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1.
Hum Mol Genet ; 33(3): 211-223, 2024 Jan 20.
Article in English | MEDLINE | ID: mdl-37819629

ABSTRACT

Duchenne muscular dystrophy (DMD) is a progressive disabling X-linked recessive disorder that causes gradual and irreversible loss of muscle, resulting in early death. The corticosteroids prednisone/prednisolone and deflazacort are used to treat DMD as the standard of care; however, only deflazacort is FDA approved for DMD. The novel atypical corticosteroid vamorolone is being investigated for treatment of DMD. We compared the pharmaceutical properties as well as the efficacy and safety of the three corticosteroids across multiple doses in the B10-mdx DMD mouse model. Pharmacokinetic studies in the mouse and evaluation of p-glycoprotein (P-gP) efflux in a cellular system demonstrated that vamorolone is not a strong P-gp substrate resulting in measurable central nervous system (CNS) exposure in the mouse. In contrast, deflazacort and prednisolone are strong P-gp substrates. All three corticosteroids showed efficacy, but also side effects at efficacious doses. After dosing mdx mice for two weeks, all three corticosteroids induced changes in gene expression in the liver and the muscle, but prednisolone and vamorolone induced more changes in the brain than did deflazacort. Both prednisolone and vamorolone induced depression-like behavior. All three corticosteroids reduced endogenous corticosterone levels, increased glucose levels, and reduced osteocalcin levels. Using micro-computed tomography, femur bone density was decreased, reaching significance with prednisolone. The results of these studies indicate that efficacious doses of vamorolone, are associated with similar side effects as seen with other corticosteroids. Further, because vamorolone is not a strong P-gp substrate, vamorolone distributes into the CNS increasing the potential CNS side-effects.


Subject(s)
Muscular Dystrophy, Duchenne , Prednisolone , Pregnadienediols , Pregnenediones , Animals , Mice , Prednisolone/therapeutic use , X-Ray Microtomography , Mice, Inbred mdx , Muscular Dystrophy, Duchenne/drug therapy , Muscular Dystrophy, Duchenne/genetics , Corticosterone/therapeutic use , Pharmaceutical Preparations
2.
Immunotherapy ; 13(12): 977-987, 2021 08.
Article in English | MEDLINE | ID: mdl-34184542

ABSTRACT

Aim: Graft-versus-host disease (GvHD) is a major complication arising in patients undergoing allogenic hematopoietic stem cell transplantation. Material & methods: We tested ruxolitinib (a selective JAK1/2 inhibitor) efficacy in three different preclinical models of GvHD. Results: Ruxolitinib, at doses that mimic clinically achievable human JAK/signal transducers and activators of transcription target inhibition, significantly reduced alloreactive T-cell activation and infiltration in the lung and skin, leading to improved outcomes in two experimental models of steroid-refractory acute and chronic GvHD. Additionally, we describe a novel humanized GvHD model in which immunodeficient NOG animals are engineered to produce human IL-15 to facilitate enhanced T- and NK cell engraftment, leading to severe GvHD. Conclusion: Ruxolitinib treatment ameliorated disease symptoms resulting from targeted immune modulation via JAK/signal transducers and activators of transcription signaling inhibition.


Subject(s)
Graft vs Host Disease/pathology , Hematopoietic Stem Cell Transplantation/adverse effects , Janus Kinase Inhibitors/pharmacology , Nitriles/pharmacology , Pyrazoles/pharmacology , Pyrimidines/pharmacology , Animals , Disease Models, Animal , Heterografts , Humans , Janus Kinase 1/antagonists & inhibitors , Janus Kinase 2/antagonists & inhibitors , Male , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Transgenic
3.
Front Pharmacol ; 12: 650295, 2021.
Article in English | MEDLINE | ID: mdl-33981229

ABSTRACT

Hyperinflammatory syndromes comprise a heterogeneous group of disorders characterized by severe inflammation, multiple organ dysfunction, and potentially death. In response to antigenic stimulus (e.g., SARS-CoV-2 infection), overactivated CD8+ T-cells and macrophages produce high levels of proinflammatory cytokines, such as IFN-γ, TNF-α, IL-6, and IL-12. Multiple inflammatory mediators implicated in hyperinflammatory syndromes utilize the Janus kinase-signal transducers and activators of transcription (JAK-STAT) cascade to propagate their biological function. Our findings demonstrate that oral ruxolitinib dosing designed to mimic clinically relevant JAK-STAT pathway inhibition significantly reduces the harmful consequences of immune overactivation in multiple hyperinflammatory models. In contrast to monoclonal antibody therapies targeting a single cytokine, ruxolitinib effectively downregulates the functional effect of multiple cytokines implicated in hyperinflammatory states, without broad immunosuppression.

4.
Clin Cancer Res ; 26(23): 6299-6309, 2020 12 01.
Article in English | MEDLINE | ID: mdl-32998963

ABSTRACT

PURPOSE: T cells engineered to express a chimeric antigen receptor (CAR) are a promising cancer immunotherapy. Such targeted therapies have shown long-term relapse-free survival in patients with B-cell leukemia and lymphoma. However, cytokine release syndrome (CRS) represents a serious, potentially life-threatening side effect often associated with CAR T-cell therapy. CRS manifests as a rapid (hyper)immune reaction driven by excessive inflammatory cytokine release, including IFNγ and IL6. EXPERIMENTAL DESIGN: Many cytokines implicated in CRS are known to signal through the JAK-STAT pathway. Here we study the effect of blocking JAK pathway signaling on CAR T-cell proliferation, antitumor activity, and cytokine levels in in vitro and in vivo models. RESULTS: We report that itacitinib, a potent, selective JAK1 inhibitor, was able to significantly and dose-dependently reduce levels of multiple cytokines implicated in CRS in several in vitro and in vivo models. Importantly, we also report that at clinically relevant doses that mimic human JAK1 pharmacologic inhibition, itacitinib did not significantly inhibit proliferation or antitumor killing capacity of three different human CAR T-cell constructs (GD2, EGFR, and CD19). Finally, in an in vivo model, antitumor activity of CD19-CAR T cells adoptively transferred into CD19+ tumor-bearing immunodeficient animals was unabated by oral itacitinib treatment. CONCLUSIONS: Together, these data suggest that itacitinib has potential as a prophylactic agent for the prevention of CAR T cell-induced CRS, and a phase II clinical trial of itacitinib for prevention of CRS induced by CAR T-cell therapy has been initiated (NCT04071366).


Subject(s)
Azetidines/pharmacology , Cytokine Release Syndrome/drug therapy , Cytokines/antagonists & inhibitors , Immunotherapy, Adoptive/adverse effects , Isonicotinic Acids/pharmacology , Janus Kinase 1/antagonists & inhibitors , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Animals , Apoptosis , Cell Proliferation , Cytokine Release Syndrome/etiology , Cytokine Release Syndrome/pathology , Female , Humans , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Inbred NOD , Mice, SCID , Precursor Cell Lymphoblastic Leukemia-Lymphoma/immunology , Precursor Cell Lymphoblastic Leukemia-Lymphoma/pathology , Tumor Cells, Cultured , Xenograft Model Antitumor Assays
5.
Nutr Res ; 82: 25-33, 2020 10.
Article in English | MEDLINE | ID: mdl-32949953

ABSTRACT

Targeting gut microbiota with probiotics has emerged as a promising nutritional approach for the prevention of obesity and metabolic syndrome. Cultured dairy products can be effectively employed for the delivery of probiotics to the gut as well as for the support of growth and survival of probiotic bacteria. The purpose of this study was to characterize the effects of probiotic-enriched pasteurized milk and dairy products (Greek-style yogurt and cottage cheese) of different origins (cow, goat, and camel) on taxonomic composition of the mouse gut microbiota. We hypothesized that cultured dairy products can be an effective vector for the delivery of probiotics to the gut because of its nutritional value, acidic nature, and long shelf-life. Mice were fed a standard low fat, plant polysaccharide-rich (LF/PP) diet supplemented with the probiotic-enriched milk and dairy products for 5 weeks. Next generation sequencing of DNA from mouse fecal samples was used to characterize the bacterial relative abundance. Mice fed a diet supplemented with camel milk demonstrated characteristic changes in the gut microbiota, which included an increase in relative abundance of order Clostridiales and genus Anaerostipes. Mice fed a diet supplemented with the probiotic-enriched cow cheese exhibited an increase in the relative abundance of order Clostridiales, family Ruminococcaceae, and family Lachnospiraceae. The results obtained and their bioinformatics analysis support the conclusion that camel milk and the probiotic cow cheese induce changes in the mouse gut microbiota, which can be characterized as potentially beneficial to health compared to the changes associated with a standard diet. These findings imply that probiotic-enriched milk and dairy products can be highly effective for the delivery and support of probiotic bacteria of the gut.


Subject(s)
Bacteria/classification , Dairy Products/microbiology , Diet , Gastrointestinal Microbiome , Milk/microbiology , Probiotics , Animals , Bacteria/growth & development , Bacteria/isolation & purification , Camelus , Cattle , Cheese/microbiology , Clostridiales/classification , Clostridiales/growth & development , Clostridiales/isolation & purification , Feces/microbiology , Female , Goats , Mice , Mice, Inbred C57BL , Weight Gain , Yogurt/microbiology
6.
Nutrients ; 12(1)2020 Jan 15.
Article in English | MEDLINE | ID: mdl-31952249

ABSTRACT

Targeting gut microbiota with synbiotics (probiotic supplements containing prebiotic components) is emerging as a promising intervention in the comprehensive nutritional approach to reducing obesity. Weight loss resulting from low-carbohydrate high-protein diets can be significant but has also been linked to potentially negative health effects due to increased bacterial fermentation of undigested protein within the colon and subsequent changes in gut microbiota composition. Correcting obesity-induced disruption of gut microbiota with synbiotics can be more effective than supplementation with probiotics alone because prebiotic components of synbiotics support the growth and survival of positive bacteria therein. The purpose of this placebo-controlled intervention clinical trial was to evaluate the effects of a synbiotic supplement on the composition, richness and diversity of gut microbiota and associations of microbial species with body composition parameters and biomarkers of obesity in human subjects participating in a weight loss program. The probiotic component of the synbiotic used in the study contained Lactobacillus acidophilus, Bifidobacterium lactis, Bifidobacterium longum, and Bifidobacterium bifidum and the prebiotic component was a galactooligosaccharide mixture. The results showed no statistically significant differences in body composition (body mass, BMI, body fat mass, body fat percentage, body lean mass, and bone mineral content) between the placebo and synbiotic groups at the end of the clinical trial (3-month intervention, 20 human subjects participating in weight loss intervention based on a low-carbohydrate, high-protein, reduced energy diet). Synbiotic supplementation increased the abundance of gut bacteria associated with positive health effects, especially Bifidobacterium and Lactobacillus, and it also appeared to increase the gut microbiota richness. A decreasing trend in the gut microbiota diversity in the placebo and synbiotic groups was observed at the end of trial, which may imply the effect of the high-protein low-carbohydrate diet used in the weight loss program. Regression analysis performed to correlate abundance of species following supplementation with body composition parameters and biomarkers of obesity found an association between a decrease over time in blood glucose and an increase in Lactobacillus abundance, particularly in the synbiotic group. However, the decrease over time in body mass, BMI, waist circumstance, and body fat mass was associated with a decrease in Bifidobacterium abundance. The results obtained support the conclusion that synbiotic supplement used in this clinical trial modulates human gut microbiota by increasing abundance of potentially beneficial microbial species.


Subject(s)
Body Composition/drug effects , Gastrointestinal Microbiome/drug effects , Obesity/drug therapy , Synbiotics , Weight Loss/drug effects , Adult , Bifidobacterium , Diet, Reducing , Dietary Supplements , Female , Humans , Lactobacillus , Male , Middle Aged
7.
Curr Protoc Pharmacol ; 86(1): e66, 2019 09.
Article in English | MEDLINE | ID: mdl-31539924

ABSTRACT

Chimeric antigen receptor (CAR) T-cell therapy against tumor antigens involves a recombinant immunoreceptor that combines an antibody-derived targeting fragment with signaling domains capable of activating T cells and fusion of this receptor domain to a costimulatory domain (typically CD28 or 4-1BB). Clinical trials of CAR T-cell therapeutics targeting CD19 antigens for relapsed or refractory B-cell malignancies have shown unparalleled results and consequently have recently been approved by the U.S. Food and Drug Administration. However, the lack of efficacy beyond B-cell malignancies, the emergence of resistance to CAR T-cell therapy due to loss of the antigenic epitope, and severe cases of cytokine release syndrome and neurotoxicity necessitate further preclinical studies. As it is very complicated to develop a single animal model that would replicate the complexity of the clinical scenario, this article focuses on transgenic models used to study human tumor-associated antigens in an immunocompetent model. © 2019 by John Wiley & Sons, Inc.


Subject(s)
Disease Models, Animal , Immunotherapy, Adoptive , Mice, Transgenic , Neoplasms/therapy , Animals , Humans , Lentivirus , Transduction, Genetic
8.
Int J Mol Sci ; 20(19)2019 Sep 26.
Article in English | MEDLINE | ID: mdl-31561568

ABSTRACT

Type 1 diabetes mellitus (T1D) is an autoimmune illness that affects millions of patients worldwide. The main characteristic of this disease is the destruction of pancreatic insulin-producing beta cells that occurs due to the aberrant activation of different immune effector cells. Currently, T1D is treated by lifelong administration of novel versions of insulin that have been developed recently; however, new approaches that could address the underlying mechanisms responsible for beta cell destruction have been extensively investigated. The strategies based on immunotherapies have recently been incorporated into a panel of existing treatments for T1D, in order to block T-cell responses against beta cell antigens that are very common during the onset and development of T1D. However, a complete preservation of beta cell mass as well as insulin independency is still elusive. As a result, there is no existing T1D targeted immunotherapy able to replace standard insulin administration. Presently, a number of novel therapy strategies are pursuing the goals of beta cell protection and normoglycemia. In the present review we explore the current state of immunotherapy in T1D by highlighting the most important studies in this field, and envision novel strategies that could be used to treat T1D in the future.


Subject(s)
Diabetes Mellitus, Type 1/immunology , Diabetes Mellitus, Type 1/therapy , Immunotherapy , Animals , Antigens/immunology , Autoimmunity , B-Lymphocytes/immunology , B-Lymphocytes/metabolism , Cell- and Tissue-Based Therapy , Cytokines/metabolism , Diabetes Mellitus, Type 1/metabolism , Humans , Immunotherapy/methods , Inflammation Mediators/metabolism , Insulin/metabolism , Insulin-Secreting Cells/cytology , Insulin-Secreting Cells/immunology , Insulin-Secreting Cells/metabolism , Mice , Stem Cell Transplantation , T-Lymphocytes/immunology , T-Lymphocytes/metabolism
10.
J Immunol ; 196(12): 5036-46, 2016 06 15.
Article in English | MEDLINE | ID: mdl-27194787

ABSTRACT

Regulatory T cells (Tregs) induced during autoimmunity often become quiescent and unable to resolve disease, suggesting inadequate activation. Resolution of established experimental autoimmune encephalomyelitis (EAE) can be achieved with myelin oligodendrocyte glycoprotein (MOG) fused to reovirus protein σ1 (MOG-pσ1), which activates Tregs, restoring protection, but requiring other regulatory cells to revitalize them. B cells have a dichotomous role in both the pathogenesis and recovery from EAE. Although inflammatory B cells contribute to EAE's pathogenesis, treatment of EAE mice with MOG-pσ1, but not OVA-pσ1, resulted in an influx of IL-10-producing B220(+)CD5(+) B regulatory cells (Bregs) enabling Tregs to recover their inhibitory activity, and in turn, leading to the rapid amelioration of EAE. These findings implicate direct interactions between Bregs and Tregs to facilitate this recovery. Adoptive transfer of B220(+)CD5(-) B cells from MOG-pσ1-treated EAE or Bregs from PBS-treated EAE mice did not resolve disease, whereas the adoptive transfer of MOG-pσ1-induced B220(+)CD5(+) Bregs greatly ameliorated EAE. MOG-pσ1-, but not OVA-pσ1-induced IL-10-producing Bregs, expressed elevated levels of B and T lymphocyte attenuator (BTLA) relative to CD5(-) B cells, as opposed to Tregs or effector T (Teff) cells, whose BTLA expression was not affected. These induced Bregs restored EAE Treg function in a BTLA-dependent manner. BTLA(-/-) mice showed more pronounced EAE with fewer Tregs, but upon adoptive transfer of MOG-pσ1-induced BTLA(+) Bregs, BTLA(-/-) mice were protected against EAE. Hence, this evidence shows the importance of BTLA in activating Tregs to facilitate recovery from EAE.


Subject(s)
B-Lymphocytes, Regulatory/immunology , Encephalomyelitis, Autoimmune, Experimental/immunology , Receptors, Immunologic/immunology , T-Lymphocytes, Regulatory/immunology , Adoptive Transfer , Animals , B-Lymphocytes, Regulatory/drug effects , B-Lymphocytes, Regulatory/physiology , CD5 Antigens/genetics , CD5 Antigens/immunology , Disease Models, Animal , Encephalomyelitis, Autoimmune, Experimental/physiopathology , Interleukin-10/biosynthesis , Interleukin-10/immunology , Leukocyte Common Antigens/genetics , Leukocyte Common Antigens/immunology , Lymphocyte Activation , Mice , Mice, Inbred C57BL , Myelin-Oligodendrocyte Glycoprotein/administration & dosage , Myelin-Oligodendrocyte Glycoprotein/immunology , Receptors, Immunologic/deficiency , Receptors, Immunologic/genetics , T-Lymphocytes, Regulatory/pathology , T-Lymphocytes, Regulatory/physiology
11.
Oncoimmunology ; 2(12): e27009, 2013 Dec 01.
Article in English | MEDLINE | ID: mdl-24498563

ABSTRACT

Peptide vaccines derived from CD8+ T-cell epitopes have shown variable efficacy in cancer patients. Thus, some peptide vaccines are capable of activating CD8+ T-cell responses, even in the absence of CD4+ T-cell epitopes or dendritic cell (DC)-activating adjuvants. However, the mechanisms underlying the clinical activity of these potent peptides are poorly understood. Using CT26 and ovalbumin-expressing B16 murine allograft tumor models, we found that the antitumor effect of helper cell-independent CD8 T-cell peptide vaccines is inhibited by the blockade of CD40 ligand (CD40L) in vivo. Furthermore, in vitro stimulation with antigenic peptides of cells derived from immunized mice induced the expression of CD40L on the surface of CD8+ T cells and fostered DC maturation, an effect that was partially inhibited by CD40L-blocking antibodies. Interestingly, CD40L blockade also inhibited CD8+ T-cell responses, even in the presence of fully mature DCs, suggesting a role for CD40L not only in promoting DC maturation but also in mediating CD8+ T-cell co-stimulation. Importantly, these potent peptides share features with bona fide CD4 epitopes, since they foster responses against less immunogenic CD8+ T-cell epitopes in a CD40L-dependent manner. The analysis of peptides used for the vaccination of cancer patients in clinical trials showed that these peptides also induce the expression of CD40L on the surface of CD8+ T cells. Taken together, these results suggest that CD40L expression induced by potent CD8+ T-cell epitopes can activate antitumor CD8+ T-cell responses, potentially amplifying the immunological responses to less immunogenic CD8+ T-cell epitopes and bypassing the requirement for CD4+ helper T cells in vaccination protocols.

12.
PLoS One ; 7(12): e50221, 2012.
Article in English | MEDLINE | ID: mdl-23226513

ABSTRACT

Lymphocytes are a key component of the immune system and their differentiation and function are directly influenced by cancer. We examined peripheral blood lymphocyte (PBL) gene expression as a biomarker of illness and treatment effect using the Affymetrix Human Gene ST1 platform in patients with metastatic renal cell carcinoma (mRCC) who received combined treatment with IL-2, interferon-?-2a and dendritic cell vaccine. We examined gene expression, cytokine levels in patient serum and lymphocyte subsets as determined by flow cytometry (FCM). Pre-treatment PBLs from patients with mRCC exhibit a gene expression profile and serum cytokine profile consistent with inflammation and proliferation not found in healthy donors (HD). PBL gene expression from patients with mRCC showed increased mRNA of genes involved with T-cell and T(REG)-cell activation pathways, which was also reflected in lymphocyte subset distribution. Overall, PBL gene expression post-treatment (POST) was not significantly different than pre-treatment (PRE). Nevertheless, treatment related changes in gene expression (post-treatment minus pre-treatment) revealed an increased expression of T-cell and B-cell receptor signaling pathways in responding (R) patients compared to non-responding (NR) patients. In addition, we observed down-regulation of T(REG)-cell pathways post-treatment in R vs. NR patients. While exploratory in nature, this study supports the hypothesis that enhanced inflammatory cytotoxic pathways coupled with blunting of the regulatory pathways is necessary for effective anti-cancer activity associated with immune therapy. This type of analysis can potentially identify additional immune therapeutic targets in patients with mRCC.


Subject(s)
Cancer Vaccines/therapeutic use , Carcinoma, Renal Cell/genetics , Dendritic Cells/immunology , Gene Expression Profiling , Interferon-alpha/therapeutic use , Interleukin-2/therapeutic use , Kidney Neoplasms/genetics , Lymphocytes/metabolism , Carcinoma, Renal Cell/blood , Carcinoma, Renal Cell/therapy , Cluster Analysis , Cytokines/blood , Female , Flow Cytometry , Humans , Kidney Neoplasms/blood , Kidney Neoplasms/therapy , Lymphocyte Subsets , Male , Middle Aged , Oligonucleotide Array Sequence Analysis , Real-Time Polymerase Chain Reaction
13.
J Autoimmun ; 37(4): 328-41, 2011 Dec.
Article in English | MEDLINE | ID: mdl-22018711

ABSTRACT

Natural killer (NK) cells and dendritic cells (DCs) have been shown to link the innate and adaptive immune systems. Likewise, a new innate cell subset, interferon-producing killer DCs (IKDCs), shares phenotypic and functional characteristics with both DCs and NK cells. Here, we show IKDCs play an essential role in the resolution of experimental autoimmune encephalomyelitis (EAE) upon treatment with the tolerizing agent, myelin oligodendrocyte glycoprotein (MOG), genetically fused to reovirus protein σ1 (termed MOG-pσ1). Activated IKDCs were recruited subsequent MOG-pσ1 treatment of EAE, and disease resolution was abated upon NK1.1 cell depletion. These IKDCs were able to kill activated CD4(+) T cells and mature dendritic DCs, thus, contributing to EAE remission. In addition, IKDCs were responsible for MOG-pσ1-mediated MOG-specific regulatory T cell recruitment to the CNS. The IKDCs induced by MOG-pσ1 expressed elevated levels of HVEM for interactions with cognate ligand-positive cells: LIGHT(+) NK and T(eff) cells and BTLA(+) B cells. Further characterization revealed these activated IKDCs being MHC class II(high), and upon their adoptive transfer (CD11c(+)NK1.1(+)MHC class II(high)), IKDCs, but not CD11c(+)NK1.1(+)MHC class II(intermediate/low) (unactivated) cells, conferred protection against EAE. These activated IKDCs showed enhanced CD107a, PD-L1, and granzyme B expression and could present OVA, unlike unactivated IKDCs. Thus, these results demonstrate the interventional potency induced HVEM(+) IKDCs to resolve autoimmune disease.


Subject(s)
Dendritic Cells/metabolism , Encephalomyelitis, Autoimmune, Experimental/immunology , Encephalomyelitis, Autoimmune, Experimental/therapy , Myelin Proteins/metabolism , Viral Proteins/metabolism , Adoptive Transfer , Animals , Antigens, Ly/metabolism , Cytotoxicity, Immunologic , Dendritic Cells/immunology , Dendritic Cells/pathology , Disease Progression , Encephalomyelitis, Autoimmune, Experimental/pathology , Encephalomyelitis, Autoimmune, Experimental/physiopathology , Immune Tolerance , Interferons/metabolism , Lymphocyte Activation , Mice , Myelin Proteins/genetics , Myelin Proteins/immunology , Myelin-Oligodendrocyte Glycoprotein , NK Cell Lectin-Like Receptor Subfamily B/metabolism , Receptors, Tumor Necrosis Factor, Member 14/metabolism , Recombinant Fusion Proteins/administration & dosage , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/immunology , Recombinant Fusion Proteins/metabolism , Recurrence , T-Lymphocyte Subsets/immunology , T-Lymphocyte Subsets/metabolism , T-Lymphocyte Subsets/pathology , T-Lymphocytes, Regulatory/immunology , T-Lymphocytes, Regulatory/metabolism , T-Lymphocytes, Regulatory/pathology , Viral Proteins/genetics , Viral Proteins/immunology
14.
PLoS One ; 6(7): e21978, 2011.
Article in English | MEDLINE | ID: mdl-21765931

ABSTRACT

γδ T cells have been postulated to act as a first line of defense against infectious agents, particularly intracellular pathogens, representing an important link between the innate and adaptive immune responses. Human γδ T cells expand in the blood of brucellosis patients and are active against Brucella in vitro. However, the role of γδ T cells in vivo during experimental brucellosis has not been studied. Here we report TCRδ(-/-) mice are more susceptible to B. abortus infection than C57BL/6 mice at one week post-infection as measured by splenic colonization and splenomegaly. An increase in TCRγδ cells was observed in the spleens of B. abortus-infected C57BL/6 mice, which peaked at two weeks post-infection and occurred concomitantly with diminished brucellae. γδ T cells were the major source of IL-17 following infection and also produced IFN-γ. Depletion of γδ T cells from C57BL/6, IL-17Rα(-/-), and GMCSF(-/-) mice enhanced susceptibility to B. abortus infection although this susceptibility was unaltered in the mutant mice; however, when γδ T cells were depleted from IFN-γ(-/-) mice, enhanced susceptibility was observed. Neutralization of γδ T cells in the absence of TNF-α did not further impair immunity. In the absence of TNF-α or γδ T cells, B. abortus-infected mice showed enhanced IFN-γ, suggesting that they augmented production to compensate for the loss of γδ T cells and/or TNF-α. While the protective role of γδ T cells was TNF-α-dependent, γδ T cells were not the major source of TNF-α and activation of γδ T cells following B. abortus infection was TNF-α-independent. Additionally, bovine TCRγδ cells were found to respond rapidly to B. abortus infection upon co-culture with autologous macrophages and could impair the intramacrophage replication of B. abortus via IFN-γ. Collectively, these results demonstrate γδ T cells are important for early protection to B. abortus infections.


Subject(s)
Brucella abortus/immunology , Brucellosis, Bovine/immunology , Brucellosis/immunology , Immunity, Innate/immunology , Receptors, Antigen, T-Cell, gamma-delta/immunology , T-Lymphocytes/immunology , Animals , Brucella abortus/growth & development , Brucellosis/prevention & control , Brucellosis, Bovine/prevention & control , Cattle , Cell Proliferation , Gene Expression Profiling , Granulocyte-Macrophage Colony-Stimulating Factor/immunology , Humans , Interferon-gamma/biosynthesis , Interleukin-17/biosynthesis , Lymphocyte Subsets/immunology , Macrophages/metabolism , Macrophages/microbiology , Mice , Mice, Inbred C57BL , Organ Size , Receptors, Antigen, T-Cell, gamma-delta/deficiency , Spleen/immunology , Spleen/microbiology , Spleen/pathology , Tumor Necrosis Factor-alpha/immunology
15.
J Leukoc Biol ; 90(6): 1043-54, 2011 Dec.
Article in English | MEDLINE | ID: mdl-21693591

ABSTRACT

Human IBD, including UC and Crohn's disease, is characterized by a chronic, relapsing, and remitting condition that exhibits various features of immunological inflammation and affects at least one/1000 people in Western countries. Polyphenol extracts from a variety of plants have been shown to have immunomodulatory and anti-inflammatory effects. In this study, treatment with APP was investigated to ameliorate chemically induced colitis. Oral but not peritoneal administration of APP during colitis induction significantly protected C57BL/6 mice against disease, as evidenced by the lack of weight loss, colonic inflammation, and shortening of the colon. APP administration dampened the mRNA expression of IL-1ß, TNF-α, IL-6, IL-17, IL-22, CXCL9, CXCL10, CXCL11, and IFN-γ in the colons of mice with colitis. APP-mediated protection requires T cells, as protection was abated in Rag-1(-/-) or TCRα(-/-) mice but not in IL-10(-/-), IRF-1(-/-), µMT, or TCRδ(-/-) mice. Administration of APP during colitis to TCRα(-/-) mice actually enhanced proinflammatory cytokine expression, further demonstrating a requirement for TCRαß cells in APP-mediated protection. APP treatment also inhibited CXCR3 expression by TCRαß cells, but not B or NK cells, in the colons of mice with colitis; however, depletion of CD4(+) or CD8(+) T cells alone did not abolish APP-mediated protection. Collectively, these results show that oral administration of APP protects against experimental colitis and diminishes proinflammatory cytokine expression via T cells.


Subject(s)
Colitis/immunology , Cytokines/antagonists & inhibitors , Dextran Sulfate/toxicity , Inflammation Mediators/physiology , Malus/physiology , Polyphenols/physiology , T-Lymphocyte Subsets/drug effects , T-Lymphocyte Subsets/pathology , Animals , Cells, Cultured , Colitis/chemically induced , Colitis/pathology , Cytokines/biosynthesis , Cytokines/genetics , Dextran Sulfate/administration & dosage , Female , Humans , Inflammation Mediators/antagonists & inhibitors , Inflammation Mediators/chemistry , Male , Malus/chemistry , Mice , Mice, Inbred C57BL , Mice, Knockout , Polyphenols/chemistry , T-Lymphocyte Subsets/immunology
16.
Eur J Immunol ; 41(2): 313-23, 2011 Feb.
Article in English | MEDLINE | ID: mdl-21268002

ABSTRACT

Stimulation of Ag-specific inducible Treg can enhance resolution of autoimmune disease. Conventional methods to induce Treg often require induction of autoimmune disease or subjection to infection. Reovirus adhesin, protein σ1 (pσ1), can successfully facilitate tolerance when fused to a tolerogen. We tested whether myelin oligodendrocyte glycoprotein (MOG) fused to pσ1 (MOG-pσ1) can stimulate Ag-specific Treg. We show that C57BL/6 mice treated nasally with MOG-pσ1 fail to induce MOG-specific Abs and delayed-type hypersensitivity (DTH) responses and resist EAE. Such resistance was attributed to stimulation of Foxp3(+) Treg, as well as Th2 cells. MOG-pσ1's protective capacity was abrogated in IL-10(-/-) mice, but restored when adoptively transferred with MOG-pσ1-induced Treg. As a therapeutic, MOG-pσ1 diminished EAE within 24 h of nasal application, unlike recombinant MOG (rMOG), pσ1, or pσ1+rMOG, implicating the importance of Ag specificity by pσ1-based therapeutics. MOG-pσ1-treated mice showed elevated IL-4, IL-10, and IL-28 production by CD4(+) T cells, unlike rMOG treated or control mice that produced elevated IFN-γ or IL-17, respectively. These data show the feasibility of using pσ1 as a tolerogen platform for Ag-specific tolerance induction and highlight its potential use as an immunotherapeutic for autoimmunity.


Subject(s)
Encephalomyelitis, Autoimmune, Experimental/immunology , Encephalomyelitis, Autoimmune, Experimental/therapy , Interleukin-10/immunology , T-Lymphocytes, Regulatory/immunology , Vaccination/methods , Adoptive Transfer , Animals , Capsid Proteins/administration & dosage , Capsid Proteins/genetics , Capsid Proteins/immunology , Cell Count , Cytokines/metabolism , Encephalomyelitis, Autoimmune, Experimental/diagnosis , Female , Glycoproteins/immunology , Hypersensitivity, Delayed/immunology , Immune Tolerance/immunology , Immunoglobulin G/blood , Immunoglobulin G/immunology , Interleukin-10/genetics , Interleukin-2 Receptor alpha Subunit/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , Myelin Proteins , Myelin-Associated Glycoprotein/administration & dosage , Myelin-Associated Glycoprotein/genetics , Myelin-Associated Glycoprotein/immunology , Myelin-Oligodendrocyte Glycoprotein , Peptide Fragments/immunology , Recombinant Fusion Proteins/administration & dosage , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/immunology , Spinal Cord/cytology , Spinal Cord/immunology , Spleen/cytology , Spleen/immunology , T-Lymphocytes, Regulatory/cytology , T-Lymphocytes, Regulatory/metabolism , T-Lymphocytes, Regulatory/transplantation , Th2 Cells/immunology , Th2 Cells/metabolism , Vaccines, Synthetic/administration & dosage , Vaccines, Synthetic/genetics , Vaccines, Synthetic/immunology
17.
Cancer Lett ; 285(1): 80-8, 2009 Nov 18.
Article in English | MEDLINE | ID: mdl-19501956

ABSTRACT

Although T central memory cells have been described as the most effective T-cell subtype against tumor growth, little is known about the requirements needed for their optimal ex vivo generation. Hence, our goal is to establish a protocol that will lead to consistent ex vivo generation of lymphocytes skewed toward a central memory phenotype. Antigen-specific T-cell lines were generated by ex vivo stimulation with Class-I and Class-II melanoma peptide pulsed dendritic cells in the presence of either IL-2 or IL-15 plus IL-21. Tumor specific lymphocytes of both central memory and effector characteristics were consistently generated from healthy donors and melanoma patients. IL15/IL21 cultures result in a cell population with a lower proportion of CD4(+)CD25(high)FoxP3(+) regulatory cells and higher number of CD8(+) and CD56(+) cells, and consequently render a higher yield of cells with a greater cytolytic activity and IFN-gamma production against melanoma cell lines.


Subject(s)
Cell Proliferation , Immunologic Memory , Immunotherapy, Adoptive , Interleukin-15/metabolism , Interleukins/metabolism , Lymphocytes, Tumor-Infiltrating/transplantation , Melanoma/therapy , T-Lymphocyte Subsets/transplantation , Antigens, Neoplasm/immunology , CD4-Positive T-Lymphocytes/immunology , CD56 Antigen/analysis , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/transplantation , Case-Control Studies , Cells, Cultured , Dendritic Cells/immunology , Feasibility Studies , Forkhead Transcription Factors/analysis , Humans , Interferon-gamma/metabolism , Interleukin-2/metabolism , Interleukin-2 Receptor alpha Subunit/analysis , Lymphocytes, Tumor-Infiltrating/immunology , Melanoma/immunology , Melanoma/pathology , Melanoma-Specific Antigens , Neoplasm Proteins/immunology , Phenotype , T-Lymphocyte Subsets/immunology , T-Lymphocytes, Cytotoxic/immunology , T-Lymphocytes, Cytotoxic/transplantation
18.
Cancer Res ; 68(18): 7684-91, 2008 Sep 15.
Article in English | MEDLINE | ID: mdl-18768667

ABSTRACT

Dendritic cells (DC) and cytokines that expand myeloid progenitors are widely used to treat cancer. Here, we show that CD11c(+)DEC205(+) DCs coexpressing alpha-smooth muscle actin and VE-cadherin home to perivascular areas in the ovarian cancer microenvironment and are required for the maintenance of tumor vasculature. Consequently, depletion of DCs in mice bearing established ovarian cancer by targeting different specific markers significantly delays tumor growth and enhances the effect of standard chemotherapies. Tumor growth restriction was associated with vascular apoptosis after DC ablation followed by necrosis, which triggered an antitumor immunogenic boost. Our findings provide a mechanistic rationale for selectively eliminating tumor-associated leukocytes to promote antitumor immunity while impeding tumor vascularization and to develop more effective DC vaccines based on a better understanding of the tumor microenvironment.


Subject(s)
Dendritic Cells/immunology , Ovarian Neoplasms/immunology , Animals , CD11c Antigen/immunology , Cell Growth Processes/immunology , Disease Progression , Female , Humans , Mice , Necrosis , Ovarian Neoplasms/drug therapy , Ovarian Neoplasms/pathology , Ovarian Neoplasms/therapy , T-Lymphocytes, Cytotoxic/immunology
19.
Blood ; 112(4): 1259-68, 2008 Aug 15.
Article in English | MEDLINE | ID: mdl-18550855

ABSTRACT

Robust T-cell responses without autoimmunity are only possible through a fine balance between activating and inhibitory signals. We have identified a novel modulator of T-cell expansion named proliferation-induced lymphocyte-associated receptor (PILAR). Surface PILAR is markedly up-regulated on CD4 and, to a lesser extent, on CD8 T cells on T-cell receptor engagement. In absence of CD28 costimulation, PILAR signaling through CD161 supports CD3 antibody-dependent and antigen-specificT-cell proliferation by increasing the expression of antiapoptotic Bcl-xL and induces secretion of T helper type 1 cytokines. These effects are abrogated by PILAR blockade with specific antibodies, which decrease surface levels of CD28. In contrast, PILAR induces apoptotic death on naive and early activated T cells if CD161 engagement is blocked. PILAR is expressed by approximately 7% to 10% of CD4 T cells in 2 samples of inflammatory synovial fluid, suggesting a potential role in the pathogenesis of joint inflammation. In addition, in the ovarian cancer microenvironment, effector T cells express PILAR, but not CD161, although expression of both can be augmented ex vivo. Our results indicate that PILAR plays a central role in modulating the extent of T-cell expansion. Manipulation of PILAR signaling may be important for treatment of autoimmune diseases and cancer.


Subject(s)
Antigens, Surface/metabolism , Cell Proliferation , Lectins, C-Type/metabolism , Lectins, C-Type/physiology , Lymphocyte Activation/immunology , Membrane Proteins/physiology , Signal Transduction , T-Lymphocytes/immunology , Autoimmune Diseases/immunology , Cells, Cultured , Chromosomes, Human, Pair 12 , Female , Humans , Immunity, Cellular , Inflammation/immunology , Lectins, C-Type/genetics , Membrane Proteins/genetics , NK Cell Lectin-Like Receptor Subfamily B , Ovarian Neoplasms/immunology , Synovial Fluid/immunology , T-Lymphocytes/chemistry , T-Lymphocytes/cytology , Up-Regulation/genetics
20.
Int J Cancer ; 121(6): 1282-95, 2007 Sep 15.
Article in English | MEDLINE | ID: mdl-17520674

ABSTRACT

Better understanding of the mechanisms that mediate spontaneous immune rejections ought to be important in the quest for improvements in immunotherapy of cancer. A set of intraperitoneal tumors of mesenchymal origin that had been chemically induced in ubiquitously expressing EGFP transgenic mice provided a model in which both T and NK cells were absolutely required for tumor rejection. Tumor cells were traceable because of being fluorescent and readily grafted in RAG1(-/-) immunodeficient mice, whereas they were rejected in a majority of syngeneic C57BL/6 and EGFP-transgenic mice. Tumor-cell clones with the highest EGFP expression tended to be rejected, but a direct involvement of EGFP as the antigen recognized for the immune rejections was ruled out. Rejections were absolutely dependent on NK cells as well as on CD4(+) and CD8(+) T lymphocytes according to selective depletion studies. Furthermore, CD8(+) and CD4(+) T lymphocytes as well as NK cells were detected in the inflammatory infiltrate that mediates tumor rejection along with some DC. The effects of IFN gamma, produced at the tumor site by T and NK lymphocytes, were only required at the malignant cell level and were necessary for tumor eradication. NK recognition of tumor cells was mediated by the NKG2D-activating receptor and blocking its function in vivo partially interfered with rejection. Therefore, complete rejection of these mesenchymal tumors requires a concerted set of activities including direct tumor-cell destruction and IFN gamma production that are mediated by both NK and T cells.


Subject(s)
Green Fluorescent Proteins/genetics , Interferon-gamma/metabolism , Killer Cells, Natural/immunology , Peritoneal Neoplasms/immunology , Receptors, Immunologic/metabolism , T-Lymphocytes/immunology , Animals , Cell Line, Tumor , Female , Flow Cytometry , Fluorescent Antibody Technique , Mesoderm/pathology , Mice , Mice, Inbred C57BL , Mice, Transgenic , NK Cell Lectin-Like Receptor Subfamily K , Neoplasms, Experimental/immunology , Neoplasms, Experimental/metabolism , Peritoneal Neoplasms/metabolism , Receptors, Natural Killer Cell
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