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Mitochondria-targeted peptide SS-31, also known as elamipretide, is a small-molecule peptide with specific mitochondria-targeting functionality. In recent years, as research into the mechanisms of aging has deepened, the critical role of mitochondrial dysfunction in the aging process has become increasingly evident. SS-31, with its unique mitochondrial targeting properties and ability to regulate mitochondrial function, holds promise for anti-aging applications.
Figure 1 Respiration and membrane potential response to ADP stimulation in isolated muscle mitochondria require supplemental cytochrome c and hexokinase clamp. ADP/ATP transport pathway and binding of ELAM to ATP Synthase and ANT.
I. Overview of the Relationship Between Mitochondria and Aging
Mitochondria, as the cell's powerhouse, are responsible for synthesizing the majority of ATP within the cell and are also involved in various metabolic regulation and signal transduction processes. As age increases, mitochondrial function gradually declines, which is one of the key markers of aging. Mitochondrial dysfunction manifests in multiple ways, including the accumulation of mutations in mitochondrial DNA (mtDNA), reduced activity of mitochondrial respiratory chain complexes, leading to insufficient energy production; increased production of reactive oxygen species (ROS) by mitochondria, triggering oxidative stress damage, which further disrupts intracellular biomolecules such as proteins, lipids, and nucleic acids, accelerating cellular aging and death. Abnormal mitochondrial function also disrupts intracellular calcium homeostasis, interfering with normal cellular physiological functions. These changes interact with each other, forming a vicious cycle that collectively drives the aging process.
II. Mechanism of Action of SS-31
Improving Mitochondrial Energy Metabolism
Enhancing ADP Sensitivity: During aging, mitochondrial sensitivity to ADP decreases, impairing ATP synthesis efficiency. Studies indicate that SS-31 can directly bind to the mitochondrial ADP transporter ANT, increasing ANT's uptake of ADP, thereby enhancing mitochondrial sensitivity to ADP. In aged muscle mitochondria, SS-31 treatment increased ADP uptake via ANT, thereby enhancing mitochondrial respiration under ADP stimulation, increasing ATP production, and improving energy metabolism in aged muscle mitochondria.
Regulation of mitochondrial respiratory chain complexes: Mitochondrial respiratory chain complexes are key components of mitochondrial energy production. During aging, the activity of respiratory chain complexes often decreases. SS-31 may maintain or enhance the activity of respiratory chain complexes by stabilizing their structural integrity or regulating the expression of related proteins. Previous studies have observed that in aging-related mitochondrial dysfunction models, the activity of respiratory chain complexes is restored after SS-31 treatment, suggesting its positive regulatory effect on the mitochondrial respiratory chain.
Figure 2 SS-31 treatment reverses cardiac aging phenotypes
Reducing oxidative stress damage
Reducing ROS production: Mitochondria are one of the primary sources of ROS within cells, and ROS production increases in mitochondria during aging. SS-31 can reduce ROS production through multiple pathways. It improves mitochondrial energy metabolism, making the mitochondrial electron transport chain more efficient and reducing electron leakage, thereby lowering ROS production. SS-31 may directly act on the mitochondrial membrane, altering its physical properties, reducing oxidative damage sites on the membrane, and inhibiting ROS production. In cardiac cells from aged mice, treatment with SS-31 significantly reduced mitochondrial ROS levels, indicating its efficacy in reducing ROS production.
Regulation of the antioxidant enzyme system: Cells contain an antioxidant enzyme system, including superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px), which are responsible for scavenging ROS. Research suggests that SS-31 may enhance cellular antioxidant defense capabilities by upregulating the expression or activity of these antioxidant enzymes. In certain cellular models, treatment with SS-31 resulted in a significant increase in the activity of SOD and GSH-Px, aiding in the timely clearance of excess ROS within cells and alleviating oxidative stress-induced damage.
Maintaining mitochondrial membrane stability
Binding to mitochondrial membrane phospholipids: SS-31 has a unique structure that allows it to preferentially bind to regions of the mitochondrial membrane rich in phosphatidylserine. Phosphatidylserine is a phospholipid specific to the mitochondrial membrane and is crucial for maintaining the structural and functional integrity of the mitochondrial membrane. During aging, phosphatidylserine is susceptible to oxidative damage, leading to abnormalities in mitochondrial membrane structure and function. After binding to phosphatidylserine, SS-31 may protect phosphatidylserine from oxidation while stabilizing mitochondrial membrane structure, preventing a decrease in mitochondrial membrane potential and the opening of the mitochondrial permeability transition pore (mPTP). In vitro experiments have shown that SS-31 effectively inhibits oxidative stress-induced decreases in mitochondrial membrane potential and mPTP opening, thereby maintaining mitochondrial membrane stability.
Regulation of membrane-associated proteins: The mitochondrial membrane contains various proteins involved in mitochondrial material transport, energy metabolism, and signal transduction processes. SS-31 may indirectly influence mitochondrial membrane stability by regulating the activity or expression of these membrane-associated proteins. It can regulate the function of voltage-dependent anion channels (VDAC), which are important channel proteins on the outer mitochondrial membrane and play a key role in maintaining material exchange and signal transmission between mitochondria and the cytoplasm. SS-31's regulation of VDAC helps maintain normal mitochondrial membrane function and stability.
III. Specific effects of SS-31 in anti-aging
Effects on the cardiovascular system
Improving cardiac function: The heart is a high-energy-demand organ, and mitochondrial function is critical for cardiac function. In aged mice, the heart often exhibits age-related issues such as diastolic dysfunction. Scientists have found that after 8 weeks of SS-31 treatment, the diastolic function of aged mice's hearts was significantly improved. This improvement was accompanied by the normalization of mitochondrial proton leakage in myocardial cells, reduced mitochondrial ROS levels, decreased cardiac protein oxidation levels, and a shift in the thiol redox state of proteins toward a more reduced state. SS-31 also increased the phosphorylation level of cMyBP-C Ser282 in myocardial cells, which is closely associated with improved cardiac diastolic function.
Figure 3 Learning impairment in aging mice induced by sleep deprivation was prevented with SS31.
Vascular protection: During aging, endothelial cell function is impaired, vascular elasticity decreases, and cardiovascular diseases are more likely to occur. In a hypertension-induced microbleeding model in the brains of aged mice, treatment with SS-31 exhibits significant vascular protective effects. It reduces mitochondrial free radical production induced by hypertension, alleviates oxidative stress damage to the vascular wall, thereby significantly delaying the onset of microbleeding and reducing its incidence. Therefore, SS-31 plays an important role in maintaining the normal structure and function of blood vessels and preventing age-related vascular diseases.
Effects on the nervous system
Alleviating cognitive dysfunction: As age increases, nervous system function gradually declines, and cognitive dysfunction such as memory impairment and reduced learning ability becomes increasingly prominent. Mitochondrial dysfunction plays a crucial role in the onset and progression of neurodegenerative diseases and cognitive dysfunction. In a model of cognitive dysfunction induced by isoflurane in aged mice, SS-31 can reverse mitochondrial dysfunction, rescue isoflurane-induced cognitive deficits. SS-31 promotes the regulation of BDNF signaling, reverses the downregulation of synaptic plasticity-related proteins such as synaptophysin, PSD-95, and p-CREB, and upregulates NR2A, NR2B, CaMKIIα, and CaMKIIβ, thereby enhancing synaptic plasticity and protecting cognitive function.
Alleviating the negative effects of sleep deprivation: Sleep deprivation is a common stressor, and its negative effects on the nervous system become more pronounced with age, leading to cognitive dysfunction and increased risk of neurodegenerative diseases. In 20-month-old aged mice, SS-31 (3 mg/kg) was administered via subcutaneous injection daily for 4 consecutive days, with 4 hours of sleep deprivation on the last two days. Results showed that sleep-deprived mice treated with SS-31 exhibited no significant impairment in learning ability, with restored brain mitochondrial ATP levels and synaptic plasticity-regulating proteins, as well as reduced levels of reactive oxygen species (ROS) and inflammatory cytokines in the hippocampus. This suggests that SS-31 may have potential therapeutic benefits in mitigating the adverse neurological effects of short-term sleep deprivation in aged mice.
Effects on the renal system: In the kidneys of aged mice, glomerulosclerosis is associated with mitochondrial damage in glomerular epithelial cells. After 8 weeks of SS-31 treatment in 26-month-old aged mice, SS-31 improved age-related mitochondrial morphology and alleviated glomerulosclerosis. Specifically, it reduced the expression of aging markers (p16, aging-associated β-Gal), increased the density of apical epithelial cells, and decreased the expression of markers of parietal epithelial cell activation (collagen IV, pERK1/2, and α-smooth muscle actin). Although SS-31 did not affect podocyte density, it reduced markers of podocyte damage (desmin), improved cytoskeletal integrity (synaptophysin), and was accompanied by higher glomerular endothelial cell density (CD31). This suggests that short-term SS-31 treatment also has a protective effect on glomerular mitochondria and improves glomerular structure.
Effects at the cellular level
Delaying cellular aging: In cell experiments, H₂O₂ was used to induce a stress-induced aging model in HEK293T cells, followed by intervention with SS-31. The results showed that the SA-β-gal positive rate was significantly reduced in the SS-31 group, indicating a decrease in cellular aging levels. Additionally, the intracellular ROS fluorescence intensity decreased, mitochondrial membrane potential increased, and ATP levels rose in the SS-31 group. Protein immunoblotting analysis revealed that the expression levels of P53, P21, and Acetyl-p53 proteins were higher in the model group compared to the control group, while the SS-31 group showed a decrease compared to the model group. Conversely, the expression level of Sirt1 protein was lower in the model group compared to the control group, while the SS-31 group showed an increase compared to the model group. Thus, it can be concluded that SS-31 can delay HEK293T cell aging by improving mitochondrial function and regulating the expression of aging-related proteins within cells.
Protection against oxidative stress-induced cellular damage: In the H₂O₂-induced oxidative stress damage model of ARPE-19 cells, treatment with SS-31 significantly increased cell survival rate, reduced intracellular ROS levels, decreased the proportion of cells with reduced mitochondrial membrane potential, markedly lowered PI positivity rate (reflecting the extent of cell death), and significantly attenuated the upregulation of RIP3 protein expression. These results indicate that SS-31 has a significant protective effect against H₂O₂-induced oxidative stress damage in ARPE-19 cells, which may aid in counteracting age-related damage.
Conclusion
The mitochondrial-targeted peptide SS-31 contributes to anti-aging. From its multifaceted regulatory mechanisms on mitochondrial function to its significant anti-aging effects at various systemic and cellular levels, it demonstrates potential as an important anti-aging strategy.
Sources
[1] Patai R, Patel K, Csik B, et al. Aging, mitochondrial dysfunction, and cerebral microhemorrhages: a preclinical evaluation of SS-31 (elamipretide) and development of a high-throughput machine learning-driven imaging pipeline for cerebromicrovascular protection therapeutic screening[J]. Geroscience, 2025.DOI:10.1007/s11357-025-01634-5.
[2] Pharaoh G, Kamat V, Kannan S, et al. The mitochondrially targeted peptide elamipretide (SS-31) improves ADP sensitivity in aged mitochondria by increasing uptake through the adenine nucleotide translocator (ANT)[J]. Geroscience, 2023,45(6):3529-3548.DOI:10.1007/s11357-023-00861-y.
[3] Chiao Y A, Zhang H, Sweetwyne M, et al. Late-life restoration of mitochondrial function reverses cardiac dysfunction in old mice[J]. Elife, 2020,9.DOI:10.7554/eLife.55513.
[1] Wu J, Dou Y, Ladiges W C. Adverse Neurological Effects of Short-Term Sleep Deprivation in Aging Mice Are Prevented by SS31 Peptide[J]. Clocks & Sleep, 2020,2(3):325-333.DOI:10.3390/clockssleep2030024.
[4] Sweetwyne M T, Pippin J W, Eng D G, et al. The mitochondrial-targeted peptide, SS-31, improves glomerular architecture in mice of advanced age[J]. Kidney International, 2017,91(5):1126-1145.DOI:10.1016/j.kint.2016.10.036.
[5] Wu J, Zhang M, Li H, et al. BDNF pathway is involved in the protective effects of SS-31 on isoflurane-induced cognitive deficits in aging mice[J]. Behavioural Brain Research, 2016,305:115-121.DOI:10.1016/j.bbr.2016.02.036.
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