Human embryonic stem cell spheroids derived exosomal hsa_circ_0003258 sequestered dual targeting of miR-502-5p on KLF4/NF-κB and GALNT4/P53 to mitigate inflammation and apoptosis in hepatic ischemia reperfusion injury

Abstract

Hepatic ischemia–reperfusion injury (HIRI) refers to excessive inflammation and metabolic dysfunction caused by liver surgery injuries. Exosomes (Exos) derived from stem cells (SCs) play an active role in mitigating HIRI, but the molecular mechanism remains largely elusive. In this study, we demonstrated that Exos derived from human embryonic stem cell spheroids (3D hESCs) could alleviate HIRI and restore liver functions by alleviating inflammation in M1 macrophages (Macs) and resisting apoptosis of hepatocytes. Regarding the molecular mechanism behind these therapeutic effects, we identified a specific circRNA, named hsa_circ_0003258, enriched in 3D hESC-Exos, plays a key role in the treatment of HIRI. The synergistic effect of hsa_circ_0003258 was identified not only to contribute to the suppression of inflammation but also concurrently inhibited hepatocyte apoptosis by using a series of in vitro assays, particularly utilizing the co-culture system of hepatocyte organoids (Hep-orgs) with M1 Macs. Conversely, silencing hsa_circ_0003258 had opposite effects. Specifically, hsa_circ_0003258 was confirmed to act as a sponge for miR-502-5p with dual targets on KLF4 of Macs and GALNT4 of hepatocytes, and jointly activating the expressions of KLF4 in Macs and GALNT4 in hepatocytes. This cascade ultimately mitigated Macs inflammation and hepatocytes apoptosis by simultaneously deactivating NF‐κB and P53 signaling pathways. In conclusion, we have illustrated that Exos derived from 3D hESCs possess the capability to attenuate HIRI. This effect is primarily achieved through the synergistic blockade of dual pathways involving hsa_circ_0003258/miR-502-5p/KLF4/NF‐κB and hsa_circ_0003258/miR-502-5p/GALNT4/P53 axes. Our findings provide insights into the precision treatment of HIRI through Exos based cell-free therapy.