Human herpesvirus infections and circulating microvesicles expressing galectin-3 binding protein in patients with systemic lupus erythematosus.

OBJECTIVES: Circulating microvesicles (MVs) expressing the type 1 interferon-inducible protein galectin-3 binding protein (G3BP) are potentially major sources of autoantigens in systemic lupus erythematosus (SLE). In this study, we explore if plasma concentrations of G3BP-expressing MVs correlate with signs of various active human herpesvirus (HHV) infections in SLE patients, suggesting a virus-induced mechanism for the generation of these vesicles. METHODS: In 49 SLE patients, the plasma levels of immunoglobulin G (IgG) against cytomegalovirus (CMV) pp52, Epstein-Barr virus (EBV) early antigen diffuse (EA/D), and HHV6 p41 were measured by ELISAs and used as humoral markers of ongoing/recently active viral infection. MVs in platelet-poor plasma were quantified and characterised by flow cytometry, with regard to the binding of Annexin V (AnxV) and the expression of G3BP. Spearman's rho and the Wilcoxon rank-sum test were applied for associative evaluation of virus serology with MV subsets, and clinical and demographic data. RESULTS: The CMV pp52-directed antibodies correlated positively with the high G3BP-expressing MVs; either low (rho=0.4, p-value=0.005) or high (rho=0.37, p-value=0.01) in AnxV-expression. Furthermore, these MV subsets were higher in individuals with high and low IgG levels against CMV pp52 and EBV EA/D, respectively, relative to subjects with low and high IgG levels against these HHV antigens. Importantly, none of the associations were explained by immunosuppressants or antimalarials. CONCLUSIONS: Ongoing/recently active CMV infection is associated with circulating MVs expressing G3BP in SLE patients, supporting a link between specific viral infections and potentially pathogenic MVs in SLE.

Increased antibody levels to stage-specific Epstein-Barr virus antigens in systemic autoimmune diseases reveal a common pathology.

The immune responses to antigens from different stages of the Epstein-Barr virus (EBV) life cycle were investigated in systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), Sjögren's syndrome (SS), and systemic sclerosis (SSc) to gain knowledge of EBV's involvement in the etiology of systemic autoimmune diseases (SADs) and for an overview of the humoral immune responses against EBV. Investigations were performed by the use of ELISA. IgM, IgA, and IgG antibody binding to 11 EBV antigens: EBNA1, EBNA2, BALF5, EAD, BALF2, EA/R, VCA p18, VCA p23, gB, gp350, and gp42 were examined in serum pools from SAD patients and healthy controls (HCs). Increased antibody levels against the 11 EBV antigens in the SAD pools were seen compared to the HC pool. Specifically, SLE was characterized by strongly increased IgA to EAD both compared to HCs and other SADs, and RA was characterized by increased IgM levels to several EBV antigens. The SADs may be partly distinguished by their differential immune responses to various antigens in the EBV life cycle. All together, these findings support an association between EBV infection and SADs.

Antibodies to expanded virus antigen panels show elevated diagnostic sensitivities in multiple sclerosis and optic neuritis.

An antigen panel consisting of Epstein-Barr, measles, mumps, varicella zoster and rubella viruses (EMMRZ) was recently presented, which may aid in the diagnosis of multiple sclerosis (MS). The aim of this study was to validate and extend the EMMRZ panel. Various candidates, such as Cytomegalovirus and John Cunningham virus were analysed in relapsing-remitting MS (RRMS) and optic neuritis (ON) samples by enzyme-linked immunosorbent assay. IgG levels were elevated in RRMS samples and correlations were found between serum and cerebrospinal fluid levels. Cohort-dependent optimized panels were obtained for RRMS and ON, which obtained the highest sensitivity when combined with the status of oligoclonal bands.

Antibodies to Epstein-Barr virus and neurotropic viruses in multiple sclerosis and optic neuritis.

Antibody indices to Measles, Mumps, Varicella Zoster (MRZ) are of diagnostic value in multiple sclerosis (MS). Here, we have investigated, if this panel could be extended to increase diagnostic value. Samples from relapsing-remitting (RR) MS and optic neuritis (ON) patients were tested for reactivity to antigens from Epstein-Barr, Varicella Zoster, Measles, Mumps and Rubella (EMMRZ) viruses. Increased IgG levels in serum and cerebrospinal fluid (CSF) were found in RRMS patients, along with a significant correlation between serum and CSF. The sensitivity of the EMMRZ panel was increased approximately 40% compared to the MRZ panel, suggesting that the EMMRZ panel may be useful in MS and ON diagnostics.

Efficient evaluation of humoral immune responses by the use of serum pools.

BACKGROUND: Collection and testing of individual serum samples are often used in research to gain knowledge about e.g. the humoral response against bacteria or virus. This is a valid but time-consuming method and might be a waste of valuable serum samples for inefficient research. So far, no study has considered using serum pools as a quick and efficient screening method to confirm or deny hypotheses. METHODS: We created serum pools from four different patient groups (systemic lupus erythematosus n=85, rheumatoid arthritis n=77, Sjögren's syndrome n=91, systemic sclerosis n=66) and one healthy control group (n=67). Each serum pool was analyzed using three well-known immunoassays: enzyme-linked immunosorbent assay (ELISA), line blot, and immunofluorescence microscopy (anti-nuclear antibody (ANA) screening). The presence of Epstein-Barr virus (EBV) EA/D-, EBNA-1-, VCA p23-, and gp350-directed antibodies was used to validate serum pools as an efficient tool for further investigations by comparison to previous findings in this area. RESULTS: The presence of EBV EA/D-, EBNA-1-, VCA p23-, and gp350-directed antibodies in each pool was consistent within the obtained ELISA and line blot results, as increased titers of IgG against the four antigens were found in all patient serum pools and also in individual sera regarding gp350. These results correspond to previous findings on individual samples from patients with these diseases. The presence of ANAs was observed in all four patient serum pools and not in the HC pool by both line blots and immunofluorescence microscopy, which corresponds with the expectations and further corroborate the application of serum pools for screenings. CONCLUSION: We developed and validated the use of serum pools that reliably and rapidly can confirm or deny hypotheses, which enables a more efficient research concentrating on the most evident factors.

Elevated Concentrations of Serum Immunoglobulin Free Light Chains in Systemic Lupus Erythematosus Patients in Relation to Disease Activity, Inflammatory Status, B Cell Activity and Epstein-Barr Virus Antibodies.

OBJECTIVE: In this study, we examined the concentration of serum immunoglobulin free light chains (FLCs) in systemic lupus erythematosus (SLE) patients and investigated its association with various disease parameters in order to evaluate the role of FLCs as a potential biomarker in SLE. Furthermore, FLCs' association with Epstein-Barr virus (EBV) antibodies was examined. METHODS: Using a nephelometric assay, κFLC and λFLC concentrations were quantified in sera from 45 SLE patients and 40 healthy controls. SLE patients with renal insufficiency were excluded in order to preclude high concentrations of serum FLCs due to decreased clearance. RESULTS: Serum FLC concentrations were significantly elevated in SLE patients compared to healthy controls (p<0.0001) also after adjusting for Ig levels (p<0.0001). The concentration of serum FLCs correlated with a global disease activity (SLE disease activity index (SLEDAI)) score of the SLE patients (r = 0.399, p = 0.007). Furthermore, concentrations of FLCs correlated with titers of dsDNA antibodies (r = 0.383, p = 0.009), and FLC levels and SLEDAI scores correlated in the anti-dsDNA-positive SLE patients, but not in anti-dsDNA-negative SLE patients. Total immunoglobulin (IgG and IgA) concentrations correlated with FLC concentrations and elevated FLC levels were additionally shown to associate with the inflammatory marker C-reactive protein and also with complement consumption determined by low C4 in SLE patients. Collectively, results indicated that elevated serum FLCs reflects increased B cell activity in relation to inflammation. SLE patients had an increased seropositivity of EBV-directed antibodies that did not associate with elevated FLC concentrations. An explanation for this could be that serum FLC concentrations reflect the current EBV activity (reactivation) whereas EBV-directed antibodies reflect the extent of previous infection/reactivations. CONCLUSION: SLE patients have elevated concentrations of serum FLCs that correlate with global disease activity scores and especially serologic markers for active disease. These findings are suggestive of circulating FLCs having potential as a new supplementary serologic biomarker in SLE.