Chemical pathology of homocysteine. I. Atherogenesis

17 Pages·2015·1.34 MB·English

ANNALS OF CLINICAL AND LABORATORY SCIENCE, Vol. 23, No. 6
Copyright © 1993, Institute for Clinical Science, Inc.
Chemical Pathology of Homocysteine
I. Atherogenesis*
KILMER S. McCULLY, M.D.
Department of Veterans Affairs Medical Center,
Providence, RI 02908
ABSTRACT
The atherogenic properties of homocysteine were discovered by obser­
vation of arteriosclerosis in children with homocystinuria caused by inher­
ited deficiency of three different enzymes. Hyperhomocysteinemia is gen­
erally recognized as an independent risk factor for coronary, cerebral, and
peripheral atherosclerosis. Hyperhomocysteinemia is caused by heterozy­
gosity for homocystinuria, micronutrient deficiency from dietary imbal­
ance, toxins, drugs, hormones, and other factors, explaining many key
observations concerning the epidemiology of atherosclerosis. The etio­
logical factors for atherosclerosis are believed to increase conversion of
methionine to homocysteine thiolactone, the reactive cyclic internal lac­
tone of homocysteine. The free amino groups of low density lipoprotein
(LDL) are thiolated by homocysteine thiolactone, causing aggregation and
increased uptake of LDL by macrophages, explaining lipid deposition in
atheromas. Homocysteine thiolactone, released from homocysteinylated
LDL within vascular wall, promotes intimal injury, oxidation of cholesterol
and unsaturated lipids, platelet aggregation, thrombogenic factors, myoin-
timal hyperplasia, deposition of sulfated glycosaminoglycans, fibrosis and
calcification of atherosclerotic plaques.
Introduction trol of growth by several newly discov­
ered amino acids containing sulfur. The
The degenerative diseases associated purpose of this review to elucidate,
with aging, particularly arteriosclerosis within the limitations of present knowl­
and cancer, present a challenge to under­ edge, the important role of the metabo­
standing of the correlations between fun­ lism of these sulfur amino acids in athero­
damental aspects of their pathogenesis. genesis, carcinogenesis, cellular and
The key to this understanding may have tissue function, degenerative diseases
arisen from an obscure line of investiga­ and aging.
tion earlier in the twentieth century that This review of the chemical pathology
was concerned with the nutritional con­ of homocysteine is divided into three
sections for publication in successive
issues of Annals of Clinical and Labora­
* Address reprint requests to: Kilmer S. McCully,
tory Science. Section I, Atherogenesis,
M.D., Pathology and Laboratory Medicine Service,
VA Medical Center, Providence RI 02908-4799. describes the discovery of methionine,
477
0091-7370/93/1100-0477 $02.00 © Institute for Clinical Science, Inc.
478 McCULLY
homocysteine, and homocysteine thiolac- that experimental methods for proving or
tone, the discovery of the homocysteine modifying these proposals can be devel­
theory of arteriosclerosis, and current oped by investigators with an interest in
knowledge concerning the role of homo- this challenging area of clinical and labo­
cysteine in the pathophysiology of ratory science.
atherogenesis. Section II, Carcinogene­
sis and Homocysteine Thiolactone Discovery of Methionine,
Metabolism, describes altered methio­ Homocysteine and
nine and homocysteine metabolism in Homocysteine Thiolactone
carcinogenesis, the synthesis of antineo­
plastic homocysteine thiolactone com­ Methionine, one of the eight essential
pounds, the effect of homocysteine thio­ amino acids of mammalian nutrition, was
lactone on normal tissues, the hypothetical discovered in broths of bacterial cultures
function of thioretinaco and thioretina- by Mueller in 1922.60 The essential
mide in oxygen metabolism, adenosine nature of methionine in nutrition, its role
triphosphate synthesis and phosphoade- in methyl group transfer, and its conver­
nosine phosphosulfate synthesis, and the sion to cystathionine, cysteine, taurine, and
importance of homocysteine thiolactone sulfate were discovered by DuVigneaud
metabolism in the growth of animals. and coworkers in the succeeding two
Section III, Cellular Function and decades.17 These studies clearly demon­
Aging, describes current knowledge of strated that, while methionine is essen­
the role of homocysteine metabolism in tial for supporting the growth of animals,
cellular respiration, cell division, growth the requirement for cysteine in support­
and function of epithelia and connective ing growth can be supplied by methio­
tissues, lipid transport and metabolism, nine, through the intermediate formation
adenosyl methionine synthesis, the aging of cystationine.
process, and degenerative diseases asso­ In the course of their work, a new sul­
ciated with aging. fur amino acid, homocysteine, was dis­
While many of the observations and covered and shown to support the growth
interpretations described in this review of animals, provided that nutrients con­
are directly supported by the evidence taining methyl groups, such as choline or
cited in the references, the reader is cau­ betaine, were supplied.5 At about the
tioned that an attempt has also been same time, the anhydride of homocys­
made to explain many of the alterations of teine, the cyclic internal lactone, homo­
homocysteine metabolism which occur cysteine thiolactone, was synthesized as
in the degenerative diseases of aging. the hydriodide salt by reaction of hydri-
This attempt has necessitated the postu­ odic acid with methionine.2
lation of a number of hypotheses, specu­
lations, and conjectures regarding the Chemical Properties of
schemes presented in Section II, Figures Homocysteine Thiolactone
4 to 11, and in Section III, Figures 2, 4,
and 5. In many respects, extensive fur­ Because of the reactive nature of its
ther experimental investigation is five membered lactone ring, homocys­
needed to establish the validity of these teine thiolactone possesses several
schemes with certainty. In defense of this important chemical properties that are
approach, many of these proposals are not shared by cysteine. Since it has one
described in sufficient biochemical, meta­ fewer methylene carbon atom than
bolic, and pathophysiological detail, so homocysteine, cysteine cannot form a sta­
HOMOCYSTEINE AND ATHEROGENESIS 479
ble internal cyclic lactone. Homocys­ ing of the second thiolactone ring to form
teine thiolactone results from the action homocysteine diketopiperazine. This
of strong acid on homocysteine, forming a substance contains two free sulfhydryl
stable, ionized, water soluble salt.73 groups that are oxidized by atmospheric
Homocysteine thiolactone is resistant to oxygen to form the insoluble homocys­
oxidation by perchloric acid, forming a teine diketopiperazine disulfide poly­
stable perchlorate salt that is soluble in mer. If homocysteine thiolactone hydro­
chloroform-methanol or ethanol.83 The chloride is hydrolyzed in the presence of
ring structure of the perchlorate salt of dichloromethane, the free base form of
homocysteine thiolactone is identical to homocysteine thiolactone is extracted
that of the hydrochloride salt, as deter­ into the dichloromethane layer and iso­
mined by X-ray diffraction crystallo- lated in pure form after drying and evap­
graphic analysis.48 In contrast, however, orating the solvent.52 The resulting color­
the disulfide dimer of homocysteine, less, viscous oil is freely soluble in both
homocystine, is susceptible to oxidation water and organic solvents. The oil has a
by hydrogen peroxide, forming homolan- pungent, sulfurous, fishy, amine odor.
thionine sulfone, homolanthionine sulf­ The oil spontaneously forms an insoluble
oxide, homocysteic acid, and sulfate.12 white solid, when allowed to stand for
The unusual chemical reactivity of one hour in the presence of air at room
homocysteine thiolactone is demon­ temperature. The nuclear magnetic reso­
strated by analysis of the products of the nance (NMR) spectrum of the oil is sim­
reaction of weak alkali with its hydro­ ilar to that of homocysteine thiolactone
chloride salt.18 Under these conditions, salts. Oxidation of this oil in aqueous solu­
oxidation fails to yield homocystine, as tion by aeration results in formation of the
expected, but instead a high melting, high melting amorphous diketopiperazine
amorphous polymeric substance with solu­ polymer. These reactions and hydrolysis
bility properties similar to those of kera­ conditions are illustrated in Figure 1.
tin is obtained. This substance was found Understanding of the scope of reactiv­
to be a polymer formed by oxidation of ity of homocysteine thiolactone was
the sulfhydryl groups of homocysteine extended in 1956, when formation of
diketopiperazine. In contrast, hydrolysis peptide bonds was shown to occur by
of homocysteine thiolactone hydrochlo­ ammoniolysis of homocysteine thiolac­
ride with strong alkali results in the tone by amines and by amino acids.3 This
quantitative formation of homocysteine, reaction introduces thiol groups into pro­
which is oxidized directly to homocys­ teins by homocysteinylation of free
tine without the intermediate formation amino groups, a process called thiola-
of homocysteine diketopiperazine poly­ tion.4 The reaction of immunoglobulin G
mer. These findings are summarized in with homocysteine thiolactone intro­
Figure 1. duces peptide bound homocysteinyl
Dimerization of homocysteine thiolac­ thiol groups, which form mercaptides by
tone in the presence of weak alkali to reaction with methyl mercury.34 Homo-
form homocysteine diketopiperazine is cysteinylated immunoglobulin G
the result of reaction of one molecule of labelled with mercury in this way can be
homocysteine thiolactone with the free visualized in tissues by electron micros­
amino group of another molecule, form­ copy.35 The same reaction was later
ing a peptide bond. The resulting homo- employed with low density lipoprotein
cyteinyl homocysteine thiolactone forms (LDL), and the affinity of thiolated LDL
a second internal peptide bond by open­ for membrane receptors of leukemic lym-
S
H2 d.
2C ate
H ic
H2CHNH2COOH + H20 H2CHNH2COOH cystine + NaCI + H2C03 NH•SCH2CH2 HN. >C homocysteine diketopiperazine polymer ysteine thiolactone are ind
C C o c
H2 H2 om H mo
> SC SC h 2CH2S H2SH of ho
OH + NaCI 101 NHCOCHNH2CH steinyl steine thiolactone NH22ch HN. >CH2C homocysteine diketopiperazine merization reactions
O cy cy h y
HSCH2CH2CHNH2C homocysteine NaHCO,-> S' homo homo hsc oxidation, and pol
s,
si
y
L cr ol
H + r
O 3 d
Na NH■» S homocysteine thiolactone hydrochloride NaOHCHpCI o homocysteine thiolactone free base cal synthesis, hy
i
CI m
H he
C
1.
OH e
O r
H2C Figu
N e
H n
C ei
CH2 cyst
2 o
H m
C o
S h
H
HOMOCYSTEINE AND ATHEROGENESIS 481
phocytes was found to be preserved.94 arises by oxidation of choline, and
These studies show that homocysteine betaine methyl transferase, a liver
thiolactone introduces peptide bound enzyme, catalyzes remethylation of
homocysteinyl groups by reaction with homocysteine to methionine by transfer
the free amino groups of diverse proteins. of the methyl groups of betaine. Several
important reactions in the metabolism of
Biochemical Functions of Methionine methionine are illustrated in Figure 2.
These nutritional and biochemical
An important derivative of methionine studies of sulfur amino acid metabolism
was discovered by Cantoni in 1952, as demonstrate the vital importance of
the product of the enzymatic reaction of methionine as an essential amino acid in
adenosine triphosphate (ATP) with protein synthesis, in methylation reac­
m ethionine to form S-adenosyl- tions, and in formation of methionine
methionine.6 This optically active sulfo­ from homocysteine and methyl donors by
nium derivative is synthesized in all liv­ transmethylation. Furthermore, methio­
ing cells by adenosyl transferase with the nine is an essential structural component
intermediate formation of tripolyphos­ of proteins, and methionyl transfer ribo­
phate.55 Adenosyl methionine transfers nucleic acid (RNA) functions as the ini­
its methyl group to a wide variety of tiator of protein synthesis by polyribo­
methyl acceptor compounds, with the somes. Methionine is the only essential
formation of adenosyl homocysteine and amino acid that contains sulfur, and cys­
methylated acceptor. Although homocys­ teine is nutritionally non-essential, since
teine is formed within cells by enzymatic is is readily formed from methionine by
hydrolysis of adenosyl homocysteine, its synthesis and catabolism of cystathionine.
concentration is very low because the Altered methionine metabolism is
reaction with adenosine favors formation encountered in malignant cells, in ath­
of adenosyl homocysteine13 and because erosclerosis, and in somatic growth of
homocysteine is actively remethylated to animals. However, the clue to under­
methionine by methyltetrahydrofolate standing of the vital importance of homo-
homocysteine methyl transferase. This cysteine thiolactone in controlling
enzyme requires methyl cobalamin for aspects of these disease processes and
activity.96 In addition, adenosyl methio­ normal cellular functions arose from an
nine, which is an allosteric activator of unusual source, the study of pathological
cystationine synthase and inhibitor of findings associated with inborn errors of
iiiethylenetelrahydrofolate reductase, sulfur amino acid metabolism. Analysis
further prevents accumulation of intra­ of metabolic and pathological abnormali­
cellular homocysteine by coordinate regu­ ties in children with homocystinuria
lation of remethylation and transsulfura­ caused by different inherited enzymatic
tion of homocysteine.79 disorders of homocysteine metabolism
Adenosyl homocysteine is a potent suggested a series of experiments in
inhibitor of the many methyl transfer organic synthesis, biochemistry physi­
enzymes that require adenosyl methio­ ology, cell biology, and experimental
nine. The ability of animals to grow with­ pathology. The results of these studies
out dietary methionine, using homocys­ have helped to elucidate the role of
teine and a methyl donor, is explained by homocysteine thiolactone metabolism in
the active remethylation of homocysteine cellular respiration, in differentiation of
to methionine by methyltetrahydrofolate normal cells and tissues, and in athero­
homocysteine methyl transferase or by sclerosis, cancer and other chronic
betaine methyl transferase. Betaine degenerative diseases.
482 McCULLY
Homocystinuria and Arteriosclerosis In 1969, a different type of homocysti­
nuria, accompanied by cystathioninuria,
In 1933, a mysterious case of carotid hypomethioninemia, and methylmalonic
arteriosclerosis and stroke in childhood aciduria, was found to be caused by
was presented at the Clinical Pathologi­ deranged cobalamin metabolism.39,57
cal Conference of the Massachusetts Pathological study of this case revealed
General Hospital.10 An eight-year-old widespread arteriosclerotic changes of
mentally retarded boy with congenital major arteries, focal infarction and meta­
dislocation of the optic lenses and coxa bolic encephalopathy of frontal and pari­
vara developed headache, vomiting, etal lobes, fatty liver, and atrophic and
obtundation, and left hemiparesis. The metaplastic changes of gastric mucosa.40,47
symptoms progressed to coma and death Since elevated blood homocysteine was
on the third hospital day. Autopsy the only metabolic abnormality found
revealed severe arteriosclerosis of the both in cystathionine synthase deficiency
carotid arteries with acute thrombosis and in methyl transferase deficiency,
and cerebral infarct. Dr. Tracy Mallory, arteriosclerotic lesions in these inborn
the pathologist who presented the errors of metabolism were attributed to
autopsy findings, commented that the the effects of homocysteine derivatives
“very marked intimal thickening [of the on arterial cells and tissues.40 This inter­
carotid artery] could be the result. . . [of pretation was subsequently supported by
a] sclerotic process such as one sees in the finding of similar arteriosclerotic
elderly people.” The disease process was lesions in a child with a third type of
considered to be of congenital origin, homocystinuria caused by deficiency of
involving optic lenses, brain, and carotid methylenetetrahydrofolate reductase.33,59
arteries. Not until 1970 was the correct The enzymatic deficiencies associated
diagnosis proven by demonstration of with the three principle types of homo­
homocystinuria and cystathionine syn­ cystinuria are illustrated in Figure 2.
thase deficiency in the relatives of the
child discussed in 1933.80 Homocysteine Theory
Homocystinuria was discovered in the of Arteriosclerosis
early 1960s.9,21,82 DuVigneaud’s group
had previously identified several patients The association of arteriosclerotic
with homocystinuria at the New York Hos­ lesions with homocysteinemia caused by
pital in the 1950s, but these cases were not different inherited disorders of homocys­
published or investigated more exten­ teine metabolism is the key observation40
sively.11 Deficiency of cystathionine syn­ that led to the development of the homo­
thase is found in liver biopsies56 and in cell cysteine theory of arteriosclerosis.42,54
cultures from the skin of patients with Since discovery of the homocysteine the­
homocystinuria.92 Pathological study of the ory, the atherogenic effect of homocys­
blood vessels in fatal cases reveals fre­ teine accumulation has been established
quent arterial and venous thrombosis of through several lines of investigation.
major branches, as well as fibrous intimal Experimentally induced homocysteine­
plaques of coronary, carotid, renal and mia in animals causes arteriosclerotic
other arteries.8,23,78 Subsequent clinical lesions similar to those observed in chil­
study showed that patients responding to dren with homocystinuria and similar to
pyridoxine therapy survived longer than those observed in individuals without
non-responders and that cardiovascular known enzymatic disorders.42 Study of
disease caused over two-thirds of the 64 patients with cerebral, coronary, and
fatalities observed in 629 patients.58 peripheral atherosclerotic disease has
HOMOCYSTEINE AND ATHEROGENESIS 483
H H H
O O O
+ H+ 2CHNH2CO 2CHNH2CO ystine 2CHNH2CO NH2COOH ysteine e disulfide
ceptor e 2CHCH 2CHCH homoc 2CHCH 2CHCH homoccystein
3chmethyl222222CHNHCHCHS-adenosineHOOCCHNHCHCHS-adenosine+ methylated acacceptorS-adenosylhomocysteineS-adenosylmethionineV+H20 \\ -H200* thioco231222COOHNH + CHBHCX)CCHNHCHCH2SH + adenosinwhomocysteinehomocysteine thiolactoneS S 2222methylH4folate HOOCCHNHCHCHSCHCHNH2COOHS^ reductase cystathionineSH4folate222HOOCCHNHCHCH2OH + HOOCCHNHCH2SH homoserinecysteine F 2. Biochemical pathways of synthesis of homocysteine thiolactone, adenosyl methionine and igurehomocysteine are indicated. Inherited deficiencies of ® cystathionine synthase, @ methyltetrahydro- folate homocysteine methyl transferase, and © methylenetetrahydrofolate reductase cause accumulation of homocystine and homocysteine cysteine disulfide in tissues and body fluids.
C 2 e
O H ^ n
HO ATP CH2CHN hionine 3() methyle
H2 et
C m
S
3
H
C
484 McCULLY
demonstrated elevated blood homocys­ greater than proteins of plant origin. The
teine levels, compared to normal con­ quantity of essential micronutrients
trols.91 Drugs, such as azaribine, metho­ required for metabolizing methionine
trexate, nitrous oxide, anti-convulsants, and preventing homocysteine accumula­
and diuretics, as well as environmental tion, especially folate, pyridoxine, cobala-
toxins such as tobacco smoke and carbon min, riboflavin, and choline or betaine, is
disulfide, which antagonize pyridoxal also correlated with decreased risk of ath­
phosphate, cause elevated blood homo­ erosclerosis. Thus, the methionine to
cysteine and increased atherogensis.46 micronutrient ratio of the diet is high in
Hormonal abnormalities, such as thyroid susceptible populations and low in resis­
deficiency and oral contraceptives, as tant populations.14 Furthermore, the
well as chronic renal failure, also cause blood homocysteine levels of South Afri­
elevated blood homocysteine 46 A ther- can indigenous populations that are resis­
molabile, inherited defect in methylene- tant to atherosclerosis are considerably
tetrahydrofolate reductase is associated lower than the blood homocysteine lev­
with homocysteinemia and coronary els of urban populations that are suscep­
heart disease.31 Approximately one- tible to the disease.87,88
fourth of individuals without known risk
factors who have carotid, peripheral, or Analysis of Blood Homocysteine
coronary atherosclerosis and homocys­
teinemia are heterozygous for cystathio­ Analysis of blood homocysteine in
nine synthase deficiency.91 A prospec­ clinical or population samples is accom­
tive study of 14,916 male physicians plished by amino acid analysis chroma­
showed that hyperhomocysteinemia is tography, high performance liquid chro­
associated with increased risk of myocar­ matography, gas chromatography,
dial infarction.85 Hyperhomocysteinemia enzymatic analysis, or other methods.46,91
is now generally accepted as an indepen­ Free homocysteine is determined as
dent risk factor for coronary heart dis­ homocysteine cysteine disulfide and
ease20 and other forms of occlusive vas­ homocystine in serum or plasma from
cular disease.32 which protein has been precipitated and
The homocysteine theory of arterio­ removed by sulfosalicyclic acid or tri­
sclerosis explains epidemiological fea­ chloroacetic acid. Homocysteine bound
tures of atherosclerosis in susceptible to protein by disulfide bonds is deter­
and in resistant populations.42,46 The mined by chromatography following
consumption of diets with predominantly reduction with dithiothreitol, sodium
animal protein from meat and dairy prod­ borohydride, or tributyl phosphine, and
ucts, together with abundant refined subsequent precipitation of plasma pro­
nutrients, sugar and fats, and highly pro­ teins. A recent method for homocysteine
cessed foods, is associated with a high analysis by high performance liquid
risk of atherosclerosis in susceptible chromatography is designed for screen­
populations. The consumption of diets ing clinical samples in hospital laborato­
with predominantly plant proteins from ries.89 Acid hydrolysis of whole serum
grains, legumes, vegetables and fruits, results in higher levels of homocysteine
together with abundant fresh, unrefined, in serum from patients with coronary
unprocessed foods without added sugar heart disease, compared to disulfide
and fats, is associated with a low risk of bound homocysteine.64,68 If a low vol­
atherosclerosis in resistant populations. ume ratio of serum or plasma to acid is
Proteins of animal origin have a methio­ employed during hydrolysis, dimin­
nine concentration two to three times ished1 or undetectable15 levels of homo­
HOMOCYSTEINE AND ATHEROGENESIS 485
cysteine are observed. The increased free base with LDL causes immediate
recovery of homocysteine by acid hydro­ aggregation and precipitation of thiolated
lysis with a high volume ratio (1:1) of LDL.43 Other lipoprotein fractions are
serum to acid66,68 is attributed to release not precipitated by homocysteine thio­
of peptide bound homocysteine from thio- lactone free base. Thiolation of LDL by
lated proteins.53 Peptide bound homo­ homocysteine thiolactone increases
cysteine originates from reaction of internalization of LDL by membrane
endogenous homocysteine thiolactone receptors, degradation and cholesterol
with amino groups of serum protein.4 accumulation within cultured macro­
Attempts to isolate free homocysteine phages.63 Thiolation of LDL by homo­
thiolactone from lipoproteins by a chloro- cysteine thiolactone is accompanied by
form-methanol extraction method were an increase in density, increase in elec­
unsuccessful, presumably because the trophoretic mobility, and a decrease in
quantities are below the limits of detec­ the number of free amino groups of apoB
tion.44 However, increased quantities of protein.63 Thus, thiolated LDL becomes
homocysteine were demonstrated in low aggregated and susceptible to spontane­
density lipoprotein (LDL), very low den­ ous precipitation because of interaction
sity lipoprotein (VLDL), and high den­ between homocysteinylated amino
sity lipoprotein (HDL) fractions of hyper- groups, probably by disulfide and by
cholesterolemic men, compared to diketopiperazine formation.63,94
normal controls, by the acid hydrolysis Chemical or biological modifications of
method.65 Homocysteine thiolactone lipoproteins, particularly LDL, are
hydrochloride added to serum was believed to increase their atherogenic-
shown to be recovered as homocysteine ity.86 Modification of LDL by acétyla­
cysteine disulfide and homocystine fol­ tion, acetoacetylation, or carbamylation
lowing acid hydrolysis.66 In hypercholes­ increases uptake and cholesterol deposi­
terolemia, the atherogenic index for tion within cultured macrophages.24
homocysteine, LDLHCy/HDLHCy, is 3.5 Incubation of LDL with cultured endo­
times greater than in controls, and the thelial cells, smooth muscle cells or mac­
atherogenic index for cholesterol, rophages results in oxidation of LDL by a
LDLchol/HDLchol, is 2.2 times greater peroxidation process involving superox­
than in controls.65 These findings show ide, hydroxyl radical, and hydroperoxyl
that in hypercholesterolemia, apprecia­ radical, generated by cultured cells in the
ble quantities of homocysteine are car­ presence of transition metal ions.28 Dur­
ried by LDL in the form of homocystei- ing this process, phosphatidyl choline is
nyl groups bound by peptide bonds to hydrolyzed to lysophosphatidyl choline,
the amino groups of apoB. The source of esterified cholesterol is decreased, and
these homocysteinyl groups is endoge­ fatty acids are oxidatively degraded to
nous homocysteine thiolactone, originat­ malondialdehyde, hydroxynonenal, and
ing from methionine in liver.83 other short chain aldehydes.61 Thiolation
of LDL by homocysteine thiolactone
Modification of LDL by does not increase susceptibility to oxida­
Homocysteine Thiolactone tive modification in vitro.63 However,
thiolated LDL is internalized by binding
Reaction of homocysteine thiolactone to LDL receptors and by phagocytosis
with LDL introduces homocysteinyl within macrophages. Release of homo­
groups by thiolation of free amino groups cysteine from internalized, thiolated
of apoB protein.94 Reaction of freshly LDL by hydrolytic degradation within
synthesized homocysteine thiolactone macrophages presumably leads to oxida­
486 McCULLY
tive degradation of lipids through effects homocystinuric patients who respond to
on cellular metabolism.63 Furthermore, pyridoxine therapy.41 These observations
low density lipoprotein of increased show that cellular homocysteine accumu­
density has increased susceptibility to lation is associate with toxic effects and
oxidative modification by macro­ that the cystathionine synthase transsul­
phages,62 presumably because of furation pathway of normal cells reduces
increased thiolation of apoB protein by toxicity by cystathionine formation from
homocysteine thiolactone.63,65 homocysteine. A related observation is
that peroxidation of LDL solutions in
Oxidation, Homocysteine vitro, in the absence of cells, is facilitated
and Atherogenesis by homocysteine, cysteine, mercapto-
ethanol or reduced glutathione, leading
Several oxidized derivatives of choles­ to binding of oxidized LDL to acetyl
terol, especially 25-hydroxy cholesterol LDL receptors and internalization by
and cholestane triol, have been shown to cultured macrophages.70 Thiol com­
be highly atherogenic.71 Purified choles­ pounds are well known facilitators of
terol is readily oxidized by atmospheric lipid peroxidation by oxygen and transi­
oxygen and is non-atherogenic in ani­ tion metal ions. In fact, men with high
mals, when protected against oxidation. stored iron and increased serum ferritin
Hence, the atherogenic effect of choles­ have an increased risk of myocardial
terol in experimental atherogenesis is infarction,76 suggesting increased athero­
attributable to its cholesterol oxide con­ genesis from peroxidation of lipoproteins
tent.29 The high incidence of atheroscle­ and cholesterol by ferric ions and by
rosis in certain immigrant populations in homocysteine released from homocystei-
England is attributed to the atherogenic nylated LDL within artery wall.63
effect of cholesterol oxides in the heated, A considerable body of evidence sug­
clarified butter in their diets.30 Other gests that progression of arteriosclerosis
processed and preserved foods contain­ is associated with and promoted by accel­
ing cholesterol, such as spray-dried egg erated oxidation of cellular and tissue
yolk, bleached butter oils and cheeses, constituents by free radical oxidants.22
and foods that are deep fried in fat, con­ Radical scavanging antioxidant nutrients,
tain angiotoxic cholesterol oxides. Sev­ such as tocopherol, selenium, caro-
eral of these cholesterol oxides have tenoids, retinoids, and ascorbate, may
been demonstrated within atheroscle­ diminish progression of arteriosclerosis
rotic plaques, but the origin of these com­ by counteracting the atherogenic effect of
pounds, whether from dietary sources or free radical oxidants. Cellular enzymes,
from oxidation of lipids within artery including superoxide dismutase, cata-
wall, is less certain.71 Many of these cho­ lase, and glutathione peroxidase, also
lesterol oxides are quite toxic to cultured counteract free radical oxidants by cataly­
cells and carcinogenic in animals, but the sis to less reactive oxygen species. These
molecular basis for this toxicity is incom­ enzymes convert superoxide to hydrogen
pletely understood. peroxide and oxygen, hydrogen peroxide
Homocysteine is toxic to cultured to water and oxygen, and hydrogen per­
endothelial cells, and the toxicity is asso­ oxide to water and oxidized glutathione,
ciated with hydrogen peroxide formation respectively. Both epidemiological and
and facilitated by copper ions.84,95 Homo­ experimental evidence suggest that sup­
cysteine is more toxic to cultured cells pression of free radical oxidation by
deficient in cystathionine synthase than antioxidant nutrients and antioxidative
normal cells, and the toxicity is reversed enzyme activity also suppresses athero-
oo
by pyridoxine in cells cultured from genesis.