Highlights • CCR6+ T helper (Th) cells dominate the cerebrospinal fluid (CSF) in MS. • CCR6+ Th17 cells are very rare. • The dominant CCR6+ T cell subset secretes IFNγ and is elevated in MS CSF. • ...The encephalitogenic cytokine GM-CSF is also secreted by these cells. • GM-CSF-only secreting Th cells, also accumulate in the CSF especially in MS.
Abstract Cerebrospinal fluid (CSF) analysis is routinely used in the diagnostic work-up of multiple sclerosis (MS), by detecting CSF-specific oligoclonal bands (OCB). More recently, several studies ...have reported CSF free light chains (FLC) as an alternative. We show that absolute CSF κFLC concentrations were highly sensitive — more than OCB testing — and specific for clinically isolated syndrome, relapsing remitting and primary progressive MS. Measurement of κFLC alone was sufficient. Our results suggest that CSF κFLC levels measured by nephelometry, if validated in a larger series, are a preferred test to OCB analysis in the diagnostic work-up of patients suspected of having MS.
Summary
Background
Several cytokines are involved in the recruitment and activation of inflammatory cells in ocular allergic diseases. The purpose of the study was to assay multiple cytokines and ...chemokines in tears, to compare subgroups of allergic conjunctivitis (AC) with controls, and in culture supernatants to determine whether conjunctival fibroblasts produce some of these cytokines.
Methods
Fifty to one hundred microlitre tears were obtained from patients with active seasonal allergic conjunctivitis (SAC; n=12), vernal keratoconjunctivitis (VKC; n=18), atopic keratoconjunctivitis (AKC; n=6) and non‐atopic controls (n=14). Primary conjunctival fibroblasts grown in vitro were stimulated with IL‐4, IL‐13 or TNF‐α for 24 h. Cell‐free tear and culture supernatants were assayed for IL‐1β, IL‐2, IL‐4, IL‐5, IL‐6, IL‐8, IL‐10, IL‐12, IL‐13, IFN‐γ, TNF‐α, eotaxin, MCP‐1 and RANTES using multiplex bead analysis. Induction of chemokine gene expression was determined by PCR.
Results
IL‐1β, IL‐2, IL‐5, IL‐6, IL‐12, IL‐13, MCP‐1 were increased in all tears groups compared with controls, with highly significant correlations between many of these molecules. In addition IL‐4, IFN‐γ, and IL‐10 were elevated in SAC and VKC, while eotaxin and TNF‐α were only increased in VKC. IL‐6, IL‐8, MCP‐1, RANTES and eotaxin were detected from fibroblasts cultures, and were all up‐regulated by TNF‐α. By PCR, fibroblasts expressed MCP‐1 transcripts constitutively, whereas IP‐10 and Mig were up‐regulated by TNF‐α.
Conclusions
Differential cytokine levels support tears as a useful indicator of immune mechanisms occurring during AC. The striking similarities in chemokine profiles between tears and fibroblasts suggest these cells as likely sources of chemokines in tears.
Abstract Spinal release of cytokines may play a critical role in the maladapted nociceptive signaling underlying chronic pain states. In order to investigate this biology, we have developed a novel ...‘high flux’ intrathecal microdialysis approach in combination with multiplex bead-based immunoassay technology to concurrently monitor the spinal release of interleukin (IL)-1β, IL-6 and tumour necrosis factor (TNF)α in rats with unilateral sciatic nerve chronic constriction injury (CCI). Intrathecal microdialysis was performed under isoflurane/N2 O anaesthesia in rats with confirmed mechanical hypersensitivity. In a first study, C-fiber strength electrical stimulation of the operated nerve in neuropathic rats was found to evoke a dramatic increase in IL-1β efflux (∼15-fold) that was significantly greater than that observed in the sham-operated group. Spinal IL-6 efflux was also responsive to primary afferent stimulation, whereas TNFα was not. In a second study, treatment with the glial inhibitor propentofylline for 7 days normalized CCI-induced mechanical hypersensitivity. In the same animals, this treatment also significantly reduced intrathecal IL-1β, IL-6 and TNFα and prevented afferent stimulation-evoked cytokine release of both IL-1β and IL-6. These results provide support for glia as the source of the majority of intrathecal IL-1β, IL-6 and TNFα that accompanies mechanical hypersensitivity in the CCI rat. Moreover, our studies demonstrate the ability of a neurone-glia signaling mechanism to dynamically modulate this release and support a role of spinal IL-1β in the phasic transmission of abnormal pain signals.
Pathological processes involved in the initiation of rheumatoid synovitis remain unclear. We undertook the present study to identify immune and stromal processes that are present soon after the ...clinical onset of rheumatoid arthritis (RA) by assessing a panel of T cell, macrophage, and stromal cell related cytokines and chemokines in the synovial fluid of patients with early synovitis. Synovial fluid was aspirated from inflamed joints of patients with inflammatory arthritis of duration 3 months or less, whose outcomes were subsequently determined by follow up. For comparison, synovial fluid was aspirated from patients with acute crystal arthritis, established RA and osteoarthritis. Rheumatoid factor activity was blocked in the synovial fluid samples, and a panel of 23 cytokines and chemokines measured using a multiplex based system. Patients with early inflammatory arthritis who subsequently developed RA had a distinct but transient synovial fluid cytokine profile. The levels of a range of T cell, macrophage and stromal cell related cytokines (e.g. IL-2, IL-4, IL-13, IL-17, IL-15, basic fibroblast growth factor and epidermal growth factor) were significantly elevated in these patients within 3 months after symptom onset, as compared with early arthritis patients who did not develop RA. In addition, this profile was no longer present in established RA. In contrast, patients with non-rheumatoid persistent synovitis exhibited elevated levels of interferon-gamma at initiation. Early synovitis destined to develop into RA is thus characterized by a distinct and transient synovial fluid cytokine profile. The cytokines present in the early rheumatoid lesion suggest that this response is likely to influence the microenvironment required for persistent RA.
Membrane microdomains known as lipid rafts have been shown recently to be involved in Fas signalling and apoptosis in T and B cell lines. Here, we have investigated further the role of lipid rafts in ...Fas-induced apoptosis in non-transformed human CD4 T cells. We show that Fas-induced apoptosis in CD4 T cells was inhibited by the lipid raft disrupter methyl-β-cyclodextrin. When lipid rafts were isolated from control and Fas ligand treated cells, we found that a small proportion of Fas was present in the raft fraction in untreated cells and that this was greatly increased upon Fas ligation. The other components of the Death Inducing Signalling Complex (DISC), FADD, and procaspase 8, were also present at higher levels in the raft fraction isolated from Fas ligand treated cells. We conclude that formation of the DISC occurs in lipid rafts and that these membrane microdomains are required for efficient Fas signalling and apoptosis.
Summary
Background Topical photodynamic therapy (PDT) is successful in the treatment of nonmelanoma skin cancers and associated precancers, but efficacy is significantly reduced in actinic keratosis ...lesions not located on the face or scalp.
Objectives To compare the changes in protoporphyrin IX (PpIX) fluorescence in lesions undergoing routine methylaminolevulinate (MAL) PDT and the clinical outcome observed 3 months after treatment in lesions located at acral and nonacral sites.
Methods This study was a noninterventional, nonrandomized, observational study, which monitored changes in PpIX fluorescence in 200 lesions during standard dermatological MAL‐PDT. These data were subsequently analysed in terms of lesions located at acral and nonacral sites.
Results Clinical clearance was significantly reduced (P < 0·01) in acral skin lesions when compared with lesions located at nonacral sites. The accumulation and destruction of PpIX fluorescence was significantly reduced in these acral lesions (P < 0·05 and P < 0·001, respectively). Specifically, lesion location at acral sites significantly reduced changes in PpIX fluorescence in actinic keratosis lesions during MAL‐PDT (P < 0·01 and P < 0·05).
Conclusions These data suggest that reduced PpIX accumulation and the subsequent reduction in PpIX photobleaching within acral lesions result in the reduced responsiveness of these lesions to MAL‐PDT. Future work should therefore aim to improve photosensitizer accumulation/photobleaching within lesions located at acral sites.
Summary
Objective The aetiology of idiopathic intracranial hypertension (IIH) is not known, but its association with obesity is well‐recognized. Recent studies have linked obesity with abnormalities ...in circulating inflammatory and adiposity related cytokines. The aim of this study was to characterize adipokine and inflammatory cytokine profiles in IIH.
Design Paired serum and cerebrospinal fluid (CSF) specimens were collected from 26 patients with IIH and compared to 62 control subjects. Samples were analysed for leptin, resistin, adiponectin, insulin, IL‐1β, IL‐6, IL‐8 (CXCL8), TNFα, MCP‐1 (CCL2), hepatocyte growth factor, nerve growth factor and PAI‐1 using multiplex bead immunoassays.
Results CSF leptin was significantly higher in patients with IIH (P = 0·001) compared to controls after correction for age, gender and body mass index (BMI). In the control population, BMI correlated with serum leptin (r = 0·34; P = 0·007) and CSF leptin (r = 0·51; P < 0·0001), but this was not the case for the IIH population. Profiles of other inflammatory cytokines and adipokines did not differ between IIH patients and controls once anthropometric factors had been accounted for.
Conclusions IIH was characterized by significantly elevated CSF leptin levels which did not correlate with BMI. We suggest that CSF leptin may be important in the pathophysiology of IIH and that obesity in IIH may occur as a result of hypothalamic leptin resistance.
Background
There continues to be uncertainty about the optimal approach to documenting bleeding data in platelet transfusion trials, with a desire to apply a common assessment tool across all trials. ...With this in mind, a consensus bleeding assessment tool (BAT) has been developed by the Biomedical Excellence for Safer Transfusion (BEST) collaborative, based on review of data collection forms used in published randomized trials and following content validation with a range of healthcare professionals at seven haematology centres through BEST members. This study aimed to evaluate reliability and reproducibility of the consensus BAT.
Methods
Replicated clinical assessments of bleeding were undertaken by participants with haematological malignancies recruited at four haematology centres in an international, multicentred, observational study. Concordance of repeat assessments was calculated for agreement in site and grade of bleeding observed.
Results
Forty patients consented to participate, and 13 trained bleeding assessors collected these data. Bleeding assessments were carried out on 113 separate days. Of all 225 bleeding assessments, 204 were compared for grade concordance, and 160 were compared for site concordance. There was very good grade concordance (83%, 95% confidence interval 74–93%) and good bleeding site concordance (69%, 95% confidence interval 57–79%) in observations of bleeding. Discordance was primarily in relation to assessing skin bleeding.
Conclusions
Alongside a structured training programme, levels of concordance for a consensus BAT were high. Researchers using assessment tools for bleeding need to balance comprehensive data collection against potential loss of accuracy for some types of bleeding, such as skin findings.
To extensively characterize the complex network of cytokines present in uveitis aqueous humor (AqH), and the relationships between cytokines and the cellular infiltrate.
AqH from noninflammatory ...control subjects and patients with idiopathic, Fuchs' heterochromic cyclitis (FHC), and herpes-viral or Behçet's uveitis were analyzed for IL-1beta, -2, -4, -5, -7, -8, -10, -12, -13, -15, TNFalpha, IFNgamma, CCL2 (MCP-1), CCL5 (RANTES), CCL11 (Eotaxin), TGFbeta2, and CXCL12 (SDF-1), using multiplex bead immunoassays. The cellular infiltrate was also determined for each sample.
Idiopathic uveitis AqH, compared with noninflammatory controls, was characterized by high levels of IL-6, IL-8, CCL2 and IFNgamma, the levels of which correlated with each other. For IL-6 and IL-8 these levels were proportional to the number of neutrophils present. By contrast, the levels of both TGFbeta2 and CXCL12 decreased in idiopathic uveitis AqH with increasing inflammation. Cluster analysis showed a degree of segregation between noninflammatory and idiopathic uveitis AqH. Further examination using random forest analysis yielded a complete distinction between these two groups. The minimum cytokines required for this classification were IL-6, IL-8, CCL2, IL-13, TNFalpha, and IL-2.
Application of multiplex bead immunoassays has allowed us to identify distinct patterns of cytokines that relate to both clinical disease and the cellular infiltrates present. Bioinformatics analysis allowed identification of cytokines that differentiate idiopathic uveitis from noninflammatory control AqH and are likely to be important for the pathogenesis of uveitis.
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