A postbiotic from Aspergillus oryzae attenuates the impact of heat stress in ectothermic and endothermic organisms.

Heat stress is detrimental to food-producing animals and animal productivity remains suboptimal despite the use of heat abatement strategies during summer. Global warming and the increase of frequency and intensity of heatwaves are likely to continue and, thus, exacerbate the problem of heat stress. Heat stress leads to the impairment of physiological and cellular functions of ectothermic and endothermic animals. Therefore, it is critical to conceive ways of protecting animals against the pathological effects of heat stress. In experiments with endothermic animals highly sensitive to heat (Bos taurus), we have previously reported that heat-induced systemic inflammation can be ameliorated in part by nutritional interventions. The experiments conducted in this report described molecular and physiological adaptations to heat stress using Drosophila melanogaster and dairy cow models. In this report, we expand previous work by first demonstrating that the addition of a postbiotic from Aspergillus oryzae (AO) into the culture medium of ectothermic animals (Drosophila melanogaster) improved survival to heat stress from 30 to 58%. This response was associated with downregulation of genes involved in the modulation of oxidative stress and immunity, most notably metallothionein B, C, and D. In line with these results, we subsequently showed that the supplementation with the AO postbiotic to lactating dairy cows experiencing heat stress decreased plasma concentrations of serum amyloid A and lipopolysaccharide-binding protein, and the expression of interleukin-6 in white blood cells. These alterations were paralleled by increased synthesis of energy-corrected milk and milk components, suggesting enhanced nutrient partitioning to lactogenesis and increased metabolic efficiency. In summary, this work provides evidence that a postbiotic from AO enhances thermal tolerance likely through a mechanism that entails reduced inflammation.

Effects of PCB 126 on thymocyte surface marker expression and immune organ development in chicken embryos.

Previous studies have shown that chicken (Gallus domesticus) embryos are sensitive to the immunotoxic effects of Ah receptor agonists. These chemicals cause atrophy of the thymus gland and bursa of Fabricius, the sites of T- and B-lymphocyte maturation, respectively. The objectives of this study were (1) to evaluate the effects of 3,3,4,4',5-pentachlorobiphenyl (PCB 126) on thymocyte phenotypes (CD4-CD8-, CD4+CD8+, CD4+CD8-, CD4-CD8+, TCRalphabeta+, and TCRgammadelta) in chicken embryos, and (2) to compare phenotype alterations with masses and cellularity of lymphoid organs. To simulate exposure in wild avian embryos, chicken eggs were injected with PCB 126 (sunflower oil carrier) into the air cell before incubation. Doses ranged from 0.051 to 0.8 ng/g egg with carrier-injected and noninjected control groups. The thymus and bursa were removed, weighed, and homogenized on d 20 of egg incubation (1 d before hatch). Thymocyte phenotypes were quantified by flow cytometry using monoclonal antibodies for CD4, CD8, TCRalphabeta (Vbeta1), and TCRgammadelta. Right thymus mass declined with dose, decreasing significantly between 0.32 and 0.8 ng/g to a size 28% lower than controls. Live lymphoid cell numbers in the right thymus dropped markedly (21% lower than controls) between 0.051 and 0.13 ng/g, with a further decrease (35% lower than controls) at higher doses. There was no significant change in the percentage of thymocytes expressing TCRalphabeta. The total number of TCRalphabeta+ thymocytes decreased with dose as a function of the declines in TCRalphabeta+ percentages and total thymocyte numbers. The percentages of all other measured phenotypes were unaltered by PCB 126. The total number of CD4+CD8+ cells, and to a lesser degree CD4-CD8+ cells, decreased in a dose-dependent manner following the pattern of total live thymocytes. The number of viable lymphoid cells in the bursa decreased to 45% lower than controls at 0.13 ng/g and fell to 76% lower than controls at 0.8 ng/g. Lymphoid atrophy occurred at doses that were 8- to 12-fold lower with full-term incubation as compared to exposure only during later stages of incubation, and the lymphoid atrophy was associated with decreased TCRalphabeta+ thymocytes at higher doses. These immunological effects were observed at concentrations of PCB 126 comparable to those found in Great Lakes herring gull eggs, after correcting for interspecies differences in sensitivity to PCB 126.

Genetic and immunologic studies of patients on procainamide.

Forty (40) patients with cardiac arrhythmias receiving procainamide (PA) therapy and 24 patients who were receiving other drugs for their cardiac disorders were investigated for class II HLA phenotypes and their DRB1*04 and DQB1*03 subtypes. Other genetic marker evaluations in the PA patients included: 1) class III MHC C4A and C4B null alleles of complement; and, 2) acetylation phenotype. Twenty (20) of the PA patients were also tested for the ability of their stimulated cells to secrete Interleukin-1 (IL-1 beta) and tumor necrosis factor (TNF alpha). We also examined the spontaneous production of these cytokines by peripheral blood leukocytes (PBL) from patients who were receiving chronic PA treatment. The results revealed no association of acetylation phenotypes with the class II HLA phenotypes nor class III MHC C4 allotypes in these patients. The results did show a significant increase in class III C4 complement allotypes in the PA patients when compared to the controls. The results also showed a significant increase in autoantibodies and DQw3 phenotypes in the PA patient group when compared to control populations. Results of spontaneous IL-1 and TNF production suggested there may be an association of select class II HLA phenotypes in some patients and this may be relevant to host responsiveness to PA treatment.

Flow cytometric evaluation of cytotoxic peripheral blood lymphocytes in acute renal graft rejection.

The presence of the S6F1+ epitope on the surface of CD8+ lymphocytes is believed to be uniquely representative of cytotoxic subpopulations. A preliminary study was conducted to evaluate the CD8+ S6F1+ peripheral lymphocytes by flow cytometry in patients undergoing renal allograft biopsy for allograft dysfunction. Lymphocytes, obtained at the time of biopsy, were analyzed by flow cytometry with CD8-FITC/S6F1-RD1 as the test monoclonal antibody and MsIgG-RD1/MsIgG-FITC as internal control. A 100% increase in S6F1+ cells over internal control was considered to be positive result. The results were correlated with the histopathologic findings in 14 instances of allograft dysfunction occurring 26.5 +/- 11.6 days posttransplantation. The histopathologic diagnosis was acute cellular rejection in eight cases, acute tubular necrosis in four, and cyclosporine nephrotoxicity in two. Flow cytometric detection of an increase in S6F1+ cells yielded a sensitivity of 87.5% and a specificity of 83.3% for the diagnosis of acute rejection. It would appear that the use of a monoclonal antibody to detect increases in the number of CD8+ S6F1+ peripheral lymphocytes is a valuable test for the detection of acute allograft rejection in the initial period after transplantation.

Anti-HLA specificity determined by serum absorption with antigen-masked lymphocytes.

Lymphocyte or platelet absorption of multispecific sera is frequently used to establish HLA antibody specificity. Absorption of such sera with cells containing the desired antigenic combinations is often limited by the availability and the HLA characteristics of fresh or frozen cell panels. The method described permits improved absorption of specific HLA antibody with a preparation of cells containing a predetermined single HLA antigen. The procedure involves the screening of the patient's active sera against a panel of frozen cells. Analysis of the positive and negative reactions allows tentative assessment of recipient serum specificities. Cells with a single available HLA antigen type are prepared for serum absorption by suspending the cells in selected antisera to block (mask) HLA surface antigens while leaving just one type of antigen free for monospecific absorption. Following absorption of the test serum with the masked cells, the absorbed serum is then retested against the original panel of frozen cells. Should the results of this screen show a loss of a single HLA activity, the suspected specificity is assigned to the recipient's serum. The use of HLA antigen-masked lymphocytes has proven useful in the definition of HLA antibody specificity.