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Photonics 10(1), 24 (2023). doi:10.3390/photonics10010024, Published by MDPI, Basel

Cefquinome (CEQ) is a novel β-lactam antibiotic that exhibits excellent antibacterial activity against Staphylococcus aureus. However, the bacterial protein targets of CEQ are unclear. To evaluate the relationship between the pharmacokinetic/pharmacodynamic (PK/PD) parameters of CEQ and strains with varying degrees of resistance and to elucidate bacterial protein responses to CEQ treatment, label-free quantitative proteomics analysis was conducted. The sensitive S. aureus ATCC6538 and the resistant 2MIC and 8MIC were tested for differentially expressed proteins. An in vitro model was treated with different concentrations of CEQ (3, 5, or 10 µg/ml) with different terminal half-lives (2.5 or 5 h) at different intervals (12 or 24 h). Differentially expressed proteins were evaluated using Gene Ontology analysis followed by KEGG pathway enrichment analysis and STRING network analysis. RT-qPCR was performed to validate the differentially expressed proteins at the molecular level. The results showed that the degree of resistance increased in a cumulative manner and increased gradually with the extension of administration time. The resistant strain would not have appeared in the model only if %T > mutant prevention concentration ≥ 50%. The expression of 45 proteins significantly changed following CEQ treatment, among which 42 proteins were obviously upregulated and 3 were downregulated. GO analysis revealed that the differentially expressed proteins were mainly present on cells and the cell membrane, participated in metabolic and intracellular processes, and had catalytic and binding activities. The RPSO, SDHB, CITZ, ADK, and SAOUHSC 00113 genes in S. aureus may play important roles in the development of resistance to CEQ. These results provided important reference candidate proteins as targets for overcoming S. aureus resistance to CEQ.

Adenosine kinase (ADK) deficiency is a very rare inborn error of methionine and adenosine metabolism. It is characterized by developmental delay, hypotonia, epilepsy, facial dysmorphism, failure to thrive, transient liver dysfunction with cholestasis, recurrent hypoglycemia, and cardiac defects. Only 26 cases (16 families) of ADK deficiency have been published since its identification in 2011. Vascular abnormalities in cervical arteries and cerebral stroke have never been reported in this condition. Here, we describe two patients with ADK deficiency and vascular tortuosity leading to stroke in one of them. ADK deficiency is a rare inborn error of methionine metabolism with a complex phenotype that might be associated with cerebrovascular abnormalities and stroke.

The binding of adenosine 5’-triphosphate (ATP) and adenosine 5’-monophosphate (AMP) to adenylate kinase (AdK) drives closure of lids over the substrate adenosyl groups. We test the hypothesis that this conformational change activates AdK for catalysis. The rate constants for Homo sapiens adenylate kinase 1 (HsAdK1)-catalyzed phosphoryl group transfer to AMP, k(cat)/K(m) = 7.0 x 10(6) M(−1) s(−1), and phosphite dianion, (k(HPi))(obs) ≤ 1 x 10(−4) M(−1) s(−1), show that the binding energy of the adenosyl group effects a ≥7.0 x 10(10)-fold rate acceleration of phosphoryl transfer from ATP. The third-order rate constant of k(cat)/K(HPi)K(EA) = 260 M(−2) s(−1) for 1-(β-d-erythrofuranosyl)adenine (EA)-activated phosphoryl transfer to phosphite dianion was determined, and the isohypophosphate reaction product characterized by (31)P NMR. The results demonstrate: (i) a ≥14.7 kcal/mol stabilization of the transition state for phosphoryl transfer by the adenosyl group of AMP and a ≥2.6 x 10(6)-fold rate acceleration from the EA-driven conformational change, and (ii) the recovery of ≥8.7 kcal/mol of this transition state stabilization for EA-activated phosphoryl transfer from ATP to phosphite.

Settlement and metamorphosis of planktonic larvae into benthic adults are critical components of a diverse range of marine invertebrate-mediated processes such as the formation of mussel beds and coral reefs, the recruitment of marine shellfisheries, and the initiation of macrobiofouling. Although larval settlement and metamorphosis induced by natural chemical cues is widespread among marine invertebrates, the mechanisms of action remain poorly understood. Here, we identified that the molecular target of adenosine (an inducer of larval settlement and metamorphosis from conspecific adults in the invasive biofouling mussel Mytilopsis sallei) is adenosine kinase (ADK). The results of transcriptomic analyses, pharmacological assays, temporal and spatial gene expression analyses, and siRNA interference, suggest that ATP-dependent phosphorylation of adenosine catalyzed by ADK activates the downstream AMPK-FoxO signaling pathway, inducing larval settlement and metamorphosis in M. sallei. This study not only reveals the role of the ADK-AMPK-FoxO pathway in larval settlement and metamorphosis of marine invertebrates but it also deepens our understanding of the functions and evolution of adenosine signaling, a process that is widespread in biology and important in medicine.

Adenosine kinase (ADK) deficiency is a rare autosomal recessive inborn error of metabolism involving the methionine and purine metabolic pathways. Prior reports show that most patients present in infancy with jaundice, hypotonia, developmental delay, and mild dysmorphic features. Characteristic biochemical findings included hypoglycemic hyperinsulinism, cholestasis, elevated liver functions, methionine, S‐adenosylhomocysteine, and S‐adenosylmethionine, with normal or mildly elevated homocysteine level. Brain imaging demonstrated atrophy, hydrocephalus, and delayed myelination. There are 26 reported patients of ADK deficiency, of which 14 patients were placed on a methionine‐restricted diet. Clinical improvement with methionine restriction was not well described. Case report We report an infant who presented at birth with persistently elevated ammonia (100‐163 μmol/L), hypoglycemia, cholestasis, and liver dysfunction. The initial metabolic and genetic work‐up was nondiagnostic, with only a mildly increased plasma methionine level (51 [

Large-scale conformational transitions in multi-domain proteins are often essential for their functions. To investigate the transitions, it is necessary to explore multiple potential pathways, which involve different intermediate structures. Here, we present a multi-basin (MB) coarse-grained (CG) structure-based Go̅ model for describing transitions in proteins with more than two moving domains. This model is an extension of our dual-basin Go̅ model in which system-dependent parameters are determined systematically using the multistate Bennett acceptance ratio method. In the MB Go̅ model for multi-domain proteins, we assume that intermediate structures may have partial inter-domain native contacts. This approach allows us to search multiple transition pathways that involve distinct intermediate structures using the CG molecular dynamics (MD) simulations. We apply this scheme to an enzyme, adenylate kinase (AdK), which has three major domains and can move along two different pathways. Using the optimized mixing parameters for each pathway, AdK shows frequent transitions between the Open, Closed, and the intermediate basins and samples a wide variety of conformations within each basin. The explored multiple transition pathways could be compared with experimental data and examined in more detail by atomistic MD simulations.

For any positive integer [Formula: see text], let [Formula: see text] be the number of solutions of the equation [Formula: see text] with integers [Formula: see text], where [Formula: see text] is the integral part of real number [Formula: see text]. Recently, Luca and Ralaivaosaona gave an asymptotic formula for [Formula: see text]. In this paper, we give an asymptotic development of [Formula: see text] for all [Formula: see text]. Moreover, we prove that the number of such partitions is even (respectively, odd) infinitely often.

The photostabilies of hexanitrostillbene (HNS) were studied at 1% wt. of photostabilizers. The photostabilizers of Chiguard BP-12 (UV- 531), ADK STAB LA-326 (UV- 326), ADK STAB LA-32 (UV-P), Irganox 1010 and Everstab LS944 were used in this study. A photoreactor including a 30 W monochromatic UV lamp was used for investigation of photodegradations of solid HNS. The changes in surface area of HNS peak in HPLC chromatogram at time intervals of 2, 4, 6 and 8 h were used for calculation of kinetic rate constants of photodegradation process. The compound of UV-P photostabilizer was found to have the greatest effect on enhancing photostability for HNS among the investigated photostabilizers. The kinetic rate constants were obtained 6.2 × 10−2 and 4.1 × 10−2 h−1, respectively, for photodegradation process of HNS in absence and in presence of UV-P.

Objective. To evaluate the effect of CLMD administration on epileptic seizures and brain injury in pentylenetetrazole- (PZT-) kindled mice. Methods. The effect of pretreatment with CLMD (5, 10, and 20 ml/kg (mg/kg) by gavage) for seven days on PTZ-induced kindling, duration and grade of kindling-induced seizures, and pathological injury in the cortex and hippocampus was evaluated. Male BALB/c mice with adenosine A1 receptor knockout were subjected to intraperitoneal injection of PTZ (35 mg/kg) once every day until kindling was successfully induced. Quantitative reverse transcription polymerase chain reaction, immunofluorescence, and western blot were performed to assess the mRNA and protein levels of p-glycoprotein (PGP), multidrug resistance-associated protein 1 (MRP1), cyclooxygenase-2 (COX-2), prostaglandin E2 (PGE2), and adenylate kinase (ADK) in the cortex and hippocampus. Results. PTZ successfully induced kindling in mice after 21 days, wherein CLMD showed an obvious dose-dependent antiepileptic effect. High-dose CLMD significantly increased the latency of epileptic seizures, decreased the sustained time of epileptic seizures and the seizure grade, and ameliorated the histopathological changes in the cortex and hippocampus. Furthermore, PTZ kindling induced significantly higher levels of PGP, MRP1, COX-2, PGE2, and ADK, but this effect was inhibited by pretreatment with CLMD in a dose-dependent manner. Conclusion. Pretreatment with CLMD attenuates PTZ-kindled convulsions and brain injury in mice. The mechanism may be related to the cyclooxygenase-2/prostaglandin E2/multidrug transporter pathway.

Endometrial adenocarcinoma (ADK) is one of the most common uterine cancer and the fourth neoplasia mortality cause in women according to the literature data. ADK is encountered in the sixth decade of life, the mean age being 63 years. Only 2-5% of cases are found in women less than 40 years old. We present the case of a 63-year-old woman admitted in our Clinic for hypogastric pain and reduced vaginal sanguinolent discharge. The genital examination and transvaginal ultrasound (US) scan were strongly suggestive for a type 0 myoma tending to be expelled through the cervical canal. Laparotomy and total hysterectomy with bilateral adnexectomy were performed. Histological examination identified a grade G1 well-differentiated endometrioid ADK without uterine wall invasion. The immunohistochemistry study for estrogen receptors expression revealed moderate and strong nuclear immunostaining in more than 70% of the tumoral cells, as well as a mosaic p16 immunoexpression, a cytokeratin 7 (CK7) immunophenotype, no p53 overexpression and low Ki67 index (estimated at 10-15%). Considering the incidental finding, the early stage and limited localization of the ADK at the myoma surface level surgery was considered sufficient as treatment but follow-up was conducted by magnetic resonance imaging (MRI) and general examination every six months. The most interesting aspect of the case was the atypical presence of an active myoma in menopause and the totally unsuspicious US appearance of the endometrium. The endometrial ADK was accidentally discovered in an unusual specimen without any other symptom.

Ubiquitination is involved in the regulation of numerous cellular functions. Research works in the ubiquitin realm rely heavily on ubiquitination assays in vitro and require large amounts of ubiquitin-activating enzyme (UBA1) and keep ATP supplies. However, UBA1 is hard to be obtained with large quantities using reported methods. We fused Escherichia coli adenylate kinase (adk) and mouse UBA1 and obtained fusion protein adk-mUBA1. The expression level of adk-mUBA1 increased about 8-fold compared with mUBA1 in an E. coli expression system, and adk-mUBA1 was easily purified to 90% purity via 2 purification steps. The purified adk-mUBA1 protein was functional for ubiquitination and could use ATP in addition to ADP as energy supply and had a higher catalytic activity than mUBA1 in cell lysis. adk-mUBA1 can be applied to preparing ubiquitin-modified substrates and kinds of ubiquitin chains in a chemical synthesis process and is a preferable application than mUBA1 in vitro ubiquitination.

[Image: see text] DNA methylation has a major role in cancer, and its inhibitors are used therapeutically. DNA methylation depends on methyl group flux through the transmethylation pathway, which forms adenosine. We hypothesized that an adenosine kinase isoform with nuclear expression (ADK-L) determines global DNA methylation in cancer cells. We quantified ADK-L expression (Western Blot) and global DNA methylation as percent 5-methyldeoxycytidine (5mdC, LC-MS/MS) in three cancer lines (HeLa, HepG2, and U373). ADK-L expression and global DNA methylation correlated positively with the highest levels in HeLa cells compared to U373 and HepG2 cells. To determine whether ADK increases global DNA methylation and to validate its potential therapeutics, we treated HeLa cells with potent ADK inhibitors MRS4203 and MRS4380 (IC(50) 88 and 140 nM, respectively). Both nucleosides, but not a structurally related poor ADK inhibitor, significantly reduced global DNA methylation in HeLa cells in a concentration-dependent manner. Thus, ADK-L is a potential target for the therapeutic manipulation of DNA methylation levels in cancer.

Increased adenosine helps limit infarct size in ischaemia/reperfusion‐injured hearts. In cardiomyocytes, 90% of adenosine is catalysed by adenosine kinase (ADK) and ADK inhibition leads to higher concentrations of both intracellular adenosine and extracellular adenosine. However, the role of ADK inhibition in myocardial ischaemia/reperfusion (I/R) injury remains less obvious. We explored the role of ADK inhibition in myocardial I/R injury using mouse left anterior ligation model. To inhibit ADK, the inhibitor ABT‐702 was intraperitoneally injected or AAV9 (adeno‐associated virus)—ADK—shRNA was introduced via tail vein injection. H9c2 cells were exposed to hypoxia/reoxygenation (H/R) to elucidate the underlying mechanisms. ADK was transiently increased after myocardial I/R injury. Pharmacological or genetic ADK inhibition reduced infarct size, improved cardiac function and prevented cell apoptosis and necroptosis in I/R‐injured mouse hearts. In vitro, ADK inhibition also prevented cell apoptosis and cell necroptosis in H/R‐treated H9c2 cells. Cleaved caspase‐9, cleaved caspase‐8, cleaved caspase‐3, MLKL and the phosphorylation of MLKL and CaMKII were decreased by ADK inhibition in reperfusion‐injured cardiomyocytes. X‐linked inhibitor of apoptosis protein (XIAP), which is phosphorylated and stabilized via the adenosine receptors A2B and A1/Akt pathways, should play a central role in the effects of ADK inhibition on cell apoptosis and necroptosis. These data suggest that ADK plays an important role in myocardial I/R injury by regulating cell apoptosis and necroptosis.

Epilepsy disease is characterized by the neuronal dysfunction or abnormal neuronal activity of the brain which is regulated by astrocytes. These are glial cells and found to be the major regulators of the brain which are guided by the occurrence of adenosine kinase (ADK) enzyme in the central nervous system (CNS). During the normal physiological environment, ADK maintains the level of adenosine in the CNS. Dysfunction of ADK levels results in accumulation of adenosine levels in the CNS that leads to the pathophysiology of the brain such as astrogliosis which is a pathological hallmark of epileptic seizures. Vicine, an alkaloid glycoside in bitter gourd juice (Momordica charantia) is found to be toxic to the human system if the bitter gourd juice is consumed more. This compound inhibits ADK enzyme activity to lead epilepsy and seizure. Here, the toxic effect of vicine targeting ADK using computational predictions was investigated. The 3-dimensional structure of ADK has been constructed using I-Tasser, which has been refined by ModRefiner, GalaxyRefine, and 3D refine and it was endorsed using PROCHECK, ERRAT, and VADAR. 3D structure of the ligand molecule has been obtained from PubChem. Molecular docking has been achieved using AutoDock 4.2 software, from which the outcome showed the effective interaction between vicine and ADK, which attains binding free energy (∆G) of - 4.13 kcal/mol. Vicine molecule interacts with the active region ARG 149 of ADK and inhibits the functions of ADK that may cause imbalance in energy homeostasis. Also, pre-ADMET results robustly propose in which vicine possesses toxicity, and meanwhile, from the Ames test, it was shown as mutagenic. Hence, the results from our study suggest that vicine was shown to be toxic that suppresses the ADK activity to undergo pathological conditions in the neuronal junctions to lead epilepsy.

Adenosine kinase (ADK) deficiency is characterized by liver disease, dysmorphic features, epilepsy and developmental delay. This defect disrupts the adenosine/AMP futile cycle and interferes with the upstream methionine cycle. We report the clinical, histological and biochemical courses of three ADK children carrying two new mutations and presenting with neonatal cholestasis and neurological disorders. One of them died of liver failure whereas the other two recovered from their liver damage. As the phenotype was consistent with a mitochondrial disorder, we studied liver mitochondrial respiratory chain activities in two patients and revealed a combined defect of several complexes. In addition, we retrospectively analyzed methionine plasma concentration, a hallmark of ADK deficiency, in a cohort of children and showed that methionine level in patients with ADK deficiency was strongly increased compared with patients with other liver diseases. ADK deficiency is a cause of neonatal or early infantile liver disease that may mimic primary mitochondrial disorders. In this context, an elevation of methionine plasma levels over twice the upper limit should not be considered as a nonspecific finding. ADK deficiency induced-liver dysfunction is most often transient, but could be life-threatening.

WOS:000789651000009; HABs involving Alexandrium pacificum have been reported in metal-contaminated ecosystems, suggesting that this distributed species adapts to and/or can tolerate the effects of metals. Modifications in soluble proteomes and PST contents were characterized in two Mediterranean A. pacificum strains exposed to mono- or polymetallic stresses (zinc, lead, copper, cadmium). These strains were isolated from two anthropized locations: Santa Giusta Lagoon (Italy, SG C10-3) and the Tarragona seaport (Spain, TAR C5-4F). In both strains, metals primarily downregulated key photosynthesis proteins. Metals also upregulated other proteins involved in photosynthesis (PCP in both strains), the oxidative stress response (HSP 60, proteasome and SOD in SG C10-3; HSP 70 in TAR C5-4F), energy metabolism (AdK in TAR C5-4F), neoglucogenesis/glycolysis (GAPDH and PEP synthase in SG C10-3) and protein modification (PP in TAR C5-4F). These proteins, possibly involved in adaptive proteomic responses, may explain the development of these A. pacificum strains in metal-contaminated ecosystems. The two strains showed different proteomic responses to metals, with SG C10-3 upregulating more proteins, particularly PCP. Among the PSTs, regardless of the metal and the strain studied, C2 and GTX4 predominated, followed by GTX5. Under the polymetallic cocktail, (i) total PSTs, C2 and GTX4 reached the highest levels in SG C10-3 only, and (ii) total PSTs, C2, GTX5 and neoSTX were higher in SG C10-3 than in TAR C5-4F, whereas in SG C10-3 under copper stress, total PSTs, GTX5, GTX1 and C1 were higher than in the controls, revealing variability in PST biosynthesis between the two strains. Total PSTs, C2, GTX4 and GTX1 showed significant positive correlations with PCP, indicating that PST production may be positively related to photosynthesis. Our results showed that the A. pacificum strains adapt their proteomic and physiological responses to metals, which may contribute to their ecological success in highly anthropized areas.

The objective of the article was to determine the variability of meningococcal house-keeping gene alleles circulating in Belarus. House-keeping genes sequencing was made by Sanger (ABI 3500). The phylogenetic analysis was done in MEGA X. SNPs were analyzed at pubMLST.org. 60 Belarusian meningococci, collected during 8 years, contain 17 alleles of abcZ gene (5.9 % first identified in Belarus – abcZ 1016) encoding 5 variants of the ABC transporter; 16 adk gene alleles – 2 variants of adenylate cyclase; 17 alleles of aroE gene (11.8 % Belarusian – aroE 944 and aroE 972) – 14 variants of shikimat dehydrogenase; 24 alleles of fumC gene (4.2 % Belarusian – fumC 988) – 4 variants of fumarate dehydratase; 18 alleles of gdh gene (16.7 % first identified in Belarus – gdh 560, gdh 985 and gdh 1083) – 4 variants of glucose-6-phosphate dehydrogenase; 18 alleles of pdhC gene – 11 variants of pyruvate dehydrogenase subunit and 20 alleles of pgm gene – 13 variants of phosphoglucomutase (5.6 and 5 % of Belarusian alleles − pdhC 888 and pgm 1099 respectively). Dominant alleles are abcZ 8 – 25 %, adk 5 – 30, aroE 6 – 28.3, fumC 17 – 30, gdh 560 – 20, pdhC 18 – 21.7, pgm 2 – 25 %. The Belarusian meningococcal population is diverse and includes both its own house-keeping gene alleles (7.7 %) and those circulating in other countries (92.3 %). The number of SNPs is varied from 29 (adk) to 125 (aroE). Single nucleotide polymorphisms are mostly synonymous and, on average, lead to amino acid substitutions in the range from 0.6 % in adenylate cyclase and up to 26.4 % in shikimat dehydrogenase.

Himalayan mountains are distinctly characterized for their unique climatic and topographic variations; therefore, unraveling the cold-adaptive mechanisms and processes of native life forms is always being a matter of concern for scientific community. In this perspective, the proteomic response of psychrophilic diazotroph Pseudomonas helmanticensis was studied towards low-temperature conditions. LC-MS-based analysis revealed that most of the differentially expressed proteins providing cold stress resistance were molecular chaperons and cold shock proteins. Enzymes involved in proline, polyamines, unsaturated fatty acid biosynthesis, ROS-neutralizing pathways, and arginine degradation were upregulated. However, proteins involved in the oxidative pathways of energy generation were severalfold downregulated. Besides these, the upregulation of uncharacterized proteins at low temperature suggests the expression of novel proteins by P. helmanticensis for cold adaptation. Protein interaction network of P. helmanticensis under cold revealed that Tif, Tig, DnaK, and Adk were crucial proteins involved in cold adaptation. Conclusively, this study documents the proteome and protein-protein interaction network of the Himalayan psychrophilic P. helmanticensis under cold stress.

Treating chronic pain using opioids, such as morphine, is hampered by the development of opioid-induced hyperalgesia (OIH; increased pain sensitivity), antinociceptive tolerance and withdrawal which can contribute to dependence and abuse. In the central nervous system, the purine nucleoside adenosine has been implicated in beneficial and detrimental actions of morphine, but the extent of their interaction remains poorly understood. Here, we demonstrate that morphine-induced OIH and antinociceptive tolerance in rats is associated with a 2-fold increase in adenosine kinase (ADK) expression in the dorsal horn of the spinal cord (DH-SC). Blocking ADK activity in the spinal cord provided greater than 90% attenuation of OIH and antinociceptive tolerance through A3 adenosine receptor (A3AR) signaling. Supplementing adenosine signaling with selective A3AR agonists blocked OIH and antinociceptive tolerance in rodents of both sexes. Engagement of A3AR in the spinal cord with an ADK inhibitor or A3AR agonist was associated with reduced DH-SC expression of the NOD-like receptor pyrin domain-containing 3 (NLRP3; 60-75%), cleaved caspase 1 (40-60%), interleukin (IL)-1β (76-80%) and tumor necrosis factor (TNF; 50-60%). In contrast, the neuroinhibitory and anti-inflammatory cytokine IL-10 increased 2-fold. In mice, A3AR agonists prevented the development of tolerance in a model of neuropathic pain and reduced naloxone-dependent withdrawal behaviors by greater than 50%. These findings suggest A3AR-dependent adenosine signaling is compromised during sustained morphine to allow the development of morphine-induced adverse effects. These findings raise the intriguing possibility that A3AR agonists may be useful adjunct to opioids to manage their unwanted effects.

AIM The aim of this study was to evaluate transfer of basic science knowledge for clinical application in our BDS programme by exploring the correlations between student performance in integrated dental science (IDS) examinations and applied dental knowledge (ADK) tests. METHODS Numeric test scores were drawn from summative IDS and ADK assessments undertaken by BDS students spanning six academic years (2013-14 to 2018-19) for two cohorts (2013 and 2014). The data included a total of 13 test scores for each cohort with four IDS tests, taken in Years 1 and 2, and nine ADK tests taken in Years 3, 4 and 5. RESULTS The sample included 120 students across both cohorts with 65 females (54%) and 55 males (46%). The correlation coefficients between the successive tests and the combinations of IDS and ADK scores were positive, all being greater than 0.48, and all being significantly correlated (P

This study investigated the efficacy of four Brucella (B.) abortus recombinant proteins, namely adenylate kinase (Adk), nucleoside diphosphate kinase (Ndk), 50S ribosomal protein (L7/L12) and preprotein translocase subunit (SecB), as a combined subunit vaccine (CSV) against B. abortus infection in BALB/c mice. Immunoblotting assay showed that these four recombinant proteins as well as pcold-TF vector reacted individually with Brucella-positive serum, but not with Brucella-negative serum. The peripheral blood CD4+ T cell population was increased in CSV-immunized mice compared to PBS and pcold-TF vector groups. In addition, CSV and pcold-TF groups displayed induced IgG1 and IgG2a antibodies production compared to PBS and RB51 group, whereas IgG2a titer was higher than IgG1 titer in CSV group. The secretion profiles of IgG1 and IgG2a production together with an enhancement of CD4+ T cell population suggested that CSV did not only induce T helper 1 (Th1) T cell immunity but also humoral immunity. Therein, Th1 T cell immunity is more predominant in eliminating intracellular bacteria B. abortus. Furthermore, CSV immunization significantly reduced the bacterial burden in the spleen as well as the spleen weight in comparison to PBS and pcold-TF groups. Altogether, combination of these antigens could be potential to induce protective immunity against B. abortus infection in animals.

Dihydronicotinamide riboside (NRH) has been suggested to act as a precursor for the synthesis of NAD+, but the biochemical pathway converting it has been unknown. Here, we show that NRH can be converted into NAD+ via a salvage pathway in which adenosine kinase (ADK, also known as AK) acts as an NRH kinase. Using isotope-labelling approaches, we demonstrate that NRH is fully incorporated into NAD+, with NMNH acting as an intermediate. We further show that AK is enriched in fractions from cell lysates with NRH kinase activity, and that AK can convert NRH into NAD+. In cultured cells and mouse liver, pharmacological or genetic inhibition of AK blocks formation of reduced nicotinamide mononucleotide (NMNH) and inhibits NRH-stimulated NAD+ biosynthesis. Finally, we confirm the presence of endogenous NRH in the liver with metabolomics. Our findings establish NRH as a natural precursor of NAD+ and reveal a new route for NAD+ biosynthesis via an NRH salvage pathway involving AK.

Objective The "adenosine hypothesis of SUDEP" (sudden unexpected death in epilepsy) predicts that a seizure-induced adenosine surge combined with impaired metabolic clearance can foster lethal apnea or cardiac arrest. Changes in adenosine receptor density and adenosine kinase (ADK) occur in surgical epilepsy patients. Our aim was to correlate the distribution of ADK and adenosine A2A and A1 receptors (A2A R and A1 R) in surgical tissue from patients with temporal lobe epilepsy and hippocampal sclerosis (TLE/HS) with SUDEP risk factors. Methods In 75 cases, patients were stratified into high-risk (n = 16), medium-risk (n = 11) and low-risk (n = 48) categories according to the frequency of generalized seizures before surgery. Using whole-slide scanning Definiens image analysis we quantified the labeling index (LI) for ADK, A2A R, and A1 R in seven regions of interest: temporal cortex, temporal lobe white matter, CA1, CA4, dentate gyrus, subiculum, and amygdala and relative to glial and neuronal densities with glial fibrillary acidic protein (GFAP) and neuronal nuclear antigen (NeuN). Results A1 R showed predominant neuronal, A2A R astroglial, and ADK nuclear labeling in all regions but with significant variation. Compared with the low-risk group, the high-risk group had significantly lower A2A R LI in the temporal cortex. In HS cases with severe neuronal cell loss and gliosis predominantly in the CA1 and CA4 regions, significantly higher A1 R was present in the amygdala in high-risk than in low-risk cases. There was no significant difference in neuronal loss or gliosis between the risk groups or differences for ADK labeling. Significance Reduced cortical A2A R suggests glial dysfunction and impaired adenosine modulation in response to seizures in patients at higher risk for SUDEP. Increased neuronal A1 R in the high-risk group could contribute to periictal amygdala dysfunction in SUDEP.

Dysregulated adenosine signaling pathway has been evidenced in the pathogenesis of breast cancer. However, the role of adenosine kinase (ADK) in tumorigenesis remains unclear while it crucially regulates the removal and availability of adenosine. ADK has two isoforms that localize to discrete subcellular spaces: i.e., nuclear, long-isoform (ADK-L) and cytosolic, short-isoform (ADK-S). We hypothesized that these two ADK isoforms would be differentially expressed in breast cancer and may contribute to divergent cellular actions in cancer. In this study, we examined the expression profiles of ADK isoforms in breast cancer tissues from 46 patient and followed up with an in vitro investigation by knocking down the expression of ADK-L or ADK-S using CRISPR gene editing to evaluate the role of ADK isoform in cancer progression and metastasis of cultured triple-negative breast cancer cell line MDA-MB-231. We demonstrated that (i) ADK-L expression level was significantly increased in breast cancer tissues versus paired normal tissues adjacent to tumor, whereas the ADK-S expression levels were not significantly different between cancerous and normal tissues; (ii) CRISPR/Cas9-mediated downregulation of ADK isoforms, led to suppressed cellular proliferation, division, and migration of cultured breast cancer cells; (iii) ADK-L knockdown significantly upregulated gene expression of matrix metalloproteinase (ADAM23, 9.93-fold; MMP9, 24.58-fold) and downregulated expression of cyclin D2 (CCND2, -30.76-fold), adhesive glycoprotein THBS1 (-8.28-fold), and cystatin E/M (CST6, -16.32-fold). Our findings suggest a potential role of ADK-L in mitogenesis, tumorigenesis, and tumor-associated tissue remodeling and invasion; and the manipulation of ADK-L holds promise as a therapeutic strategy for aggressive breast cancer.

Emerging evidence implicates the sphingosine-1-phosphate (S1P) receptor subtype 1 (S1PR1) in the development of neuropathic pain. Continued investigation of the signaling pathways downstream of S1PR1 are needed to support development of S1PR1 antagonists. In rodents, intrathecal (i.th.) injection of SEW2871, a selective S1PR1 agonist, activates the nod-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome, increases interleukin-1β (IL-1β) and causes behavioral hypersensitivity. I.th. injection of a IL-1β receptor antagonist blocks SEW2871-induced hypersensitivity, suggesting that IL-1β contributes to S1PR1’s actions. Interestingly, previous studies have suggested that IL-1β increases the expression/activity of adenosine kinase (ADK), a key regulator of adenosine signaling at its receptors (ARs). Increased ADK expression reduces adenosine signaling whereas inhibiting ADK restores the action of adenosine. Here, we show that SEW287-induced behavioral hypersensitivity is associated with increased expression of ADK in astrocytes of the dorsal horn of the spinal cord (DH-SC). Moreover, the ADK inhibitor, ABT702, blocks SEW2871-induced hypersensitivity. These findings link ADK activation to S1PR1. If SEW2871-induced pain is mediated by IL-1β, which in turn activates ADK and leads to mechano-allodynia, then blocking ADK should attenuate IL-1β effects. In support of this idea, recombinant rat (rrIL-1β)-induced allodynia was blocked by at least 90% with ABT702, functionally linking ADK to IL-1β. Moreover, the selective A3AR antagonist, MRS1523, prevents the ability of ABT702 to block SEW2871 and IL-1β-induced allodynia, implicating A3AR signaling in the beneficial effects exerted by ABT702. Our findings provide novel mechanistic insight into how S1PR1 signaling in the spinal cord produces hypersensitivity through IL1-b and ADK activation.

Long-term accumulation of adenosine (Ado) in tumor tissues helps to establish the immunosuppressive tumor microenvironment and to promote tumor development. Regulation of Ado metabolism is particularly pivotal for blocking Ado-mediated immunosuppression. The activity of adenosine kinase (ADK) for catalyzing the phosphorylation of Ado plays an essential role in regulating Ado metabolism. Specifically, accumulated Ado in the tumor microenvironment occupies the active site of ADK, inhibiting the phosphorylation of Ado. Phosphate can protect ADK from inactivation and restore the activity of ADK. Herein, calcium phosphate-reinforced iron-based metal-organic frameworks (CaP@Fe-MOFs) are designed to reduce Ado accumulation and to inhibit Ado-mediated immunosuppressive response in the tumor microenvironment. CaP@Fe-MOFs are found to regulate the Ado metabolism by promoting ADK-mediated phosphorylation and relieving the hypoxic tumor microenvironment. Moreover, CaP@Fe-MOFs can enhance the antitumor immune response via Ado regulation, including the increase of T lymphocytes and dendritic cells and the decrease of regulatory T lymphocytes. Finally, CaP@Fe-MOFs are used for cancer treatment in mice, alleviating the Ado-mediated immunosuppressive response and achieving tumor suppression. This study may offer a general strategy for blocking the Ado-mediated immunosuppression in the tumor microenvironment and further for enhancing the immunotherapy efficacy in vivo.

The ongoing SARS-CoV-2 pandemic requires efficient and safe antiviral treatment strategies. Drug repurposing represents a fast and low-cost approach to the development of new medical treatment options. The direct antiviral agent remdesivir has been reported to exert antiviral activity against SARS-CoV-2. Whereas remdesivir only has a very short half-life time and a bioactivation, which relies on pro-drug activating enzymes, its plasma metabolite GS-441524 can be activated through various kinases including the adenosine kinase (ADK) that is moderately expressed in all tissues. The pharmacokinetics of GS-441524 argue for a suitable antiviral drug that can be given to patients with COVID-19. Here, we analyzed the antiviral property of a combined treatment with the remdesivir metabolite GS-441524 and the antidepressant fluoxetine in a polarized Calu-3 cell culture model against SARS-CoV-2. The combined treatment with GS-441524 and fluoxetine were well-tolerated and displayed synergistic antiviral effects against three circulating SARS-CoV-2 variants in vitro in the commonly used reference models for drug interaction. Thus, combinatory treatment with the virus-targeting GS-441524 and the host-directed drug fluoxetine might offer a suitable therapeutic treatment option for SARS-CoV-2 infections.

The adenosine hypothesis of schizophrenia posits that reduced availability of the neuromodulator adenosine contributes to dysregulation of dopamine and glutamate transmission and the symptoms associated with schizophrenia. It has been proposed that increased expression of the enzyme adenosine kinase (ADK) may drive hypofunction of the adenosine system. While animal models of ADK overexpression support such a role for altered ADK, the expression of ADK in schizophrenia has yet to be examined. In this study, we assayed ADK gene and protein expression in frontocortical tissue from schizophrenia subjects. In the dorsolateral prefrontal cortex (DLPFC), ADK-long and -short splice variant expression was not significantly altered in schizophrenia compared to controls. There was also no significant difference in ADK splice variant expression in the frontal cortex of rats treated chronically with haloperidol-decanoate, in a study to identify the effect of antipsychotics on ADK gene expression. ADK protein expression was not significantly altered in the DLPFC or anterior cingulate cortex (ACC). There was no significant effect of antipsychotic medication on ADK protein expression in the DLPFC or ACC. Overall, our results suggest that increased ADK expression does not contribute to hypofunction of the adenosine system in schizophrenia and that alternative mechanisms are involved in dysregulation of this system in schizophrenia.

Severe reduction in the β‐cell number (collectively known as the β‐cell mass) contributes to the development of both type 1 and type 2 diabetes. Recent pharmacological studies have suggested that increased pancreatic β‐cell proliferation could be due to specific inhibition of adenosine kinase (ADK). However, genetic evidence for the function of pancreatic β‐cell ADK under physiological conditions or in a pathological context is still lacking. In this study, we crossed mice carrying LoxP‐flanked Adk gene with Ins2‐Cre mice to acquire pancreatic β ‐cell ADK deficiency (Ins2‐Cre±Adkfl/fl) mice. Our results revealed that Ins2‐Cre+/‐Adkfl/fl mice showed improved glucose metabolism and β‐cell mass in younger mice, but showed normal activity in adult mice. Moreover, Ins2‐Cre±Adkfl/fl mice were more resistant to streptozotocin (STZ) induced hyperglycaemia and pancreatic β‐cell damage in adult mice. In conclusion, we found that ADK negatively regulates β‐cell replication in young mice as well as under pathological conditions, such as STZ induced pancreatic β‐cell damage. Our study provided genetic evidence that specific inhibition of pancreatic β‐cell ADK has potential for anti‐diabetic therapy.