Induction of Regenerative Cardiac Cells from Cardiomyocytes Using a Two-Drug Combination: In Vitro and In Vivo Evidence

Alapfogalmak

A novel two-drug combination, CHIR99021 and A-485 (2C), effectively reprograms human cardiomyocytes into regenerative cardiac cells (RCCs) in vitro and improves cardiac function in mice following myocardial infarction.

Kivonat

Bibliographic Information:

Wang, Z., et al. (2023). Pharmacologically inducing regenerative cardiac cells by small molecule drugs. Unknown Journal.

Research Objective:

This study investigates the potential of a two-drug combination, CHIR99021 and A-485 (2C), to induce the dedifferentiation of cardiomyocytes (CMs) into regenerative cardiac cells (RCCs) and evaluate its therapeutic efficacy in cardiac regeneration.

Methodology:

• Human embryonic stem cell (hESC)-derived CMs and neonatal rat CMs were treated with 2C in vitro.
• Lineage tracing experiments were conducted to confirm the origin of RCCs.
• In vivo studies involved administering 2C to neonatal and adult mice, followed by myocardial infarction (MI) induction.
• Cardiac function was assessed using echocardiography and magnetic resonance imaging (MRI).
• Bulk RNA sequencing (RNA-seq) and single-cell RNA sequencing (scRNA-seq) were performed to analyze gene expression changes.
• Chromatin immunoprecipitation sequencing (ChIP-seq) was used to investigate epigenetic modifications.

Key Findings:

• 2C treatment effectively induced the dedifferentiation of hESC-derived CMs into RCCs, characterized by sarcomere disassembly, ISL1 expression, and the ability to differentiate into CMs, endothelial cells, and smooth muscle cells.
• Lineage tracing confirmed that 2C-induced RCCs originated from TNNT2+ CMs.
• In vivo administration of 2C induced RCCs in neonatal and adult mice hearts.
• Prophylactic and therapeutic treatment with 2C significantly improved cardiac function and survival rates in mice following MI.
• Mechanistically, CHIR99021 activated Wnt signaling and promoted embryonic gene expression, while A-485 modulated histone acetylation to suppress CM-specific genes and enhance RCC-specific gene expression.

Main Conclusions:

• The 2C combination effectively reprograms CMs into RCCs, representing a promising therapeutic strategy for cardiac regeneration.
• CHIR99021 and A-485 synergistically induce CM dedifferentiation through transcriptional and epigenetic modifications.

Significance:

This study provides compelling evidence for the feasibility of pharmacologically inducing cardiac regeneration by reprogramming endogenous CMs into RCCs.

Limitations and Future Research:

• The study acknowledges the limited proliferative capacity of 2C-induced RCCs compared to natural cardiac progenitors.
• Further research is needed to optimize culture conditions or treatment protocols to enhance RCC proliferation.
• Investigating the long-term effects of 2C treatment on cardiac function and structure is crucial.

Statisztikák

A-485's optimal working concentration was determined to be 0.5 µM.
2C treatment resulted in a ∼3-fold increase in ISL1 expression at both mRNA and protein levels.
There was a notable increase (roughly 1.4-fold) in the number of re-differentiated CMs with clear TNNT2 staining relative to the initial CMs before 2C treatment.
Approximately 0.6% of K9-derived mCherry-negative CMs were permanently labeled with EGFP following 6 days of tamoxifen treatment.
Mice treated with 2C post-MI displayed recovered cardiac function as assessed by left ventricular ejection fraction (LVEF).
2C treatment significantly increased H3K9Ac enrichment at 4,485 gene promoters and H3K27Ac at 2,560 gene promoters, with corresponding decreases in 346 and 1,834 genes, respectively.

Idézetek

"This proof-of-concept discovery demonstrates that a simple combination of small molecule drugs can endow the mammalian heart with regenerative capacity by reprogramming CMs into RCCs."
"This study demonstrates that the synergistic interaction between these two molecules is essential for the reprogramming of endogenous CMs into RCCs, as evidenced by the absence of RCCs in postnatal rat hearts treated with either molecule alone."
"These insights contribute significantly to the understanding of cardiac lineage plasticity and highlight the therapeutic potential of induced multipotent cardiovascular progenitors for heart repair and regeneration."

Főbb Kivonatok

Pharmacologically inducing regenerative cardiac cells by small molecule drugs

by Zhou,W., He,... : www.biorxiv.org 10-26-2023

https://www.biorxiv.org/content/10.1101/2023.10.24.563872v3

Mélyebb kérdések

Could the 2C drug combination be used to regenerate other tissues beyond cardiac muscle?

While the study specifically focuses on the 2C drug combination's efficacy in regenerating cardiac muscle, the findings suggest a broader potential for tissue regeneration. This is because the underlying mechanism involves the reprogramming of terminally differentiated cells, like cardiomyocytes, into an earlier, more plastic state resembling embryonic progenitor cells. These induced pluripotent stem cell-like cells, termed regenerative cardiac cells (RCCs) in the study, exhibit the capacity to differentiate into various cardiovascular lineages, including cardiomyocytes, smooth muscle cells, and endothelial cells.
Extrapolating these findings to other tissues, the 2C combination, or similar epigenetic modifiers, could potentially:

Induce dedifferentiation in other terminally differentiated cell types: The study highlights the role of CHIR99021 in activating the Wnt signaling pathway and A-485 in modulating histone acetylation, both crucial for cellular reprogramming. These pathways are fundamental to development and cell fate decisions across various tissues. Therefore, targeting these pathways with similar drug combinations could potentially induce dedifferentiation in other cell types, like fibroblasts in the skin or skeletal muscle cells.
Promote regeneration in tissues with limited regenerative capacity: Many adult mammalian tissues, like the heart, have limited regenerative potential. The 2C combination's success in inducing a regenerative response in the heart suggests that similar strategies could be explored for tissues like the nervous system, kidneys, or lungs, which also suffer from limited regenerative capacity.
Enhance the generation of induced pluripotent stem cells (iPSCs): The study's findings on cellular reprogramming could be leveraged to improve iPSC generation protocols. By incorporating molecules like CHIR99021 and A-485, the efficiency and speed of reprogramming somatic cells into iPSCs could be enhanced, facilitating research and therapeutic applications.
However, several factors warrant careful consideration:

Tissue specificity: The optimal drug combinations and treatment durations may vary significantly across different tissues. Extensive research is needed to determine the efficacy and safety of such approaches for each specific tissue type.
Off-target effects: Targeting fundamental pathways like Wnt signaling and histone acetylation could have unintended consequences on other cellular processes and tissues. Thorough investigation is crucial to minimize potential off-target effects.
Tumorigenicity: Inducing cellular plasticity raises concerns about uncontrolled cell proliferation and potential tumor formation. Rigorous safety assessments are paramount to ensure the long-term safety of such regenerative therapies.
In conclusion, while the 2C drug combination holds promise for broader tissue regeneration applications, extensive research is necessary to translate these findings into safe and effective therapies for other tissues beyond cardiac muscle.

What are the potential long-term risks and benefits of inducing cardiac regeneration using pharmacological agents?

Pharmacologically inducing cardiac regeneration represents a paradigm shift in treating heart disease, offering potential long-term benefits and risks that warrant careful consideration.
Potential Long-Term Benefits:

Improved Cardiac Function: By regenerating functional cardiomyocytes, pharmacological agents could lead to sustained improvement in cardiac contractility, restoring heart function and improving quality of life for patients with heart failure.
Reduced Scar Tissue: Promoting cardiac regeneration could minimize scar tissue formation after injury, preserving the heart's structural integrity and preventing long-term complications like arrhythmias and heart failure.
Personalized Treatment Strategies: Understanding the molecular mechanisms underlying drug-induced regeneration could pave the way for personalized therapies tailored to individual patient profiles, optimizing treatment efficacy and minimizing adverse effects.
Reduced Healthcare Burden: Successful cardiac regeneration could reduce the need for invasive procedures like heart transplants and mechanical assist devices, potentially alleviating the significant healthcare burden associated with heart disease.
Potential Long-Term Risks:

Tumorigenicity: Inducing cellular plasticity raises concerns about uncontrolled cell proliferation and the potential for tumor formation. Long-term monitoring and safety assessments are crucial to mitigate this risk.
Arrhythmias: Newly regenerated cardiomyocytes might not integrate seamlessly into the existing cardiac conduction system, potentially leading to arrhythmias and disrupting heart rhythm.
Off-Target Effects: Pharmacological agents targeting fundamental cellular pathways could have unintended consequences on other organ systems, leading to unforeseen long-term side effects.
Immune Response: The immune system might recognize newly generated cardiomyocytes as foreign, triggering an immune response and potentially leading to inflammation and rejection.
Mitigating Risks and Maximizing Benefits:

Thorough preclinical studies: Rigorous testing in animal models is essential to assess long-term safety, efficacy, and potential for tumorigenicity before human trials.
Controlled drug delivery: Developing targeted drug delivery systems could help confine the regenerative effects to the heart, minimizing off-target effects on other organs.
Personalized medicine approaches: Tailoring treatment strategies based on individual patient characteristics and genetic profiles could optimize efficacy and minimize adverse effects.
Long-term monitoring: Patients receiving regenerative therapies require close monitoring for potential complications like tumor development, arrhythmias, and immune reactions.
In conclusion, while pharmacologically induced cardiac regeneration holds immense promise for treating heart disease, a balanced approach considering both the potential long-term benefits and risks is crucial. Thorough research, careful patient selection, and long-term monitoring are paramount to ensure the safety and efficacy of these novel therapies.

How might this research influence the development of personalized regenerative therapies for heart disease?

This research, demonstrating the feasibility of pharmacologically inducing cardiac regeneration using the 2C drug combination, holds significant implications for developing personalized regenerative therapies for heart disease. Here's how:
1. Identifying Patient Subgroups:

Genetic Profiling: Analyzing patients' genetic makeup can identify those with variations that might influence their response to the 2C treatment. This could involve studying genes related to the Wnt signaling pathway, histone acetylation, or those involved in cardiac development and regeneration.
Molecular Diagnostics: Developing biomarkers or molecular signatures that predict the likelihood of successful regeneration in response to 2C could help identify patients who would benefit most from this therapy.
2. Tailoring Treatment Regimens:

Dosage Optimization: Genetic and molecular information could guide personalized dosage adjustments of CHIR99021 and A-485, maximizing efficacy while minimizing potential side effects.
Combination Therapies: Combining 2C with other regenerative approaches, such as cell therapy or biomaterial scaffolds, could be tailored based on individual patient needs and disease severity.
3. Monitoring Treatment Response:

Imaging Techniques: Advanced imaging techniques like cardiac MRI and echocardiography can track changes in cardiac structure and function over time, allowing for personalized assessment of treatment efficacy.
Liquid Biopsies: Analyzing blood or other bodily fluids for specific biomarkers could provide real-time information on the regenerative process, enabling adjustments to treatment strategies as needed.
4. Expanding the Regenerative Toolkit:

Drug Discovery: The study's findings on the synergistic effects of CHIR99021 and A-485 could guide the development of novel small-molecule drugs with enhanced regenerative potential and improved safety profiles.
Cell-Based Therapies: The insights gained from understanding how 2C reprograms cardiomyocytes could be applied to enhance the generation and efficacy of cell-based therapies for heart regeneration.
5. Addressing Ethical Considerations:

Informed Consent: As personalized regenerative therapies emerge, ensuring patients fully understand the potential benefits, risks, and limitations of these treatments is crucial.
Access and Equity: Developing strategies to ensure equitable access to these potentially transformative therapies for all patients, regardless of socioeconomic background, is paramount.
In conclusion, this research provides a foundation for developing personalized regenerative therapies for heart disease. By integrating genetic profiling, molecular diagnostics, and tailored treatment strategies, we can move towards a future where heart disease treatment is individualized, maximizing therapeutic benefit while minimizing risks for each patient.