Homocysteine, Vitamins, and Prevention of Vascular Disease

Transcription

1 MILITARY MEDICINE, 169, 4:325, 2004 Homocysteine, Vitamins, and Prevention of Vascular Disease Guarantor: Kilmer S. McCully, MD Contributor: Kilmer S. McCully, MD Within the past four decades, the efforts of investigators worldwide have established the amino acid homocysteine as an important factor in arteriosclerosis and diseases of aging. After its discovery in 1932, homocysteine was demonstrated to be an important intermediate in the metabolism of amino acids. However, little was known about the broader biomedical significance of homocysteine until 1962, when children with mental retardation, accelerated growth, dislocated ocular lenses, and frequent vascular thrombosis were found to excrete homocysteine in the urine. My study of two patients with homocystinuria caused by different inherited enzymatic disorders in 1968 disclosed advanced widespread arteriosclerotic plaques in both cases. This discovery led to the conclusion that homocysteine causes vascular disease by a direct effect on the cells and tissues of the arteries. This interpretation suggests that homocysteine is important in the pathogenesis of arteriosclerosis in persons with hereditary, dietary, environmental, hormonal, metabolic, and other factors predisposing them to hyperhomocysteinemia. Within the past decade, many major clinical and epidemiological studies have proven that hyperhomocysteinemia is a potent independent risk factor for vascular disease. According to the homocysteine theory of arteriosclerosis, insufficient dietary intake of the B vitamins, folic acid and pyridoxine, caused by losses of these nutrients during processing of foods, leads to elevation of blood homocysteine and vascular disease in the general population. The dramatic decline in cardiovascular mortality since the 1960s in the United States is attributed to fortification of the food supply by synthetic pyridoxine and folic acid. The recent Swiss Heart Study showed that B vitamins slowed restenosis in patients with coronary arteriosclerosis treated with angioplasty. Currently, more than 20 prospective, worldwide, interventional trials involving at least 100,000 participants are examining whether lowering plasma homocysteine levels with supplemental B vitamins will prevent mortality and morbidity from arteriosclerotic vascular disease. Discovery of the Homocysteine Theory of Arteriosclerosis n 1932, the American biochemist Vincent DuVigneaud discovered a new sulfur amino acid by treating methionine with I sulfuric acid. The molecular structure of this amino acid is similar to cysteine except for one extra carbon atom, hence the name homocysteine. Subsequent biochemical investigations established homocysteine as an important intermediate in sulfur amino acid metabolism and transmethylation reactions. Homocysteine was also found to support the growth of animals when fed in a diet containing a source of methyl groups, such as choline or methyl folate, and containing no methionine or cysteine. Chief, Pathology and Laboratory Medicine Service, Veterans Affairs Medical Center, 1400 Veterans of Foreign Wars Parkway, West Roxbury, MA and Director, Boston Area Consolidated Laboratories, VA Boston Healthcare System. This manuscript was received for review in September 2003 and was accepted for publication in October However, little was known about the broader biomedical significance of homocysteine until 1962, when children with mental retardation, accelerated growth, osteoporosis, skeletal abnormalities, dislocated ocular lenses, and propensity to thrombosis of arteries and veins were found to excrete homocysteine in the urine. Investigation of the biochemical abnormalities in these children with homocystinuria revealed deficient activity of cystathionine synthase, a pyridoxal phosphate-dependent enzyme, in most cases. As a result of this inherited enzyme deficiency, these children excrete homocystine in the urine, and the amino acids methionine and homocystine accumulate to high levels in the plasma. During Human Genetics Rounds at Massachusetts General Hospital in 1968, I learned of the fascinating case report of an 8-year-old boy with mental retardation, skeletal abnormalities, and dislocated ocular lenses who had died of a stroke in This case report was uncovered in 1965 by pediatricians who had identified homocystinuria in a niece of the boy. My review of the pathological findings in this archival case revealed the cause of death to be carotid arteriosclerosis with thrombosis and cerebral infarct. Through study of the original slides and tissues in this case, I also found that arteriosclerotic plaques were scattered through the arteries of the major organs. However, the possible connection between homocysteine and arteriosclerotic plaques was unclear from study of this case. Later in the year, I was fortunate to learn of another case of homocystinuria in a 2-month-old boy who had died several months earlier. This child was studied intensively at Massachusetts General Hospital, the National Institutes of Health, and Brandeis University. The investigators demonstrated deficient activity of the enzyme methionine synthase, a folate- and cobalamin-dependent enzyme. Because of this enzyme abnormality, the child excreted homocystine, cystathionine, and methyl malonic acid in the urine. The concentrations of homocystine and cystathionine in the plasma were elevated, and the concentration of methionine in the plasma was decreased. This is the first case in the medical literature of cobalamin C disease. My review of the pathological findings in the 1968 case with methionine synthase deficiency disclosed widespread and astonishingly advanced arteriosclerotic plaques throughout the arteries of the major organs. Because of the difference in the enzymatic and metabolic abnormalities in these two cases of homocystinuria, I was able to conclude that the amino acid homocysteine causes arteriosclerotic plaques by a direct effect on the cells and tissues of the arteries. Several years later, in 1974, investigators in Chicago demonstrated similar arteriosclerotic plaques in a child who had died with methylenetetrahydrofolate reductase deficiency and homocystinuria, the third major type of homocystinuria. Their findings independently corroborated my conclusion that homocysteine is an atherogenic amino acid. In my publication of these two interesting cases of homo- 325

2 326 Homocysteine, Vitamins, and Prevention of Vascular Disease cystinuria in 1969, I pointed out that this interpretation explains the finding of arteriosclerotic plaques in two important animal models of arteriosclerosis. 1 First, feeding a diet that is deficient in pyridoxine (vitamin B 6 ) causes arteriosclerotic plaques in monkeys. I suggested that deficiency of pyridoxine may cause elevation of blood levels of homocysteine, causing the arteriosclerotic plaques. Second, feeding a diet that is deficient in methyl group donors such as choline causes arteriosclerotic plaques in rats. I suggested that failure of transmethylation in these animals would lead to elevation of blood levels of homocysteine. Subsequent experiments have proven these two suggestions to be true. Thus, the metabolic abnormalities in these two animal models closely parallel the cause of homocystinuria in these two reported cases of human homocystinuria. Finally, in my publication from 1969, I suggested that elevation of blood homocysteine is likely to be of importance in the pathogenesis of arteriosclerosis in the general population without these rare inherited diseases of homocysteine metabolism. Thus, patients with hereditary, dietary, environmental, hormonal, metabolic, or other predispositions to arteriosclerosis are likely to have elevated blood levels of homocysteine, causing plaques by damage to the lining cells and tissues of arteries. This formulation of the homocysteine theory of arteriosclerosis attributes the primary vascular damage to homocysteine and the deposition of cholesterol and lipids to a secondary effect of the disease. Within the past decade, hundreds of major prospective and retrospective clinical and epidemiological studies have proven the underlying validity of the homocysteine theory by showing that hyperhomocysteinemia is a potent independent risk factor for vascular disease. Molecular Basis of Atherogenesis If homocysteine is atherogenic in children with different inherited enzymatic disorders, then administration to animals should demonstrate this effect experimentally. A series of experiments with rabbits showed that administration of homocysteine by parenteral injection or by dietary feeding results in arteriosclerotic plaques in aorta and peripheral arteries. High doses cause prominent plaques that closely resemble fibrous plaques in human arteriosclerosis and in homocystinuria. Some of the animals develop venous thrombosis and pulmonary embolism, effects that are found in patients with homocystinuria. The plaques are of the fibrotic type without cholesterol or lipid deposition, but feeding cholesterol or fats and cholesterol to animals produces fibrolipid plaques with prominent lipid deposits. Other investigators working with baboons or pigs obtained similar results. The molecular basis for production of arteriosclerotic plaques is related to the effect of homocysteine on cellular degeneration, damage to arterial intima, cellular growth, connective tissue formation, deposition of lipoproteins, and enhancement of blood coagulation. In each of these critical processes of atherogenesis, homocysteine plays a key role. Experiments with cell cultures taken from the skin of a child with homocystinuria show that an abnormal aggregated extracellular matrix results from binding of excess sulfate groups to the macromolecules. In the same cell cultures, a new pathway for conversion of the sulfur atom to sulfate was demonstrated by using radioactively labeled homocysteine. In this pathway, the sulfur of homocysteine is bound to retinoic acid (vitamin A acid) and is then oxidized to sulfite and sulfate for conversion to the sulfating coenzyme phosphoadenosine phosphosulfate. As the result of binding of homocysteine thiolactone to the carboxyl groups of extracellular proteoglycans, excess sulfate binding converts the macromolecules from a helical to a random coil configuration. Deposition of such a sulfated extracellular matrix is a characteristic observation within early arteriosclerotic plaques. Arteriosclerotic plaques also contain fragmented and degenerative elastic fibers. Other investigators have shown that homocysteine activates lysosomal elastase, leading to destruction and degeneration of elastic laminae. In addition, homocysteine causes cultured smooth muscle cells to produce excess collagen, a constituent of fibrous tissue. This finding explains the fibrosis that is characteristically found in human and experimental arteriosclerotic plaques. Arterial smooth muscle cells proliferate in arteriosclerotic plaques because homocysteine activates cyclins, signaling proteins that mediate cell division. The final result is a fibrous plaque with connective tissue changes of fibrosis, glycosaminoglycan deposition, destruction of elastic fibers, and calcification. Because of the accelerated growth of many children with homocystinuria, the growth-promoting properties of homocysteine were investigated in normal and hypophysectomized animals. Homocysteine causes accelerated growth of normal rabbits and guinea pigs, and homocysteic acid increases the growth of hypophysectomized rats that are treated with thyroxine. Assay of the plasma of the rats treated with homocysteic acid and thyroxine showed a release of insulin-like growth factor from the liver. This plasma factor causes growth by increasing the sulfation of epiphyseal cartilage in skeletal growth, among other physiological effects. Other investigators showed that homocystine increases sulfation of cartilage fragments when added directly to the assay for insulin-like growth factor. These experiments show that homocysteine mediates the accelerated skeletal growth in children with homocystinuria and also mediates the growth of smooth muscle cells in developing arteriosclerotic plaques. The initial phase in formation of arteriosclerotic plaques is characterized by endothelial and intimal damage, as shown in the rapidly advancing plaques in homocystinuria. Cellular apoptosis has been found by many investigators to be related to effects of homocysteine in endothelial cells, in neurons, and other cell types in culture and in experimental animals. In experiments with the free base form of homocysteine thiolactone, the skin of mice became ulcerated and inflamed when painted on the skin surface. Microscopic study shows inflammation, necrosis, and stromal and epithelial proliferation with angiogenesis, hypertrophy of nerves, and intravascular coagulation. In some animals, epithelial dysplasia is accompanied by early invasive squamous cell carcinoma of skin. These inflammatory, proliferative, and prothrombotic effects on tissues may be related to oxidant stress within cells affected by homocysteine. 2 Lipoproteins, including low-density lipoprotein (LDL) and high-density lipoprotein, contain homocysteine that is bound to apob protein by peptide bonds. The ratio of homocysteine within LDL to homocysteine within high-density lipoprotein is higher in patients with hypercholesterolemia than in normal controls.

3 Homocysteine, Vitamins, and Prevention of Vascular Disease Reaction of homocysteine thiolactone with normal human LDL in vitro causes increased density, increased electrophoretic mobility, and aggregation and precipitation of LDL particles. These homocysteinylated LDL particles are phagocytosed by cultured human macrophages to form foam cells. Many investigators believe that foam cell formation is a key feature of the atherogenic process. Deposition of cholesterol and lipids within arteriosclerotic plaques probably occurs by a similar process in vivo. Epidemiological and Observational Evidence The first human study of homocysteine in vascular disease in 1976 showed that oral methionine loading causes increased levels of homocystine and homocysteine cysteine mixed disulfide in the plasma of patients with coronary heart disease. Many subsequent studies showed that persons with coronary, cerebral, or peripheral arteriosclerosis have elevated levels of homocysteine in the blood. 3 In patients with early onset of arteriosclerosis, elevation of homocysteine is a more potent risk factor than cholesterol elevation and is similar in strength to the effect of smoking. The European Concerted Action Study showed that hyperhomocysteinemia potentiates the effect of hypertension, smoking, and cholesterol elevation on cardiovascular risk. The Hordaland Homocysteine Study showed that homocysteine levels are correlated with multiple known risk factors for coronary heart disease, such as smoking, lack of exercise, increasing age, lack of dietary fruits and vegetables, male gender, postmenopausal status, hypertension, cardiac hypertrophy, and excess coffee consumption. The Framingham Heart Study showed that the homocysteine levels of elderly participants are related to dietary deficiencies of vitamin B 6 or folate or to malabsorption of vitamin B 12. About two-thirds of participants were deficient in one of these three B vitamins, leading to hyperhomocysteinemia. The degree of carotid arteriosclerosis was shown to parallel the degree of homocysteine elevation in the same group of participants. The Physicians Health Study showed that physicians with elevated homocysteine levels have an increased risk of myocardial infarction over a 5-year period. Retrospective studies and most prospective studies have shown a correlation between homocysteine levels and risk of cardiovascular disease. A meta-analysis of these studies suggested that elevation of the homocysteine level is responsible for 10% of the U.S. mortalities from cardiovascular disease. Recent meta-analysis of 72 studies in which the genetic polymorphism of methylenetetrahydrofolate reductase was determined concluded that the association between homocysteine elevation and cardiovascular disease is causal. On this basis, lowering of the homocysteine level in the population by 3 M/L would be expected to reduce coronary disease risk by 16%, deep vein thrombosis by 25%, and stroke by 24%. The Nutrition Canada Study showed that a low plasma level of folate is associated with an increased risk of mortality from coronary heart disease. The Nurses Health Study showed that decreased dietary folate or vitamin B 6 is associated with increased risk of cardiovascular mortality and morbidity. Dietary intake of less than 350 g per day of folate or 3 mg per day of vitamin B 6 increased cardiovascular risk, and moderate alcohol intake significantly reduced risk at all levels of B vitamin intake. The Third National Health and Nutrition Examination Survey study shows that increased plasma homocysteine level is associated with increased risk of myocardial infarction. Survival studies from Israel and Oregon show that elevation of blood homocysteine is directly correlated with risk of mortality. A study from Norway showed that survival of patients with angiographically proven coronary heart disease is inversely related to blood homocysteine levels. A study of stored blood samples from the 1970s in different countries showed that mortality from cardiovascular disease is directly related to homocysteine levels. Countries such as Spain, France, and Japan with low mortality rates have levels of 7 to 8 M/L homocysteine. Countries such as Finland, Northern Ireland, and Northern Germany with high mortality rates have levels of 10 to 11 M/L homocysteine. These studies suggest that blood levels of homocysteine below 8 M/L are associated with decreased mortality from vascular disease. Homocysteine Theory of Arteriosclerosis 327 As formulated during the period of 1969 to 1975, the homocysteine theory of arteriosclerosis implicates the elevation of blood homocysteine levels as the key factor in the genesis of arteriosclerotic disease in the general population. 4 Insufficient dietary intake of the B vitamins, folic acid, vitamin B 6, and vitamin B 12 leads to the elevation of blood homocysteine levels. The dietary sufficiency of B 6 and folate is inadequate in populations consuming processed foods because these sensitive vitamins are destroyed by heat, milling of grains, extraction of sugar or oils, chemical additives, or other traditional methods of food processing. Dietary vitamin B 12 is usually adequate, and the vitamin is stable in most forms of food processing. However, insufficient absorption of B 12 becomes a problem in persons in the older decades of life because of loss of gastric acid and intrinsic factor, secondary to atrophy of gastric mucosa and infection by Helicobacter pylori. The only source of homocysteine is from metabolism of the essential amino acid, methionine, in the liver. The methionine content of animal proteins is two to three times greater than that of plant proteins. Metabolic regulation by pyridoxal phosphate, methyl tetrahydrofolate, and methyl cobalamin controls the production of homocysteine from methionine. Fresh fruits and vegetables contain abundant pyridoxine and folate, and populations consuming these foods tend to have lower homocysteine levels and lower rates of vascular disease than populations consuming processed foods that are deficient in these vitamins. The only dietary sources of vitamin B 12 are foods of animal origin, such as meat, fish, and dairy products. Because of this, vegans who eat no meat or dairy foods have a higher level of homocysteine compared with those consuming these foods. Deficiency of dietary vitamin B 6 causes elevation of blood homocysteine levels after a meal containing protein, and deficiency of dietary folate or vitamin B 12 causes elevation of fasting blood homocysteine levels. Dietary betaine may help to prevent the elevation of plasma homocysteine through a separate enzymatic pathway of transmethylation of homocysteine to methionine in the liver. Dietary protein is also a factor in controlling homocysteine levels, as shown by studies from Maryland and West Africa. Protein deprivation leads to the elevation of homocysteine levels and increased dietary protein leads to lower homocysteine levels. The reason for this observation is that methionine is a

4 328 Homocysteine, Vitamins, and Prevention of Vascular Disease precursor of adenosyl methionine, a metabolic regulator of methionine metabolism in the liver. Dietary protein deprivation causes decreased synthesis of adenosyl methionine, leading to decreased conversion of homocysteine to cystathionine. Abundant dietary protein causes increased synthesis of adenosyl methionine, leading to increased conversion of homocysteine to cystathionine. The amounts of dietary or supplemental B vitamins that are needed to prevent hyperhomocysteinemia have been studied in clinical and interventional trials. Generally, 400 g of folic acid, 3 mg of vitamin B 6, and 3 g of vitamin B 12 are sufficient to maintain a desirable homocysteine level less than 8 M/L in a healthy, young person. However, in a nutritionally depleted older person with vascular disease, higher intakes of these B vitamins may be necessary to attain a desirable level of blood homocysteine. Larger intakes of these vitamins through supplementation are generally quite safe, as no toxicity has been reported from moderate doses of these B vitamins. Folate can mask the neurological abnormalities associated with vitamin B 12 deficiency, but cases with inadequate B 12 are extremely rare because folate is almost always combined with B 12 in multivitamin preparations. Reports of neuropathy from megadoses of pyridoxine are very rare and may be caused by contaminants of the synthetic vitamin because neuropathy is not observed in patients with homocystinuria or carpal tunnel syndrome treated with doses of 50 to 500 mg per day for a period of years. No toxicity of vitamin B 12 has ever been reported for humans or animals, even in extremely large doses. Dietary choline, a constituent of wheat germ, vegetables, meats, liver, eggs, and seafood, is converted to betaine for transmethylation of homocysteine to methionine in liver. The daily requirement of dietary choline is about 550 mg per day. Betaine, a constituent of wheat germ, spinach, beets, liver, and seafood, lowers plasma homocysteine when given in doses of 1 g or more per day. The Nurses Health Study showed that dietary intakes of 400 g of folic acid and 3 mg of pyridoxine are necessary to minimize mortality and morbidity from cardiovascular disease. These figures agree with the results of the Framingham Heart Study, which show that these amounts of dietary folic acid and pyridoxine are necessary to keep homocysteine levels in the desirable range. The Food and Drug Administration requirement for folate fortification of enriched refined grain foods in 1998 increased the plasma folate levels and decreased the homocysteine levels of the U.S. population. In addition, rates of neural tube birth defects have significantly declined in the United States since folate fortification because of control of homocysteine levels in mothers and infants. The current Recommended Dietary Intake for pyridoxine is 1.4 to 1.7 mg per day for men, women, and pregnant mothers. Pyridoxine is not added to fortified grain products, and the current dietary intake of approximately 1.5 mg per day is too low to prevent vascular disease, judging from the results of the Nurses Health Study. Factors other than dietary deficiency are implicated in production of hyperhomocysteinemia. About 12% of the population contains a genetic variant of the enzyme methylenetetrahydrofolate reductase, which leads to hyperhomocysteinemia and an increased dietary requirement for folate. Other genetic polymorphisms involving methionine synthase and cystathionine synthase are being evaluated for effects on production of hyperhomocysteinemia. To date, molecular geneticists have identified over 200 different alleles affecting homocysteine metabolism. Another common condition leading to the elevation of blood homocysteine is chronic renal failure and end-stage renal disease. These individuals commonly have homocysteine levels of 20 to 30 M/L, and the annual cardiovascular mortality in this group approaches 25% per year. Hypothyroidism leads to elevation of blood homocysteine and an increased risk of cardiovascular disease. Treatment with thyroxine restores blood homocysteine to normal levels. Antifolate drugs, such as methotrexate or dilantin, cause elevation of homocysteine levels. Nitrous oxide, used as a dental anesthetic, antagonizes vitamin B 12, leading to hyperhomocysteinemia. Sulfasalazine, which is frequently used in inflammatory bowel disease, antagonizes vitamin B 6, causing hyperhomocysteinemia. The antimetabolite, azaribine, used in treatment of refractory psoriasis, also causes hyperhomocysteinemia by antagonizing vitamin B 6. Azaribine was recalled by the Food and Drug Administration because of numerous cases of vascular thrombosis, stroke, and myocardial infarction that were attributed to homocysteine elevation. Many other commonly used drugs, such as diuretics and antiepileptic agents, cause mild elevation of blood homocysteine. Lipid-lowering drugs, including clofibrate and gemfibrozil, cause elevation of homocysteine, but statin drugs do not affect blood homocysteine levels. Homocysteine, Vitamins, and Prevention of Vascular Disease As reported by the Centers for Disease Control and Prevention, the mortality from coronary heart disease has declined more than twofold since the mid-1960s. At a national conference held at the National Institutes of Health in 1979, the factors that may be responsible for this dramatic decline were examined in detail. Dietary fat and serum cholesterol levels have changed very little during this time period, and smoking, exercise, coronary care units, hypertension, and other suspected factors could not account for the decline. Starting in the 1960s, synthetic pyridoxine (vitamin B 6 ) was added to the U.S. food supply in the form of enriched cereals and vitamin supplements. Several years later, synthetic folic acid was also added to the food supply, and in 1998, the Food and Drug Administration mandated the addition of folic acid to enriched grain foods. Studies by the Framingham Heart Study show that serum folate levels increased and serum homocysteine levels declined after fortification. The addition of these key B vitamins to the food supply accounts for the decrease in cardiovascular mortality by lowering plasma homocysteine levels in a susceptible population. In an autopsy study of 200 consecutive deaths among veterans, two-thirds of the cases with the most severe arteriosclerosis had no evidence of hypertension, renal failure, elevated serum cholesterol, or diabetes. Large-scale clinical studies, on the other hand, indicate that about 80% of patients with coronary heart disease have at least one of the four most significant risk factors for vascular disease: smoking, elevated cholesterol, diabetes, or hypertension. Estimates of the fraction of the population with elevated homocysteine levels vary from 10% to 40%, depending upon the definition of normal levels. In the United

5 Homocysteine, Vitamins, and Prevention of Vascular Disease States today, the mean homocysteine level for the population is 8.9 M, as determined by the Third National Health and Nutrition Examination Survey. In most studies, the predictive value of plasma homocysteine is independent of the traditional risk factors. Certainly, serum cholesterol is predictive of vascular disease only in men under age 45, as women and older men reveal no correlation between serum cholesterol and vascular disease risk. There is no doubt that supplemental B vitamins will lower homocysteine levels in the majority of subjects with or without vascular disease. In the Warsaw study in 1989, moderate doses of vitamin B 6, folic acid, vitamin B 12, riboflavin, choline, and troxerutin (citrus bioflavonoid) not only lowered homocysteine levels significantly, but serum cholesterol and apob protein of LDL were also significantly lowered. Current interventional studies have found that modest doses of these vitamins, 500 g of folic acid, 10 mg of vitamin B 6, and 100 g of vitamin B 12, will lower plasma homocysteine levels to the desirable range in the majority of participants. In older individuals, or in persons with renal failure, higher doses may be necessary to lower plasma homocysteine levels to the desirable range. At the present time more than 20 prospective, worldwide, interventional trials involving at least 100,000 participants are examining whether lowering plasma homocysteine levels by supplemental B vitamins will prevent mortality and morbidity from arteriosclerotic vascular disease. At the recent Fourth International Conference on Homocysteine Metabolism, held in Switzerland, the problem of determining the efficacy of this approach in fortified and unfortified populations was discussed. The opinion was expressed that analysis of multiple trials involving large numbers of participants will be needed to draw significant conclusions about the possible role of reducing elevated homocysteine levels in prevention of vascular disease. In a current trial sponsored by the Department of Veterans Affairs, 2,000 patients with renal failure are being studied to determine whether high-dose folic acid, pyridoxine, and vitamin B 12 will reduce mortality and morbidity from arteriosclerotic disease. In London, Ontario, Canada interventional studies with moderate doses of B vitamins in patients with cerebral arteriosclerosis showed that the plaque area stopped progressing and began to regress, as determined by quantitative ultrasound imaging of carotid arteries. In the Swiss Heart Study, interventional studies of B vitamins in patients with angiographically demonstrated coronary heart disease showed that restenosis of stented and nonstented arteries was slowed by doses of 1 mg of folic acid, 400 g ofb 12, and 10 mg of B 6 compared with placebo controls. In addition, adverse cardiac events were significantly decreased in 1 year of follow-up observation. The effect is most pronounced in small coronary arteries, where restenosis is more significant that in larger stented arteries. Multiple studies of dementia by the Framingham Heart Study, the OPTIMA study of Oxford, England, and the Nun Study have shown a correlation of elevation of blood homocysteine with risk of Alzheimer s disease, cognitive decline, and vascular dementia. In the Framingham Study, participants in the highest quartile of blood homocysteine over 15 M were significantly more susceptible to development of Alzheimer s disease than those in lower quartiles. Interventional studies in elderly subjects have demonstrated some protection against cognitive decline by B vitamins, especially B 12 and folic acid. References McCully KS: Vascular pathology of homocysteinemia: implications for the pathogenesis of arteriosclerosis. Am J Pathol 1969; 56: s 2. McCully KS: Chemical pathology of homocysteine. I. Atherogenesis. Ann Clin Lab Sci 1993; 23: McCully KS: Homocysteine and vascular disease. Nat Med 1996; 2: McCully KS: Homocysteine, folate, vitamin B6 and cardiovascular disease. J Am Med Assoc 1998; 279: Two Air Force attendees share a moment during a meeting break