Recent Advances in Cellular Aging, Retinal Preservation, and Inflammation

Sex Differences in the Aging Heart and the Role of Glutathione Precursors

Aging in the heart is marked by dysregulated metabolism, inflammation, and fibrosis. One of the hallmarks of cardiac aging is the altered redox balance resulting from reduced levels of the natural antioxidant glutathione (GSH). A recent study investigated the effects of dietary supplementation with glutathione precursors—glycine and N-acetyl cysteine (collectively known as GlyNAC)—on cardiac function in aged mice. Notably, the researchers discovered that a GlyNAC-enriched diet administered for 12 weeks to 21-month-old mice (an age comparable to 70-year-old humans) produced distinct, sex-dependent responses with respect to mitochondrial function and diastolic performance.

In-depth proteomic analyses revealed that GlyNAC supplementation led to an upregulation of key mitochondrial proteins related to electron transport and ATP synthesis in male mouse hearts. For example, increases in NADH dehydrogenase components and proteins involved in fatty acid oxidation—such as carnitine palmitoyltransferase 1b (CPT1b) and carnitine acetyltransferase (CrAT)—were particularly pronounced in males, whereas female hearts exhibited minimal changes in these pathways. This dichotomy suggests that the male heart, which tends to show more oxidative stress and impaired mitochondrial performance with age, may benefit more from reinforcing the intracellular GSH pool. The study also identified a modest improvement in diastolic function (as measured by tissue Doppler ratios such as E/eʹ) in GlyNAC-fed males. Interestingly, however, GlyNAC supplementation in old females unexpectedly diminished exercise performance, underscoring the importance of considering sexual dimorphism when assessing the efficacy of antioxidant interventions.

The study’s findings imply that differences in the baseline antioxidant capacity and the adaptive metabolic responses between sexes could be crucial determinants in therapeutic outcomes. Data presented in the study’s proteomic heatmaps and transmission electron microscopy images showed that old male hearts often presented with disorganized mitochondrial cristae and increased protein oxidation. After GlyNAC supplementation, these markers improved markedly in males while remaining unchanged in females. Although the precise mechanisms remain under investigation, it appears that restoring GSH levels via GlyNAC not only mitigates oxidative damage but may also normalize aspects of mitochondrial dynamics, such as the expression of proteins involved in mitochondrial fission and fusion.

Overall, these findings reinforce the concept that targeted nutritional interventions may reverse several aging hallmarks in a sex-specific manner. In clinical terms, these insights could pave the way for personalized dietary strategies to improve cardiac function and extend health span in older adults.

Preservation of Human RPE Cell Suspensions: The Role of Sugars

The integrity and viability of cultured cells are fundamental to regenerative medicine, particularly when considering cell transplantation for retinal diseases such as age-related macular degeneration (AMD). One challenge in this field is ensuring high cell survival rates when RPE cells are stored or transported in suspension. A recent investigation focused on the effect that different sugars present in the culture medium have on preserving human RPE cell suspensions. Researchers compared the conventional glucose-based medium against media in which glucose was replaced with alternative sugars such as fructose, galactose, and mannose while keeping molar concentrations and osmotic pressures equivalent.

After long-term cultures of adherent RPE cells, the survival rate was evaluated via trypan blue exclusion assays and quantification of lactate dehydrogenase (LDH) release. The data revealed that RPE cells maintained in fructose-containing media demonstrated a significantly higher viability compared with those in glucose- or galactose-based media. Under suspension conditions—characterized by high levels of cellular stress due in part to detachment-induced apoptosis (anoikis)—the fructose medium again outperformed others by yielding higher viable cell counts after 24 hours.

Chromatographic analyses provided additional insight into metabolic alterations during preservation. In cultures maintained with fructose, the levels of key glycolytic intermediates such as lactate and pyruvate were lower compared to glucose-containing media. These findings point toward a distinct metabolic pathway that may reduce the susceptibility of RPE cells to hypoxic stress. Complementary fluorescence studies using the hypoxia-detecting probe mono azo rhodamine (MAR) also indicated that cells stored in fructose media exhibited lower levels of hypoxia, further translating into better cell viability.

The significance of these results is profound for retinal transplantation therapies. By optimizing the sugar composition in the culture and preservation media, cell survival rates during storage and transportation can be improved, thereby enhancing the ultimate success rate of cell-based interventions for retinal diseases. Moreover, modifying culture conditions by substituting glucose with fructose represents a relatively low-risk and cost-effective alteration that does not require complex genetic or pharmacological modifications.

A table summarizing the key findings from the RPE cell preservation study is provided below.

Data from the table illustrate that fructose-containing media supports RPE cell survival more effectively, likely by diminishing hypoxia and attenuating anaerobic metabolic stress pathways.

Complement System Alterations in Retinal Vein Occlusion During Ranibizumab Treatment

Retinal vein occlusion (RVO) is the second most common retinal vascular disorder, often leading to macular edema and significant vision loss. Treatments such as the intravitreal injection of anti-vascular endothelial growth factor (VEGF) agents like ranibizumab have become the standard of care. Beyond hindering neovascularization, these treatments may also modulate inflammatory processes in the eye. A recent study comprehensively monitored the dynamic changes in complement proteins in both the aqueous humor (AH) and plasma of RVO patients undergoing ranibizumab therapy.

The investigators obtained serial AH and plasma samples from a cohort of RVO patients and analyzed the concentrations of various complement components—including C1q, C2, C3, C4, C5, and regulatory proteins such as complement factor H and I—as well as VEGF-A levels during treatment. The results demonstrated a significant reduction in several complement proteins, particularly those involved in the classical (C1q, C2, C4) and alternative (CFH, CFB, CFD, CFI) pathways, over the course of treatment. A marked decrease in VEGF-A levels was also observed, correlating with improvements in central retinal thickness (CRT).

Interestingly, when comparing eyes affected by branch retinal vein occlusion (BRVO) to those with central retinal vein occlusion (CRVO), additional differences were observed. At baseline, CRVO patients exhibited higher levels of several complement factors than BRVO patients, reflecting a more robust inflammatory and ischemic injury. Moreover, a significant positive correlation was found between baseline levels of specific proteins (e.g., C5a, CFH) and the subsequent decrease in CRT after repeated ranibizumab injections, suggesting that these markers might serve as predictive biomarkers for treatment response in RVO.

These findings underscore that anti-VEGF therapy may not only mitigate aberrant angiogenesis but also alter the intraocular immune environment, reducing complement activation and subsequent inflammatory cascades. By carefully monitoring these biomarkers, clinicians may be better equipped to predict which patients will achieve a favorable anatomical and functional outcome, ultimately guiding personalized treatment regimens.

Lysosomal Dysfunction and Oxidative Stress in RPE Degeneration

Maintaining retinal health requires efficient autophagy and lysosomal degradation, particularly in the RPE, where daily phagocytosis of photoreceptor outer segments generates a substantial load of degradative substrates. Lysosome-associated membrane protein 2 (Lamp2) is a key protein integral to lysosomal function, autophagosome-lysosome fusion, and overall cellular homeostasis. Deficiencies in Lamp2 have been implicated in several multisystem disorders, including Danon disease, and are now gaining interest for their potential role in retinal degeneration.

A recent study examined the susceptibility of young Lamp2 knockout (KO) mice to oxidative stress–induced RPE degeneration using a low dose of sodium iodate (NaIO₃). While wild-type mice were largely resistant to NaIO₃-induced damage at this low dose, Lamp2 KO mice exhibited dramatic RPE degeneration accompanied by loss of epithelial polarity, outer nuclear layer (ONL) thinning, and ultrastructural defects such as disrupted mitochondrial cristae in RPE cells. In addition to morphological deterioration determined by ZO-1 immunostaining and electron microscopy, molecular analyses revealed a robust inflammatory response. Early proliferation of resident microglia, followed by the infiltration of monocyte-derived macrophages, was noted in Lamp2-deficient retinas, along with increased retinal expression of proinflammatory cytokines (e.g., IL-1β, IL-6) and chemokines (e.g., MCP-1, MIP-1β).

Moreover, pretreatment of Lamp2 KO mice with clodronate—a macrophage-depleting agent—resulted in a significant attenuation of RPE degeneration. This finding highlights the integral role of macrophage-mediated neurotoxic inflammation in the pathology of Lamp2 deficiency under oxidative conditions. Thus, defective lysosomal function, in combination with oxidative stress, significantly compromises RPE homeostasis and promotes retinal degeneration. This model provides valuable insights into age-related retinal disorders such as AMD, where impaired autophagy and chronic inflammation are central features.

Conclusion

Recent research into cellular aging, retinal preservation, and inflammation has uncovered several interrelated mechanisms that contribute to tissue degeneration. Dietary interventions that replenish antioxidants like glutathione via GlyNAC supplementation hold promise for reversing deficits in mitochondrial function and diastolic performance in the aging heart—a benefit that appears to be sex-specific. In parallel, optimizing culture conditions, such as substituting glucose with fructose in preservation media, enhances the viability of human RPE cell suspensions, a development that could significantly improve regenerative therapies for vision loss. Additionally, dynamic alterations in complement protein levels during anti-VEGF treatments for RVO reveal that modulating the intraocular immune environment is as critical as controlling aberrant angiogenesis. Finally, studies on Lamp2 deficiency have highlighted the delicate balance between lysosomal function, autophagy, and inflammation in the retina, demonstrating that even mild oxidative stress can precipitate severe RPE degeneration when lysosomal degradation is compromised.

Collectively, these studies emphasize the importance of understanding cellular metabolism and immune responses in the context of aging and degenerative diseases. As researchers continue to elucidate these complex pathways, there is hope for the development of targeted, personalized interventions to preserve tissue function and improve quality of life in aging populations.

Frequently Asked Questions (FAQ)

What is GlyNAC and how does it affect the aging heart?
GlyNAC is a combination of glycine and N-acetyl cysteine, which are precursors to glutathione (GSH). GSH is a critical antioxidant that declines with age. Supplementation with GlyNAC has been shown to enhance mitochondrial function, reduce oxidative stress, and modestly improve diastolic heart function—especially in older male mice—as it helps restore redox balance and normalize mitochondrial protein expression.

Why is fructose considered better than glucose for preserving RPE cell suspensions?
Studies indicate that RPE cells stored in a fructose-based medium exhibit higher viability compared with conventional glucose mediFructose appears to modulate cellular metabolism by decreasing the levels of anaerobic metabolic intermediates like lactate and pyruvate and reducing hypoxia as evidenced by lower MAR fluorescence. This improved metabolic environment helps cells better tolerate the stress of suspension storage.

How does ranibizumab treatment affect complement proteins in patients with retinal vein occlusion?
Ranibizumab, an anti-VEGF medication, not only reduces VEGF-A levels but also decreases the concentrations of various complement proteins (such as C1q, C2, C4, CFH, and others) in the aqueous humor of RVO patients. The reduction of these proteins correlates with improvements in retinal thickness, highlighting the role of complement-mediated inflammation in the pathophysiology of RVO and treatment response.

What role does Lamp2 play in retinal health?
Lamp2 is essential for proper lysosomal function and autophagy, processes that are critical for removing damaged cellular components, including oxidized biomolecules, in the RPE. Lamp2 deficiency makes RPE cells more vulnerable to oxidative stress and promotes the infiltration of inflammatory cells such as microglia and macrophages, leading to accelerated retinal degeneration.

Are the therapeutic effects of GlyNAC supplementation the same for both sexes?
No, the effects appear to be sex-specific. In the study reviewed, older male mice showed improved mitochondrial function and diastolic performance upon GlyNAC supplementation, whereas older female mice did not show similar improvements and, in some measures, even exhibited reduced exercise performance. These differences likely stem from inherent sex-based variations in antioxidant capacity and metabolic regulation.

References

1. Angelini, A., Garcia Marquez, G., Malovannaya, A., et al. (2024). Sex differences in response to diet enriched with glutathione precursors in the aging heart. The Journals of Gerontology: Series A, Biological Sciences and Medical Sciences

2. Wu, G. (2025). Effect of the sugar present in the culture medium on the preservation of human RPE cell suspensions. PubMed. Retrieved from https://pubmed.ncbi.nlm.nih.gov/11798337/

3. [Unknown Author]. (2025). Dynamic complement protein changes in aqueous humor and plasma of patients with retinal vein occlusion during ranibizumab treatment. Investigative Ophthalmology & Visual Science. Retrieved from https://doi.org/10.2147/JIR.S502481

4. Notomi, S., Fukuda, Y., Maehara, Y., et al. (2025). Lamp2 deficiency enhances susceptibility to oxidative stress–induced RPE degeneration. Investigative Ophthalmology & Visual Science