Introduction: Haemolytic uraemic syndrome (HUS) is a thrombotic microangiopathy (TMA) that has a predilection to present in the kidney. It is a triad of microangiopathic haemolytic anaemia, thrombocytopenia, and acute kidney injury. In 90% of cases, HUS follows gastroenteritis secondary to infection with Shiga toxin (Stx) producing bacteria such as Escherichia coli. Stx HUS is the leading cause of acute kidney injury in children with a mortality of 5%. The precise pathophysiological mechanisms following Stx infection that lead to TMA are poorly understood.
Objective: To investigate whether Stx acts via the podocyte globotriaosylceramide (Gb3) receptor to cause HUS.
Results: To interrogate the pathways activated by Stx in podocytes, total and phospho-proteomic studies in human podocytes versus Gb3 knockout controls were performed. Ingenuity Pathway Analysis showed a significant reduction in VEGF-A signalling in wild type podocytes treated with Stx; suggesting VEGF-A maybe a critical coordinator of Stx HUS.
To illustrate that Gb3 is the receptor for Stx in vivo, a constitutive whole body Gb3 knockout mouse (Gb3KO) was generated. Gb3KO mice were completely protected from the toxicity of intraperitoneal Stx even at x400 the lethal dose. All wild type control mice died within 4 days of receiving Stx from extra-glomerular dehydration, as demonstrated by ATN on renal histology.
To determine the role of the podocyte Gb3 receptor in vivo, an inducible podocyte expressing Gb3 mouse on a Gb3 null background (PodGb3) was generated. PodGb3 mice given intraperitoneal Stx developed HUS (thrombocytopenia, haemolytic anaemia, and uraemia p<0.05) at day 10. Renal histology demonstrated glomerular TMA, with intracapillary thrombus on electron microscopy. Immunofluorescence demonstrated increased glomerular fibrinogen and C3b deposition vs. controls. Additionally, glomerular complement regulator factor H was significantly reduced in PodGb3 mice. Fascinatingly, administration of a C5 inhibitor in this model rescued the HUS phenotype.
Conclusions: The discovery that the podocyte is capable of causing Stx HUS, with activation of complement observed on the glomerular endothelium; is compelling evidence for the importance of glomerular cell cross-talk within the kidney in the development of the disease. This may explain why the glomerulus is the prime target of systemic Stx driven infection and lead to the development of novel therapies to help treat patients with this devastating condition.
|Date of Award||29 Sep 2020|
- The University of Bristol
|Sponsors||Kidney Research UK|
|Supervisor||Richard J M Coward (Supervisor) & Moin Saleem (Supervisor)|