N-acetylcysteine
(NAC)
NAC is a thiol-containing amino
acid; it is readily absorbed and is non-toxic. It has many beneficial
effects; perhaps the most surprising is the likely ability of
this molecule to destroy chlamydial elementary bodies. It is
also a powerful antioxidant, and replenishes intracellular glutathione.
It may also be effective in inactivating fungal gliotoxins. These
properties are reviewed below.
Destroying the elementary
bodies: one of the many possible benefits of N-acetylcysteine
Chlamydiae are apparently unique amongst Gram negative organisms
in possessing cysteine-rich proteins on the surface membrane
of the spore-like EB. Cysteine is an amino acid containing a
sulfhydryl ( S-H ) group; a disulphide bond ( S-S
) can form between two cysteine molecules. Such cross-linking
bonds are common within and between the EB surface proteins.
They are thought to preserve the integrity and shape of the EB.
Furthermore, these disulphide bonds are accessible for cleavage.
Why do chlamydial EBs have this surface structure? Structure,
in the realm of the prokaryote, implies function: there is no
room for ornament or unnecessary redundancy. Evolution works
with swift efficiency at this scale. Reduction of these bonds
disrupts the integrity of the EB's surface, and may be the mechanism
by which the EB rapidly opens within the endosome. Rapidity is
important, because the chlamydial body, now truly inside the
host cell, must avoid fusion with the host lysosome or it will
be killed. Raulston and co-workers have shown that the disulphide
bonds between the chlamydial surface proteins must be opened
up just prior to or during attachment to the host cell. [Raulston JE et al.,
Surface accessibility of the 70-kilodalton Chlamydia trachomatis
heat shock protein following reduction of outer membrane protein
disulfide bonds. Infect Immun. 2002; 70(2): 535-43.] The mechanisms of chlamydial attachment
to the host cell membrane; its entry into the cell; its ability
to rapidly open and to subvert host cellular activities; its
ability to evade host-defences and to track actively across the
host cytoplasm taking what it needs from various host organelles
as if it were in a department store are fascinating
to consider. The parasite probably uses an intricate system of
microscopic syringes and needles (the type III secretion system)
to inject subversional proteins into host cytoplasm to induce
endosomal formation. (Other bacteria, easier to study - for instance,
Shigella flexneri - do this by injecting proteins which
cause the host-cell surface to ruffle, overarch and then seal
behind the parasite.) Once the EB is in the endosome, time is
of the essence; it has only a limited amount of preformed proteins
and it must get to work metabolizing as fast as possible.
It is reasonable to think that, were the EB coat to be opened
up before achieving attachment to a nutrient-rich host cell,
the unprotected EB would perish through starvation.
Destruction of EBs may be very important, as there is evidence
that they may accumulate in extracellular spaces awaiting their
chance to enter host cells - this may be analogous to the 'acellular
(or extracellular) load' seen in HIV infections [Chuck Stratton, personal
communication.]
Penicillamine (dimethyl cysteine), which reduces disulphide bonds,
inactivates EBs in vitro and prevents the initiation of
infection in vivo. Amoxycillin, an inexpensive b-lactam
antibiotic, is also effective in inactivating EBs in vivo;
penicillamine is one of its major metabolites. [Chuck Stratton, personal communication.]
N-acetyl cysteine, which, being a thiol antioxidant, is a good
candidate for the reduction of disulphide bonds in EB coating
proteins. It is readily available as a health supplement without
the need for prescription.
The NAC Test
One indirect indicator of chronic infection with this organism
is the N-acetyl cysteine test. This relies on the ability of
NAC to rupture the extracellular Elementary Body by opening up
surface disulphide bonds in the organisms geodesic coat,
as described above. The EB opens and perishes. The release of
naked bacterial components causes local inflammatory symptoms.
Because EBs are more numerous in primary respiratory infections,
the acellular load of EBs is likely to be highest around respiratory
structures. In a positive NAC test the daily administration of
2.4 G of NAC will cause, after a few days, sinusitis-like symptoms,
with watery mucous; also a cough productive of a clear, moderately
viscous sputum. Systemic symptoms 'NAC flu' may
also occur. If symptoms are severe, the dose of NAC may be cut
down to 600mg and slowly built up as may be tolerated. Symptoms
wane, sometimes quickly, after a few days if the chlamydial load
is small; if the load is large they may continue for a month
or more as the EBs are destroyed and their remains removed by
the immune system. As far as I am aware, NAC is unlikely to produce
die-off reactions with any other genus.
NAC's other valuable properties
NAC's ability to replenish depleted glutathione in an animal
model with defective transporter neuronal proteins is dramatic.
[Aoyama
K, Suh SW, Hamby AM, Liu J, Chan WY, Chen Y, Swanson RA. Neuronal
glutathione deficiency and age-dependent neurodegeneration in
the EAAC1 deficient mouse. Nat Neurosci. 2005 Nov 27;
{Epub ahead of print}]
The mice with this gene-defect aged prematurely and lost brain
substance. They had depleted glutathione levels in their neurones.
The authors comment in an interview: For several days, a group
of gene-deficient mice were fed N-acetylcysteine, an oral form
of cysteine that is readily taken up by neurons. When their neuron
slices were compared with slices from untreated gene-deficient
mice, it was found that N-acetylcysteine "had completely
corrected the biochemical defect" in their neurons, recounts
Swanson. "Their glutathione levels were normal, their ability
to withstand hydrogen peroxide toxicity was normal, and the oxidants
we saw in the neurons in response to oxidative challenges were
normal." This is remarkable. Major oxidative stress is a
cause of cell-death in MS. NAC's antioxidant benefits may not
be limited to the replenishment of depleted glutathione. [Gosset P, Wallaert
B, Tonnel AB, Fourneau C. Thiol regulation of the production
of TNF-alpha, IL-6 and IL-8 by human alveolar macrophages. Eur
Respir J. 1999 Jul;14(1):98-105.]
NAC may prove useful for other reasons, having been credited
with many beneficial effects:
replenishing glutathione, a major intracellular antioxidant;
see above.
preventing further new infections with C.
pneumoniae and thus averting the risk of MS relapse which
this can produce.
ameliorating intracellular infections (such
as influenza A) which are known to precede MS relapse;
modulating the immune response, making cells
more resistant to the effects of proinflammatory cytokines such
as TNF-a.
is itself a major antioxidant. It is non-toxic
in standard supplemental doses.
chelates heavy metals, which are known to
accentuate oxidative stress.
may moderate the effect of endotoxins, which
are made by chlamydiae. |
NAC may be of particular importance
in demyelinating conditions. Not only may it help moderate lipid
peroxidation as an antioxidant, it may also moderate the induction
of ceramide production by TNF alpha and consequent cell-death.[Singh I, Pahan K,
Khan M, Singh AK. Cytokine-mediated induction of ceramide production
is redox-sensitive. Implications to proinflammatory cytokine-mediated
apoptosis in demyelinating diseases. J Biol Chem. 1998 Aug 7;273(32):20354-62.] These authors also found that thiol
depletion of itself could induce ceramide production independently
of TNFalpha. Furthermore, they comment: 'NAC, which has been
used to block the cytokine-induced ceramide production in this
study and to inhibit cytokine-mediated induction of inducible
nitric oxide synthase in a previous study, is a nontoxic pharmaceutical
drug that enters the cell readily and serves both as a scavenger
of ROS and a precursor of GSH, the major intracellular thiol.
Therefore, the use of reductants such as NAC or other thiol compounds
may be beneficial in restoring cellular redox and in inhibition
of cytokine-mediated induction of inducible nitric oxide synthase
and breakdown of sphingomyelin thus reducing NO-mediated cytotoxicity
as well as ceramide-mediated apoptosis in neuroinflammatory diseases.'
Persons with a high EB load
may experience a variety of symptoms as the EBs are ruptured
and endotoxin and other bacterial material are released. Common
symptoms include pain round the eyes and over the sinuses; wheezing
and sputum production. This is not surprising, as the organism
will have been in the respiratory tract - producing EBs- for
a long time before it was forced to become a persistent intracellular
form. Other symptoms depend on the location of the EBs, but joint
pain, pain in soft tissues and abdominal discomfort have been
experienced. Symptoms tend to last a month or so.
(NAC is used medically to liquefy the sputum of patients with
lung diseases characterized by overproduction of mucus; this
makes expectoration easier. This liquefying action is carried
out by reduction of the disulphide bonds between protein molecules
in the sputum; the more of these bonds the more viscid the sputum.)
N-acetyl cysteine and Gliotoxin,
a poisonous metabolite secreted by Candida sp.
N-acetyl cysteine may also be
helpful in preventing the establishment of fungal infections
such as candidiasis. Candida (and fungi in other and often distant
genera) produce a toxin called gliotoxin [Shah DT, Larsen B. Clinical isolates
of yeast produce a gliotoxin-like substance. Mycopathologia.
1991 Dec;116(3):203-8.]

This is one member of a large family of mycotoxins which cause
damage to the immune system, provoking caspase-mediated apoptosis
in monocytes and macrophages. Gliotoxin molecules contain a highly
active surface disulphide bond which, on being reduced, damages
host proteins by altering their structure. Gliotoxin may also
deplete host glutathione, removing host antioxidant potential
and increasing free radical damage; the molecule has been shown
to oscillate between oxidized and reduced forms with the production
of unstable peroxides. From the yeast's point of view the production
of gliotoxin may be fundamental to the establishment of chronic
colonization; locally high levels are found, for instance, in
the genital tract of women with severe vaginal candidiasis [Shah DT, Glover DD,
Larsen B. In situ mycotoxin production by Candida albicans
in women with vaginitis. Gynecol Obstet Invest. 1995;
39(1): 67-9.] Gliotoxin
is a potent neurotoxin, and may alter gut motility. It impairs
the efficiency of host polymorphonuclear neutrophils [Shah DT, Jackman S,
Engle J, Larsen B. Effect of gliotoxin on human polymorphonuclear
neutrophils. Infect Dis Obstet Gynecol. 1998; 6(4): 168-75.] There is some evidence that intestinal
gliotoxin may cause dysfunction of the gut barrier by damaging
enterocytes [Upperman
JS, Potoka DA et al., Mechanism of intestinal-derived fungal
sepsis by gliotoxin, a fungal metabolite. J Pediatr Surg.
2003 Jun; 38(6): 966-70.]
Gliotoxin has been shown to be a virulence factor in other fungal
infections such as invasive aspergillosis.
N-acetyl cysteine, on theoretical grounds, may be expected to
neutralize gliotoxin by opening the disulphide bond, and indeed
has been shown to be protective in vitro. [Zhou X, Zhao A, Goping
G, Hirszel P. Gliotoxin-induced cytotoxicity proceeds via apoptosis
and is mediated by caspases and reactive oxygen species in LLC-PK1
cells. Toxicol Sci. 2000 Mar; 54(1): 194-202.] It probably acts by opening the gliotoxin
disulphide bond. Upperman and co-workers [reference above] found
that dithiothreitol, which also opens available disulphide bonds,
protected gut cells from gliotoxin-mediated apoptosis. Not all
orally taken N-acetyl cysteine is absorbed; that which remains
in the gut may be useful in counteracting gliotoxin produced
by intestinal candida. It should be noted that N-acetyl cysteine
does not possess intrinsic activity against the yeast itself;
Candida albicans is not inhibited even by high concentrations
(> 100 mg/L) of NAC [Wheldon D: unpublished data.]
Gardiner and co-authors
provide a very good overview of gliotoxin and related mycotoxins
[Gardiner
DM, Waring P, Howlett BJ. The epipolythiodioxopiperazine (ETP)
class of fungal toxins: distribution, mode of action, functions
and biosynthesis. Microbiology. 2005 Apr;151 (Pt 4): 1021-32.] the full text of which is available
as a pdf file.
Caveat There is confusion over the term gliotoxin.
The term was first used in the 1930s to describe a toxic metabolite
of the fungus Gliocladium fimbriatum; the name
is derived from the name of the organism. This toxin is the small
molecule with the heterocyclic nucleus spanned by a disulphide
bond as described above. Unfortunately the same name gliotoxin
has also been applied to a protein, toxic to the glial
(gliocyte) classes astrocytes and oligodendrocytes, which
has been found in the CSF of persons with MS [Menard A, Amouri R, Dobransky T, et
al., A gliotoxic factor and multiple sclerosis. J Neurol Sci.
1998 Feb 5; 154 (2) : 209-21.]
These two toxins have categorically different structures and
have nothing in common except their name. Despite this, Internet
articles have appeared which assume a common identity and which
forward the assertion that mycotoxins have been demonstrated
in the CSF of persons with MS.
Other sulphur-containing organic molecules have anti-candidal
activity; the best known being allicin, which is found in garlic
[Yamada
Y, Azuma K. Evaluation of the in vitro antifungal activity of
allicin. Antimicrob Agents Chemother. 1977 Apr;11(4):743-9.]
(On a personal note I have to say I love garlic. So, fortunately,
does Sarah. However, it has got me into trouble. A friend of
mine and I - both aged 11 - were hauled in front of the headmaster
one day. Our crime? Introducing wild garlic into our dinner to
remedy the dull, repetitive school-food blandness. Our accuser,
the housekeeper, bristled with indignation. I can see her face
now as she held out the pungent evidence. Forty-five years have
passed, and I'm pleased to say that garlic is much more acceptable
in the anglophone world.)
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page revised 15th January
2008
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