CINC-2 and miR-199a-5p in EVs secreted by transplanted Thy1+ cells activate hepatocytic progenitor cell growth in rat liver regeneration

Abstract

Background Small hepatocyte-like progenitor cells (SHPCs) are hepatocytic progenitor cells that transiently form clusters in rat livers treated with retrorsine (Ret) that underwent 70% partial hepatectomy (PH). We previously reported that transplantation of Thy1+ cells obtained from d-galactosamine-treated livers promotes SHPC expansion, thereby accelerating liver regeneration. Extracellular vesicles (EVs) secreted by Thy1+ cells induce sinusoidal endothelial cells (SECs) and Kupffer cells (KCs) to secrete IL17B and IL25, respectively, thereby activating SHPCs through IL17 receptor B (RB) signaling. This study aimed to identify the inducers of IL17RB signaling and growth factors for SHPC proliferation in EVs secreted by Thy1+ cells (Thy1-EVs). Methods Thy1+ cells isolated from the livers of rats treated with d-galactosamine were cultured. Although some liver stem/progenitor cells (LSPCs) proliferated to form colonies, others remained as mesenchymal cells (MCs). Thy1-MCs or Thy1-LSPCs were transplanted into Ret/PH-treated livers to examine their effects on SHPCs. EVs were isolated from the conditioned medium (CM) of Thy1-MCs and Thy1-LSPCs. Small hepatocytes (SHs) isolated from adult rat livers were used to identify factors regulating cell growth in Thy1-EVs. Results The size of SHPC clusters transplanted with Thy1-MCs was significantly larger than that of SHPC clusters transplanted with Thy1-LSPCs (p = 0.02). A comprehensive analysis of Thy1-MC-EVs revealed that miR-199a-5p, cytokine-induced neutrophil chemoattractant-2 (CINC-2), and monocyte chemotactic protein 1 (MCP-1) were candidates for promoting SHPC growth. Additionally, miR-199a-5p mimics promoted the growth of SHs (p = 0.02), whereas CINC-2 and MCP-1 did not. SECs treated with CINC-2 induced Il17b expression. KCs treated with Thy1-EVs induced the expression of CINC-2, Il25, and miR-199a-5p. CM derived from SECs treated with CINC-2 accelerated the growth of SHs (p = 0.03). Similarly, CM derived from KCs treated with Thy1-EVs and miR-199a-5p mimics accelerated the growth of SHs (p = 0.007). In addition, although miR-199a-overexpressing EVs could not enhance SHPC proliferation, transplantation of miR-199a-overexpressing Thy1-MCs could promote the expansion of SHPC clusters. Conclusion Thy1-MC transplantation may accelerate liver regeneration owing to SHPC expansion, which is induced by CINC-2/IL17RB signaling and miR-199a-5p via SEC and KC activation. Graphical Abstract

“Small Hepatocytes” in the Liver

“…In addition, activated KCs by transplanted hepatic Thy1 + cells produced cytokineinduced neutrophil chemoattractant-2 (CINC-2) and miR-199a-5p, which promoted Il17b expression in SECs and SHPC proliferation, respectively. Furthermore, the conditioned medium of SECs treated with CINC-2 promoted SH proliferation [194]. These results indicate that hepatic Thy1 transplantation into Ret/PH-treated livers promotes SHPC proliferation via the IL-17RB signaling pathway and that miR-199a-5p may act directly on SHPCs to promote proliferation.…”

“…[171] miR-125b-5p [194] miR-199a-5p [194] Inhibitors Growth factors Activin A [195,196] TGF-β [195] Cytokines Interleukin 1β (IL-1β) [195] Others * Dexamethasone [193] Autocrine Stimulators Growth factors TGF-α [191] Follistatin [196] Inhibitor Activin B [196] * SHPCs.…”

“…We have reported that cell transplantation into Ret/PH-treated rat livers affects SHPC proliferation [158,171,194]. Isolated Thy1-positive mesenchymal cells (hepatic Thy1) from GalN-treated rat liver and rat BM-MCs could stimulate liver regeneration of the recipients as well as hepatomegaly due to SHPC expansion.…”

“…Hepatic Thy1 consists of two types of cells, fibroblast-like mesenchymal cells (Thy1-MCs) and epithelial-like cells, but only Thy1-MCs can stimulate SHPC proliferation, while the latter cannot. However, epithelial-like cells contain LSPCs that differentiate into SHs [158,194].…”