Modulation Of Liver Regeneration Via Myeloid PTEN Deficiency

Liver regeneration is the compensatory hyperplasia of the liver in response to injury.During this process,the innate immune system,particularly Kupffer cells and NK cells,plays a fundamental role.Several lines of evidence suggest that Kupffer cells support liver regeneration,particularly based on secretion of TNF-a and IL-6,which facilitate activation of Stat3 in hepatocytes.Kupffer cells are also critical for initiating the proliferation of liver progenitor cells following liver injury.In contrast,NK cells negatively modulate liver regeneration.This effect appears secondary to the secretion of IFN-y,which activates Statl and antagonizes Stat3 activation in hepatocytes,thus inhibiting hepatocyte proliferation.A recent study suggests that NK cells are the most important source of IFN-y after partial hepatectomy,and deficiency of the co-inhibitory receptor TIGIT on NK cells leads to over-activation of NK cells and thus potentially impedes liver regeneration.The phosphatase protein PTEN was originally identified as a tumor suppressor protein,and is commonly mutated or deleted in a wide variety of tumors.PTEN is a lipid phosphatase that can negatively modulate the PI3K-Akt signaling pathway,one of the most important drivers of cell survival and proliferation.In addition,PTEN is also important in the regulation of immune cell function.A recent study found that PTEN was a positive regulator of TLR4 signaling in murine peritoneal macrophages,thus affecting their secretion of various inflammatory cytokines.PTEN can also regulate the expression of several genes required for M2 polarization in peritoneal macrophages and modulate inflammatory cytokine production in the liver.Nevertheless,the role of PTEN in liver regeneration is unclear.We propose that a better understanding of the interplay of Kupffer cells and NK cells is essential to understanding the molecular events that modulate liver regeneration.To better investigate the regulation of liver regeneration by Kupffer cells and NK cells,we mainly took advantage of LysMcre/+PTENf/f mice(PTEN deficiency specifically in myeloid cells,PTENmKO mice)and the control PTENf/f mice and their 2/3 partial hepatectomy(2/3 PHx)surgery to implement our research.Our results are listed as follows:Characteristics of liver Kupffer cells after PHxWe found that,as previously reported,Kupffer cell depletion by clodronate liposomes significantly compromised the liver regeneration rate.Moreover,as peritoneal macrophages were reported to modulate liver repair during sterile inflammation,to exclude the effect of peritoneal macrophages on liver regeneration,the peritoneal cavity was washed with PBS 24h before 2/3 PHx.Meanwhile,these mice were treated with PBS-liposomes or clodronate liposomes,respectively.The result showed a similar trend,as Kupffer cell depletion in these peritoneal cavity washed mice also significantly compromised the liver regeneration rate.Furthermore,even though the number of Kupffer cells did not change significantly,the PTEN expression level increased dramatically in Kupffer cells after 2/3 PHx.The increase in PTEN level was accompanied by a more activated M1-related phenotype of Kupffer cells,as reflected by the down-regulation of CD206,and up-regulation of CD 11c,MHC-II,and CD80.PTENmKO mice show a more prominent liver-regenerating capacity after PHxWe further used PTENmKO mice and littermate PTENf/f mice following PHx to investigate the role of PTEN in Kupffer cells during liver regeneration.Compared with PTENf/f control mice,PTENmKO mice showed an enhanced liver regeneration rate,as indicated by immunohistochemical staining of PCNA and Ki-67 of liver tissues,mitosis rate reflected by HE staining,and the ratio of liver weight to body weight at various time points after PHx.PTEN-deficient Kupffer cells demonstrate an M2-like polarization state and compromised NK cell-activating abilityFurther evidence demonstrated that PTENmKO Kupffer cells showed significantly higher levels of phosphorylated Akt and FoxO1 compared with those of PTENf/f mice,rendering their polarization towards an M2 state.Moreover,NK cells from PTENmKO showed less activation state,which was reflected by their IFN-? secretion ability and activation-associated surface markers.Further investigations demonstrated that the M2-polarized Kupffer cells in PTENmKO mice expressed less surface markers associated with NK cell activation,and produced less IL-12,indicating they were responsible for the less activated state of NK cells from these mice.PTEN-deficient Kupffer cells had increased growth factor-producing capacityUsing real-time PCR analysis,we found that the expression levels of pdgf,hgf and osm were all significantly higher in PHx treated PTENmKO Kupffer cells compared with those of PTENf/f control mice 48h post-PHx.In accordance with this,significantly higher levels of phosphorylated Stat3(p-Stat3),a downstream signaling molecule,were observed in PTENmKO mice at this time.This direct mitogenic role of rupffer cell was corroborated in vitro,because conditioned medium from PTENmKO mouse-derived Kupffer cells more potently stimulated mouse hepatocyte cell line growth.In conclusion,the results presented here demonstrated that PTEN is pivotal in inhibiting M2-like polarization of kupffer cells after PHx,resulting in NK cell activation,thus inhibiting the process of liver regeneration.Moreover,PTEN also prevents growth factor secretion from kupffer cells.These effects of myeloid PTEN combined to play a comprehensive role for suppression of liver regeneration.Considering the convenience of kupffer cell-targeted drugs for the reason of their strong phagocytosing ability,it is very promising to develop new therapies aimed at interfering PTEN expression in kupffer cells after liver resection surgery clinically.