Adenosine Levels Research Articles

Adenosine accumulation in the blood of newborn mice weakens antimicrobial host defenses.

Pediatric intensive care patients are particularly susceptible to severe bacterial infections because of ineffective neutrophil responses. The reasons why neutrophils of newborns are less responsive than those of adults are not clear. Because adenosine triphosphate (ATP) and adenosine (ADO) tightly regulate neutrophils, we studied whether the ATP and ADO levels in the blood of newborn mice could impair the function of their neutrophils. We observed significant changes in plasma ATP and ADO levels throughout the lifespan of mice. ADO levels in newborns were significantly higher than in older mice, while ATP levels were significantly lower. These changes were particularly striking in newborn and juvenile mice with ATP and ADO levels of about 80 and 600 nM in newborns versus 130 and 190 nM in juveniles, respectively. The ratios of the ATP versus ADO levels of newborns were (with 0.2) significantly lower than those of juveniles (1.4) and adults (0.5). These low ATP/ADO ratios correlated with significantly weakened neutrophil activation responses following in vitro stimulation with a formyl peptide receptor agonist and a markedly higher morbidity and mortality rate of newborns following bacterial infection. We found that enhanced AMP hydrolysis via CD73, a lack of ADO breakdown by adenosine deaminase, and reduced ADO uptake by nucleoside transporters are responsible for the low ATP/ADO ratios in blood of newborn mice. We conclude that the extracellular ADO accumulation in newborn mice impairs inflammatory responses and reduces the ability of neutrophils to mount effective antimicrobial defenses against bacterial infections.

Illuminating the impact of CD38-induced adenosine formation in B-cell lymphoma

The expression of CD38 by cancer cells may mediate an immune-suppressive effect by producing Extracellular Adenosine (ADO) acting through G-protein-coupled cell surface receptors on cellular components and tumor cells. This can increase PD-1 expression and interaction with PD-L1, suppressing CD8 + cytotoxic T cells. This study examines the impact of heightened CD38 expression and extracellular ADO on various hematological and clinical parameters in patients with mature B-cell lymphoma, alongside their correlation with the soluble counterparts of the PD-1/PD-L1 axis. Our study was conducted on 90 patients, CD38-positive and CD38-negative (measured by flow cytometry), with mature B-cell lymphoma divided into CLL and B-NHL subtypes. Their serum ADO, soluble PD-1, and PD-L1 levels were measured using a sandwich ELISA. Our study revealed a positive correlation between CD38 expression, sADO, sPD-1, and sPD-L1 in mature B-cell lymphoma patients. CD38-positive patients had higher sADO, sPD-1, and sPD-L1 levels. Higher CD38 expression and extracellular ADO negatively affected HB level and PLT count and positively correlated with the higher risk stratification in mature B-cell lymphoma patients. This study explored the potential impact of CD38 expression and elevated extracellular ADO on B-cell lymphoma alongside their link with the PD-1/PD-L1 axis. Our findings underscore the influence of extracellular ADO on the neoplastic process of mature B-cell lymphoma. We also propose targeting the CD38-induced-ADO formation pathway, which could serve as a promising therapeutic immune target with multifaceted effects within mature B-cell neoplasms.

Antagonists of CD39 and CD73 potentiate doxycycline repositioning to induce a potent antitumor immune response.

Studies have reported that cellular metabolism at the tumor-immune microenvironment (TiME) serves as a critical checkpoint and perturbs/supports anti-cancer immunity. Extra cellular ATP (eATP) may mediate anti-cancer immune response; however, its catabolism by ectonucleotidase generates immunosuppressive adenosine. In the presented work, we have tried to repurpose doxycycline with or without an antagonist of ectonucleotidase for mitigating ATP metabolism and immunosuppression. In this methodology eATP and adenosine levels were quantified. Bone marrow-derived M1 and M2 polarized macrophages were maintained in tumor mimicking condition (TMC). Total/CD4+Tcells were co-cultured with macrophages to understand the impact of doxycycline and/or antagonist of ectonucleotidase on T cell/subset differentiation. Preclinical efficacy of doxycycline and/or ectonucleotidase antagonist and their synergy was scored in 4 T1-induced breast carcinoma. We found that Doxycycline manipulated macrophage polarization by decreasing the frequency of CD206+M2 macrophages, which resulted in enhanced CD4+ directed CD8+ T cell response. Doxycycline alleviated the expression of CD39 and CD73, rescuing ATP catabolism. Doxycycline delayed tumor growth by enhancing F4/80+ CD86+ M1 macrophages and subsequently anti-tumor Tbet+ CD4+Tcells, attenuating the frequency of FOXP3+ regulatory T cells, which was cooperatively supported by ARL67156 and AMPCP (CD39 and CD73 antagonist).A synergy was observed with ARL67156 and AMPC Pensuring a possibility of using doxycycline alone or in combination with an antagonist of ectonucleotidase to present adenosine-mediated immunosuppression. Subsequently, our finding indicated that prospective usage of doxycycline as a novel metabolic checkpoint blocker (IMB) against ectonucleotidase and may be modified/delivered appropriately as a monotherapy or in combination with antagonists of ectonucleotidases as an IMB.

Salinity stress impairs disease resistance in white shrimp, Penaeus vannamei through AMPK pathway, ameliorated by dietary glucose-mediated energy homeostasis.

This study presents a comprehensive examination of the physiological adaptations of white shrimp (Penaeus vannamei) to low-salinity conditions and evaluates the effects of supplementing dietary glucose on disease resistance. Compared to the control group, shrimp cultured at a salinity of 4psu exhibit significantly elevated expression levels of adenosine 5'-monophosphate-activated protein kinase (AMPK) in the hepatopancreas, which leads to increased energy expenditure and a corresponding reduction in resistance to infection by Vibrio alginolyticus. The suppression of AMPK via dsAMPK treatment markedly enhances disease resistance. Moreover, shrimp raised in low salinity conditions exhibit downregulation of mTOR-associated molecules, including Lipin-1 and hypoxia-inducible factor 1-α (HIF-1α), both of which are essential for immune regulation. Metabolic assessments revealed reduced ATP levels and disrupted ATP/AMP and ATP/ADP ratios, indicating energy imbalance under low salinity stress. Notably, supplementing the diet with 1% glucose significantly increased glycogen reserves and ATP content, stabilized hemolymph glucose levels, and upregulated glycolysis-related genes, thereby optimizing energy metabolism and enhancing resilience to stress. This study underscores that AMPK activation in response to low salinity conditions leads to increased energy expenditure, which in turn lowers disease resistance. Furthermore, it underscores the critical role of strategic dietary management in maintaining energy homeostasis and improving disease resistance in white shrimp under stressful environmental conditions associated with climate change, offering valuable insights for aquaculture nutrition strategies.

Review on Allopurinol in the Treatment of Gout

Gout and pain are synonymous, and a study in this issue of the BJP reports a novel anti-nociceptive effect of allopurinol, the drug most commonly used to treat gout. Allopurinol works by inhibiting xanthine oxidase (XO), the enzyme responsible for converting hypoxanthine to uric acid which is deposited as crystals in the joints of gout sufferers. Hypoxanthine is a metabolite of, and a possible precursor to, adenosine. find that acute inhibition of XO with allopurinol produces a modest adenosine A1 receptor-mediated anti-nociceptive effect in common tests of chemical and thermal nociception in mice. A concomitant increase in cerebrospinal fluid levels of adenosine supports their hypothesis that inhibiting XO increases adenosine levels via salvage from hypoxanthine. Elevating endogenous adenosine levels by inhibiting metabolism is a well-established strategy for producing anti-nociception in many preclinical models, but inhibiting XO is likely to be particularly beneficial in some chronic pain states because of the pro-nociceptive reactive oxygen species that are produced by XO activity. Thus, allopurinol may have unexpected benefits in pain associated with chronic inflammation, diabetes and vascular dysfunction.

Investigation of the anti-Huanglongbing effects using antimicrobial lipopeptide and phytohormone complex powder prepared from Bacillus amyloliquefaciens MG-2 fermentation

Global citrus production has been severely affected by citrus Huanglongbing (HLB) disease, caused by Candidatus Liberibacter asiaticus (Clas), and the development of effective control methods are crucial. This study employed antimicrobial lipopeptide and phytohormone complex powder (L1) prepared from the fermentation broth of the endophytic plant growth promoting bacterium (PGPB) of Bacillus amyloliquefaciens strain MG-2 to treat Candidatus Liberibacter asiaticus (CLas)-infected ‘Citrus reticulata ‘Chun Jian’ plants. Real-time fluorescence quantitative polymerase chain reaction (qPCR) and PCR were employed for disease detection. The results revealed that after 15 spray-drench treatments with L1 solution, the HLB infection rate decreased from 100 to 50%, the bacterial titer decreased by 51.9% compared with a 27.9% decrease in the control group. L1 treatment triggered the production of reactive oxygen species, increased lignin content, and increased defense enzyme activities (p < 0.05). Defense-related gene expression significantly increased within 12 h of treatment. In addition, L1 application also promoted plant growth, as evidenced by higher transpiration rates and net photosynthetic rates as well as increased leave or root density. Root flora analysis revealed that the abundances of Burkholderia_thailandensis, unclassified_g_Burkholderia-Caballeronia-Paraburkholderia, unclassified_g__Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium, and Pseudomonas_mosselii were 1.64, 1.46, 5.84, and 6.93 times greater, respectively, than those in the control group. The levels of phenylpropanoids, polyketides, lipids, lipid-like molecules, organic acids, and derivatives, significantly increased following L1 treatment (FC > 2, p < 0.05). Additionally, salicylic acid, dihydrojasmonic acid, and isopentenyl adenosine levels in leaves markedly increased. High-performance liquid chromatography (HPLC) confirmed that L1 contained surfactin, iturin and fengycin cyclic-lipopeptides (CLPs) as well as indole-3-acetic acid (IAA), 3-indolebutyric acid (IBA), indole-3-carboxylic acid and indole-3-carboxaldehyde auxins, N6-entopentenyladenine and t-zeatin-riboside cytokinins, abscisic acid, 1-aminocyclicpanecarboxylic acid, salicylic acid, and gibberellin A1, A3 and A4 phytohormones. These findings provide insight into multiple mechanisms by which endophytic Bacillus PGPB L1 is able to combat HLB disease, to promote citrus plant growth, and to optimize the root flora for soil health which offering an innovative strategy for sustainable management of this severe disease and improving citrus plant growth and productivity

Abstract IA013: Discovery of the dual A2A/A2B receptor antagonist MK-1088 for the treatment of solid tumors

Abstract In the tumor microenvironment (TME), adenosine levels have been shown to be elevated relative to normal tissues. This increase in adenosine levels renders an immunosuppressive effect via direct effects on T cells via agonism of the A2A receptor and indirect effects via agonism of both A2A and A2B receptors on myeloid cells. With these observations as a backdrop, we sought to develop a dual A2A/A2B receptor antagonist with properties that would enable maintenance of high levels of target engagement of the A2A and A2B receptors even at trough concentration. Drawing on our organization’s significant prior experience developing A2A receptor antagonists for the potential treatment of Parkinson’s disease, we developed MK-1088, a highly potent A2A/A2B dual receptor antagonist that was purposefully designed to possess excellent selectivity over the related A1 and A3 receptors. This presentation will detail the discovery and development strategy that was employed to identify molecules that met the profile exemplified by this molecule. A single ascending dose study of MK-1088 in healthy human volunteers demonstrated our ability to achieve, at trough concentration, >99% target engagement (TE) at the A2A receptor and >90% TE at the A2B receptor. Details of the pharmacokinetics, safety, and tolerability from this study will be highlighted. Citation Format: Duane E. DeMong, Sheila Ranganath, Jared Cumming, Matthew Larsen, Yonglian Zhang, Christopher Plummer, Amjad Ali, Anthony Palmieri, Evan Barry, Pierre Daublain, Pranav Gupta, Manash Chatterjee, Jeremy Presland, Sebastian Schneider, Paul Ciaccio, Daniel Tatosian, Aaron Sather, Ben Turnbull, Steven Silverman, Harry Chobanian, Harini Krishnamurthy, Richard Wnek, Stephen Crowley, Alita Miller, Mark Ayers, Marlene Hinton, Jill Chrencik, Sylvie Rottey, Jennifer O'Neil. Discovery of the dual A2A/A2B receptor antagonist MK-1088 for the treatment of solid tumors [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Optimizing Therapeutic Efficacy and Tolerability through Cancer Chemistry; 2024 Dec 9-11; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(12_Suppl):Abstract nr IA013.

Abstract PR001: Discovery of Etrumadenant, a first-in-class dual A2a and A2b adenosine receptor antagonist for cancer immunotherapy

Abstract High levels of adenosine are generated in the tumor microenvironment (TME) by sequential hydrolysis of extracellular ATP by the ecto-nucleotidases CD39 (ATP→AMP) and CD73 (AMP→adenosine). Adenosine activates A2a and A2b adenosine receptors on immune cells, resulting in immunosuppression. Since A2aR is expressed by a variety of lymphocytes, the suppressive effects of adenosine on this cell type can be potentially reversed by blocking the A2aR. In contrast, myeloid cells that have comparable expression of A2aR and A2bR require dual A2a/bR blockade for full reversal of the immunosuppressive effects of adenosine. The first generation of adenosine antagonists deployed in oncology were repurposed CNS drug candidates with properties that were deemed less than optimal for anti-tumor efficacy (e.g., A2aR-selective, good BBB permeability, not optimized for minimal plasma protein binding, etc.). We sought to design a potent and selective dual antagonist of A2aR and A2bR to effectively block the high concentrations of adenosine found in tumors, with diminished brain penetration, and minimal non-specific binding to plasma proteins. Using a pharmacophore mapping approach, we discovered potent A2aR antagonists with favorable PK profile in rats, good selectivity, and moderate potency against A2bR. Systematic SAR studies led to improvement in A2bR potency, which ultimately resulted in Etrumadenant (AB928). Etrumadenant inhibits both A2aR and A2bR with similar potencies (KB: 1.4 nM and 2 nM, respectively), and is highly selective against other targets in the adenosine pathway with minimal penetration across the blood brain barrier. Etrumadenant provides superior dendritic cell activity compared to A2aR antagonist alone. Its combination with immunogenic chemotherapy enhances immune activity and suppresses tumor growth and metastasis in vivo. These data support our rational design of a dual A2a and A2b adenosine receptor antagonist to impact adenosine biology on multiple cell types within the TME. Etrumadenant is currently undergoing phase 2 clinical trials in colorectal and lung cancer patients. Citation Format: Ehesan U. Sharif, Dillon H. Miles, Brandon R. Rosen, Joel Beatty, Jenna L. Jeffrey, Laurent P. P. Debien, Rhiannon Thomas-Tran, Debashis Mandal, Dan Direnzo, Sachie Marubayashi, Kristen Zhang, Ferdie Soriano, Elaine Ginn, Divyank Soni, Pei-Yu Chen, Lixia Jin, Steve W. Young, Matt J. Walters, Manmohan R. Leleti, Jay P. Powers. Discovery of Etrumadenant, a first-in-class dual A2a and A2b adenosine receptor antagonist for cancer immunotherapy. [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Optimizing Therapeutic Efficacy and Tolerability through Cancer Chemistry; 2024 Dec 9-11; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(12_Suppl):Abstract nr PR001

The JNK Signaling Pathway Regulates Seizures Through ENT1 in Pilocarpine-Induced Epilepsy Rat Model.

The study investigates whether the expression and function of ENT1 can be regulated by inhibiting the JNK signaling pathway, thereby altering the levels of extracellular adenosine and glutamate in neurons, and subsequently affecting the progression of epilepsy. The adult male SD rats were randomly divided into four groups: EP + SP600125 group, EP + DMSO group, EP group, and normal control group. The expression levels of ENT1, p-JNK, and JNK in the hippocampus of rats from each experimental group were detected using Western blotting technology. The expression and localization of ENT1 and p-JNK in the CA1, CA3, and DG areas of the hippocampus were detected by immunohistochemical staining and immunofluorescence staining. Microdialysis combined with liquid chromatography-mass spectrometry was used to determine the concentrations of adenosine and glutamate in the extracellular fluid of hippocampus in each experimental group. This study showed that the JNK-specific inhibitor SP600125 could reduce ENT1 expression and seizure intensity in experimental rats. Statistical analysis confirmed that adenosine and glutamate levels in the extracellular fluid of the hippocampus increased significantly after seizures in rats, and the JNK-specific inhibitor SP600125 could increase adenosine levels in the extracellular fluid but decrease glutamate levels. The JNK-specific inhibitor SP600125 can specifically inhibit the JNK signaling pathway and reduce the expression of ENT1 transporter. The mechanism is related to the transport of adenosine from the extracellular space to the intracellular space by ENT1 during epileptic states. Inhibition of ENT1 can increase the concentration of adenosine in the extracellular fluid of the hippocampus. The increase in adenosine concentration stopped glutamate from being released and reduced the amount of glutamate in the outside of the cell.

Extracellular adenosine oppositely regulates the purinome machinery in glioblastoma and mesenchymal stem cells.

Glioblastoma (GB) is a lethal brain tumor that rapidly adapts to the dynamic changes of the tumor microenvironment (TME). Mesenchymal stem/stromal cells (MSCs) are one of the stromal components of the TME playing multiple roles in tumor progression. GB progression is prompted by the immunosuppressive microenvironment characterized by high concentrations of the nucleoside adenosine (ADO). ADO acts as a signaling molecule through adenosine receptors (ARs) but also as a genetic and metabolic regulator. Herein, the effects of high extracellular ADO concentrations were investigated in a human glioblastoma cellular model (U343MG) and MSCs. The modulation of the purinome machinery, i.e., the ADO production (CD39, CD73, and adenosine kinase [ADK]), transport (equilibrative nucleoside transporters 1 (ENT1) and 2 (ENT2)), and degradation (adenosine deaminase [ADA]) were investigated in both cell lines to evaluate if ADO could affect its cell management in a positive or negative feed-back loop. Results evidenced a different behavior of GB and MSC cells upon exposure to high extracellular ADO levels: U343MG were less sensitive to the ADO concentration and only a slight increase in ADK and ENT1 was evidenced. Conversely, in MSCs, the high extracellular ADO levels reduced the ADK, ENT1, and ENT2 expression, which further sustained the increase of extracellular ADO. Of note, MSCs primed with the GB-conditioned medium or co-cultured with U343MG cells were not affected by the increase of extracellular ADO. These results evidenced how long exposure to ADO could produce different effects on cancer cells with respect to MSCs, revealing a negative feedback loop that can support the GB immunosuppressive microenvironment. These results improve the knowledge of the ADO role in the maintenance of TME, which should be considered in the development of therapeutic strategies targeting adenosine pathways as well as cell-based strategies using MSCs.

CD39 activities in the treated acupoints contributed to the analgesic mechanism of acupuncture on arthritis rats.

Our previous work had identified that at the acupuncture point (acupoint), acupuncture-induced ATP release was a pivotal event in the initiation of analgesia. We aimed to further elucidate the degradation of ATP by CD39. Acupuncture was administered at Zusanli acupoint on arthritis rats, and pain thresholds of the hindpaws were determined. Pharmacological tools or adeno-associated viruses were administered at the acupoints to interfere with targeting signals. Protein expression was determined with qRT-PCR, WB, or immunofluorescent labeling. Cultured keratinocytes, HaCaT line, were subjected to hypotonic shock to simulate needling stimulation. Extracellular ATP and adenosine levels were quantified using luciferase-luciferin assay and ELISA, respectively. Acupuncture-induced prompt analgesia was impaired by inhibiting CD39 activities to prevent the degradation of ATP to AMP but was mimicked by using CD39 agonists. Acupuncture-induced ATP accumulation exhibited synchronous changes. Similarly, acupuncture analgesia was hindered by suppressing CD73 to prevent the conversion of AMP to adenosine. Furthermore, the acupuncture effect was replicated by agonism at P2Y2Rs but inhibited by antagonism at them. Acupuncture upregulated CD73 and P2Y2Rs but not CD39. Immunofluorescent labeling demonstrated that keratinocytes were a primary site for these proteins. Shallow acupuncture also demonstrated antinociception. In vitro tests showed that hypotonic shock induced HaCaT cells to release ATP and adenosine, which was impaired by suppressing CD39 and CD73, respectively. Finally, agonism at P2Y2Rs promoted ATP release and [Ca2+]i rise. CD39 at the acupoints contributes to the analgesic mechanism of acupuncture. It may facilitate adenosine signaling in conjunction with CD73 or provide an appropriate ATP milieu for P2Y2Rs. Skin tissue may be one of the scenes for these signalings.

Exploring the mechanism of action of huoermai essential oil for plateau insomnia based on the camp/CREB/BDNF/gabaergic pathway

Ethnopharmacological relevanceThe traditional Huoermai therapy is a treatment for insomnia used by the Tibetan people living on the Tibetan plateau in China. This therapy involves the use of Myristica fragrans Houtt. and Carum carvi L., along with fomentation and massage, and has shown significant clinical effects. However, the mechanism of how Huoermai therapy treats plateau insomnia needs further clarification. Aim of the studyThis study aimed to investigate the mechanism of action of Huoermai essential oil (HEO) in treating plateau insomnia, focusing on the cAMP/CREB/BDNF/GABAergic pathway. MethodsThe major components of Huoermai essential oil were identified by Gas chromatography-mass spectrometry (GC-MS) for subsequent network pharmacology analysis. Proteomics techniques were employed to pinpoint disparities in brain tissue protein expression in a mouse model of plateau insomnia following Huoermai therapy administration, in conjunction with network pharmacology to forecast pathways related to hypoxia and insomnia. Plateau insomnia mouse model was established and the therapeutic impact of Huoermai essential oil was evaluated. Hematoxylin & Eosin staining(HE) was conducted to observe pathological damage to the cortex, hippocampus, thalamus and hypothalamus structures. Changes in serotonin (5-HT), melatonin (MT), adenosine (AD), cyclic adenosine monophosphate (cAMP) and malondialdehyde (MDA) levels in mouse brain tissue were gauged through enzyme-linked immunosorbent assay (ELISA) to assess sleep status and oxidative stress levels in mice. Molecular docking was employed to anticipate the target binding energy of Huoermai essential oil constituents. ELISA and Western Blot (WB) were used to ascertain the expression of cAMP/CREB/BDNF/GABAergic pathway. ResultsThe results indicated that HEO positively impacted intermittent hypobaric hypoxia-induced plateau insomnia in mice. Histological examination results showed that HEO ameliorated neuronal damage in specific regions of the brain affected by plateau insomnia, such as the cortex, hippocampus, thalamus, and hypothalamus. Through GC-MS analysis, 56 volatile oil components were identified. Subsequently, a combined network pharmacology and proteomics analyses led to selecting the cAMP/CREB/BDNF/GABAergic pathway for further study. ELISA experiments demonstrated that HEO treatment increased GABA and MT levels while significantly reducing 5-HT and adenosine levels in brain tissue of mice with plateau insomnia. WB results revealed that HEO ameliorated plateau insomnia by suppressing the hyperactivation of the cAMP pathway, increasing brain-derived neurotrophic factor (BDNF) levels and B-cell lymphoma-2 (BCL-2) expression, and alleviating hypoxia-induced oxidative stress. Moreover, molecular docking results showed strong binding affinity of all pharmacological components to their targets and proteins in the brain. ConclusionThese results indicate that HEO significantly prolongs sleep duration in plateau insomniac mice and treats plateau insomnia by modulating levels of sleep-related regulators, modulating the cAMP pathway, increasing GABA receptor expression, and improving neuronal survival and anti-apoptosis.

Chronic corticosterone exposure causes anxiety- and depression-related behaviors with altered gut microbial and brain metabolomic profiles in adult male C57BL/6J mice

Chronic exposure to glucocorticoids in response to long-term stress is thought to be a risk factor for major depression. Depression is associated with disturbances in the gut microbiota composition and peripheral and central energy metabolism. However, the relationship between chronic glucocorticoid exposure, the gut microbiota, and brain metabolism remains largely unknown. In this study, we first investigated the effects of chronic corticosterone exposure on various domains of behavior in adult male C57BL/6J mice treated with the glucocorticoid corticosterone to evaluate them as an animal model of depression. We then examined the gut microbial composition and brain and plasma metabolome in corticosterone-treated mice. Chronic corticosterone treatment resulted in reduced locomotor activity, increased anxiety-like and depression-related behaviors, decreased rotarod latency, reduced acoustic startle response, decreased social behavior, working memory deficits, impaired contextual fear memory, and enhanced cued fear memory. Chronic corticosterone treatment also altered the composition of gut microbiota, which has been reported to be associated with depression, such as increased abundance of Bifidobacterium, Turicibacter, and Corynebacterium and decreased abundance of Barnesiella. Metabolomic data revealed that long-term exposure to corticosterone led to a decrease in brain neurotransmitter metabolites, such as serotonin, 5-hydroxyindoleacetic acid, acetylcholine, and gamma-aminobutyric acid, as well as changes in betaine and methionine metabolism, as indicated by decreased levels of adenosine, dimethylglycine, choline, and methionine in the brain. These results indicate that mice treated with corticosterone have good face and construct validity as an animal model for studying anxiety and depression with altered gut microbial composition and brain metabolism, offering new insights into the neurobiological basis of depression arising from gut-brain axis dysfunction caused by prolonged exposure to excessive glucocorticoids.

Adenosine regulates depressive behavior in mice with chronic social defeat stress through gut microbiota

Major depressive disorder (MDD) is recognized as the most prevalent affective disorder worldwide. Metagenomic studies increasingly support a critical role for dysbiosis of gut microbiota in the development of depression. Previous studies have demonstrated that adenosine alleviates gut dysbiosis, suggesting that elevating adenosine levels could be a novel intervention for MDD; however, the mechanisms underlying this effect remain unclear. This study utilized 16S rRNA gene sequencing, fecal microbiota transplantation (FMT) and ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to test the hypothesis that increased adenosine alleviates depressive behaviors in male mice subjected to chronic social defeat stress (CSDS) through alterations to gut microbiota. The data showed that depression-susceptible (SUS) mice exhibited gut dysbiosis, and FMT from SUS mice increased depression-like behaviors in healthy recipients. In SUS mice, adenosine supplementation ameliorated both depression-like behaviors and abnormalities in gut microbiota, and co-administration of probiotics and adenosine not only mitigated depression-like behaviors but also enhanced gut barrier integrity. By including 83 depressed adolescents and 67 healthy controls, this study found that the level of short-chain fatty acids (SCFAs) in the depression group was reduced, this finding parallels reductions seen in SUS mice and in recipient mice after FMT from SUS donors. Conversely, supplementation with either adenosine or probiotics led increased SCFAs concentrations in the serum of SUS mice. These findings suggest that adenosine may alleviate depression-like behaviors in CSDS mice by modulating the gut microbiota. This effect is likely associated with increased serum SCFAs, metabolites produced by the gut microbiota, following adenosine supplementation.This article is part of the Special Issue on "Personality Disorders".

Adenosine-Dependent Arousal Induced by Astrocytes in a Brainstem Circuit.

Astrocytes play a crucial role in regulating sleep-wake behavior. However, how astrocytes govern a specific sleep-arousal circuit remains unknown. Here, the authors show that parafacial zone (PZ) astrocytes responded to sleep-wake cycles with state-differential Ca2+ activity, peaking during transitions from sleep to wakefulness. Using chemogenetic and optogenetic approaches, they find that activating PZ astrocytes elicited and sustained wakefulness by prolonging arousal episodes while impeding transitions from wakefulness to non-rapid eye movement (NREM) sleep. Activation of PZ astrocytes specially induced the elevation of extracellular adenosine through the ATP hydrolysis pathway but not equilibrative nucleoside transporter (ENT) mediated transportation. Strikingly, the rise in adenosine levels induced arousal by activating A1 receptors, suggesting a distinct role for adenosine in the PZ beyond its conventional sleep homeostasis modulation observed in the basal forebrain (BF) and cortex. Moreover, at the circuit level, PZ astrocyte activation induced arousal by suppressing the GABA release from the PZGABA neurons, which promote NREM sleep and project to the parabrachial nucleus (PB). Thus, their study unveils a distinctive arousal-promoting effect of astrocytes within the PZ through extracellular adenosine and elucidates the underlying mechanism at the neural circuit level.

Impact of Vitamin D deficiency on immunological and metabolic responses in women with recurrent pregnancy loss: focus on VDBP/HLA-G1/CTLA-4/ENTPD1/adenosine-fetal-maternal conflict crosstalk

Background and aimRecurrent pregnancy loss (RPL), also known as recurrent implantation failure (RIF), is a distressing condition affecting women characterized by two or more consecutive miscarriages or the inability to carry a pregnancy beyond 20 weeks. Immunological factors and genetic variations, particularly in Vit D Binding Protein (VDBP), have gained attention as potential contributors to RPL. This study aimed to provide insight into the immunological, genetic, and metabolic networks underlying RPL, placing a particular emphasis on the interactions between VDBP, HLA-G1, CTLA-4, ENTPD1, and adenosine-fetal-maternal conflict crosstalk.MethodsA retrospective study included 198 women with three or more consecutive spontaneous abortions. Exclusion criteria comprised uterine abnormalities, endocrine disorders, parental chromosomal abnormalities, infectious factors, autoimmune diseases, or connective tissue diseases. Immunological interplay was investigated in 162 female participants, divided into two groups based on their Vit D levels: normal Vit D-RPL and low Vit D-RPL. Various laboratory techniques were employed, including LC/MS/MS for Vit D measurement, ELISA for protein detection, flow cytometry for immune function analysis, and molecular docking for protein–ligand interaction assessment.ResultsGeneral characteristics between groups were significant regarding Vit D and glucose levels. Low Vit D levels were associated with decreased NK cell activity and downregulation of HLA-G1 and HLA-G5 proteins, while CTLA-4 revealed upregulation. VDBP was significantly downregulated in the low Vit D group. Our findings highlight the intricate relationship between Vit D status and adenosine metabolism by the downregulation of SGLT1, and NT5E, key components of adenosine metabolism, suggests that Vit D deficiency may disrupt the regulation of adenosine levels, leading to an impaired reproductive outcome. HNF1β, a negative regulator of VDBP, was upregulated, while HNF1α, a positive regulator, was downregulated in low Vit D women after RPL. Molecular docking analysis revealed crucial residues involved in the interaction between Vit D and HNF1β.ConclusionCollectively, these findings underscore the importance of Vit D in modulating immune function and molecular pathways relevant to pregnancy maintenance, highlighting the need for further research to elucidate the mechanisms and potential therapeutic interventions for improving pregnancy outcomes in individuals with Vit D deficiency and RPL.

Dynamic changes in serum adenosine and the adenosine metabolism-based signature for prognosis in HER2-positive metastatic breast cancer patients

AimsAdenosine metabolism in the breast cancer microenvironment is critical for tumor immunity. However, the prognostic significance of adenosine in breast cancer remains unclear. We aimed to dynamically monitor serum adenosine levels in patients with HER2-positive metastatic breast cancer (MBC) patients and to explore its predictive significance in trastuzumab therapy. MethodsThe sequencing and clinical data were downloaded from TCGA and GSE176078. Adenosine-related differentially expressed genes was analyzed by “DESeq2” package. Multivariate Cox and lasso-penalized Cox regressions were used to construct prognostic risk signatures. The risk scores were calculated from the identified expression of the hub genes. Bioinformatic analyses were performed using R with related packages. We also enrolled the metastatic breast cancer patients with HER2-positive from in our center and classified them into different groups according to the clinical outcomes assessed by enhanced CT. The adenosine levels were dynamically detected, and the difference in immune microenvironment between the subgroups was assessed by the immune cells that were recorded in our center. ResultsA total of 109 breast cancer patients with HER2-positive MBC were enrolled, and the expressions of 22 adenosine-related genes were filtered and matched from the TCGA database. The survival model based on the 15 differentially expressed genes was established, and the risk scores of each patient were the prognostic risk factors. Single-cell transcriptome sequencing data identified transcriptomic differences in patients with HER2-positive breast cancer. We also confirmed the predictive value of serum adenosine in the clinical progression of HER2-positive MBC patients. The different immune microenvironment between the subgroups supported the reliability of the predictive ability of adenosine in HER2-positive MBC patients. ConclusionsThe dynamic change of adenosine is a predictive biomarker for monitoring disease progression. The adenosine metabolism-based signature has the potential application in the prognosis of HER2-positive MBC patients.

Spatially restricted ecto-5'-nucleotidase expression promotes the growth of uterine leiomyomas by modulating Akt activity.

Found in as many as 80% of women, uterine leiomyomas are a frequent cause of abnormal uterine bleeding, pelvic pain, and infertility. Despite their significant clinical impact, the mechanisms responsible for driving leiomyoma growth remain poorly understood. After obtaining IRB permission, expression of ecto-5'-nucleotidase (NT5E, CD73) was assessed in matched specimens of myometrium and leiomyoma by real-time qPCR, Western blot, and immunohistochemistry (IHC). Adenosine concentrations were measured by enzyme-linked assay. Primary cultures were used to assess the impact of adenosine and/or adenosine receptor agonists on proliferation, apoptosis, and patterns of intracellular signaling invitro. When compared to matched specimens of healthy myometrium, uterine leiomyomas were characterized by reduced CD73 expression. Largely limited to thin-walled vascular structures and the pseudocapsule of leiomyomas despite diffuse myometrial distribution. Restricted intra-tumoral CD73 expression was accompanied by decreased levels of intra-tumoral adenosine. Invitro, incubation of primary leiomyoma cultures with adenosine or its hydrolysis-resistant analog 2-chloro-adenosine (2-CL-AD) inhibited proliferation, induced apoptosis, and reduced proportion of myocytes in S- and G2-M phases of the cell cycle. Decreased proliferation was accompanied by reduced expression of phospho-Akt, phospho-Cdk2-Tyr15, and phospho-Histone H3. Enforced expression of the A2B adenosine receptor (ADORA2B) and ADORA2B-selective agonists similarly suppressed proliferation and inhibited Akt phosphorylation. Collectively, these observations broadly implicate CD73 and reducedextracellular concentrations ofadenosine as key regulators of leiomyoma growth and potentiallyidentify novel strategies for clinically managing these common tumors.

A1 adenosine receptor evoked Ca2+ signals in astrocytes

Abstract In addition to neurons, glial cells make up the nervous system. These glial cells are divided into macroglial cells - including astrocytes - and microglial cells. Astrocytes have several important functions in the central nervous system, such as synthesizing transmitters and releasing them, thereby activating the surrounding neurons and astrocytes. Astrocytes do not generate action potentials, but they can transmit information to neighboring cells and react to external stimuli by employing Ca2+ dynamics. Increased neuronal activity can lead to an increased extracellular level of adenosine, reflected by a change in the metabolic state. Adenosine is a naturally occurring nucleoside distributed throughout the body as a metabolic intermediary. Its action is mediated by binding to adenosine receptors, such as the G-protein coupled receptors (GPCRs) A1 adenosine receptor. The binding of adenosine to the specific receptor type activates different signaling pathways in astrocytes. In our work, we studied Ca2+ signals generated upon adenosine A1 receptor activation by adding the pathway to the computational astrocyte model by Oschmann et.al (2017). We investigated the influence of the high-affinity A1 adenosine receptor in various astrocyte domains on the regulated Ca2+ release from the endoplasmic reticulum (ER) of astrocytes and concluded that the receptor-dependent pathway contributes to Ca2+ signals mainly in the soma.

Design and construction of an artificial labor-division consortium for phenanthrene degradation with three-functional modules

Polycyclic Aromatic Hydrocarbons (PAHs) are highly toxic organic pollutants. Phenanthrene often serves as a model compound for studying PAHs biodegradation. In this work, we firstly engineered Escherichia coli M01 containing seven phenanthrene degradation genes and combined it with existing engineered strains E. coli M2 and M3 to form an artificial three-bacteria consortium, named M0123, which exhibited a degradation ratio of 64.66% for 100 mg/L of phenanthrene over 8 days. Subsequently, we constructed engineered Pseudomonas putida KTRL02 which could produce 928.49 mg/L rhamnolipids and integrated it with M0123, forming a four-bacteria consortium with an impressive 81.62% phenanthrene degradation ratio. Assessment of extracellular adenosine levels during the degradation process indicated high cellular energy demand in the four-bacteria consortium. Then, we introduced Bacillus subtilis RH33, a riboflavin-producing strain, as an energy-supplying bacterium, to create a five-bacteria consortium, which exhibited an 88.19% degradation ratio for phenanthrene. The NADH/NAD+ ratio in the five-bacteria consortium during the degradation process was monitored, which was consistently higher than that of the four-bacteria consortium over the eight-day period, indicating a higher overall intracellular reduction capacity. Furthermore, the five-bacteria consortium displayed good tolerance to phenanthrene, even achieving a degradation ratio of 79.38% for 500 mg/L of phenanthrene. This study demonstrates that designing and constructing artificial consortia from the functional perspective and various angles can effectively enhance the degradation of phenanthrene after the addition of the energy-supplying bacterium. This study demonstrates that designing and constructing artificial labor-division consortia from the functional perspective and various angles can effectively enhance the degradation of phenanthrene.