Glycogen Engineering Improves the Starvation Resistance of Mesenchymal stem cells and their Therapeutic Efficacy in Pulmonary Fibrosis
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Curated by eLife
eLife Assessment
This valuable study presents a novel approach to enhance the therapeutic potential of mesenchymal stromal cells (MSCs) by genetically modifying their glycogen synthesis pathway, resulting in increased glycogen accumulation and improved cell survival under starvation conditions, particularly in the context of experimental pulmonary fibrosis. The methods and findings are generally solid and could be strengthened by investigating the kinetics of persistence, the immunomodulatory effects, and the underlying improved mechanism of action of MSCs in this pulmonary fibrosis model. If confirmed, this approach could suggest potential methods to improve the therapeutic functionality of MSCs in cell therapy strategies.
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Abstract
Abstract
Mesenchymal stem cells (MSCs) are widely used in regenerative medicine, including the treatment of pulmonary fibrosis. However, implanted MSCs disappear within days, constraining therapeutic efficacy, which is largely attributed to nutrient deprivation. In this study, we established glycogen metabolism engineering strategies in mammalian cells. By expressing a functionally optimized glycogen synthase (GYSmut), MSCs could accumulate large amounts of glycogen rapidly as a reserve substance. Glycogen engineering significantly improved the survival of MSCs during starvation both in vitro and in vivo, enhancing cell viability post-implantation and their therapeutic efficacy in pulmonary fibrosis. Glycogen-engineered MSCs may serve as chassis cells for further applications. Our research highlights the importance of glucose metabolism regulation in cell-based therapy and demonstrates the great potential for the metabolic engineering of MSCs and other therapeutic cells.
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eLife Assessment
This valuable study presents a novel approach to enhance the therapeutic potential of mesenchymal stromal cells (MSCs) by genetically modifying their glycogen synthesis pathway, resulting in increased glycogen accumulation and improved cell survival under starvation conditions, particularly in the context of experimental pulmonary fibrosis. The methods and findings are generally solid and could be strengthened by investigating the kinetics of persistence, the immunomodulatory effects, and the underlying improved mechanism of action of MSCs in this pulmonary fibrosis model. If confirmed, this approach could suggest potential methods to improve the therapeutic functionality of MSCs in cell therapy strategies.
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Reviewer #1 (Public review):
Summary:
This study provides the first evidence that glucose availability, previously shown to support cell survival in other models, is also a key determinant for post-implantation MSC survival in the specific context of pulmonary fibrosis. To address glucose depletion in this context, the authors propose an original, elegant, and rational strategy: enhancing intracellular glycogen stores to provide transplanted MSCs with an internal energy reserve. This approach aims to prolong their viability and therapeutic functionality after implantation.
Strengths:
The efficacy of this metabolic engineering strategy is robustly demonstrated both in vitro and in an orthotopic mouse model of pulmonary fibrosis.
Comments and questions for clarification:
(1) Glycogen biosynthesis typically involves several enzymes. In …
Reviewer #1 (Public review):
Summary:
This study provides the first evidence that glucose availability, previously shown to support cell survival in other models, is also a key determinant for post-implantation MSC survival in the specific context of pulmonary fibrosis. To address glucose depletion in this context, the authors propose an original, elegant, and rational strategy: enhancing intracellular glycogen stores to provide transplanted MSCs with an internal energy reserve. This approach aims to prolong their viability and therapeutic functionality after implantation.
Strengths:
The efficacy of this metabolic engineering strategy is robustly demonstrated both in vitro and in an orthotopic mouse model of pulmonary fibrosis.
Comments and questions for clarification:
(1) Glycogen biosynthesis typically involves several enzymes. In this context, could the authors comment on the effect of overexpressing a single enzyme - especially a mutant version - on the structure or quality of the glycogen synthesized?
(2) Regarding the in vitro starvation experiments (Figure 2C), what oxygen conditions (pO₂) were used? Are these conditions physiologically relevant and representative of the in vivo lung microenvironment?
(3) In the in vitro model, how many hours does it take for the intracellular glycogen reserve to be completely depleted under starvation conditions?
(4) For the in vivo model, is there a quantitative analysis of the survival kinetics of the transplanted cells over time for each group? This would help to better assess the role and duration of glycogen stores as an energy buffer after implantation.
(5) Finally, the study was performed in male mice only. Could sex differences exist in the efficacy or metabolism of the engineered MSCs? It would be helpful to discuss whether the approach could be expected to be similarly effective in female subjects.
(6) The number of mice for each group and time point should be specified.
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Reviewer #2 (Public review):
Summary:
In this article, the authors investigate enhancing the therapeutic and regenerative properties of mesenchymal stem cells (MSCs) through genetic modification, specifically by overexpressing genes involved in the glycogen synthesis pathway. By creating a non-phosphorylatable mutant form of glycogen synthase (GYSmut), the authors successfully increased glycogen accumulation in MSCs, leading to significantly improved cell survival under starvation conditions. The study highlights the potential of glycogen engineering to improve MSC function, especially in inflammatory or energy-deficient environments. However, critical gaps in the study's design, including the lack of validation of key findings, limited differentiation assessments, and missing data on MSC-GYSmut resistance to reactive oxygen species …
Reviewer #2 (Public review):
Summary:
In this article, the authors investigate enhancing the therapeutic and regenerative properties of mesenchymal stem cells (MSCs) through genetic modification, specifically by overexpressing genes involved in the glycogen synthesis pathway. By creating a non-phosphorylatable mutant form of glycogen synthase (GYSmut), the authors successfully increased glycogen accumulation in MSCs, leading to significantly improved cell survival under starvation conditions. The study highlights the potential of glycogen engineering to improve MSC function, especially in inflammatory or energy-deficient environments. However, critical gaps in the study's design, including the lack of validation of key findings, limited differentiation assessments, and missing data on MSC-GYSmut resistance to reactive oxygen species (ROS), necessitate further exploration.
Strengths:
(1) Novel Approach: The study introduces an innovative method of enhancing MSC function by manipulating glycogen metabolism.
(2) Increased Glycogen Storage: The genetic modification of GYS1, resulting in GYSmut, significantly increased glycogen accumulation, leading to improved MSC survival under starvation, which has strong implications for enhancing MSC therapeutic properties in energy-deficient environments.
(3) Potential Therapeutic Impact: The findings suggest significant therapeutic potential for MSCs in conditions that require improved survival, persistence, and immunomodulation, especially in inflammatory or energy-limited settings.
(4) In Vivo Validation: The in vivo murine model of pulmonary fibrosis demonstrated the improved survival and persistence of MSC-GYSmut, supporting the translational potential of the approach.
Weaknesses:
(1) Lack of Differentiation Assessments: The study did not evaluate key MSC differentiation pathways, including chondrogenic and osteogenic differentiation. The absence of analysis of classical MSC surface markers and multipotency limits the understanding of the full potential of MSC-GYSmut.
(2) Missing Validation of RNA Sequencing Data: Although RNA sequencing data revealed promising transcriptomic changes in chondrogenesis and metabolic pathways, these findings were not experimentally validated, limiting confidence.
(3) Lack of ROS Resistance Analysis: Resistance to reactive oxygen species (ROS), an important feature for MSCs under regenerative conditions, was not assessed, leaving out a critical aspect of MSC function.
(4) Inconsistencies in In Vivo Data: There is a discrepancy between the number of animals shown in the figures and the graph (three individuals vs. five animals), as well as missing details on how luciferase signal intensity was quantified, requiring further clarification.
(5) Limited Exploration of Immunosuppressive Properties: The study did not address the immunosuppressive functions of MSC-GYSmut, which are critical for MSC-based therapies in clinical settings.
Conclusion:
The study presents an exciting new direction for enhancing MSC function through glycogen metabolism engineering. While the results show promise, key experiments and validations are missing, and several areas, such as differentiation capacity, ROS resistance, and immunosuppressive properties, require further investigation. Addressing these gaps would solidify the conclusions and strengthen the potential clinical applications of MSC-GYSmut in regenerative medicine.
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