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Homocysteine:- a naturally occurring amino acid
Homocysteine is a naturally occurring amino acid produced as part of the body’s
methylation process. The level of homocysteine in the plasma is increasingly being
recognised as a risk factor for disease and seen as a predictor of potential health problems
such as cardiovascular disease and Alzheimer’s.
The complex metabolism of homocysteine within the body is highly dependent on vitamin
derived cofactors, and deficiencies in vitamin B12, folic acid and vitamin B6 are associated
with raised homocysteine levels. Other factors thought to raise levels are poor diet, poor
lifestyle - especially smoking and high coffee and alcohol intake, some prescription drugs
(such as proton pump inhibitors), diabetes, rheumatoid arthritis and poor thyroid function.
There is no consensus about the upper reference limits for plasma homocysteine
concentrations although the ‘normal’ range for healthy individuals is considered to be
between 5 and 15 µmol/L. However levels as low as 6.3 µmol/L are thought to confer an
increased risk and each 5 µmol/L can increase the risk of coronary heart disease events by
approximately 20%.
The good news is that homocysteine levels can be tested and high homocysteine levels
can, in many cases, be normalised through diet and vitamin supplementation. The most
important nutrients that help lower homocysteine levels are folate, the vitamins B12, B6 and
B2, zinc and trimethylglycine (TMG).
This article discusses the causes and impacts of high homocysteine levels and the
importance of homocysteine measurement, whilst highlighting some of the limitations
associated with sample handling and homocysteine testing, and vitamin supplementation to
normalise homocysteine levels.
Background
The importance of homocysteine as a risk factor is becoming much more familiar to us. A
constantly increasing number of studies have been published that show homocysteine to be
a predictor of potential health problems. It is clear now that raised plasma homocysteine
concentrations both predict and precede the development of cardiovascular disease
including stroke. A study published in the British Medical Journal showed clearly
homocysteine level in blood plasma predicts risk of death from cardiovascular disease in
older people even better than any conventional measure of risk including cholesterol, blood
pressure or smoking.
Raised levels of homocysteine are also linked to Alzheimer’s, dementia, declining memory,
poor concentration and judgment and lowered mood. Women with high homocysteine levels
find it harder to conceive and are at risk from repeated early miscarriage. High
homocysteine has also been linked to migraines, and those with conditions such as
diabetes and osteoporosis are at increased risk of raised homocysteine levels.
Homocysteine has therefore been shown to play a crucial role as a key marker for disease
development determining longevity and health throughout a person’s life.
Why is homocysteine harmful?
Homocysteine is a naturally occurring amino acid produced as part of the methylation
process. It has the formula C4H9NO2S and is a derivative of protein that is found in blood
plasma when body chemistry is out of balance. It is a homologue of the amino acid
cysteine, differing by an additional methylene (-CH2-) group. Homocysteine is not obtained
from the diet, instead, it is biosynthesized from methionine via a multi-step process that
probably occurs in every cell of the body (Figure 1).
Methionine Metabolism
Methionine is an amino acid, ingested as a component of food protein, and is found
primarily in meats, eggs, dairy products, fish, chicken, seeds, nuts and some vegetables.
Methionine is activated to S-adenosylmethionine (SAM) by the enzyme methionine
adenosyltransferase. Circulating levels of homocysteine are usually low due to its rapid
metabolism via one of two pathways: a cobalamin (vitamin B12) and folate dependent remethylation pathway that regenerates methionine, or a pyridoxal 5’ phosphate (PLP, vitamin
B6) dependent trans-sulphuration pathway that converts homocysteine into cysteine.
The complex metabolism of homocysteine within the body is highly dependent on vitamin
derived cofactors, and deficiencies in vitamin B12, folic acid and vitamin B6 are associated
with hyperhomocysteinaemia. The reason homocysteine accumulates in the body causing
cell damage and the onset of major disease, is because the biochemical transformation
process is not working properly, usually due to lack of these needed vitamins. If these
pathways are lacking the required vitamins and minerals, dangerous homocysteine levels
and potential ill health can result.
The proposed mechanisms by which hyperhomocysteinaemia can cause harm such as
vascular damage, cognitive impairment, neurological complications, congenital defects and
pregnancy complications are common to all these conditions. A detailed review of these
mechanisms is outside the scope of this article, however, raised homocysteine is
associated with damage to the arteriesand one mechanism by which homocysteine is
thought to cause this damage is by interfering with the way cells use oxygen, resulting in a
build-up of damaging free radicals. Oxidation triggers many diseases including heart
disease, strokes, cancers and autoimmune diseases.
Reactive chemical forms such as free radicals can oxidize low-density lipoproteins
producing oxy-cholesterols and oxidized fats and proteins within developing arterial
plaques. This oxidation injury, along with changes in nitric oxide metabolism, an important
regulator and mediator of numerous processes in the nervous, immune and cardiovascular
systems, and decreased methylation, appears to contribute to the damage caused. Indeed,
methylation defects and impaired DNA repair caused by disturbed folate metabolism are
suggested to contribute to carcinogenesis.
Homocysteine also stimulates the growth of smooth muscle cells, causing deposition of
extracellular matrix and collagen, which causes a thickening and hardening of artery walls.
Overall though, the exact mechanisms involved in the increased risk of ill health with raised
homocysteine still remain a mystery in many respects, and more studies are needed to
elucidate the exact associations.
What causes raised homocysteine levels?
Many factors are thought to raise levels of homocysteine; among them are poor diet, poor
lifestyle especially smoking and high coffee and alcohol intake, some prescription drugs,
diabetes, rheumatoid arthritis and poor thyroid function. Raised levels are also associated
with chronic inflammatory diseases in general, and some intestinal disorders such as
coeliac and Crohn’s diseases. Levels increase with age and higher levels are more
common in men than women. Levels of homocysteine can increase with oestrogen
deficiency and with some long term medications, including corticosteroids. Strict
vegetarians and vegans may also be at risk and people who suffer from stress. As with
cholesterol, family history and genetic make-up can play a part in causing raised levels as
can obesity and lack of exercise. Even people with an active, healthy lifestyle may still be at
risk, if there is a family history of high levels of homocysteine or disease.
A rapidly increasing number of variations of the genes that regulate the enzymes that are
involved in methionine metabolism have been identified. Reduction in the activity of genes
such as the one that regulates the enzyme methyl-enetetrahydrofolate reductase (MTHFR)
increases mean homocysteine levels. This gene is present in its homozygous form in about
10% of most European populations but the frequency varies widely geographically and
between different ethnic populations.
Testing for homocysteine
The measurement of homocysteine, or more correctly the measurement of total
homocysteine that includes the sum of the concentrations of free and bound homocysteine
in the blood plasma, is technologically difficult. After blood sampling the blood cells produce
and release homocysteine resulting in an increase in measured levels which equates to
about 10% per hour at room temperature. Thus it is vital to collect blood plasma (using
EDTA or heparin to prevent coagulation) and to centrifuge the sample, to remove the blood
cells from the plasma, within 30 minutes of collection; even the most efficient transport from
clinic or GP practice to laboratory centrifuge rarely renders this feasible. Collection of blood
serum is not appropriate due to the relatively long clotting time required. Once separated
from the blood cells the homocysteine is stable in plasma for at least 4 days at room
temperature, and much longer refrigerated or frozen.
Some manufacturers now provide blood collection vacutainers, which contain a stabiliser
solution that can help delay the urgent need for centrifugation. However, the most effective
device on the market for the collection and immediate separation of blood plasma for
homocysteine measurement is the unique and patented plasma separation system
manufactured by one laboratory in the UK; the only device of its kind on the market. This
device enables immediate separation of blood plasma, which means that a finger prick
blood sample can be collected using a lancet in the comfort of the clinic or even at home.
The collection device is easy to use and immediately separates the blood cells and collects
the plasma in an absorbent pad that can be posted back to the laboratory for testing. There
is also a visible in-built indicator that defines the volume collected so that the exact amount
of blood required (just a couple of drops) is collected. The laboratory claims that when using
the plasma separator device for collection of samples there is no degradation in the
homocysteine for up to 10 days, even when stored at 37ºC.
There are several other important factors to consider when blood sampling. As methionine,
ingested as protein in the diet, is the only source of homocysteine, it would be expected that
food intake would cause increased concentrations of homocysteine in plasma. Indeed a
protein rich meal can increase the total homocysteine levels significantly and so it is
generally recommended that the person taking the test should be overnight fasted at the
time of blood sampling.
Urine homocysteine measurement is not useful as renal excretion does not seem to be an
important route of elimination. Only about 1% of the homocysteine filtered by the glomeruli
is normally found in the urine. The rest is reabsorbed and metabolized. In saying that, some
GPs do offer urine homocysteine measurement, as it is easy to collect, however, results
should be interpreted with caution as they are only likely to pick up extreme
hyperhomocysteineaemia.
Test methods
The increased need in clinical chemistry laboratories for methods of homocysteine
determination, in correlation with cardiovascular diseases and nutritional deficient status,
has led to the development of different analytical methods for measurement.
Methods include amino acid analysis, high performance liquid chromatography (HPLC),
capillary electrophoresis, gas chromatography-mass spectrometry (GC-MS) and
immunoassay; HPLC being the gold standard reference method. Different total
homocysteine measurement methods do usually give comparable results, however, there is
variation among methods and between laboratories and so caution should be used in
comparing values obtained from different laboratories. In saying that, studies have shown
that single homocysteine measurements can classify persons with respect to their average
plasma homocysteine level quite well. However, due to the inter-method and interlaboratory variability it is important that any monitoring of homocysteine concentrations over
time, and with treatment, should be carried out using the same method and the same
laboratory. Choosing laboratory testing services is a critical process.
"Normal" reference ranges
There is no consensus about the upper reference limits for plasma homocysteine
concentrations. Among apparently healthy individuals “normal” concentrations commonly
range from 5 to 15µmol/L. However, studies on targeted segments of the population have
shown that the upper limit of 15µmol/L is far too high in well-nourished populations without
obvious vitamin deficiency. It is clear now that each increase of 5µmol/L in homocysteine
level increases the risk of coronary heart disease events by approximately 20%,
independently of traditional coronary heart disease risk factors. In addition it is well
documented that risk for coronary artery disease is represented by a continuum of
homocysteine concentrations with a substantial risk occurring between 10 and 15µmol/L.
Some report that any homocysteine measurement over 6.3µmol/L represents an increased
risk.
There is little doubt that homocysteine measurement will become a routine screening tool
for risk assessment in the future, but that current reference limits will change from being
based on values from a presumed normal healthy population towards a baseline where
‘normal’ is the optimum level. Overall, the careful monitoring of homocysteine levels with
treatment is recommended.
Treatment
The good news is that high homocysteine levels can, in many cases, be normalised, and
the advice regarding diet and vitamin supplementation have been shown to be very
effective in reducing plasma homocysteine levels. It is clear though that vitamin
supplementation can normalize homocysteine levels even when serum vitamin levels are
within the normal range, or even in the high range. Metabolic, environmental and genetic
factors make it virtually impossible to determine individual nutritional requirements without
first carrying out a homocysteine test; the test result can then define the diet and
supplementation regime required. The most important nutrients that help lower
homocysteine levels are folate, the vitamins B12, B6 and B2, zinc and trimethylglycine
(TMG).
Conclusion
This article has provided an overview of the importance of homocysteine measurement,
whilst highlighting some of the limitations associated with sample handling and
homocysteine testing. There will always be people who are at high risk of raised
homocysteine, but the only way to find out definitively is to perform a validated
homocysteine test, and then monitor levels during treatment.
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