Advanced glycation end products in diabetic and non-diabetic human subjects suffering from cataract.
Sample Size: 50 cataracts patients with T2DM, 50 without T2DM
Protocol: ELISA and western blotting was used to quantify AGE and N- (carboxyethyl) lysine (CEL) in cataractous lenses.
Statistical Analysis: Student’s t-test
Type of study: Cross-sectional
Results: AGE and CEL levels were significantly higher in diabetic patients with cataracts compared to control, p-value: 0.001.
Discussion/Conclusion: The level of AGE formation contributes to cataract formation. Cataract formation occurs earlier in diabetic patients.
Hashim Z, Zarina S. Advanced glycation end products in diabetic and non-diabetic human subjects suffering from cataract. Age (Dordr). 2011 Sep;33(3):377-84. doi: 10.1007/s11357-010-9177-1. Epub 2010 Sep 15. PMID: 20842534; PMCID: PMC3168597.
Correlation Between Vitreous Advanced Glycation End Products, and D-dimer with Blood HbA1c Levels in Proliferative Diabetic Retinopathy
Type of Study: This was a cross sectional study that was done in vivo.
Sample Size: 47 patients
Methods: There were two groups in this study and were named controlled and uncontrolled. The controlled group had a Hb1Ac level that was greater than 7 % whereas the uncontrolled group had a Hb1Ac level that was less than 7%. There were also statistical studies that were done afterwards to assess the correlation between the Hb1Ac levels, the AGE levels in the virtuous humor, and also the levels of D-dimers.
Statistical Tests ran/Results: There was positive correlation (p<.05) of AGE levels in the patients who had uncontrolled hyperglycemia and controlled hyperglycemia. There was also significant correlation between the amount of AGE in the blood and the amount of AGE in the vitreous humor. There were no significant levels of difference between the amount D-dimer between the uncontrolled vs controlled diabetes. They used a calculation to predict the amount of AGEs that would be present in the eye from the amount of AGE in the blood using this equation vitreous AGEs = -1.442+ (1.740xblood HbA1c).
Discussion/Conclusion: A complication that develops as a result of diabetes (both Type and Type 2 ) is called diabetic retinopathy. It occurs as a result of tissue damage to the Retina. In this study , there were two groups who had uncontrolled hyperglycemia and controlled hyperglycemia. This distinction was made on the basis of HB1aC levels. The study found that there were increased levels of AGE in people who had uncontrolled hyperglycemia. There was also correlation between the amount of AGE that was in the blood and the amount of AGE in the eye. Advanced End Glycation products may be a target of synthesizing a cure for diabetes.
Loho T, Venna V, Setiabudy RD, et al. Correlation Between Vitreous Advanced Glycation End Products, and D-dimer with Blood HbA1c Levels in Proliferative Diabetic Retinopathy. Acta Med Indones. 2018;50(2):132-137.
Lens autofluorescence ratio as a noninvasive marker of peripheral diabetic neuropathy.
Impact factor: 3.007
Level of evidence: IV
Sample size: 160 T2DM patients
Protocol: All subjects underwent a lens fluorescence ratio (LFR) measurement. Diabetic peripheral neuropathy (DPN) was defined as the presence of neuropathic pain or feet sensory loss (or both). Neurothesiometer, monofilament test, and DN4 test results were used to diagnose DPN.
Statistical Analysis: 1-sample Kolmogorov–Smirnov test was used to assess the distribution of the data. Numerical variables in different patients were compared using either the t test or Mann–Whitney test. Correlation analyses were performed using the Pearson or Spearman correlation test. Linear regression analysis was used to show a detailed relation of parameters. Categorical variables were analyzed by the chi-squared test. Statistical significance was defined as p < 0.05.
Study Type: Cross sectional
Results: The LFR of 43 patients was higher than the expected levels. According to the DN4 questionnaire, 35 of 160 patients had neuropathic pain. 37 patients had higher vibration perception thresholds than expected. The monofilament test showed that 42 patients seemed to be affected by DPN. All of the tests individually showed that patients with higher LFR had more problems related to DPN. High LFR had a sensitivity of 50% and a specificity of 81% in the diagnosis of DPN. Although there was no significant difference in fasting blood glucose levels, HbA1c levels were higher and diabetes duration was longer in patients with higher LFR.
Conclusion: Diabetic neuropathy occurs as a consequence of chronic hyperglycemia, which leads to peripheral nerve injury through the enhanced formation of AGEs, increased cytokine release, activation of protein kinase C, and oxidative stress. AGEs accumulate in lens crystallins, nerve myelin, and skin collagen irreversibly. Lens autofluorescence is increased in patients with higher AGE levels, but because there is no clinical device that can measure this, this study used LFR to see if it is diagnostic for DPN.
Sertbas M, Sertbas Y, Uner OE, Elarslan S, Okuroglu N, Ak F, Dayan A, Ozdemir A. Lens autofluorescence ratio as a noninvasive marker of peripheral diabetic neuropathy. Pol Arch Intern Med. 2019 Mar 29;129(3):175-180. doi: 10.20452/pamw.4449. Epub 2019 Feb 14. PMID: 30762026.
Increased retinopathy occurrence in type 1 diabetes patients with increased serum levels of the advanced glycation endproduct hydroimidazolone
Impact Factor: 3.362
Summary: A cross sectional study was done in a Scandinavian ophthalmology clinic that looked at 61 outpatient T1DM. The goal of the study was to determine how advanced end glycation products on diabetic retinopathy. They took blood samples to evaluate AGE’s level and pictures on the retina which compared between patients with retinopathy and those without. The study found that increased levels of AGE’s was significant in patients with retinopathy and therefore associated (P value of 0.022). They found that this conclusion aligns with other studies that were done on T2DM.
Fosmark DS, Berg JP, Jensen AB, Sandvik L, Agardh E, Agardh CD, Hanssen KF. Increased retinopathy occurrence in type 1 diabetes patients with increased serum levels of the advanced glycation endproduct hydroimidazolone. Acta Ophthalmol. 2009 Aug;87(5):498-500. doi: 10.1111/j.1755-3768.2008.01300.x. Epub 2008 Jul 9. PMID: 18631328.
Decrease in the glyceraldehyde derived advanced glycation end products in the sera of patients with Vogt-Koyanagi-Harada disease
Impact factor: 3.611
Level of evidence: IV
Type of study: prospective cohort, in vivo
Sample size: 31 patients with active VKH. 33 healthy control patients.
Methods: Serum samples were obtained from 31 VKH patients--20 of which were treated with systemic corticosteroids--and 33 healthy control patients. A competitive enzyme linked immunosorbent assay using AGE-2 polyclonal antibody was used to determine serum AGE-2 levels.
Statistical analysis: Mann-Whitney U test and paired t-test
Results: The mean AGE-2 level in the sera of patients with VKH disease was 4.91 U/ml, which was significantly lower than that of the healthy control subjects (8.32 U/ml). The average serum AGE-2 level significantly increased to 13.49 U/ml after the patients were treated with systemic corticosteroids. The fasting plasma glucose levels of the corticosteroid treatment group were measured before and after as well.
Conclusion: Most VKH patients have glucose intolerance at the onset of the disease, which usually recovers after treatment with systemic corticosteroids. This explains why higher levels of fasting plasma glucose were found at the acute disease phase, while lower levels of plasma glucose were found after systemic corticosteroid administration. This pattern of glucose intolerance has also been seen in active rheumatoid arthritis which improved after corticosteroid treatment as well. Overall, this study suggests AGE-2’s involvement in VKH onset.
Kitamura M, Kitaichi N, Takeuchi M, Kitamei H, Namba K, Yamagishi SI, Iwabuchi K, Onoé K, Ohno S. Decrease in the glyceraldehyde derived advanced glycation end products in the sera of patients with Vogt-Koyanagi-Harada disease. Br J Ophthalmol. 2005 Nov;89(11):1407-9. doi: 10.1136/bjo.2005.072678. PMID: 16234440; PMCID: PMC1772931.
Parainflammation associated with advanced glycation endproduct stimulation of RPE in vitro: implications for age-related degenerative diseases of the eye
Impact Factor: 3.67
Level of evidence: V
Type of study: review
Summary: The purpose of this study is to determine the effects of AGE stimulation on pro and anti-inflammatory pathways in the primary culture of human retinal pigment epithelial cells. The researchers identified the cellular pathways activated by AGE stimulation by exploring the changes in genome expression and validated several gene products by suspension array technology.
AGEs accumulate in drusen associated with AMD and promote parainflammation. AGE stimulation of RPE cells trigger inflammation associated gene changes. Additional pathways triggered by AGE include proteasome degradation and caspase signaling. Interferon inducible products CXCL11 and RSAD2 are highly upregulated in RPE after AGE stimulation.
Important methods: RPE cells were isolated from human fetal donor eyes. AGEs were obtained by modification of bovine serum albumin using a Maillard reaction from published protocols. The RPE cells were treated with the AGE concentration. In the experiments, researchers studied the response of healthy RPE cells after 24h of AGE stimulation.
Statistical tests ran: Statistical comparisons of western blot data were determined by homoscedastic one-tailed Student’s t-test. Differences were considered statistically significant when p < 0.001.
Results: The results indicated that AGEs trigger an adaptive response from RPE cells. The data reflects that AGE can elicit a complex response from RPE cells and are one of the key players in the outer retina and AMD. The data may reflect an “adequate” immune response from healthy RPE cells to AGE stimulation. If RPE cells were exposed to AGE chronically, there may be a tipping point where the stresses overwhelm RPE cell’s adaptive mechanisms leading to dysfunction and death. Evidence of a chronic inflammatory response in which several of the proinflammatory mediators studied here was recently shown in an immunohistochemical study of the postmortem eye with drusen.
Limitations: The AGEs were from BSA which is non-physiologic and may not necessarily correlate with the exact physiological activities of AGE found in human plasma.
Lin, T., Walker, G. B., Kurji, K., Fang, E., Law, G., Prasad, S. S., Kojic, L., Cao, S., White, V., Cui, J. Z., & Matsubara, J. A. (2013). Parainflammation associated with advanced glycation endproduct stimulation of RPE in vitro: implications for age-related degenerative diseases of the eye. Cytokine, 62(3), 369–381. https://doi.org/10.1016/j.cyto.2013.03.027
Receptor for advanced glycation end products is upregulated in optic neuropathy of Alzheimer's disease
Impact Factor: 5.32
Level of evidence: V
Type of study and any information related to it: case control study
Sample size: 10 donor patients with AD (Alzheimer's disease) (n=3) non Alzheimer's disease control (n=13)
Summary: The purpose of this study was to investigate the role of RAGE (receptor for advanced glycation end products) and the pathogenesis of Alzheimer's disease optic neuropathy. It is thought that RAGE can play a role in signal transduction, leading to amplification and perpetuation of inflammatory processes. Both qualitative observation and quantitative analysis using imaging software were used to document the extent of RAGE in neural tissues. In the AD samples the intensity and extent of RAGE expression was more prominent compared to controls.
Important methods: The brain tissue samples were stained with hematoxylin eosin. The samples were incubated with primary antibodies for RAGE specific antibodies. The intensity of the immunoreactivity in cells at the hippocampus were scored on a scale of 0-3, 0= no staining and 3= strong staining. Additional hippocampal samples were homogenized in a SDS lysis buffer and collected for western blot analysis.
The optic nerve specimens were obtained and immersion-fixed in 10% neutral buffered formalin following enucleation of the eyes. The tissues were then processed for paraffin embedding and placed on microscope slides for immunohistochemistry. The samples were then examined using indirect immunostaining for the human RAGE antigen. The tissue was incubated with a monoclonal anti-human RAGE antibody. The purpose of incubating tissue with RAGE antibody was to determine the association of RAGE with astrocytes
Statistical tests ran: Statistical analysis were performed using the Wilcoxon rank sum test, fisher’s exact test, Pearson test and spearman test. Two-tailed P values of .05 or less were considered statistically significant.
Results: The quantitative analysis confirmed that the expression of RAGE was significantly greater in the AD group compared to the controls. The extent of RAGE immunolabeling was 6.78 +/- 2.99% in the AD optic nerves and .57 +/- in .3% in the control optic nerves. THe analysis determined that the intensity of RAGE increased in both AD and control groups. The increase in RAGE was significantly greater in the AD group compared to the controls. In the control group, there was no significant correlation between age and RAGE. However in the AD group, there was a significant correlation between RAGE and gender, duration of AD, pathologic stage, clinical dementia rating score or mini mental state examination score.
Conclusion: This study demonstrates that there is an increased expression of RAGE in AD optic nerves compared to controls. The study also showed that RAGE increases steadily with age but much more rapidly with AD. RAGE plays a role in inflammatory and immune systems. RAGE mediated signaling induces the secretion of proinflammatory cytokines which could be a causative agent in the pathophysiology of AD optic neuropathy. These findings suggest that RAGE expression in the microvasculature could be a major pathogenic event in AD optic neuropathy by increasing the inward flux of amyloid- beta into the optic nerves.
Limitations: The sample size is small. This is a study done on autopsy optic nerve specimens.
Wang MY, Ross-Cisneros FN, Aggarwal D, Liang CY, Sadun AA. Receptor for advanced glycation end products is upregulated in optic neuropathy of Alzheimer's disease. Acta Neuropathol. 2009 Sep;118(3):381-9. doi: 10.1007/s00401-009-0513-4. Epub 2009 Mar 11. PMID: 19277685; PMCID: PMC2867613.
Measurement of lens autofluorescence can distinguish subject with diabetes from those without.
Impact Factor: 2.7
Sample Size:178 subjects
Study Type: Cross Sectional In Vivo Study
Summary: This study was aimed at creating a tool that aids in the detection and the diagnosis of Diabetes in patients. A ClearPath DS -120 Lens Fluoresence Biomicroscope was used to correct for age and subtract the value of the normal patients. The sensitivity of the test was found to be 67% and the specificity was found to be 94%. The standards that were used were from the National Institute of Standards and Technology. As a patient gets older, and is diabetic, the amount of fluorescence that is seen in his eye is measured increases. This means the amount of advanced age glycation product that is seen in the eye is increased with more severe cases of diabetes. This tool can be used for early detection of diabetes for more efficient and earlier diagnosis of the disease.
Cahn F, Burd J, Ignotz K, Mishra S. Measurement of Lens Autofluorescence Can Distinguish Subjects With Diabetes From Those Without. J Diabetes Sci Technol. 2014 Jan;8(1):43-49. doi: 10.1177/1932296813516955. Epub 2014 Jan 1. PMID: 24876536; PMCID: PMC4454118.