Horizontal RNA transfer mediates platelet-induced hepatocyte proliferation.

Liver regeneration is stimulated by blood platelets, but the molecular mechanisms involved are largely unexplored. Although platelets are anucleate, they do contain coding or regulatory RNAs that can be functional within the platelet or, after transfer, in other cell types. Here, we show that platelets and platelet-like particles (PLPs) derived from the megakaryoblastic cell line MEG-01 stimulate proliferation of HepG2 cells. Platelets or PLPs were internalized within 1 hour by HepG2 cells and accumulated in the perinuclear region of the hepatocyte. Platelet internalization also occurred following a partial hepatectomy in mice. Annexin A5 blocked platelet internalization and HepG2 proliferation. We labeled total RNA of MEG-01 cells by incorporation of 5-ethynyluridine (EU) and added EU-labeled PLPs to HepG2 cells. PLP-derived RNA was detected in the cytoplasm of the HepG2 cell. We next generated PLPs containing green fluorescent protein (GFP)-tagged actin messenger RNA. PLPs did not synthesize GFP, but in coculture with HepG2 cells, significant GFP protein synthesis was demonstrated. RNA-degrading enzymes partly blocked the stimulating effect of platelets on hepatocyte proliferation. Thus, platelets stimulate hepatocyte proliferation via a mechanism that is dependent on platelet internalization by hepatocytes followed by functional transfer of RNA stored in the anucleate platelet. This mechanism may contribute to platelet-mediated liver regeneration.

Transient von Willebrand factor-mediated platelet influx stimulates liver regeneration after partial hepatectomy in mice.

BACKGROUND & AIMS: In addition to their function in thrombosis and haemostasis, platelets play an important role in the stimulation of liver regeneration. It has been suggested that platelets deliver mitogenic cargo to the regenerating liver, and accumulation of platelets in the regenerating liver has been demonstrated. We studied kinetics of platelet influx in the regenerating liver and investigated the signal that initiates platelet influx. METHODS: We visualized platelets in the liver remnant after partial hepatectomy in mice using intravital microscopy and assessed liver regeneration by examination of liver/body weight ratio and the number of proliferating hepatocytes examined by immunohistochemistry. RESULTS: We demonstrated rapid but transient platelet influx into the liver remnant after a partial liver resection. Liver regeneration in thrombocytopenic mice was substantially impaired as evidenced by a reduced liver-to-body weight ratio and decreased numbers of proliferating hepatocytes at day 3 compared to mice with normal platelet counts. In contrast, liver regeneration was only mildly impaired when thrombocytopaenia was induced 2 hours after partial liver resection. Platelet influx into the liver remnant was virtually absent in the presence of an antibody to von Willebrand factor (VWF) suggesting that VWF release from liver sinusoidal endothelial cells mediates platelet influx. Additionally, liver regeneration in mice deficient in VWF was markedly impaired. CONCLUSIONS: A rapid but transient VWF-dependent platelet influx into the liver remnant drives platelet-mediated liver regeneration.

Evidence against a role for platelet-derived molecules in liver regeneration after partial hepatectomy in humans.

Background and Aim: Blood platelets have been shown to stimulate liver regeneration after partial hepatectomy in animal models and humans, but the molecular mechanisms involved are unclear. It has been proposed that growth factors and angiogenic molecules stored within platelets drive platelet-mediated liver regeneration, but little direct evidence in support of this mechanism is available. Methods: We assessed levels of relevant platelet-derived proteins (vascular endothelial growth factor, hepatocyte growth factor, fibroblast growth factor, platelet-derived growth factor, thrombospondin, and endostatin) in platelet-rich and platelet-poor plasma taken at various perioperative time points from patients undergoing a (extended) right partial hepatectomy (n = 17) or a pylorus-preserving pancreatico-duodenectomy (n = 10). In addition, we collected intraoperative samples from the efferent and afferent liver veins prior to and after completion of liver resection. Twenty-four healthy controls were included to establish reference ranges for the various tests. Results and Conclusions: Although we demonstrate perioperative changes in platelet and plasma levels of the proteins assessed, the changes observed in patients undergoing partial hepatectomy largely mirror the changes observed in patients undergoing a pylorus-preserving pancreatico-duodenectomy. In addition, no change in the growth factor levels in platelet-rich plasma between afferent and efferent liver veins was observed. Thus, the absence of an intra- or postoperative consumption of platelet-derived proteins in patients undergoing partial hepatectomy argues against a role of release of these molecules in stimulation of liver regeneration. Relevance for patients: In depth knowledge of the mechanism underlying platelet-mediated liver regeneration may facilitate development of targeted therapeutic interventions for patients with failing liver regeneration, which for example may occur following a partial hepatectomy. Although the prevailing dogma is that platelet stimulate liver regeneration by release of growth factors stored within platelets, data in this manuscript argue against this mechanism and suggest other pathways to be responsible.

Vitamin E Attenuates the Progression of Non-Alcoholic Fatty Liver Disease Caused by Partial Hepatectomy in Mice.

BACKGROUND AND AIM: The progression of non-alcoholic fatty liver disease (NAFLD) likely involves a 'multiple hit' mechanism. We hypothesized that partial hepatectomy, a procedure performed frequently in patients with NAFLD, would accelerate the progression of disease. METHODS: C57BL/6JolaHsd mice were fed a choline-deficient L-amino acid-defined diet (CD-AA) or a choline-sufficient L-amino acid-defined control diet (CS-AA). Part of the mice in the CD-AA group received a diet enriched in vitamin E (~20 mg /day). Two weeks after the start of the diet, mice underwent a partial hepatectomy or a sham operation. RESULTS: In the CD-AA group, NAFLD activity scores were significantly higher at 7 days after partial hepatectomy compared to the sham operated mice (3.7 ± 1.3 vs. 1.8 ± 0.7; P<0.05). In addition, TBARS, a measure for oxidative stress, in liver tissue of the CD-AA group were significantly higher at day 1, 3 and 7 after partial hepatectomy compared to the sham operated mice (P<0.05). Vitamin E therapy significantly reduced TBARS level at day 7 after partial hepatectomy compared to the CD-AA diet group (P< 0.05). Vitamin E suppletion reduced NAFLD activity score at day 7 after partial hepatectomy compared to the CD-AA group (2.3 ± 0.8 vs. 3.8 ± 1.0; P<0.05). CONCLUSION: Partial hepatectomy accelerates the progression of NAFLD. Disease progression induced by partial hepatectomy is substantially attenuated by vitamin E.