RESUMEN
BACKGROUND: Enoyl-CoA hydratase short-chain 1 (ECHS1) is an enzyme involved in the metabolism of branched chain amino acids and fatty acids. Mutations in the ECHS1 gene lead to mitochondrial short-chain enoyl-CoA hydratase 1 deficiency, resulting in the accumulation of intermediates of valine. This is one of the most common causative genes in mitochondrial diseases. While genetic analysis studies have diagnosed numerous cases with ECHS1 variants, the increasing number of variants of uncertain significance (VUS) in genetic diagnosis is a major problem. METHODS: Here, we constructed an assay system to verify VUS function for ECHS1 gene. A high-throughput assay using ECHS1 knockout cells was performed to index these phenotypes by expressing cDNAs containing VUS. In parallel with the VUS validation system, a genetic analysis of samples from patients with mitochondrial disease was performed. The effect on gene expression in cases was verified by RNA-seq and proteome analysis. RESULTS: The functional validation of VUS identified novel variants causing loss of ECHS1 function. The VUS validation system also revealed the effect of the VUS in the compound heterozygous state and provided a new methodology for variant interpretation. Moreover, we performed multiomics analysis and identified a synonymous substitution p.P163= that results in splicing abnormality. The multiomics analysis complemented the diagnosis of some cases that could not be diagnosed by the VUS validation system. CONCLUSIONS: In summary, this study uncovered new ECHS1 cases based on VUS validation and omics analysis; these analyses are applicable to the functional evaluation of other genes associated with mitochondrial disease.
Asunto(s)
Enfermedades Mitocondriales , Humanos , Fenotipo , Enfermedades Mitocondriales/diagnóstico , Enfermedades Mitocondriales/genética , Mutación/genética , Enoil-CoA Hidratasa/genética , Enoil-CoA Hidratasa/metabolismo , Pruebas GenéticasRESUMEN
BACKGROUND: Ferroptosis, nonapoptotic cell death mediated by free radical reactions and driven by the oxidative degradation of lipids, is a therapeutic target because of its role in organ damage, including AKI. Ferroptosis-causing radicals that are targeted by ferroptosis suppressors have not been unequivocally identified. Because certain cytochrome P450 substrate drugs can prevent lipid peroxidation via obscure mechanisms, we evaluated their antiferroptotic potential and used them to identify ferroptosis-causing radicals. METHODS: Using a cell-based assay, we screened cytochrome P450 substrate compounds to identify drugs with antiferroptotic activity and investigated the underlying mechanism. To evaluate radical-scavenging activity, we used electron paramagnetic resonance-spin trapping methods and a fluorescence probe for lipid radicals, NBD-Pen, that we had developed. We then assessed the therapeutic potency of these drugs in mouse models of cisplatin-induced AKI and LPS/galactosamine-induced liver injury. RESULTS: We identified various US Food and Drug Administration-approved drugs and hormones that have antiferroptotic properties, including rifampicin, promethazine, omeprazole, indole-3-carbinol, carvedilol, propranolol, estradiol, and thyroid hormones. The antiferroptotic drug effects were closely associated with the scavenging of lipid peroxyl radicals but not significantly related to interactions with other radicals. The elevated lipid peroxyl radical levels were associated with ferroptosis onset, and known ferroptosis suppressors, such as ferrostatin-1, also functioned as lipid peroxyl radical scavengers. The drugs exerted antiferroptotic activities in various cell types, including tubules, podocytes, and renal fibroblasts. Moreover, in mice, the drugs ameliorated AKI and liver injury, with suppression of tissue lipid peroxidation and decreased cell death. CONCLUSIONS: Although elevated lipid peroxyl radical levels can trigger ferroptosis onset, some drugs that scavenge lipid peroxyl radicals can help control ferroptosis-related disorders, including AKI.
Asunto(s)
Lesión Renal Aguda/prevención & control , Reposicionamiento de Medicamentos , Ferroptosis/efectos de los fármacos , Depuradores de Radicales Libres/farmacología , Peroxidación de Lípido/efectos de los fármacos , Animales , Células Cultivadas , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , RatasRESUMEN
BACKGROUND: Cardiorenal syndrome is a major cause of mortality in patients with chronic kidney disease (CKD). However, the involvement of detrimental humoral mediators in the pathogenesis of cardiorenal syndrome is still controversial. Trimethylamine-N-oxide (TMAO), a hepatic metabolic product of trimethylamine generated from dietary phosphatidylcholine or carnitine derived by the gut microbiota, has been linked directly with progression of cardiovascular disease and renal dysfunction. Thus, targeting TMAO may be a novel strategy for the prevention of cardiovascular disease and chronic kidney disease. METHODS: Linaclotide, a guanylate cyclase C agonist, was administered to adenine-induced renal failure (RF) mice and changes in renal function and levels of gut-derived uremic toxins, as well as the gut microbiota community, were analyzed using metabolomic and metagenomic methods to reveal its cardiorenal effect. RESULTS: Linaclotide decreased the plasma levels of TMAO at a clinically used low dose of 10 µg/kg in the adenine-induced RF mouse model. At a high concentration of 100 µg/kg, linaclotide clearly improved renal function and reduced the levels of various uremic toxins. A reduction in TMAO levels following linaclotide treatment was also observed in a choline-fed pro-atherosclerotic model. Linaclotide ameliorated renal inflammation and fibrosis and cardiac fibrosis, as well as decreased the expression of collagen I, transforming growth factor-ß, galectin-3 (Gal-3) and ST2 genes. Plasma levels of Gal-3 and ST2 were also reduced. Because exposure of cardiomyocytes to TMAO increased fibronectin expression, these data suggest that linaclotide reduced the levels of TMAO and various uremic toxins and may result in not only renal, but also cardiac, fibrosis. F4/80-positive macrophages were abundant in small intestinal crypts in RF mice, and this increased expression was decreased by linaclotide. Reduced colonic claudin-1 levels were also restored by linaclotide, suggesting that linaclotide ameliorated the 'leaky gut' in RF mice. Metagenomic analysis revealed that the microbial order Clostridiales could be responsible for the change in TMAO levels. CONCLUSION: Linaclotide reduced TMAO and uremic toxin levels and could be a powerful tool for the prevention and control of the cardiorenal syndrome by modification of the gut-cardio-renal axis.
Asunto(s)
Adenina/toxicidad , Síndrome Cardiorrenal/tratamiento farmacológico , Microbioma Gastrointestinal/efectos de los fármacos , Guanilato Ciclasa/química , Agonistas de la Guanilato Ciclasa C/farmacología , Péptidos/farmacología , Insuficiencia Renal Crónica/tratamiento farmacológico , Animales , Síndrome Cardiorrenal/inducido químicamente , Síndrome Cardiorrenal/metabolismo , Síndrome Cardiorrenal/patología , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Fibrosis/inducido químicamente , Fibrosis/tratamiento farmacológico , Fibrosis/metabolismo , Fibrosis/patología , Inflamación/inducido químicamente , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Inflamación/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Insuficiencia Renal Crónica/inducido químicamente , Insuficiencia Renal Crónica/metabolismo , Insuficiencia Renal Crónica/patologíaRESUMEN
Mitochondrial dysfunction causes increased oxidative stress and depletion of ATP, which are involved in the etiology of a variety of renal diseases, such as CKD, AKI, and steroid-resistant nephrotic syndrome. Antioxidant therapies are being investigated, but clinical outcomes have yet to be determined. Recently, we reported that a newly synthesized indole derivative, mitochonic acid 5 (MA-5), increases cellular ATP level and survival of fibroblasts from patients with mitochondrial disease. MA-5 modulates mitochondrial ATP synthesis independently of oxidative phosphorylation and the electron transport chain. Here, we further investigated the mechanism of action for MA-5. Administration of MA-5 to an ischemia-reperfusion injury model and a cisplatin-induced nephropathy model improved renal function. In in vitro bioenergetic studies, MA-5 facilitated ATP production and reduced the level of mitochondrial reactive oxygen species (ROS) without affecting activity of mitochondrial complexes I-IV. Additional assays revealed that MA-5 targets the mitochondrial protein mitofilin at the crista junction of the inner membrane. In Hep3B cells, overexpression of mitofilin increased the basal ATP level, and treatment with MA-5 amplified this effect. In a unique mitochondrial disease model (Mitomice with mitochondrial DNA deletion that mimics typical human mitochondrial disease phenotype), MA-5 improved the reduced cardiac and renal mitochondrial respiration and seemed to prolong survival, although statistical analysis of survival times could not be conducted. These results suggest that MA-5 functions in a manner differing from that of antioxidant therapy and could be a novel therapeutic drug for the treatment of cardiac and renal diseases associated with mitochondrial dysfunction.
Asunto(s)
Ácidos Indolacéticos/farmacología , Túbulos Renales/citología , Mitocondrias/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Miocitos del Músculo Liso/efectos de los fármacos , Fenilbutiratos/farmacología , Animales , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
Mitochondria are key organelles implicated in a variety of processes related to energy and free radical generation, the regulation of apoptosis, and various signaling pathways. Mitochondrial dysfunction increases cellular oxidative stress and depletes ATP in a variety of inherited mitochondrial diseases and also in many other metabolic and neurodegenerative diseases. Mitochondrial diseases are characterized by the dysfunction of the mitochondrial respiratory chain, caused by mutations in the genes encoded by either nuclear DNA or mitochondrial DNA. We have hypothesized that chemicals that increase the cellular ATP levels may ameliorate the mitochondrial dysfunction seen in mitochondrial diseases. To search for the potential drugs for mitochondrial diseases, we screened an in-house chemical library of indole-3-acetic-acid analogs by measuring the cellular ATP levels in Hep3B human hepatocellular carcinoma cells. We have thus identified mitochonic acid 5 (MA-5), 4-(2,4-difluorophenyl)-2-(1H-indol-3-yl)-4-oxobutanoic acid, as a potential drug for enhancing ATP production. MA-5 is a newly synthesized derivative of the plant hormone, indole-3-acetic acid. Importantly, MA-5 improved the survival of fibroblasts established from patients with mitochondrial diseases under the stress-induced condition, including Leigh syndrome, MELAS (myopathy encephalopathy lactic acidosis and stroke-like episodes), Leber's hereditary optic neuropathy, and Kearns-Sayre syndrome. The improved survival was associated with the increased cellular ATP levels. Moreover, MA-5 increased the survival of mitochondrial disease fibroblasts even under the inhibition of the oxidative phosphorylation or the electron transport chain. These data suggest that MA-5 could be a therapeutic drug for mitochondrial diseases that exerts its effect in a manner different from anti-oxidant therapy.
Asunto(s)
Adenosina Trifosfato/metabolismo , Supervivencia Celular/efectos de los fármacos , Descubrimiento de Drogas , Fibroblastos/efectos de los fármacos , Ácidos Indolacéticos/química , Ácidos Indolacéticos/farmacología , Enfermedades Mitocondriales/tratamiento farmacológico , Fenilbutiratos/farmacología , Análisis de Varianza , Línea Celular Tumoral , Supervivencia Celular/fisiología , Fibroblastos/fisiología , Humanos , Fosforilación Oxidativa , Fenilbutiratos/química , Bibliotecas de Moléculas PequeñasRESUMEN
MRPS23 is a nuclear gene encoding a mitochondrial ribosomal protein. A patient with a mitochondrial disorder was found to carry a variant in MRPS23. More cases are necessary to establish MRPS23 as a mitochondrial disease gene. Of 5134 exomes performed in our center, we identified five independent patients who had similar clinical manifestations and were homozygous for the same germline variant c.119C>T; p.P40L in MRPS23. Detailed clinical findings, mitochondrial enzyme activity assays from cultured skin fibroblasts, PCR-Sanger-sequencing, and variant age estimation were performed. Their available family members were also studied. Eight members homozygous for the MRPS23 p.P40L were identified. All were from Hmong hilltribe. Seven presented with alteration of consciousness and recurrent vomiting, while the eighth who was a younger brother of a proband was found pre-symptomatically. Patients showed delayed growth and development, hearing impairment, hypoglycemia, lactic acidosis, and liver dysfunction. In vitro assays of cultured fibroblasts showed combined respiratory chain complex deficiency with low activities of complexes I and IV. PCR-Sanger-sequencing confirmed the variant, which was estimated to have occurred 1550 years ago. These results establish the MRPS23-associated mitochondrial disorder inherited in an autosomal recessive pattern and provide insight into its clinical and metabolic features.
Asunto(s)
Acidosis Láctica , Enfermedades Mitocondriales , Masculino , Humanos , Enfermedades Mitocondriales/genética , Mitocondrias/genética , Mitocondrias/metabolismo , Proteínas Ribosómicas/genética , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Acidosis Láctica/genéticaRESUMEN
Childhood cerebral ALD is a rapidly progressive and neurodegenerative disorder for which HSCT is the curative therapy if carried out at early stages. We successfully treated two patients of childhood cerebral ALD by CBT with RIC. The proband was a seven-yr-old boy whose brain MRI severity score (Loes score) was 14.5. Unrelated CBT was performed in five wk. To minimize conditioning regimen-related neurotoxicity, the combination of fludarabine (125 mg/m(2)), melphalan (140 mg/m(2)), and 4 Gy of brain-sparing TBI was used. The second patient was a six-yr-old brother of the proband. Four wk after the detection of a single small lesion (Loes score 1), he received unrelated CBT with the same RIC as the proband. In both patients, the engraftment was fast and stable, and severe complications were not observed. Furthermore, gadolinium-enhanced inflammation on brain MRI rapidly disappeared after CBT. Now, 20 and 13 months have passed after CBT, respectively, and both patients are neurologically stable. The RIC we used was sufficient for stable engraftment of cord blood and also tolerable even to the patient with advanced ALD. RIC-CBT should be considered for the patients with cerebral ALD at advanced stages, as well as those at early stages.
Asunto(s)
Adrenoleucodistrofia/cirugía , Trasplante de Células Madre de Sangre del Cordón Umbilical , Acondicionamiento Pretrasplante/métodos , Adrenoleucodistrofia/diagnóstico , Encéfalo/patología , Niño , Humanos , Masculino , Melfalán/administración & dosificación , Agonistas Mieloablativos/administración & dosificación , Índice de Severidad de la Enfermedad , Vidarabina/administración & dosificación , Vidarabina/análogos & derivados , Irradiación Corporal TotalRESUMEN
The m.14453G > A mutation in MT-ND6 has been described in a few patients with mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes or Leigh syndrome.However, the clinical spectrum and molecular characteristics are unclear.Here, we present four infantile-onset patients with m.14453G > A-associated Leigh syndrome. All four patients had brainstem lesions with basal ganglia lesions, and two patients had cardiac manifestations. Decreased ND6 protein expression and immunoreactivity were observed in patient-derived samples. There was no clear correlation between heteroplasmy levels and onset age or between heteroplasmy levels and phenotype; however, infantile onset was associated with Leigh syndrome.
Asunto(s)
Enfermedad de Leigh , Encefalomiopatías Mitocondriales , ADN Mitocondrial/genética , Heteroplasmia , Humanos , Enfermedad de Leigh/genética , Mutación , ProbabilidadRESUMEN
BACKGROUND: Hypophosphatasia (HPP) is a rare inherited metabolic disorder caused by mutations in the ALPL gene, which encodes tissue nonspecific alkaline phosphatase. The severity of HPP is widely diverse from the perinatal form to the adult mild form. The former represents the most severe form and was earlier associated with high mortality due to pneumonia which was caused by severe hypomineralization of the bones-such as chest deformity and fractured ribs-and muscle weakness. Enzyme replacement therapy using asfotase alfa (AA) was approved in 2015 in Japan for treating patients with HPP and has improved their pulmonary function and life prognosis. There are several practical and ethical challenges related to using orphan drugs for a rare disorder in a publicly funded healthcare system. Sharing experiences about their application is essential towards formulating guidelines to assist clinicians with decisions about their initiation and withdrawal. We report the details of AA experience in ten cases of pediatric-onset HPP in nine families from January 2015 to November 2019 (median [interquartile range] age 11.0 [7.6-12.5] years; 60% male). This is a study of a single-center cohort describing the clinical course of patients with HPP, mainly consisting of the mild childhood form of HPP, treated with AA in Japan. RESULTS: One case of perinatal form of HPP, two cases of benign prenatal form, and seven cases of childhood form were observed. The most common symptom at onset was pain. All patients had low serum alkaline phosphatase levels as compared to the age-matched reference range before the commencement of AA. All HPP patients seem to have responded to AA treatment, as evidenced by pain alleviation, increased height standard deviation, improvement in respiratory condition and 6-min walk test result improvement, disappearance of kidney calcification, alleviation of fatigue, and/or increases in bone mineralization. There were no serious adverse events, but all patients had an injection site reaction and skin changes at the injection sites. Genetic analysis showed that eight out of ten patients had compound heterozygosity. CONCLUSIONS: AA may be effective in patients with mild to severe pediatric-onset forms of HPP.
Asunto(s)
Hipofosfatasia , Adulto , Fosfatasa Alcalina/genética , Niño , Femenino , Humanos , Hipofosfatasia/complicaciones , Hipofosfatasia/tratamiento farmacológico , Inmunoglobulina G , Japón , Masculino , Dolor/tratamiento farmacológico , Enfermedades Raras/tratamiento farmacológico , Proteínas Recombinantes de FusiónRESUMEN
OBJECTIVE: Neonatal-onset mitochondrial disease has not been fully characterised owing to its heterogeneity. We analysed neonatal-onset mitochondrial disease in Japan to clarify its clinical features, molecular diagnosis and prognosis. DESIGN: Retrospective observational study from January 2004 to March 2020. SETTING: Population based. PATIENTS: Patients (281) with neonatal-onset mitochondrial disease diagnosed by biochemical and genetic approaches. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Disease types, initial symptoms, biochemical findings, molecular diagnosis and prognosis. RESULTS: Of the 281 patients, multisystem mitochondrial disease was found in 194, Leigh syndrome in 26, cardiomyopathy in 38 and hepatopathy in 23 patients. Of the 321 initial symptoms, 236 occurred within 2 days of birth. Using biochemical approaches, 182 patients were diagnosed by mitochondrial respiratory chain enzyme activity rate and 89 by oxygen consumption rate. The remaining 10 patients were diagnosed using a genetic approach. Genetic analysis revealed 69 patients had nuclear DNA variants in 36 genes, 11 of 15 patients had mitochondrial DNA variants in five genes and four patients had single large deletion. The Cox proportional hazards regression analysis showed the effects of Leigh syndrome (HR=0.15, 95% CI 0.04 to 0.63, p=0.010) and molecular diagnosis (HR=1.87, 95% CI 1.18 to 2.96, p=0.008) on survival. CONCLUSIONS: Neonatal-onset mitochondrial disease has a heterogenous aetiology. The number of diagnoses can be increased, and clarity regarding prognosis can be achieved by comprehensive biochemical and molecular analyses using appropriate tissue samples.
Asunto(s)
Enfermedad de Leigh , Enfermedades Mitocondriales , ADN Mitocondrial/genética , Humanos , Recién Nacido , Enfermedad de Leigh/diagnóstico , Enfermedad de Leigh/genética , Enfermedades Mitocondriales/diagnóstico , Enfermedades Mitocondriales/genética , Mutación , PronósticoRESUMEN
BACKGROUND: Cardiomyopathy is a risk factor for poor prognosis in pediatric patients with mitochondrial disease. However, other risk factors including genetic factors related to poor prognosis in mitochondrial disease has yet to be fully elucidated. METHODS AND RESULTS: Between January 2004 and September 2019, we enrolled 223 consecutive pediatric mitochondrial disease patients aged <18 years with a confirmed genetic diagnosis, including 114 with nuclear gene mutations, 89 patients with mitochondrial DNA (mtDNA) point mutations, 11 with mtDNA single large-scale deletions and 9 with chromosomal aberrations. Cardiomyopathy at baseline was observed in 46 patients (21%). Hazard ratios (HR) and 95% confidence intervals (CI) were calculated for all-cause mortality. Over a median follow-up of 36 months (12-77), there were 85 deaths (38%). The overall survival rate was significantly lower in patients with cardiomyopathy than in those without (p < 0.001, log-rank test). By multivariable analysis, left ventricular (LV) hypertrophy (HR = 4.6; 95% CI: 2.8-7.3), neonatal onset (HR = 2.9; 95% CI: 1.8-4.5) and chromosomal aberrations (HR = 2.9; 95% CI: 1.3-6.5) were independent predictors of all-cause mortality. Patients with LV hypertrophy with neonatal onset and/or chromosomal aberrations had higher mortality (100% in 21 patients) than those with LV hypertrophy alone (71% in 14 patients). CONCLUSION: In pediatric patients with mitochondrial disease, cardiomyopathy was common (21%) and was associated with increased mortality. LV hypertrophy, neonatal onset and chromosomal aberrations were independent predictors of all-cause mortality. Prognosis is particularly unfavorable if LV hypertrophy is combined with neonatal onset and/or chromosomal aberrations.
Asunto(s)
Cardiomiopatías , Enfermedades Mitocondriales , Cardiomiopatías/diagnóstico , Cardiomiopatías/genética , Niño , Antecedentes Genéticos , Humanos , Hipertrofia Ventricular Izquierda , Recién Nacido , Enfermedades Mitocondriales/diagnóstico , Enfermedades Mitocondriales/epidemiología , Enfermedades Mitocondriales/genética , Pronóstico , Factores de RiesgoRESUMEN
Sodium-dependent glucose cotransporters (SGLTs) have attracted considerable attention as new targets for type 2 diabetes mellitus. In the kidney, SGLT2 is the major glucose uptake transporter in the proximal tubules, and inhibition of SGLT2 in the proximal tubules shows renoprotective effects. On the other hand, SGLT1 plays a role in glucose absorption from the gastrointestinal tract, and the relationship between SGLT1 inhibition in the gut and renal function remains unclear. Here, we examined the effect of SGL5213, a novel and potent intestinal SGLT1 inhibitor, in a renal failure (RF) model. SGL5213 improved renal function and reduced gut-derived uremic toxins (phenyl sulfate and trimethylamine-N-oxide) in an adenine-induced RF model. Histological analysis revealed that SGL5213 ameliorated renal fibrosis and inflammation. SGL5213 also reduced gut inflammation and fibrosis in the ileum, which is a primary target of SGL5213. Examination of the gut microbiota community revealed that the Firmicutes/Bacteroidetes ratio, which suggests gut dysbiosis, was increased in RF and SGL5213 rebalanced the ratio by increasing Bacteroidetes and reducing Firmicutes. At the genus level, Allobaculum (a major component of Erysipelotrichaceae) was significantly increased in the RF group, and this increase was canceled by SGL5213. We also measured the effect of SGL5213 on bacterial phenol-producing enzymes that catalyze tyrosine into phenol, following the reduction of phenyl sulfate, which is a novel marker and a therapeutic target for diabetic kidney disease DKD. We found that the enzyme inhibition was less potent, suggesting that the change in the microbial community and the reduction of uremic toxins may be related to the renoprotective effect of SGL5213. Because SGL5213 is a low-absorbable SGLT1 inhibitor, these data suggest that the gastrointestinal inhibition of SGLT1 is also a target for chronic kidney diseases.
Asunto(s)
Adenina/toxicidad , Microbioma Gastrointestinal/efectos de los fármacos , Insuficiencia Renal/inducido químicamente , Insuficiencia Renal/tratamiento farmacológico , Transportador 1 de Sodio-Glucosa/antagonistas & inhibidores , Sorbitol/análogos & derivados , Animales , Glucemia/efectos de los fármacos , Glucemia/metabolismo , Microbioma Gastrointestinal/fisiología , Ratones , Ratones Endogámicos C57BL , Insuficiencia Renal/metabolismo , Sorbitol/farmacología , Sorbitol/uso terapéuticoRESUMEN
Unlike complete deficiency of hypoxanthine phosphoribosyltransferase (HPRT) (i.e., Lesch-Nyhan syndrome), partial HPRT deficiency causes HPRT-related hyperuricemia without neurological symptoms. Herein, we describe a 22-year-old man without neurological symptoms that presented gout, hyperuricemia (serum urate level, 12.2 mg/dL), multiple renal microcalculi, and a family history of juvenile gout that was exhibited by his brother and grandfather. Genetic testing revealed a novel missense mutation, c.103G>A (p.V35M), in the HPRT1 gene, and biochemical testing (conducted using the patient's erythrocytes) showed that the patient retained only 12.4% HPRT enzymatic activity compared to that exhibited by a healthy control subject. We thus diagnosed the patient with HPRT-related hyperuricemia caused by partial HPRT deficiency. After his serum urate level was controlled via treatment with febuxostat, his gout did not recur. Thus, this study emphasizes that HPRT deficiency should be considered as a potential cause of familial juvenile gout, even in the absence of neurological symptoms.
Asunto(s)
Gota/genética , Hiperuricemia/genética , Hipoxantina Fosforribosiltransferasa/deficiencia , Hipoxantina Fosforribosiltransferasa/genética , Enfermedades Renales/genética , Febuxostat/administración & dosificación , Febuxostat/uso terapéutico , Gota/complicaciones , Gota/diagnóstico , Gota/tratamiento farmacológico , Supresores de la Gota/administración & dosificación , Supresores de la Gota/uso terapéutico , Humanos , Hiperuricemia/sangre , Hiperuricemia/diagnóstico , Hiperuricemia/tratamiento farmacológico , Hiperuricemia/etiología , Cálculos Renales/diagnóstico , Cálculos Renales/etiología , Cálculos Renales/patología , Enfermedades Renales/diagnóstico , Enfermedades Renales/tratamiento farmacológico , Masculino , Mutación Missense/genética , Resultado del Tratamiento , Adulto JovenRESUMEN
The detoxification of ammonia to urea requires a functional hepatic urea cycle, which consists of six enzymes and two mitochondrial membrane transporters. The initial step of the urea cycle is catalyzed by carbamyl phosphate synthetase 1 (CPS1). CPS1 deficiency (CPS1D) is a rare autosomal recessive disorder. N-Carbamylglutamate (NCG), a deacylase-resistant analogue of N-acetylglutamate, can activate CPS1. We describe the therapeutic course of a patient suffering from neonatal onset CPS1D with compound heterozygosity for the c.2359C > T (p.Arg787*) and c.3559G > T (p.Val1187Phe) variants in CPS1, treated with NCG. She presented with hyperammonemia, which reached 944 µmol/L at the age of 2 days. The ammonia concentration decreased after treatment with continuous hemodiafiltration, NCG, sodium benzoate, sodium phenylbutyrate, L-arginine, vitamin cocktail (vitamin B1, vitamin B12, vitamin C, vitamin E, biotin), l-carnitine, coenzyme Q10, and parenteral nutrition. Her ammonia and glutamine levels remained low; thus, protein intake was increased to 1.2 g/kg/day. Furthermore, the amount of sodium benzoate and sodium phenylbutyrate were reduced. She remained metabolically stable and experienced no metabolic crisis following treatment with oral NCG, sodium benzoate, sodium phenylbutyrate, citrulline, vitamin cocktail, l-carnitine, and coenzyme Q10 until she underwent liver transplantation at 207 days of age. She had no neurological complications at the age of 15 months. Ammonia and glutamine levels of the patient were successfully maintained at a low level via NCG treatment with increased protein intake, which led to normal neurological development. Thus, undiagnosed urea cycle disorders should be treated rapidly with acute therapy including NCG, which should be maintained until a genetic diagnosis is reached. It is essential to prevent metabolic crises in patients with CPS1D until liver transplantation to improve their prognoses.
RESUMEN
Sporadic inclusion body myositis (sIBM) is the most common idiopathic inflammatory myopathy, and several reports have suggested that mitochondrial abnormalities are involved in its etiology. We recruited 9 sIBM patients and found significant histological changes and an elevation of growth differential factor 15 (GDF15), a marker of mitochondrial disease, strongly suggesting the involvement of mitochondrial dysfunction. Bioenergetic analysis of sIBM patient myoblasts revealed impaired mitochondrial function. Decreased ATP production, reduced mitochondrial size and reduced mitochondrial dynamics were also observed in sIBM myoblasts. Cell vulnerability to oxidative stress also suggested the existence of mitochondrial dysfunction. Mitochonic acid-5 (MA-5) increased the cellular ATP level, reduced mitochondrial ROS, and provided protection against sIBM myoblast death. MA-5 also improved the survival of sIBM skin fibroblasts as well as mitochondrial morphology and dynamics in these cells. The reduction in the gene expression levels of Opa1 and Drp1 was also reversed by MA-5, suggesting the modification of the fusion/fission process. These data suggest that MA-5 may provide an alternative therapeutic strategy for treating not only mitochondrial diseases but also sIBM.
Asunto(s)
Ácidos Indolacéticos/uso terapéutico , Mitocondrias Musculares/metabolismo , Miositis por Cuerpos de Inclusión/tratamiento farmacológico , Fenilbutiratos/uso terapéutico , Adenosina Trifosfato/biosíntesis , Anciano , Anciano de 80 o más Años , Butionina Sulfoximina/farmacología , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , ADN Mitocondrial/genética , Evaluación Preclínica de Medicamentos , Dinaminas/biosíntesis , Dinaminas/genética , Femenino , Factores de Crecimiento de Fibroblastos/sangre , Fibroblastos/efectos de los fármacos , GTP Fosfohidrolasas/biosíntesis , GTP Fosfohidrolasas/genética , Factor 15 de Diferenciación de Crecimiento/biosíntesis , Factor 15 de Diferenciación de Crecimiento/sangre , Factor 15 de Diferenciación de Crecimiento/genética , Humanos , Ácidos Indolacéticos/farmacología , Masculino , Persona de Mediana Edad , Mitocondrias Musculares/patología , Mioblastos/efectos de los fármacos , Mioblastos/metabolismo , Mioblastos/ultraestructura , Miositis por Cuerpos de Inclusión/metabolismo , Miositis por Cuerpos de Inclusión/patología , Consumo de Oxígeno , Fenilbutiratos/farmacología , Especies Reactivas de Oxígeno/metabolismo , Estudios RetrospectivosRESUMEN
Mitochondrial disease is a genetic disorder in which individuals suffer from energy insufficiency. The various clinical phenotypes of mitochondrial disease include Leigh syndrome (LS), myopathy encephalopathy lactic acidosis and stroke-like episodes (MELAS). Thus far, no curative treatment is available, and effective treatment options are eagerly awaited. We examined the cell protective effect of an existing commercially available chemical library on fibroblasts from four patients with LS and MELAS and identified apomorphine as a potential therapeutic drug for mitochondrial disease. We conducted a cell viability assay under oxidative stress induced by L-butionine (S, R)-sulfoximine (BSO), a glutathione synthesis inhibitor. Among the chemicals of library, 4 compounds (apomorphine, olanzapine, phenothiazine and ethopropazine) rescued cells from death induced by oxidative stress much more effectively than idebenone, which was used as a positive control. The EC50 value showed that apomorphine was the most effective compound. Apomorphine also significantly improved all of the assessed oxygen consumption rate values by the extracellular flux analyzer for fibroblasts from LS patients with complex I deficiency. In addition, the elevation of the Growth Differentiation Factor-15 (GDF-15), a biomarker of mitochondrial disease, was significantly reduced by apomorphine. Among 441 apomorphine-responsive genes identified by the microarray, apomorphine induced the expression of genes that inhibit the mammalian target of rapamycin (mTOR) activity and inflammatory responses, suggesting that apomorphine induced cell survival via a new potential pathway. In conclusion, apomorphine rescued fibroblasts from cell death under oxidative stress and improved the mitochondrial respiratory activity and appears to be potentially useful for treating mitochondrial disease.
Asunto(s)
Apomorfina/farmacología , Apoptosis/efectos de los fármacos , Fibroblastos/metabolismo , Enfermedad de Leigh/metabolismo , Síndrome MELAS/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Adolescente , Adulto , Biomarcadores/metabolismo , Preescolar , Femenino , Fibroblastos/patología , Factor 15 de Diferenciación de Crecimiento/metabolismo , Humanos , Recién Nacido , Enfermedad de Leigh/tratamiento farmacológico , Enfermedad de Leigh/patología , Síndrome MELAS/tratamiento farmacológico , Síndrome MELAS/patología , Masculino , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
Diabetic kidney disease is a major cause of renal failure that urgently necessitates a breakthrough in disease management. Here we show using untargeted metabolomics that levels of phenyl sulfate, a gut microbiota-derived metabolite, increase with the progression of diabetes in rats overexpressing human uremic toxin transporter SLCO4C1 in the kidney, and are decreased in rats with limited proteinuria. In experimental models of diabetes, phenyl sulfate administration induces albuminuria and podocyte damage. In a diabetic patient cohort, phenyl sulfate levels significantly correlate with basal and predicted 2-year progression of albuminuria in patients with microalbuminuria. Inhibition of tyrosine phenol-lyase, a bacterial enzyme responsible for the synthesis of phenol from dietary tyrosine before it is metabolized into phenyl sulfate in the liver, reduces albuminuria in diabetic mice. Together, our results suggest that phenyl sulfate contributes to albuminuria and could be used as a disease marker and future therapeutic target in diabetic kidney disease.
Asunto(s)
Albuminuria/etiología , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Tipo 1/complicaciones , Diabetes Mellitus Tipo 2/complicaciones , Nefropatías Diabéticas/sangre , Microbioma Gastrointestinal/fisiología , Ésteres del Ácido Sulfúrico/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Albuminuria/sangre , Albuminuria/tratamiento farmacológico , Albuminuria/patología , Animales , Animales Modificados Genéticamente , Estudios de Cohortes , Diabetes Mellitus Experimental/sangre , Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Experimental/orina , Diabetes Mellitus Tipo 1/sangre , Diabetes Mellitus Tipo 2/sangre , Nefropatías Diabéticas/etiología , Nefropatías Diabéticas/patología , Perros , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Femenino , Humanos , Células de Riñón Canino Madin Darby , Masculino , Metabolómica/métodos , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Transportadores de Anión Orgánico/genética , Podocitos/metabolismo , Podocitos/patología , Ratas , Estreptozocina/toxicidad , Ésteres del Ácido Sulfúrico/sangre , Tirosina Fenol-Liasa/antagonistas & inhibidores , Tirosina Fenol-Liasa/metabolismo , Adulto JovenRESUMEN
Focal segmental glomerulosclerosis (FSGS) is caused by various etiologies, with mitochondrial dysfunction being one of the causes. FSGS is known to be associated with mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS), which is a subclass of mitochondrial disease. However, it has rarely been reported in other mitochondrial disease subclasses. Here, we reported a 20-year-old man diagnosed with FSGS associated with chronic progressive external ophthalmoplegia (CPEO) due to mitochondrial DNA (mtDNA) 3243A>G mutation. He presented with left ptosis, short stature, mild sensorineural deafness, and cardiac conduction block. A renal biopsy sample showed segmental sclerosis and adhesions between capillaries and Bowman's capsule, indicating FSGS. Electron microscopy demonstrated abnormal aggregated mitochondria in podocytes, and the basement membrane and epithelial cells of Bowman's capsule. Skeletal muscle biopsy also showed accumulation of abnormal mitochondria. mtDNA analysis identified heteroplasmic mtDNA 3243A>G mutation with no large-scale deletions. From these findings, we diagnosed the case as CPEO with multi-organ involvement including FSGS. Our report demonstrates that CPEO, as well as MELAS, can be associated with FSGS. Because mitochondrial disease presents with a variety of clinical symptoms, atypical cases with non-classical manifestations are observed. Thus, mitochondrial disease should be considered as an underlying cause of FSGS with systemic manifestations even with atypical phenotypes.
Asunto(s)
ADN Mitocondrial/genética , Glomeruloesclerosis Focal y Segmentaria/complicaciones , Glomeruloesclerosis Focal y Segmentaria/genética , Mutación , Oftalmoplejía Externa Progresiva Crónica/complicaciones , Oftalmoplejía Externa Progresiva Crónica/genética , Biopsia , Progresión de la Enfermedad , Glomeruloesclerosis Focal y Segmentaria/patología , Humanos , Riñón/patología , Masculino , Músculo Esquelético/patología , Oftalmoplejía Externa Progresiva Crónica/patología , Podocitos/patología , Adulto JovenRESUMEN
Renal fibrosis is closely related to chronic inflammation and is under the control of epigenetic regulations. Because the signaling of transforming growth factor-ß1 (TGF-ß1) and tumor necrosis factor-α (TNF-α) play key roles in progression of renal fibrosis, dual blockade of TGF-ß1 and TNF-α is desired as its therapeutic approach. Here we screened small molecules showing anti-TNF-α activity in the compound library of indole derivatives. 11 out of 41 indole derivatives inhibited the TNF-α effect. Among them, Mitochonic Acid 35 (MA-35), 5-(3, 5-dimethoxybenzyloxy)-3-indoleacetic acid, showed the potent effect. The anti-TNF-α activity was mediated by inhibiting IκB kinase phosphorylation, which attenuated the LPS/GaIN-induced hepatic inflammation in the mice. Additionally, MA-35 concurrently showed an anti-TGF-ß1 effect by inhibiting Smad3 phosphorylation, resulting in the downregulation of TGF-ß1-induced fibrotic gene expression. In unilateral ureter obstructed mouse kidney, which is a renal fibrosis model, MA-35 attenuated renal inflammation and fibrosis with the downregulation of inflammatory cytokines and fibrotic gene expressions. Furthermore, MA-35 inhibited TGF-ß1-induced H3K4me1 histone modification of the fibrotic gene promoter, leading to a decrease in the fibrotic gene expression. MA-35 affects multiple signaling pathways involved in the fibrosis and may recover epigenetic modification; therefore, it could possibly be a novel therapeutic drug for fibrosis.
Asunto(s)
Indoles/farmacología , Enfermedades Renales/metabolismo , Enfermedades Renales/patología , Transducción de Señal/efectos de los fármacos , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Línea Celular , Modelos Animales de Enfermedad , Matriz Extracelular/metabolismo , Fibrosis , Hepatitis/tratamiento farmacológico , Hepatitis/etiología , Hepatitis/metabolismo , Hepatitis/patología , Histonas/metabolismo , Humanos , Quinasa I-kappa B/metabolismo , Enfermedades Renales/tratamiento farmacológico , Enfermedades Renales/etiología , Lipopolisacáridos/efectos adversos , Masculino , Metilación , Ratones , Modelos Biológicos , Fosforilación/efectos de los fármacos , Proteína smad3/metabolismoRESUMEN
Mitochondrial dysfunction increases oxidative stress and depletes ATP in a variety of disorders. Several antioxidant therapies and drugs affecting mitochondrial biogenesis are undergoing investigation, although not all of them have demonstrated favorable effects in the clinic. We recently reported a therapeutic mitochondrial drug mitochonic acid MA-5 (Tohoku J. Exp. Med., 2015). MA-5 increased ATP, rescued mitochondrial disease fibroblasts and prolonged the life span of the disease model "Mitomouse" (JASN, 2016). To investigate the potential of MA-5 on various mitochondrial diseases, we collected 25 cases of fibroblasts from various genetic mutations and cell protective effect of MA-5 and the ATP producing mechanism was examined. 24 out of the 25 patient fibroblasts (96%) were responded to MA-5. Under oxidative stress condition, the GDF-15 was increased and this increase was significantly abrogated by MA-5. The serum GDF-15 elevated in Mitomouse was likewise reduced by MA-5. MA-5 facilitates mitochondrial ATP production and reduces ROS independent of ETC by facilitating ATP synthase oligomerization and supercomplex formation with mitofilin/Mic60. MA-5 reduced mitochondria fragmentation, restores crista shape and dynamics. MA-5 has potential as a drug for the treatment of various mitochondrial diseases. The diagnostic use of GDF-15 will be also useful in a forthcoming MA-5 clinical trial.