Immune Mechanisms Underpinning Long COVID: Collegium Internationale Allergologicum Update 2024.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can result in a prolonged multisystem disorder termed long COVID, which may affect up to 10% of people following coronavirus disease 2019 (COVID-19). It is currently unclear why certain individuals do not fully recover following SARS-CoV-2 infection. SUMMARY: In this review, we examine immunological mechanisms that may underpin the pathophysiology of long COVID. These mechanisms include an inappropriate immune response to acute SARS-CoV-2 infection, immune cell exhaustion, immune cell metabolic reprogramming, a persistent SARS-CoV-2 reservoir, reactivation of other viruses, inflammatory responses impacting the central nervous system, autoimmunity, microbiome dysbiosis, and dietary factors. KEY MESSAGES: Unfortunately, the currently available diagnostic and treatment options for long COVID are inadequate, and more clinical trials are needed that match experimental interventions to underlying immunological mechanisms.

Immunological Patient Stratification in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a complex disease characterized by profound fatigue, post-exertional malaise (PEM), and neurocognitive dysfunction. Immune dysregulation and gastrointestinal symptoms are commonly observed in ME/CFS patients. Despite affecting approximately 0.89% of the general population, the underlying pathophysiological mechanisms remain poorly understood. This study aimed to elucidate the relationship between immunological characteristics and intestinal barrier function in ME/CFS patients. ME/CFS patients were stratified into two groups based on their immune competence. After documentation of detailed medical records, serum and plasma samples were collected for the assessment of inflammatory immune mediators and biomarkers for intestinal barrier integrity by ELISA. We found reduced complement protein C4a levels in immunodeficient ME/CFS patients suggesting a subgroup-specific innate immune dysregulation. ME/CFS patients without immunodeficiencies exhibit a mucosal barrier leakage, as indicated by elevated levels of Lipopolysaccharide-binding protein (LBP). Stratifying ME/CFS patients based on immune competence enabled the distinction of two subgroups with different pathophysiological patterns. The study highlights the importance of emphasizing precise patient stratification in ME/CFS, particularly in the context of defining suitable treatment strategies. Given the substantial health and socioeconomic burden associated with ME/CFS, urgent attention and research efforts are needed to define causative treatment approaches.

Real-life evaluation of molecular multiplex IgE test methods in the diagnosis of pollen associated food allergy.

BACKGROUND: Diagnosis of food allergies is challenging, as combining information from specific IgE (sIgE)-sensitization pattern and skin prick tests (SPTs) with clinical history is necessary for a personalized management of allergic patients. The aim of this study was to compare two molecular tests, the ImmunoCAP ISAC (ISAC) and the Allergy Explorer, version 2 (ALEX2 ) in the context of pollen food syndrome (PFS) diagnosis in a real-life scenario, to assess the benefit of multiplex testing in PFS patients. METHODS: Diagnosis of food allergy was performed in 53 patients. Allergen-sIgE concentrations were measured with ISAC and ALEX2 . Results for sIgE were statistically compared with each other, with SPT results and with clinical presentation of the patients. RESULTS: Using ISAC as reference test for sIgE measurements, the average sensitivity of ALEX2 for PR-10 allergens was 83.2% and the average specificity 88.0%. If only low sIgE concentrations were included, the sensitivity was 60.8% and the specificity 91.1%. Apple and hazelnut sensitizations were confirmed in most patients by concordance of sIgE and SPT results. Significant correlations were shown between clinical symptoms and Mal d 1- and Gly m 4-sIgE levels measured by both tests and for Cor a 1-sIgE levels measured by ALEX2 . In eight patients, profilin related symptoms were supported by Hev b 8-sensitization. CONCLUSION: Multiplex testing is beneficial to understand patient-specific individual sensitization profiles and to providing personalized management recommendations. In the future, custom-designed test kits might enable reducing costs of multiplex testing for specific patient groups without compromising the diagnostic value.

DMTMM-mediated methylamidation for MALDI mass spectrometry analysis of N-glycans with structurally conserved sialic acid residues in biological fluids "via direttissima".

Characterization of serum glycoprotein N-glycans with matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) in positive-ion mode needs a derivatization step to stabilize and neutralize the negative charge on sialic acids. The acidic sugars are attached to the end of glycoproteins, glycolipids or gangliosides. Here, we present a method for sialic acid stabilization via modification based on derivatization of carboxylic acid group activated with 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMTMM) with methylamine. DMTMM substitutes in many processes N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide (EDC/NHS) as activation reagent due to its better performance and higher stability in water. Glycosylated proteins are used as solid phase support for glycan derivatization and purification from excess of derivatization reagents. We evaluated our glycan analysis method in murine sera and intestinal lavages. The stabilization of sialic acid enables a complete conservation of the glycan structures, in contrast to other methods where sialic acids are partially lost. In BALB/c mouse sera, we detected predominantly mono- and di-sialylated N-glycans with mostly N-Glycolylneuraminic acid (Neu5Gc) and only trace amounts of N-Acetyl neuraminic acid (Neu5Ac). BALB/c mouse intestinal lavages glycoproteins contained asialo N-glycans. DMTMM-mediated methylamidation of N-glycans for MALDI mass spectrometry analysis is a fast and cheap method for structurally conserved glycan derivatization.

Evaluation of Immune Dysregulation in an Austrian Patient Cohort Suffering from Myalgic Encephalomyelitis/Chronic Fatigue Syndrome.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a severe multi-systemic disease characterized by debilitating fatigue that is not relieved by rest. The causes of the disease are still largely unexplained, and no causative treatment is currently available. Changes in the immune response are considered as fundamental in the development of ME/CFS. Thus, we aimed to evaluate the immunological profile of ME/CFS patients in a retrospective data analysis. As part of the routine workup for ME/CFS patients, a differential blood count, leukocyte subtyping, and quantification of immunoglobulins and IgG subclasses, as well as a complement analysis, was performed. Out of 262 ME/CFS patients, 64.9% had a reduction or deficiency in at least one of the listed immune parameters. In contrast, 26.3% showed signs of immune activation or inflammation. A total of 17.6% of the ME/CFS patients had an unclassified antibody deficiency, with IgG3 and IgG4 subclass deficiencies as the most common phenotypes. Reduced MBL (mannose-binding lectin) levels were found in 32% of ME/CFS patients, and MBL deficiency in 7%. In summary, the present results confirmed the relevance of immune dysfunction in ME/CFS patients underlining the involvement of a dysfunctional immune response in the disease. Thus, immune parameters are relevant disease biomarkers, which might lead to targeted therapeutic approaches in the future.

The Impact of Dietary Sphingolipids on Intestinal Microbiota and Gastrointestinal Immune Homeostasis.

The large surfaces of gastrointestinal (GI) organs are well adapted to their diverse tasks of selective nutritional uptake and defense against the external environment. To maintain a functional balance, a vast number of immune cells is located within the mucosa. A strictly regulated immune response is required to impede constant inflammation and to maintain barrier function. An increasing prevalence of GI diseases has been reported in Western societies over the past decades. This surge in GI disorders has been linked to dietary changes followed by an imbalance of the gut microbiome, leading to a chronic, low grade inflammation of the gut epithelium. To counteract the increasing health care costs associated with diseases, it is paramount to understand the mechanisms driving immuno-nutrition, the associations between nutritional compounds, the commensal gut microbiota, and the host immune response. Dietary compounds such as lipids, play a central role in GI barrier function. Bioactive sphingolipids (SLs), e.g. sphingomyelin (SM), sphingosine (Sph), ceramide (Cer), sphingosine-1- phosphate (S1P) and ceramide-1-phosphate (C1P) may derive from dietary SLs ingested through the diet. They are not only integral components of cell membranes, they additionally modulate cell trafficking and are precursors for mediators and second messenger molecules. By regulating intracellular calcium levels, cell motility, cell proliferation and apoptosis, SL metabolites have been described to influence GI immune homeostasis positively and detrimentally. Furthermore, dietary SLs are suggested to induce a shift in the gut microbiota. Modes of action range from competing with the commensal bacteria for intestinal cell attachment to prevention from pathogen invasion by regulating innate and immediate defense mechanisms. SL metabolites can also be produced by gut microorganisms, directly impacting host metabolic pathways. This review aims to summarize recent findings on SL signaling and functional variations of dietary SLs. We highlight novel insights in SL homeostasis and SL impact on GI barrier function, which is directly linked to changes of the intestinal microbiota. Knowledge gaps in current literature will be discussed to address questions relevant for understanding the pivotal role of dietary SLs on chronic, low grade inflammation and to define a balanced and healthy diet for disease prevention and treatment.

Gastric Enzyme Supplementation Inhibits Food Allergy in a BALB/c Mouse Model.

Impaired gastric digestion due to suppressed gastric acidity enhances the risk for food allergy development. In the current study, we aimed to evaluate the impact of a supported gastric digestion via application of a pharmaceutical gastric enzyme solution (GES) on food allergy development and allergic reactions in a BALB/c mouse model. The ability of the GES to restore hypoacidic conditions was tested in mice treated with gastric acid suppression medication. To evaluate the impact on allergic symptoms, mice were orally sensitized with ovalbumin (OVA) under gastric acid suppression and subjected to oral challenges with or without GES. The immune response was evaluated by measurement of antibody titers, cytokine levels, mucosal allergy effector cell influx and regulatory T-cell counts. Clinical response was objectified by core body temperature measurements after oral OVA challenge. Supplementation of GES transiently restored physiological pH levels in the stomach after pharmaceutical gastric acid suppression. During oral sensitization, supplementation of gastric enzymes significantly reduced systemic IgE, IgG1 and IgG2a levels and allergic symptoms. In food allergic mice, clinical symptoms were reduced by co-administration of the gastric enzyme solution. Support of gastric digestion efficiently prevents food allergy induction and alleviates clinical symptoms in our food allergy model.

Immune Effects of the Nitrated Food Allergen Beta-Lactoglobulin in an Experimental Food Allergy Model.

Food proteins may get nitrated by various exogenous or endogenous mechanisms. As individuals might get recurrently exposed to nitrated proteins via daily diet, we aimed to investigate the effect of repeatedly ingested nitrated food proteins on the subsequent immune response in non-allergic and allergic mice using the milk allergen beta-lactoglobulin (BLG) as model food protein in a mouse model. Evaluating the presence of nitrated proteins in food, we could detect 3-nitrotyrosine (3-NT) in extracts of different foods and in stomach content extracts of non-allergic mice under physiological conditions. Chemically nitrated BLG (BLGn) exhibited enhanced susceptibility to degradation in simulated gastric fluid experiments compared to untreated BLG (BLGu). Gavage of BLGn to non-allergic animals increased interferon-γ and interleukin-10 release of stimulated spleen cells and led to the formation of BLG-specific serum IgA. Allergic mice receiving three oral gavages of BLGn had higher levels of mouse mast cell protease-1 (mMCP-1) compared to allergic mice receiving BLGu. Regardless of the preceding immune status, non-allergic or allergic, repeatedly ingested nitrated food proteins seem to considerably influence the subsequent immune response.