Focal areas of tumor necrosis were observed. In addition, most cells were of epithelioid type, but some lesions also showed areas with fusiform cells, another sign of aggressiveness. None of the animals developed spleen or lung metastasis (only about 1 mm peritoneal nodules were detected at the injection site in most animals). In parallel to the MRI-assessed HCC growth, ISC recipients also tended to have higher levels of cancer cell proliferation in the 10-min Epigenetics inhibitor ischemia experiments. Two weeks after transplantation, 42 [34–59]% of the cancer cells were Ki67 positive in the ISCs vs. 40 [27–55]% in the controls (p = 0.12) and 34 [18–57]% in the ISC/Rs (p = 0.013). Because serotonin has been suggested as an important mediator of normal hepatocyte and HCC cell proliferation, serotonin and its receptors were assessed [12] and [13]. The 12-h serum serotonin levels were similar between groups in the 10-min ischemia experiment (controls 22 [14–46] ng/ml, ISC 19 [14–28] ng/ml, ISC/R 30 [22–36] ng/ml, ISC vs. control p = 0.62, ISC vs. ISC/R p = 0.11 and ISC/R vs. control p = 0.37). In addition, the levels of expression of the serotonin receptors htr2a and htr2b were also similar between groups (data not shown). At 12 h, VEGF serum levels, VEGF and Flt1 (tyrosine kinase receptor for vascular endothelial growth factor) liver expressions were also similar between groups (data not shown). ISC recipients demonstrated a stronger pro-inflammatory serum cytokine profile 12 h after transplantation in the 10-min experiment. IL1β was higher in ISCs (372 [271–517] pg/ml) vs. controls (155 [102–385] pg/ml) and ISC/Rs (234 [171–313] pg/ml; ISC vs. control p = 0.020, ISC vs. ISC/R p = 0.0005, ISC/R vs. control p = 0.14, Fig. 3A). IL6 also tended to be higher in ISC and ISC/R recipients, but did not reach statistical significance ( Fig. 3B). By contrast, IL10 was higher in the ISC/R group, suggesting an anti-inflammatory effect of this treatment (ISC/R 177 [77–372] pg/ml, controls 18 [11–26] pg/ml, and ISC 25 [9–138] pg/ml; ISC vs. control p = 0.51, ISC vs. ISC/R p = 0.0014, ISC/R vs. control p = 0.0002, Fig. 3C). TNF-α was undetectable in the serum of all groups 12 h after transplantation. The level of expression was first tested using the “Rat Endothelial Cell Biology” RT2 Profiler PCR Array (SaBiosciences, Frederick, MD) (Supplementary Fig. 1), and subsequent specific PCR assessments were performed for selected genes in the 10-min experiments. Hmox1, Hif1a, and Serpine1 (plasminogen activator inhibitor-1), have been previously shown to be up-regulated in case of liver hypoxia and injury [14], [15] and [16]. The levels of these genes were lower in the ISC/R recipient grafts 12 h after transplantation ( Fig. 4A–C). Hmox1 level was 0.41 [0.31–0.87] in the ISC/R vs. 1.23 [0.70–3.03] in the controls and 2.51 [0.53–7.26] in the ISC (ISC/R vs. controls p = 0.0006, ISC/R vs. ISC p <0.0001). Hif1a level was 0.65 [0.45–0.78] in the ISC/R vs.