RESUMEN
BACKGROUND: Lung sound analysis parameters have been reported to be useful biomarkers for evaluating airway condition. We developed an automatic lung sound analysis software program for infants and children based on lung sound spectral curves of frequency and power by leveraging machine learning (ML) technology. METHODS: To put this software program into clinical practice, in Study 1, the reliability and reproducibility of the software program using data from younger children were examined. In Study 2, the relationship between lung sound parameters and respiratory flow (L/s) was evaluated using data from older children. In Study 3, we conducted a survey using the ATS-DLD questionnaire to evaluate the clinical usefulness. The survey focused on the history of wheezing and allergies, among healthy 3-year-old infants, and then measured lung sounds. The clinical usefulness was evaluated by comparing the questionnaire results with the results of the new lung sound parameters. RESULTS: In Studies 1 and 2, the parameters of the new software program demonstrated excellent reproducibility and reliability, and were not affected by airflow (L/s). In Study 3, infants with a history of wheezing showed lower FAP0 and RPF75p (p < 0.001 and p = 0.025, respectively) and higher PAP0 (p = 0.001) than healthy infants. Furthermore, infants with asthma/asthma-like bronchitis showed lower FAP0 (p = 0.002) and higher PAP0 (p = 0.001) than healthy infants. CONCLUSIONS: Lung sound parameters obtained using the ML algorithm were able to accurately assess the respiratory condition of infants. These parameters are useful for the early detection and intervention of childhood asthma.
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Asma , Ruidos Respiratorios , Programas Informáticos , Humanos , Ruidos Respiratorios/fisiopatología , Asma/fisiopatología , Asma/diagnóstico , Lactante , Masculino , Preescolar , Femenino , Reproducibilidad de los Resultados , Aprendizaje Automático , Encuestas y Cuestionarios , NiñoRESUMEN
Paired box 2 (Pax2) is a transcription factor essential for kidney development and is reactivated in proximal tubular epithelial cells (PTECs) during recovery from kidney injury. However, the role of Pax2 in this process is still unknown. Here the role of Pax2 reactivation during injury was examined in the proliferation of PTECs using an ischemia-reperfusion injury (IRI) mouse model. Kidney proximal tubule-specific Pax2 conditional knockout mice were generated by mating kidney androgen-regulated protein-Cre and Pax2 flox mice. The degree of cell proliferation and fibrosis was assessed and a Pax2 inhibitor (EG1) was used to evaluate the role of Pax2 in the hypoxic condition of cultured PTECs (O2 5%, 24 hours). The number of Pax2-positive cells and Pax2 mRNA increased after IRI. Sirius red staining indicated that the area of interstitial fibrosis was significantly larger in knockout mice 14 days after IRI. The number of Ki-67-positive cells (an index of proliferation) was significantly lower in knockout than in wild-type mice after IRI, whereas the number of TUNEL-positive cells (an index of apoptotic cells) was significantly higher in knockout mice four days after IRI. Expression analyses of cell cycle-related genes showed that cyclin-dependent kinase 4 (CDK4) was significantly less expressed in the Pax2 knockout mice. In vitro data showed that the increase in CDK4 mRNA and protein expression induced by hypoxia was attenuated by EG1. Thus, Pax2 reactivation may be involved in PTEC proliferation by activating CDK4, thereby limiting kidney fibrosis.
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Lesión Renal Aguda , Daño por Reperfusión , Lesión Renal Aguda/patología , Animales , Proliferación Celular , Quinasa 4 Dependiente de la Ciclina/metabolismo , Células Epiteliales/metabolismo , Fibrosis , Riñón/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , ARN Mensajero/metabolismo , Daño por Reperfusión/patologíaRESUMEN
Hyperphosphatemia is a common complication in patients with advanced chronic kidney disease (CKD) as well as an increased risk of cardiovascular mortality; however, the molecular mechanisms of phosphate-mediated kidney injury are largely unknown. Autophagy is a lysosomal degradation system, which plays protective roles against kidney diseases. Here, we studied the role of autophagy in kidney proximal tubular cells (PTECs) during phosphate overload. Temporal cessation of autophagy in drug-induced PTEC-specific autophagy-deficient mice that were fed high phosphate diet induced mild cytosolic swelling and an accumulation of SQSTM1/p62-and ubiquitin-positive protein aggregates in PTECs, indicating that phosphate overload requires enhanced autophagic activity for the degradation of increasing substrate. Morphological and biochemical analysis demonstrated that high phosphate activates mitophagy in PTECs in response to oxidative stress. PTEC-specific autophagy-deficient mice receiving heminephrectomy and autophagy-deficient cultured PTECs exhibited mitochondrial dysfunction, increased reactive oxygen species production, and reduced ATP production in response to phosphate overload, suggesting that high phosphate-induced autophagy counteracts mitochondrial injury and maintains cellular bioenergetics in PTECs. Thus, potentiating autophagic activity could be a therapeutic option for suppressing CKD progression during phosphate overload.
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Autofagia , Riñón/patología , Mitocondrias/patología , Fosfatos/toxicidad , Animales , Autofagia/efectos de los fármacos , Citoprotección , Células Epiteliales/efectos de los fármacos , Células Epiteliales/patología , Túbulos Renales Proximales/patología , Ratones Endogámicos C57BL , Mitocondrias/efectos de los fármacos , MitofagiaRESUMEN
BACKGROUND: Evidence of a protective role of autophagy in kidney diseases has sparked interest in autophagy as a potential therapeutic strategy. However, understanding how the autophagic process is altered in each disorder is critically important in working toward therapeutic applications. METHODS: Using cultured kidney proximal tubule epithelial cells (PTECs) and diabetic mouse models, we investigated how autophagic activity differs in type 1 versus type 2 diabetic nephropathy. We explored nutrient signals regulating starvation-induced autophagy in PTECs and used autophagy-monitoring mice and PTEC-specific autophagy-deficient knockout mice to examine differences in autophagy status and autophagy's role in PTECs in streptozotocin (STZ)-treated type 1 and db/db type 2 diabetic nephropathy. We also examined the effects of rapamycin (an inhibitor of mammalian target of rapamycin [mTOR]) on vulnerability to ischemia-reperfusion injury. RESULTS: Administering insulin or amino acids, but not glucose, suppressed autophagy by activating mTOR signaling. In db/db mice, autophagy induction was suppressed even under starvation; in STZ-treated mice, autophagy was enhanced even under fed conditions but stagnated under starvation due to lysosomal stress. Using knockout mice with diabetes, we found that, in STZ-treated mice, activated autophagy counteracts mitochondrial damage and fibrosis in the kidneys, whereas in db/db mice, autophagic suppression jeopardizes kidney even in the autophagy-competent state. Rapamycin-induced pharmacologic autophagy produced opposite effects on ischemia-reperfusion injury in STZ-treated and db/db mice. CONCLUSIONS: Autophagic activity in PTECs is mainly regulated by insulin. Consequently, autophagic activity differs in types 1 and 2 diabetic nephropathy, which should be considered when developing strategies to treat diabetic nephropathy by modulating autophagy.
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Autofagia/efectos de los fármacos , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Nefropatías Diabéticas/prevención & control , Lisosomas/metabolismo , Sirolimus/farmacología , Aminoácidos/farmacología , Animales , Células Cultivadas , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/fisiopatología , Diabetes Mellitus Tipo 2/fisiopatología , Nefropatías Diabéticas/fisiopatología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Insulina/farmacología , Túbulos Renales Proximales/citología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Sensibilidad y Especificidad , Estreptozocina/farmacologíaRESUMEN
Urinary tract infection is a bacterial infection that commonly occurs in children. Vesicoureteral reflux is a major underlying precursor condition of urinary tract infection, and an important disorder in the field of pediatric urology. Vesicoureteral reflux is sometimes diagnosed postnatally in infants with fetal hydronephrosis diagnosed antenatally. Opinions vary regarding the diagnosis and treatment of vesicoureteral reflux, and diagnostic procedures remain debatable. In terms of medical interventions, options include either follow-up observation in the hope of possible spontaneous resolution of vesicoureteral reflux with growth/development or provision of continuous antibiotic prophylaxis based on patient characteristics (age, presence/absence of febrile urinary tract infection, lower urinary tract dysfunction and constipation). Furthermore, there are various surgical procedures with different indications and rationales. These guidelines, formulated and issued by the Japanese Society of Pediatric Urology to assist medical management of pediatric vesicoureteral reflux, cover the following: epidemiology, clinical practice algorithm for vesicoureteral reflux, syndromes (dysuria with vesicoureteral reflux, and bladder and rectal dysfunction with vesicoureteral reflux), diagnosis, treatment (medical and surgical), secondary vesicoureteral reflux, long-term prognosis and reflux nephropathy. They also provide the definition of bladder and bowel dysfunction, previously unavailable despite their close association with vesicoureteral reflux, and show the usefulness of diagnostic tests, continuous antibiotic prophylaxis and surgical intervention using site markings.
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Hidronefrosis , Infecciones Urinarias , Reflujo Vesicoureteral , Profilaxis Antibiótica , Niño , Humanos , Lactante , Estudios Retrospectivos , Infecciones Urinarias/diagnóstico , Infecciones Urinarias/etiología , Infecciones Urinarias/prevención & control , Reflujo Vesicoureteral/complicaciones , Reflujo Vesicoureteral/diagnóstico , Reflujo Vesicoureteral/terapiaRESUMEN
BACKGROUND: In order to determine the optimal breathing method for childhood lung sound analyses, it is important to study the effect of airflow on the parameters of lung sounds. METHODS: Sixty-one well-controlled children with atopic asthma (median; 12 years) participated. After confirming that there was no wheezing or respiratory symptoms, the lung sound spectrums of the inspiratory flow before and after inhalation of a ß2 stimulant were analyzed. At the same time, their lung function was measured by a spirogram and the forced oscillation technique. RESULTS: Before ß2 agonist inhalation, the area under the entire curve (AT) and 99% frequency (F99) in the lung sound of inspiratory flow around 2.0L/s due to slightly strong breathing were significantly higher than the lung sound of inspiratory flow around 1.0L/s due to rest breathing. However, no marked differences were observed in the lung sound parameters based on the lung sound spectrum. The improvement in the lung sound parameters after ß2 agonist inhalation was clearer at an inspiratory flow around 1.0L/s than that around 2.0L/s. CONCLUSION: The present study showed that changes after ß2 agonist inhalation and the correlation with the lung function parameters were clear during resting breathing. This method may be used for the long-term montoring of children with asthma.
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Asma/diagnóstico , Pruebas de Función Respiratoria , Ruidos Respiratorios , Agonistas de Receptores Adrenérgicos beta 2 , Asma/fisiopatología , Niño , Humanos , Pulmón/fisiopatologíaAsunto(s)
Bronquiolitis Viral , Bronquiolitis , Infecciones por Virus Sincitial Respiratorio , Humanos , Lactante , Broncodilatadores/uso terapéutico , Ruidos Respiratorios/etiología , Bronquiolitis/diagnóstico , Bronquiolitis/tratamiento farmacológico , Pulmón , Administración por Inhalación , Infecciones por Virus Sincitial Respiratorio/tratamiento farmacológicoRESUMEN
Excessive fat intake contributes to the progression of metabolic diseases via cellular injury and inflammation, a process termed lipotoxicity. Here, we investigated the role of lysosomal dysfunction and impaired autophagic flux in the pathogenesis of lipotoxicity in the kidney. In mice, a high-fat diet (HFD) resulted in an accumulation of phospholipids in enlarged lysosomes within kidney proximal tubular cells (PTCs). In isolated PTCs treated with palmitic acid, autophagic degradation activity progressively stagnated in association with impaired lysosomal acidification and excessive lipid accumulation. Pulse-chase experiments revealed that the accumulated lipids originated from cellular membranes. In mice with induced PTC-specific ablation of autophagy, PTCs of HFD-mice exhibited greater accumulation of ubiquitin-positive protein aggregates normally removed by autophagy than did PTCs of mice fed a normal diet. Furthermore, HFD-mice had no capacity to augment autophagic activity upon another pathologic stress. Autophagy ablation also exaggerated HFD-induced mitochondrial dysfunction and inflammasome activation. Moreover, renal ischemia-reperfusion induced greater injury in HFD-mice than in mice fed a normal diet, and ablation of autophagy further exacerbated this effect. Finally, we detected similarly enhanced phospholipid accumulation in enlarged lysosomes and impaired autophagic flux in the kidneys of obese patients compared with nonobese patients. These findings provide key insights regarding the pathophysiology of lipotoxicity in the kidney and clues to a novel treatment for obesity-related kidney diseases.
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Autofagia/fisiología , Dieta Alta en Grasa/efectos adversos , Enfermedades Renales/etiología , Riñón/metabolismo , Metabolismo de los Lípidos , Lisosomas/fisiología , Animales , Autofagia/efectos de los fármacos , Riñón/efectos de los fármacos , Metabolismo de los Lípidos/efectos de los fármacos , Lisosomas/efectos de los fármacos , Masculino , Ratones , Ácido Palmítico/farmacologíaRESUMEN
BACKGROUND: Kawasaki disease is a common systemic vasculitis that leads to coronary artery lesions. Besides its antihypertensive effects, losartan can modulate inflammation in cardiovascular disease. We examined whether losartan can attenuate coronary inflammation in a murine model of Kawasaki disease. METHODS AND RESULTS: Five-wk-old C57/BL6J male mice were intraperitoneally injected with Lactobacillus casei cell wall extract to induce coronary inflammation and divided into four groups: placebo, intravenous immunoglobulin (IVIG), losartan, and IVIG+losartan. After 2 wk, mice were harvested. The coronary perivasculitis was significantly attenuated by losartan but not by IVIG alone, and further dramatic attenuation by IVIG+losartan was observed. The frequency of Lactobacillus casei cell wall extract-induced myocarditis (80%) was markedly lowered by losartan (22%) and IVIG+losartan (0%). Furthermore, interleukin (IL)-6 mRNA was markedly attenuated by IVIG+losartan. Serum levels of IL-6, TNF-α, MCP-1, and IL-10 after Lactobacillus casei cell wall extract injection were slightly decreased by IVIG or losartan. Moreover, IL-1ß, IL-10, and MCP-1 levels were significantly decreased by IVIG+losartan. CONCLUSION: The addition of losartan to IVIG strongly attenuated the severity of coronary perivasculitis and the incidence of myocarditis, along with suppressing systemic/local cytokines as well as the activated macrophage infiltration. Therefore, losartan may be a potentially useful additive drug for the acute phase of Kawasaki disease to minimize coronary artery lesions.
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Antiarrítmicos/uso terapéutico , Antiinflamatorios/uso terapéutico , Losartán/uso terapéutico , Síndrome Mucocutáneo Linfonodular/tratamiento farmacológico , Vasculitis/tratamiento farmacológico , Animales , Pared Celular , Quimiocina CCL2/sangre , Modelos Animales de Enfermedad , Inmunohistoquímica , Inflamación , Infusiones Intravenosas , Interleucina-10/sangre , Interleucina-1beta/sangre , Interleucina-6/sangre , Lacticaseibacillus casei , Macrófagos/citología , Masculino , Ratones , Ratones Endogámicos C57BL , Factor de Necrosis Tumoral alfa/sangreRESUMEN
The renal tissue renin-angiotensin system is activated in chronic kidney diseases. We previously demonstrated that intrarenal ANG II is synthesized primarily from liver-derived angiotensinogen filtered through the glomerulus and that podocyte injury increases the passage of angiotensinogen into the tubular lumen and generation of ANG II. In the present study, we tested the effect of cessation of glomerular filtration by ureteral obstruction on renal ANG II generation in kidneys with podocyte injury under two experimental conditions. Ureteral obstruction is known to activate the renin-angiotensin system in nonproteinuric kidneys. Transgenic mice expressing hCD25 in podocyte (NEP25) were injected with 1.25 or 10 ng/g body wt of LMB2, a hCD25-targeted immunotoxin, subjected to unilateral ureteral ligation on the following day, and euthanized 7 and 4 days later, respectively. In both experiments, compared with the kidney in untreated wild-type mice, renal angiotensinogen protein, as assessed by immunostaining and Western blot analysis, was increased in the contralateral unobstructed kidney. However, it was markedly decreased in the obstructed kidney. Whereas intrarenal ANG II content was increased in the contralateral kidney compared with the untreated kidney (248 ± 83 vs. 106 ± 21 and 298 ± 185 vs. 64.8 ± 20 fmol/g kidney, respectively), this increase was suppressed in the obstructed kidney (161 ± 75 and 113 ± 34 fmol/g kidney, respectively), a pattern opposite to what we expected in obstructed kidneys without podocyte injury. Thus, our study indicates that the major source of increased renal ANG II in podocyte injury is filtered angiotensinogen.
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Angiotensina II/metabolismo , Angiotensinógeno/metabolismo , Enfermedades Renales/metabolismo , Podocitos/metabolismo , Sistema Renina-Angiotensina , Obstrucción Ureteral/metabolismo , Albuminuria/metabolismo , Albuminuria/patología , Albuminuria/fisiopatología , Angiotensinógeno/genética , Animales , Anticuerpos Monoclonales , Modelos Animales de Enfermedad , Regulación hacia Abajo , Exotoxinas , Femenino , Tasa de Filtración Glomerular , Humanos , Subunidad alfa del Receptor de Interleucina-2/genética , Subunidad alfa del Receptor de Interleucina-2/metabolismo , Enfermedades Renales/inducido químicamente , Enfermedades Renales/genética , Enfermedades Renales/patología , Enfermedades Renales/fisiopatología , Ratones Endogámicos C57BL , Ratones Transgénicos , Podocitos/patología , ARN Mensajero/metabolismo , Sistema Renina-Angiotensina/genética , Transducción de Señal , Factores de Tiempo , Obstrucción Ureteral/patología , Obstrucción Ureteral/fisiopatologíaRESUMEN
Metabolic acidosis, a common complication of CKD, causes mitochondrial stress by undefined mechanisms. Selective autophagy of impaired mitochondria, called mitophagy, contributes toward maintaining cellular homeostasis in various settings. We hypothesized that mitophagy is involved in proximal tubular cell adaptations to chronic metabolic acidosis. In transgenic mice expressing green fluorescent protein-tagged microtubule-associated protein 1 light chain 3 (GFP-LC3), NH4Cl loading increased the number of GFP puncta exclusively in the proximal tubule. In vitro, culture in acidic medium produced similar results in proximal tubular cell lines stably expressing GFP-LC3 and facilitated the degradation of SQSTM1/p62 in wild-type cells, indicating enhanced autophagic flux. Upon acid loading, proximal tubule-specific autophagy-deficient (Atg5-deficient) mice displayed significantly reduced ammonium production and severe metabolic acidosis compared with wild-type mice. In vitro and in vivo, acid loading caused Atg5-deficient proximal tubular cells to exhibit reduced mitochondrial respiratory chain activity, reduced mitochondrial membrane potential, and fragmented morphology with marked swelling in mitochondria. GFP-LC3-tagged autophagosomes colocalized with ubiquitinated mitochondria in proximal tubular cells cultured in acidic medium, suggesting that metabolic acidosis induces mitophagy. Furthermore, restoration of Atg5-intact nuclei in Atg5-deficient proximal tubular cells increased mitochondrial membrane potential and ammoniagenesis. In conclusion, metabolic acidosis induces autophagy in proximal tubular cells, which is indispensable for maintaining proper mitochondrial functions including ammoniagenesis, and thus for adapted urinary acid excretion. Our results provide a rationale for the beneficial effect of alkali supplementation in CKD, a condition in which autophagy may be reduced, and suggest a new therapeutic option for acidosis by modulating autophagy.
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Acidosis/fisiopatología , Túbulos Renales Proximales/fisiopatología , Mitofagia , Acidosis/metabolismo , Acidosis/patología , Compuestos de Amonio/metabolismo , Animales , Autofagia , Células Cultivadas , Transporte de Electrón , Femenino , Túbulos Renales Proximales/metabolismo , Túbulos Renales Proximales/patología , Masculino , Potencial de la Membrana Mitocondrial , Ratones TransgénicosRESUMEN
Intrarenal angiotensin II is increased in kidney diseases independently of plasma angiotensin II and is thought to promote progressive deterioration of renal architecture. Here we investigated the mechanism of enhanced renal angiotensin II generation in kidney glomerular diseases. For this, kidney- or liver-specific angiotensinogen gene (Agt) knockout was superimposed on the mouse model of inducible podocyte injury (NEP25). Seven days after induction of podocyte injury, renal angiotensin II was increased ninefold in NEP25 mice with intact Agt, accompanied by increases in urinary albumin and angiotensinogen excretion, renal angiotensinogen protein, and its mRNA. Kidney Agt knockout attenuated renal Agt mRNA but not renal angiotensin II, renal, or urinary angiotensinogen protein. In contrast, liver Agt knockout markedly reduced renal angiotensin II to 18.7% of that of control NEP25 mice, renal and urinary angiotensinogen protein, but not renal Agt mRNA. Renal angiotensin II had no relationship with renal Agt mRNA, or with renal renin mRNA, which was elevated in liver Agt knockouts. Kidney and liver dual Agt knockout mice showed phenotypes comparable to those of liver Agt knockout mice. Thus, increased renal angiotensin II generation upon severe podocyte injury is attributed to increased filtered angiotensinogen of liver origin resulting from loss of macromolecular barrier function of the glomerular capillary wall that occurs upon severe podocyte injury.
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Angiotensina II/metabolismo , Angiotensinógeno/metabolismo , Enfermedades Renales/metabolismo , Túbulos Renales Proximales/metabolismo , Hígado/metabolismo , Podocitos/metabolismo , Albuminuria/genética , Albuminuria/metabolismo , Albuminuria/patología , Angiotensinógeno/deficiencia , Angiotensinógeno/genética , Angiotensinógeno/orina , Animales , Anticuerpos Monoclonales , Modelos Animales de Enfermedad , Exotoxinas , Genotipo , Enfermedades Renales/inducido químicamente , Enfermedades Renales/genética , Enfermedades Renales/patología , Túbulos Renales Proximales/patología , Proteínas de la Membrana/metabolismo , Ratones Noqueados , Fenotipo , Podocitos/patología , ARN Mensajero/metabolismo , Sodio/metabolismo , Factores de Tiempo , Regulación hacia ArribaRESUMEN
Background: It is speculated that the coronavirus disease 2019 (COVID-19) pandemic-associated reduction in the prevalence of respiratory tract infections has influenced the incidence of asthma in young children. Objectives: We investigated an association between the reduction in viral infections and the reduction in asthma in young children. Methods: The subjects were infants born in the early stages of the COVID-19 pandemic in Japan, which began in February 2020. A questionnaire survey related to asthma and allergy was conducted at 18 months and 3 years of age. These results were compared to those of age-matched infants during the nonpandemic period. Results: There were no epidemics of viral infectious diseases until the target child was 18 months old. At 18 months, the incidence of asthma/asthmatic bronchitis diagnosed by physicians in pandemic children was significantly lower than that in nonpandemic children. In 3-year-olds, no marked difference was observed between nonpandemic infants and pandemic children, except for an increase in respiratory syncytial virus infection in pandemic children. In a comparative study of the same children at ages 18 months and 3 years, an increased prevalence of asthma/asthmatic bronchitis was observed in pandemic children. Furthermore, the incidence of asthma after respiratory syncytial virus infection in pandemic infants was significantly lower than that in nonpandemic children. Conclusion: The COVID-19 pandemic-associated reduction in respiratory tract infections may have reduced the incidence of asthma in early childhood, and respiratory syncytial virus infection after 18 months of age had little effect on the onset of asthma. These results indicate the importance of preventing respiratory tract infections in early infancy.
RESUMEN
Autophagy is a highly conserved bulk protein degradation pathway involved in cellular homeostasis. Although emerging evidence indicates involvement of autophagy in various conditions, efforts to clarify the role of autophagy in renal tubules are beginning to be elucidated. In the present study, we examined the hypothesis that autophagy guards against acute kidney injury (AKI) by modulating several deteriorative pathways that lead to tubular cell death using a cisplatin-induced model of AKI. Cisplatin treatment of GFP-LC3 (green fluorescent protein-microtubule-associated protein 1 light chain 3) transgenic mice induced autophagy in kidney proximal tubules in a time-dependent manner. Proximal tubule-specific autophagy-deficient mice exhibited more severe cisplatin-induced AKI than did control mice, as assessed via kidney function and morphologic findings. In addition, cisplatin induced more severe DNA damage and p53 activation, concomitant with an increase in apoptotic cell number, and a massive accumulation of protein aggregates in autophagy-deficient proximal tubules. Cisplatin treatment significantly increased reactive oxygen species-producing damaged mitochondria in immortalized autophagy-deficient proximal tubular cells when compared with autophagy-retrieved control cells. In conclusion, autophagy guards kidney proximal tubules against AKI, possibly by alleviating DNA damage and reactive oxygen species production and by eliminating toxic protein aggregates. Enhancing autophagy may provide a novel therapeutic option to minimize AKI.
Asunto(s)
Lesión Renal Aguda/inducido químicamente , Antineoplásicos/toxicidad , Autofagia/fisiología , Cisplatino/toxicidad , Túbulos Renales Proximales/fisiopatología , Lesión Renal Aguda/fisiopatología , Animales , Apoptosis/fisiología , Daño del ADN/fisiología , Proteínas Fluorescentes Verdes/metabolismo , Masculino , Ratones , Ratones Transgénicos , Proteínas Asociadas a Microtúbulos/metabolismo , Mitocondrias/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Factor de Transcripción TFIIH , Factores de Transcripción/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Ubiquitina/metabolismoRESUMEN
BACKGROUND: Mice carrying the null-mutated Foxc1 gene frequently develop an anomalous double collecting system. These mice provide an ideal opportunity to specify the role of ectopic budding in the development of congenital anomalies of the kidney and urinary tract. METHODS: Tissue specimens were collected from Foxc1(ch/ch) mutants at several embryonic stages and at birth. The upper and lower pole kidneys were qualitatively and quantitatively examined by histology, in situ hybridization and immunohistochemistry. RESULTS: Upper pole kidneys of newborn Foxc1(ch/ch) mice were significantly more hypoplastic and contained significantly fewer glomeruli than their lower pole counterparts. On embryonic day 14.5, the stage immediately before the formation of the first urine, the upper pole kidney was already smaller than the lower pole kidney. Neither histology nor immunostaining for kidney markers showed dysplastic regions in either kidney of newborn Foxc1(ch/ch) mice. Of note, expression of Foxc1 was restricted to maturing podocytes and was not detectable in any intermediate structure of nephron development in the nephrogenic zone. CONCLUSION: Ectopic budding alone results only in kidney hypoplasia but not dysplasia. The development of dysplasticity in the maturing kidney involves gene(s) that function beyond the initial budding stage within the metanephros.
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Factores de Transcripción Forkhead/genética , Riñón/anomalías , Riñón/embriología , Mutación , Animales , Regulación del Desarrollo de la Expresión Génica , Riñón/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
Angiotensin II content in the kidney is much higher than in the plasma, and it increases more in kidney diseases through an uncertain mechanism. Because the kidney abundantly expresses angiotensinogen mRNA, transcriptional dysregulation of angiotensinogen within the kidney is one potential cause of increased renal angiotensin II in the setting of disease. Here, we observed that kidney-specific angiotensinogen knockout mice had levels of renal angiotensinogen protein and angiotensin II that were similar to those levels of control mice. In contrast, liver-specific knockout of angiotensinogen nearly abolished plasma and renal angiotensinogen protein and renal tissue angiotensin II. Immunohistochemical analysis in mosaic proximal tubules of megalin knockout mice revealed that angiotensinogen protein was incorporated selectively in megalin-intact cells of the proximal tubule, indicating that the proximal tubule reabsorbs filtered angiotensinogen through megalin. Disruption of the filtration barrier in a transgenic mouse model of podocyte-selective injury increased renal angiotensin II content and markedly increased both tubular and urinary angiotensinogen protein without an increase in renal renin activity, supporting the dependency of renal angiotensin II generation on filtered angiotensinogen. Taken together, these data suggest that liver-derived angiotensinogen is the primary source of renal angiotensinogen protein and angiotensin II. Furthermore, an abnormal increase in the permeability of the glomerular capillary wall to angiotensinogen, which characterizes proteinuric kidney diseases, enhances the synthesis of renal angiotensin II.
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Angiotensina II/metabolismo , Angiotensinógeno/metabolismo , Riñón/metabolismo , Hígado/metabolismo , Angiotensina II/deficiencia , Angiotensina II/genética , Angiotensinógeno/deficiencia , Angiotensinógeno/genética , Animales , Riñón/patología , Hígado/patología , Proteína 2 Relacionada con Receptor de Lipoproteína de Baja Densidad/deficiencia , Proteína 2 Relacionada con Receptor de Lipoproteína de Baja Densidad/genética , Proteína 2 Relacionada con Receptor de Lipoproteína de Baja Densidad/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Modelos Animales , Fenotipo , ARN Mensajero/metabolismoRESUMEN
Obesity is a major risk factor for end-stage kidney disease. We previously found that lysosomal dysfunction and impaired autophagic flux contribute to lipotoxicity in obesity-related kidney disease, in both humans and experimental animal models. However, the regulatory factors involved in countering renal lipotoxicity are largely unknown. Here, we found that palmitic acid strongly promoted dephosphorylation and nuclear translocation of transcription factor EB (TFEB) by inhibiting the mechanistic target of rapamycin kinase complex 1 pathway in a Rag GTPase-dependent manner, though these effects gradually diminished after extended treatment. We then investigated the role of TFEB in the pathogenesis of obesity-related kidney disease. Proximal tubular epithelial cell-specific (PTEC-specific) Tfeb-deficient mice fed a high-fat diet (HFD) exhibited greater phospholipid accumulation in enlarged lysosomes, which manifested as multilamellar bodies (MLBs). Activated TFEB mediated lysosomal exocytosis of phospholipids, which helped reduce MLB accumulation in PTECs. Furthermore, HFD-fed, PTEC-specific Tfeb-deficient mice showed autophagic stagnation and exacerbated injury upon renal ischemia/reperfusion. Finally, higher body mass index was associated with increased vacuolation and decreased nuclear TFEB in the proximal tubules of patients with chronic kidney disease. These results indicate a critical role of TFEB-mediated lysosomal exocytosis in counteracting renal lipotoxicity.
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Dieta Alta en Grasa , Exocitosis , Lípidos , Insuficiencia Renal Crónica , Animales , Humanos , Ratones , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/metabolismo , Dieta Alta en Grasa/efectos adversos , Exocitosis/genética , Riñón/metabolismo , Riñón/patología , Lípidos/toxicidad , Lisosomas/metabolismo , Obesidad/metabolismo , Insuficiencia Renal Crónica/metabolismoRESUMEN
Autophagy is a bulk protein degradation system that likely plays an important role in normal proximal tubule function and recovery from acute ischemic kidney injury. Using conditional Atg5 gene deletion to eliminate autophagy in the proximal tubule, we determined whether autophagy prevents accumulation of damaged proteins and organelles with aging and ischemic renal injury. Autophagy-deficient cells accumulated deformed mitochondria and cytoplasmic inclusions, leading to cellular hypertrophy and eventual degeneration not observed in wildtype controls. In autophagy-deficient mice, I/R injury increased proximal tubule cell apoptosis with accumulation of p62 and ubiquitin positive cytoplasmic inclusions. Compared with control animals, autophagy-deficient mice exhibited significantly greater elevations in serum urea nitrogen and creatinine. These data suggest that autophagy maintains proximal tubule cell homeostasis and protects against ischemic injury. Enhancing autophagy may provide a novel therapeutic approach to minimize acute kidney injury and slow CKD progression.
Asunto(s)
Lesión Renal Aguda/prevención & control , Autofagia , Túbulos Renales Proximales/patología , Riñón/irrigación sanguínea , Daño por Reperfusión/prevención & control , Animales , Proteína 5 Relacionada con la Autofagia , Hipertrofia , Riñón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Asociadas a Microtúbulos/fisiologíaRESUMEN
Recently, we identified a novel mechanism of lipotoxicity in the kidney proximal tubular cells (PTECs); lipid overload stimulates macroautophagy/autophagy for the renovation of plasma and organelle membranes to maintain the integrity of the PTECs. However, this autophagic activation places a burden on the lysosomal system, leading to a downstream suppression of autophagy, which manifests as phospholipid accumulation and inadequate acidification in lysosomes. Here, we investigated whether pharmacological correction by eicosapentaenoic acid (EPA) supplementation could restore autophagic flux and alleviate renal lipotoxicity. EPA supplementation to high-fat diet (HFD)-fed mice reduced several hallmarks of lipotoxicity in the PTECs, such as phospholipid accumulation in the lysosome, mitochondrial dysfunction, inflammation, and fibrosis. In addition to improving the metabolic syndrome, EPA alleviated renal lipotoxicity via several mechanisms. EPA supplementation to HFD-fed mice or the isolated PTECs cultured in palmitic acid (PA) restored lysosomal function with significant improvements in the autophagic flux. The PA-induced redistribution of phospholipids from cellular membranes into lysosomes and the HFD-induced accumulation of SQSTM1/p62 (sequestosome 1), an autophagy substrate, during the temporal and genetic ablation of autophagy were significantly reduced by EPA, indicating that EPA attenuated the HFD-mediated increases in autophagy demand. Moreover, a fatty acid pulse-chase assay revealed that EPA promoted lipid droplet (LD) formation and transfer from LDs to the mitochondria for beta-oxidation. Noteworthy, the efficacy of EPA on lipotoxicity is autophagy-dependent and cell-intrinsic. In conclusion, EPA counteracts lipotoxicity in the proximal tubule by alleviating autophagic numbness, making it potentially suitable as a novel treatment for obesity-related kidney diseases.Abbreviations: 4-HNE: 4-hydroxy-2-nonenal; ACTB: actin beta; ADGRE1/F4/80: adhesion G protein-coupled receptor E1; ATG: autophagy-related; ATP: adenosine triphosphate; BODIPY: boron-dipyrromethene; BSA: bovine serum albumin; cKO: conditional knockout; CML: N-carboxymethyllysine; COL1A1: collagen type I alpha 1 chain; COX: cytochrome c oxidase; CTRL: control; DGAT: diacylglycerol O-acyltransferase; EPA: eicosapentaenoic acid; FA: fatty acid; FFA: free fatty acid; GFP: green fluorescent protein; HFD: high-fat diet; iKO: inducible knockout; IRI: ischemia-reperfusion injury; LAMP1: lysosomal-associated membrane protein 1; LD: lipid droplet; LRP2: low density lipoprotein receptor-related protein 2; MAP1LC3: microtubule-associated protein 1 light chain 3; MTORC1: mechanistic target of rapamycin kinase complex 1; OA: oleic acid; PAS: periodic-acid Schiff; PPAR: peroxisome proliferator activated receptor; PPARGC1/PGC1: peroxisome proliferator activated receptor, gamma, coactivator 1; PTEC: proximal tubular epithelial cell; ROS: reactive oxygen species; RPS6: ribosomal protein S6; SDH: succinate dehydrogenase complex; SFC/MS/MS: supercritical fluid chromatography triple quadrupole mass spectrometry; SQSTM1/p62: sequestosome 1; TFEB: transcription factor EB; TG: triglyceride; TUNEL: terminal deoxynucleotidyl transferase dUTP nick end labeling.
Asunto(s)
Lesión Renal Aguda/tratamiento farmacológico , Autofagia/efectos de los fármacos , Dieta Alta en Grasa/efectos adversos , Ácido Eicosapentaenoico/farmacología , Ácido Eicosapentaenoico/uso terapéutico , Lesión Renal Aguda/inducido químicamente , Animales , Riñón/efectos de los fármacos , Túbulos Renales Proximales/efectos de los fármacos , Lisosomas/efectos de los fármacos , Ratones , Ratones Transgénicos , Fosfolípidos/metabolismoRESUMEN
OBJECTIVE: Using a commercially available breath sound analyzer, the airway reversibility in asthmatic children during healthy periods was investigated. METHODS: Fifty samples of 34 children with asthma (median age, 11 years; range, 6-16 years) who visited our hospital and whose lung function was normal were included. The breath sound parameters, the frequency limiting 99% of the power spectrum (F99) and spectrum curve indices, the total area under the curve of the dBm data (A3/AT and B4/AT) and the ratio of power and frequency at 50% and 75% of the highest frequency of the power spectrum (RPF75 and RPF50) were evaluated before and after ß2 agonist inhalation. RESULTS: The values of spectrum curve indices were significantly increased after ß2 agonist inhalation. The changes in these parameters were more marked than the changes in the FOT parameters. The changes in A3/AT and B4/AT were significantly related to two FOT parameters: R5-R20 and X5. CONCLUSIONS: Our study suggested that significant changes in breath sound parameters were present in asthmatic children during the period of good control. A breath sound analysis may be useful for assessing the airway condition of asthmatic children during long-term management.