Papel de las vesículas extracelulares en el deterioro cognitivo y motor en hiperamonemia y encefalopatía hepática. Mecanismos moleculares e implicaciones terapéuticas

Resumen

Background: Appearance of minimal hepatic encephalopathy is associated with changes in the peripheral immune system which are transferred to brain, leading to neuroinflammation which leads to cognitive and motor impairment. The mechanisms by which changes in the immune system induce cerebral alterations remain unclear. Extracellular vesicles (EVs) seem to play a role in this process in certain pathologies. The aim of this work was to assess whether EVs play a role in the induction by chronic hyperammonemia of neuroinflammation in cerebellum and hippocampus and in the induction of motor and cognitive impairment. Extracellular vesicles from mesenchymal stem cells (MSCs) reduce neuroinflammation in some pathological conditions. This thesis also aimed to assess if treatment of hyperammonemic rats with EVs from MSCs reduces neuroinflammation and restores cognitive and motor function and to study the mechanisms involved. Methods: We isolated EVs from plasma of control or hyperammonemic rats and characterized the differences in protein cargo by proteomics and Western blot. We assessed whether injection of EVs to normal rats induces neuroinflammation in cerebellum and hippocampus. by immunohistochemistry and Western blot Effects on motor and cognitive function were assessed using the beam walking, object location memory, object recognition memory and Y Maze tests. Mechanistic effects were analyzed ex vivo in cerebellar and hippocampal slices. Treatment of EVs from MSCs was performed in vivo by i.v. injection and ex vivo in slices. Learning and memory were assessed using the following tests: object location, object recognition, Y maze and radial maze. Results: Hyperammonemia increases EVs amount and alters their protein cargo. TNFα and its receptor TNFR1 are increased in EVs in hyperammonemia. Injected EVs reached the hippocampus and the cerebellum. Injection of EVs from hyperammonemic, but not from control rats, induced motor incoordination and cognitive impairment, mediated by neuroinflammation, microglia and astrocytes activation. Ex vivo studies showed that this effect is mediated by the TNFa present in the vesicles. Hyperammonemia induced neuroinflammation in hippocampus and cerebellum and impaired learning, motor coordination and memory in the tests performed. Treatment with EVs reduced microglia and astrocytes activation and restored performance of hyperammonemic rats in all the behavioral tests. Ex vivo studies showed that these beneficial effects were dependent on TGFb contained in the EVs. Conclusions: Plasma EVs from hyperammonemic rats carry molecules necessary and sufficient to trigger neuroinflammation in cerebellum and hippocampus and the mechanisms leading to motor incoordination and cognitive impairment. EVs from MSCs reduce neuroinflammation in cerebellum and hippocampus and restore cognitive and motor function in hyperammonemic rats. EVs from MSCs may be useful to improve cognitive function in patients with MHE.