Hepatic injury signals drive proteomic reprogramming of the mesenchymal stem cells secretome to counteract liver damage

Paracetamol (APAP) overdose remains a leading cause of acute liver failure (ALF), a life-threatening condition with limited therapeutic options. The secretome of mesenchymal stem cells (MSCs) contain immunomodulatory and pro-regenerative factors that promote endogenous liver regeneration in ALF. In vivo , MSCs respond dynamically to injury microenvironments, adapting their paracrine activity to counteract damage. Accordingly, in vitro priming has been explored to reproduce injury-associated cues to enhance MSC therapeutic efficacy. This study aimed at uncovering the regenerative response of MSCs to hepatic pathological conditions, in specific to hepatic injury-associated signals. Human neonatal MSCs (hnMSCs) were primed with SOS signals from APAP-injured HepG2 cells. The resulting secretome (CM-pMSC) was analyzed for proteomic composition and evaluated for biological activity in APAP-injured hepatocyte-like cells (HLCs). CM-pMSC significantly increased APAP-injured HLCs viability, reduced reactive oxygen species, recovered APAP-impaired urea production and restored HLCs’ gene expression of proliferation- and APAP toxicity-related genes, namely ccnd1 , wnt2 , cyp2e1 , ask1 and gss . Proteomic analysis revealed that CM-pMSC was enriched in proteins associated with response to stress, immunoregulation, extracellular matrix (ECM) organization and metabolism regulation. This included proteins involved in key phases of hepatocyte proliferation, e.g. , IL-6, HGF, TGF-β1, as well as proteins relevant to the modulation of the regenerative milieu, such as MMPs, ORM1, GDF15, SERPINA1, APOA1, APOE. CM-pMSC promoted a regenerative response in APAP-injured HLCs, potentially linked to proliferation, oxidative stress control, immunomodulation, ECM remodeling, and metabolic regulation. These findings support the development of targeted MSC secretome-based therapies for acute liver injury.