NAD+ (nicotinamide adenine dinucleotide), a core coenzyme in cellular metabolism, is involved in multiple physiological processes including redox reactions, energy metabolism and signal transduction. Dysregulation of its metabolism is closely associated with the development and progression of various diseases. In recent years, the therapeutic potential of NAD+ precursor supplements in disease models has been widely verified, yet the functional specificity and regulatory mechanisms of NAD+ pools at the subcellular level remain to be elucidated.
In December 2025, a research team from the University of Pennsylvania published a study titled Hepatocyte mitochondrial NAD+ content is limiting for liver regeneration in Nature Metabolism. This study for the first time confirmed that hepatocyte mitochondrial NAD+ content is a key limiting factor for liver regeneration, and mitochondrial NAD+ transport mediated by the transporter SLC25A51 plays a core regulatory role in this process, providing a completely new perspective for the targeted therapy of liver diseases.

Subcellular Compartmentalization of NAD+: A Core Basis for Functional Specificity
Intracellular NAD+ exhibits a strict subcellular compartmentalized distribution, with distinct NAD+ pools in mitochondria, cytoplasm, nucleus and other regions, each participating in unique physiological processes. Previous studies have shown that systemic NAD+ precursor supplementation can improve liver regeneration efficiency, but it is impossible to distinguish which specific subcellular pool plays a key role.
This study used stable isotope labeling combined with subcellular fractionation technology (NAD-SILEC) to confirm that the dynamic changes of mitochondrial NAD+ pools are highly correlated with the rate of liver regeneration, while cytoplasmic NAD+ levels show no significant fluctuations during the regeneration process.
This finding reveals the regional specificity of NAD+ function — the sufficient supply of only the mitochondrial NAD+ pool can meet the energy metabolism and biosynthesis requirements for liver regeneration, providing a theoretical basis for the precise regulation of NAD+ metabolism.
SLC25A51: Key mediator of NAD+transport in hepatocyte mitochondria
The selective permeability of the mitochondrial membrane dictates that NAD+ must enter the matrix through specific transporters to exert its effects. Previous studies have speculated that SLC25A47 may be the major NAD+ transporter in hepatocyte mitochondria, but this study overturned this hypothesis through a series of functional verification experiments:

• Gene silencing and overexpression experiments: In HeLa cells and HepG2 hepatocytes, silencing SLC25A51 significantly reduced mitochondrial NAD+ content, while overexpressing SLC25A47 had no obvious effect on mitochondrial NAD+ levels;

• Gene knockout mouse verification: Liver mitochondrial NAD+ content in hepatocyte-specific SLC25A47 knockout mice remained unchanged, further confirming that it does not possess NAD+ transport function;

• Transport activity assay: Using a PARylation detection system based on mitochondria-targeted PARP1 catalytic domain (mitoPARP1cd), it was confirmed that SLC25A51 can efficiently mediate NAD+ entry into mitochondria, while SLC25A47 lacks this activity.

The above results clearly demonstrate that SLC25A51 is the major functional mediator of NAD+ transport in hepatocyte mitochondria, and its expression level directly determines the size of the mitochondrial NAD+ pool.
Molecular mechanism of mitochondrial NAD+regulation of liver regeneration
Through two models — SLC25A51 gene heterozygous deletion (Slc25a51+/-) and hepatocyte-specific overexpression — the study systematically elucidated the core mechanism by which mitochondrial NAD+ regulates liver regeneration:
1. Inhibitory Effect of Slc25a51 Heterozygous Deletion
Slc25a51+/- mice exhibit a significant reduction in liver mitochondrial NAD+ content, and show obvious defects in regeneration after partial hepatectomy (PHx):

• Decreased hepatocyte proliferation rate and delayed recovery of liver-to-body weight ratio;
• Impaired respiratory function dependent on mitochondrial respiratory chain complex I and reduced fatty acid oxidation efficiency;
• Insufficient hepatic ATP production and increased triglyceride accumulation during regeneration.

2. Promotive Effect of SLC25A51 Overexpression
Adeno-associated virus (AAV)-mediated hepatocyte-specific overexpression of SLC25A51 can significantly increase mitochondrial NAD+ content, leading to the following effects:

• Significantly improved liver regeneration efficiency, with markedly increased hepatocyte mitotic index and the proportion of Ki-67 positive cells;
• Reduced lipid accumulation during regeneration, activated the PPAR signaling pathway, and upregulated the expression of genes related to fatty acid synthesis and metabolism;
• Enhanced mitochondrial energy metabolism, increased hepatic ATP and nucleoside triphosphate levels, and maintained the energy charge required for regeneration.

These results indicate that mitochondrial NAD+ provides essential support for liver regeneration through two core pathways: optimizing energy metabolism and regulating lipid homeostasis.
Clinical Translational Value and Research Prospects
The findings of this study hold important clinical translational significance:

1. New therapeutic targets for liver diseases: Targeted regulation of SLC25A51 can precisely elevate mitochondrial NAD+ levels, providing new therapeutic strategies for liver cirrhosis, liver injury and post-liver transplantation repair;
2. Optimization direction for NAD+ supplements: The clinical efficacy of existing NAD+ precursor supplements is inconsistent, which may be related to the ability of drugs to effectively enter mitochondria. In the future, mitochondria-targeted NAD+ delivery systems can be developed to improve treatment efficiency;
3. Expanded research on metabolism-related diseases: Abnormal mitochondrial NAD+ metabolism may be involved in the development and progression of non-alcoholic fatty liver disease, liver cancer and other diseases, and SLC25A51 is expected to become a potential diagnostic marker and therapeutic target for related diseases.

Through rigorous experimental design, this study clarifies the core role of the hepatocyte mitochondrial NAD+ pool in liver regeneration, reveals the SLC25A51-mediated NAD+ transport mechanism, and provides a brand-new perspective for the precise regulation of NAD+ metabolism. With in-depth analysis of the mitochondrial NAD+ metabolic network, it is expected to develop more efficient and specific therapeutic regimens for liver diseases, promoting the translation of NAD+-related therapies from basic research to clinical application.
References
Mukherjee S, Velázquez Aponte RA, Perry CE, Lee WD, Janssen KA, Niere M, Adzika GK, Lu MJ, Chan HR, Zou X, Chen B, Bye N, Xiao T, Yook JS, Salik O, Frederick DW, Gaspar RB, Doan KV, Davis JG, Rabinowitz JD, Wallace DC, Snyder NW, Kajimura S, Cambronne XA, Ziegler M, Baur JA. Hepatocyte mitochondrial NAD+ content is limiting for liver regeneration. Nat Metab. 2025 Dec;7(12):2424-2437.