A.G.E.s & My Bones research

Increased concentration of two different advanced glycation end-products detected by enzyme immunoassays with new monoclonal antibodies in sera of patients with rheumatoid arthritis.

Impact factor: 2.4

Level of evidence: II

Sample Size/Population: 45 RA patients, 40 patients with SLE, 48 patients with OA of the knee, and 51 healthy volunteers. All RA patients were treated with infliximab in combination with methotrexate. Additionally, 40/45 RA patients received a constant dosage of glucocorticoids. Sera was collected after treatment with infliximab.

Protocol: Two immunoassays were developed and based on new monoclonal antibodies to advanced glycation end- products. Antibody 103-E3 reacts with an unidentified antigen while antibody 8-C1 responds to Nε-(carboxyethyl)lysine (CEL). The monoclonal antibodies were used to measure levels of advanced glycation end-products in sera of patients with rheumatoid arthritis, systemic lupus erythematosus, osteoarthritis, and healthy controls. The correlation between AGE levels and the Disease Activity Score 28 (DAS28), age, disease duration, CRP, anti-CCP, rheumatoid factor and treatment with corticosteroids were also calculated.

Statistical Analysis: two cohorts compared with Mann-Whitney U and three compared with Kruskal-Wallis followed by Dunn’s post-hoc. Relationship was determined via Spearman’s correlation (patients with samples whose antigen level was below ELISA detection were included). P value: 0.05. 

Study Type: Case Control

Results: Levels of both glycations products were significantly higher in rheumatoid arthritis patients compared to those with systemic lupus erythematosus, osteoarthritis, or the healthy controls. The levels of Nε-(carboxyethyl)lysine and103-E3 antigen did not show a correlating DAS28-scored rheumatoid arthritis activity. The levels of Nε-(carboxyethyl)lysine and103-E3 antigen also did not correlate with age, gender, corticosteroid treatment, or levels of CRP, anti-CCP antibodies, and rheumatoid factor in sera; therefore, The increase in the two AGEs in the sera of patients with rheumatoid arthritis could not be explained by rheumatoid arthritis activity or by inflammation.

Conclusion: Presumes a link between AGE formation and the initiation of autoimmunity.

Vytásek R, Sedová L, Vilím V. Increased concentration of two different advanced glycation end-products detected by enzyme immunoassays with new monoclonal antibodies in sera of patients with rheumatoid arthritis. BMC Musculoskelet Disord. 2010 May 3;11:83. doi: 10.1186/1471-2474-11-83. PMID: 20433772; PMCID: PMC2881016.

Intake of high-fructose corn syrup sweetened soft drinks, fruit drinks and apple juice is associated with prevalent arthritis in US adults, aged 20-30 years.

Impact factor: 4.357 

Level of evidence: VI

Sample Size:  1209 adults aged 20-30y 

Important methods: Exposure variables considered were high EFF drinks, including HFCS sweetened soft drink, and other HFCS sweetened soft drinks, fruit drinks and apple juice. Diet soda and diet fruit juice were analyzed for comparison. The results were based on self-reported arthritis. 

Study Type: Cross-sectional 

Statistical Analysis: Rao Scott X^2 to test for significance of differences in arthritis prevalence by intake frequency; Weight variables used to account for non-response and oversampling of various age and ethnic groups; P-value: 0.05. 

Results: Consumption of beverages with high levels of EFF was significantly correlated with arthritis in US young adults. Young adults reporting intake of EFF greater than 5 times per week had three times higher odds of arthritis as those who only had 1-3 per month.

Limitations: The exposures and outcome are simultaneously assessed, and follow-up longitudinal studies are needed. Arthritis was also self-reported, so it could contain bias. 

Discussion/Conclusion: Regular consumption of High EFF beverages is associated with arthritis in young adults. Unabsorbed excess free fructose in the GI tract may interact with dietary proteins to form AGEs that could play a rold in gut and joint inflammation in young adults. 

DeChristopher LR, Uribarri J, Tucker KL. Intake of high-fructose corn syrup sweetened soft drinks, fruit drinks and apple juice is associated with prevalent arthritis in US adults, aged 20-30 years. Nutr Diabetes. 2016 Mar 7;6(3):e199. doi: 10.1038/nutd.2016.7. PMID: 26950480; PMCID: PMC4817078.  

Physicochemical studies on glycation-induced structural changes in human IgG.

Impact factor: 3.09

Level of evidence: V

Study Type: In vitro

Protocol: Human IgG mixed with PBS was incubated with 5mM glucose in sterile conditions at 37 °C for 5-25 days. After 25 days, the solutions were dialyzed against PBS to remove excess glucose. Absorbance spectroscopy, electrophoresis, fluorescence studies, circular dichoism (CD) measurements, FTIR spectroscopy, thermal denaturation, determination of protein-bound carbonyl groups as well as thiobarbituric acid assay were completed. 

Statistical Analysis: Statistical significance was a p value of < 0.05. The data are presented as mean ± Standard deviation (SD) and statistical significance was determined by Student’s t test.

Results: Formation of AGEs was confirmed by fluorescence spectroscopy. An increase of 84.2% in FI was seen in AGE-IgG when compared to the native form of IgG. CD spectrum of native IgG showed a characteristic spectrum of mostly β-sheeted protein, but CD spectrum of AGE-IgG indicated significant loss of its β structure upon glycation. FTIR spectra of native and AGE-IgG were analyzed for the position of amide I and amide II bands. AGE-IgG showed a shift to the left for each band indicating an alteration in IgG structure on glycation. Thermal denaturation showed that AGE-IgG had a higher melting temperature (74°C) than native IgG (70.5°C) indicating thermostability of the glycated IgG. Protein carbonyl groups are a known biomarker for oxidative stress, and AGE-IgG had an almost threefold increase in carbonyl level compared to native IgG.

Conclusion: IgG are serum proteins rich in lysine residues making them an ideal target for glycation. This study showed extensive damage to IgG when glycated when analyzed using various physiochemical techniques. AGE-IgG samples showed hyperchromicity that could be explained by the unfolding and fragmentation upon glycation. AGE-IgG also showed increased mobility in PAGE due to a change in charged residues in the protein structure. Native IgG showed a largly negative dichroism due to its β-sheet structure, and the AGE-IgG showed a loss in β-sheet structure with an increase in dichroism. The AGE-IgG is more thermostable than the native IgG and could be due to cross-links. This study shows that in vitro treatment of human IgG with physiological concentrations of glucose causes alterations in the immunoglobulin resulting in the formation of AGEs. It has been reported that patients with rheumatoid arthritis that have severe systemic symptoms have AGE-IgG and elevated IgM and IgA antibodies against the AGE-IgG. AGE-IgGs may play a role in the induction of circulating autoantibodies in RA. Being more aware of AGE content in foods could decrease the amount of circulating AGE-IgG.

Ahmad S, Moinuddin, Khan RH, Ali A. Physicochemical studies on glycation-induced structural changes in human IgG. IUBMB Life. 2012 Feb;64(2):151-6. doi: 10.1002/iub.582. PMID: 22241644. 

Advanced Glycation End-products and Bone Fractures

Impact Factor: 3.998 

Evidence level: V 

Study type: Review article  

Discussion:  In this review article, the author surveys the current evidence for an association between AGE levels and bone fracture risk. The author notes there is evidence that measuring AGE levels in bone may serve as a useful biomarker for fracture risk, especially in the aging population, those with diabetes, and those undergoing menopause. Of particular importance as a biomarker, the AGE crosslink compound pentosidine has been identified in bone samples and is believed to contribute to bone fragility. Current methods used to measure AGE levels in bone most commonly include HPLC or fluorometric assay, but the author notes easier methods are being investigated, namely, the use of serum and urine samples. 

Vashishth D. Advanced Glycation End-products and Bone Fractures. IBMS Bonekey. 2009;6(8):268-278. doi:10.1138/20090390

The effect of aminoguanidine (AG) and pyridoxamine (PM) on ageing human cortical bone.

Impact factor: 3.532 

Level of evidence: Level VI

Type of study and any information related to it: This was a cross-sectional study seeking to determine the inhibitory effect that aminoguanidine (AG) and pyridoxamine (PM) have on AGE formation in cortical bone. An in vitro model was used to study the efficacy of AG and PM. 

Sample size: 20 mid-diaphyseal tibial cortical bone segments were taken from female cadavers with an age range from 57 to 97 years old. 

Important methods: The tibial cortical bone segments were randomly assigned to four groups: control, glucose only, glucose and AG, and glucose and PM. Cortical bone segments were cut into 4 mm*4mm*4mm segments, treated with acid hydrolysis, and fluorescence in a multimode plate reader was used to assess AGE levels. Bone segments were treated with either glucose, glucose and AG, or glucose and PM for seven days. The biomechanical property of the bone was assessed by indentation distance increase (IDI). The IDI is measured by placing a reference probe on the surface of the cortical bone after treatment and the probe initiates a crack. 

Statistical tests ran: Differences between experimental groups were analyzed using an ANOVA. A p-value of less than 0.05 was considered statistically significant. 

Results: There was a statistically significant reduction in AGE content after treatment with AG and PM when compared with the glucose only group. There was no statistically significant difference in AGE content between the control group and the AG and PM treatment groups. After three days of treatment with glucose only the IDI increased significantly compared to the control group. Treatment with both AG and PM significantly reduced IDI compared to the glucose alone group. There was no statistically significant difference in IDI between the control group and the AG and PM treatment groups. The study also demonstrated that elevated AGE content is associated with decreased IDI. 

Discussion/Conclusion: AGE accumulation is one of many changes that occurs in adult bone matrix over time. Cross linking from AGEs with bone matrix decreases the quality of the matrix and may negatively affect bone remodeling. This study demonstrated that AG and PM, which cleave AGE cross links, can improve the biomechanical property of bone. AG and PM could potentially be used as a therapy to help older adults improve bone quality and function and prevent bone matrix deterioration over time. Limitations: Firstly, this study was conducted in vitro, therefore, in vivo tests in animal and human models will be necessary to demonstrate the efficacy and safety of AG and PM. Secondly, the small sample size and limited treatment period (7-days) merit future research to confirm the results of this study and help establish what the effects of long-term AG and PM treatment are.

Abar, O., Dharmar, S., & Tang, S. Y. (2018). The effect of aminoguanidine (AG) and pyridoxamine (PM) on ageing human cortical bone. Bone & joint research, 7(1), 105-110.

Effects of long-term physical activity and diet on skin glycation and Achilles tendon structure.

Impact factor: 4.546 

Level of evidence: Level VI

Type of study and any information related to it: This is a cross-sectional study investigating the effects that long-term physical activity and diet have on skin glycation and Achilles tendon thickness. 

Sample size: 194 regularly exercising males who competed in the 2017 European Masters Athletics Championships Stadia in Aarhus, Denmark and 34 untrained controls were selected for the study. 

Important methods: Selection criteria for the athletes were that they had been training at least twice a week for the past 10 years and were at least 18 years of age. Twenty-four different nationalities were represented in the study. Participants filled out a questionnaire including items on physical activity and diet. AGEs levels were estimated by measuring skin autofluorescence (SAF) using an AGE reader (DiagnOptics B.V., Groningen, The Netherlands). Ultrasonography was used to measure the anteroposterior thickness of the Achilles tendon at the proximal and distal ends of the tendon. 

Statistical tests ran: A Welch’s t-test was used to determine differences in basic characteristics and diet between athletes and control groups. Relationships between SAF and participant characteristics were analyzed by multiple linear regressions using backward elimination. A p-value of less than 0.05 was considered statistically significant. 

Results: Athletes consumed more vegetables, fruit and fish compared to the control group. There was no significant difference between SAF in the athletic and control groups; however, SAF was shown to independently decrease with training years. The athletic group was shown to have increased Achilles tendon thickness when compared to the control group; however, there was no significant association between SAF levels and Achilles tendon thickness. Participants who indicated having a more “Western” diet were associated with higher SAF readings. The association of lower SAF levels with “Western” diet and training years were present in both athletic and control groups. 

Discussion/Conclusion: The results from this study were unexpected in that they did not show a significant difference between SAF measurements in the athletic and control groups. Additionally, there was no relationship between Achilles tendon thickness and SAF levels. The study did show that training years and diet were independently associated with decreased SAF levels. This suggests that long term athletes who exercise consistently do not have lower glycation rates when compared to controls. This study does demonstrate that consistent physical activity, regardless of whether you are a lifelong athlete, and a “healthier” diet consisting of fruit, vegetables, and fish can lead to lower levels of AGEs in your skin. 

Limitations: This was a single cross-sectional study with a relatively small sample size; therefore, a causal relationship cannot be established. The length of the questionnaire was limited, and the researchers did not measure any of the variables related to physical activity directly. Additionally, the nature of the recruiting process made it impossible to blind investigators to the participants' groupings.

Hjerrild, J. N., Wobbe, A., Stausholm, M. B., Larsen, A. E., Josefsen, C. O., Malmgaard-Clausen, N. M., ... & Couppé, C. (2019). Effects of long-term physical activity and diet on skin glycation and achilles tendon structure. Nutrients, 11(6), 1409. 

Elevated serum advanced glycation end products and poor grip strength in older community-dwelling women.

Impact factor:  2.094 

Level of Evidence: Level IV 

Study type: Cross-sectional study 

Sample size: 599 women, age 65 and older 

Intro:  Recent studies have shown an association between advanced glycation end products (AGE) and inflammatory diseases such as diabetes. This study aims to look at the association between AGE with poor muscle strength.  It has been observed that a pentosidine, a specific AGE product, is increased by more than 200% in skeletal muscle of older adults compared to younger adults.   

Methods:  Standardized questionnaires were administered in the participant's home by trained interviewers. Mini-Mental State Examination (MMSE) was recorded. Geriatric Depression Scale, consisting of 30 items with a yes or no response, was assessed at each visit, and a score of 14 or higher indicating moderate to high level of depressive symptomatology. Serum carboxymethyl-lysine (CML), a dominant AGE, circulating soluble form of receptor for advanced glycation end products (sRAGE), and endogenous secretory receptor for advanced glycation end product (esRAGE) and grip strength were measured in 559 moderately to severely disabled women, age 65 and older   

Statistical Analysis:  Linear regression analysis was used to examine the relationship between serum CML, sRAGE, and esRAGE and other factors with grip strength as a continuous outcome variable. Variables that were at a level of significance of p < .10 in univariate analyses were included in the multivariate models. An MMSE score of <24 was defined as cognitive impairment.   

Results: Serum sRAGE and esRAGE were not significantly associated with grip strength. It is also possible that larger sample size and power are needed to examine the association between circulating RAGE and skeletal muscle strength. It is also possible that larger sample size and power are needed to examine the association between circulating RAGE and skeletal muscle strength.     

Conclusion: Women with high serum AGEs have greater muscle weakness. Further studies are needed to determine whether AGEs, a potentially modifiable risk factor, are associated with physical performance and disability in older adults.  

Dalal M, Ferrucci L, Sun K, Beck J, Fried LP, Semba RD. Elevated serum advanced glycation end products and poor grip strength in older community-dwelling women. J Gerontol A Biol Sci Med Sci. 2009 Jan;64(1):132-7. doi: 10.1093/gerona/gln018. Epub 2009 Jan 31. PMID: 19182228; PMCID: PMC2645474. 

Advanced glycation end products are associated with physical activity and physical functioning in the older population.

Impact factor: 5.236 

Level of evidence: Level II 

Type of study and any information related to it: The relationship between advanced glycation end products (AGEs) and age-related decline in physical activity and physical functioning in older populations were investigated using cross-sectional data from the LifeLines Cohort study. The LifeLines study is a large population-based investigation on how various factors (e.g. genomic, phenotypic, environmental) impact aging and the development of chronic diseases. Data for the LifeLines study were collected from 2006-2013 and the participants were from the northern part of the Netherlands. 

Sample size: Drenth et al. analyzed data on 5,624 participants aged 65 years and older who had their AGE levels assessed. 

Important methods: AGEs were assessed by skin autofluorescence (SAF) which was measured with an AGE Reader device. Each participant’s number of physically active days per week was determined using the Short Questionnaire to Assess Health-Enhancing Physical Activity (SQAUSH). Physical functioning (total score) was assessed by the RAND-36 questionnaire. Scores from the SQUASH were also used to determine whether a participant complies with the Dutch Physical Activity (DPA) guidelines which require a moderately intensive activity for 30 minutes at least 5 days a week. 

Statistical tests ran: ANOVA and chi-square tests were used to test differences in continuous and categorical variables between low (SAF-AGE ≤ 2.19 AU), middle (2.19> <2.56AU) and high (SAF-AGE ≥2.56 AU) groups. Multiple linear regression analysis was used to estimate the association between AGEs and physical functioning and physical activity. A p-value of less than 0.05 was considered statistically significant. 

Results: The study demonstrated a significant association between AGE levels and the number of physically active days, physical functioning, and compliance with DPA guidelines. 

Discussion/Conclusion: This study indicates that there may be an association between elevated levels of AGEs and lower levels of physical activity and physical functioning in older adults. The study indicated that with one unit of AGE increase, the number of physically active days in a week were 20% less. The study also suggested that elevated levels of AGEs may be associated with decreased physical functioning and functional motor decline as individuals with higher AGEs were less likely to meet the DPA guidelines. 

Limitations: While the SQUASH and RAND-36 are reliable and valid measures of physical activity and physical functioning the study did not directly measure these variables. This may have led to a misrepresentation of the impact that AGEs had on physical activity and physical functioning. 

Drenth H, Zuidema SU, Krijnen WP, et al. Advanced Glycation End Products Are Associated With Physical Activity and Physical Functioning in the Older Population. The Journals of gerontology. Series A, Biological Sciences and Medical Sciences. 2018 Oct;73(11):1545-1551. DOI: 10.1093/gerona/gly108. PMID: 29718128. 

The Contribution of Advanced Glycation End product (AGE) accumulation to the decline in motor function.

Impact factor: 2.447 

Level of evidence: V 

Type of study and any information related to it: This review article highlighted eight studies that investigated the relationship between advanced glycation end products (AGEs) and how they relate to physical functioning, physical performance, and musculoskeletal outcomes. 

Sample size:  Eight studies were included in the review. Six of the studies used a cross-sectional design, one used a longitudinal design, and one used a mixed longitudinal and cross-sectional design. 1154 studies were identified and only eight studies made it through the selection process.

Important methods: Articles were found on the Embase, Pubmed, Cinahl, and Web of science databases. Only articles that measured the relationship between AGEs and motor function were included in the review. Studies were excluded if they were disease or diet/weight related, were narrative reviews, conference abstracts or editorial comments, or if they did not describe relevant outcomes. Each paper was read by two reviewers and the articles were rated as “good” (valid and applicable with sufficient statistical analysis), “moderate” (validity was rated as doubtful and sufficient statistical analysis), or “low”. 

Statistical tests ran: Risk ratios (RR), Odds ratios (OR), Hazard ratio’s (HR), correlation coefficients, and group differences were presented as they were in the studies. The magnitude of the effect AGEs had on outcomes were either presented as association coefficients or effect sizes (ES). Cohen’s benchmarks were used to define small (ES= 0.20), medium (ES= 0.50), and large (ES= 0.80) effect sizes. Meta-analysis was not performed.  

Results: The motor function outcomes that were covered in the eight studies were walking ability, activities of daily living (ADL), upper extremity function, muscular strength and mass, and physical frailty as defined by the Fried criteria. The Fried criteria describe physical frailty as having three or more of the following: exhaustion, decreased muscle mass, low physical activity, slow walking speed, and unintentional weight loss. Studies assessed AGEs using circulating carboxymethyl lysine (CML), circulating Pentosidine, or by measuring skin auto-fluorescence (AF).   

Two different studies investigated the relationship between plasma CML and slow walking speeds. In one report elevated CML levels in community dwelling elderly were significantly associated with decreased walking speed which was defined as being less than 0.79 m/s (OR= 1.87, 95 % CI 1.15–3.04, P= 0.01). Another study observed that subjects in the highest quartile for serum CML in moderately to severely disabled female subjects was significantly associated with decreased walking speed which was defined as being less than 0.4 m/s (HR’s between 1.46 and 1.63, all p < 0.05). 

One study examined the relationship between circulating CML levels and the likelihood that participants in the Cardiovascular Health study would develop an ADL complication over a 14-year period. The ADL activities were walking around the home, getting out of bed or a chair, dressing, bathing, eating, and toileting. High levels of serum CML at baseline were associated with a 10% higher incidence of developing an initial ADL complication (HR= 1.10, p < 0.01). Additionally, elevated CML levels were associated with a 5% higher incidence of having a disability (HR= 1.05, p= 0.03). 

One study compared upper extremity function, assessed using the disability of the arm, shoulder and hand (DASH) questionnaire, to AGEs measured via AF. The participants responses to the DASH questionnaire (scores ranged from 0 to 100) were found to be significantly correlated to their AF levels, suggesting that higher levels of AGEs in the skin correlated with increased dysfunction of the upper extremity (r= 0.51, P= 0.009). 

Four studies investigated the effect AGEs had on muscle strength and power. One study explored the association between handgrip strength and circulating CML in moderate to severely disabled elderly women. Women with elevated CML had a decreased mean grip strength compared to the groups with lower levels of CML (Beta= −1.31, SE= 0.61, P= 0.03). Another study examined the relationship between skeletal muscle CML/RAGE levels (internal abdominal oblique) in a group of healthy subjects divided into self-reported weight maintainers (WM), weight gainers (WG), and elderly (E). The study found that increased skeletal muscle CML/RAGE levels were associated with decreased handgrip strength (r= −0.54, p < 0.05). One study measured AF levels in healthy adult males and found that leg extension power was 10% lower in subjects in subjects in the highest tertile for AF levels (ES= 0.44, P= 0.04). Additionally, it was found that AF was not related to shoulder flexor muscle strength (r= 0.07, P=0.7). 

The relationship of skeletal muscle mass and Pentosidine was investigated using a group of postmenopausal women. Serum Pentosidine levels were negatively correlated with lower extremity muscle mass (beta= -0.18, P= 0.071) and relative skeletal muscle mass index (beta= -0.27, P= 0.008). No relationship was found between Pentosidine and the muscle mass of the upper extremity (r= -0.11, P= 0.219). 

The relationship between physical frailty and CML was examined using a group of healthy and older adults, with physical frailty being assessed by the Fried criteria discussed above. Mean CML levels were significantly higher in men with low physical activity, increased exhaustion and decreased grip strength and this subsequently led to a significant association with physical frailty (OR= 1.24, 95 % CI:1.07–1.45, p= 0.04). No significant relationship between physical activity, exhaustion, and grip strength were found for females. 

Discussion/Conclusion: From this review it is clear that elevated levels of AGEs are associated with a decline in walking ability, lower extremity ADL, decreased muscle strength and mass and increased physical frailty. AGEs have the potential to be used as a biomarker to assess the risk for a decline in motor function in individuals. While most participants in the 8 studies reviewed in this paper were 64 years of age or older (n= 5433), two of the studies included adults between 37 and 56 years of age (n= 387), which could mean that increased AGE levels in midlife could be used to predict future motor decline. Most research to date has been observational or cross-sectional so a causal relationship between AGEs and motor decline cannot be established. Therefore, future research aimed at exploring the pathological and physiological changes that AGEs have on tissue and how that impacts motor function is needed before AGEs can be used as a marker for decline in motor function. Specifically, studies should consider how AGE induced changes in joints, muscle, tendons, ligaments and the nervous system directly relate to a decline in motor function. Future research could also assess if reducing the amount of AGEs in subjects leads to improved motor function and what modalities (e.g. functional training, strength training) are best suited for preventing AGE related motor decline. 

Drenth, Hans, et al. "The Contribution of Advanced Glycation End product (AGE) accumulation to the decline in motor function." European review of aging and physical activity 13.1 (2016): 1-13. 

The potential role of advanced glycation end products (AGEs) and soluble receptors for AGEs (sRAGE) in the pathogenesis of adult-onset Still's disease.

Impact factor: 2.398 

Level of evidence: IV 

Type of study: cross sectional 

Sample size: 52 adult-onset Still’s disease (AOSD) patients, 36 systemic lupus erythematosus (SLE) patients, and 16 healthy controls (HC). 

Methods: ELISA to determine plasma levels of AGEs and sRAGE in all patients. Their associations with activity parameters and disease courses were evaluated. 

Statistical analysis: Kruskal-Wallis test was used for between-group comparison of AGEs or sRAGE levels. P-value determined using the Mann–Whitney U test. The correlation coefficient was obtained using nonparametric Spearman’s rank correlation test. A logistic regression analysis was used to evaluate the effects of clinical characteristics/disease activity parameters on levels of AGEs or sRAGE in the AOSD group. Wilcoxon signed rank test was used to compare AGE and sRAGE levels during follow-up in AOSD patients after effective therapy. p < 0.05 was considered significant. 

Results: Significantly higher median levels of AGEs were observed in active AOSD patients and active SLE patients than those in HC. AGEs levels were positively correlated with activity scores, ferritin levels, and CRP levels in AOSD patients. Significantly lower median levels of sRAGE were observed in active AOSD patients and active SLE patients compared with HC. Plasma sRAGE levels were negatively correlated with AOSD activity scores. Significantly higher AGEs levels were observed in AOSD patients with polycyclic or chronic articular patterns compared to monocyclic. With treatment, AGEs levels decreased and sRAGE levels increased as disease activity declined. 

Conclusion: Increased AGEs and decreased sRAGE, which were significantly correlated with disease activity parameters, may be involved in AOSD pathogenesis. This was the first study to have evidence showing that plasma levels of AGEs and sRAGE may be linked with disease outcome in people with AOSD. The elevation of AGEs levels and depression of sRAGE levels in active AOSD patients, suggests their pathogenic role in AOSD.

Chen DY, Chen YM, Lin CC, Hsieh CW, Wu YC, Hung WT, Chen HH, Lan JL. The potential role of advanced glycation end products (AGEs) and soluble receptors for AGEs (sRAGE) in the pathogenesis of adult-onset still's disease. BMC Musculoskelet Disord. 2015 May 9;16:111. doi: 10.1186/s12891-015-0569-3. PMID: 25956266; PMCID: PMC4436789.

Plantar fasciitis in patients with type 1 and type 2 diabetes: A contemporary cohort study.

Impact Factor: 2.781

Level of Evidence: VI

Sample Size: 722,439 (Type 1 diabetes- 7148; Type 2 diabetes- 61,632; no diabetes- 653,659). 

Protocol: Five year prevalence study using the electronic medical record of a large tertiary healthcare system. The database was screened for all patients above 18 years of age with plantar fasciitis between 2011 and 2016. Patients were divided by diabetes status: Type 1, Type 2, no diabetes. The prevalence of plantar fasciitis in patients with and without diabetes was compared, while accounting for the influence of other variables like age and BMI.

Statistical Analysis: Fisher’s exact test, Cochran-Mantel-Haenszel Statistics. 

Study Type: Case-control, retrospective. 

Results: Prevalence of plantar fasciitis overall, in the Type 2 group, Type 1 group, and no diabetes group were 0.85%, 1.31%, 0.92%, and 0.80%, respectively.  The Type 2 diabetes group showed a significantly higher prevalence of plantar fasciitis compared to Type 1 and no diabetes, respectively (p=0.0054 and p<0.0001). Females showed a higher prevalence of plantar fasciitis than males across all groups (p<0.0001).

Conclusion: AGEs have been associated with the development of plantar fasciitis. Hyperglycemia in diabetic patients could be a cause of AGEs in this population. The rates of plantar fasciitis were seen to be higher in diabetic patients pointing to the role of diabetes in AGE formation leading to plantar fasciitis. This was a retrospective study using database data, so better controlled studies on this topic would be needed to make a direct correlation.

Priesand SJ, Schmidt BM, Ang L, et al. Plantar fasciitis in patients with type 1 and type 2 diabetes: A contemporary cohort study. J Diabetes Complications. 2019;33(10):107399. doi:10.1016/j.jdiacomp.2019.06.004

Bone microarchitecture, biomechanical properties, and advanced glycation end-products in the proximal femur of adults with type 2 diabetes. 

Impact Factor: 4.147

Level of Evidence: IV

Sample Size: Serum and proximal femur specimens were taken from 20 type II diabetic patients and 33 non diabetic patients undergoing total hip replacement surgery. 

Protocol: A section from the femoral neck was imaged by microcomputed tomography (microCT), tested by cyclic reference point indentation, and quantified for AGE content. A trabecular core taken from the femoral head was imaged by microCT and subjected to uniaxial unconfined compression tests.

Statistical Analysis: N/A

Study Type: Clinical study 

Results: T2D subjects had greater HbA1c (+23%, p ≤ 0.0001), but no difference in cortical tissue mineral density, cortical porosity, or trabecular microarchitecture compared to non-diabetics. Cyclic reference point indentation revealed that creep indentation distance (+18%, p ≤ 0.05) and indentation distance increase (+20%, p ≤ 0.05) were greater in cortical bone from T2D than in non-diabetics, but no other indentation variables differed. Trabecular bone mechanical properties were similar in both groups, except for yield stress, which tended to be lower in T2D than in non-diabetics. Neither serum pentosidine nor serum total AGEs were different between groups. Cortical, but not trabecular, bone AGEs tended to be higher in T2D subjects (21%, p = 0.09). 

Conclusions: Serum AGEs and pentosidine were positively correlated with cortical and trabecular bone AGEs. This study presents new data on biomechanical properties and AGEs in adults with T2D, which are needed to better understand mechanisms contributing to diabetic skeletal fragility.

Lamya Karim, Julia Moulton, Miranda Van Vliet, Kelsey Velie, Ann Robbins, Fatemeh Malekipour, Ayesha Abdeen, Douglas Ayres, Mary L Bouxsein. Bone microarchitecture, biomechanical properties, and advanced glycation end-products in the proximal femur of adults with type 2 diabetes. DOI: 10.1016/j.bone.2018.05.030

Hyperglycemia-related advanced glycation end-products is associated with the altered phosphatidylcholine metabolism in osteoarthritis patients with diabetes

Impact factor: 2.740 

Level of evidence: IV 

Type of study: Cross sectional 

Sample size: 84 patients with knee osteoarthritis (OA) (46 with DM and 38 without DM) 

Methods: Blood samples were collected after overnight fast of at least 8 h, and synovial fluid (SF) samples were collected during joint replacement surgeries. Metabolic profiling in both plasma and SF were performed by the Waters XEVO TQ MS system. Correlation between AGEs and phosphatidylcholine acyl-alkyl C34:3 (PC ae C34:3) and C36:3 (PC ae C36:3) was measured. Liquid chromatography and mass spectrometry were used to measure methylglyoxal (MG) and hydroimidazolone (MG-H1) in plasma and SF. 

Statistical analysis: Comparisons of the 4 AGEs between OA with and without DM were tested by t-test. The correlation between each of the 4 AGEs and PC ae C34:3 and PC ae C36:3 were measured by Spearman correlation coefficient. Statistical significance was defined as p < 0.05. Confounders including age, sex, BMI, and hexose level (>90% is glucose) were adjusted by linear regression modeling. 

Results: The difference in hexose levels between DM and non-DM patients were significant. There was no significant difference in plasma levels of MG, MG-H1, CML, and CEL between OA patients with and without DM. However, MG and MG-H1, but not CML and CEL in SF were significantly higher in OA patients with DM than in those without. The correlation became even stronger after adjusting for confounders. MG-H1 in SF was negatively correlated with PC ae C34:3 and PC ae C36:3 after the adjustment of age, BMI, sex and hexose level.  

Conclusion: Production of non-protein bound AGEs was increased in the OA-affected joint of DM patients. This is due to changes in phosphatidylcholine metabolism, and might be responsible for the association between OA and DM. 

Zhang W, Randell EW, Sun G, Likhodii S, Liu M, Furey A, Zhai G. Hyperglycemia-related advanced glycation end-products are associated with the altered phosphatidylcholine metabolism in osteoarthritis patients with diabetes. PLoS One. 2017 Sep 12;12(9):e0184105. doi: 10.1371/journal.pone.0184105. PMID: 28898260; PMCID: PMC5595284. 

Increased concentration of two different advanced glycation end-products detected by enzyme immunoassays with new monoclonal antibodies in sera of patients with rheumatoid arthritis.

Impact factor: 1.88  

Level of evidence: IV 

Type of study: case control  

Important methods: 2 immunoassays were used to detect 2 antibodies, 103-E3 and 8-C1. 103-E3 antibodies react with unidentified antigen formed in the reaction of protein and ribose. 8-C1 responds to Nepsilon. These monoclonal antibodies measure levels of advanced glycation end-products in patients with rheumatoid arthritis, systemic lupus erythematosus, osteoarthritis, and healthy controls.  

Statistical tests ran: Does not state but based on graphs, data is not normally distributed.  

Results: Levels of both glycation products were significantly higher in sera of patients with rheumatoid arthritis when compared with sera of patients with systemic lupus erythematosus, osteoarthritis, or the healthy controls. Neither the level of Nepsilon-(carboxyethyl)lysine nor the level of the 103-E3 antigen in rheumatoid arthritis sera correlated with the DAS28-scored rheumatoid arthritis activity. The levels of both antigens in rheumatoid arthritis sera did not correlate with age, gender, corticosteroid treatment, or levels of CRP, anti-CCP antibodies, and rheumatoid factor in sera. 

Discussion/Conclusion: Levels of pentosidine (AGE) are higher in patients with rheumatoid arthritis. This study showed that other AGEs are also increased in people with rheumatoid arthritis, although the levels did not correlate with age, gender, treatment, CRP, anti-CCP, antibodies and RF. The increase could not be explained by either the rheumatoid arthritis activity or by inflammation.  

Vytásek R, Sedová L, Vilím V. Increased concentration of two different advanced glycation end-products detected by enzyme immunoassays with new monoclonal antibodies in sera of patients with rheumatoid arthritis. BMC Musculoskelet Disord. 2010;11:83. Published 2010 May 3. doi:10.1186/1471-2474-11-83

A role for advanced glycation end products in diminished bone healing in type 1 diabetes.

Type of Study: Rat Study/ RTC  

Sample Size: 20 BALB/cBYJ mice  

Statistical test: ANOVA Methods: There were two groups of  mice in this study with an AGE protein ligand called carboxymethylysine and the effect that it has after a craniotomy and the effects that it has on the healing. The healing was then monitored in both groups of mice through histology samples, the expressiveness of osteoblastic cells, and also the amount of healing in individual lesions. The amount of bone mineral content, osteopenia, the rate of fracture was also measured. 

Discussion and Conclusion: The diabetic rats who were found to have higher levels of AGE levels in their system were reported to have only 40% of the healing that was found in nondiabetic rats. In nondiabetic rats, that were treated with the AGE serum, were said to have a decreased amount of bone healing from about 63-42 %. These results may indicate that the levels of AGEs may be an area of concern when it comes to healing of diabetes.  

Santana RB, Xu L, Chase HB, Amar S, Graves DT, Trackman PC. A role for advanced glycation end products in diminished bone healing in type 1 diabetes. Diabetes. 2003;52(6):1502-1510. doi:10.2337/diabetes.52.6.1502

Identification of the advanced glycation end products N(epsilon)-carboxymethyllysine in the synovial tissue of patients with rheumatoid arthritis.

Type of Study: This was an in vivo cross sectional study  

Sample Size: 10 patients with Rheumatoid Arthritis and 8 patients that were controls (4 with osteoarthritis) 

Methods: Synovial tissue samples were taken from the patients who were diagnosed with Rheumatoid Arthritis. These samples were frozen. These samples were then given rabbit-anti-CML-IgG and goat-anti rabbit-IgG and then was visualized with streptavidin-alkaline phosphatase. 

Discussion and Conclusion: The CML marker, which is a marker for N(epsilon)-carboxymethyllysine, is a marker for inflammation and stress in the body. After the synovial tissue samples were taken from these patients, they were analyzed for the presence of CML. In all of the patients that had Rheumatoid Arthritis, all of the patients had elevated levels of the CML marker. In all of the patients who also suffered from Osteoarthritis, they also had elevated levels of the CML marker.  

Drinda S, Franke S, Canet CC, Petrow P, Bräuer R, Hüttich C, Stein G, Hein G. Identification of the advanced glycation end products N(epsilon)-carboxymethyllysine in the synovial tissue of patients with rheumatoid arthritis. Ann Rheum Dis. 2002 Jun;61(6):488-92. doi: 10.1136/ard.61.6.488. PMID: 12006318; PMCID: PMC1754129.

Advanced Glycation Endproducts and Bone Material Strength in Type 2 Diabetes

Impact Factor: 5.605 

Level: 2 

Summary: A cross sectional study was done to determine the role of advanced end glycation products in bone deterioration which is a known complication of type 2 diabetes. The study was done on 16 postmenopausal women with T2DM and 19 healthy controls also postmenopausal. They measured the level of AGE’s by skin autofluorescence and used the bone material strength index. In t2dm they did see an association of 0.006 between reduced bone material strength, whereas there was no association with the controls. The SAF accounted for approximately 26% of the age-adjusted variance in bone material strength in t2dm ( P value of 0.03) this suggests that AGE’s can impair the bone material strength in post menopausal T2Dm women.  

Furst JR, Bandeira LC, Fan WW, Agarwal S, Nishiyama KK, McMahon DJ, Dworakowski E, Jiang H, Silverberg SJ, Rubin MR. Advanced Glycation Endproducts and Bone Material Strength in Type 2 Diabetes. J Clin Endocrinol Metab. 2016 Jun;101(6):2502-10. doi: 10.1210/jc.2016-1437. Epub 2016 Apr 26. PMID: 27115060; PMCID: PMC4891790. 

Relationship of shoulder activity and skin intrinsic fluorescence with low level shoulder pain and disability in people with type 2 diabetes

Impact Factor: 2.74 

Level: IV 

Summary: A study was done with 81 participants 52 T2DM and 29 controls were examined to determine the role of advanced end glycation in shoulder mobility and pain. The AGEs were measured by skin autofluorescence. The study concluded that T2DM patients had 23% less mobility (p value of 0.01) as well as a greater skin autofluorescence levels (p value of 0.01). However, they concluded that more research was needed to determine the effects of therapy and intervention on the joint mobility overall.   

Sorensen CJ, Hastings MK, Lang CE, McGill JB, Clark BR, Bohnert KL, Mueller MJ. Relationship of shoulder activity and skin intrinsic fluorescence with low level shoulder pain and disability in people with type 2 diabetes. J Diabetes Complications. 2017 Jun;31(6):983-987. doi: 10.1016/j.jdiacomp.2017.03.005. Epub 2017 Mar 18. PMID: 28392042; PMCID: PMC5438882. 

Advanced Glycation End products in Bone Material Properties in Type 2 Diabetic Mice 

Impact Factor: 2.4   

Level of Evidence: V  

Sample Size: 12 female mice   

Study type: Rat Study   

Statistical Analysis: Kruskal- Wallis nonparametric tests   

Methods: 12 female mice were taken with severe onset of Type 1 Diabetes. The mice were all the same age. The femoral bones were taken from each of mice for analysis and then were stored in about 70% alcohol. Raman Microscopic Analysis was used in order to focus the laser beam on the periosteum.   

Results: The mice who had diabetes had a decrease in the mineral to matrix ratio. This suggests that high levels of AGE might be associated with bone deterioration. AGE might be a therapeutic strategy and target of focus when it comes to the strengthening of bones and the prevention of fractures.   

Furst JR, Bandeira LC, Fan WW, et al. Advanced Glycation Endproducts and Bone Material Strength in Type 2 Diabetes. J Clin Endocrinol Metab. 2016;101(6):2502-2510. doi:10.1210/jc.2016-1437

Association of accumulated advanced glycation end-products with a high prevalence of sarcopenia and dynapenia in patients with type 2 diabetes 

Impact factor: 3.761 

Level of evidence: IV 

Type of study: cross-sectional 

Sample size: 166 patients with type 2 diabetes, age 30 and older.  

Methods: Skin autofluorescence as a marker of AGEs, limb skeletal muscle mass index, grip strength, knee extension strength and gait speed were assessed. A diagnostic survey questionnaire was used to estimate physical activity in each patient. 

Statistical analysis: Intergroup comparisons of non‐sarcopenia and dynapenia, sarcopenia, dynapenia were measured using an unpaired one‐way analysis of variance for continuous variables, and a χ2‐test for categorical variables. Univariate and multivariate logistic regression analyses were used to calculate the cross‐sectional association of a low SMI, low grip strength, slow gait speed, low knee extension strength, sarcopenia, and dynapenia. The odds ratios and 95% confidence intervals were calculated using logistic regression models. Pearson's correlation coefficient and multiple regression analyses were used for correlation analysis between skin autofluorescence and other clinical parameters. P-value < 0.05 was considered statistically significant. 

Results: 7.2% of participants were diagnosed with sarcopenia and 13.9% with dynapenia. Patients ≥65 years had a significantly higher rate of sarcopenia, dynapenia, higher rate of low SMI, low grip strength, and low gait speed values than those who were less than 65 years old. Sarcopenic patients were older with longer duration of diabetes, and had a higher rate of incidental falls, skin AF and serum pentosidine, and lower BMI and body fat than did control patients. Dynapenic patients were also older with longer duration of diabetes, and had higher incidence of falls and skin AF, but had higher BMI and body fat than did control patients. Dynapenic patients had higher BMI, body fat and visceral fat area, and had a lower skin AF than sarcopenic patients did. 

Conclusion: Sarcopenia was observed in 7.2% of participants, and dynapenia was observed in 13.9% of participants. Skin autofluorescence was significantly higher in patients with sarcopenia and dynapenia. Sarcopenia and dynapenia are associated with incidental falls in patients with type 2 diabetes. AGE accumulation was significantly associated with low muscle strength and mass, and related to sarcopenia and dynapenia prevalence in type 2 diabetes. 

Mori H, Kuroda A, Ishizu M, Ohishi M, Takashi Y, Otsuka Y, Taniguchi S, Tamaki M, Kurahashi K, Yoshida S, Endo I, Aihara KI, Funaki M, Akehi Y, Matsuhisa M. Association of accumulated advanced glycation end-products with a high prevalence of sarcopenia and dynapenia in patients with type 2 diabetes. J Diabetes Investig. 2019 Sep;10(5):1332-1340. doi: 10.1111/jdi.13014. Epub 2019 Feb 19. PMID: 30677242; PMCID: PMC6717916.