited insulin-like growth factor 1 (IGF-1)- and epidermal growth factor (EGF)-mediated proliferation of breast and prostate cancer cells, respectively, and enhanced the induction of apoptosis by increasing the activity of caspase-3,-8, and -9 in both breast and prostate cancer cells. In addition, treatment of breast and prostate cancer cells with WEV+NP or WEV alone revealed that the combination of WEV with NP robustly decreased the phosphorylation of AKT, ERK, and IκBα; decreased the expression of cyclin D1, surviving, and the antiapoptotic Bcl-2 family members Bcl-2, Bcl-XL, and Mcl-1; markedly increased the expression of cyclin B1 and the proapoptotic Bcl-2 family members Bak, Bax, and Bim; altered the mitochondrial membrane potential; and subsequently sensitized tumor cells to growth arrest. Our data reveal the therapeutic potential of the nanoparticle-sustained delivery of snake venom against different cancer cell types.

Accumulating research indicates that B cells are involved in anti-tumor immunity. Chronic alcohol consumption is associated with decreased survival of cancer patients. The effect of alcohol consumption on B cells in tumor-bearing hosts is unknown. Results in melanoma-bearing mice showed that chronic alcohol consumption did not alter the percentage and number of B cells in bone marrow, spleen, and lymph nodes but dramatically decreased B cells in the peripheral blood. Alcohol consumption did not alter the development of B cells in the bone marrow and did not affect follicular B cells in the spleen; however, it increased T1 B cells and decreased marginal zone B cells in the spleen. Alcohol consumption also decreased mature B cells in the blood. It did not alter the chemotactic capacity of plasma to facilitate migration of splenocytes or the chemotactic response of splenocytes to CXCL13 and CCL21. However, the response of splenocytes to sphingosine-1-phosphate was impaired in alcohol-consuming, melanoma-bearing mice. The expression of sphingosine-1-phosphate receptor-1 (S1PR1) and sphingosine-1-phosphate lyase-1 (SPL1) in splenocytes was downregulated. Taken together, these results indicate that chronic alcohol consumption decreases peripheral blood B cells by compromising B cell egress from the spleen. The downregulation of S1PR1 and SPL1 expression in alcohol-consuming, melanoma-bearing mice could be associated with compromised egress of B cells from the spleen.

Elevated CXCL13 within the central nervous system (CNS) correlates with humoral responses in several neuroinflammatory diseases, yet its role is controversial. During coronavirus encephalomyelitis CXCL13 deficiency impaired CNS accumulation of memory B cells and antibody-secreting cells (ASC) but not naïve/early-activated B cells. However, despite diminished germinal center B cells and follicular helper T cells in draining lymph nodes, ASC in bone marrow and antiviral serum antibody were intact in the absence of CXCL13. The data demonstrate that CXCL13 is not essential in mounting effective peripheral humoral responses, but specifically promotes CNS accumulation of differentiated B cells.

BACKGROUND

Current guidelines regarding Lyme neuroborreliosis [LNB] require the presence of intrathecal Borrelia burgdorferi-specific antibody production for the definite diagnosis of LNB. However, about 20% of early stage infections present without an elevated antibody index. Moreover, intrathecal B. burgdorferi specific antibody synthesis may persist long after successful therapy of LNB. Recently published data indicate that CXCL13 seems to be a promising diagnostic tool for early stage LNB. In addition, CXCL13 might be suitable for treatment monitoring.

METHODS

We report on a 39-year-old male patient from southern Germany, who has been suffering from subfebrile body temperatures and meningeal headache for six weeks. On the second day after hospital admission he developed peripheral palsy of the VII. cranial nerve. Cerebrospinal fluid (CSF) analysis showed granulocytic pleocytosis, elevated total protein and blood-CSF barrier dysfunction. Differential diagnostics for granulocytic pleocytosis were unremarkable. Only a second lumbar puncture, on day 6 after admission, revealed a lymphocytic pleocytosis. Serologic testing pointed to clear intrathecal Borrelia specific IgG antibody production. Interestingly, no anti-OspC antibodies were detectable. DNA of the rare Borrelia garinii OspA-type 7 could be amplified from the first CSF sample. The monitoring of CXCL13 in all CSF samples documented a fast decrease from 5000 pg/ml to 450 pg/ml after appropriate antibiotic treatment.

CONCLUSIONS

CXCL13 is a novel biomarker with high sensitivity and specificity for acute LNB. Our data show, that CXCL13 might be helpful in unclear cases and support the presumption that it might be a valuable tool for treatment monitoring. Anti-OspC antibody negativity is a rare observation, given the need of OspC for infection of the human hosts. Most likely this is due to a lack of sensitivity of OspC immunoblots that are unable to detect rare OspC variants.

During brain development, the immune system mediates neurogenesis, gliogenesis and synapse formation. However, it remains unclear whether peripheral lymphocytes contribute to brain development. Here we identified the subtypes of lymphocytes that are present in neonatal mouse brains and investigated their functions. We found that B-1a cells, a subtype of B cells, were abundant in the neonatal mouse brain and infiltrated into the brain in a CXCL13-CXCR5-dependent manner. B-1a cells promoted the proliferation of oligodendrocyte-precursor cells (OPCs) in vitro, and depletion of B-1a cells from developing brains resulted in a reduction of numbers of OPCs and mature oligodendrocytes. Furthermore, neutralizing Fcα/μR, the receptor for the Fc region of IgM secreted by B-1a cells, inhibited OPC proliferation and reduced the proportion of myelinated axons in neonatal mouse brains. Our results demonstrate that B-1a cells infiltrate into the brain and contribute to oligodendrogenesis and myelination by promoting OPC proliferation via IgM-Fcα/μR signaling.

BACKGROUND

Chemokine CXC ligand 13 (CXCL13) has been implicated in perivascular inflammation and pulmonary vascular remodeling in patients with idiopathic pulmonary artery hypertension (IPAH). We wondered whether CXCL13 may also play a role in chronic thromboembolic pulmonary hypertension (CTEPH) and whether serum levels of CXCL13 might serve as biomarkers in these conditions.

METHODS

Lung tissue from patients with IPAH or CTEPH was immunostained for CXCL13. Serum samples were obtained from patients with IPAH (n = 42) or CTEPH (n = 50) and from healthy controls (n = 13). Serum CXCL13 concentrations were measured by enzyme-linked immunosorbent assay technology and were evaluated for associations with markers of disease severity and survival.

RESULTS

CXCL13 was expressed in pulmonary vascular lesions and lymphocytes of patients with IPAH and inoperable CTEPH, respectively. Serum CXCL13 was elevated in patients compared to healthy controls [median, interquartile range, 83 (55,114) pg/ml versus 40 (28, 48) pg/ml; p < 0.001]. Serum CXCL13 showed only weak and inconsistent correlations with markers of inflammation or disease severity. In both populations, patients with serum CXCL13 above the median of the respective groups did not have a higher risk of death than patients with lower serum CXCL13.

CONCLUSIONS

CXCL13 was overexpressed in pulmonary vascular lesions of patients with IPAH and CTEPH, and increased serum concentrations were found in patients with IPAH and CTEPH, suggesting a potential pathogenic role of CXCL13 in both diseases. However, given the weak associations between serum CXCL13 and markers of disease severity and outcome, CXCL13 is unlikely to become a promising biomarker in these patient populations.

In human inflammatory sites, PD-1hiCXCR5-CD4+ T cells are involved in the formation of ectopic lymphoid-like structures (ELSs) by the secretion of chemokine CXCL13, but how the transcription of CXCL13 is regulated in CD4+ T cells is still unclear. Here we show that Sox4 is a key transcription factor for CXCL13 production in human CD4+ T cells under inflammatory conditions. In vitro TGF-β+, IL-2-neutralizing culture conditions give rise to PD-1hiCXCR5-CD4+ T cells that preferentially express CXCL13, and transcriptome analysis and lentiviral overexpression indicate Sox4 association with the CXCL13 transcription. In vivo, Sox4 is significantly upregulated in synovial CD4+ T cells, when compared with blood CD4+ T cells, from patients with rheumatoid arthritis (RA), and further correlates with ELS formation in RA synovium. Overall, our studies suggest that Sox4 contributes to CXCL13 production and ELS formation at inflammatory sites in humans.

The chemokine receptor CXCR5 is primarily expressed on B cells and Tfh cells and facilitates their migration towards B cell follicles. In the present study we investigated the role of the CXCL13/CXCR5 axis in the pathogenesis of rheumatoid arthritis (RA) and specifically addressed the impact of CXCR5-mediated T and B cell migration in this disease. Employing collagen-induced arthritis (CIA) we identify CXCR5 as an absolutely essential factor for the induction of inflammatory autoimmune arthritis. Cxcr5-deficient mice and mice selectively lacking Cxcr5 on T cells were completely resistant to CIA, showed impaired germinal center responses and failed to mount an IgG1 antibody response to collagen II. Selective ablation of CXCR5 expression in B cells also led to suppression of CIA owing to diminished GC responses in secondary lymphoid organs (SLO) and impaired anti-collagen II antibody production. Chimeric mice harboring Cxcr5-proficient and Cxcr5-deficient immune cells revealed SLO and not the synovial tissue as the compartment where CXCR5-mediated cell migration induces autoimmune inflammation in arthritis. Thus our data demonstrate that CXCR5-mediated co-localization of Tfh cells and B cells in SLOs is absolutely essential for the induction of RA and identify CXCR5 and Tfh cells as promising therapeutic targets for the treatment of RA.

BACKGROUND

Sandhoff disease is a lysosomal storage disorder characterized by the absence of β-hexosaminidase and storage of GM2 ganglioside and related glycolipids. We have previously found that the progressive neurologic disease induced in Hexb(-/-) mice, an animal model for Sandhoff disease, is associated with the production of pathogenic anti-glycolipid autoantibodies.

RESULTS

In our current study, we report on the alterations in the thymus during the development of mild to severe progressive neurologic disease. The thymus from Hexb(-/-) mice of greater than 15 weeks of age showed a marked decrease in the percentage of immature CD4(+)/CD8(+) T cells and a significantly increased number of CD4(+)/CD8(-) T cells. During involution, the levels of both apoptotic thymic cells and IgG deposits to T cells were found to have increased, whilst swollen macrophages were prominently observed, particularly in the cortex. We employed cDNA microarray analysis to monitor gene expression during the involution process and found that genes associated with the immune responses were upregulated, particularly those expressed in macrophages. CXCL13 was one of these upregulated genes and is expressed specifically in the thymus. B1 cells were also found to have increased in the thy mus. It is significant that these alterations in the thymus were reduced in FcRγ additionally disrupted Hexb(-/-) mice.

CONCLUSIONS

These results suggest that the FcRγ chain may render the usually poorly immunogenic thymus into an organ prone to autoimmune responses, including the chemotaxis of B1 cells toward CXCL13.

Both T cells and B cells are implicated in the pathology of multiple sclerosis (MS), but how these cells cooperate to drive disease remains unclear. Recent studies using experimental autoimmune encephalomyelitis (EAE) demonstrated that the TH17 pathway is correlated with increased numbers of ectopic B-cell follicles in the central nervous system (CNS). As follicular T helper (TFH) cells are regulators of B cell responses, we sought to examine the role of TFH cells in EAE induced by the transfer of myelin-specific TH17 cells (TH17-EAE). In this study, we first confirmed previous reports that B-cells are a major cell type infiltrating the CNS during TH17-EAE. In addition, we found that B cells contribute to the severity of TH17-EAE. Class-switched B-cells in the CNS were positively correlated with disease and, strikingly, the severity TH17-EAE was diminished in B cell deficient mice. We next focused on the role TFH cells play in TH17-EAE. We found substantial numbers of CXCR5+PD1+CD4+ TFH cells in the CNS tissue of TH17-EAE mice and that at the peak of disease, the number of infiltrating TFHs was correlated with the number of infiltrating B-cells. Using congenic CD45.1+ donor mice and CD45.2+ recipient mice, we determined that the TFH cells were recipient-derived, whereas IL-17+ cells were donor-derived. We assessed whether myelin-specific TFH cells are capable of inducing EAE in recipient mice and found that transferring TFH cells failed to induce EAE. Finally, we tested the effects of blocking TFH trafficking in TH17-EAE using an antagonistic antibody against CXCL13, the chemokine ligand for CXCR5 on TFH cells. We found anti-CXCL13 treatment significantly reduced TH17-EAE disease. This treatment blocked CD4+ T cells from entering the CNS, but had no effect on infiltration of B cells. Strikingly, this antibody treatment had no measurable effect on TH17 disease in B cell-deficient mice. These data demonstrate that infiltrating TFH cells are a key cell type that contributes to an inflammatory B cell response in TH17-EAE and provide evidence for targeting TFH cells as a treatment for neuro-autoimmune diseases like MS.