Genetic Deficiency of Indoleamine 2,3-dioxygenase Aggravates Vascular but Not Liver Disease in a Nonalcoholic Steatohepatitis and Atherosclerosis Comorbidity Model.

Nonalcoholic steatohepatitis (NASH) is a chronic liver disease that increases cardiovascular disease risk. Indoleamine 2,3-dioxygenase-1 (IDO1)-mediated tryptophan (Trp) metabolism has been proposed to play an immunomodulatory role in several diseases. The potential of IDO1 to be a link between NASH and cardiovascular disease has never been investigated. Using Apoe-/-and Apoe-/-Ido1-/- mice that were fed a high-fat, high-cholesterol diet (HFCD) to simultaneously induce NASH and atherosclerosis, we found that Ido1 deficiency significantly accelerated atherosclerosis after 7 weeks. Surprisingly, Apoe-/-Ido1-/- mice did not present a more aggressive NASH phenotype, including hepatic lipid deposition, release of liver enzymes, and histopathological parameters. As expected, a lower L-kynurenine/Trp (Kyn/Trp) ratio was found in the plasma and arteries of Apoe-/-Ido1-/- mice compared to controls. However, no difference in the hepatic Kyn/Trp ratio was found between the groups. Hepatic transcript analyses revealed that HFCD induced a temporal increase in tryptophan 2,3-dioxygenase (Tdo2) mRNA, indicating an alternative manner to maintain Trp degradation during NASH development in both Apoe-/- and Apoe-/-Ido1-/mice-. Using HepG2 hepatoma cell and THP1 macrophage cultures, we found that iron, TDO2, and Trp degradation may act as important mediators of cross-communication between hepatocytes and macrophages regulating liver inflammation. In conclusion, we show that Ido1 deficiency aggravates atherosclerosis, but not liver disease, in a newly established NASH and atherosclerosis comorbidity model. Our data indicate that the overexpression of TDO2 is an important mechanism that helps in balancing the kynurenine pathway and inflammation in the liver, but not in the artery wall, which likely determined disease outcome in these two target tissues.

Endangered Lymphocytes: The Effects of Alloxan and Streptozotocin on Immune Cells in Type 1 Induced Diabetes.

Alloxan (ALX) and streptozotocin (STZ) are extensively used to induce type 1 diabetes (T1D) in animal models. This study is aimed at evaluating the differences in immune parameters caused by ALX and STZ. T1D was induced either with ALX or with STZ, and the animals were followed for up to 180 days. Both ALX and STZ induced a decrease in the total number of circulating leukocytes and lymphocytes, with an increase in granulocytes when compared to control mice (CT). STZ-treated mice also exhibited an increase in neutrophils and a reduction in the lymphocyte percentage in the bone marrow. In addition, while the STZ-treated group showed a decrease in total CD3+, CD4-CD8+, and CD4+CD8+ T lymphocytes in the thymus and CD19+ B lymphocytes in the pancreas and spleen, the ALX group showed an increase in CD4-CD8+ and CD19+ only in the thymus. Basal levels of splenic interleukin- (IL-) 1ß and pancreatic IL-6 in the STZ group were decreased. Both diabetic groups showed atrophy of the thymic medulla and degeneration of pancreatic islets of Langerhans composed of inflammatory infiltration and hyperemia with vasodilation. ALX-treated mice showed a decrease in reticuloendothelial cells, enhanced lymphocyte/thymocyte cell death, and increased number of Hassall's corpuscles. Reduced in vitro activation of splenic lymphocytes was found in the STZ-treated group. Furthermore, mice immunized with ovalbumin (OVA) showed a more intense antigen-specific paw edema response in the STZ-treated group, while production of anti-OVA IgG1 antibodies was similar in both groups. Thereby, important changes in immune cell parameters in vivo and in vitro were found at an early stage of T1D in the STZ-treated group, whereas alterations in the ALX-treated group were mostly found in the chronic phase of T1D, including increased mortality rates. These findings suggest that the effects of ALX and STZ influenced, at different times, lymphoid organs and their cell populations.

Immune-pineal axis protects rat lungs exposed to polluted air.

Environmental pollution in the form of particulate matter <2.5 µm (PM2.5 ) is a major risk factor for diseases such as lung cancer, chronic respiratory infections, and major cardiovascular diseases. Our goal was to show that PM2.5 eliciting a proinflammatory response activates the immune-pineal axis, reducing the pineal synthesis and increasing the extrapineal synthesis of melatonin. Herein, we report that the exposure of rats to polluted air for 6 hours reduced nocturnal plasma melatonin levels and increased lung melatonin levels. Melatonin synthesis in the lung reduced lipid peroxidation and increased PM2.5 engulfment and cell viability by activating high-affinity melatonin receptors. Diesel exhaust particles (DEPs) promoted the synthesis of melatonin in a cultured cell line (RAW 264.7 cells) and rat alveolar macrophages via the expression of the gene encoding for AANAT through a mechanism dependent on activation of the NFκB pathway. Expression of the genes encoding AANAT, MT1, and MT2 was negatively correlated with cellular necroptosis, as disclosed by analysis of Gene Expression Omnibus (GEO) microarray data from the human alveolar macrophages of nonsmoking subjects. The enrichment score for antioxidant genes obtained from lung gene expression data (GTEx) was significantly correlated with the levels of AANAT and MT1 but not the MT2 melatonin receptor. Collectively, these data provide a systemic and mechanistic rationale for coordination of the pineal and extrapineal synthesis of melatonin by a standard damage-associated stimulus, which activates the immune-pineal axis and provides a new framework for understanding the effects of air pollution on lung diseases.

Diabetes Mellitus and Liver Surgery: The Effect of Diabetes on Oxidative Stress and Inflammation.

Diabetes mellitus (DM) is a metabolic disorder characterized by hyperglycaemia and high morbidity worldwide. The detrimental effects of hyperglycaemia include an increase in the oxidative stress (OS) response and an enhanced inflammatory response. DM compromises the ability of the liver to regenerate and is particularly associated with poor prognosis after ischaemia-reperfusion (I/R) injury. Considering the growing need for knowledge of the impact of DM on the liver following a surgical procedure, this review aims to present recent publications addressing the effects of DM (hyperglycaemia) on OS and the inflammatory process, which play an essential role in I/R injury and impaired hepatic regeneration after liver surgery.

Insulin Influences LPS-Induced TNF-α and IL-6 Release Through Distinct Pathways in Mouse Macrophages from Different Compartments.

BACKGROUND/AIMS: Diabetic subjects are more susceptible to infections, which is partially due to insulin deficiency and hyperglycemia. We hypothesized that insulin influences cytokine release by macrophages from diabetic C57BL/6 mice stimulated with lipopolysaccharides (LPS). METHODS: Bone marrow-derived macrophages (BMDM) and tissue-specific macrophages from diabetic (alloxan 60 mg/kg, i.v.) male C57BL/6 mice were stimulated by LPS (100 ng/mL) and/or treated by insulin (1 mU/mL). RESULTS: Using BMDM from diabetic mice, we showed that LPS induced an increase in TNF-α and IL-6 release and p38, SAPK/JNK, ERK 1/2, and Akt (308-Thr and 473-Ser) phosphorylation but not in PKCα/ß II and delta. Insulin increased TNF-α and IL-6 secretion in LPS-stimulated macrophages as well as p-p38, p-SAPK/JNK, p-ERK 1/2, p-PI3K (p55) and p-Akt (473-Ser) expression. Furthermore, PI3-kinase inhibition by wortmannin decreased TNF-α release, and inhibition by LY294002 decreased both TNF-α and IL-6 levels after LPS-insulin treatment. PD98059, which inhibits the ERK upstream activators MAPK kinase (MKK) 1 and MKK2, reduced the effect promoted by insulin in BMDM stimulated by LPS In tissue-specific macrophages, insulin reduced LPS-induced TNF-α, IL-6 and IL-1ß secretion in alveolar and peritoneal macrophages. CONCLUSION: These data suggest that insulin through the modulation of PI3-kinase and ERK 1/2 pathways drive different responses in macrophages, thereby enhancing our understanding of the plasticity of these cells.

The role and effects of glucocorticoid-induced leucine zipper in the context of inflammation resolution.

Glucocorticoid (GC)-induced leucine zipper (GILZ) has been shown to mediate or mimic several actions of GC. This study assessed the role of GILZ in self-resolving and GC-induced resolution of neutrophilic inflammation induced by LPS in mice. GILZ expression was increased during the resolution phase of LPS-induced pleurisy, especially in macrophages with resolving phenotypes. Pretreating LPS-injected mice with trans-activator of transcription peptide (TAT)-GILZ, a cell-permeable GILZ fusion protein, shortened resolution intervals and improved resolution indices. Therapeutic administration of TAT-GILZ induced inflammation resolution, decreased cytokine levels, and promoted caspase-dependent neutrophil apoptosis. TAT-GILZ also modulated the activation of the survival-controlling proteins ERK1/2, NF-κB and Mcl-1. GILZ deficiency was associated with an early increase of annexin A1 (AnxA1) and did not modify the course of neutrophil influx induced by LPS. Dexamethasone treatment resolved inflammation and induced GILZ expression that was dependent on AnxA1. Dexamethasone-induced resolution was not altered in GILZ(-/-) mice due to compensatory expression and action of AnxA1. Our results show that therapeutic administration of GILZ efficiently induces a proapoptotic program that promotes resolution of neutrophilic inflammation induced by LPS. Alternatively, a lack of endogenous GILZ during the resolution of inflammation is compensated by AnxA1 overexpression.

Insulin Modulates Liver Function in a Type I Diabetes Rat Model.

BACKGROUND/AIMS: Several studies have been performed to unravel the association between diabetes and increased susceptibility to infection. This study aimed to investigate the effect of insulin on the local environment after cecal ligation and puncture (CLP) in rats. METHODS: Diabetic (alloxan, 42 mg/kg i.v., 10 days) and non-diabetic (control) male Wistar rats were subjected to a two-puncture CLP procedure and 6 h later, the following analyses were performed: (a) total and differential cell counts in peritoneal lavage (PeL) and bronchoalveolar lavage (BAL) fluids; (b) quantification of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL- 6, IL-10 and cytokine-induced neutrophil chemoattractant (CINC)-1 and CINC-2 in the PeL and BAL fluids by enzyme-linked immunosorbent assay (ELISA); (c) total leukocyte count using a veterinary hematology analyzer and differential leukocyte counts on stained slides; (d) biochemical parameters (urea, creatinine, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP) by colorimetric analyses); and (e) lung, kidney, and liver morphological analyses (hematoxylin and eosin staining). RESULTS: Relative to controls, non-diabetic and diabetic CLP rats exhibited an increased in the concentration of IL-1ß, IL-6, IL-10, CINC-1, and CINC-2 and total and neutrophil in the PeL fluid. Treatment of these animals with neutral protamine Hagedorn insulin (NPH, 1IU and 4IU, respectively, s.c.), 2 hours before CLP procedure, induced an increase on these cells in the PeL fluid but it did not change cytokine levels. The levels of ALT, AST, ALP, and urea were higher in diabetic CLP rats than in non-diabetic CLP rats. ALP levels were higher in diabetic sham rats than in non-diabetic sham rats. Treatment of diabetic rats with insulin completely restored ALT, AST, and ALP levels. CONCLUSION: These results together suggest that insulin attenuates liver dysfunction during early two-puncture CLP-induced peritoneal inflammation in diabetic rats.

Insulin influences autophagy response distinctively in macrophages of different compartments.

BACKGROUND/AIMS: Diabetes mellitus (DM) is characterized by hyperglycemia, associated to a lack or inefficiency of the insulin to regulate glucose metabolism. DM is also marked by alterations in a diversity of cellular processes that need to be further unraveled. In this study, we examined the autophagy pathway in diabetic rat macrophages before and after treatment with insulin. METHODS: Bone marrow-derived macrophages (BMM), bronchoalveolar lavage (BAL) and splenic tissue of diabetic male Wistar rats (alloxan, 42 mg/kg, i.v., 10 days) and control rats (physiological saline, i.v.). Some diabetic rats were given neutral protamine Hagedorn insulin (4 IU, s.c.) 8 h before experiments. For characterization of the model and evaluation of the effect of insulin on the autophagic process, the following analyzes were performed: (a) concentrations of cytokines: interleukin (IL)-1ß, tumor necrosis factor (TNF)-α, IL-6, IL-4, IL-10, cytokine-induced neutrophil chemoattractant (CINC)-1 and CINC-2 in the BAL supernatant was measured by ELISA; (b) characterization of alveolar macrophage (AM) of the BAL as surface antigens (MHCII, pan-macrophage KiM2R, CD11b) and autophagic markers (protein microtubule-associated light chain (LC)3, autophagy protein (Atg)12 by flow cytometry and confocal microscopy (c) study of macrophages differentiated from the bone marrow by flow cytometry and confocal microscopy (d) histology of the spleen by immunohistochemistry associated with confocal microscopy. RESULTS: Interestingly, insulin exerted antagonistic effects on macrophages from different tissues. Macrophages from bronchoalveolar lavage (BAL) enhanced their LC3 autophagosome bound content after treatment with insulin whereas splenic macrophages from red pulp in diabetic rats failed to enhance their Atg 12 levels compared to control animals. Insulin treatment in diabetic rats did not change LC3 content in bone marrow derived macrophages (BMM). M1 and M2 macrophages behaved accordingly to the host they were derived from. Diabetic M1 BMM had their LC3 vesicle-bound content diminished and M2 BMM enhanced their LC3 levels and insulin treatment failed to rescue autophagy to control levels. Insulin normalizes CINC-2 level but does not modulate autophagy markers. CONCLUSION: Taking these results together, diabetic macrophages derived from different compartments show different levels of autophagy markers compared to healthy animals, therefore, they suffer distinctively in the absence of insulin.

Sepsis-induced lung inflammation is modulated by insulin.

BACKGROUND: We have previously shown that diabetic rats are more susceptible to sepsis, but that the Acute lung injury (ALI) secondary to sepsis is less intense than in non-diabetics. In the present study, we further investigated the ALI-secondary to sepsis in diabetic rats and the effect of insulin treatment. METHODS: Diabetes was induced in male Wistar rats by alloxan and sepsis by cecal ligation and puncture surgery (CLP). Some diabetic rats were given neutral protamine Hagedorn (NPH) insulin (4 IU, s.c.) 2 h before CLP. Six h later, the lungs were examined for edema, cell infiltration and prostaglandin-E2 (PGE2) levels in the bronchoalveolar lavage (BAL). RESULTS: The results confirmed that leukocyte infiltration and edema were milder in diabetic rats with sepsis. After insulin treatment, the lung inflammation in diabetics increased to levels comparable to the non-diabetics. The BAL concentration of PGE2 was also lower in diabetics with sepsis, and increased after insulin treatment. Sepsis was followed by early fibroblast activation in the lung parenchyma, evaluated by increased transforming growth factor (TGF)-ß and smooth muscle actin (α-SMA) expression, as well as an elevated number of cells with myofibroblasts morphology. These events were significantly lower in diabetic rats and increased after insulin treatment. CONCLUSION: The results show that insulin modulates the early phase of inflammation and myofibroblast differentiation in diabetic rats.

Effects of low-dose rapamycin on lymphoid organs of mice prone and resistant to accelerated senescence.

Aging is a complex, natural, and irreversible phenomenon that subjects the body to numerous changes in the physiological process, characterized by a gradual decline in the organism's homeostatic mechanisms, closely related to immunosenescence. Here, we evaluated the regulation of immunosenescence in lymphoid organs of senescence-accelerated prone 8 (SAM-P8) and senescence-accelerated resistant 1 (SAM-R1) mice treated with a low dose of rapamycin (RAPA). Mice were treated with a dose of 7.1 µg/kg RAPA for 2 months and had body mass and hematological parameters analyzed prior and during treatment. Cellular and humoral parameters of serum, bone marrow, thymus, and spleen samples were evaluated by ELISA, histology, and flow cytometry. Changes in body mass, hematological parameters, cell number, and in the secretion of IL-1ß, IL-6, TNF-α, IL-7, and IL-15 cytokines were different between the 2 models used. In histological analyses, we observed that SAM-P8 mice showed faster thymic involution than SAM-R1 mice. Regarding the T lymphocyte subpopulations in the spleen, CD4+ and CD8+ T cell numbers were higher and lower, respectively, in SAM-P8 mice treated with RAPA, with the opposite observed in SAM-R1. Additionally, we found that the low dose of RAPA used did not trigger changes that could compromise the immune response of these mice and the administered dose may have contributed to changes in important lymphocyte populations in the adaptive immune response and the secretion of cytokines that directly collaborate with the maturation and proliferation of these cells.

The influence of high glucose conditions on macrophages and its effect on the autophagy pathway.

Introduction: Macrophages are central cells in mediating the inflammatory response. Objective and Methods: We evaluated the effect of high glucose conditions on the inflammatory profile and the autophagy pathway in Bone-Marrow Derived Macrophages (BMDM) from diabetic (D-BMDM) (alloxan: 60mg/kg, i.v.) and non-diabetic (ND-BMDM) C57BL/6 mice. BMDM were cultured in medium with normal glucose (5.5 mM), or high glucose (25 mM) concentration and were primed with Nigericin (20µM) stimulated with LPS (100 ng/mL) at times of 30 minutes; 2; 4; 6 and 24 hours, with the measurement of IL-6, IL-1ß and TNF-α cytokines. Results: We have further identified changes in the secretion of pro-inflammatory cytokines IL-6, IL-1ß and TNF-α, where BMDM showed increased secretion of these cytokines after LPS + Nigericin stimulation. In addition, changes were observed in the autophagy pathway, where the increase in the autophagic protein LC3b and Beclin-1 occurred by macrophages of non-diabetic animals in hyperglycemic medium, without LPS stimulation. D-BMDM showed a reduction on the expression of LC3b and Beclin-1, suggesting an impaired autophagic process in these cells. Conclusion: The results suggest that hyperglycemia alters the inflammatory pathways in macrophages stimulated by LPS, playing an important role in the inflammatory response of diabetic individuals.

The role of captopril in leukotriene deficient type 1 diabetic mice.

T1D can be associated with metabolic disorders and several impaired pathways, including insulin signaling, and development of insulin resistance through the renin-angiotensin system (RAS). The main precursor of RAS is angiotensinogen (Agt) and this system is often linked to autophagy dysregulation. Dysregulated autophagy has been described in T1D and linked to impairments in both glucose metabolism, and leukotrienes (LTs) production. Here, we have investigated the role of RAS and LTs in both muscle and liver from T1D mice, and its effects on insulin and autophagy pathways. We have chemically induced T1D in 129sve and 129sve 5LO-/- mice (lacking LTs) with streptozotocin (STZ). To further inhibit ACE activity, mice were treated with captopril (Cap). In muscle of T1D mice, treatment with Cap increased the expression of RAS (angiotensinogen and angiotensin II receptor), insulin signaling, and autophagy markers, regardless of the genotype. In the liver of T1D mice, the treatment with Cap increased the expression of RAS and insulin signaling markers, mostly when LTs were absent. 5LO-/- T1D mice showed increased insulin sensitivity, and decreased NEFA, after the Cap treatment. Cap treatment impacted both insulin signaling and autophagy pathways at the mRNA levels in muscle and liver, indicating the potential role of ACE inhibition on insulin sensitivity and autophagy in T1D.

Briefs on insulin and innate immune response.

Insulin is a pivotal regulator of glucose metabolism and exerts an important anabolic function throughout the body. Insulin commands the glucose uptake by the cells and might control the processes in which there is need for energy such as mitogenesis and gene transcription. In certain conditions, diabetes mellitus for example, when insulin is diminished, the homeostasis of many tissues and organs are broken what can lead to a higher mortality due to an enhanced susceptibility to infections. This vulnerability to infections can partially be explained by a change in response to inflammation. In fact, diabetic animals and patients show a deficient inflammatory response. Many animal models have shown that neutrophils chemotaxis and recruitment are dampened and macrophages from diabetic patients have low phagocytic and microbicidal activities. In most cases, once insulin therapy is introduced, clinical symptoms and signs can be reverted. In addition, there are a number of studies trying to demystify pathways under insulin command. Researchers are also trying to understand how insulin is able to keep inflammatory response under control, restores innate immune cells ability to fight against pathogens and harmlessly activates adaptive immunity response. This review provides an overview on how inflammatory response is driven in the absence of insulin in diabetes and discusses recent findings on the influence of insulin on innate imune response. At the end, some signaling pathways are also highlighted and important enzymes and proteins that control DNA transcription are presented.

Modulation of lung allergic response by renal ischemia and reperfusion injury.

The Th1/Th2 balance represents an important factor in the pathogenesis of renal ischemia-reperfusion injury (IRI). In addition, IRI causes a systemic inflammation that can affect other tissues, such as the lungs. To investigate the ability of renal IRI to modulate pulmonary function in a specific model of allergic inflammation, C57Bl/6 mice were immunized with ovalbumin/albumen on days 0 and 7 and challenged with an ovalbumin (OA) aerosol on days 14 and 21. After 24 h of the second antigen challenge, the animals were subjected to 45 minutes of ischemia. After 24 h of reperfusion, the bronchoalveolar lavage (BAL) fluid, blood and lung tissue were collected for analysis. Serum creatinine levels increased in both allergic and non-immunized animals subjected to IRI. However, BAL analysis showed a reduction in the total cells (46%) and neutrophils (58%) compared with control allergic animals not submitted to IRI. In addition, OA challenge induced the phosphorylation of ERK and Akt and the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in lung homogenates. After renal IRI, the phosphorylation of ERK and expression of COX-2 and iNOS were markedly reduced; however, there was no difference in the phosphorylation of Akt between sham and ischemic OA-challenged animals. Mucus production was also reduced in allergic mice after renal IRI. IL-4, IL-5 and IL-13 were markedly down-regulated in immunized/challenged mice subjected to IRI. These results suggest that renal IRI can modulate lung allergic inflammation, probably by altering the Th1/Th2 balance and, at least in part, by changing cellular signal transduction factors.

Effects of captopril on glucose metabolism and autophagy in liver and muscle from mice with type 1 diabetes and diet-induced obesity.

Impaired metabolic functions underlie the pathophysiology of diabetes and obesity. The renin-angiotensin system (RAS) is one pathway related to the pathophysiology of both diseases. RAS activation in metabolically active tissues exerts pro-inflammatory effects via angiotensin II (Ang II), linked to dysfunction in cellular processes such as autophagy, which is associated with obesity and diabetes. Here, we determined whether RAS is involved in metabolic dysregulations in a Type 1 Diabetes (T1D) mouse model, treated with captopril, and in an obesity mouse model (Agt-Tg) that overexpresses angiotensinogen (Agt) in adipose tissue. T1D mice had lower plasma leptin, resistin and higher non-esterified fatty acids (NEFA) compared to wild type (Wt) mice, even under captopril treatment. Further, mRNA levels for Agt, At1, Insr, and Beclin1 were upregulated in muscle and liver of T1D mice with captopril compared to Wt. Moreover, autophagy markers LC3 and p62 proteins were decreased, regardless of captopril treatment in the liver from T1D mice. In obese Wt mice, captopril increased muscle Irs1 gene levels. Further, captopril reduced mRNA levels of At1, Insr, Ampk, Beclin1, Atg12, and Lc3 in the liver from both Wt and Agt-Tg mice, while Agt, At1, Insr, and Atg12 expression was reduced in Agt-Tg mice without captopril treatment. Irs1 expression was decreased in the liver from obese Wt mice treated with captopril. Our results suggest that captopril treatment upregulates components of RAS, insulin signaling, and autophagy in both muscle and liver, indicating potential utility of captopril in targeting both insulin sensitivity and autophagy in diabetes and obesity.

Blockade of caspase cascade overcomes malaria-associated acute respiratory distress syndrome in mice.

Malaria is an enormous burden on global health that caused 409,000 deaths in 2019. Severe malaria can manifest in the lungs, an illness known as acute respiratory distress syndrome (ARDS). Not much is known about the development of malaria-associated ARDS (MA-ARDS), especially regarding cell death in the lungs. We had previously established a murine model that mimics various human ARDS aspects, such as pulmonary edema, hemorrhages, pleural effusion, and hypoxemia, using DBA/2 mice infected with Plasmodium berghei ANKA. Here, we explored the mechanisms and the involvement of apoptosis in this syndrome. We found that apoptosis contributes to the pathogenesis of MA-ARDS, primarily as facilitators of the alveolar-capillary barrier breakdown. The protection of pulmonary endothelium by inhibiting caspase activation could be a promising therapeutic strategy to prevent the pathogenicity of MA-ARDS. Therefore, intervention in the programmed death cell mechanism could help patients not to develop severe malaria.

XPR1 Mediates the Pancreatic ß-Cell Phosphate Flush.

Glucose-stimulated insulin secretion is the hallmark of the pancreatic ß-cell, a critical player in the regulation of blood glucose concentration. In 1974, the remarkable observation was made that an efflux of intracellular inorganic phosphate (Pi) accompanied the events of stimulated insulin secretion. The mechanism behind this "phosphate flush," its association with insulin secretion, and its regulation have since then remained a mystery. We recapitulated the phosphate flush in the MIN6m9 ß-cell line and pseudoislets. We demonstrated that knockdown of XPR1, a phosphate transporter present in MIN6m9 cells and pancreatic islets, prevented this flush. Concomitantly, XPR1 silencing led to intracellular Pi accumulation and a potential impact on Ca2+ signaling. XPR1 knockdown slightly blunted first-phase glucose-stimulated insulin secretion in MIN6m9 cells, but had no significant impact on pseudoislet secretion. In keeping with other cell types, basal Pi efflux was stimulated by inositol pyrophosphates, and basal intracellular Pi accumulated following knockdown of inositol hexakisphosphate kinases. However, the glucose-driven phosphate flush occurred despite inositol pyrophosphate depletion. Finally, while it is unlikely that XPR1 directly affects exocytosis, it may protect Ca2+ signaling. Thus, we have revealed XPR1 as the missing mediator of the phosphate flush, shedding light on a 45-year-old mystery.

AeMOPE-1, a Novel Salivary Peptide From Aedes aegypti, Selectively Modulates Activation of Murine Macrophages and Ameliorates Experimental Colitis.

The sialotranscriptomes of Aedes aegypti revealed a transcript overexpressed in female salivary glands that codes a mature 7.8 kDa peptide. The peptide, specific to the Aedes genus, has a unique sequence, presents a putative secretory nature and its function is unknown. Here, we confirmed that the peptide is highly expressed in the salivary glands of female mosquitoes when compared to the salivary glands of males, and its secretion in mosquito saliva is able to sensitize the vertebrate host by inducing the production of specific antibodies. The synthetic version of the peptide downmodulated nitric oxide production by activated peritoneal murine macrophages. The fractionation of a Ae. aegypti salivary preparation revealed that the fractions containing the naturally secreted peptide reproduced the nitric oxide downmodulation. The synthetic peptide also selectively interfered with cytokine production by murine macrophages, inhibiting the production of IL-6, IL-12p40 and CCL2 without affecting TNF-α or IL-10 production. Likewise, intracellular proteins associated with macrophage activation were also distinctively modulated: while iNOS and NF-κB p65 expression were diminished, IκBα and p38 MAPK expression did not change in the presence of the peptide. The anti-inflammatory properties of the synthetic peptide were tested in vivo on a dextran sulfate sodium-induced colitis model. The therapeutic administration of the Ae. aegypti peptide reduced the leukocytosis, macrophage activity and nitric oxide levels in the gut, as well as the expression of cytokines associated with the disease, resulting in amelioration of its clinical signs. Given its biological properties in vitro and in vivo, the molecule was termed Aedes-specific MOdulatory PEptide (AeMOPE-1). Thus, AeMOPE-1 is a novel mosquito-derived immunobiologic with potential to treat immune-mediated disorders.