Autoantibody screening in Guillain-Barré syndrome.

BACKGROUND: Guillain-Barré syndrome (GBS) is an acute inflammatory neuropathy with a heterogeneous presentation. Although some evidences support the role of autoantibodies in its pathogenesis, the target antigens remain unknown in a substantial proportion of GBS patients. The objective of this study is to screen for autoantibodies targeting peripheral nerve components in Guillain-Barré syndrome. METHODS: Autoantibody screening was performed in serum samples from all GBS patients included in the International GBS Outcome study by 11 different Spanish centres. The screening included testing for anti-ganglioside antibodies, anti-nodo/paranodal antibodies, immunocytochemistry on neuroblastoma-derived human motor neurons and murine dorsal root ganglia (DRG) neurons, and immunohistochemistry on monkey peripheral nerve sections. We analysed the staining patterns of patients and controls. The prognostic value of anti-ganglioside antibodies was also analysed. RESULTS: None of the GBS patients (n = 100) reacted against the nodo/paranodal proteins tested, and 61 (61%) were positive for, at least, one anti-ganglioside antibody. GBS sera reacted strongly against DRG neurons more frequently than controls both with IgG (6% vs 0%; p = 0.03) and IgM (11% vs 2.2%; p = 0.02) immunodetection. No differences were observed in the proportion of patients reacting against neuroblastoma-derived human motor neurons. Reactivity against monkey nerve tissue was frequently detected both in patients and controls, but specific patterns were only detected in GBS patients: IgG from 13 (13%) patients reacted strongly against Schwann cells. Finally, we confirmed that IgG anti-GM1 antibodies are associated with poorer outcomes independently of other known prognostic factors. CONCLUSION: Our study confirms that (1) GBS patients display a heterogeneous repertoire of autoantibodies targeting nerve cells and structures; (2) gangliosides are the most frequent antigens in GBS patients and have a prognostic value; (3) further antigen-discovery experiments may elucidate other potential antigens in GBS.

LRP1/CD91 is highly expressed in monocytes from patients with vitiligo - even after repigmentation.

Vitiligo pathophysiology is mediated by antigen-specific cytotoxic T cells. Environmental stressors cause susceptible melanocytes to secrete damage-associated molecular patterns (DAMPs). DAMPs are recognized by receptors such as the endocytic low-density lipoprotein receptor-related protein (LRP1/CD91), expressed in antigen-presenting cells, which activate self-reactive CD8+ T cells, leading to melanocyte destruction. Within this response, interferon gamma triggers production of cytokine CXCL10, recruiting more activated T cells causing further melanocytic damage. We hypothesized that expression of LRP1/CD91 was higher in vitiligo patients compared to non-vitiligo individuals. And further that levels/expression of CXCL10 in plasma were linked to disease severity. We enrolled forty individuals in this study: 18 patients with vitiligo and 22 healthy volunteers. We assessed LRP1/CD91 expression and plasma CXCL10 in patients with vitiligo and healthy volunteers. Additionally, vitiligo patients received combined treatment for 16 weeks following which the said parameters were reassessed. Vitiligo Area Scoring Index was calculated before and after treatment for these patients. Analysis of LRP1/CD91 MFI values in monocytes from vitiligo patients showed high surface levels of LRP1/CD91 than from healthy volunteers (10.50 ± 0.77 vs. 6.55 ± 0.77 MFI units, p < 0.001). This expression did not change after treatment. Plasma levels of CXCL10 were higher in vitiligo patients than healthy volunteers (93.78 ± 7.73 vs. 40.17 ± 6.25 pg/ml). The patients with a good clinical response to treatment had a parallel reduction in plasma CXCL10 levels (105.8 ± 18.44 vs. 66.13 ± 4.87 pg/ml) before and after treatment. LRP1/CD91 expression may reflect susceptibility to vitiligo. Plasma levels of CXCL10 can represent a biomarker for monitoring treatment response. LRP1 and CXCL10 may represent therapeutic targets.

Binding of Platelets to Lymphocytes: A Potential Anti-Inflammatory Therapy in Rheumatoid Arthritis.

Soluble factors released from platelets can modulate the immune response of leukocytes. We and others have recently found that T lymphocytes with bound platelets have reduced proliferation and IFN-γ and IL-17 production. Thus, we speculate that if we induce the binding of platelets to lymphocytes, we will be able to regulate the inflammatory response. When we cocultured platelets with lymphocytes at different ratios, we were able to increase the percentage of lymphocytes with bound platelets. The coculture of platelets with lymphocytes in the presence of stimulation decreased the production of IFN-γ and TNF-α, T cell proliferation, and the expression of CD25, PD-L1, and SLAM. However, this coculture increased CD39 expression. All of these effects were dependent on the dose of platelets and operated indistinctly with platelets from different healthy donors. When platelets were cocultured in the same compartment with lymphocytes, we observed less IFN-γ and TNF-α production and T lymphocyte proliferation than in cultures with platelets separated from lymphocytes by a 0.4-µm pore size filter. The binding of platelets to lymphocytes was blocked with anti-P-selectin Abs, and when this occurred we observed higher IFN-γ and TNF-α production than in nonblocked conditions. The cocultures of platelets with synovial fluid cells from rheumatoid arthritis patients reduced inflammatory cytokine production and increased IL-10 production. These results suggest that platelet binding to lymphocytes effectively regulates T lymphocyte function. This mechanism could be easily applied to reduce inflammatory responses.

Anti-NF155 chronic inflammatory demyelinating polyradiculoneuropathy strongly associates to HLA-DRB15.

BACKGROUND: The aim of the research is to study the human leukocyte antigen (HLA) class II allele frequencies in chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) associated with anti-neurofascin 155 (NF155) antibodies. METHODS: Thirteen anti-NF155+ and 35 anti-NF155 negative (anti-NF155neg) CIDP patients were included in a case-control study. The frequencies of the DRB1 HLA allele were analyzed in all patients while DQ frequencies were only studied in patients sharing the DRB1*15 allele. In silico HLA-peptide binding and NF155 antigenicity, predictions were performed to analyze overlap between presented peptides and antigenic regions. RESULTS: DRB1*15 alleles (DRB1*15:01 and DRB1*15:02) were present in 10 out of 13 anti-NF155+ CIDP patients and in only 5 out of 35 anti-NF155neg CIDP patients (77 vs 14%; OR = 20, CI = 4.035 to 99.13). DRB1*15 alleles appeared also in significantly higher proportions in anti-NF155+ CIDP than in normal population (77 vs 17%; OR = 16.9, CI = 4.434 to 57.30). Seven anti-NF155+ CIDP patients (53%) and 5 anti-NF155neg CIDP patients had the DRB1*15:01 allele (OR = 7, p = 0.009), while 3 anti-NF155+ CIDP patients and none of the anti-NF155neg CIDP patients had the DRB1*15:02 allele (OR = 23.6, p = 0.016). In silico analysis of the NF155 peptides binding to DRB1*15 alleles showed significant overlap in the peptides presented by the 15:01 and 15:02 alleles, suggesting functional homology. CONCLUSIONS: DRB1*15 alleles are the first strong risk factor associated to a CIDP subset, providing additional evidence that anti-NF155+ CIDP patients constitute a differentiated disease within the CIDP syndrome.

Expression of toll-like receptors 2 and 4 in subjects with asthma by total serum IgE level.

BACKGROUND: Emerging data suggest that innate immunity may play a role in asthma, particularly the toll-like receptors (TLRs). Some studies pointed to an involvement of TLRs 2 and 4 in the pathogenesis of allergic asthma, and other studies related TLRs to IgE. However, there are not any studies that have comprehensively evaluated the expression of TLRs 2 and 4 in inflammatory cells, in peripheral blood and induced sputum specimens from asthmatic patients, according to their total serum IgE. METHODS: We studied 44 asthmatic patients (15 with high total serum IgE and 29 with normal total serum IgE). On a single visit, all patients underwent: induced sputum, pulmonary function tests, determination of exhaled nitric oxide fraction, venipuncture for blood analysis and skin prick allergy tests. The induced sputum cellularity was analyzed by flow cytometry, where expression of TLRs 2 and 4 was studied using fluorochrome-conjugated monoclonal antibodies. RESULTS: Asthmatic patients with high total serum IgE showed, a higher percentage of macrophages expressing TLR4 (42.99 % ± 22.49) versus asthmatic patients with normal total serum IgE (28.84 % ± 15.16) (P = 0.048). Furthermore, we observed a correlation (but weak) between the percentage of macrophages expressing TLR4 in induced sputum and the total serum IgE level (R = 0.314; P = 0.040). CONCLUSION: Asthmatic subjects with high total serum IgE show increased macrophage expression of TLR4 in induced sputum. This outcome may result from a link between innate immunity and IgE-mediated, adaptive immune responses in asthma, and point to TLR4 as a potential therapeutic target.

VSL#3 probiotic treatment decreases bacterial translocation in rats with carbon tetrachloride-induced cirrhosis.

BACKGROUND & AIMS: Probiotics can prevent pathological bacterial translocation in cirrhosis by modulating intestinal microbiota and improving gut barrier and immune disturbances. To evaluate the effect of probiotic VSL#3 on bacterial translocation, intestinal microbiota, gut barrier and inflammatory response in rats with experimental cirrhosis. METHODS: Forty-six Sprague-Dawley rats with CCl4 -induced cirrhosis were randomized into two groups: VSL#3 group (n = 22) that received VSL#3 in drinking water, and water group (n = 24) that received water only. Treatment began at week 6 of cirrhosis induction and continued until laparotomy, performed 1 week after development of ascites or at week 20. A control group included 11 healthy rats. At this study end, we evaluated bacterial translocation, intestinal flora, intestinal barrier (ileal claudin-2 and 4, ß-defensin-1, occludin and malondialdehyde as index of oxidative damage) and serum cytokines. RESULTS: Mortality during this study was similar in the VSL#3 group (10/22, 45%) and the water group (10/24, 42%) (P = 1). The incidence of bacterial translocation was 1/12 (8%) in the VSL#3 group, 7/14 (50%) in the water group (P = 0.03 vs. VSL#3 group) and 0/11 in the control group (P = 0.008 vs. water group). The concentration of ileal and caecal enterobacteria and enterococci was similar in the two groups of cirrhotic rats. The ileal occludin concentration was higher and ileal malondialdehyde and serum levels of TNF-α were lower in the VSL#3 group than in the water group (P < 0.05). CONCLUSIONS: VSL#3 decreases bacterial translocation, the pro-inflammatory state and ileal oxidative damage and increases ileal occludin expression in rats with experimental cirrhosis.

Rituximab-induced interleukin-15 reduction associated with clinical improvement in rheumatoid arthritis.

Rituximab therapy alters all aspects of B-cell participation in the disturbed immune response of rheumatoid arthritis patients. To determine the impact of B-cell depletion on other immune compartments, we analysed levels of soluble and surface interleukin-15 (IL-15) along with the frequency of IL-15-related subsets after rituximab treatment. We then studied the correlation of observed changes with clinical activity. Heparinized blood samples from 33 rheumatoid arthritis patients were collected on days 0, 30, 90 and 180 after each of three rituximab cycles. Serum cytokine levels were determined by ELISA. Interleukin-15 trans-presentation was analysed by cytometry. Flow cytometry with monoclonal antibodies was performed to analyse circulating cell subsets. Interleukin-15 was detected in the serum of 25 patients before initiating the treatment. Rituximab then progressively reduced serum IL-15 (138 ± 21 pg/ml at baseline, 48 ± 18 pg/ml after third cycle, P = 0·03) along with IL-17 (1197 ± 203 pg/ml at baseline, 623 ± 213 pg/ml after third cycle, P = 0·03) and tended to increase the frequency of circulating regulatory T cells (3·1 ± 1 cells/µl at baseline, 7·7 ± 2 cells/µl after third cycle). Rituximab also significantly decreased IL-15 trans-presentation on surface monocytes of patients negative for IL-15 serum (mean fluorescence intensity: 4·82 ± 1·30 at baseline, 1·42 ± 0·69 after third cycle P = 0·05). Reduction of serum IL-15 was associated with decrease in CD8(+)  CD45RO(+) /RA(+) ratio (1·17 ± 0·21 at baseline, 0·36 ± 0·06 at third cycle, P = 0·02). DAS28, erythrocyte sedimentation rate and C-reactive protein correlated significantly with CD8(+)  CD45RO(+) /RA(+) ratio (R = 0·323, R = 0·357, R = 0·369 respectively, P < 0·001). Our results suggest that sustained clinical improvement after rituximab treatment is associated with IL-15/memory T-cell-related mechanisms beyond circulating B cells.

Randomized pilot study: effects of an exercise programme and leucine supplementation in patients with cirrhosis.

BACKGROUND: Physical exercise could improve functional limitations, muscle mass, and health-related quality of life (HRQoL) in patients with cirrhosis. AIM: The purpose of this study was to evaluate the efficacy and safety of an exercise programme and leucine supplementation to increase exercise capacity, muscle mass, and HRQoL in patients with cirrhosis. PATIENTS AND METHODS: Seventeen outpatients with cirrhosis were randomized to an exercise group (n = 8) or a control group (n = 9) in a pilot study. The programme of moderate exercise was performed for 12 weeks under supervision of a physiotherapist. All patients received oral leucine (10 g/day) during the study. At baseline and at the end of the study, we determined exercise capacity (6-min walk and 2-min step tests), anthropometric measurements, and HRQoL by Short Form-36 (SF-36) questionnaire. We also analyzed safety regarding complications of cirrhosis, liver and renal function, inflammatory response and oxidative stress. RESULTS: In the exercise group, exercise capacity improved, as shown by the increase in the 6-min walk test from 365 (160-420) to 445 m (250-500) (p = 0.01), and in the 2-min step test (p = 0.02). Lower thigh circumference also increased, from 41 (34-53) to 46 cm (36-56) (p = 0.02), and the domains of SF-36 general health (p = 0.03), vitality (p = 0.01) and social function (p = 0.04) improved significantly. In the control group, no statistically significant changes were observed in any of the parameters. We did not observe complications of cirrhosis in either group during the study. CONCLUSIONS: A programme of moderate physical exercise together with leucine supplements in patients with cirrhosis is safe and improves exercise capacity, leg muscle mass and HRQoL.

Diagnostic accuracy of serum calprotectin measured by CLIA and EIA in juvenile idiopathic arthritis: a proof-of-concept study.

Objective: C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) are used to assess disease activity in juvenile idiopathic arthritis (JIA). However, because these biomarkers do not always differentiate between active and inactive disease, there is a need for alternative markers such as serum calprotectin (sCal). The main aim of this proof-of-concept study was to assess the diagnostic accuracy of sCal in patients with JIA. Secondary aims were to identify the optimal sCal cut-off levels to define active disease and evaluate the association between these biomarkers and disease activity status. Methods: Serum samples were obtained from 25 pediatric patients with JIA. Serum calprotectin levels were determined by two different assays, the QUANTA FLASH chemiluminescence immunoassay (CLIA) from Inova Diagnostics and the solid-phase enzyme immunoassay (EIA) from Bühlmann Laboratories. Diagnostic accuracy was assessed for sCal CLIA, sCal EIA, CRP, and ESR. The results obtained by the CLIA and EIA methodologies were compared. We also evaluated the association between the individual each biomarkers (sCal CLIA, sCal EIA, CRP, and ESR) and disease activity (according to JADAS-27 criteria and the ACR criteria modified by Anink and colleagues). Results: For both sCal assays (CLIA and EIA), the optimal cut-off level (ROC analysis) was the same (2.3 µg/ml). Serum calprotectin levels measured by CLIA and EIA were strongly correlated with each other (Kendall's tau-b, 0.71; p < 0.001). Compared to ESR and CRP, sCal CLIA and EIA were both more accurate (i.e., greater sensitivity) in identifying patients with active disease. By contrast, ESR and CRP were more effective in identifying patients in remission (i.e., better specificity). Conclusion: This proof-of-concept study shows that determination of serum calprotectin levels with CLIA or EIA can accurately identify the presence of active disease in patients with JIA.

Inter-laboratory comparison of routine autoantibody detection methods for autoimmune neuropathies and myasthenia gravis.

Serological tests are important to detect autoantibodies (autoAbs) in patients with autoimmune neuropathies (AN) and myasthenia gravis (MG) as they are biomarkers for diagnosis, stratification, treatment selection, and monitoring. However, tests to detect autoAbs frequently lack proper standardization and results differ across diagnostic laboratories. We compared results for tests routinely performed in Spanish diagnostic laboratories to detect AN and MG autoAbs. In the Spanish Society of Immunology Autoimmunity Group national workshop, serum samples from 13 patients with AN or MG were tested for anti-ganglioside, anti-myelin-associated glycoprotein (MAG), anti-nicotinic acetylcholine receptor (AChR), and anti-muscle-specific kinase (MuSK) autoAbs using reference methods and were distributed for analysis to 27 participating laboratories using their routine methods. Overserved were inter-laboratory variability and worryingly low sensitivity, especially for anti-ganglioside immunoglobulin G and anti-MAG autoAb detection. This pilot study reflects autoAbs detection state of the art in AN and MG testing in leading diagnostic laboratories in Spain, highlighting the need for standardization prior to clinical use.

Prognostic significance of lymphocytic foci composition in minor salivary gland biopsies for severe disease flare and severity in Sjögren's syndrome: a 3-year follow-up cohort study.

Introduction: This was an ambispective cohort study evaluating the prognostic significance of lymphocytic foci and its lymphoid composition in minor salivary gland biopsy (MSGB) for short-term disease flare and severity in Sjögren's syndrome (SS). Methods: The inclusion criteria comprised individuals meeting the ACR/EULAR 2016 criteria who underwent MSGB with an infiltration of more than 50 lymphocytes and received clinical diagnosis between September 2017 and December 2018. Patients with inadequate biopsy samples were excluded. The number of lymphocytic foci and their lymphoid composition in MSGB were assessed using immunofluorescence staining. Major organ damage and improvements in the EULAR Sjögren's Syndrome Disease Activity Index (ESSDAI) were measured. Statistical analyses, including Cox and linear regressions, were conducted. Results: A total of 78 patients with at least one lymphocytic focus were included in the study. The presence of higher T-cell counts in lymphocytic foci in MSGB was associated with severe disease flare, and a logarithmic transformation of T-cell count indicated increased risk (HR 1.96, 95% CI 0.91-4.21). Improvements in the ESSDAI were associated with higher total lymphocyte count and T- and B-cell numbers in the lymphoid composition of the lymphocytic foci. Seropositive patients exhibited higher T CD4+ cell numbers. Correlation analysis showed negative associations between age and lymphocytic foci and the T-cell count. Positive correlations were observed between antinuclear antibody (ANA) titers and total lymphocyte numbers. Discussion: Patients with a higher number of T cells in the lymphocytic infiltrates of lymphocytic foci may have a two-fold risk of severe disease flare. The number of B cells and T CD4+ cells in the lymphocytic infiltrates of lymphocytic foci showed a weak but positive relation with the ESSDAI improvement during follow-up. Age and seropositivity appeared to influence the lymphoid composition of the lymphocytic foci.

Anti-TIF1γ antibodies (anti-p155) in adult patients with dermatomyositis: comparison of different diagnostic assays.

BACKGROUND: A new myositis-specific autoantibody (anti-p155) directed against transcriptional intermediary factor 1 γ (TIF1γ) has been described as a good marker of cancer-associated myositis (CAM). OBJECTIVE: To analyse the feasibility of detecting this autoantibody in patient serum samples using new assays with commercially available recombinant TIF1γ. METHODS: The study included 90 Spanish patients with dermatomyositis (DM), classified as clinically amyopathic DM, CAM, or DM without cancer. Anti-TIF1γ antibodies were detected by ELISA and immunoblot techniques and compared with anti-p155 antibody detection by protein immunoprecipitation assays with radiolabelled HeLa cells. The κ coefficient was used to compare the agreement between the different tests. RESULTS: Serum samples from 23 (25.6%) and 20 (22.2%) patients with DM recognised TIF1γ by ELISA and immunoblot, respectively. ELISA (κ=0.91) and immunoblot (κ=0.88) showed excellent agreement with immunoprecipitation analysis (anti-p155). Good concordance (κ=0.91) was also seen between ELISA and immunoblot. CONCLUSIONS: Excellent agreement was found between anti-p155 detected by immunoprecipitation and anti-TIF1γ detected by ELISA or immunoblot. These data indicate that identification of this autoantibody can be reliably performed in a standard laboratory setting, with potential application in clinical practice for cancer screening in adult patients with DM.

Functional consequences of CD36 downregulation by TLR signals.

TLR recognition activates the secretion of pro- and anti-inflammatory cytokines and it also modulates the expression of crucial molecules involved in phagocytosis and antimicrobial activity. Scavenger receptors can act as TLR co-receptors or facilitate antigen loading. However, it remains unknown whether TLR can modulate the expression of these scavenger receptors. We stimulated human peripheral blood mononuclear cells (PBMC) with TLR2 (Pam3CSK4 and FSL1) and TLR4 ligand lipopolysaccharide (LPS) and then analyzed CD36 expression on different monocyte subpopulations by flow cytometry. TLR2 and TLR4 ligands can downregulate CD36 on the surface of monocytes, guiding the protein to intracellular compartments. Even though TLR-activation induced TNFα, IL-10 and IL-6 production, only recombinant TNFα was able to downregulate CD36. Neutralizing anti-TNFα antibodies showed that the Pam3CSK4 and FSL1-induced downregulation was partially mediated by TNFα but not by IL-6 or IL-10. However, LPS-induced downregulation could have also been caused by direct TLR4 targeting and signaling, and/or mediated by other unknown factors. CD36 downregulation reduced the capability of monocytes to phagocyte apoptotic neutrophils. In conclusion, modulation of scavenger receptor expression by TLR targeting on monocytes has functional consequences. Characterization this complex regulation may help us to understand this innate response and develop specific therapeutic drugs for each mechanism.

Clinical and serological features of acute sensory ataxic neuropathy with antiganglioside antibodies.

There is as yet no consensus for considering pure acute sensory ataxic neuropathy (ASAN) as a variant of Guillain-Barré syndrome (GBS). Reactivity against gangliosides sharing disialosyl epitopes has been reported in these patients. The aim of this study was to determine the spectrum of reactivity against gangliosides in ASAN and to define the clinical pattern. From our database we identified patients with suspicion of ASAN. We defined ASAN as the presence of ataxia of peripheral origin with loss of proprioception, and areflexia, absence of ophthalmoplegia and no or minimal muscle weakness. Patients who met these criteria were retrospectively reviewed for their spectrum of reactivity against gangliosides and clinical features. We identified 12 patients fulfilling pre-defined criteria for ASAN. Reactivity against gangliosides containing disialosyl epitopes was present in seven patients. Concomitant reactivity against other gangliosides was present in 6/7 patients. All patients presented good prognosis and an antecedent illness was present in nine. Our results support the previously described clinico-immunological association between ASAN and disialosyl specificity, and widen the spectrum of reactivity against gangliosides. The acute presentation with a monophasic course, and good prognosis in all cases, together with transient immunoglobulin G antiganglioside antibodies and infectious antecedent in 7/12 patients support the inclusion of ASAN as a GBS variant.

Early differentiated CD28+ CD27+ T lymphocytes as a biomarker for short and long-term outcomes in older patients with pneumonia.

This study tested the hypothesis that a more senescent immune system would predict a worse outcome in older patients hospitalized for community-acquired pneumonia (CAP). CAP has long been responsible for high rates of mortality and readmissions among older people. Although immunosenescence is a key factor in the increased susceptibility to infections, there are no related biomarkers currently available in clinical practice. In this context, the aim of this prospective study was to identify immunosenescence-related biomarkers to predict outcomes in patients older than 65 years hospitalized for CAP. We evaluated 97 patients admitted to our hospital for CAP in 2019 and 2020. All patients were followed for 1 year. Our findings showed that elevated levels of early differentiated CD28+ CD27+ T cells at admission were associated with better short (2 months) and long-term (1 year) outcomes in terms of mortality and readmissions. Early differentiated CD28+ CD27+ CD4+ T cell counts were even better long-term predictors. In conclusion, early differentiated CD28+ CD27+ T cells could be useful biomarkers to identify high-risk older patients with CAP, helping clinicians with risk stratification and follow-up.

Krebs von den Lungen-6 glycoprotein circulating levels are not useful as prognostic marker in COVID-19 pneumonia: A large prospective cohort study.

Coronavirus disease 2019 (COVID-19) has rapidly expanded worldwide. Currently, there are no biomarkers to predict respiratory worsening in patients with mild to moderate COVID-19 pneumonia. Small studies explored the use of Krebs von de Lungen-6 circulating serum levels (sKL-6) as a prognostic biomarker of the worsening of COVID-19 pneumonia. We aimed at a large study to determine the prognostic value of sKL-6 in predicting evolving trends in COVID-19. We prospectively analyzed the characteristics of 836 patients with COVID-19 with mild lung disease on admission. sKL-6 was obtained in all patients at least at baseline and compared among patients with or without respiratory worsening. The receiver operating characteristic curve was used to find the optimal cutoff level. A total of 159 (19%) patients developed respiratory worsening during hospitalization. Baseline sKL-6 levels were not higher in patients who had respiratory worsening (median {IQR} 315.5 {209-469} vs. 306 {214-423} U/ml p = 0.38). The last sKL-6 and the change between baseline and last sKL-6 were higher in the respiratory worsening group (p = 0.02 and p < 0.0001, respectively). The best sKL-6 cutoff point for respiratory worsening was 497 U/ml (area under the curve 0.52; 23% sensitivity and 85% specificity). sKL-6 was not found to be an independent predictor of respiratory worsening. A conditional inference tree (CTREE) was not useful to discriminate patients at risk of worsening. We found that sKL-6 had a low sensibility to predict respiratory worsening in patients with mild-moderate COVID-19 pneumonia and may not be of use to assess the risk of present respiratory worsening in inpatients with COVID-19 pneumonia.

Growth Differentiation Factor 15 (GDF-15): A Novel Biomarker Associated with Poorer Respiratory Function in COVID-19.

It is essential to find new biomarkers for severity stratification of patients with coronavirus disease (COVID-19). Growth differentiation factor 15 (GDF-15) is upregulated in pathological conditions that involve inflammation and/or oxidative stress. We determined circulating levels of GDF-15 and correlated them with clinical and laboratory parameters reflecting severity in 84 patients with COVID-19, finding that GDF-15 levels were higher in both patients than in 20 healthy controls and were higher in patients with poorer respiratory function. GDF-15 levels also correlated with interleukin-6, C-reactive protein, ferritin and D-dimer levels and with neutrophilia and lymphopenia. Of all the analysed biomarkers, GDF-15 showed the best area under the receiver operating characteristics curve in identifying patients with poor respiratory function. In conclusion, our data support GDF-15 as a biomarker associated with pulmonary impairment in COVID-19 and so can potentially be useful in stratifying COVID-19 cases by severity.

Bulbar involvement in patients with antiganglioside antibodies against NeuNAc(alpha2-3)Gal.

BACKGROUND: Reactivity against terminal NeuNAc(alpha2-3)Gal, common to several gangliosides such as GD1a, GT1b and GM3, has rarely been reported. The authors recently described a patient with a clinical picture of acute relapsing sensory ataxic neuropathy and bulbar involvement in whom they demonstrated concomitant reactivity against NeuNAc(alpha2-3)Gal and disialosyl epitopes. The authors suggested a correlation between NeuNAc(alpha2-3)Gal reactivity and bulbar involvement. AIM: To determine the frequency of reactivity against terminal NeuNAc(alpha2-3)Gal in acute and chronic immune-mediated disorders, and its possible correlation with bulbar involvement. METHODS: The authors retrospectively reviewed reactivity in the serum of more than 3000 consecutive patients with acute and chronic disorders in which antiganglioside antibodies were studied. The authors selected those patients who were simultaneously positive, by ELISA or thin-layer chromatography, for IgG or IgM antibodies anti-GM3, GD1a and GT1b, and reviewed their clinical features. RESULTS: Reactivity against NeuNAc(alpha2-3)Gal, shared by GM3, GD1a and GT1b gangliosides, was detected in 10 patients: isolated in one patient, and concomitant with reactivity against other gangliosides in the remaining patients. Reactivity against NeuNAc(alpha2-3)Gal was frequently associated (8/10) with symptoms suggestive of bulbar involvement, such as dysphagia, dysarthria or dysphonia. Severe respiratory failure requiring mechanical ventilation was observed in four patients. CONCLUSIONS: Reactivity against the NeuNAc(alpha2-3)Gal epitope is rare and is generally found in association with reactivity against the disialosyl epitope. It can be detected in patients with acute or chronic disorders and could be a serological marker of clinical bulbar involvement and, to a lesser extent, associated with the development of severe respiratory failure.

Anti-HMGCR Specificity of HALIP: A Confirmatory Study.

A distinctive new indirect immunofluorescence pattern in liver tissue has been associated with anti-HMGCR autoantibodies. It is known as HALIP (HMGCR Associated Liver Immunofluorescence Pattern). In this study, we furthered the original studies to demonstrate the association of anti-HMGCR antibodies with the HALIP. Human anti-HMGCR antibodies from patients' sera were purified and incubated with rat triple tissue (kidney/stomach/liver). A characteristic HALIP was observed. Additionally, a colocalization assay of human anti-HMGCR antibodies with rabbit polyclonal anti-HMGCR antibodies showed colocalization of both immunofluorescence patterns. This study confirms that the HALIP is due to human anti-HMGCR antibodies.

High Levels of Platelet-Lymphocyte Complexes in Patients with Psoriasis Are Associated with a Better Response to Anti-TNF-α Therapy.

Psoriasis is currently considered to be an immune-mediated disease whose patho-mechanisms involve platelet activation, which seems to correlate with the activity of the disease. Platelet activation is associated with the formation of platelet-lymphocyte complexes (PLyC), although their significance remains unknown. Moreover, biological treatments that target tumor necrosis factor-α (TNF-α) reduce platelet activation. To clarify the significance of PLyC, we compared their levels in patients with psoriasis with those of healthy donors and determined whether platelet binding modifies the secretion of IL-17A by T helper cells. Finally, we assessed the effect of anti-TNF-α treatment on PLyC in responder and non-responder patients with psoriasis. Ours results demonstrated an increase in PLyC in patients with psoriasis. Moreover, the percentage of IL-17-secreting cells was observed to be higher in the platelet-lymphocyte complex population, and these cells tended to secrete greater amounts of IL-17A. Psoriasis patients treated with anti-TNF-α normalized platelet-lymphocyte complex values, and the basal percentage of platelet-T helper lymphocyte complexes was significantly higher in the responder group. In conclusion, PLyC are increased in psoriasis patients, and the number of complexes decreases in response to anti-TNF-α treatment, specifically in the responder group of patients. This finding suggests that PLyC are a prognostic biomarker of response to anti-TNF-α therapy, but prospective studies are necessary to verify these results in patients with psoriasis.