Cathepsin D interacts with adenosine A2A receptors in mouse macrophages to modulate cell surface localization and inflammatory signaling.

Adenosine A2A receptor (A2AR)-dependent signaling in macrophages plays a key role in the regulation of inflammation. However, the processes regulating A2AR targeting to the cell surface and degradation in macrophages are incompletely understood. For example, the C-terminal domain of the A2AR and proteins interacting with it are known to regulate receptor recycling, although it is unclear what role potential A2AR-interacting partners have in macrophages. Here, we aimed to identify A2AR-interacting partners in macrophages that may effect receptor trafficking and activity. To this end, we performed a yeast two-hybrid screen using the C-terminal tail of A2AR as the "bait" and a macrophage expression library as the "prey." We found that the lysosomal protease cathepsin D (CtsD) was a robust hit. The A2AR-CtsD interaction was validated in vitro and in cellular models, including RAW 264.7 and mouse peritoneal macrophage (IPMΦ) cells. We also demonstrated that the A2AR is a substrate of CtsD and that the blockade of CtsD activity increases the density and cell surface targeting of A2AR in macrophages. Conversely, we demonstrate that A2AR activation prompts the maturation and enzymatic activity of CtsD in macrophages. In summary, we conclude that CtsD is a novel A2AR-interacting partner and thus describe molecular and functional interplay that may be crucial for adenosine-mediated macrophage regulation in inflammatory processes.

VEGF-A from Granuloma Macrophages Regulates Granulomatous Inflammation by a Non-angiogenic Pathway during Mycobacterial Infection.

Many autoimmune and infectious diseases are characterized by the formation of granulomas which are inflammatory lesions that consist of spatially organized immune cells. These sites protect the host and control pathogens like Mycobacterium tuberculosis (Mtb), but are highly inflammatory and cause pathology. Using bacille Calmette-Guerin (BCG) and Mtb infection in mice that induce sarcoid or caseating granulomas, we show that a subpopulation of granuloma macrophages produces vascular endothelial growth factor (VEGF-A), which recruits immune cells to the granuloma by a non-angiogenic pathway. Selective blockade of VEGF-A in myeloid cells, combined with granuloma transplantation, shows that granuloma VEGF-A regulates granulomatous inflammation. The severity of granuloma-related inflammation can be ameliorated by pharmaceutical or genetic inhibition of VEGF-A, which improves survival of mice infected with virulent Mtb without altering host protection. These data show that VEGF-A inhibitors could be used as a host-directed therapy against granulomatous diseases like tuberculosis and sarcoidosis, thereby expanding the value of already existing and approved anti-VEGF-A drugs.

Interplay between colonic inflammation and tachykininergic pathways in the onset of colonic dysmotility in a mouse model of diet-induced obesity.

BACKGROUND: The murine model of high fat diet (HFD)-induced obesity is characterized by an increment of intestinal permeability, secondary to an impairment of mucosal epithelial barrier and enteric inflammation, followed by morphofunctional rearrangement of the enteric nervous system. The present study investigated the involvement of abdominal macrophages in the mechanisms underlying the development of enteric dysmotility associated with obesity. METHODS: Wild type C57BL/6J mice were fed with HFD (60% kcal from fat) or normocaloric diet (NCD, 18% kcal from fat) for 8 weeks. Groups of mice fed with NCD or HFD were treated with clodronate encapsulated into liposomes to deplete abdominal macrophages. Tachykininergic contractions, elicited by electrical stimulation or exogenous substance P (SP), were recorded in vitro from longitudinal muscle colonic preparations. Substance P distribution was examined by confocal immunohistochemistry. The density of macrophages in the colonic wall was examined by immunohistochemical analysis. Malondialdehyde (MDA, colorimetric assay) and IL-1ß (ELISA assay) levels were also evaluated. RESULTS: MDA and IL-1ß levels were increased in colonic tissues from HFD-treated animals. In colonic preparations, electrically evoked tachykininergic contractions were enhanced in HFD mice. Immunohistochemistry displayed an increase in substance P immunoreactivity in myenteric ganglia, as well as in the muscular layers of colonic cryosections from obese mice. Macrophage depletion in HFD mice was associated with a significant reduction of colonic inflammation. In addition, the decrease in macrophage density attenuated the morphofunctional alterations of tachykininergic pathways observed in obese mice. CONCLUSION: Obesity elicited by HFD determines a condition of colonic inflammation, followed by a marked rearrangement of motor excitatory tachykininergic enteric nerves. Macrophage depletion counteracted the morphofunctional changes of colonic neuromuscular compartment, suggesting a critical role for these immune cells in the onset of enteric dysmotility associated with obesity.

Macrophage P2X4 receptors augment bacterial killing and protect against sepsis.

The macrophage is a major phagocytic cell type, and its impaired function is a primary cause of immune paralysis, organ injury, and death in sepsis. An incomplete understanding of the endogenous molecules that regulate macrophage bactericidal activity is a major barrier for developing effective therapies for sepsis. Using an in vitro killing assay, we report here that the endogenous purine ATP augments the killing of sepsis-causing bacteria by macrophages through P2X4 receptors (P2X4Rs). Using newly developed transgenic mice expressing a bioluminescent ATP probe on the cell surface, we found that extracellular ATP levels increase during sepsis, indicating that ATP may contribute to bacterial killing in vivo. Studies with P2X4R-deficient mice subjected to sepsis confirm the role of extracellular ATP acting on P2X4Rs in killing bacteria and protecting against organ injury and death. Results with adoptive transfer of macrophages, myeloid-specific P2X4R-deficient mice, and P2rx4 tdTomato reporter mice indicate that macrophages are essential for the antibacterial, antiinflammatory, and organ protective effects of P2X4Rs in sepsis. Pharmacological targeting of P2X4Rs with the allosteric activator ivermectin protects against bacterial dissemination and mortality in sepsis. We propose that P2X4Rs represent a promising target for drug development to control bacterial growth in sepsis and other infections.

Lithocholic acid, a bacterial metabolite reduces breast cancer cell proliferation and aggressiveness.

Our study aimed at finding a mechanistic relationship between the gut microbiome and breast cancer. Breast cancer cells are not in direct contact with these microbes, but disease could be influenced by bacterial metabolites including secondary bile acids that are exclusively synthesized by the microbiome and known to enter the human circulation. In murine and bench experiments, a secondary bile acid, lithocholic acid (LCA) in concentrations corresponding to its tissue reference concentrations (< 1 µM), reduced cancer cell proliferation (by 10-20%) and VEGF production (by 37%), aggressiveness and metastatic potential of primary tumors through inducing mesenchymal-to-epithelial transition, increased antitumor immune response, OXPHOS and the TCA cycle. Part of these effects was due to activation of TGR5 by LCA. Early stage breast cancer patients, versus control women, had reduced serum LCA levels, reduced chenodeoxycholic acid to LCA ratio, and reduced abundance of the baiH (7α/ß-hydroxysteroid dehydroxylase, the key enzyme in LCA generation) gene in fecal DNA, all suggesting reduced microbial generation of LCA in early breast cancer.

Adenosine receptors differentially regulate type 2 cytokine production by IL-33-activated bone marrow cells, ILC2s, and macrophages.

Group 2 innate lymphoid cells (ILC2s) represent a rapid source of type 2 cytokines, such as IL-5 and IL-13, and play an important role in orchestrating type 2 immune response. Adenosine is an endogenous purine nucleoside, a catabolite of ATP that binds and activates ≥1 of 4 transmembrane G protein-coupled cell-surface adenosine receptors (ARs)-A1, A2A, A2B, and A3. Here, we studied the role of ARs in the regulation of cytokine production by ILC2s. We found that A2BARs suppress the production of both IL-5 and IL-13 by ILC2s, whereas A2AARs augment IL-5 production and fail to affect IL-13 release. Combined stimulation of all ARs led to the suppression of both IL-5 and IL-13 production, which indicated that A2BARs dominate A2AARs. Both pre- and post-transcriptional processes may be involved in the AR modulation of ILC2 IL-5 and IL-13 production. Thus, we identify adenosine as a novel negative regulator of ILC2 activation.-Csóka, B., Németh, Z. H., Duerr, C. U., Fritz, J. H., Pacher, P., Haskó, G. Adenosine receptors differentially regulate type 2 cytokine production by IL-33-activated bone marrow cells, ILC2s, and macrophages.

A2A adenosine receptors control pancreatic dysfunction in high-fat-diet-induced obesity.

Adenosine, a key extracellular signaling mediator, regulates several aspects of metabolism by activating 4 G-protein-coupled receptors, the A1, A2A, A2B, and A3 adenosine receptors (ARs). The role of A2AARs in regulating high-fat-diet (HFD)-induced metabolic derangements is unknown. To evaluate the role of A2AARs in regulating glucose and insulin homeostasis in obesity, we fed A2AAR-knockout (KO) and control mice an HFD for 16 wk to initiate HFD-induced metabolic disorder. We found that genetic deletion of A2AARs caused impaired glucose tolerance in mice fed an HFD. This impaired glucose tolerance was caused by a decrease in insulin secretion but not in insulin sensitivity. Islet size and insulin content in pancreata of A2AAR-deficient mice were decreased compared with control mice after consuming an HFD. A2AAR-KO mice had decreased expression of the ß-cell-specific markers pdx1, glut2, mafA, and nkx6.1 and increased expression of the dedifferentiation markers sox2 and hes1. Ex vivo islet experiments confirmed the role of A2AARs in protecting against decreased insulin content and release caused by HFD. Other experiments with bone marrow chimeras revealed that inflammation was not the primary cause of decreased insulin secretion in A2AAR-KO mice. Altogether, our data showed that A2AARs control pancreatic dysfunction in HFD-induced obesity.-Csóka, B., Töro, G., Vindeirinho, J., Varga, Z. V., Koscsó, B., Németh, Z. H., Kókai, E., Antonioli, L., Suleiman, M., Marchetti, P., Cseri, K., Deák, Á., Virág, L., Pacher, P., Bai, P., Haskó, G. A2A adenosine receptors control pancreatic dysfunction in high-fat-diet-induced obesity.

Colonic motor dysfunctions in a mouse model of high-fat diet-induced obesity: an involvement of A2B adenosine receptors.

Adenosine A2B receptors (A2BR) regulate several enteric functions. However, their implication in the pathophysiology of intestinal dysmotility associated with high-fat diet (HFD)-induced obesity has not been elucidated. We investigated the expression of A2BR in mouse colon and their role in the mechanisms underlying the development of enteric dysmotility associated with obesity. Wild-type C57BL/6J mice were fed with HFD (60% kcal from fat) or normocaloric diet (NCD; 18% kcal from fat) for 8 weeks. Colonic A2BR localization was examined by immunofluorescence. The role of A2BR in the control of colonic motility was examined in functional experiments on longitudinal muscle preparations (LMPs). In NCD mice, A2BR were predominantly located in myenteric neurons; in HFD animals, their expression increased throughout the neuromuscular layer. Functionally, the A2BR antagonist MRS1754 enhanced electrically induced NK1-mediated tachykininergic contractions in LMPs from HFD mice, while it was less effective in tissues from NCD mice. The A2B receptor agonist BAY 60-6583 decreased colonic tachykininergic contractions in LMPs, with higher efficacy in preparations from obese mice. Both A2BR ligands did not affect contractions elicited by exogenous substance P. Obesity is related with a condition of colonic inflammation, leading to an increase of A2BR expression. A2BR, modulating the activity of excitatory tachykininergic nerves, participate to the enteric dysmotility associated with obesity.

Extracellular ATP protects against sepsis through macrophage P2X7 purinergic receptors by enhancing intracellular bacterial killing.

Extracellular ATP binds to and signals through P2X7 receptors (P2X7Rs) to modulate immune function in both inflammasome-dependent and -independent manners. In this study, P2X7(-/-) mice, the pharmacological agonists ATP-magnesium salt (Mg-ATP; 100 mg/kg, EC50 ≈ 1.32 mM) and benzoylbenzoyl-ATP (Bz-ATP; 10 mg/kg, EC50 ≈ 285 µM), and antagonist oxidized ATP (oxi-ATP; 40 mg/kg, IC50 ≈ 100 µM) were used to show that P2X7R activation is crucial for the control of mortality, bacterial dissemination, and inflammation in cecal ligation and puncture-induced polymicrobial sepsis in mice. Our results with P2X7(-/-) bone marrow chimeric mice, adoptive transfer of peritoneal macrophages, and myeloid-specific P2X7(-/-) mice indicate that P2X7R signaling on macrophages is essential for the protective effect of P2X7Rs. P2X7R signaling protects through enhancing bacterial killing by macrophages, which is independent of the inflammasome. By using the connexin (Cx) channel inhibitor Gap27 (0.1 mg/kg, IC50 ≈ 0.25 µM) and pannexin channel inhibitor probenecid (10 mg/kg, IC50 ≈ 11.7 µM), we showed that ATP release through Cx is important for inhibiting inflammation and bacterial burden. In summary, targeting P2X7Rs provides a new opportunity for harnessing an endogenous protective immune mechanism in the treatment of sepsis.

Adenosine signalling in diabetes mellitus--pathophysiology and therapeutic considerations.

Adenosine is a key extracellular signalling molecule that regulates several aspects of tissue function by activating four G-protein-coupled receptors, A1, A2A, A2B and A1 adenosine receptors. Accumulating evidence highlights a critical role for the adenosine system in the regulation of glucose homeostasis and the pathophysiology of type 1 diabetes mellitus (T1DM) and type 2 diabetes mellitus (T2DM). Although adenosine signalling is known to affect insulin secretion, new data indicate that adenosine signalling also contributes to the regulation of ß-cell homeostasis and activity by controlling the proliferation and regeneration of these cells as well as the survival of ß cells in inflammatory microenvironments. Furthermore, adenosine is emerging as a major regulator of insulin responsiveness by controlling insulin signalling in adipose tissue, muscle and liver; adenosine also indirectly mediates effects on inflammatory and/or immune cells in these tissues. This Review critically discusses the role of the adenosine-adenosine receptor system in regulating both the onset and progression of T1DM and T2DM, and the potential of pharmacological manipulation of the adenosinergic system as an approach to manage T1DM, T2DM and their associated complications.

CD39 improves survival in microbial sepsis by attenuating systemic inflammation.

Sepsis remains the leading cause of morbidity and mortality in critically ill patients. Excessive inflammation is a major cause of organ failure and mortality in sepsis. Ectonucleoside triphosphate diphosphohydrolase 1, ENTPDase1 (CD39) is a cell surface nucleotide-metabolizing enzyme, which degrades the extracellular purines ATP and ADP, thereby regulating purinergic receptor signaling. Although the role of purinergic receptor signaling in regulating inflammation and sepsis has been addressed previously, the role of CD39 in regulating the host's response to sepsis is unknown. We found that the CD39 mimic apyrase (250 U/kg) decreased and knockout or pharmacologic blockade with sodium polyoxotungstate (5 mg/kg; IC50 ≈ 10 µM) of CD39 increased mortality of mice with polymicrobial sepsis induced by cecal ligation and puncture. CD39 decreased inflammation, organ damage, immune cell apoptosis, and bacterial load. Use of bone marrow chimeric mice revealed that CD39 expression on myeloid cells decreases inflammation in septic mice. CD39 expression is upregulated during sepsis in mice, as well as in both murine and human macrophages stimulated with Escherichia coli. Moreover, E. coli increases CD39 promoter activity in macrophages. Altogether, these data indicate CD39 as an evolutionarily conserved inducible protective pathway during sepsis. We propose CD39 as a novel therapeutic target in the management of sepsis.

A2B adenosine receptor induces protective antihelminth type 2 immune responses.

The type 2 immune response evoked by intestinal nematode parasites contributes to worm expulsion and tolerance to associated tissue damage. We investigated whether this host response is affected by blocking signaling by the putative endogenous danger signal adenosine, which can be released during inflammation and host cell damage. Specific blockade of the A2B adenosine receptor (A2BAR) inhibited worm elimination and the development of innate and adaptive components of the type 2 primary and memory response. Infected mice lacking A2BAR exhibited decreased M2 macrophage and eosinophil recruitment and reduced IL-4 and IL-13 cytokine production. Additionally, shortly after infection, upregulation of the alarmin IL-33, which drives type 2 immunity, and activation of innate lymphoid type 2 (ILC2) cells was inhibited, while exogenous IL-33 restored ILC2 cell activation and type 2 cytokine expression. Thus, adenosine acts as a danger-associated molecular pattern (DAMP) that initiates helminth-induced type 2 immune responses through A2BAR.

Adenosine and inflammation: what's new on the horizon?

Adenosine contributes to the maintenance of tissue integrity by modulating the immune system. Encouraging results have emerged with adenosine receptor ligands for the management of several inflammatory conditions in preclinical and clinical settings. However, therapeutic applications of these drugs are sometimes complicated by the occurrence of serious adverse effects. The scientific community is making intensive efforts to design novel adenosine receptor ligands endowed with greater selectivity or to develop innovative compounds acting as allosteric receptor modulators. In parallel, research is focusing on novel pharmacological entities (designated as adenosine-regulating agents) that can increase, in a site- and event-specific manner, adenosine concentrations at the inflammatory site, thereby minimizing the adverse systemic effects of adenosine.

A2B adenosine receptors prevent insulin resistance by inhibiting adipose tissue inflammation via maintaining alternative macrophage activation.

Obesity causes increased classical and decreased alternative macrophage activation, which in turn cause insulin resistance in target organs. Because A2B adenosine receptors (ARs) are important regulators of macrophage activation, we examined the role of A2B ARs in adipose tissue inflammation and insulin resistance. A2B AR deletion impaired glucose and lipid metabolism in mice fed chow but not a high-fat diet, which was paralleled by dysregulation of the adipokine system, and increased classical macrophage activation and inhibited alternative macrophage activation. The expression of alternative macrophage activation-specific transcriptions factors, including CCAAT/enhancer-binding protein-ß, interferon regulatory factor 4, and peroxisome proliferator-activated receptor-γ, was decreased in adipose tissue of A2B AR-deficient mice. Furthermore, in in vitro studies, we found that stimulation of A2B ARs suppressed free fatty acid-induced deleterious inflammatory and metabolic activation of macrophages. Moreover, AR activation upregulated the interleukin-4-induced expression of CCAAT/enhancer-binding protein-ß, interferon regulatory factor 4, and peroxisome proliferator-activated receptor-γ in macrophages. Altogether, our results indicate that therapeutic strategies targeting A2B ARs hold promise for preventing adipose tissue inflammation and insulin resistance.

Adenosine augments IL-10-induced STAT3 signaling in M2c macrophages.

The alternatively activated macrophage phenotype induced by IL-10 is called M2c. Adenosine is an endogenous purine nucleoside that accumulates in the extracellular space in response to metabolic disturbances, hypoxia, inflammation, physical damage, or apoptosis. As adenosine is known to regulate classically activated M1 and IL4- and IL-13-activated M2a macrophages, the goal of the present study was to explore its effects on M2c macrophages. We found that adenosine augmented the IL-10-induced expression of TIMP-1 and arginase-1 by the mouse macrophage cell line RAW 264.7 and by mouse BMDMs. The effects of AR stimulation on IL-10-induced TIMP-1 or arginase-1 expression were lacking in A2BAR KO macrophages. The role of A2BAR on TIMP-1 production of RAW 264.7 cells was confirmed with specific agonist BAY606583 and antagonist PSB0788. AR stimulation augmented IL-10-induced STAT3 phosphorylation in macrophages, and pharmacological inhibition or silencing of STAT3 using siRNA reduced the stimulatory effect of AR stimulation on TIMP-1 production. In contrast to its stimulatory effect on IL-10-induced STAT3 activation, adenosine inhibited IL-6-induced STAT3 phosphorylation and SAA3 expression. In conclusion, adenosine enhances IL-10-induced STAT3 signaling and M2c macrophage activation.

Stimulation of A2B adenosine receptors protects against trauma-hemorrhagic shock-induced lung injury.

Inflammation is responsible for secondary organ failure after trauma and hemorrhagic shock (T/HS). Adenosine, acting through four G protein-coupled cell surface receptors, A1, A2A, A2B, and A3, exerts a number of tissue protective and anti-inflammatory effects. The goal of the present study was to test the effect of A2B adenosine receptor stimulation on T/HS-induced organ injury and inflammation in rats. Rats after T/HS were resuscitated with Ringer's lactate containing the A2B receptor agonist BAY 60-6583 or its vehicle. We found that BAY 60-6583 decreased T/HS-induced lung permeability and plasma creatine kinase levels but failed to affect T/HS-induced lung neutrophil infiltration and IκBα expression and plasma alanine aminotransferase levels. Thus, we conclude that stimulation of A2B receptors protects against T/HS-induced lung and muscle injury.

Adenosine promotes alternative macrophage activation via A2A and A2B receptors.

Adenosine has been implicated in suppressing the proinflammatory responses of classically activated macrophages induced by Th1 cytokines. Alternative macrophage activation is induced by the Th2 cytokines interleukin (IL)-4 and IL-13; however, the role of adenosine in governing alternative macrophage activation is unknown. We show here that adenosine treatment of IL-4- or IL-13-activated macrophages augments the expression of alternative macrophage markers arginase-1, tissue inhibitor of matrix metalloproteinase-1 (TIMP-1), and macrophage galactose-type C-type lectin-1. The stimulatory effect of adenosine required primarily A(2B) receptors because the nonselective adenosine receptor agonist 5'-N-ethylcarboxamidoadenosine (NECA) increased both arginase activity (EC(50)=261.8 nM) and TIMP-1 production (EC(50)=80.67 nM), and both pharmacologic and genetic blockade of A(2B) receptors prevented the effect of NECA. A(2A) receptors also contributed to the adenosine augmentation of IL-4-induced TIMP-1 release, as both adenosine and NECA were less efficacious in augmenting TIMP-1 release by A(2A) receptor-deficient than control macrophages. Of the transcription factors known to drive alternative macrophage activation, CCAAT-enhancer-binding protein ß was required, while cAMP response element-binding protein and signal transducer and activator of transcription 6 were dispensable in mediating the effect of adenosine. We propose that adenosine receptor activation suppresses inflammation and promotes tissue restitution, in part, by promoting alternative macrophage activation.

Adenosine augments IL-10 production by microglial cells through an A2B adenosine receptor-mediated process.

Microglia are activated by pathogen-associated molecular patterns and produce proinflammatory cytokines, such as TNF-α, IL-6, and IL-12, and the anti-inflammatory cytokine IL-10. Adenosine is an endogenous purine nucleoside and a ligand of four G protein-coupled adenosine receptors (ARs), which are the A(1)AR, A(2A)AR, A(2B)AR, and A(3)AR. ARs have been shown to suppress TNF-α production by microglia, but their role in regulating IL-10 production has not been studied. In this study, we demonstrate that adenosine augments IL-10 production by activated murine microglia while suppressing the production of proinflammatory cytokines. Because the order of potency of selective AR agonists in inducing IL-10 production was NECA > IB-MECA > CCPA ≥ CGS21680, and the A(2B)AR antagonist MRS1754 prevented the effect of NECA, we conclude that the stimulatory effect of adenosine on IL-10 production is mediated by the A(2B)AR. Mechanistically, adenosine augmented IL-10 mRNA accumulation by a transcriptional process. Using mutant IL-10 promoter constructs we showed that a CREB-binding region in the promoter mediated the augmenting effect of adenosine on IL-10 transcription. Chromatin immunoprecipitation analysis demonstrated that adenosine induced CREB phosphorylation at the IL-10 promoter. Silencing CREB using lentivirally delivered short hairpin RNA blocked the enhancing effect of adenosine on IL-10 production, confirming a role for CREB in mediating the stimulatory effect of adenosine on IL-10 production. In addition, adenosine augmented IL-10 production by stimulating p38 MAPK. Collectively, our results establish that A(2B)ARs augment IL-10 production by activated murine microglia.

Ecto-5'-nucleotidase (CD73) decreases mortality and organ injury in sepsis.

The extracellular concentrations of adenosine are increased during sepsis, and adenosine receptors regulate the host's response to sepsis. In this study, we investigated the role of the adenosine-generating ectoenzyme, ecto-5'-nucleotidase (CD73), in regulating immune and organ function during sepsis. Polymicrobial sepsis was induced by subjecting CD73 knockout (KO) and wild type (WT) mice to cecal ligation and puncture. CD73 KO mice showed increased mortality in comparison with WT mice, which was associated with increased bacterial counts and elevated inflammatory cytokine and chemokine concentrations in the blood and peritoneum. CD73 deficiency promoted lung injury, as indicated by increased myeloperoxidase activity and neutrophil infiltration, and elevated pulmonary cytokine levels. CD73 KO mice had increased apoptosis in the thymus, as evidenced by increased cleavage of caspase-3 and poly(ADP-ribose) polymerase and increased activation of NF-κB. Septic CD73 KO mice had higher blood urea nitrogen levels and increased cytokine levels in the kidney, indicating increased renal dysfunction. The increased kidney injury of CD73 KO mice was associated with augmented activation of p38 MAPK and decreased phosphorylation of Akt. Pharmacological inactivation of CD73 in WT mice using α, ß-methylene ADP augmented cytokine levels in the blood and peritoneal lavage fluid. These findings suggest that CD73-derived adenosine may be beneficial in sepsis.