Using stem cell-derived artificial organelles to improve neural oxidative phosphorylation imbalances

In humans, the main molecular energy carrier is adenosine triphosphate (ATP). It is critical for cellular functions all over the body, especially in the brain. Its synthesis, which occurs in the mitochondria, through oxidative phosphorylation (OXPHOS) is a complex, multi-step process. When one or more of the elements of this process malfunction, it may result in imbalances in ATP production or the over production of reactive oxygen species (ROS). This can cause oxidative damage in several cellular biomolecules including proteins and DNA and may eventually end in cell death. In the brain, this imbalance can cause problems which can lead to the loss of neurons, resulting in the damage seen in some degenerative neurological disorders.

Treating problems with their root in errors in ATP synthesis has been a challenge given the complexity of OXPHOS. However, recently Chinese researchers from the Dalian Stem Cell and Precision Medicine Innovation Research Institute have designed a scientific framework ‘Engine Repair Theory’ that uses self-assembling organelles (SAOs), derived from neural stem cells, that can carry out the main pathways of ATP production to address mitochondrial imbalances.

The framework “offers a transformative approach in the field of cell-free organelle-based therapy, particularly focusing on the use of neural stem cell-derived oxidative phosphorylating artificial organelles (SAOs) to restore mitochondrial energy homeostasis and mitigate oxidative stress,” said Jing Liu, one of the authors of the paper and a professor at the Dalian Stem Cell and Precision Medicine Innovation Research Institute.

Their research was published in Nature Communications on Sept. 8.

While the ATP synthesis pathway OXPHOS is a very useful therapeutic target, its complexity has made it a difficult target to hit. One promising area of research is in the use of self-assembling biomimetic organelles that can mimic specific biological processes. SAOs can be made self-assembling using the property of self-assembly found naturally in cell membranes. This approach has shown promise in other biological systems.

The researchers designed a rigorous manufacturing process to ensure high quality, consistent organelles. They grew homogeneous cellular clones, introduced disrupted cell membranes from neural stem cells to induce self-assembly and enriched them with OXPHOS components to create SAOs capable of synthesizing ATP.

They tested the cell populations selected to guarantee they were homogenous. This allows for more certainty in how the organelles will work as there is no variation to confound the results. Mouse neural tissue was treated with the SAOs and then tested, and they found an increase in ATP production. This increase was proof that the SAOs were able to work as functioning units and to undergo OXPHOS.

In vitro testing was done using several types of neural cells that had undergone one of three different types of oxidative stress. In the cell cultures that had been treated with the SAOs, there was reduced cell damage and lowered ROS levels in the mitochondria. The organelles also promoted cell regeneration via neural differentiation, maturation and function.

The in vivo tests used rats with middle cerebral artery occlusion (MCO), which occurs when there is a block in blood flow in the cerebral artery. In rats treated with SAO, there was clear evidence that treatment promoted regeneration and repair of several different cell types, increased OXPHOS production and restored cellular metabolic equilibrium. It reduced cell damage caused by the occlusion and reduced ROS levels in the mitochondria. “These findings suggest that the SAOs alleviated the neurological deficit symptoms in rats,” said Jiayi Wang, first author and a researcher at Stem Cell Clinical Research Center, The First Affiliated Hospital of Dalian Medical University. Finally, safety tests done in vivo showed that SAO had a good safety profile when looking at tumorigenicity, associated immune response and toxicity.

“The key takeaway from this study is the successful demonstration of the ‘Engine Repair Theory,’” where multi-target modulation of OXPHOS imbalance is pivotal for repairing neural damage caused by ischemia-reperfusion. The development of a high-throughput, homogeneous production process for SAOs enables precise regulation of mitochondrial energy metabolism and OXPHOS homeostasis. This represents a significant breakthrough in cell-free organelle-based therapy, offering a novel approach for treating neurodegenerative conditions. “The research lays the foundation for using artificial organelles to replace or supplement traditional stem cell therapies, bringing renewed hope for efficient and targeted neural repair,” Liu said.

Looking forward to where this research may lead, “the ultimate goal is to advance the development of next-generation cell-free therapies, facilitating a paradigm shift from traditional stem cell-based treatments to organelles-focused therapeutic approaches that offer precise, efficient, and safer solutions,” Liu said.

Other contributors include Mengke Zhao, Meina Wang, Dong Fu, Lin Kang, Yu Xu, Liming Shen, Shilin Jin, and Liang Wang, all are from the Stem Cell Clinical Research Center, The First Affiliated Hospital of Dalian Medical University and the 2National Local Joint Engineering Laboratory, The First Affiliated Hospital of Dalian Medical University, and 3National Genetic Test Center, The First Affiliated Hospital of Dalian Medical University and the 4Liaoning Key Laboratory of Frontier Technology of Stem Cell and Precision Medicine, Dalian Innovation Institute of Stem Cell and Precision Medicine, Dalian City, Liaoning Province, PR China.

This work was supported by the National Nature Science Foundation of China (82072953), National Health and Family Planning Commission and Food and Drug Administration (CMR-20161129- 1003), Science and Technology Program of Liaoning Province ([2021] 49), Outstanding Young Talents of Dalian City (2021RJ12) and Science and Technology Personnel Innovation of Dalian city ([2022]25). We acknowledge the use of images from Servier Medical Art, licensed under CC BY 4.0. The images were modified for this publication. For original images and licensing details, please visit https://smart. servier.com. We would like to thank Prof. Bingbing Sun (Dalian University of Technology) and Prof. Ran Zhang (Chinese Academy of Medical Sciences) for help with the mass manufacture SAOs and cell function studies.