LAUSR

Orthotopic liver transplantation is the only effective cure for endstage liver disease. Unfortunately, the current need for liver transplants by far eclipses the availability of suitable grafts, resulting in the death of many waitlisted patients, as well as those who are not eligible for transplant. Therefore, regenerative therapies restoring liver function in these patients are desperately needed. Hepatocyte transplantation has the potential to restore metabolic capacity in dysfunctional livers, bridging patients with endstage liver disease to transplantation. Successful engraftment and subsequent expansion of hepatocytes may even enable the liver to regenerate and prevent the need for transplantation. However, generating sufficient numbers of hepatocytes and enabling their functional engraftment to rescue patients with impaired liver function remains a challenge.[1] Because mature primary human hepatocytes (PHHs) do not readily proliferate in culture, it is challenging to supply the high number of PHHs necessary to treat a patient with metabolic disease or acute liver failure.[1] Therefore, researchers have generated a variety of different hepatocytelike cells, including induced pluripotent stem cell (iPS)– derived hepatocytes (iHEPs)[2] and various complex liver organoid systems.[3] Direct reprogramming of human fibroblasts into functional and expandable hepatocytes may even allow syngeneic hepatocyte transplantation without having to take a detour through an iPS stage.[4] However, these hepatocytelike cells show reduced metabolic capacity when compared to PHHs and often resemble a rather immature progenitor state.[2– 4] PHHs can only be extensively expanded following dedifferentiation in defined culture media (proliferating human hepatocytes [ProliHHs]).[5] Unfortunately, their proliferative capacity comes at the expense of maturity and metabolic capacity.[5] Recent spatial transcriptomic profiling and lineage tracing experiments in mice further suggest an inverse correlation between the proliferative and metabolic capacity of hepatocytes in vivo.[6] Another key challenge is limited engraftment of hepatocytelike cells or ProliHHs following transplantation.[2– 5] Unfortunately, animal studies demonstrated that the innate immune system cleared the majority of transplanted hepatocytes, regardless of whether they were allogeneic or syngeneic.[7] A new study from Wang et al. is shedding light on the mechanisms preventing successful transplantation of expanded hepatocytes and identifies potential mitigation strategies.[8] First, the authors assessed whether longterm culture, which is necessary to obtain the number of hepatocytes required for transplantation into patients, affected engraftment. They transplanted shortterm cultured ProliHHs (scProliHHs) and longterm cultured ProliHHs (lcProliHHs) into fumarylacetoacetate hydrolase (Fah)−/− Rag2−/− IL2rg−/− (FRG) mice [9] and assessed engraftment rates. FRG mice enable transplantation of human cells through suppression of T cells and natural killer cells and provide a competitive advantage for transplanted ProliHHs by inducing damage in the resident hepatocyte population through Fah deletion.[9] While the size of engrafted cell clones was similar, there was a dramatic reduction in the number of clones per liver lobe in lcProliHHs when compared to scProliHHs, suggesting that longterm expansion of ProliHHs reduced their engraftment capacity while retaining their proliferative potential. The lab has previously shown that ProliHHs are dedifferentiated when compared with PHHs, as indicated by reduced metabolic function and an increase in progenitor cell markers.[5] RNA expression profiling now revealed that lcProliHHs further dedifferentiated when compared to scProliHHs, suggesting that longterm culture reduced hepatocyte maturity. Interestingly, lcProliHHs showed increased RNA expression as well as protein secretion of several chemokines responsible for the recruitment of monocytes, macrophages, and neutrophils. In addition, multiple proinflammatory factors, damageassociated molecule patterns, and cytokines activating macrophages were upregulated in lcProliHHs when compared to scProliHHs. Collectively, the authors termed these factors dedifferentiationassociated inflammatory factors (DAIFs). Because FRG mice retain the innate immune response mediated by macrophages and neutrophils[9] Received: 1 April 2022 | Accepted: 1 April 2022