Augmented Innate and Adaptive Immune Responses Under Conditions of Diabetes–Filariasis Comorbidity

Metainflammation, as seen in chronic diabetes subjects, impairs immunity and increases the susceptibility to infections. In the present study, the effect of diabetes on immune response against filariasis was studied. Both toll-like receptor (TLR)-mediated and crude antigen-induced immune responses were quantified, in whole blood cultures from filariasis-infected subjects (LF+), with and without diabetes. Blood cultures were stimulated with TLR ligands (TLR2 and TLR4) or filarial antigen or were left unstimulated (control) for 18 h. Cytokine, chemokine, and defensin secretion was quantified by ELISA. Expression of HLA-DR, B7-1, B7-2, activation marker (CD69), and Th (Th1, Th2, Th17, and Th9) phenotypes was quantified by flow cytometry. Expression of immunomodulatory effectors (Cox-2, HO-1, IDO-1, and p47Phox) and Th-polarizing transcription factors (T-bet, GATA3, and ROR-γt) was quantified by quantitative PCR. Secretion of IL-27, IL-1Ra, IL-12, IL-33, IL-9, and SDF-1 was increased under diabetes conditions with increased Th9 polarization and increased expression of Cox-2 and IDO. Overall, diabetes was found to augment both TLR-mediated and antigen-induced inflammation, which can promote chronic pathology in LF+ subjects.

Immunophenotyping of blood cells of experimental animals immunized with Brucella abortus thermoextracts

Aim. To study the subpopulational structure of blood cells of the experimental animals immunized with thermoextracts (TE) of Brucella abortus in L- and S-form. Materials and methods. Total 100 certified («Vector», Novosibirsk) outbred mice were immunized with B. abortus I-206 TE in L- and S-form in 20 μg protein dose. After 1, 3, 7, 14 and 21 days of observation the phenotypes (CD45, CD3, CD4, CD8, CD19, CD69) of blood cells were detected. Results. General regularities were revealed after injection of the experimental preparations. So, B. abortus TE in L- and S-form caused the immune response that increased granulocyte number and expression of early activation marker CD69 by T- and B-lymphocytes of blood in early period of observation (1-3 days), decrease in general B-lymphocyte content in late periods of observation (7-21 days). Thus, mice received B. abortus ТE in L-form demonstrated authentically higher CD69 expression of blood lymphocyte subpopulations than mice received B. abortus ТE in S-form. Distinctions in formation of humoral immune response were revealed that probably was connected with alteration of Brucella chemical composition in the course of L-transformation. Conclusion. The investigation established that B. abortus TE in L- or S-form caused immunological reorganization in the experimental animal organisms. On the basis of the fin

Lymph Node Derived CLL Cells Have a More Activated Phenotype and Better Antigen Presentation Capabilities Compared To Those From The Peripheral Blood

It is now well established that CLL is a highly proliferative disease with replication restricted to lymphoid tissue microenvironments. There is good evidence that interactions involving activated CD4+ T cells influence proliferation of the neoplastic B-cell clone. In this study we investigated the paradox that, whilst chronically activated T cells are found at these sites, in-vitro studies have shown that peripheral blood (PB) CLL cells suppress T cell activation. We hypothesized that the explanation for these contradictory findings is that lymph node (LN) CLL cells have increased antigen-presenting capacity compared to those in the PB. Accordingly, we obtained paired PB and LN fine needle aspirate (FNA) samples from a series of CLL patients and compared the expression of molecules associated with activation, antigen presentation and the ability to stimulate a third party mixed lymphocyte reaction (MLR). Using multi color flow cytometry, CD5+/CD19+ CLL cells were gated for expression of CD80, CD86, HLA-DR, CD25 and CD69. LN CLL cells had higher expression of markers associated with co-stimulation: CD80 (p=0.002), CD86 (p=0.037, n=6), antigen presentation: HLA-DR (p=0.04) and activation: CD25 (p=0.005) and CD69 (p=0.0018). We next used an MLR to determine the functional significance of these findings. Irradiated CLL cells from PB and LN were mixed at co-culture ratio’s of 1:1 and 1:10 and the proliferation/activation of CD4+ T cells measured using the expression of Ki67, CD69, and HLA-DR, and thymidine incorporation. T cell expression of the proliferation marker ki67 and activation marker CD69 was higher when co-cultured with LN CLL cells compared to PB derived cells. 1:1 co-culture ratio, mean Ki67 expression induced by LN CLL cells was 16.1% , SD ± 8.0 compared to 11.5%, SD± 7.7 induced by PB CLL cells (p=0.04) and at 1:10 ratio 8.4% , SD ± 3.6 compared to 3.9%, SD± 3.8 (p=0.02). Similarly, T cell expression of CD69 induced by LN CLL cells at 1:1 ratio was 37.0% , SD ± 15.5 compared to 26.6%, SD± 13.8 induced by PB-CLL cells (p=0.025) and at a 1:10 ratio 14.7% , SD ± 7.7 compared to 11.0%, SD± 7.4 (p=0.0027). LN-derived CLL cells also induced higher levels of T cell HLA-DR expression: 1:1 ratio 30.0%, SD ± 14.7 compared to 21.1%, SD± 9.1 (p=0.02) and at a 1:10 ratio 12.3%, SD ± 5.0 compared to 8.4%, SD± 3.7 (p=0.03). In accordance with these findings, T cell proliferation, measured by thymidine incorporation, was significantly higher in LN-CLL co-cultures compared to PB: 1:1 ratio, LN 2252, SD ± 1549 compared to PB 1615, SD± 1302 (p=0.008) and at 1:10 ratio, LN 910, SD ± 746 compared to, PB 416, SD± 366 (p=0.052). These data support our hypothesis that LN CLL cells have an increased capacity to activate T cells compared to those from the PB. We next investigated whether migration through the endothelium might play a role in promoting the antigen presenting function of CLL cells and hypothesized that this increased antigen presentation capacity of LN-CLL cells would be associated with markers of adhesion and migration. In keeping with this concept, we established that LN-CLL cells had higher expression of CD49d (p=0.008) and CD38 (p=0.018) and reduced CXCR4 (p=0.02) when compared to PB-CLL. To demonstrate this functionally we used a novel in vitro circulation system to compare the phenotype of cells that migrate through the endothelium with those that remain in circulation. CLL cells that migrated into the extravascular compartment in this system had a similar phenotype to LN CLL cells with elevated levels of CD49d and CD38 (P = 0.02 and P = 0.009 respectively), increased levels of CD80 (p=0.0014) and CD86 (p=0.015) and a trend towards increased expression of the activation marker CD69 (p=0.08). CLL cells that remained in the circulating compartment had a phenotype comparable to PB CLL cells. These results provide an explanation for the fact that CLL lymph nodes contain significant numbers of activated T cells, despite the known inhibitory effects of the tumor. Since activated T cells are in turn thought to promote tumor proliferation, our findings suggest that reciprocal interactions between CLL cells and LN T cells might play an important role in disease progression. In addition, our results show that transendothelial migration appears to activate CLL cells. Whether this, or other interactions such as stimulation of the B cell receptor, are responsible for the enhanced antigen presenting capacity of LN CLL cells remains to be determined. Disclosures: No relevant conflicts of interest to declare.

Sex-based effects on the distribution of NK cell subsets in response to exercise and carbohydrate intake in adolescents

Carbohydrate (CHO) supplementation and female sex independently influence the natural killer (NK) cell response to acute exercise. Consequently, this study sought to elucidate sex-based differences in the distribution of NK cell subsets (i.e., CD56dimand CD56bright) in response to exercise and CHO intake. Twenty-two healthy 14-yr-old girls ( n = 11) and boys ( n = 11) cycled for 60 min at 70% maximal oxygen consumption while drinking 6% CHO (CT) or flavored water (WT). Blood was collected at rest, during exercise (30 and 60 min), and during recovery (30 and 60 min) to identify CD3−CD56dimand CD3−CD56brightNK cells. The activation marker CD69 was also determined on CD3−CD56+cells. CD56dimresponses, expressed as proportions or cell counts, were greater ( P ≤ 0.01) in girls by 67 and 105%, respectively. CD56brightcell counts ( P = 0.006), but not CD56brightproportions ( P = 0.89), were greater in girls by 82%. Both CD56dimand CD56brightsubset responses, expressed as proportions or cell counts, were lower ( P ≤ 0.01) in CT vs. WT by 33–36%. The CD56bright-to-CD56dimratio decreased at 30 min of exercise but increased during recovery ( P < 0.001), with no effect of sex or CHO. Regardless of trial, CD3−CD56+cells expressed ∼18% higher levels of CD69 during recovery in girls but not boys ( P = 0.03), despite similar proportions and counts of CD69+cells. These results demonstrate sex-based differences in the distribution of NK cell subsets and activation status in response to exercise, but not CHO intake, and further support the need to control for sex in exercise immunology studies.