| Acute kidney injury(AKI)is a serious public health burden in all countries of the world.The global incidence of AKI is steadily increasing,especially for the severely ill hospitalized patients or for those undergoing major surgeries.Due to its complex pathogenesis,current therapy is still limited to mechanical support such as dialysis,which is costly and predisposes to the progression of CKD(chronic kidney disease).Consequently,the development of strategies for alleviating the severity and progression of AKI is urgently needed.Ischemia-reperfusion(I/R)injury commonly causes and constantly worsens an existing AKI.Extracellular vesicles(EVs)released by mesenchymal stem cells(MSCs)have exhibited regenerative capability in the renal I/R injury mouse models.However,the in vivo fate and behaviors of MSC-derived EVs(MSC-EVs)in renal I/R injury has yet to be established.Moreover,precise targets in renal cells responsible for the regenerative effects of MSC-EVs and the associated regulatory mechanisms are presently undefined.Here,we first demonstrate the use of an aggregation-induced emission luminogens(AIEgens),DPA-SCP,for efficiently labeling and real-time tracking of MSC-EVs.Our in vitro results showed that both EVs and AIEgen-labeled EVs(AIE-EVs)are typical round to oval vesicles enclosed by a double membrane,and the diameter is around 120 nm.The mean size of AIE-EVs was comparable to the size of unlabeled EVs.There is no significant difference in protein contents between AIE-EVs and EVs.By confocal laser scanning microscopy(CLSM),we confirmed that AIE-EVs can be internalized into the renal tubular epithelial cells(TECs).Subsequently,in vivo imaging results showed that DPA-SCP realized non-invasively and precisely report the behaviors of MSC-EVs for 72 hours and we observed that MSC-EVs specifically accumulated in the injured kidney and taken up by TECs.Compared with commercial EV fluorescence probe PKH26,DPA-SCP exhibited high labeling efficiency and stronger optical stability.Moreover,DPA-SCP did not affect the anti-inflammatory and anti-apoptotic renal protective affects of MSC-EVs,exhibiting superior spatiotemporal resolution and tracking ability in I/R induced AKI mice.Mitochondria maintain a variety of cellular functions and process,such as the level of ROS and cell apoptosis.The most important function of mitochondria is the production of ATP,thus supplying the energy source for kideny repair and regeneration.Some investigators have emphasized that mitochondrial dysfunction has a role in the early stages and progression of renal diseases.In the present paper,we revealed that the accumulated MSC-EVs stimulated mitochondrial antioxidant defense and ATP production via activating the Keapl-Nrf2 signaling pathway,which protected TECs against oxidative insult by reducing mitochondrial fragmentation,normalizing mitochondrial membrane potential and increasing mitochondrial DNA copy number.Increased microRNA-200a-3p expression in renal TECs induced by MSC-EVs was identified as a regulatory mechanism contributing to the protective actions on mitochondria as well as stimulating the renal signal transduction pathways.In conclusion,we have developed an AIEgen,DPA-SCP,to directly label EVs without influencing their native biological structures and functions.The biocompatible DPA-SCP allows real-time visualizing and monitoring EV trafficking,and exhibits superior brightness,negligible photobleaching and excellent photostability to the most popular commercial EV trackers.We also demonstrated that MSC-EVs accumulated in the renal tubules during renal I/R injury and promoted the recovery of kidney function via activating Keapl-Nrf2 signaling pathway and enhancing mitochondrial function of TECs.Our data will provide evidence and expand the success of the clinical translation of MSC-EV for AKI patients. |
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