Targeting adrenergic receptors to mitigate invariant natural killer T cells-induced acute liver injury.

Invariant Natural Killer T (iNKT) cell activation by α-galactosylceramide (αGC) potentiates cytotoxic immune responses against tumors. However, αGC-induced liver injury is a limiting factor for iNKT-based immunotherapy. Although adrenergic receptor stimulation is an important immunosuppressive signal that curbs tissue damage induced by inflammation, its effect on the antitumor activity of invariant Natural Killer T (iNKT) cells remains unclear. We use mouse models and pharmacological tools to show that the stimulation of the sympathetic nervous system (SNS) inhibits αGC-induced liver injury without impairing iNKT cells' antitumoral functions. Mechanistically, SNS stimulation prevents the collateral effect of TNF-α production by iNKT cells and neutrophil accumulation in hepatic parenchyma. Our results suggest that the modulation of the adrenergic signaling can be a complementary approach to αGC-based immunotherapy to mitigate iNKT-induced liver injury without compromising its antitumoral activity.

Tissue-specific metabolic profile drives iNKT cell function during obesity and liver injury.

Invariant natural killer T (iNKT) cells are a distinct population of lymphocytes characterized by their reactivity to glycolipids presented by CD1d. iNKT cells are found throughout the body, and little is known about their tissue-specific metabolic regulation. Here, we show that splenic and hepatic iNKT cells are metabolically comparable and rely on glycolytic metabolism to support their activation. Deletion of the pyruvate kinase M2 (Pkm2) gene in splenic and hepatic iNKT cells impairs their response to specific stimulation and their ability to mitigate acute liver injury. In contrast, adipose tissue (AT) iNKT cells exhibit a distinctive immunometabolic profile, with AMP-activated protein kinase (AMPK) being necessary for their function. AMPK deficiency impairs AT-iNKT physiology, blocking their capacity to maintain AT homeostasis and their ability to regulate AT inflammation during obesity. Our work deepens our understanding on the tissue-specific immunometabolic regulation of iNKT cells, which directly impacts the course of liver injury and obesity-induced inflammation.

Sleep Disturbance during Infection Compromises Tfh Differentiation and Impacts Host Immunity.

Although the influence of sleep quality on the immune system is well documented, the mechanisms behind its impact on natural host immunity remain unclear. Meanwhile, it has been suggested that neuroimmune interactions play an important role in this phenomenon. To evaluate the impact of stress-induced sleep disturbance on host immunity, we used a murine model of rapid eye movement sleep deprivation (RSD) integrated with a model of malaria blood-stage infection. We demonstrate that sleep disturbance compromises the differentiation of T follicular helper cells, increasing host susceptibility to the parasite. Chemical inhibition of glucocorticoid (Glcs) synthesis showed that abnormal Glcs production compromised the transcription of Tfh-associated genes resulting in impaired germinal center formation and humoral immune response. Our data demonstrate that RSD-induced abnormal activation of the hypothalamic-pituitary-adrenal axis drives host susceptibility to infection. Understanding the impact of sleep quality in natural resistance to infection may provide insights for disease management.

The axis IL-10/claudin-10 is implicated in the modulation of aggressiveness of melanoma cells by B-1 lymphocytes.

B-1 lymphocytes are known to increase the metastatic potential of B16F10 melanoma cells via the extracellular signal-regulated kinase (ERK) pathway. Since IL-10 is associated with B-1 cells performance, we hypothesized that IL-10 could be implicated in the progression of melanoma. In the present work, we found that the C57BL/6 mice, inoculated with B16F10 cells that were co-cultivated with B-1 lymphocytes from IL-10 knockout mice, developed fewer metastatic nodules than the ones which were injected with the melanoma cells that were cultivated in the presence of wild-type B-1 cells. The impairment of metastatic potential of the B16F10 cells was correlated with low activation of the ERK signaling pathway, supporting the idea that the production of IL-10 by B-1 cells influences the behavior of the tumor. A microarray analysis of the B-1 lymphocytes revealed that IL-10 deficiency is associated with down-regulation of the genes that code for claudin-10, a protein that is involved in cell-to-cell contact and that has been linked to lung adenocarcinoma. In order to determine the impact of claudin-10 in the cross-talk between B-1 lymphocytes and the B16F10 tumor cells, we took advantage of small interfering RNA. The silencing of claudin-10 gene in B-1 lymphocytes inhibited activation of the ERK pathway and abrogated the B-1-induced aggressive behavior of the B16F10 cells. Thus, our findings suggest that the axis IL-10/claudin-10 is a promising target for the development of therapeutic agents against aggressive melanoma.

Oxidative stress and modification of renal vascular permeability are associated with acute kidney injury during P. berghei ANKA infection.

Malaria associated-acute kidney injury (AKI) is associated with 45% of mortality in adult patients hospitalized with severe form of the disease. However, the causes that lead to a framework of malaria-associated AKI are still poorly characterized. Some clinical studies speculate that oxidative stress products, a characteristic of Plasmodium infection, as well as proinflammatory response induced by the parasite are involved in its pathophysiology. Therefore, we aimed to investigate the development of malaria-associated AKI during infection by P. berghei ANKA, with special attention to the role played by the inflammatory response and the involvement of oxidative stress. For that, we took advantage of an experimental model of severe malaria that showed significant changes in the renal pathophysiology to investigate the role of malaria infection in the renal microvascular permeability and tissue injury. Therefore, BALB/c mice were infected with P. berghei ANKA. To assess renal function, creatinine, blood urea nitrogen, and ratio of proteinuria and creatininuria were evaluated. The products of oxidative stress, as well as cytokine profile were quantified in plasma and renal tissue. The change of renal microvascular permeability, tissue hypoxia and cellular apoptosis were also evaluated. Parasite infection resulted in renal dysfunction. Furthermore, we observed increased expression of adhesion molecule, proinflammatory cytokines and products of oxidative stress, associated with a decrease mRNA expression of HO-1 in kidney tissue of infected mice. The measurement of lipoprotein oxidizability also showed a significant increase in plasma of infected animals. Together, our findings support the idea that products of oxidative stress, as well as the immune response against the parasite are crucial to changes in kidney architecture and microvascular endothelial permeability of BALB/c mice infected with P. berghei ANKA.

The polysaccharide fraction of Propionibacterium acnes modulates the development of experimental focal segmental glomerulosclerosis.

The pathogenesis of focal segmental glomerulosclerosis (FSGS) appears to be associated with type-2 cytokines and podocyte dysfunction. In this study, we tested the hypothesis that immunization with the polysaccharide fraction of Propionibacterium acnes (PS), a pro-Th1 agonist, may subvert the type-2 profile and protect podocytes from adriamycin-induced glomerulosclerosis. Adriamycin injection resulted in albuminuria and increased serum creatinine in association with loss of glomerular podocin and podoplanin expression, which is consistent with podocyte dysfunction. Renal tissue analysis revealed the expression of transcripts for GATA3 and fibrogenic-related proteins, such as TGF-ß, tissue inhibitor of metalloproteinase-1 (TIMP-1) and metalloproteinase 9 (MMP9). In association with the expression of fibrogenic transcripts, we observed peri-glomerular expression of α-smooth muscle actin (α-SMA), indicating epithelial-to-mesenchymal transition, and increased expression of proliferating cell nuclear antigen (PCNA) in tubular cells, suggesting intense proliferative activity. Previous immunization with PS inhibited albuminuria and serum creatinine in association with the preservation of podocyte proteins and inhibition of fibrogenic transcripts and the expression of α-SMA and PCNA proteins. Tissue analysis also revealed that PS treatment induced expression of mRNA for GD3 synthase, which is a glycosiltransferase related to the synthesis of GD3, a ganglioside associated with podocyte physiology. In addition, PS treatment inhibited the influx of inflammatory CD8(pos) and CD11b(pos) cells to kidney tissue. Finally, PS treatment on day 4 post-ADM, a period when proteinuria was already established, was able to improve renal function. Thus, we demonstrate that the PS fraction of P. acnes can inhibit FSGS pathogenesis, suggesting that immunomodulation can represent an alternative approach for disease management.

Regulatory T cells accumulate in the lung allergic inflammation and efficiently suppress T-cell proliferation but not Th2 cytokine production.

Foxp3(+)CD25(+)CD4(+) regulatory T cells are vital for peripheral tolerance and control of tissue inflammation. In this study, we characterized the phenotype and monitored the migration and activity of regulatory T cells present in the airways of allergic or tolerant mice after allergen challenge. To induce lung allergic inflammation, mice were sensitized twice with ovalbumin/aluminum hydroxide gel and challenged twice with intranasal ovalbumin. Tolerance was induced by oral administration of ovalbumin for 5 consecutive days prior to OVA sensitization and challenge. We detected regulatory T cells (Foxp3(+)CD25(+)CD4(+) T cells) in the airways of allergic and tolerant mice; however, the number of regulatory T cells was more than 40-fold higher in allergic mice than in tolerant mice. Lung regulatory T cells expressed an effector/memory phenotype (CCR4(high)CD62L(low)CD44(high)CD54(high)CD69(+)) that distinguished them from naive regulatory T cells (CCR4(int)CD62L(high)CD44(int)CD54(int)CD69(-)). These regulatory T cells efficiently suppressed pulmonary T-cell proliferation but not Th2 cytokine production.

Vitronectin dictates intraglomerular fibrinolysis in immune-mediated glomerulonephritis.

During human glomerulonephritis, the severity of injuries correlates with glomerular fibrin deposits, which are tightly regulated by the intraglomerular fibrinolytic system. Here, we evaluated the role of vitronectin (VTN; also known as complement S protein), the principal cofactor of the plasminogen activator inhibitor-1 (PAI-1), in a mouse model of acute glomerulonephritis. We found that in mice subjected to nephrotoxic serum, the absence of VTN resulted in a lower glomerular PAI-1 activity and a higher glomerular fibrinolytic activity. Challenged VTN(-/-) mice displayed significantly less fibrin deposits, proteinuria, and renal failure than their wild-type counterparts. Notably, this protective effect afforded by VTN deficiency was still observed after a C3 depletion. Finally, the injection of VTN(+/+) serum in VTN(-/-) mice induced the glomerular deposition of VTN, increased PAI-1 deposition, decreased glomerular fibrinolytic activity, and aggravated glomerular injury. As in mice, abundant glomerular VTN deposits were also observed in patients with severe glomerulonephritis. Here, we show that plasma-exchange therapy, admittedly beneficial in this clinical context, induces a significant depletion in circulating VTN, which might modulate PAI-1 activity locally and accelerate the clearance of fibrin deposits in the glomeruli. Collectively, these results demonstrate that VTN exerts a deleterious role independently from complement, by directing PAI-dependent fibrinolysis in the glomerular compartment.

Critical role of ROR-γt in a new thymic pathway leading to IL-17-producing invariant NKT cell differentiation.

Invariant natural killer T (iNKT) cells constitute a subpopulation of T cells that recognize glycolipids presented by CD1d molecules. They are characterized by their prompt production of interleukin-4 (IL-4) and interferon-gamma (IFN-gamma), which enables them to modulate diverse immune responses. Recently, we enlarged this concept by identifying a distinct IL-17-producing iNKT cell subset, named iNKT17 cells. The mechanisms leading to the acquisition of this new iNKT cell activity are unknown. Herein we show that IL-17-producing iNKT cells are already present in the thymus, predominantly among a subset regarded so far as an immature stage of thymic iNKT cell development, the CD1d tetramer(pos)CD44(pos)NK1.1(neg)CD4(neg) cells. Using EGFP reporter mice, we demonstrate that the transcription factor ROR-gammat is critical for the thymic differentiation of this subset because only ROR-gammat(pos) iNKT cells are capable of massively secreting IL-17. Moreover, IL-17-producing CD1d tetramer(pos)CD44(pos)NK1.1(neg)CD4(neg) thymic iNKT cells have reached a mature differentiation stage because they fail to generate other cell subsets in fetal thymic organ culture. Conversely, thymic ROR-gammat(neg) iNKT cell precursors give rise to progeny, but acquire neither ROR-gammat expression nor the ability to secrete IL-17. In conclusion, our findings demonstrate an alternative thymic pathway leading to the development of iNKT17 cells that requires ROR-gammat expression.

Identification of an IL-17-producing NK1.1(neg) iNKT cell population involved in airway neutrophilia.

Invariant natural killer T (iNKT) cells are an important source of both T helper type 1 (Th1) and Th2 cytokines, through which they can exert beneficial, as well as deleterious, effects in a variety of inflammatory diseases. This functional heterogeneity raises the question of how far phenotypically distinct subpopulations are responsible for such contrasting activities. In this study, we identify a particular set of iNKT cells that lack the NK1.1 marker (NK1.1(neg)) and secrete high amounts of interleukin (IL)-17 and low levels of interferon (IFN)-gamma and IL-4. NK1.1(neg) iNKT cells produce IL-17 upon synthetic (alpha-galactosylceramide [alpha-GalCer] or PBS-57), as well as natural (lipopolysaccharides or glycolipids derived from Sphingomonas wittichii and Borrelia burgdorferi), ligand stimulation. NK1.1(neg) iNKT cells are more frequent in the lung, which is consistent with a role in the natural immunity to inhaled antigens. Indeed, airway neutrophilia induced by alpha-GalCer or lipopolysaccharide instillation was significantly reduced in iNKT-cell-deficient Jalpha18(-/-) mice, which produced significantly less IL-17 in their bronchoalveolar lavage fluid than wild-type controls. Furthermore, airway neutrophilia was abolished by a single treatment with neutralizing monoclonal antibody against IL-17 before alpha-GalCer administration. Collectively, our findings reveal that NK1.1(neg) iNKT lymphocytes represent a new population of IL-17-producing cells that can contribute to neutrophil recruitment through preferential IL-17 secretion.

Nitric oxide paradox in asthma.

Asthma results from allergen-driven intrapulmonary Th2 response, and is characterized by intermittent airway obstruction, airway hyperreactivity (AHR), and airway inflammation. Accumulating evidence indicates that inflammatory diseases of the respiratory tract are commonly associated with elevated production of nitric oxide (NO). It has been shown that exhaled NO may be derived from constitutive NO synthase (NOS) such as endothelial (NOS 3) and neural (NOS 1) in normal airways, while increased levels of NO in asthma appear to be derived from inducible NOS2 expressed in the inflamed airways. Nevertheless, the functional role of NO and NOS isoforms in the regulation of AHR and airway inflammation in human or experimental models of asthma is still highly controversial. In the present commentary we will discuss the role of lipopolysaccharides contamination of allergens as key element in the controversy related to the regulation of NOS2 activity in experimental asthma.

Nitric oxide paradox in asthma

Asthma results from allergen-driven intrapulmonary Th2 response, and is characterized by intermittent airway obstruction, airway hyperreactivity (AHR), and airway inflammation. Accumulating evidence indicates that inflammatory diseases of the respiratory tract are commonly associated with elevated production of nitric oxide (NO). It has been shown that exhaled NO may be derived from constitutive NO synthase (NOS) such as endothelial (NOS 3) and neural (NOS 1) in normal airways, while increased levels of NO in asthma appear to be derived from inducible NOS2 expressed in the inflamed airways. Nevertheless, the functional role of NO and NOS isoforms in the regulation of AHR and airway inflammation in human or experimental models of asthma is still highly controversial. In the present commentary we will discuss the role of lipopolysaccharides contamination of allergens as key element in the controversy related to the regulation of NOS2 activity in experimental asthma.

Decreased nasal tolerance to allergic asthma in mice fed an amino acid-based protein-free diet.

Intranasal (i.n.) administration of soluble proteins induces a state of specific unresponsiveness to subsequent immunization, known as nasal tolerance. It is thought that newborns are less susceptible to nasal tolerance induction. Recently, we have shown that feeding adult animals with a protein-free diet (Aa) resulted in their arrest at an immature immunological profile. Here, we examined the effects of the Aa diet on the development of nasal tolerance to ovalbumin (OVA) in a murine model of allergic asthma. Nasal OVA administration suppressed almost totally the OVA-induced asthma-like responses (airway eosinophilia, type 2 cytokine production, and OVA-specific IgE antibodies) in chow- or casein-fed BALB/c mice. In contrast, in Aa-fed animals the suppression of asthma-like responses by nasal OVA was partial, being effective in suppressing airway eosinophilia, but not airway type 2 cytokine or OVA-specific IgE response. We conclude that animals fed the Aa diet are more resistant to the induction of nasal tolerance. Our animal model may mimic the features of the immune system of human infants.