E effects of rIP-10 were compatible to iPS alone (Fig. 5B). Combined treatment of rIP-10 and iPS had no additive beneficial effects in injured mice. The application of anti-IP-10 neutralizing antibody attenuated the protective effects of iPS (Fig. 5C). In addition, the Ki67 or BrdU staining revealed that the proliferation of hepatocytes at portal regions after iPS infusion was significantly reduced by the anti-IP-10 neutralizing antibody (Fig. 5D).Localization of iPS in the Injured LiverFrom above results, iPS outperformed the iHL in promotion of hepatocyte regeneration. Therefore, we further examined the engraftment of the transplanted iPS. To examine the localization 1676428 of iPS in the liver, we labeled iPS with a red fluorescence dye, DiI, before infusion. Under fluorescent microscopic observation, theIP-10 in Liver Injury Post iPS TransplantationFigure 1. iPS and hepatocytes transplantation reduced hepatic injury. (A) Mean AST and ALT levels in mice receiving PBS (open bars), iPS (gray bars), and iHL (solid bars) following CCl4 treatment (n = 6, *P,0.05 vs. PBS, #P,0.05 vs. iPS). (B) Representative liver sections from CCl4-injuredIP-10 in Liver 
Injury Post iPS Transplantationmice that received vehicle, iPS or iHL infusion. Necrotic area were quantified and the percentage were shown (n = 5, *p,0.05 vs. vehicle). (C) At 48 h post CCl4 treatment, hepatocyte proliferation of vehicle (PBS), iHL, iPS was measured by Ki67 immunostaining and BrdU incorporation assay (n = 6, *p,0.05 vs. PBS, #p,0.05 vs. iPS). doi:10.1371/journal.pone.0050577.gIPS Improved the Survival of Repetitive Injured MiceTo evaluate the survival effects of iPS and IP-10, the 72-hour survival rate was evaluated in repetitive CCl4-injured 
mice, to which two additional doses of CCl4 (given at 24 and 48 hours) were given after the first dose. Half of the repetitive injured mice were randomized into two groups to receive either iPS, or rIP-10 (5 ng) treatment. Both rIP-10 and IPS groups had significantly higher 72-hour survival rates (100 and 85.7 , respectively) when compared to the untreated group (53.3 , P,0.05) (Fig. 5E). No significant difference was noted between iPS and rIP-10 groups.DiscussionAcute massive or chronic persistent liver injuries can lead to liver 4 IBP failure. Developing a cell-based treatment or alternative therapeutic stratagem to reduce damage, prevent progression, and restore liver function is of important clinical relevance. This study demonstrated that the intravenously administered iPS reduced the intensity of injury and promoted hepatocyte proliferation. Thetransplanted iPS secreted IP-10 and help to increase hepatic IP-10 levels. The protective effect of iPS was attenuated by anti-IP-10 neutralizing antibody. In addition, applying rIP-10 protected hepatocytes and mice from CCl4 injury and improved their survival. These results demonstrated that iPS transplantation facilitated liver damage repair and promoted hepatocyte regeneration in order to restore liver function. Hepatic IP-10 was an important factor that mediated the beneficial effect of iPS in acute liver injury. Because iPS have the potential to proliferate Nafarelin manufacturer indefinitely and differentiated into different cell types, hepatocytes generated from iPS can be a valuable alternative source of primary hepatocytes [7,12]. However, it is unknown if the hepatocytes derived from iPS can provide adequate function better than iPS in the recipients. To answer this question, we compared the therapeutic effects o.E effects of rIP-10 were compatible to iPS alone (Fig. 5B). Combined treatment of rIP-10 and iPS had no additive beneficial effects in injured mice. The application of anti-IP-10 neutralizing antibody attenuated the protective effects of iPS (Fig. 5C). In addition, the Ki67 or BrdU staining revealed that the proliferation of hepatocytes at portal regions after iPS infusion was significantly reduced by the anti-IP-10 neutralizing antibody (Fig. 5D).Localization of iPS in the Injured LiverFrom above results, iPS outperformed the iHL in promotion of hepatocyte regeneration. Therefore, we further examined the engraftment of the transplanted iPS. To examine the localization 1676428 of iPS in the liver, we labeled iPS with a red fluorescence dye, DiI, before infusion. Under fluorescent microscopic observation, theIP-10 in Liver Injury Post iPS TransplantationFigure 1. iPS and hepatocytes transplantation reduced hepatic injury. (A) Mean AST and ALT levels in mice receiving PBS (open bars), iPS (gray bars), and iHL (solid bars) following CCl4 treatment (n = 6, *P,0.05 vs. PBS, #P,0.05 vs. iPS). (B) Representative liver sections from CCl4-injuredIP-10 in Liver Injury Post iPS Transplantationmice that received vehicle, iPS or iHL infusion. Necrotic area were quantified and the percentage were shown (n = 5, *p,0.05 vs. vehicle). (C) At 48 h post CCl4 treatment, hepatocyte proliferation of vehicle (PBS), iHL, iPS was measured by Ki67 immunostaining and BrdU incorporation assay (n = 6, *p,0.05 vs. PBS, #p,0.05 vs. iPS). doi:10.1371/journal.pone.0050577.gIPS Improved the Survival of Repetitive Injured MiceTo evaluate the survival effects of iPS and IP-10, the 72-hour survival rate was evaluated in repetitive CCl4-injured mice, to which two additional doses of CCl4 (given at 24 and 48 hours) were given after the first dose. Half of the repetitive injured mice were randomized into two groups to receive either iPS, or rIP-10 (5 ng) treatment. Both rIP-10 and IPS groups had significantly higher 72-hour survival rates (100 and 85.7 , respectively) when compared to the untreated group (53.3 , P,0.05) (Fig. 5E). No significant difference was noted between iPS and rIP-10 groups.DiscussionAcute massive or chronic persistent liver injuries can lead to liver failure. Developing a cell-based treatment or alternative therapeutic stratagem to reduce damage, prevent progression, and restore liver function is of important clinical relevance. This study demonstrated that the intravenously administered iPS reduced the intensity of injury and promoted hepatocyte proliferation. Thetransplanted iPS secreted IP-10 and help to increase hepatic IP-10 levels. The protective effect of iPS was attenuated by anti-IP-10 neutralizing antibody. In addition, applying rIP-10 protected hepatocytes and mice from CCl4 injury and improved their survival. These results demonstrated that iPS transplantation facilitated liver damage repair and promoted hepatocyte regeneration in order to restore liver function. Hepatic IP-10 was an important factor that mediated the beneficial effect of iPS in acute liver injury. Because iPS have the potential to proliferate indefinitely and differentiated into different cell types, hepatocytes generated from iPS can be a valuable alternative source of primary hepatocytes [7,12]. However, it is unknown if the hepatocytes derived from iPS can provide adequate function better than iPS in the recipients. To answer this question, we compared the therapeutic effects o.