Homocysteine: One More Piece In The Puzzle

Homocysteine: One More Piece In The Puzzle?

Dr. S.M.Sadikot (Mumbai)
Consultant in Endocrinology,
Diabetes and Metabolic Disorders,
Jaslok Hospital and Research Centre, Mumbai.

Atherosclerosis is too complex to be explained by any one mechanism or cause. To date, many factors have been identified which have been shown to be a risk factor for the development of the atheroma plaque. At the same time, it is now being increasingly appreciated that the traditional risk factors for cardiovascular disease (CVD) may account for only one half to two thirds of the actual risk. In the past, when a patient was considered at risk for atherosclerotic heart disease (AHD), the treating physicians began management by "rounding up the usual suspects": smoking, obesity, hypercholesterolemia, family history, physical inactivity, diabetes mellitus, hypertension, and other comorbidities. Yet these risk factors cannot account for all CVD. It is easy to blame it on the "genes" and it is accepted that a genetic tendency does play a very important role in predisposing a person to cardiovascular disease. It becomes important to try and identify other risk factors, especially those that can be easily modified or corrected. Some of the factors whose role is being seriously investigated include estrogen deficiency, lipoprotein(a), plasma fibrinogen, plasminogen-activator inhibitor type 1, endogenous tissue plasminogen activator (tPA), C-reactive protein , and homocysteine.

In the late 1960s, McCully became the first to hypothesize that elevated plasma homocysteine concentrations could cause atherosclerosis. Dr.Kilmer McCully, then a Harvard pathologist, observed severe atherosclerosis in two young children with rare diseases marked by very high homocysteine levels. Children with this defect typically succumb at an early age to complications of arteriosclerosis. If great excesses of homocysteine can cause this, McCully reasoned, could moderately elevated levels contribute to heart disease in middle-aged and older people?

Although not accepted as a theory in the early days, since then, numerous case-control and cross-sectional studies have yielded significant epidemiologic evidence to validate this relationship.

Researchers in at least three large, well-known studies have examined the complex association between hyperhomocysteinemia and CVD -- and ways to weaken that association. In one arm of the Physicians' Health Study, 14,916 male physicians without known atherosclerosis were followed for an average of five years after baseline homocysteine measurement. In an analysis of data from 271 men who subsequently developed MI and paired controls, researchers found a relative risk for MI of 3.1 among men with homocysteine levels in the highest 5%, compared with those in the lowest 90%; that risk rose to 3.4 after adjustment for diabetes, hypertension, and other potential confounders.

In a second arm of the Physicians' Health Study, Chasan-Taber et al prospectively examined plasma levels of folate and vitamin B6 in relation to subsequent MI occurrence. Over 7.5 years, 333 men experienced MI; in a comparison with paired controls, the researchers found that men with the lowest 20% of folate levels had a relative risk for developing MI of 1.4 (and those with the lowest 20% of vitamin B6 values, 1.5), compared with those in the remaining 80%. During the first half of follow-up, the authors note, men with homocysteine levels in the top 5% had a nearly threefold increase in risk of MI. Though "compatible" with the hypothesis that inadequate dietary intake of folate and vitamin B6 contribute to risk of MI, these data were not considered statistically significant.

Ten-year stroke incidence and all-cause and CVD mortality have been examined in participants from the Framingham Heart Study cohort whose nonfasting plasma total homocysteine levels were measured at baseline examinations. In the stroke substudy, 165 incident strokes occurred among 1,947 subjects who had been divided into quartiles by homocysteine level, lowest to highest. After adjusting for age, sex, systolic blood pressure, diabetes, smoking, and history of atrial fibrillation and coronary heart disease, the investigators determined that, compared with quartile 1, participants' relative risk for stroke rose to 1.32 in quartile 2, 1.44 in quartile 3, and 1.82 in quartile 4.

In the second Framingham substudy, 653 deaths, including 244 CVD deaths, occurred over 10 years. Persons with baseline homocysteine levels above 14.26 µmol/L had an estimated relative risk for all-cause mortality of 2.18, and for CVD death of 2.17, when compared with subjects whose homocysteine levels were lower than 14.26 µmol/L.

A Tufts University study of the elderly adds to rapidly mounting evidence that blood levels of the amino acid, homocysteine, can predict odds for having a stroke or heart attack. The study showed that the higher the homocysteine level, the greater the chance of carotid artery obstruction, a warning sign of increased risk for both stroke and coronary heart disease. Another finding was that odds for carotid blockage also rose as levels of folic acid and vitamin B6 dropped. Based on their results, the authors propose clinical trials of the vitamins to determine whether fatal and nonfatal vascular disease in the elderly can be reduced. The study, headed by Jacob Selhub, Ph.D., of the Human Nutrition Research Center at Tufts, was published in the February 2, 1995 issue of the New England Journal of Medicine (Vol.332: pp. 286-291).

Using noninvasive ultrasound imaging to measure the degree of carotid artery narrowing, Selhub and his group examined 418 men and 623 women who participated in the Framingham Heart Study. The subjects, ranging in age from 67 to 96, were divided into two groups. The first included people in whom no more than 24 percent of a carotid artery was obstructed. The second group consisted of those with a carotid blockage of at least 25 percent, a cutoff point above which stroke and coronary heart disease rates have been shown to rise.

The more dangerous obstructions were detected in 43 percent of men and 34 percent of women. An examination of the relationship between these blockages and the subjects' blood level of homocysteine strongly implicated homocysteine as an independent risk factor for vascular disease. Among men in the study, the odds for carotid blockage were more than twice as high in the 25 percent of the group with the highest homocysteine levels as in the bottom quartile. Disease risk increased gradually as homocysteine levels rose. Although women's risk did not increase with moderately elevated readings and somewhat fewer women with the highest levels had carotid blockages, the link between homocysteine levels and vascular disease was also statistically significant.

An interesting observation in both sexes was that risk began to rise at levels as low as 11.4 umol per liter, which were previously considered normal. This finding suggests that, as in the case of cholesterol readings, norms may need to be redefined, say the authors.

Among 27 studies of homocysteine and vascular disease cited by the University of Washington review was a Harvard project involving 15,000 physicians (JAMA, vol. 268, pp.877 -81). The research, reported in 1992, showed that although relatively few of the doctors had coronaries, those in the five percent of the group with the highest homocysteine readings had a 3.4 fold increase in heart attack risk. Also cited was a 1995 Tufts University study of over 1,000 elderly men and women, which showed that high homocysteine levels raised odds for significant carotid artery obstruction. (New England Journal of Medicine, vol.332, pp.286-291). A carotid blockage is considered a warning sign of above-average risk for both stroke and coronary artery disease.

The Washington researchers concluded that a 5 u.mol/L increment in homocysteine level raises coronary artery disease risk as much as a 20 mg/dL rise in cholesterol. No one has yet proven how homocysteine causes atherosclerosis, but scientists suspect it may do its harm during one or more steps in the process that transforms a healthy blood vessel into the site of a heart attack. The arteries of animals injected with homocysteine showed changes that may lay the groundwork for the buildup of atherosclerotic plaques. There is also evidence suggesting that homocysteine stimulates proliferation of blood vessel cells that help form plaques and that it encourages clotting.

A large multi-center European trial, published in the June 11, 1997, issue of the Journal of the American Medical Association, found that among men and women younger than age 60, the overall risk of coronary and other vascular disease was 2.2 times higher in those with plasma total homocysteine levels in the top fifth of the normal range compared with those in the bottom four-fifths. This risk was independent of other risk factors, but was notably higher in smokers and persons with high blood pressure.

A Norwegian study, published in the July 24, 1997, issue of the New England Journal of Medicine, found that among 587 patients with coronary heart disease, the risk of death after four to five years was proportional to plasma total homocysteine levels. The risk rose from 3.8 percent in those with the lowest levels (below 9 µmol per liter) to 24.7 percent with the highest levels (greater than 15 µmol per liter).

Other evidence suggests that homocysteine may have an effect on atherosclerosis by damaging the inner lining of arteries, and promoting blood clots. However, a direct causal link hasn't been established.

New data comes from an analysis of 1,041 men and women ages 67 to 96 involved in the Framingham Heart Study. That study, now nearly 50 years old, is an ongoing analysis of cardiovascular risk factors in the community of Framingham, Mass. Researchers from several institutions including Tufts University and Boston University conducted ultrasound tests on the carotid arteries of the study participants to determine the degree of vessel narrowing. This is considered a good indication of the degree of arteriosclerosis, or artery thickening. After obtaining the ultrasound measurements, the researchers then used statistical methods to determine the potential correlation between significant vessel narrowing (at least 25 percent of the inner diameter of the neck arteries) and blood levels of homocysteine and related vitamins. The analysis revealed that participants with the highest homocysteine levels were twice as likely to have arteriosclerosis as those with the lowest levels of homocysteine. The odds decreased as plasma levels of folate and, to a lesser extent, vitamin B6 increased. In addition, the risk of significant narrowing increased at homocysteine levels previously considered to be normal.

Given these findings, the next step was to establish a direct association between increased intake of folate and vitamins B6 and B12 and reduction in risk of CVD morbidity and mortality. According to homocysteine pioneer McCully, this relation was demonstrated in a substudy of the Nurses' Health Study reported in 1998 by Rimm et al. During 14 years' follow-up of 80,082 women (among whom 658 incident cases of nonfatal MI and 281 cases of fatal coronary heart disease [CHD] occurred), the researchers determined intake of folate and vitamin B6 from detailed food frequency questionnaires and examined extreme quintiles.

Among women in the highest quintile of folate intake, the age-adjusted relative risk of a CHD event was 0.55, compared with those in the lowest quintile; the risk was 0.49 in the highest quintile for vitamin B6 intake, compared with the lowest. Among regular users of multiple vitamins, the age-adjusted relative risk of CHD incidence was 0.69, compared with nonusers. Each 100µg/d increase in folate, the researchers calculated, meant a 5.8% reduction in CHD risk.

Some of the studies quoted above which have shown a link between CVD and homocysteine levels have also shown a negative relationship between levels of folate and CVD.

Moreover, Canadian researchers have demonstrated that therapy with these nutrients can reverse the progression of carotid atherosclerosis -- in patients who had failed to respond to treatment for other CVD risk factors. At the Robarts Research Institute in London, Ontario, Hackam et al compared rates of progression of carotid plaque (as measured by ultrasonography; see Figures 1 and 2) in 101 patients with vascular disease, before and after a daily regimen of 2.5 mg of folic acid, 25 mg vitamin B6, and 250 µg of vitamin B12. Irrespective of their baseline homocysteine levels (ie, above or below 14 µmol/L), patients experienced "a rapid rate of [carotid plaque] progression before treatment and a slight regression after treatment." Plaque regression was most significant in patients older than 65 years. In patients with vascular disease, the researchers suggest, "the plasma homocyst(e)ine level to treat may be <9 µmol/L."

Canadian research has linked low blood levels of the B vitamin, folic acid, to increased odds for fatal coronary heart disease. A study of more than 5,000 people found that those in the quarter of the group with the lowest folate levels were 69 percent more likely to die of a coronary problem than those in the quartile with the greatest stores of the vitamin. The study underscores the need for clinical trials to determine whether increasing folic acid intake can prevent heart disease, says Meir Stampfer, M.D., Dr. P.H. of the Harvard School of Public Health, in an editorial accompanying the report. The study was published in the June 26, 1996 issue of JAMA (Vol 275, pp. 1893 -1896).

The research, led by Howard Morrison, Ph.D., of the Cancer Bureau in Ottawa, Ontario, involved men and women, aged 35 to 79, who reported no history of coronary heart disease. The subjects' blood levels of folic acid were measured from 1970 to 1972. The group was then followed through 1985. During the 15-year period, the researchers identified 165 people who died of coronary heart disease. The data show that as folic acid levels dropped, risk of death rose in a stepwise fashion. Folate appeared most protective in women and in people under age 65. An interesting finding in all age groups was that risk increased even at folic acid levels that are presently considered normal. This observation suggests the need to redefine the norms, note the authors.

New studies suggesting a protective role for folic acid continue to appear. In 1996, Canadian investigators reported that among more than 5,000 men and women who participated in a national nutrition survey, those in the quarter of the group with the lowest folic acid levels were 69 percent more likely to die of a coronary problem than those in the quartile with the greatest stores of the vitamin (JAMA, vol.275, pp.1893-95). There is even evidence that high risk patients have the most to gain.

In 1995, a University of Utah study compared over 160 men and women who had evidence of early familial coronary artery disease with a comparable group who did not have the disease. The patients, who had already had either a heart attack, bypass surgery or balloon angioplasty, showed "a considerably greater sensitivity" to the blood's concentration of the B vitamin, according to the researchers (Arteriosclerosis, Thrombosis and Vascular Biology).

How does increased levels of homocysteine in the blood predispose to atheroma formation? Atherosclerosis begins with low-density lipoprotein (LDL) filtering into the lining of the artery and becoming entrapped and oxidized in the intima. There, uptake of oxidized LDL by macrophages occurs and foam cells are produced. As foam cells accumulate in the intima, they combine with intercellular lipid to form fatty streaks, which are gradually converted into fibrous plaques in a process similar to that of scar formation. Ultimately, fibrous plaques can be transformed into complicated atherosclerotic lesions, which underlie most adverse clinical events.

At elevated levels, homocysteine can block production of nitric oxide in the cells of the blood vessel walls, making the vessels less pliable and allowing plaque to build up. Several different mechanisms have been proposed to explain the apparent association between homocysteine and atherosclerotic vascular disease.

Table 1. Possible Mechanisms Of Homocysteine-Mediated Atherogenesis

1. Endothelial dysfunction
2. Endothelial cell injury
3. Promotion of smooth muscle cell proliferation
4. Enhanced platelet aggregation
5. Increased binding of lipoprotein(a) to fibrin
6. Generation of free radical species
7. Stimulation of low-density lipoprotein oxidation
8. Procoagulant effects

Homocysteine is a naturally occurring amino acid, derived from methionine and produced in small amounts by the human body. It is metabolized by transsulfuration (which depends on vitamin B6) and remethylation (which relies on folate [folic acid] and vitamin B12). According to a 1999 science advisory from the American Heart Association (AHA) Nutrition Committee, plasma concentrations of homocysteine between 5 and 15 µmol/L are considered normal. Elevated homocysteine levels are referred to as hyperhomocysteinemia -- moderate, between 16 and 30 µmol/L; intermediate, between 31 and 100 µmol/L; and severe, higher than 100 µmol/L. The authors of the 1999 AHA science advisory consider a basal homocysteine level below 10 µmol/L "a reasonable therapeutic goal for subjects at increased risk." For significant reduction of serum homocysteine levels (and, presumably, "favorable impact on CHD rates"),

Homocysteine, an intermediate in protein metabolism, is involved in conversion of the amino acid methionine to cysteine or in remethylation to form methionine (Figure 1).

Mechanisms Of Homocysteine-Mediated Atherogenesis

FIGURE 1. Homocysteine metabolism. An intermediate in protein metabolism, homocysteine is involved in conversion of the amino acid methionine to cysteine or in remethylation to form methionine (SAM=S-adenosyl methionine; SAH=S-adenosyl homocysteine; MS=methionine synthase; MTHFR=methylene tetrahydrofolate reductase; CBS=cystathionine b-synthase).

Increases in homocysteine concentrations are often the result of decreased activity of key enzymes involved in either of these metabolic pathways.

The most common inherited form of hyperhomocysteinemia results from an alteration in the gene encoding the enzyme methylene tetrahydrofolate reductase. A mutation in the methylene tetrahydrofolate reductase gene leading to mild to moderate hyperhomocysteinemia has been found in 15 percent of patients with premature cerebrovascular disease.

Less often, the cause of hyperhomocysteinemia is heterozygous cystathionine b-synthase deficiency. Homocystinuria is a rare, but severe homozygous form of cystathionine b-synthase deficiency in which total homocysteine concentrations generally exceed 100 µmol per L but can reach 500 µmol per L if the disorder is untreated. Individuals with this inherited disorder are known to have very premature coronary artery disease.

Hyperhomocysteinemia can be acquired as the result of dietary deficiencies of folate, vitamin B12 and/or vitamin B6. These nutrients are necessary cofactors for the optimal function of methylene tetrahydrofolate reductase and cystathionine b-synthase. Deficiencies in the absorption or transport of these vitamins can also cause hyperhomocysteinemia.

Certain drugs, especially vitamin antagonists such as methotrexate and anticonvulsants, can cause hyperhomocysteinemia. Notable homocysteine elevations can also occur in illnesses such as chronic kidney disease or hypothyroidism.

Common causes of hyperhomocysteinemia are listed in Table 1.

Causes of Hyperhomocysteinemia

  • Acquired causes
  • Vitamin deficiencies
  • Folic acid
  • Vitamin B
  • Vitamin B12
  • Chronic diseases
  • Chronic renal failure
  • Hypothyroidism
  • Psoriasis
  • Malignancies (including acute lymphoblastic leukemia)
  • Medications
  • Anticonvulsants
  • Methotrexate
  • Nitrous oxide
  • Inherited causes
  • Cystathionine b-synthase deficiency
  • Methylene tetrahydrofolate reductase deficiency or defect
  • Methionine synthase defect
  • Vitamin B12 transport defect
  • Vitamin B12 coenzyme synthesis defect

There is adequate evidence that folic acid supplementation can decrease the elevated homocysteine levels in the blood. Folic acid is thought to protect against heart disease because it breaks down homocysteine and allows it to be cleared from the blood stream. Rimm advocates folate intake of at least 400 µg/d. To this, the science advisory authors add, patients should be encouraged to take 2 mg/d of vitamin B6 and 6 µg/d of vitamin B12 in vitamin-fortified foods and/or vitamin supplements. Older people often malabsorb food-bound B12, and thus it is important to assess for vitamin B12 deficiency in elderly patients. Treatment with folic acid can obscure unsuspected vitamin B12 deficiency and allow progression of neurologic complications.

How does folic acid and Vitamins B6 and B12 decrease blood homocysteine levels? They help in breaking down homocysteine and thus allowing it to be cleared from the bloodstream.


The optimal total dose of folic acid appears to be within the range of 650 to 1,000 µg per day. In the United States, the average folic acid intake is approximately 200 µg per day. Although many recommend getting the required folic acid from natural or even fortified foods, this has been shown to increase the folic acid intake to less than 400 ug per day. It is currently recommended that people increase their intake of folate-rich foods and that they consider adding a daily multivitamin containing 400 µg of folic acid. This recommendation is especially important for pregnant women and for persons considered to be at high risk for cardiovascular disease.

In fact, many authorities feel that high-risk patients with homocysteine levels above 12 µmol per L should take a daily multivitamin containing 400 µg of folic acid plus an additional 800 µg of folic acid per day. The homocysteine concentration should be rechecked in eight weeks to assess the response to therapy. If the homocysteine level returns to normal, the supplemental folic acid may be discontinued. In such cases, the multivitamin should be continued, and the homocysteine concentration should be rechecked in approximately eight to 12 weeks to ensure that the level is stable. If the homocysteine level is not less than 12 µmol per L after eight weeks of supplementation, the folic acid supplementation can be increased to 2 mg per day for an additional eight weeks, with repeat homocysteine testing performed at the end of treatment. Persistently elevated homocysteine levels warrant a careful assessment of patient compliance or testing for other possible causes of hyperhomocysteinemia.

The authors of one study concluded that with an increased intake of approximately 200 µg of folic acid per day, homocysteine levels could be reduced by 4 µmol per L in an average population. Regardless of the cause of hyperhomocysteinemia, most patients should derive some benefit from folic acid supplementation via the conversion of homocysteine back to methionine. Homocysteine levels usually decrease after a few weeks of therapy and normalize within six to eight weeks. At the time of the initial evaluation, a serum B12 level should be obtained to ensure that intake of this vitamin is adequate before folic acid supplementation is initiated.

Folic acid supplements are safe, especially when the dosage is less than 5 mg per day. Current evidence indicates no significant concerns with folic acid or other vitamin B supplementation, except for the possible masking of vitamin B12 deficiency. The neurologic defects associated with vitamin B12 deficiency persist with folic acid supplementation. Vitamin B12 deficiency can still be detected clinically by the history and physical examination, along with serum methylmalonic acid or vitamin B12 measurement, factors that are not altered by folic acid supplementation. Hypersensitivity reactions to folic acid supplements are quite rare.