LINUS PAULING INSTITUTE RESEARCH REPORT

Régis Moreau

Browning: The Dark Side of Sugar

Régis Moreau, Ph.D.
LPI Research Associate

Summary: Sugar can react with proteins in the body in a process called protein glycation. Oxidants can further chemically modify these glycated proteins, producing “advanced glycation endproducts” or AGE. The accumulation of these modified proteins has been implicated in diabetes and aging. In rats, the administration of a substance called aminoguanidine inhibited AGE formation and improved kidney function, as well as other age-related pathologies.

The first scientific account of non-enzymatic reactions involving the browning of sugar was published by the French biochemist Louis Camille Maillard in 1912. He reported that aqueous solutions of amino acids and glucose, or sugar, turned progressively yellow-brown when heated or stored under physiological conditions. This phenomenon, called protein glycation, involves the reaction of a sugar, such as glucose or fructose, with the amino group of proteins to form what is known as a Schiff base. While Maillard’s prediction that this reaction occurs in the human body, particularly in diabetic patients, went unnoticed for the next 50 years, the Maillard reaction became of major interest to researchers in food science and technology. Since the time our ancestors controlled fire, the value of cooking has been recognized for improving the flavor and digestibility of food. It also became apparent that cooked food could be stored longer than raw food. Over time, cooking practices have developed into an art. Food manufacturers have long realized that solutions of amino acids and sugar should not be heated or mixed together. Yet, thermal processes are used in the food industry to improve texture, color, and flavor, and to sterilize and pasteurize, enabling longer shelf-life and improving product safety. While the food industry exploits the Maillard reaction to improve existing products and to develop new products, unfortunately, not all Maillard products endow foods with positive characteristics, such as better flavor and color. For the industry, the goal is to maximize food flavor through the heat-induced Maillard reaction without impairing nutritive value or creating carcinogenic heterocyclic amines, which are formed during high temperature cooking of foods like meat and chicken that contain high amounts of protein. Well-known industrial applications of the Maillard reaction include the production of caramel, cola, coffee, brewed products, infant formulas, and baked products.

In contrast to the steady progress in understanding the chemistry and significance of the Maillard reaction in food technology, the potential harmful effects of the Maillard reaction in biological systems were largely overlooked until the discovery in 1971 by Trivelli of non-enzymatically glycated hemoglobin in diabetic patients. Glycation of proteins has been linked to a growing number of diseases and conditions,such as diabetes, cataract, and aging. Twenty years ago, protein glycation was considered responsible for diabetic complications. Gradually, the glycation hypothesis has evolved into the “advanced glycation endproducts” (or AGE) hypothesis, which focuses on non-enzymatic chemical modifications occurring subsequent to glycation and referred to as glycoxidation. As the name suggests, oxidants contribute to this process. Chemically, the Schiff bases formed from the reaction between sugar and amino acids in proteins can change into another compound called the Amadori product. Degradation of the Amadori product to so-called carbonyl compounds leads to irreversible sugar-derived protein compounds called adducts and protein crosslinks, which collectively are known as AGE. This mechanism—the formation, accumulation, and tissue toxicity of AGE—has emerged as one of the most coherent explanations for the development of many of the pathological complications of diabetes and normal aging.

Protein glycoxidation is of great interest in geriatric research because protein modifications are recognized as markers of and contributors to the aging process. While glycation and subsequent glycoxidation occur over a period of weeks, thereby affecting long-lived proteins like collagen, elastin, myelin, and crystallin, conditions like diabetes and aging, where oxidants and reactive carbonyls are elevated, promote interactions with shorter-lived proteins that impair specific functions. Since aging is associated with impaired handling of blood sugar and increased oxidative stress and DNA damage, the cellular constituents of older adults are at elevated risk for glycation/ glycoxidation. This risk becomes especially relevant in tissues with a slow rate of regeneration, such as the brain, blood vessels, and the heart. For this reason, organs and tissues of the cardiovascular system represent an ideal model to examine the extent of protein glycoxidation during aging. Moreover, mitochondria in the heart that produce chemical energy are very active metabolically and produce oxidants constantly. The production of these oxidants increases as we age. The subcellular mitochondria, also known as the “power house” of the cell, utilize sugars and lipids to generate the energy necessary for the heart to beat.

Nε-(carboxymethyl)lysine (CML) is the most abundant AGE in proteins in the body and appears to be a useful biomarker of aging. In our experiments, we found that the extent of protein-associated CML in the cardiovascular system (serum, aorta, and heart) of old rats (28-months old) was substantially greater than that of young rats (4-months old). Since there was an age-associated increase in total CML in serum proteins, we further examined the particular proteins being modified. Albumin was the predominant protein adducted by CML in the circulation. Since the extent to which albumin was modified did not change between young and old rats, this suggests that all potential sites in the protein are already modified at the age of 4 months and that other serum proteins must be modified by CML in an age-dependent manner. Further analyses showed that the CML-modified serum proteins included transferrin and the heavy and light chains of immunoglobins in the immune system and that old rats exhibited higher levels of CML associated with these proteins than did their young counterparts. Because serum immunoglobulins increased in old rats concurrently with CML modifications, the increase of these adducts may result from an accumulation of moderately modified immunoglobulins rather than from an increase in the ratio of adduct to protein. The accumulation of modified immunoglobulins indicated that old rats showed signs of primary amyloidosis, a disease of the immune system in which plasma cells produce excess amounts of antibodies that accumulate in the blood and, subsequently, in almost all organs. If excess antibodies are deposited in the kidneys, renal function may become impaired and lead to decreased waste clearance. In an attempt to dilute accumulating waste, old rats typically drink more water, as we observed in this study.

If we can prevent damage or remove existing damage caused by protein glycoxidation and maintain tissue integrity, we may be able to improve the health and vigor of the elderly. Based on our knowledge of the Maillard reaction, we administered a modified amino acid to rats as a means to trap carbonyl compounds and stop AGE formation. The trapping agent was aminoguanidine (AG), a compound not normally found in the body that inhibits AGE formation in the test tube. Previous animal studies have shown the beneficial effects of AG in ameliorating or preventing the complications caused by diabetes. In those studies, AG was able to either prevent or retard glomerular sclerosis, arterial stiffening, atherogenesis, and cardiac hypertrophy. We think that the potential usefulness of AG could be extended to normal aging, which is considered to share pathophysiological attributes with diabetes, such as insulin resistance and dyslipidemia. In our trial, AG was administered to old rats in the drinking water for 3 months, beginning at the age of 25 months. The animals had free access to food and water throughout the trial. We found that AG ameliorated the signs of aging exhibited by the untreated old rats. Specifically, AG significantly lowered CML content by 15, 44, and 28% in the serum, aorta, and heart, respectively. AG was more effective in lowering CML in the aorta and heart than in serum, reflecting the tissue compartmentalization and flux of CML, first originating in the circulation then migrating to and diluting out of organs. AG prevented the accumulation of immunoglobulins in the serum that we observed in untreated old rats and also normalized water consumption, which was elevated in untreated old rats. AG directly improves renal ultrastructure and function, thus facilitating rapid clearance of AGE-peptides, and also normalizes blood chemistry and water intake. We did not find any significant difference in mitochondrial proteins between young and old rats treated with AG, probably because glucose is excluded from mitochondria and thus does not chemically react with mitochondrial proteins. While glucose is an important precursor of CML, other sources, such as products from the degradation of lipids and the metabolism of amino acids, also play a role. Nevertheless, mitochondrial protein-associated CML could not be detected in rat hearts, perhapsdue to the rapid turnover of cardiac mitochondria and their proteins every 6-10 days.

Our results showed that old rats exhibit elevated CML in serum, aorta, and heart proteins, with possible negative repercussion on kidney function. In the serum, modified proteins included albumin, transferrin, and immunoglobulins. AG administration for 3 months significantly inhibited CML-protein formation in the aorta and heart, lowered serum content of CML-modified immunoglobulins, and normalized water intake. Thus, the extent of tissue protein glycoxidation that results in physiological impairment may be decreased through pharmacological and, perhaps, dietary interventions. The measurement of CML in tissues of the cardiovascular system may be of practical importance for assessing the effectiveness of various strategies for intervention in the aging process. We believe that the use of aminoguanidine as a pharmacological tool to prevent some of the deleterious consequences of aging in humans should be further investigated.

This work was supported by the Collins Medical Trust and the National Institute on Aging.

Last updated November 2004

 

Micronutrient Research for Optimum Health

 
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