In a new study published in the Journal of Hepatology, researchers from the San Raffaele Telethon Institute for Gene Therapy (SR-Tiget) show that only 15–20% of hepatocytes in newborn mice — dubbed clonogenic hepatocytes — are responsible for generating over 90% of the adult liver mass.
These findings have major implications for pediatric gene therapy. Understanding the dynamics of this subset of hepatocytes early in life can help scientists to achieve more effective and durable correction of inherited liver diseases through in vivo gene engineering strategies such as lentiviral gene transfer and genome editing.
Latest-Generation Technologies Reveal Cellular Blueprints
The team combined cutting-edge single-cell and spatial transcriptomics, clonal tracing, and mathematical modeling to analyze how hepatocytes proliferate and mature after birth. This approach allowed them to identify not only the clonogenic cells but also the molecular signals and tissue niches that regulate their activity.
"Using spatial transcriptomics, we captured the precise localization and transcriptional identity of hepatocytes during postnatal liver growth," explains Dr. Michela Milani, co-first author of the study. "It gave us an unprecedented view into how different hepatocyte subsets proliferate and mature."
Timing and Cell Identity Shape Gene Therapy Success
The researchers found that gene editing by homology-directed repair (HDR) is significantly enriched within the clonogenic hepatocytes, thus resulting in an expansion of the proportion of the gene-edited liver area at the end of the organ growth. On the other hand, lentiviral vectors distribute more evenly across hepatocyte populations, but their efficiency is affected by the maturation state and lobular position of the cells — particularly the emergence of peri-central (PC) identity after weaning, which is associated with reduced vector permissiveness due to elevated proteasome activity.
"Knowing that the liver becomes less permissive to gene transfer in certain zones as it matures helps us refine not just what cells to target, but also when to treat," says Dr. Francesco Starinieri, co-first author.
Pre-treatment with a proteasome inhibitor restored lentiviral transduction in adult hepatocytes, suggesting that this barrier can be pharmacologically modulated to improve gene delivery in mature livers.
A Shared Niche for Growth and Proliferation
Another key finding is the discovery of a specialized instructive tissue niche in neonatal livers, where clonogenic hepatocytes are found in close proximity to hematopoietic progenitor cells. This spatial co-localization suggests a shared pool of growth signals and might open up new possibilities for regenerative medicine.
Toward Durable Pediatric Gene Therapies
"This study extends our understanding of how the liver grows and matures and how we can intervene early in life to durably correct genetic diseases," says Dr. Alessio Cantore , senior and corresponding author. "By identifying the specific hepatocytes that fuel liver growth — and how they respond to gene delivery — we can now rationally design more effective and lasting therapies for children."
Dr. Cantore recently received a prestigious Consolidator Grant from the European Research Council (ERC) to further investigate liver tissue dynamics mechanisms and translate them into therapeutic applications.
The research was conducted in collaboration with Dr. Andrés Muro of the International Centre for Genetic Engineering and Biotechnology (ICGEB) , Trieste, and involved multidisciplinary contributions from molecular biologists, physicists, and bioinformaticians at SR-Tiget and the San Raffaele Scientific Institute.
This research was mainly supported by Fondazione Telethon, the Italian Ministry of Health, and the EU Horizon 2020 Program.
