Escudero Lirola, Esther

Background: Antibodies to lipids are part of the first line of defense against microorganisms and regulate the pro/anti-inflammatory balance. Viruses modulate cellular lipid metabolism to enhance their replication, and some of these metabolites are proinflammatory. We hypothesized that antibodies to lipids would play a main role of in the defense against SARS-CoV-2 and thus, they would also avoid the hyperinflammation, a main problem in severe condition patients. Methods: Serum samples from COVID-19 patients with mild and severe course, and control group were included. IgG and IgM to different glycerophospholipids and sphingolipids were analyzed using a high-sensitive ELISA developed in our laboratory. A lipidomic approach for studying lipid metabolism was performed using ultra-high performance liquid chromatography coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry (UHPLC-ESI-QTOF-MS). Results: Mild and severe COVID-19 patients had higher levels of IgM to glycerophosphocholines than control group. Mild COVID-19 patients showed higher levels of IgM to glycerophosphoinositol, glycerophosphoserine and sulfatides than control group and mild cases. 82.5% of mild COVID-19 patients showed IgM to glycerophosphoinositol or glycerophosphocholines plus sulfatides or glycerophosphoserines. Only 35% of severe cases and 27.5% of control group were positive for IgM to these lipids. Lipidomic analysis identify a total of 196 lipids, including 172 glycerophospholipids and 24 sphingomyelins. Increased levels of lipid subclasses belonging to lysoglycerophospholipids, ether and/or vinyl-ether-linked glycerophospholipids, and sphingomyelins were observed in severe COVID-19 patients, when compared with those of mild cases and control group. Conclusion: Antibodies to lipids are essential for defense against SARS-CoV-2. Patients with low levels of anti-lipid antibodies have an elevated inflammatory response mediated by lysoglycerophospholipids. These findings provide novel prognostic biomarkers and therapeutic targets.

We developed an ELISA assay demonstrating the high prevalence of serum IgM to phosphatidylcholine (IgM-PC) in the first stages of multiple sclerosis (MS). We aimed to analyze the role of serum IgM-PC as a biomarker of response to treatment. Paired serum samples from 95 MS patients were obtained before (b.t) and after (a.t) treatment with disease modifying therapies. Patients were classified as non-responders or responders to treatment, according to classical criteria. Serum IgM-PC concentration was analyzed using our house ELISA assay. The level of serum IgM-PC b.t was higher in patients treated later with natalizumab than in those treated with Copaxone (p = 0.011) or interferon-β (p = 0.009). Responders to natalizumab showed higher concentration of serum IgM-PC b.t than those who did not respond to it (p = 0.019). The 73.3% of patients with the highest level of serum IgM-PC b.t responded to natalizumab. IgM-PC level decreased a.t in both cases, non-responders and responders to natalizumab. IgM-PC levels a.t did not decrease in non-responders to interferon-β, but in responders to it the IgM-PC level decreased (p = 0.007). Serum IgM-PC could be a biomarker of response to natalizumab or interferon-β treatment. Further studies would be necessary to validate these results.

The detection of intrathecal IgA synthesis (IAS) in multiple sclerosis (MS) could be underestimated. To assess it, we develop a highly sensitive assay based on isoelectric focusing (IEF). 151 MS patients and 53 controls with different neurological diseases were recruited. IgA concentration was analyzed using a newly developed in house ELISA. IgA oligoclonal bands to detect IAS were determined by IEF. Most individuals showed an IgA concentration within normal range in serum samples (90.69%) but 31.37% of individuals had a IgA concentration below the normal range in the cerebrospinal fluid (CSF). No significant differences were observed between MS and control groups, neither in CSF nor in serum. The new IEF was more sensitive than those previously described (0.01 mg/dl of IgA), and clearly identified patients with and without IAS, that was not related with IgA concentration. Using IEF, MS patients showed higher percentage of IAS-IEF (43.00%) than the control group (16.98) (p = 0.001). The incidence was especially higher in patients with clinically isolated syndrome (66.00%). The new IFE demonstrated a higher percentage of IAS in MS patients than assumed in the past. The presence of IAS-IEF in MS is higher than in other neurological diseases.

Antibodies and oxidative stress are hallmarks of multiple sclerosis (MS) lesions. We aimed to clarify the relation between them, their role in MS patients and to investigate their specificity, comparing MS with classical neurodegenerative diseases (ND). Brain samples from 14 MS cases, 6 with ND and 9 controls (without neurological diseases). Immunohistochemistry assays were used to detect oxidized lipids (EO6), IgG and IgM, oligodendrocytes (Olig2), axons (NF, neurofilament) and cellular (TUNEL) and axonal damage (APP, amyloid precursor protein). We did not observe EO6 in controls. All samples from MS patients showed EO6 in oligodendrocytes and axons within lesions. We did not detect co-localization between EO6 and antibodies. Neither did we between EO6 and TUNEL or APP. 94.4% of TUNEL-positive cells in normal appearing white matter were also stained for IgG and 75.5% for IgM. IgM, but not IgG, co-localized with APP. EO6 was associated with axonal damage in amyotrophic lateral sclerosis (ALS). We did not observe association between antibodies and cellular or axonal damage in ND patients. MS patients showed a higher number of B cells and plasma cells in the lesions and meninges than controls. The number of B cells and plasma cells was associated with the presence of antibodies and with the activity of the lesions. We observed a main role of B lymphocytes in the development of MS lesions. Antibodies contribute to the oligodendrocyte and axonal damage in MS. Oxidative stress was associated with axonal damage in ALS.

Multiple sclerosis (MS) is a demyelinating and neurodegenerative disease of the central nervous system. It affects young people, and a considerable percentage of patients need the help of a wheelchair in 15 years of evolution. Currently, there is not a specific technique for the diagnosis of MS. The detection of oligoclonal IgG bands (OIgGBs) is the most sensitive assay for it, but it is not standardizable, only reference laboratories develop it, and uses cerebrospinal fluid. To obtain this sample, a lumbar puncture is necessary, an invasive proceeding with important side effects. It is important to develop and implement standard assays to obtain a rapid diagnosis because the earlier the treatment, the better the evolution of the disease. There are numerous modifying disease therapies, which delay the progression of the disease, but they have important side effects, and a considerable percentage of patients give up the treatment. In addition, around 40% of MS patients do not respond to the therapy and the disease progresses. Numerous researches have been focused on the characterization of predictive biomarkers of response to treatment, in order to help physicians to decide when to change to a second-line treatment, and then the best therapeutic option. Here, we review the new biomarkers for the diagnosis and response to treatment in MS. We draw attention in a new assay, the detection of serum IgM to phosphatidylcholine, that showed a similar sensitivity as OIgGBs and predicts the response to disease modifying treatments.

Objective: To evaluate the value of serum immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies reactive with phosphatidylcholine (PC) and lactosylceramide (LC) as biomarkers in MS. Methods: We developed an ultrasensitive ELISA technique to analyze serum IgG and IgM antibodies to LC and PC, which we used to analyze samples from 362 patients with MS, 10 patients with non-MS myelin diseases (Non-MSMYDs), 11 patients with nonmyelin neurologic diseases (Non-MYNDs), and 80 controls. MS serum samples included clinically isolated syndrome (CIS, n = 17), relapsing-remitting MS (RRMS, n = 62), secondary progressive MS (SPMS, n = 50), primary progressive MS (PPMS, n = 37), and benign MS (BENMS, n = 36). Results: We detected higher levels of serum IgM antibodies to PC (IgM-PC) in MS than control samples; patients with CIS and RRMS showed higher IgM-PC levels than patients with SPMS, PPMS, and BENMS and controls. MS and control samples did not differ in serum levels of IgM antibodies reactive with LC, nor in IgG antibodies reactive with LC or PC. Conclusions: Serum IgM-PC antibodies are elevated in patients with MS, particularly during the CIS and RRMS phases of the disease. Thus, serum IgM-PC is a candidate biomarker for early inflammatory stages of MS. Classification of evidence: This study provides Class III evidence that serum antibodies to PC are elevated in patients with MS. The study is rated Class III because of the case control design and the risk of spectrum bias: antibody levels in patients with MS were compared with healthy controls.