Abstract
Tumor cells educate platelets by modulating their RNA profiles and phenotypes. These platelets are termed "tumor educated platelets: TEPs", and this concept is widely accepted. We have previously reported that platelets are activated in microvessels within liver tumors, leading to the progression of hepatocellular carcinoma (HCC). When platelets are activated, platelet-derived microvesicles (PDMV) containing platelet-derived nucleic acids and proteins are released. The contents in these are known to vary depending on the various pathological conditions of the host. It is generally believed that when cells take up extracellular vesicles, they are affected by the action of the encapsulated substances. However, it is still unclear what biological properties of PDMV are released by TEPs and how they are involved in carcinogenesis. We hypothesized that the release of PDMV by activated platelets in liver tissue and the uptake of its contents by normal hepatocytes and HCC cells may be involved in the progression of hepatocarcinogenesis, and we analyzed their role in carcinogenesis.
The rat HCC model was prepared according to the Solt-Farber protocol. Briefly, male F344 rats were injected intraperitoneally with diethylnitrosamine at 4 weeks of age, then fed a powdered diet containing 2-acetylaminofluorene to induce liver damage, followed by partial hepatectomy. The rats were then kept under normal conditions for more than one month. Abdominal ultrasonography was performed on rats, and those with obvious tumors with a diameter of 2 mm or more were used for the experiments. Peripheral blood was collected from the tail veins of normal rats or HCC model rats, and platelets were isolated. The platelets were labeled with the fluorescent substance CMRA, activated in vitro with thrombin, and the PDMV in the supernatant was collected by polyethylene glycol precipitation and used as CMRA-labeled PDMV in the experiments.
When PDMV was added to primary cultured rat hepatocytes and cultured for 24 hours, fluorescence was observed in the hepatocytes. The fluorescence was also observed in hepatocytes in the portal region of liver tissue collected 24 hours after injection of CMRA-labeled PDMV from the mesenteric vein of rats. These indicate that PDMV released into the liver tissue is taken up by hepatocytes. Peripheral blood platelets from normal rats or HCC model rats were isolated and PDMV was purified from each (N-PDMV and HCC-PDMV). These PDMVs were added to cultures of normal rat hepatocytes and hepatocellular carcinoma cell lines to analyze their effects on cell survival. N-PDMV did not affect the viability of normal hepatocytes, but mildly enhanced the survival of HCC cells. On the other hand, HCC-PDMV inhibited the viability of normal hepatocytes, but promoted viability of HCC cells more strongly than N-PDMV.
These results indicate that the contents of PDMV differ between normal and HCC rats, and in particular that PDMV from rats with HCC is taken up by normal and HCC cells and is associated with selective survival of HCC cells.
Disclosures
No relevant conflicts of interest to declare.
Author notes
Asterisk with author names denotes non-ASH members.
