RESUMO
The hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is a rare autosomal recessive inborn error of the urea cycle caused by mutations in the SLC25A15 gene. Besides the well-known metabolic complications, patients often present intercurrent infections associated with acute hyperammonemia and metabolic decompensation. However, it is currently unknown whether intercurrent infections are associated with immunological alterations besides the known metabolic imbalances. Herein, we describe the case of a 3-years-old girl affected by the HHH syndrome caused by two novel SLC25A15 gene mutations associated with immune phenotypic and functional alterations. She was admitted to the hospital with an episode of recurrent otitis, somnolence, confusion, and lethargy. Laboratory tests revealed severe hyperammonemia, elevated serum levels of liver transaminases, hemostasis alterations, hyperglutaminemia and strikingly increased orotic aciduria. Noteworthy, serum protein electrophoresis showed a reduction in the gamma globulin fraction. Direct sequencing of the SLC25A15 gene revealed two heterozygous non-conservative substitutions in the exon 5: c.649G>A (p.Gly217Arg) and c.706A>G (p.Arg236Gly). In silico analysis indicated that both mutations significantly impair protein structure and function and are consistent with the patient clinical status confirming the diagnosis of HHH syndrome. In addition, the immune analysis revealed reduced levels of serum IgG and striking phenotypic and functional alterations in the T and B cell immune compartments. Our study has identified two non-previously described mutations in the SLC25A15 gene underlying the HHH syndrome. Moreover, we are reporting for the first time functional and phenotypic immunologic alterations in this rare inborn error of metabolism that would render the patient immunocompromised and might be related to the high frequency of intercurrent infections observed in patients bearing urea cycle disorders. Our results point out the importance of a comprehensive analysis to gain further insights into the underlying pathophysiology of the disease that would allow better patient care and quality of life.
Assuntos
Hiperamonemia , Distúrbios Congênitos do Ciclo da Ureia , Sistemas de Transporte de Aminoácidos Básicos/genética , Pré-Escolar , Feminino , Humanos , Hiperamonemia/complicações , Hiperamonemia/diagnóstico , Proteínas de Transporte da Membrana Mitocondrial , Ornitina/deficiência , Qualidade de Vida , Distúrbios Congênitos do Ciclo da Ureia/complicações , Distúrbios Congênitos do Ciclo da Ureia/diagnóstico , Distúrbios Congênitos do Ciclo da Ureia/genéticaRESUMO
Hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is caused by deficiency of ornithine translocase leading to predominant tissue accumulation and high urinary excretion of ornithine (Orn), homocitrulline (Hcit) and ammonia. Although affected patients commonly present neurological dysfunction manifested by cognitive deficit, spastic paraplegia, pyramidal and extrapyramidal signs, stroke-like episodes, hypotonia and ataxia, its pathogenesis is still poorly known. Although astrocytes are necessary for neuronal protection. Therefore, in the present study we investigated the effects of Orn and Hcit on cell viability (propidium iodide incorporation), mitochondrial function (thiazolyl blue tetrazolium bromide-MTT-reduction and mitochondrial membrane potential-ΔΨm), antioxidant defenses (GSH) and pro-inflammatory response (NFkB, IL-1ß, IL-6 and TNF-α) in unstimulated and menadione-stressed cortical astrocytes that were previously shown to be susceptible to damage by neurotoxins. We first observed that Orn decreased MTT reduction, whereas both amino acids decreased GSH levels, without altering cell viability and the pro-inflammatory factors in unstimulated astrocytes. Furthermore, Orn and Hcit decreased cell viability and ΔΨm in menadione-treated astrocytes. The present data indicate that the major compounds accumulating in HHH syndrome impair mitochondrial function and reduce cell viability and the antioxidant defenses in cultured astrocytes especially when stressed by menadione. It is presumed that these mechanisms may be involved in the neuropathology of this disease.
Assuntos
Astrócitos/efeitos dos fármacos , Citrulina/análogos & derivados , Mitocôndrias/efeitos dos fármacos , Ornitina/farmacologia , Sistemas de Transporte de Aminoácidos Básicos/efeitos dos fármacos , Animais , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Astrócitos/metabolismo , Morte Celular/efeitos dos fármacos , Citrulina/farmacologia , Hiperamonemia/tratamento farmacológico , Hiperamonemia/metabolismo , Masculino , Mitocôndrias/metabolismo , Ornitina/deficiência , Ornitina/metabolismo , Ratos Wistar , Fator de Necrose Tumoral alfa/metabolismo , Distúrbios Congênitos do Ciclo da Ureia/tratamento farmacológico , Distúrbios Congênitos do Ciclo da Ureia/metabolismoRESUMO
Hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is an inborn error of metabolism caused by a defect in the transport of ornithine (Orn) into mitochondrial matrix leading to accumulation of Orn, homocitrulline (Hcit), and ammonia. Affected patients present a variable clinical symptomatology, frequently associated with cerebellar symptoms whose pathogenesis is poorly known. Although in vitro studies reported induction of oxidative stress by the metabolites accumulating in HHH syndrome, so far no report evaluated the in vivo effects of these compounds on redox homeostasis in cerebellum. Therefore, the present work was carried out to investigate the in vivo effects of intracerebellar administration of Orn and Hcit on antioxidant defenses (reduced glutathione concentrations and the activities of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, and glucose-6-phosphate dehydrogenase), lipid oxidation (malondialdehyde concentrations), as well as on the activity of synaptic Na(+), K(+)-ATPase, an enzyme highly vulnerable to free radical attack, in the cerebellum of adolescent rats. Orn significantly increased malondialdehyde levels and the activities of all antioxidant enzymes, and reduced Na(+), K(+)-ATPase activity. In contrast, glutathione concentrations were not changed by Orn treatment. Furthermore, intracerebellar administration of Hcit was not able to alter any of these parameters. The present data show for the first time that Orn provokes in vivo lipid oxidative damage, activation of the enzymatic antioxidant defense system, and reduction of the activity of a crucial enzyme involved in neurotransmission. It is presumed that these pathomechanisms may contribute at least partly to explain the neuropathology of cerebellum abnormalities and the ataxia observed in patients with HHH syndrome.
Assuntos
Cerebelo/efeitos dos fármacos , Hiperamonemia/etiologia , Ornitina/deficiência , Ornitina/farmacologia , ATPase Trocadora de Sódio-Potássio/metabolismo , Sinapses/efeitos dos fármacos , Distúrbios Congênitos do Ciclo da Ureia/etiologia , Animais , Antioxidantes/metabolismo , Cerebelo/metabolismo , Glutationa/metabolismo , Homeostase/efeitos dos fármacos , Hiperamonemia/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Malondialdeído/metabolismo , Ornitina/administração & dosagem , Ornitina/metabolismo , Oxirredução/efeitos dos fármacos , Ratos , Ratos Wistar , Maturidade Sexual/fisiologia , Sinapses/metabolismo , Distúrbios Congênitos do Ciclo da Ureia/metabolismoRESUMO
AIMS: Cerebellar ataxia is commonly observed in hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome, an inherited metabolic disorder biochemically characterized by ornithine (Orn), homocitrulline (Hcit) and ammonia accumulation. Since the pathophysiology of cerebellum damage in this disorder is still unknown, we investigated the effects of Hcit and Orn on important parameters of redox and energy homeostasis in cerebellum of young rats. MATERIAL AND METHODS: We determined thiobarbituric acid-reactive substance (TBA-RS) levels, carbonyl content, nitrate and nitrite production, hydrogen peroxide production, GSH concentrations, sulfhydryl content, as well as activities of respiratory chain complexes I-IV, creatine kinase, Na(+),K(+)-ATPase, aconitase and α-ketoglutarate dehydrogenase. KEY FINDINGS: Orn and Hcit significantly increased TBA-RS levels (lipid oxidation), that was totally prevented by melatonin and reduced glutathione (GSH). We also found that nitrate and nitrite production was not altered by any of the metabolites, in contrast to hydrogen peroxide production which was significantly enhanced by Hcit. Furthermore, GSH concentrations were significantly reduced by Orn and Hcit and sulfhydryl content by Orn, implying an impairment of antioxidant defenses. As regards energy metabolism, Orn and Hcit provoked a significant reduction of aconitase activity, without altering the other parameters. Furthermore, Orn-elicited reduction of aconitase activity was totally prevented by GSH, indicating that the critical groups of this enzyme were susceptible to oxidation caused by this amino acid. SIGNIFICANCE: Taken together, our data indicate that redox homeostasis is disturbed by the major metabolites accumulating in HHH syndrome and that this mechanism may be implicated in the ataxia and cerebellar abnormalities observed in this disorder.
Assuntos
Cerebelo/metabolismo , Citrulina/análogos & derivados , Homeostase/efeitos dos fármacos , Hiperamonemia/metabolismo , Ornitina/farmacologia , Distúrbios Congênitos do Ciclo da Ureia/metabolismo , Aconitato Hidratase/metabolismo , Animais , Cerebelo/patologia , Citrulina/farmacologia , Creatina Quinase/metabolismo , Transporte de Elétrons , Glutationa/metabolismo , Peróxido de Hidrogênio/metabolismo , Hiperamonemia/patologia , Complexo Cetoglutarato Desidrogenase/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Nitratos/metabolismo , Nitritos/metabolismo , Ornitina/deficiência , Ornitina/metabolismo , Oxirredução/efeitos dos fármacos , Ratos , Ratos Wistar , ATPase Trocadora de Sódio-Potássio/metabolismo , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo , Distúrbios Congênitos do Ciclo da Ureia/patologiaRESUMO
Hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is an autosomal recessive disorder caused by a defect in the mitochondrial ornithine transporter, leading to accumulation of ornithine (Orn), homocitrulline (Hcit) and ammonia. Progressive neurological regression whose pathogenesis is not well established is common in this disease. The present work investigated the in vivo effects of intracerebroventricular administration of Orn and Hcit on important parameters of oxidative stress and energy metabolism in cerebral cortex from young rats. Orn and Hcit significantly increased thiobarbituric acid-reactive substances values and carbonyl formation, indicators of lipid and protein oxidative damage, respectively. Furthermore, N-acetylcysteine and the combination of the free radical scavengers ascorbic acid plus α-tocopherol attenuated the lipid oxidation and totally prevented the protein oxidative damage provoked by Orn and Hcit, suggesting that reactive species were involved in these effects. Hcit, but not Orn administration, also decreased glutathione concentrations, as well as the activity of catalase and glutathione peroxidase, indicating that Hcit provokes a reduction of brain antioxidant defenses. As regards to the parameters of energy metabolism, we verified that Orn and Hcit significantly inhibited the citric acid cycle function (inhibition of CO(2) synthesis from [1-(14)C] acetate), the aerobic glycolytic pathway (reduced CO(2) production from [U-(14)C] glucose) and complex I-III activity of the respiratory chain. Hcit also inhibited the activity of aconitase, an enzyme very susceptible to free radical attack. Taken together, our data indicate that mitochondrial homeostasis is disturbed by Orn and especially by Hcit. It is presumed that the impairment of brain bioenergetics and the oxidative damage induced by these metabolites may possibly contribute to the brain deterioration and neurological symptoms affecting patients with HHH syndrome.