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Establishment of Three Different Glycation-Damage Cell Models and Analysis of Their Action Mechanism
Xinya Qian1, Chencan Cao1, Lei Liu1
1College of Light Industry Science and Engineering, Beijing Technology and Business University, Higher Education Garden, Liangxiang, Fangshan District, Beijing 102488, China.
Abstract:
Background/Objective: With rising per capita sugar consumption, skin glycation-related issues including dullness, homeostasis disruption and accelerated wrinkling have gained widespread attention. However, globally standardized and rigorous evaluation criteria for anti-glycation efficacy remain lacking. This study aimed to establish stage-specific glycation injury cell models and elucidate the stage-dependent molecular mechanisms of glycation-induced fibroblast damage, providing a standardized reference for anti-glycation efficacy assessment. Methods: Three glycation injury models were constructed in human foreskin fibroblasts (HFF-1): early-stage (glucose-induced), intermediate-stage (glyoxal-induced), and late-stage (advanced glycation end products (AGEs)-induced). Core biomarkers including Nε-(carboxymethyl)lysine (CML), collagen type I (Col I) and elastin (ELN) were used to optimize modeling conditions via Cell Counting Kit-8 (CCK-8) and enzyme-linked immunosorbent assay (ELISA). Untargeted metabolomics based on ultra-high-performance liquid chromatography (UHPLC)-Q Exactive Orbitrap was applied to identify differential metabolites and perturbed pathways, following Metabolomics Standards Initiative (MSI) Level 2 identification criteria. Results: Optimal conditions were determined as 50 mmol/L glucose for 48 h, 0.5 mmol/L glyoxal for 48 h, and 200 μg/mL AGEs for 24 h. A total of 319, 34 and 148 differential metabolites were identified in the three groups, respectively. Six key pathways were significantly perturbed. Early and intermediate models shared similar mechanisms (purine metabolism disturbance), while the late model showed distinct alterations in pyrimidine, nicotinate, arachidonic acid and steroid hormone metabolism. Conclusions: Three stable stage-specific glycation models were successfully established in HFF-1 cells. Significant differences in metabolic profiles and mechanisms exist across the three stages, providing a rational basis for model selection and theoretical support for anti-glycation efficacy evaluation.
Insights
This study established stage-specific skin glycation cell models to understand aging mechanisms. Findings reveal distinct metabolic pathways affected by early, intermediate, and late-stage glycation, aiding anti-glycation product development.
Area of Science:
- Dermatology and Cellular Biology
- Metabolomics and Biochemistry
Background:
- Rising sugar consumption contributes to skin aging issues like dullness and wrinkles.
- Lack of standardized methods hinders effective anti-glycation efficacy assessment.
- Glycation-induced fibroblast damage requires stage-specific investigation.
Purpose of the Study:
- Establish distinct cell models for early, intermediate, and late-stage glycation.
- Elucidate stage-dependent molecular mechanisms of glycation in fibroblasts.
- Provide a standardized reference for evaluating anti-glycation efficacy.
Main Methods:
- Constructed three glycation models in human foreskin fibroblasts (HFF-1) using glucose, glyoxal, and advanced glycation end products (AGEs).
- Optimized modeling conditions using Cell Counting Kit-8 (CCK-8) and ELISA for biomarkers like Nε-(carboxymethyl)lysine (CML).
- Employed untargeted metabolomics (UHPLC-Q Exactive Orbitrap) for differential metabolite and pathway identification.
Main Results:
- Determined optimal conditions for early (glucose), intermediate (glyoxal), and late-stage (AGEs) glycation models.
- Identified 319, 34, and 148 differential metabolites across the three stages, respectively.
- Revealed distinct metabolic perturbations: early/intermediate models showed purine metabolism disturbance, while the late model affected pyrimidine, nicotinate, arachidonic acid, and steroid hormone metabolism.
Conclusions:
- Successfully established three stable, stage-specific glycation models in HFF-1 cells.
- Demonstrated significant differences in metabolic profiles and mechanisms across glycation stages.
- Provided a basis for selecting appropriate models and theoretical support for anti-glycation efficacy evaluation.
