Detection and isolation of auto-reactive human antibodies from primary B cells

The isolation of human monoclonal antibodies (hmAb) has emerged as a versatile platform in a wide variety of contexts ranging from vaccinology to therapeutics. In particular, the presence of high titers of circulating auto-antibodies is implicated in the pathology and outcome of autoimmune diseases. Therefore, the molecular characterization of these hmAb provides an avenue to understanding the pathogenesis of autoimmune diseases. Additionally, the phenotype of the auto-reactive B cells may have direct relevance for therapeutic intervention. In this report, we describe a high-throughput single-cell assay, microengraving, for the screening, characterization and isolation of anti-citrullinated protein antibodies (ACPA) from peripheral blood mononuclear cells (PBMC) of rheumatoid arthritis (RA) patients. Stimulated B cells are profiled at the single-cell level in a large array of sub-nanoliter nanowells (?105), assessing both the phenotype of the cells and their ability to secrete cyclic-citrullinated peptide (CCP)-specific antibodies. Single B cells secreting ACPA are retrieved by automated micromanipulation, and amplification of the immunoglobulin (Ig) heavy and light chains is performed prior to recombinant expression. The methodology offers a simple, rapid and low-cost platform for isolation of auto-reactive antibodies from low numbers of input cells and can be easily adapted for isolation and characterization of auto-reactive antibodies in other autoimmune diseases

Targeting type I interferons in systemic lupus erythematous

Systemic lupus erythematosus (SLE) is a complex autoimmune disease with systemic clinical manifestations including, but not limited to, rash, inflammatory arthritis, serositis, glomerulonephritis, and cerebritis. Treatment options for SLE are expanding and the increase in our understanding of the immune pathogenesis is leading to the development of new therapeutics. Autoantibody formation and immune complex formation are important mediators in lupus pathogenesis, but an important role of the type I interferon (IFN) pathway has been identified in SLE patients and mouse models of lupus. These studies have led to the development of therapeutics targeting type I IFN and related pathways for the treatment of certain manifestations of SLE. In the current narrative review, we will discuss the role of type I IFN in SLE pathogenesis and the potential translation of these data into strategies using type I IFN as a biomarker and therapeutic target for patients with SLE

Effect of dietary supplementation of crude microalgal extracts on growth performance, survival and disease resistance of Lates calcarifer (Bloch, 1790) larvae

The present study aimed to assess growth performance and health status of Lates calcarifer larvae fed with crude microalgal extracts. Larvae weighing 5.96 ± 0.39 mg were randomly divided into four experimental groups in triplicates, with each replicate having 35 larvae. Four diets formulated to contain 55% protein and 12% lipid, were prepared with pigment rich microalgal extracts at 0% (T1) and 1% each of crude phycocyanin (T2), chlorophyll (T3) and astaxanthin (T4) extracts. The experiment was carried out in 30 l plastic tanks by feeding experimental diets to satiation. There was no significant difference (p<0.05) in growth performance in larval groups fed different experimental diets. Percentage survival during the feeding experiment was significantly higher in the astaxanthin fed group (77.14±1.65) and lower in chlorophyll fed group (36.19±0.95) compared to control group. Cannibalism (%) was significantly reduced in the treatments fed astaxanthin (7.61±0.95) and phycocyanin (8.57±0.3) pigmented diets compared to chlorophyll fed group (13.33±0.95) and control group. Bacterial challenge with pathogenic Vibrio alginolyticus revealed highest resistance in astaxanthin and phycocyanin fed larvae. The study confirms that astaxanthin and phycocyanin at 1% dietary levels improve survival and disease resistance in Asian seabass larvae but do not affect growth performance

Glycolytic requirement for NK cell cytotoxicity and cytomegalovirus control

NK cell activation has been shown to be metabolically regulated in vitro; however, the role of metabolism during in vivo NK cell responses to infection is unknown. We examined the role of glycolysis in NK cell function during murine cytomegalovirus (MCMV) infection and the ability of IL-15 to prime NK cells during CMV infection. The glucose metabolism inhibitor 2-deoxy-ᴅ-glucose (2DG) impaired both mouse and human NK cell cytotoxicity following priming in vitro. Similarly, MCMV-infected mice treated with 2DG had impaired clearance of NK-specific targets in vivo, which was associated with higher viral burden and susceptibility to infection on the C57BL/6 background. IL-15 priming is known to alter NK cell metabolism and metabolic requirements for activation. Treatment with the IL-15 superagonist ALT-803 rescued mice from otherwise lethal infection in an NK-dependent manner. Consistent with this, treatment of a patient with ALT-803 for recurrent CMV reactivation after hematopoietic cell transplant was associated with clearance of viremia. These studies demonstrate that NK cell-mediated control of viral infection requires glucose metabolism and that IL-15 treatment in vivo can reduce this requirement and may be effective as an antiviral therapy