RESUMO
Acanthamoeba castellanii, a free-living amoeba, can be pathogenic to humans causing a corneal infection named Acanthamoeba keratitis (AK). The mannose-binding protein (MBP) is well established as the major factor related to Acanthamoeba pathogenesis. However, additional factors that participate in the adhesion process and protect trophozoites from cytolytic effects caused by host immune responses remain unknown. Ectonucleotidases, including 3'-nucleotidase/nuclease (3'-NT/NU), a bifunctional enzyme that was recently reported in A. castellanii, are frequently related to the establishment of parasitic infections. We verified that trophozoites can hydrolyze 3'-AMP, and this activity is similar to that observed in other protists. The addition of 3'-AMP increases the adhesion of trophozoites to LLC-MK2 epithelial cells, and this stimulation is completely reversed by DTT, an inhibitor of ecto-3'-nucleotidase activity. Lesions in corneal cells caused by AK infection may elevate the extracellular level of 3'-AMP. We believe that ecto-3'-nucleotidase activity can modulate the host immune response, thus facilitating the establishment of parasitic infection. This activity results from the generation of extracellular adenosine, which can bind to purinergic receptors present in host immune cells. Positive feedback may occur in this cascade of events once the ecto-3'-nucleotidase activity of trophozoites is increased by the adhesion of trophozoites to LLC-MK2 cells.
Assuntos
Acanthamoeba castellanii , Adenosina , Adesão Celular , Trofozoítos , Acanthamoeba castellanii/enzimologia , Adenosina/metabolismo , Linhagem Celular , Animais , Nucleotidases/metabolismo , Células Epiteliais/parasitologiaRESUMO
Nucleotidases contribute to the regulation of inflammation, coagulation, and cardiovascular activity. Exercise promotes biological adaptations, but its effects on nucleotidase activities and expression are unclear. The objective of this study was to review systematically the effects of exercise on nucleotidase functionality in healthy and unhealthy subjects. The MEDLINE, EMBASE, Cochrane Library, and Web of Science databases were searched to identify, randomized clinical trials, non-randomized clinical trials, uncontrolled clinical trials, quasi-experimental, pre-, and post-interventional studies that evaluated the effects of exercise on nucleotidases in humans, and was not limited by language and date. Two independent reviewers performed the study selection, data extraction, and assessment of risk of bias. Of the 203 articles identified, 12 were included in this review. Eight studies reported that acute exercise, in healthy and unhealthy subjects, elevated the activities or expression of nucleotidases. Four studies evaluated the effects of chronic training on nucleotidase activities in the platelets and lymphocytes of patients with metabolic syndrome, chronic kidney disease, and hypertension and found a decrease in nucleotidase activities in these conditions. Acute and chronic exercise was able to modify the blood plasma and serum levels of nucleotides and nucleosides. Our results suggest that short- and long-term exercise modulate nucleotidase functionality. As such, purinergic signaling may represent a novel molecular adaptation in inflammatory, thrombotic, and vascular responses to exercise.
Assuntos
Exercício Físico , Hipertensão , Terapia por Exercício , Humanos , Nucleotidases , NucleotídeosRESUMO
BACKGROUND: Acupuncture is a treatment for neuropathic pain, but its mechanism remains unclear. Previous studies showed that analgesia was induced in rats with neuropathic pain when their spinal cord adenosine content increased after electroacupuncture (EA); however, the mechanism behind this electroacupuncture-induced increase has not been clarified. OBJECTIVE: This study aimed to determine the role that ecto-5'-nucleotidase plays in EA-induced analgesia for neuropathic pain. METHODS: We performed electroacupuncture at the Zusanli acupoint on the seventh day after establishing a rat model of neuropathic pain induced through chronic constriction injuries. We observed the mechanical withdrawal threshold and thermal pain threshold and detected the expression of ecto-5'-nucleotidase in the spinal cord using Western blot. Chronic constriction injury rat models were intraperitoneally injected with α,ß-methyleneadenosine 5'-diphosphate, an ecto-5'-nucleotidase inhibitor, 30 min before electroacupuncture. The adenosine content of the spinal cord was detected using high-performance liquid chromatography. Lastly, the adenosine A1 receptor agonist N6-cyclopentyladenosine was intrathecally injected into the lumbar swelling of the rats, and the mechanical withdrawal and thermal pain thresholds were reevaluated. RESULTS: Analgesia and increased ecto-5'-nucleotidase expression and adenosine content in the spinal cord were observed 1 h after electroacupuncture. α,ß-methyleneadenosine 5'-diphosphate was able to inhibit upregulation of adenosine content and electroacupuncture-induced analgesia. After administration of N6-cyclopentyladenosine, electroacupuncture-induced analgesia was restored. CONCLUSIONS: Our results suggest that electroacupuncture at Zusanli can produce analgesia in chronic constriction injury rat models, possibly via the increased ecto-5'-nucleotidase expression induced through electroacupuncture, thus leading to increased adenosine expression in the spinal cord.
Assuntos
Analgesia , Eletroacupuntura , Neuralgia , 5'-Nucleotidase/metabolismo , Adenosina , Animais , Neuralgia/terapia , Nucleotidases , Ratos , Ratos Sprague-Dawley , Medula Espinal/metabolismoRESUMO
Background: Naja atra is a venomous snake species medically relevant in China. In the current study, we evaluated the composition and toxicological profile of venom collected from farm-raised N. atra. Methods: Venom was collected from third-generation captive bred N. atra on a snake farm in Hunan Province, China. The venom was analyzed using sodium dodecyl sulfate polyacrylamide gel electrophoresis and nano-liquid chromatography with electrospray ionization tandem mass spectrometry. In addition, hemolytic activity, median lethal dose, serum biochemical and histopathological parameters were accessed. Results: N. atra venom proteome was dominated by phospholipase A2 (46.5%) and three-finger toxins (41.4 %), and a set of common low relative abundance proteins, including cysteine-rich secretory proteins (4.7%), NGF-beta (2.4%), snake venom metalloproteinase (1.5%), glutathione peroxidase (0.6%), vespryn (0.3%), and 5ʹ-nucleotidases (0.2%) were also found. Furthermore, the venom exhibited direct hemolytic activity, neurotoxicity, myotoxicity, and high lethal potency in mice, with a subcutaneous median lethal dose of 1.02 mg/kg. Histopathological analysis and serum biochemical tests revealed that venom caused acute hepatic, pulmonary and renal injury in mice. Conclusion: This study revealed the composition and toxicity of venom collected from farm-raised N. atra, thereby providing a reference for the analysis of venom samples collected from captive-born venomous snakes in the future.(AU)
Assuntos
Animais , Venenos de Serpentes/toxicidade , Fosfolipases A2 , Naja naja , Miotoxicidade , NucleotidasesRESUMO
Early life stressors, such as social isolation (SI), can disrupt brain development contributing to behavioral and neurochemical alterations in adulthood. Purinergic receptors and ectonucleotidases are key regulators of brain development in embryonic and postnatal periods, and they are involved in several psychiatric disorders, including schizophrenia. The extracellular ATP drives purinergic signaling by activating P2X and P2Y receptors and it is hydrolyzed by ectonucleotidases in adenosine, which activates P1 receptors. The purpose of this study was to investigate if SI, a rodent model used to replicate abnormal behavior relevant to schizophrenia, impacts purinergic signaling. Male Wistar rats were reared from weaning in group-housed or SI conditions for 8 weeks. SI rats exhibited impairment in prepulse inhibition and social interaction. SI presented increased ADP levels in cerebrospinal fluid and ADP hydrolysis in the hippocampus and striatum synaptosomes. Purinergic receptor expressions were upregulated in the prefrontal cortex and downregulated in the hippocampus and striatum. A2A receptors were differentially expressed in SI prefrontal cortex and the striatum, suggesting distinct roles in these brain structures. SI also presented decreased ADP, adenosine, and guanosine levels in the cerebrospinal fluid in response to D-amphetamine. Like patients with schizophrenia, uric acid levels were prominently increased in SI rats after D-amphetamine challenge. We suggest that the SI-induced deficits in prepulse inhibition might be related to the SI-induced changes in purinergic signaling. We provide new evidence that purinergic signaling is markedly affected in a rat model relevant to schizophrenia, pointing out the importance of purinergic system in psychiatry conditions.
Assuntos
Receptores Purinérgicos , Transdução de Sinais , Isolamento Social , Difosfato de Adenosina/líquido cefalorraquidiano , Animais , Comportamento Animal , Estimulantes do Sistema Nervoso Central/farmacologia , Dextroanfetamina/farmacologia , Masculino , Nucleotidases/metabolismo , Ratos , Ratos Wistar , Receptor A2A de Adenosina/metabolismo , Receptores Purinérgicos P2X/metabolismo , Receptores Purinérgicos P2Y/metabolismo , Reflexo de Sobressalto , Psicologia do Esquizofrênico , Comportamento Social , Isolamento Social/psicologia , DesmameRESUMO
BACKGROUND: Alzheimer's disease (AD) is a chronic, progressive neurodegenerative disease. Recent studies have reported the close association between cognitive function in AD and purinergic receptors in the central nervous system. In the current study, we investigated the effect of CD73 inhibitor α, ß-methylene ADP (APCP) on cognitive impairment of AD in mice, and to explore the potential underlying mechanisms. RESULTS: We found that acute administration of Aß142 (i.c.v.) resulted in a significant increase in adenosine release by using microdialysis study. Chronic administration of APCP (10, 30 mg/kg) for 20 d obviously mitigated the spatial working memory impairment of Aß142-treated mice in both Morris water maze (MWM) test and Y-maze test. In addition, the extracellular adenosine production in the hippocampus was inhibited by APCP in Aß-treated mice. Further analyses indicated expression of acetyltransferase (ChAT) in hippocampus of mice of was significantly reduced, while acetylcholinesterase (AChE) expression increased, which compared to model group. We observed that APCP did not significantly alter the NLRP3 inflammasome activity in hippocampus, indicating that anti-central inflammation seems not to be involved in APCP effect. CONCLUSIONS: In conclusion, we report for the first time that inhibition of CD73 by APCP was able to protect against memory loss induced by Aß142 in mice, which may be due to the decrease of CD73-driven adenosine production in hippocampus. Enhancement of central cholinergic function of the central nervous system may also be involved in the effects of APCP.
Assuntos
Animais , Masculino , Camundongos , Difosfato de Adenosina/análogos & derivados , Doenças Neurodegenerativas/prevenção & controle , Hipocampo , Nucleotidases/antagonistas & inibidores , Acetilcolinesterase , Difosfato de Adenosina/administração & dosagem , Doença de Alzheimer/prevenção & controle , Teste do Labirinto Aquático de Morris , Camundongos Endogâmicos C57BLRESUMO
Progressive diabetic nephropathy (DN) and loss of renal function correlate with kidney fibrosis. Crosstalk between TGF-ß and adenosinergic signaling contributes to the phenotypic transition of cells and to renal fibrosis in DN models. We evaluated the role of TGF-ß on NT5E gene expression coding for the ecto-5`-nucleotidase CD73, the limiting enzyme in extracellular adenosine production. We showed that high d-glucose may predispose HK-2 cells towards active transcription of the proximal promoter region of the NT5E gene while additional TGF-ß results in full activation. The epigenetic landscape of the NT5E gene promoter was modified by concurrent TGF-ß with occupancy by the p300 co-activator and the phosphorylated forms of the Smad2/3 complex and RNA Pol II. Transcriptional induction at NT5E in response to TGF-ß was earlier compared to the classic responsiveness genes PAI-1 and Fn1. CD73 levels and AMPase activity were concomitantly increased by TGF-ß in HK-2 cells. Interestingly, we found increased CD73 content in urinary extracellular vesicles only in diabetic patients with renal repercussions. Further, CD73-mediated AMPase activity was increased in the urinary sediment of DN patients. We conclude that the NT5E gene is a target of the profibrotic TGF-ß cascade and is a traceable marker of progressive DN.
Assuntos
5'-Nucleotidase/genética , Nefropatias Diabéticas/genética , Fibrose/genética , Fator de Crescimento Transformador beta/genética , Adenosina/biossíntese , Biomarcadores/metabolismo , Linhagem Celular , Nefropatias Diabéticas/patologia , Proteína p300 Associada a E1A/genética , Epigênese Genética/genética , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Fibrose/patologia , Proteínas Ligadas por GPI/genética , Regulação da Expressão Gênica , Humanos , Rim/metabolismo , Rim/patologia , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/patologia , Nucleotidases/genética , Regiões Promotoras Genéticas/genética , RNA Polimerase II/genéticaRESUMO
The cholinergic, purinergic and oxidative stress systems were related to nervous system damage in some pathologies, as well as being involved in pro-inflammatory and anti-inflammatory pathways. The objective was to investigate changes in purinergic, cholinergic systems and oxidative stress related to the neuropathology of listeriosis. Gerbils were used as experimental models. The animals were divided in two groups: control and infected. The animals were orally infected with 5â¯×â¯108 CFU/animal of the pathogenic strain of Listeria monocytogenes. Collected of material was 6 and 12th days post-infection (PI). Infected animals showed moderate mixed inflammatory infiltrates in the liver. The spleen and brain was used for PCR analyses, confirming infection by L. monocytogenes. Increase in number of total leukocytes because of an increase in lymphocytes in infected (Pâ¯<â¯0.001). ATP and ADP hydrolysis by NTPDase was lower at 6 and 12th days PI in infected animals than in the control group. ADA (adenosine deaminase) activity was higher on the 6th day PI (Pâ¯<â¯0.05) and decreased on the 12th day PI (Pâ¯<â¯0.05) in infected animals. AChE (acetylcholinesterase) activity did not differ between groups on the 6th day PI; however, activity decreased in infected group on the 12th day PI (Pâ¯<â¯0.05). On the 12th day PI, an increase of oxygen-reactive species levels and lower catalase and superoxide dismutase activities in the infected group was observed, characterizing a situation of cerebral oxidative stress. The inflammatory and oxidative mechanisms are present in listeriosis in asymptomatic animals, and that ectonucleotidases and cholinesterase's are involved in immunomodulation.
Assuntos
Listeria monocytogenes/patogenicidade , Listeriose/metabolismo , Listeriose/patologia , Acetilcolinesterase/metabolismo , Adenosina Desaminase/metabolismo , Animais , Encéfalo/enzimologia , Encéfalo/microbiologia , Encéfalo/patologia , Catalase/metabolismo , DNA Bacteriano/isolamento & purificação , Doenças Transmitidas por Alimentos/microbiologia , Gerbillinae , Hematócrito , Intestino Delgado/patologia , Contagem de Leucócitos , Listeria monocytogenes/genética , Listeriose/enzimologia , Listeriose/transmissão , Fígado/patologia , Nucleotidases/metabolismo , Estresse Oxidativo , Espécies Reativas de Oxigênio/análise , Baço/microbiologia , Baço/patologia , Superóxido Dismutase/metabolismoRESUMO
Leishmaniasis is one of the most significant of the neglected tropical diseases, with 350 million people in 98 countries worldwide living at risk of developing one of the many forms of the disease. During the transmission of the parasite from its vector to the vertebrate host, neutrophils are rapidly recruited to the site of the sandfly bite. Using different strategies, neutrophils can often kill a large number of parasites. However, some parasites can resist neutrophil-killing mechanisms and survive until macrophage arrival at the infection site. One of the strategies for neutrophil-mediated killing is the production of neutrophil extracellular traps (NETs). Because of its ecto-localized nuclease activity, the enzyme 3'-nucleotidase/nuclease (3'NT/NU), present in different Leishmania species, was recently identified as part of a possible parasite escape mechanism from NET-mediated death. Previous studies showed that 3'NT/NU also plays an important role in the establishment of Leishmania infection by generating extracellular adenosine that favors the parasite and macrophage interaction. This study aims to deepen the knowledge about 3'NT/NU, mainly with respect to its nuclease activity that is little studied in the current literature. For this, we cloned, expressed and purified the recombinant La3'NT/NU and have confirmed its contribution to the parasite escape from NET-mediated killing.
Assuntos
Desoxirribonucleases/imunologia , Armadilhas Extracelulares/imunologia , Leishmania/enzimologia , Leishmaniose/imunologia , Neutrófilos/imunologia , Nucleotidases/imunologia , Proteínas de Protozoários/imunologia , Clonagem Molecular , Desoxirribonucleases/genética , Armadilhas Extracelulares/parasitologia , Humanos , Leishmania/genética , Leishmania/imunologia , Leishmaniose/parasitologia , Nucleotidases/genética , Proteínas de Protozoários/genéticaRESUMO
The incidence of papillary thyroid carcinoma (PTC) has been increasing, which raised the interest in its molecular pathways. Although the high expression of ecto-5'-nucleotidase (NT5E) gene expression and NT5E enzymatic activity in several types of cancer is associated with tumor progression, its role in PTC remains unknown. Here, we investigated the AMP hydrolysis in human normal thyroid cells and PTC cells, in primary culture, and the association of NT5E expression with clinical aspects of PTC patients. AMPase activity was higher in thyroid cells isolated from PTC, as compared to normal thyroid (Pâ¯=â¯0.0063). Significant correlation was observed between AMPase activity and NT5E levels in primary thyroid cell cultures (râ¯=â¯0.655, Pâ¯=â¯0.029). NT5E expression was higher in PTC than in the adjacent non-malignant thyroid tissue (Pâ¯=â¯0.0065) and were positively associated with metastatic lymph nodes (Pâ¯=â¯0.0007), risk of recurrence (Pâ¯=â¯0.0033), tumor size (Pâ¯=â¯0.049), and nodular hyperplasia in the adjacent thyroid parenchyma, when compared to normal thyroid or lymphocytic thyroiditis (Pâ¯=â¯0.0146). After adjusting for potential confounders, the malignant/non-malignant paired expression ratio of NT5E mRNA was independently associated with metastatic lymph nodes (Pâ¯=â¯0.0005), and tumor size (P=0.0005). In addition, the analysis of PTC described in the TCGA database also showed an association between higher expression of NT5E and metastatic lymph nodes, and tumor microinvasion. These results support the hypothesis that NT5E have a role in PTC microenvironment and might be a potential target for PTC therapy.
Assuntos
5'-Nucleotidase/metabolismo , Linfonodos/enzimologia , Linfonodos/patologia , Metástase Linfática/patologia , Câncer Papilífero da Tireoide/enzimologia , Câncer Papilífero da Tireoide/patologia , 5'-Nucleotidase/genética , Linhagem Celular Tumoral , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Pessoa de Meia-Idade , Nucleotidases/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Câncer Papilífero da Tireoide/genética , Glândula Tireoide/patologiaRESUMO
Evidences show that purinergic signaling is involved in processes associated with health and disease, including noncommunicable, neurological, and degenerative diseases. These diseases strike from children to elderly and are generally characterized by progressive deterioration of cells, eventually leading to tissue or organ degeneration. These pathological conditions can be associated with disturbance in the signaling mediated by nucleotides and nucleosides of adenine, in expression or activity of extracellular ectonucleotidases and in activation of P2X and P2Y receptors. Among the best known of these diseases are atherosclerosis, hypertension, cancer, epilepsy, Alzheimer's disease (AD), Parkinson's disease (PD), and multiple sclerosis (MS). The currently available treatments present limited effectiveness and are mostly palliative. This review aims to present the role of purinergic signaling highlighting the ectonucleotidases E-NTPDase, E-NPP, E-5'-nucleotidase, and adenosine deaminase in noncommunicable, neurological, and degenerative diseases associated with the cardiovascular and central nervous systems and cancer. In conclusion, changes in the activity of ectonucleotidases were verified in all reviewed diseases. Although the role of ectonucleotidases still remains to be further investigated, evidences reviewed here can contribute to a better understanding of the molecular mechanisms of highly complex diseases, which majorly impact on patients' quality of life.
Assuntos
Doenças Cardiovasculares/enzimologia , Neoplasias/enzimologia , Doenças Neurodegenerativas/enzimologia , Nucleotidases/metabolismo , Receptores Purinérgicos/metabolismo , Animais , Humanos , Doenças não Transmissíveis , Qualidade de Vida , Transdução de SinaisRESUMO
It is recognized that the purinergic system, through the activities of ectonucleoside triphosphate diphosphohydrolase (E-NTPDase), ecto-5'-nucleotidase (E-5'-nucleotidase), and ecto-adenosine deaminase (E-ADA), is involved in the regulation and modulation of the physiological and pathological events linked to hemostasis. This occurs due to the role of adenosine diphosphate (ADP) in the activation and recruitment of platelets, and the role of adenosine (Ado) in the inhibition of platelet activation. Thus, here we aimed to evaluate whether Aeromonas caviae infection impairs the ecto-enzymes of the purinergic system in fish thrombocytes and the involvement of this system in the hemorrhagic septicemia. The total number of fish thrombocytes decreased in infected animals compared to uninfected animals. Regarding the ecto-enzymes of the purinergic system, the E-NTPDase and E-5'-nucleotidase activities increased in infected animals compared to uninfected animals, while the E-ADA activity decreased. These findings show that adenine nucleotide hydrolysis is modified in the thrombocytes of fish experimentally infected with A. caviae, which impairs the coagulation process due the excessive hydrolysis of ADP, a molecule linked with activation and recruitment of thrombocytes at the site of vascular injury, and augmentation on Ado levels, a molecule linked with inhibitory effects on platelet activation and aggregation. In summary, the purinergic system might contribute to the occurrence of hemorrhagic frames in fish infected with A. caviae.
Assuntos
Aeromonas caviae/patogenicidade , Plaquetas/metabolismo , Ativação Enzimática , Infecções por Bactérias Gram-Negativas/veterinária , Nucleotidases/metabolismo , 5'-Nucleotidase/metabolismo , Adenosina/fisiologia , Adenosina Desaminase , Animais , Brasil , Peixes-Gato/microbiologia , Doenças dos Peixes/microbiologia , Peixes , Hidrólise , PirofosfatasesRESUMO
The aim of this study was to verify the effect of diphenyl diselenide (PhSe)2 on hepatic nucleotidases and on the concentration of purines in mice infected by Toxoplasma gondii. The animals were divided into four groups: Group A (uninfected), Group B (uninfected and treated with (PhSe)2), Group C (infected), and Group D (infected and treated with (PhSe)2). The inoculation (groups C and D) was performed with 50 cysts of T. gondii (ME-49 strain). Mice from groups B and D were treated with 5 µmol kg-1 of (PhSe)2. Liver tissue from infected mice showed less severe inflammation, elevated ATP/ADO ratio, elevated NTPDase, 5'nucleotidase, and ADA activities compared to the uninfected group (Group A; P < 0.05). However, infected and treated mice showed decreased ATP levels and elevated ADO levels, as well as higher NTPDase and 5'nucleotidase activities and decreased ADA activity in the hepatic tissue compared to the infected group (P < 0.05). Moreover, the (PhSe)2 treatment of infected mice reduced the hepatic inflammation and showed an immunomodulatory effect on ectonucleotidases of hepatic lymphocytes, which it returned to basal levels. Therefore, chronic infection by T. gondii induces hepatic inflammation in mice, and it is possible that purine levels and nucleotidase activities in hepatic tissue are related to the pathogenesis of the infection in this tissue. The treatment with (PhSe)2 was able to reverse the hepatic inflammation in mice chronically infected, possibly due to the modulation of purinergic enzymes that produce an anti-inflammatory profile through the purinergic system in the liver tissue.
Assuntos
Derivados de Benzeno/farmacologia , Inflamação/patologia , Fígado/efeitos dos fármacos , Fígado/patologia , Compostos Organosselênicos/farmacologia , Toxoplasmose/patologia , Animais , Camundongos , Nucleotidases/efeitos dos fármacos , Nucleotidases/metabolismo , Purinas/metabolismoRESUMO
Toxoplasma gondii, an intracellular protozoan, may cause chronic infection in the brain tissue of the host inducing a systemic pro-inflammatory profile. Chronic infections can induce numerous physiological changes, such as alterations in the immune and oxidative profiles. Diphenyl diselenide (PhSe)2, an organoselenium compound, has shown antioxidant and immunomodulatory activities in recent studies. So, the aim of this study was to investigate the activity of purinergic enzymes and reactive oxygen species (ROS) in serum and spleen of mice chronically infected by T. gondii, untreated and treated with (PhSe)2. For this experiment, were divided into four groups: Group A (healthy mice), Group B (healthy mice treated with (PhSe)2), Group C (infected mice) and Group D (infected mice treated with (PhSe)2). Group C and group D were infected via oral route with ME49 Toxoplasma gondii strain. Groups B and D were treated subcutaneously with 5 µmol kg-1 of (PhSe)2. Chronic T. gondii infection induced splenomegaly and physiological changes in the spleen and raised histologic inflammatory markers, ROS levels and the activity of purinergic enzymes activity such as NTPDase, 5´nucleotidase and ADA. In serum, the infection increased 5´nucleotidase and ADA activities. (PhSe)2per se has managed to decrease ROS levels and ADA activity and increase NTPDase and 5´nucleotidase in spleen. In infected mice, treatment with (PhSe)2 reversed splenomegaly, reduced histological inflammatory markers, ROS levels and ADA activity in the spleen. Our results prove that chronic toxoplasmosis can induce splenomegaly, heightens ROS levels and purinergic enzyme activity in mice. These results suggest that (PhSe)2 is a potential therapy for the alterations found in the spleen in chronic T. gondii infection.
Assuntos
Derivados de Benzeno/uso terapêutico , Nucleotidases/sangue , Compostos Organosselênicos/uso terapêutico , Baço/patologia , Toxoplasmose Animal/tratamento farmacológico , 5'-Nucleotidase/sangue , 5'-Nucleotidase/metabolismo , Difosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Derivados de Benzeno/farmacologia , Feminino , Inflamação/tratamento farmacológico , Camundongos , Nucleotidases/metabolismo , Compostos Organosselênicos/farmacologia , Espécies Reativas de Oxigênio/sangue , Espécies Reativas de Oxigênio/metabolismo , Baço/efeitos dos fármacos , Baço/enzimologia , Baço/metabolismo , Toxoplasmose Animal/enzimologia , Toxoplasmose Animal/patologiaRESUMO
The enzymatic activities of NTPDase, 5'-nucleotidase and adenosine deaminase (ADA) are important in regulating the concentration of adenine nucleotides, molecules known to be involved on platelet aggregation. Fasciolosis causes coagulation disorders that have not been completely elucidated. Taking into consideration the association between the purinergic system and hemostasis, this study aimed to evaluate the enzymatic activities of NTPDase (hydrolyze ATP and ADP), 5'-nucleotidase (hydrolyze AMP) and ADA (deamination of adenosine) in platelets from cattle experimentally infected by Fasciola hepatica on days 20, 40, 60 and 80 post-infection (PI). For this study, 10 healthy Friesian steers were separated into two groups: the group A (n = 5) was used as uninfected control, and the group B was composed of steers experimentally infected by F. hepatica (n = 5). The number of platelets did not differ between groups in the periods evaluated. Reduction of NTPDase (p < 0.05) hydrolysing ATP (days 20, 40 and 60 PI), and ADP (days 40, 60 and 80 PI), and on 5'-nucleotidase hydrolyzing AMP (days 40 and 60 PI) was observed. A reduction (p < 0.05) in ADA activity on day 20 PI, as well as an increase (p < 0.05) in ADA activity on days 40 and 60 PI was observed when compared to the control. Based on these results, we can conclude that ATP, ADP and AMP hydrolysis and adenosine deamination were altered in platelets of cattle infected by F. hepatica. Considering the importance of the purinergic system in hemostasis, it is believed that those changes may contribute to the coagulation impairment observed in acute fasciolosis described in the literature.
Assuntos
Adenosina Desaminase/sangue , Plaquetas/enzimologia , Doenças dos Bovinos/parasitologia , Fasciolíase/veterinária , Nucleotidases/sangue , Animais , Bovinos , Doenças dos Bovinos/sangue , Doenças dos Bovinos/enzimologia , Fasciola hepatica/fisiologia , Fasciolíase/sangue , Fasciolíase/enzimologia , Fezes/parasitologia , Fígado/parasitologia , Fígado/patologia , Masculino , Contagem de Ovos de Parasitas/veterinária , Contagem de Plaquetas/veterináriaAssuntos
Anormalidades Craniofaciais/genética , Nanismo/genética , Homozigoto , Instabilidade Articular/genética , Mutação de Sentido Incorreto , Ossificação Heterotópica/genética , Pentosiltransferases/genética , Polidactilia/genética , Criança , Pré-Escolar , Consanguinidade , Anormalidades Craniofaciais/patologia , Nanismo/patologia , Feminino , Humanos , Lactente , Recém-Nascido , Instabilidade Articular/patologia , Nucleotidases/genética , Ossificação Heterotópica/patologia , Polidactilia/patologia , UDP Xilose-Proteína XilosiltransferaseRESUMO
The protozoan parasite Leishmania amazonensis is the etiological agent of cutaneous leishmaniasis. During its life cycle, the flagellated metacyclic promastigote forms are transmitted to vertebrate hosts by sandfly bites, and they develop into amastigotes inside macrophages, where they multiply. L. amazonensis possesses a bifunctional enzyme, called 3'-nucleotidase/nuclease (3'NT/NU), which is able to hydrolyze extracellular 3'-monophosphorylated nucleosides and nucleic acids. 3'NT/NU plays an important role in the generation of extracellular adenosine and has been described as a key enzyme in the acquisition of purines by trypanosomatids. Furthermore, it has been observed that 3'NT/NU also plays a valuable role in the establishment of parasitic infection. In this context, this study aimed to investigate the modulation of the 3'-nucleotidase (3'NT) activity of L. amazonensis by several nucleotides. It was observed that 3'NT activity is inhibited by micromolar concentrations of guanosine and guanine nucleotides. The inhibition promoted by 5'-GMP on the 3'NT activity of L. amazonensis is reversible and uncompetitive because the addition of the inhibitor decreased the kinetic parameters Km and Vmax. Finally, we found that the addition of 5'-GMP is able to reverse the stimulation promoted by 3'-AMP in a macrophage-parasite interaction assay. The determination of compounds that can inhibit the 3'NT activity of Leishmania is very important because this enzyme does not occur in mammals, making it a potential therapeutic target.
Assuntos
Guanosina Difosfato/farmacologia , Guanosina Monofosfato/farmacologia , Guanosina Trifosfato/farmacologia , Leishmania mexicana/enzimologia , Nucleotidases/antagonistas & inibidores , Animais , Cinética , Leishmania mexicana/efeitos dos fármacos , Macrófagos/parasitologia , Camundongos , Nucleotidases/metabolismo , Células RAW 264.7RESUMO
Leishmaniasis is a widespread neglected tropical disease caused by parasites of the Leishmania genus. These parasites express the enzyme 3'-nucleotidase/nuclease (3'NT/NU), which has been described to be involved in parasite nutrition and infection. Bacteria that express nucleases escape the toxic effects of neutrophil extracellular traps (NETs). Hence, we investigated the role of 3'NT/NU in Leishmania survival of NET-mediated killing. Promastigotes of Leishmania infantum were cultured in high-phosphate (HP) or low-phosphate (LP) medium to modulate nuclease activity. We compared the survival of the two different groups of Leishmania during interaction with human neutrophils, assessing the role of neutrophil extracellular traps. As previously reported, we detected higher nuclease activity in parasites cultured in LP medium. Both LP and HP promastigotes were capable of inducing the release of neutrophil extracellular traps from human neutrophils in a dose- and time-dependent manner. LP parasites had 2.4 times more survival than HP promastigotes. NET disruption was prevented by the treatment of the parasites with ammonium tetrathiomolybdate (TTM), a 3'NT/NU inhibitor. Inhibition of 3'NT/NU by 3'-AMP, 5'-GMP, or TTM decreased promastigote survival upon interaction with neutrophils. Our results show that Leishmania infantum induces NET release and that promastigotes can escape NET-mediated killing by 3'-nucleotidase/nuclease activity, thus ascribing a new function to this enzyme.
Assuntos
Leishmania infantum/enzimologia , Neutrófilos/parasitologia , Nucleotidases/fisiologia , Sobrevivência Celular/fisiologia , Espaço Extracelular , Humanos , Leishmaniose Visceral , Fosfatos/farmacologiaRESUMO
Ecto-enzymes can be defined as membrane-bound proteins that have their active site facing the extracellular millieu. In trypanosomatids, the physiological roles of these enzymes remain to be completed elucidated; however, many important events have already been related to them, such as the survival of parasites during their complex life cycle and the successful establishment of host infection. This chapter focuses on two remarkable classes of ecto-enzymes: ecto-nucleotidases and ecto-phosphatases, summarizing their occurrence and possible physiological roles in Leishmania and Trypanosoma genera. Ecto-nucleotidases are characterized by their ability to hydrolyze extracellular nucleotides, playing an important role in purinergic signaling. By the action of these ecto-enzymes, parasites are capable of modulating the host immune system, which leads to a successful parasite infection. Furthermore, ecto-nucleotidases are also involved in the purine salvage pathway, acting in the generation of nucleosides that are able to cross plasma membrane via specialized transporters. Another important ecto-enzyme present in a vast number of pathogenic organisms is the ecto-phosphatase. These enzymes are able to hydrolyze extracellular phosphorylated substrates, releasing free inorganic phosphate that can be internalized by the cell, crossing the plasma membrane through a Pi-transporter. Ecto-phosphatases are also involved in the invasion and survival of parasite in the host cells. Several alternative functions have been suggested for these enzymes in parasites, such as participation in their proliferation, differentiation, nutrition and protection. In this context, the present chapter provides an overview of recent discoveries related to the occurrence of ecto-nucleotidase and ecto-phosphatase activities in Leishmania and Trypanosoma parasites.
Assuntos
Leishmania/enzimologia , Nucleotidases/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo , Trypanosoma/enzimologia , Animais , Transdução de SinaisRESUMO
Hyperargininemia is an inborn error of metabolism (IEM) characterized by tissue accumulation of arginine (Arg). Mental retardation and other neurological features are common symptoms in hyperargininemic patients. Considering purinergic signaling has a crucial role from the early stages of development and underlying mechanisms of this disease are poorly established, we investigated the effect of Arg administration on locomotor activity, morphological alterations, and extracellular nucleotide hydrolysis in larvae and adult zebrafish. We showed that 0.1 mM Arg was unable to promote changes in locomotor activity. In addition, 7-day-post-fertilization (dpf) larvae treated with Arg demonstrated a decreased body size. Arg exposure (0.1 mM) promoted an increase in ATP, ADP, and AMP hydrolysis when compared to control group. These findings demonstrated that Arg might affect morphological parameters and ectonucleotidase activities in zebrafish larvae, suggesting that purinergic system is a target for neurotoxic effects induced by Arg.