Th17/1-Biased Inflammatory Environment Involved in the Response of Epithelial Cells to Antigen Stimuli in Nasal Polyps

T Cell Subsets and Associated Cytokines in Chronic Graft-Versus-Host-Disease Using G-CSF Primed Bone Marrow in Allogeneic Hematopoietic Stem Cell Transplantation

Blood ◽

10.1182/blood.v128.22.4580.4580 ◽

2016 ◽

Vol 128 (22) ◽

pp. 4580-4580

Author(s):

Monica M Rivera Franco ◽

Eucario Leon Rodriguez ◽

Diana Gomez Martin ◽

Javier Merayo Chalico ◽

Jorge Alcocer Varela

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Stem Cell ◽

Th17 Cells ◽

Mononuclear Cells ◽

Chronic Gvhd ◽

The Other ◽

Hematopoietic Stem ◽

Graft Versus Host ◽

Ifn Γ

Abstract Background Graft versus host disease (GVHD) is the major complication of allogeneic hematopoietic stem cell transplantation. It is characterized by an imbalance between the effector and regulatory arms of the immune system which results in the over production of inflammatory cytokines. Regulatory T (T regs) cells and T helper 17 (Th17) cells are two recently described lymphocyte subsets with opposing actions. Both can develop from naïve CD4+ T cell precursors under the influence of TGFβ1. Th17 lymphocytes, are key effector cells in rodent models of human diseases including GVHD. The other subset, T regs, is essential for dominant immunologic tolerance. At our institution, patients transplanted using G-CSF primed bone marrow (G-BM), have a lower incidence of acute and chronic GVHD when compared to those transplanted with peripheral blood and not primed bone marrow. Some microenvironment characteristics of this hematopoietic stem cells (HSC) source remain unknown, as well as the difference between Tregs, Th17 and cytokine levels in patients who develop GVHD and those who do not. Objective To analyze the characteristics of thirty-eight G-BM donor samples, identifying lymphocytes subsets and associated cytokines, and comparing patients who developed chronic GVHD (cGVHD) and those who did not. Materials and Methods A prospective analysis was performed in 38 G-BM samples from donors from 1999 to 2016. Mononuclear cells were defrosted, counted, and viability was evaluated. A 24 hour resting with RPMI, and posterior activation with PMA (50 ng/ml) for 48 hours was performed. Cells were harvested and cytokines were evaluated by flow cytometry (CBA assay). From each sample, one million mononuclear cells were permeabilized, fixed, and stained with CD4-FITC, IL17A-PE, IFN-γ APC, and IL-4 PECy7, for their posterior phenotipication by flow cytometry. The samples were obtained in a BD LSR Fortessa cytometry, and analyzed with the Flow-Jo software. Patients (recipients) information was analyzed using SPSS v.21. Results GVHD incidence was reported as following: Three (8%) patients developed acute GVHD (2 grade II, and 1 grade IV), 11 patients (29%) developed chronic GVHD (9% extensive, and 91% limited), and 24 patients did not present either. Mononuclear cells from G-BM from donors of patients who developed cGVHD showed a pro inflammatory response, characterized by an increased concentration of IL-17A (15.5 vs 0.71 pg/mL, p=0.013), TNF-α (80.27 vs 0.13 pg/mL, p=0.001), and IL-6 (4953.6 vs 11.75 pg/mL, p=0.025), after a mitogenic stimulation, compared to cells from donors of patients who did not developed GVHD. On the other hand, a decreased IL-10 production (2.62 vs 52.81 pg/mL, p=0.001) was documented in mononuclear cells from donors of patients who developed chronic GVHD, compared to donor cells of patients who did not. No significant difference in the production of IL-2, IL-4, and IFN-γ was observed. There was no difference in Th1 and Th2 between both groups, but mononuclear cells from donors of patients who developed chronic GVHD had a higher percentage of Th17 (1.02% vs 0.46%, p<0.001), and less Tregs (0.88% vs 1.95%, p<0.001), compared to those who did not developed GVHD. Conclusions Patients who develop cGVHD (29%) are characterized by a pro inflammatory response with an increased production of IL-17A, IL-6, and IFN-γ, and also a major percentage of Th17 cells. Also, a decreased suppressive response was documented with reduced IL-10 and Tregs levels. The low incidence of cGVHD show that G-CSF primed bone marrow is an excellent source for allogeneic HSC transplantations, and would be useful to compare these results with other HSC sources. Disclosures No relevant conflicts of interest to declare.

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Functional Characterization of CD4+ T-Cells in Aplastic Anemia (AA)

Blood ◽

10.1182/blood.v118.21.1340.1340 ◽

2011 ◽

Vol 118 (21) ◽

pp. 1340-1340 ◽

Cited By ~ 1

Author(s):

Shahram Y Kordasti ◽

Judith C. W. Marsh ◽

Sufyan Al-Khan ◽

Jie Jiang ◽

Alexander E Smith ◽

...

Keyword(s):

T Cells ◽

Treatment Group ◽

High Throughput ◽

Cd4 T Cells ◽

Th17 Cells ◽

Absolute Number ◽

Th2 Cells ◽

Th1 Cells ◽

Ifn Γ ◽

Pre Treatment

Abstract Abstract 1340 We have examined the role of CD4+ T-cells in the pathogenesis of AA in 63 patients, 48 of whom were analyzed at diagnosis and 15 following immunosuppressive therapy (IST). Absolute numbers of CD4+ regulatory T cells (Tregs, defined as CD3+CD4+CD25highCD27+Foxp3+) were lower in pre-treatment AA patients compared to 10 healthy donors (HDs) (5.5 × 106 v 1.4 × 107)(p=0.01). In patients with severe (SAA) and very severe AA (VSAA), the absolute number and frequency of Tregs were lower than non-severe AA (NSAA) (4.4 × 106/L v 1 × 107/L)(p=0.01) and HDs (4.4 × 106/L v 3 × 107/L) (p<0.001). Absolute numbers of Th1 and Th2 cells in all pre-treatment patients were higher compared to HDs (6.4 × 107/L v 1.8 × 107/L)(p=0.03) for Th1 and (2.6 × 107/L v 2.4 × 106/L)(p=0.006) Th2 cells. Although mean percentages of AA Th17 cells were higher than in HDs (1.5% v 0.15%)(p=0.04), differences in absolute numbers were not significant. Absolute numbers of Th2 and Th17 cells were increased in SAA (1.3 × 107/L v 7.4 × 106/L for Th2)(p=0.01) compared to NSAA (5.7 × 106/L v 2.15 × 106/L for Th17)(p=0.02). Ratios of Th1/Tregs (p=0.003), Th2/Tregs (p=0.02), and Th17/Tregs (p=0.001) were higher in SAA and VSAA compared to NSAA. Percentage of both activated (CD4+CD45RA−CD25highFoxp3high) and resting (CD4+CD45RA+ CD25highFoxp3low) Tregs was decreased in AA patients, compared to HDs (p=0.004 and p=0.01), whereas cytokine secreting Tregs (CD4+CD45RA−CD25high Foxp3low) were increased in AA (p<0.003). Sorted Tregs from AA patients did not suppress cytokine secretion by autologous or HD T effectors (Te) cells in 1:1 co-cultures, whereas IL-2 and IFN-γ secretion by AA Te (CD4+CD25lowCD127high) was suppressible by allogeneic Tregs from HDs, confirming Tregs dysfunction. AA Tregs did not inhibit either CD154 or CD69 expression on Te cells. Tregs from AA patients secreted significantly more IFN-γ, TNF-α and IL-17 (p=0.02, p=0.02 and p=0.01, respectively) after 4 hours stimulation with PMA/Ionomycine compared to HDs. Expression levels of FoxP3, ROR□c and T-bet in AA Tregs was normal. IFN-γ secreting cells (Th1) were enriched using enrichment kit then further enriched by FACS sorting. CDR3 region products of TCR Vβ-chain were amplified using Vβ specific forward and Cβ reverse primers. CDR3 PCR products from AA patients and HDs were subjected 454 sequencing (Roche GS FLX titanium). Sequencing was performed to yield an average ‘depth’ in excess of 1000 clonally reads (1000x) for each sample specific CDR3 PCR amp icon. Reads were processed using Roche Amp icon Variant Analyzer software (AVA). Diversity of TCR receptors (measured by spectratyping and confirmed by high throughput deep sequencing) in AA Th1 cells was lower than HDs (p=0.037), as shown by the percentage and number of consensus clusters in total sequence reads. Interestingly, percentages of the most dominant CDR3 clones, revealed by high throughput sequencing, were higher in AA compared to HDs, regardless of spectratyping pattern. Global gene expression of Tregs was compared in 3 pre-IST AA patients and 5 HDs. A unique gene signature consisting of 86 genes that were significant was identified. There were 8 down regulated genes (fold change) in the pre-treatment group; PIN4 (−4.1), OR2T12 (−3.3), AMAC1 (−2.73), PERP (−2.69), UTS2 (−2.27), RNF139 (−2.13), COMMD9 (−2.09) and LOC100128356 (−2.01). The top 10 of 78 up-regulated genes in the pre-treatment group were HBB (19.5), PSME2 (13.8), CSDA (13.07), FAM127A (7.78), EXOSC1 (7.73), BPGM (7.43), CYSLTR1 (7.17), CHPT1 (6.96) and PLAC8 (6.71). qPCR analysis for CSDA, HBB, PSMiE2, PERP, PIN4, and UTS2 confirmed a similar trend to the microarray results. Interestingly absolute number of Tregs, and Th2/Treg ratio were higher in 10 IST responsive patients compared to 5 non-responsive patients (p=0.005 and 0.02, respectively). We show that expansion of Th1, Th2, Th17, and decreased/skewed Tregs immunophenotype and function are a consistent and defining feature of SAA and VSAA. Clonal expansion of Th1 cells is likely to be antigen driven and the presence of dysfunctional Tregs aggravates this autoimmune response. Increases of Tregs, and Th2/Treg ratios following IST predicts a favourable response to this treatment. Disclosures: No relevant conflicts of interest to declare.

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PBMC-06- Intracellular Cytokine Staining (ICS) of IFN-gamma, IL-4 and IL-17 on Human Peripheral Blood Mononuclear Cells (PBMC) v1

10.17504/protocols.io.byrppv5n ◽

2021 ◽

Author(s):

Alessia Furgiuele ◽

Emanuela Rasini ◽

Maria Giulia Albizzati ◽

Alessandra Luini ◽

Marco Ferrari ◽

...

Keyword(s):

Flow Cytometry ◽

T Cells ◽

Peripheral Blood Mononuclear Cells ◽

Peripheral Blood ◽

Mononuclear Cells ◽

Brefeldin A ◽

Intracellular Cytokine ◽

Peripheral Blood Mononuclear ◽

Blood Mononuclear Cells ◽

Ifn Γ

This present protocol is developed to analyze the frequency of IFN-γ-, IL-4- and IL-17-producing CD4+T cells, identified from ex vivo human peripheral blood mononuclear cells (PBMC). The frequencies of cytokine producing cells derived from activation of PBMC was induced trough the stimulus phorbol 12-myristate 13-acetate (PMA) and ionomycin. According onpreviously published protocols concentrations of stimulating substances were in the range from 10, to 50 ng/ml for PMA and 1 µg/ml for ionomycin (Gupta and Maecker, 2015; Foster et al., 2007; Freer and Rindi, 2013; https://www.bdbiosciences.com/content/bdb/paths/generate-tds-document.in.560751.pdf). The PMA concentrations of 10, 20 and 50 ng/ml were tested and finally the PMA concentration of 10 ng/ml was chosen since it was sufficient to obtain a frequency of cytokines comparable to that obtained with higher stimulus concentrations. PMA/ionomycin and brefeldin A are incubate together for a time of 5 h (Gupta and Maecker, 2015, Foster et al., 2007, Freer and Rindi, 2013, https://www.bdbiosciences.com/content/bdb/paths/generate-tds-document.in.560751.pdf). The protein secretion inhibitor brefeldin A, was used at the concentration of 10 µg/ml (Gupta and Maecker, 2015; Foster et al., 2007; Freer and Rindi, 2013). Cell concentrations may vary in a range from 2.5 x106 to 10 x106 cells/ml (Maecker, 2004; Freer and Rindi, 2013a; https://www.bdbiosciences.com/content/bdb/paths/generate-tds-document.in.560751.pdf). Concentration of 1x106 cells/ml, 4x106 cells/ml and 8x106cells/ml were tested. Cell tritation have shown a higher functional response proportional to the cell concentration when exposed to a fixed concentration of stimulants. Cell concentration of 8 milions/ml was selected in order to obtain the higher percentage of IFN-γ-, IL-4- and IL-17-producing CD4+T cells. In conclusion the present protocol provides that, for a optimal optimal percentage of IFN-γ-, IL-4- and IL-17-producing CD4+T cells as assessed by flow cytometry (Table 1), PBMC in a concentration 8 milions/ml were stimulated with PMA 10 ng/ml and ionomycin 1 µg/ml, and cultured for 5 h in presence of brefeldin A 10 µg/ml according to the procedure described in detail below. References Baran, J., Kowalczyk, D., Ozog, M., Zembala, M., 2001. Three-color flow cytometry detection of intracellular cytokines in peripheral blood mononuclear cells: Comparative analysis of phorbol myristate acetate-ionomycin and phytohemagglutinin stimulation. Clin. Diagn. Lab. Immunol. 8, 303–313. https://doi.org/10.1128/CDLI.8.2.303-313.2001 Foster, B., Prussin, C., Liu, F., Whitmire, J.K., Whitton, J.L., 2007. Detection of intracellular cytokines by flow cytometry. Curr. Protoc. Immunol. Chapter 6. https://doi.org/10.1002/0471142735.im0624s78 Freer, G., Rindi, L., 2013. Intracellular cytokine detection by fluorescence-activated flow cytometry: Basic principles and recent advances. Methods 61, 30–38. https://doi.org/10.1016/j.ymeth.2013.03.035 Gupta, S., Maecker, H., 2015. Intracellular Cytokine Staining (ICS) on Human Lymphocytes or Peripheral Blood Mononuclear Cells (PBMCs). BIO-PROTOCOL 5. https://doi.org/10.21769/bioprotoc.1442 Maecker, H.T., 2004. Cytokine flow cytometry. Methods Mol. Biol. 263, 95–108. https://doi.org/10.1385/1-59259-773-4:095 https://www.bdbiosciences.com/content/bdb/paths/generate-tds-document.us.560751.pdf BEFORE STARTING with this procedure Moreover, work under laminar flow hood when you are processing samples from the beginning to the end of the culture. Make sure you are using, sterile culture medium and sterile plastic disposable as well.

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Induction of Autoimmune Thyroiditis by Depletion of CD4+CD25+ Regulatory T Cells in Thyroiditis-Resistant IL-17, But Not Interferon-γ Receptor, Knockout Nonobese Diabetic-H2h4 Mice

Endocrinology ◽

10.1210/en.2011-1356 ◽

2011 ◽

Vol 152 (11) ◽

pp. 4448-4454 ◽

Cited By ~ 20

Author(s):

Ichiro Horie ◽

Norio Abiru ◽

Hikaru Sakamoto ◽

Yoichiro Iwakura ◽

Yuji Nagayama

Keyword(s):

T Cells ◽

Regulatory T Cells ◽

Immune Responses ◽

Autoimmune Thyroiditis ◽

Th17 Cells ◽

Intracellular Cytokine Staining ◽

Th1 Cells ◽

Treg Depletion ◽

Nonobese Diabetic ◽

Ifn Γ

Iodine-induced experimental autoimmune thyroiditis in the nonobese diabetic (NOD)-H2h4 mouse is a prototype of animal models of Hashimoto's thyroiditis in humans. Recent studies have shown the resistance to thyroiditis of NOD-H2h4 mice genetically deficient for either IL-17 or interferon (IFN)-γ, implicating both of T helper type 1 (Th1) and Th17 immune responses in disease pathogenesis. However, we hypothesized that robust induction of a single arm of effector T cells (either Th1 or Th17) might be sufficient for inducing thyroiditis in NOD-H2h4 mice. To address this hypothesis, enhanced immune responses consisting of either Th1 or Th17 were induced by anti-CD25 antibody-mediated depletion of regulatory T cells (Treg) in thyroiditis-resistant IL-17 knockout (KO) or IFN-γ receptor (IFN-γR) KO, respectively, NOD-H2h4 mice. Depletion of Treg in IL-17 KO mice (i.e. Th1 enhancement) elicited antithyroglobulin autoantibodies and thyroiditis. Immunohistochemical analysis of the thyroid glands revealed the similar intrathyroidal lymphocyte infiltration patterns, with CD4+ T and CD19+ B cells being dominant between the wild-type and Treg-depleted IL-17 KO mice. In contrast, Treg-depleted IFN-γR KO mice remained thyroiditis resistant. Intracellular cytokine staining assays showed differentiation of Th1 cells in IL-17 KO mice but not of Th17 cells in IFN-γR KO mice. Our findings demonstrate that a robust Th1 immune response can by itself induce thyroiditis in otherwise thyroiditis-resistant IL-17 KO mice. Thus, unlike Th17 cells in IFN-γR KO mice, Th1 cells enhanced by Treg depletion can be sustained and induce thyroiditis.

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Preferential Accumulation of Activated Th1 Cells Not Only in Rheumatoid Arthritis But Also in Osteoarthritis Joints

The Journal of Rheumatology ◽

10.3899/jrheum.101355 ◽

2011 ◽

Vol 38 (8) ◽

pp. 1569-1575 ◽

Cited By ~ 36

Author(s):

HISAKATA YAMADA ◽

YASUHARU NAKASHIMA ◽

KEN OKAZAKI ◽

TARO MAWATARI ◽

JUN-ICHI FUKUSHI ◽

...

Keyword(s):

Rheumatoid Arthritis ◽

T Cells ◽

Cd4 T Cells ◽

Th17 Cells ◽

Mononuclear Cells ◽

Qualitative Difference ◽

Flow Cytometric Analysis ◽

Interferon Γ ◽

Th1 Cells ◽

Activation Markers

Objective.It was previously found that Th1 but not Th17 cells were predominant in the joints of rheumatoid arthritis (RA). To verify whether this is a unique feature of CD4 T cells in RA joints, we performed comparative flow cytometric analysis of CD4 T cells in RA and osteoarthritis (OA) joints.Methods.Mononuclear cells were isolated from peripheral blood (PB), synovial membrane (SM), and synovial fluid (SF) from a total of 18 RA and 12 OA patients. The expression of surface molecules and cytokine production of CD4 T cells was examined by a flow cytometer.Results.Most CD4 T cells in RA joints expressed memory/activation markers, such as CD45RO, HLA-DR, and CD69. CCR5 was highly expressed on CD4 T cells in SF but not in PB or SM. With regard to Th17-related molecules, CD4 T cells expressing CCR6 were not enriched in either SF or SM. In contrast, CD161-positive cells were abundant in the joint, many of which, however, produced interferon-γ but not interleukin 17A. Virtually all T cells in OA joints, although much less numerous than in RA joints, expressed activation markers. Th1 cells were predominant in both OA and RA joints, while there were a few Th17 cells. The frequency of Th17 cells in the joint tended to be lower in OA than RA.Conclusion.There was a quantitative but not qualitative difference in CD4 T cells, including the expression of activation markers and cytokine profiles, between RA and OA joints.

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Epigenetic reprogramming ameliorates type 1 diabetes by decreasing the generation of Th1 and Th17 subsets and restoring self-tolerance in CD4+ T cells

10.1101/2021.07.24.453657 ◽

2021 ◽

Author(s):

Vasu Patel ◽

Arathi Jayaraman ◽

Sundararajan Jayaraman

Keyword(s):

T Cells ◽

Th17 Cells ◽

Ex Vivo ◽

Autoimmune Diabetes ◽

Trichostatin A ◽

Nod Mice ◽

Th1 Cells ◽

Dependent Manner ◽

Self Tolerance ◽

Ifn Γ

The histone modifier Trichostatin A (TSA) ameliorated diabetes and repressed IFN-γ and IL-17A expression in prediabetic female NOD mice. Purified CD4+ cells could be polarized ex vivo into Th1 and Th17 subsets, which comparably transferred diabetes into NOD.scid mice. Polarized Th1 cells were devoid of IL-17A-producing cells and did not transdifferentiate into Th17 cells in an immunodeficient environment. However, Th17 cells had contaminant Th1 cells, which expressed IFN-γ upon adoptive transfer into lymphopenic recipients. Notably, TSA treatment abrogated the transfer of diabetes by CD4+ T-cells cultured under Th1 or Th17 polarizing conditions accompanied by the absence of Ifng and Il17a expression in NOD.scid recipients. Significantly, the histone modifier restored the ability of CD4+ but not CD8+ T-cells to undergo CD3-mediated apoptosis ex vivo in a caspase-dependent manner. Thus, the histone modifier afforded protection against autoimmune diabetes by negative regulation of signature lymphokines and restitution of self-tolerance in CD4+ T cells.

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Dulaglutide Modulates the Development of Tissue-Infiltrating Th1/Th17 Cells and the Pathogenicity of Encephalitogenic Th1 Cells in the Central Nervous System

International Journal of Molecular Sciences ◽

10.3390/ijms20071584 ◽

2019 ◽

Vol 20 (7) ◽

pp. 1584 ◽

Cited By ~ 1

Author(s):

Hsin-Ying Chiou ◽

Ming-Wei Lin ◽

Pi-Jung Hsiao ◽

Chun-Lin Chen ◽

Shiang Chiao ◽

...

Keyword(s):

Central Nervous System ◽

T Cells ◽

Nervous System ◽

Th17 Cells ◽

Mononuclear Cells ◽

Clinical Manifestations ◽

Vital Role ◽

Th1 Cells ◽

The Central Nervous System ◽

Glucagon Like Peptide 1

GLP-1 (glucagon-like peptide-1) has been reported to play a vital role in neuroprotection. Experimental autoimmune encephalomyelitis (EAE) is a well-established animal model widely used to study human multiple sclerosis, a chronic demyelination disease in the central nervous system (CNS). Recently, important studies have designated that the signaling axis of GLP-1 and its receptor controls the clinical manifestations and pathogenesis of EAE. However, it is elusive whether GLP-1 receptor signaling regulates the phenotype of autoreactive T cells in the CNS. We administered dulaglutide, a well-established GLP-1 receptor agonist (GLP-1 RA), to treat EAE mice prophylactically or semi-therapeutically and subsequently analyzed the mononuclear cells of the CNS. In this study, dulaglutide treatment significantly alleviates the clinical manifestations and histopathological outcomes of EAE. Dulaglutide decreases incidences of encephalitogenic Th1/Th17 cells and Th1 granulocyte-macrophage-colony-stimulating factor (GM-CSF) expression in the CNS. Administration of dulaglutide failed to control the chemotactic abilities of encephalitogenic Th1 and Th17 cells; however, prophylactic treatment considerably decreased the populations of dendritic cells and macrophages in the CNS parenchyma. These results obtained indicate that dulaglutide modulates the differentiation of encephalitogenic Th1/Th17 and the pathogenicity of Th1 cells by influencing antigen presenting cells quantities, providing mechanism insight on T cells regulation in ameliorating EAE by GLP-1.

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Increased Th1/Th17 Responses Contribute to Low-Grade Inflammation in Age-Related Macular Degeneration

Cellular Physiology and Biochemistry ◽

10.1159/000484907 ◽

2017 ◽

Vol 44 (1) ◽

pp. 357-367 ◽

Cited By ~ 10

Author(s):

Jiajia Chen ◽

Wenzhan Wang ◽

Qiuming Li

Keyword(s):

T Cells ◽

Macular Degeneration ◽

Cd4 T Cells ◽

Th17 Cells ◽

Retinal Pigment ◽

Age Related Macular Degeneration ◽

Low Grade ◽

Th1 Cells ◽

Age Related ◽

Ifn Γ

Background/Aims: Age-related macular degeneration (AMD) is the primary cause of senior blindness in developed countries. Mechanisms underlying initiation and development of AMD remained known. Methods: We examined the CD4+ T cell compartments and their functions in AMD patients. Results: AMD patients presented significantly higher frequencies of interferon (IFN)-γ-expressing and interleukin (IL)-17-expressing CD4+ T cells than healthy controls. The levels of IFN-γ and IL-17 expression by CD4+ T cells were significantly higher in AMD patients. These IFN-γ-expressing Th1 cells and IL-17-expressing Th17 cells could be selectively enriched by surface CCR3+ and CCR4+CCR6+ expression, respectively. Th1 and Th17 cells from AMD patients promoted the differentiation of monocytes toward M1 macrophages, which were previously associated with retinal damage. Th1 and Th17 cells also increased the level of MHC class I expression in human retinal pigment epithelial (RPE)-1 cells, while Th1 cells increased the frequency of MHC class II-expressing RPE-1 cells. These proinflammatory effects were partly, but not entirely, induced by the secretion of IFN-γ and IL-17. Conclusions: This study demonstrated an enrichment of Th1 cells and Th17 cells in AMD patients. These Th1 and Th17 cells possessed proinflammatory roles in an IFN-γ- and IL-17-dependent fashion, and could potentially serve as therapeutic targets.

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Donor Th17 and Th1 Contribute to Chronic Graft-Versus-Host Disease

Blood ◽

10.1182/blood.v116.21.727.727 ◽

2010 ◽

Vol 116 (21) ◽

pp. 727-727

Author(s):

Hisakazu Nishimori ◽

Haruko Sugiyama ◽

Koichiro Kobayashi ◽

Yoshiko Yamasuji ◽

Sachiyo Kadohisa ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

T Cells ◽

Th1 Cells ◽

Flow Cytometry Analysis ◽

Spleen Cells ◽

Graft Versus Host ◽

Donor T Cells ◽

Ifn Γ ◽

Deficient Mice

Abstract Abstract 727 Chronic graft-versus-host disease (cGVHD) remains a major cause of late death and morbidity after allogeneic hematopoietic cell transplantation, and the treatment of cGVHD remains challenging. All-trans retinoic acid (ATRA), a potent derivative of vitamin A, can regulate immune responses. Am80, which has biological activity approximately 10 times more potent than that of ATRA by binding to RARα and RARβ, but not RARγ, also reduces the severity and progression of immune disease models, including contact dermatitis, collagen-induced arthritis, allergic encephalomyelitis, and atherosclerosis models. Am80 has been reported to suppress the differentiation of Th17 and Th1 cells. Previously, we demonstrated that the administration of ATRA or Am80, significantly decreased skin fibrosis and alopecia, a dominant feature of cutaneous cGVHD, using a well-characterized experimental model of cGVHD: B10.D2 (H−2d) → BALB/c (H−2d). Flow cytometry analysis of the peripheral lymph nodes (PLNs) on day 16 showed significant reductions in Th1, Th17, and Foxp3+ regulatory T cells (Tregs) in Am80-treated recipients, as compared to controls, whereas no reduction in Th2 cells was observed (Nishimori et al., 2009 ASH). To further investigate the roles of Th17 cells and other Th subsets (Th1 and Th2) in cGVHD, we first assessed the kinetics of Th cytokines in the B10.D2 (H-2d) → BALB/c (H-2d) model of cGVHD, which reflects the clinical and pathological symptoms of human cGVHD. BALB/c mice were subjected to sublethal irradiation (6.75 Gy) and injected with 8 × 106 T-cell-depleted bone marrow cells and 8 × 106 spleen cells or 2 × 106 CD90+ spleen T cells from B10.D2 mice. In the early phase (day 14) of post bone marrow transplant (BMT), donor T cells produced significantly less IL-17 in the PLNs of recipients in allogeneic BMT than in syngeneic BMT (1.68±0.09% vs. 4.30±0.53%, p<0.01), while in the late phase (day 28) donor T cells that produced IL-17 were increased significantly in allogeneic recipients, compared with syngeneic controls (7.76±0.27% vs. 1.61±0.16%, p < 0.005). Both IL-17−/IFN-γ+ and IL-17/IFN-γ double positive cells (Th17/Th1 cells) were constantly detected more frequently in allogeneic recipients (p<0.05). Next, we evaluated whether Th17 contributes in a model utilizing IL-17-deficient mice on a B10.D2 background as donor mice; these mice were backcrossed for more than six generations from the original knock-out (KO) mice. Upon transfer of IL-17−/− B10.D2 donor T cells in allogeneic BMT models, skin cGVHD was significantly ameliorated, as compared to recipients of wild-type (WT) T cells, as shown in Figure (p=0.02). Histopathological examination of the skin showed significantly reduced cGVHD pathology in recipients of IL-17−/− donors (3.17±1.09 vs. 8.50±0.84, p=0.006). Flow cytometry analysis of the PLNs, mesenteric lymph nodes (MLNs), and spleen cells in the early phase (day 14) showed no differences in Th1, Th2, and Tregs, while recipients of IL-17−/− had significantly fewer Th1 (p<0.05) and Th17/Th1 (p<0.01) cells on day 35. Furthermore, IFN-γ deficient mice on a B10.D2 background were also backcrossed from the original KO mice and cGVHD was evaluated. BMT from IFN-γ−/− donors significantly improved the clinical cGVHD score, compared with WT donors (p<0.05). Allogeneic recipients given anti-IFN-γ mAb (500 μg/mouse) on days 0, 5, 10, and 15 post BMT had significantly less severe cGVHD, compared with control antibody recipients (p<0.05). Taken together, Th17, especially in the late-chronic phase, and Th1 contribute to the development of cGVHD by promoting the production of proinflammatory cytokines. Targeting Th17 and Th1 may be a promising strategy for preventing and treating cGVHD. Disclosures: No relevant conflicts of interest to declare.

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The Effects of Sishen Wan on T Cell Responses in Mice Models of Ulcerative Colitis Induced by Dextran Sodium Sulfate

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2021/9957709 ◽

2021 ◽

Vol 2021 ◽

pp. 1-11

Author(s):

Ke Li ◽

Jiamin Dong ◽

Dongyu Ge ◽

Mengjia Li ◽

Hehe Ye ◽

...

Keyword(s):

Ulcerative Colitis ◽

T Cells ◽

T Cell ◽

Sodium Sulfate ◽

Th17 Cells ◽

T Cell Responses ◽

Dextran Sodium Sulfate ◽

Th1 Cells ◽

Cell Responses ◽

Ifn Γ

Currently, it is unclear whether Sishen Wan (SSW) could modulate the balance of Th1 cells, Th17 cells, and Tregs and we evaluated the effects of SSW on T cell responses in mice models of ulcerative colitis (UC). The mice models of acute UC (4% dextran sodium sulfate (DSS), 8 days) and chronic UC (3% DSS, 16 days) with SSW were assayed. Colon tissues were collected for immunohistochemical analysis, enzyme linked immunosorbent assay (ELISA), and flow cytometry (FCM). The expressions of cytokines associated with Tregs, transcription factors of Th17 cells, the frequencies of Th1 cells, Th17 cells, and Tregs, and the functional plasticity of Th17 cells were detected. The frequency of IFN-γ+ T cells was not changed significantly with SSW treatment in acute DSS. In chronic models, the frequency of IFN-γ+ T cells was downregulated with SSW. Meanwhile, the levels of RORγt and the frequency of IL-17A+ Th17 cells showed no significant differences after SSW treatment. Despite no significant effect on the transdifferentiation of Th17 cells in chronic UC models, SSW transdifferentiated Th17 cells into IL-10+ Th17 cells and downregulated IFN-γ+ Th17 cells/IL-10+ Th17 cells in acute DSS. Moreover, there were no significant changes of cytokines secreted by Tregs in acute DSS after SSW treatment, but SSW facilitated the expressions of IL-10 and IL-35, as well as development of IL-10+ Tregs in chronic DSS. SSW showed depressive effects on the immunoreaction of Th17 cells and might promote the conversion of Th17 cells into IL-10+ Th17 cells in acute UC, while it inhibited the excessive reaction of Th1 cells, facilitated the development of Tregs, and enhanced the anti-inflammatory effects in chronic UC.

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