RESUMEN
Background Microalbuminuria is a common clinical symptom that manifests in the early stages of diabetic kidney disease (DKD) and is also the main feature of glomerular endothelial cells (GECs) injury. There is increasing evidence that the transcytosis of albumin across GECs is closely related to the formation of albuminuria. Our previous studies have shown that angiopoietin 2 (ANGPT2) can inhibit albumin transcytosis across renal tubular epithelial cells by activating caveolin 1 (CAV1) phosphorylation during high glucose (HG) exposure. The role of ANGPT2 in albumin transcytosis across GECs remains unclear. Losartan significantly reduces albuminuria, but the mechanism has not been clarified. Methods We established an in vitro albumin transcytosis model to investigate the change in albumin transcytosis across human renal glomerular endothelial cells (hrGECs) under normal glucose (NG), high glucose (HG) and losartan intervention. We knocked down ANGPT2 and CAV1 to evaluate their roles in albumin transcytosis across hrGECs and verified the relationship between them. In vivo, DKD mouse models were established and treated with different doses of losartan. Immunohistochemistry and Western blot were used to detect the expression of ANGPT2 and CAV1. Results In vitro, the transcytosis of albumin across hrGECs was significantly increased under high glucose stimulation, and losartan inhibited this process. The expression of ANGPT2 and CAV1 were both increased in hrGECs under HG conditions and losartan intervention reduced the expression of them. Moreover, ANGPT2 downregulation reduced albumin transcytosis in hrGECs by regulating CAV1 expression. In vivo, the expression of ANGPT2 and CAV1 in the glomerulus was both increased significantly in DKD mice. Compared with DKD mice, losartan treatment reduced albuminuria and decreased the expression of ANGPT2 and CAV1 in a dose-dependent manner (AU)
Antecedentes La microalbuminuria es un síntoma clínico común que se manifiesta en las fases tempranas de la enfermedad renal diabética (ERD), y también es característica del daño de las células endoteliales glomerulares (GEC). Existe evidencia creciente en cuanto a que la transcitosis de la albúmina a través de las GEC está estrechamente relacionada con la formación de albuminuria. Nuestros estudios previos reflejaron que angiopoyetina 2 (ANGPT2) puede inhibir la transcitosis de la albúmina a través de las células epiteliales tubulares renales activando la fosforilación de caveolina 1 (CAV1) durante la exposición a hiperglucemia (HG). El rol de ANGPT2 en la transcitosis de la albúmina a través de las GEC resulta incierto. Losartan reduce considerablemente la albuminuria, aunque no se ha esclarecido el mecanismo. Métodos Establecimos un modelo in vitro de transcitosis de la albúmina para investigar el cambio de dicho mecanismo a través de las células endoteliales glomerulares renales humanas (hrGEC) en condiciones de glucosa normal (GN), hiperglucemia (HG) e intervención de losartan. Realizamos breakdown de ANGPT2 y CAV1 para evaluar sus roles en la transcitosis de la albúmina a través de las hrGEC, y verificamos la relación entre ellas. Se establecieron modelos in vivo de ratones con ERD, tratados con diferentes dosis de losartan. Se utilizaron pruebas de inmunohistoquímica e inmunotransferencia para detectar la expresión de ANGPT2 y CAV1. Resultados In vitro, la transcitosis de la albúmina a través de hrGEC se incrementó considerablemente en condiciones de estimulación de la hiperglucemia, inhibiendo losartan este proceso. La expresión de ANGPT2 y CAV1 se incrementó en las hrGEC en condiciones de HG, reduciendo la intervención de losartan la expresión de ambas (AU)
Asunto(s)
Animales , Masculino , Ratones , Diabetes Mellitus Experimental/metabolismo , Glomérulos Renales/metabolismo , Albúminas/metabolismo , Transcitosis , Angiopoyetinas/metabolismo , Ratones Endogámicos C57BL , Caveolinas/farmacología , Losartán/farmacología , Modelos AnimalesRESUMEN
BACKGROUND: Microalbuminuria is a common clinical symptom that manifests in the early stages of diabetic kidney disease (DKD) and is also the main feature of glomerular endothelial cells (GECs) injury. There is increasing evidence that the transcytosis of albumin across GECs is closely related to the formation of albuminuria. Our previous studies have shown that angiopoietin 2 (ANGPT2) can inhibit albumin transcytosis across renal tubular epithelial cells by activating caveolin 1 (CAV1) phosphorylation during high glucose (HG) exposure. The role of ANGPT2 in albumin transcytosis across GECs remains unclear. Losartan significantly reduces albuminuria, but the mechanism has not been clarified. METHODS: We established an in vitro albumin transcytosis model to investigate the change in albumin transcytosis across human renal glomerular endothelial cells (hrGECs) under normal glucose (NG), high glucose (HG) and losartan intervention. We knocked down ANGPT2 and CAV1 to evaluate their roles in albumin transcytosis across hrGECs and verified the relationship between them. In vivo, DKD mouse models were established and treated with different doses of losartan. Immunohistochemistry and Western blot were used to detect the expression of ANGPT2 and CAV1. RESULTS: In vitro, the transcytosis of albumin across hrGECs was significantly increased under high glucose stimulation, and losartan inhibited this process. The expression of ANGPT2 and CAV1 were both increased in hrGECs under HG conditions and losartan intervention reduced the expression of them. Moreover, ANGPT2 downregulation reduced albumin transcytosis in hrGECs by regulating CAV1 expression. In vivo, the expression of ANGPT2 and CAV1 in the glomerulus was both increased significantly in DKD mice. Compared with DKD mice, losartan treatment reduced albuminuria and decreased the expression of ANGPT2 and CAV1 in a dose-dependent manner. CONCLUSIONS: ANGPT2 exacerbated albumin transcytosis across GECs by increasing CAV1 expression during HG exposure, thereby increasing albuminuria. Losartan reduces albumin transcytosis and albuminuria formation in DKD by inhibiting the upregulation of ANGPT2 under HG conditions. Our findings suggest that ANGPT2 and CAV1 may be novel therapeutic targets for diabetic albuminuria. In addition, we provide new evidence to elaborate on the mechanism of losartan in the development of DKD.