RESUMO
The Cystic Fibrosis Transmembrane conductance Regulator (CFTR) protein is a chloride channel localized at the apical plasma membrane of epithelial cells. We previously described that syntaxin 8, an endosomal SNARE (Soluble N-ethylmaleimide-sensitive factor Attachment protein REceptor) protein, interacts with CFTR and regulates its trafficking to the plasma membrane and hence its channel activity. Syntaxin 8 belongs to the endosomal SNARE complex which also contains syntaxin 7, vti1b and VAMP8. Here, we report that these four endosomal SNARE proteins physically and functionally interact with CFTR. In LLC-PK1 cells transfected with CFTR and in Caco-2 cells endogenously expressing CFTR, we demonstrated that endosomal SNARE protein overexpression inhibits CFTR activity but not swelling- or calcium-activated iodide efflux, indicating a specific effect upon CFTR activity. Moreover, co-immunoprecipitation experiments in LLC-PK1-CFTR cells showed that CFTR and SNARE proteins belong to a same complex and pull-down assays showed that VAMP8 and vti1b preferentially interact with CFTR N-terminus tail. By cell surface biotinylation and immunofluorescence experiments, we evidenced that endosomal SNARE overexpression disturbs CFTR apical targeting. Finally, we found a colocalization of CFTR and endosomal SNARE proteins in Rab11-positive recycling endosomes, suggesting a new role for endosomal SNARE proteins in CFTR trafficking in epithelial cells.
Assuntos
Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Células Epiteliais/metabolismo , Proteínas Qa-SNARE/metabolismo , Proteínas Qb-SNARE/metabolismo , Proteínas R-SNARE/metabolismo , Proteínas SNARE/metabolismo , Animais , Linhagem Celular , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Endossomos/metabolismo , Células Epiteliais/citologia , Humanos , Iodetos/metabolismo , Transporte Proteico/fisiologia , Proteínas Qa-SNARE/genética , Proteínas Qb-SNARE/genética , Proteínas R-SNARE/genética , Interferência de RNA , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas SNARE/genética , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismoRESUMO
BACKGROUND: Whether continuous renal replacement therapy is better than intermittent haemodialysis for the treatment of acute renal failure in critically ill patients is controversial. In this study, we compare the effect of intermittent haemodialysis and continuous venovenous haemodiafiltration on survival rates in critically ill patients with acute renal failure as part of multiple-organ dysfunction syndrome. METHODS: Our prospective, randomised, multicentre study took place between Oct 1, 1999, and March 3, 2003, in 21 medical or multidisciplinary intensive-care units from university or community hospitals in France. Guidelines were provided to achieve optimum haemodynamic tolerance and effectiveness of solute removal in both groups. The two groups were treated with the same polymer membrane and bicarbonate-based buffer. 360 patients were randomised, and the primary endpoint was 60-day survival based on an intention-to-treat analysis. FINDINGS: Rate of survival at 60-days did not differ between the groups (32% in the intermittent haemodialysis group versus 33% in the continuous renal replacement therapy group [95 % CI -8.8 to 11.1,]), or at any other time. INTERPRETATION: These data suggest that, provided strict guidelines to improve tolerance and metabolic control are used, almost all patients with acute renal failure as part of multiple-organ dysfunction syndrome can be treated with intermittent haemodialysis.
Assuntos
Injúria Renal Aguda/terapia , Hemodiafiltração/métodos , Diálise Renal/métodos , Idoso , Feminino , Hemodiafiltração/efeitos adversos , Humanos , Unidades de Terapia Intensiva , Tempo de Internação , Masculino , Pessoa de Meia-Idade , Insuficiência de Múltiplos Órgãos/mortalidade , Diálise Renal/efeitos adversosRESUMO
Type 1 plasminogen activator inhibitor (PAI-1) is the primary inhibitor of tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA). Whereas PAI-1 is not expressed in normal kidneys, it is strongly induced in glomerular diseases and thus could promote the local accumulation of fibrin. To study the role of PAI-1 in the development of inflammatory glomerular injury, passive antiglomerular basement membrane (GBM) glomerulonephritis (GN) was induced in PAI-1 knockout mice and in wild-type mice of the same genetic background. Unexpectedly, PAI-1 deficiency was associated with an early and severe exacerbation of glomerular injury: Infiltration by CD4 T cells, proportion of fibrinous crescents, and renal function impairment were significantly more pronounced in PAI-1 -/- mice. Interestingly, activation of transforming growth factor (TGF)- beta, which is known to be dependent on the PA/plasmin system in vitro, was dramatically enhanced in the kidneys in the absence of PAI-1. Moreover, administration of neutralizing antibodies against TGF-beta significantly attenuated the disease in PAI-1 -/- mice. This suggests that the poor outcome of GN in PAI-1 -/- mice is consecutive to an uncontrolled activation of TGF-beta and confers PAI-1 with a new, immunomodulatory role.
