Non-invasive evaluation of advanced glycation end products in hair as early markers of diabetes and aging

Abstract

Continuous metabolic monitoring is essential for assessing lifestyle-related disease risks. Hair, an easily accessible tissue, allows for long-term metabolic evaluation, with glycated proteins linked to diabetic complications found in hair. We established a mass spectrometry system to detect advanced glycation end products (AGEs) in hair samples from humans and rats, assessing their variations with aging and disease. Hair samples were hydrolyzed and processed using a cation-exchange column for mass spectrometric analysis. Regardless of temperature variations, the levels of AGEs [N^ε-(carboxymethyl)lysine (CML), and methylglyoxal-derived hydroimidazolone-1 (MG-H1)] in human hair remained stable for one week. Age and CML levels, or AGEs z-scores combined with CML and CEL levels in human hair samples, were positively correlated. In streptozotocin-induced insulin-deficient diabetic model (DM) rats, hair CEL and MG-H1 levels were higher than in non-diabetic rats. Receiver operating characteristic curve analysis showed an area under the curve of 1 for hair CEL and MG-H1 levels. Serum and hair CML levels were positively correlated. Hair AGE levels vary more between DM and non-DM rats than serum AGE levels. They remain stable under heat treatment and correlate with age, indicating that hair analysis is an effective non-invasive method for assessing metabolic fluctuations.

Keywords: Advanced glycation end products; Diabetes; Hair; Mass spectrometry; Non-invasive method.

Fig. 1

Structures of AGEs and the position of the deuterium atom. The asterisk (*) indicates the position of the deuterium atom. (a) N^ε-(carboxymethyllysine) (CML), (b) N^ε-(carboxyethyllysine (CEL), (c) methylglyoxal-derived hydroimidazolone-1 (MG-H1).

Fig. 2

Advanced glycation end products (AGEs) in human hair stored at various temperatures. (a) Levels of N^ε-(carboxymethyl)lysine (CML), (b) levels of N^ε-(carboxyethyl)lysine (CEL), (c) levels of methylglyoxal-derived hydroimidazolone-1 (MG-H1), (d) lysine (Lys) levels, (e) arginine (Arg) levels. Pink: 40 °C, green: 4 °C, blue: −80 °C. n = 3. Statistical analysis: Wilcoxon rank-sum test; * P < 0.05.

Fig. 3

Correlation between AGEs levels in hair and age in healthy participants. Hair AGEs levels of 30 healthy females and their ages. (a) Hair N^ε-(carboxymethyl)lysine (CML), (b) Hair N^ε-(carboxyethyl)lysine (CEL), (c) Hair methylglyoxal-derived hydroimidazolone-1 (MG-H1), (d) Z-score with Hair CML + CEL levels and ages. Statistical analysis: Spearman’s analysis was performed with a significance level set at P < 0.05. Crosses inside the circles indicate non-significant correlations.Green and pink text indicate positive and negative correlations, respectively.

Fig. 4

Comparison of AGEs levels in serum and hair in diabetes (DM)-induced and non-diabetic (control) rats at 12 weeks. (a) Serum N^ε-(carboxymethyl)lysine (CML), (b) Hair CML, (c) Serum N^ε-(carboxyethyl)lysine (CEL), (d) Hair CEL, (e) Serum methylglyoxal-derived hydroimidazolone-1 (MG-H1), (f) Hair MG-H1 levels. Control (n = 7) and DM (n = 6). Data are shown as box plots, with bold horizontal bars indicating median values. Statistical analysis: Wilcoxon rank-sum test; * P < 0.05; ** P < 0.01; *** P < 0.001.

Fig. 5

Receiver operating characteristic (ROC) analysis of hair AGE levels in rats. ROC analysis was performed to assess the discriminatory ability of hair AGE levels and to determine the optimum cutoff point. The box plot shows individual sample values from the control (green, non-DM) and diabetes-induced (red, DM) rat groups. (a) Hair N^ε-(carboxyethyl)lysine (CEL) and (b) Hair methylglyoxal-derived hydroimidazolone-1 (MG-H1) levels.

Fig. 6

Correlation between hair and serum AGE levels in rats. Statistical analysis: Spearman’s analysis; significance level was set at P < 0.05. Green and pink text indicate positive and negative correlations, respectively.