肝细胞调控肝窦内皮细胞影响肝再生及其机制研究

Liver regeneration is an intricate but evolutionarily conserved process involving the interactions of various hepatic cell types, which contribute to hepatocytes proliferation and hepatovasular mass reconstitution. However, it remains unclear how hepatocytes coordinate with liver sinusoidal endothelial cells (LSECs) to initiate and restore liver regeneration. Here, we showed that, whatever in 70 % partial hepatectomy (PHx) or carbon tetrachloride (CCl4)-induced liver injury model, hepatocytes initiate proliferation and LSECs follow up. Even in liver biopsies from patients with fibrosis/cirrhosis, increasing hepatocytes accompanied by endothelial cells were also observed. Specific blockage of hepatocyte proliferation by oncostatin M (OSM) antibody or c-Met inhibitor (JNJ-38877605) obviously blocks later LSECs proliferation after PHx or CCl4 exposure. Various liver injury-induced hepatocyte proliferation were accompanied by Gankyrin expression. In PHx or iterative CCl4 injection model, Gankyrin deficiency in hepatocytes impairs the early burst of hepatocyte proliferation through reductive response to HGF, and resultant reconstitution of the hepatovascular mass. Remarkably, in vitro co-cultures, gankyrin-absent hepatocyte lessened endothelial cell proliferation, strongly indicating some unknown cytokine or growth factor secreted from hepatocyte to indirectly regulate endothelial cell growth. Next, this unknown cytokine or the growth factor will be screened by multicytokines analysis and verified in vitro. Once solid evidence is got, the trangenic mouse about this cytokine will be introduced or constructed to confirm its role in vivo. In addition, given LSEC progenitor cells (SPCs) as a main source of LSECs during hepatectomy, the effect of hepatocyte on SPCs recruiting or grafting in the liver will be further studied. Collectively, we believe that these data will confirm our hypothesis but also be helpful for liver transplantation.

肝再生过程是不同类型的细胞在多种细胞/生长因子及众多基因共同调控下，进行增殖与分化以恢复正常体积和功能。其中，肝窦内皮细胞早于肝细胞活化，却晚于肝细胞增殖的时相差异调控机制尚不明确。早期快速启动的肝细胞增殖缓解了肝脏受损后面临的代谢压力，然而血管形成滞后以及肝细胞快速增殖后相对缺血缺氧的环境，对新生肝细胞的存活产生了应激压力，肝细胞需要驱动新生肝内血管，以满足自身的生存需求。因此，探讨肝细胞是否调控肝窦内皮细胞影响肝再生将有助于深化认识肝再生过程特别是肝脏重建机理。我们前期在Gankyrin肝细胞敲除小鼠模型上发现，肝再生过程中肝细胞内快速上调的Gankyrin可调节肝内皮细胞再生。我们将在明确该调控机制基础上，进一步通过特定细胞/生长因子肝细胞敲除小鼠，探索肝再生中肝细胞调控肝窦内皮细胞再生机理，肝细胞是否调控内皮前体细胞趋化、定植以及肝窦内皮细胞功能，阐述肝细胞调控下肝内血管重建机制。

肝再生过程是不同类型的细胞在多种细胞/生长因子及众多基因共同调控下，进行增殖与分化以恢复正常体积和功能。其中，肝窦内皮细胞早于肝细胞活化，却晚于肝细胞增殖的时相差异调控机制尚不明确。早期快速启动的肝细胞增殖缓解了肝脏受损后面临的代谢压力，然而血管形成滞后以及肝细胞快速增殖后相对缺血缺氧的环境，对新生肝细胞的存活产生了应激压力，肝细胞需要驱动新生肝内血管，以满足自身的生存需求。因此，探讨肝细胞是否调控肝窦内皮细胞影响肝再生将有助于深化认识肝再生过程特别是肝脏重建机理。我们前期在Gankyrin肝细胞敲除小鼠模型上发现，肝再生过程中肝细胞内快速上调的Gankyrin可调节肝内皮细胞再生。我们将在明确该调控机制基础上，进一步通过Met和OSRMR肝细胞敲除小鼠，探索肝再生中肝细胞调控肝窦内皮细胞再生机理；发现了肝细胞可通过降低PEDF，促进VEGFA分泌，一方面促使内皮前体细胞趋化、定植分化为肝窦内皮细胞，另一方面VEGFA促进了内皮细胞的增殖，阐述了肝细胞调控下肝内血管重建机制。为了排除肝内其他非实质细胞的影响，我们进一步借助类器官培养体系验证了肝细胞对肝窦内皮细胞的调控作用及其机制，明确了PEDF可作为提高肝切除术患者术后再生恢复的干预靶点。另一方面，我们还研究了Kupffer细胞在肝再生进程中的作用，发现了Kupffer细胞除了参与再生启动之外，还参与了再生进程中肝细胞增殖的维持，但不影响再生终止；机制上表现为Kupffer细胞可通过分泌OSM，抑制TGF-β通路活化，进而维持再生进程中肝细胞增殖。综上，我们探讨了肝细胞和Kupffer细胞对肝再生的影响，不仅为阐明肝再生中细胞间的互作机制提供了重要线索，也为行肝切除术后再生恢复干预提供了实验性依据。