Download Vitamin D and tuberculosis

Emerging Science
Vitamin D and tuberculosis
Patricia Chocano-Bedoya and Alayne G Ronnenberg
Tuberculosis is highly prevalent worldwide, accounting for nearly two million deaths
annually. Vitamin D influences the immune response to tuberculosis, and vitamin D
deficiency has been associated with increased tuberculosis risk in different
populations. Genetic variability may influence host susceptibility to developing
active tuberculosis and treatment response. Studies examining the association
between genetic polymorphisms, particularly the gene coding for the vitamin D
receptor (VDR), and TB susceptibility and treatment response are inconclusive.
However, sufficient evidence is available to warrant larger epidemiologic studies
that should aim to identify possible interactions between VDR polymorphisms and
vitamin D status.
© 2009 International Life Sciences Institute
INTRODUCTION
According to the World Health Organization (WHO),
there were 9.2 million incident cases and 1.7 million
deaths from tuberculosis (TB) worldwide in 2006.1 The
current treatment of tuberculosis involves the use of multiple drugs on a daily dose for 6–8 months. Interruption
of the treatment leads to multidrug-resistant tuberculosis, a problem that has already been reported in over 100
countries in the world.1 Better treatments and vaccines
for TB are currently under development to battle against
the reemergence of tuberculosis and especially against
new forms of multidrug-resistant tuberculosis.
Mycobacterium tuberculosis, the etiologic agent of
TB, is a facultative intracellular bacterial parasite that can
be spread by inhalation of a minimal dose (1–5 bacilli).
The first immune defense response to TB infection begins
with the innate immune system, involving the epithelial
cells and alveolar macrophages in the airways. This initial
response is strengthened by recruitment of neutrophils,
which are among the first cells to arrive at the site of
infection.2,3
Macrophages phagocytize the bacilli, but the normal
destruction of bacilli by macrophages can be interrupted
by the defense mechanisms of the mycobacteria. One of
the potential pathways through which the mycobacteria
prevent their own destruction involves glycosylated
phosphatidylinositol lipoarabinomannan, a compound of
the mycobacterial cell membrane. Lipoarabinomannan is
translocated to the phagosome wall, interrupting the
normal maturation of the phagosome and its further
fusion with the lysosome.2,4 Another potential mycobacterial defense mechanism involves inhibition of Ca2+
signaling events, which are also required for phagosome
maturation.5 Thus protected from host defenses, the
viable mycobacteria reproduce inside the macrophages
and can also migrate to other tissues. However, a localized
inflammatory response promotes the recruitment of T
lymphocytes, which leads to the formation of a granuloma5 to wall off the spread of the infection. The TB
infection is usually contained inside the granuloma, and
the infection may remain dormant, or latent, for many
years. However, immunodeficiency secondary to an event
such as coinfection with human immunodeficiency
virus (HIV) or malnutrition, can lead to activation of the
disease.6,7
Although tuberculosis is a highly infective disease,
only 1 in 10 infected persons may become sick with
active TB.1 The susceptibility to active disease can be
influenced by environmental and genetic factors or by
Affiliations: P Chocano-Bedoya and AG Ronnenberg are with the Department of Nutrition, School of Public Health and Health Sciences,
University of Massachusetts, Amherst, Massachusetts, USA.
Correspondence: AG Ronnenberg, Department of Nutrition, School of Public Health and Health Sciences, University of Massachusetts,
209 Chenoweth Lab, 100 Holdsworth Way, Amherst, MA 01003, USA. E-mail: [email protected], Phone: +1-413-545-1076,
Fax: +1-413-545-1074.
Key words: gene-nutrient interaction, innate immunity, tuberculosis, vitamin D deficiency, vitamin D receptor polymorphism
doi:10.1111/j.1753-4887.2009.00195.x
Nutrition Reviews® Vol. 67(5):289–293
289
gene-environment interactions. Genetic variability may
influence not only host susceptibility to active TB but also
host response to treatment. Polymorphisms of many different gene candidates have been studied,8 and the gene
for the vitamin D receptor (VDR) is of great interest.9–14
ROLE OF VITAMIN D IN TUBERCULOSIS
Even before the discovery of the etiologic cause of tuberculosis by Robert Koch in 1903, vitamin D from cod liver
oil7 and from exposure to sun or radiation was used to
treat tuberculosis.13,15 Several recent studies in different
populations have associated a deficiency in vitamin D
with increased risk of tuberculosis.16–21 In a recent metaanalysis by Nnoaham and Clarke,21 a pooled effect size of
vitamin D was estimated to be 0.68 (95% CI 0.43–0.93),
indicating that vitamin D levels were 0.68 SD lower in
persons with tuberculosis than in controls. However,
these findings cannot be considered conclusive since the
association may be confounded by important variables,
such as smoking and sunlight exposure, which were not
accounted for in the analysis.
It is well-established that immune cells can produce
the hormonally active metabolite of vitamin D. Macrophages and other immune cells can express 1a-hydoxylase,
the enzyme that converts circulating 25-hydroxyvitamin
D3 into 1,25-dihydroxyvitamin D3, the active form of
vitamin D.7 Moreover, M. tuberculosis infection activates
Toll-like receptors (TLR1/2) that mediate the activation
of different cells in the innate immune system and their
expression of cytokines and antimicrobial peptides.5,22
Recent observations indicate that activation of this cell
surface receptor also upregulates expression of both the
1-a-hydroxylase enzyme and the VDR in monocytes and
macrophages, leading to both increased levels of active
vitamin D and increased potential binding of 1,25dihydroxyvitamin D with the VDR.7,22
Although the biological mechanisms through which
vitamin D modulates the immune system to fight Mycobacterium infection are still under study,3,15,23 two possible
mechanisms have emerged as the most likely. For
instance, 1,25-dihydroxyvitamin D3 appears to reduce the
viability of M. tuberculosis by enhancing the fusion of the
phagosome and lysosome in infected macrophages.4,7,13
The capacity of Mycobacterium infection to prevent macrophage maturation and formation of the phagolysosome
is completely reversed in the presence of 1,25dihydroxyvitamin D3.4 The pathways used to promote
vitamin D-induced phagolysosome formation are independent of the classical interferon-gamma (IFN-g)dependent macrophage activation and involve products
of phosphatidylinositol-3-kinases (PI3K),4,24 which help
to regulate the transport of endosomes to lysosomes.5
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In addition, 1,25-dihydroxyvitamin D may enhance
the production of LL-37, an antimicrobial peptide of the
cathelicidin family.3,22,24 Antimicrobial peptides, such as
defensins and cathelicidins, are involved as a first line of
defense in the prevention of infections, including tuberculosis.3 Although cathelicidins are widely distributed in
mammals, LL-37 is the only member of the cathelicidin
family that has been identified in humans, where it is
found in alveolar macrophages, lymphocytes, neutrophils, and epithelial cells.3,7 In addition to having direct
bactericidal activity, LL-37 also modulates the immune
response by attracting monocytes, T cells, and neutrophils to the site of infection.7 The presence of 1,25dihydroxyvitamin D3 in neutrophils and macrophages
upregulates in a dose-dependent manner the hCAP-18
gene that codes for LL-37,7,24 which suggests that 1,25dihydroxyvitamin D induction of LL-37 may play a role
in host defense against TB infection.
VITAMIN D RECEPTOR GENE POLYMORPHISMS
AND TUBERCULOSIS
The vitamin D receptor (VDR) gene is found in the
chromosomal 12q13 region. VDR gene polymorphisms
are commonly found in many population groups,
although the prevalence of certain VDR genotypes
varies among different populations. Most of the gene
polymorphisms studied are based only on restriction
fragment length polymorphism (RFLP) analysis, which
does not identify the functional effects of these
changes.13,25,26 Genetic alterations of the VDR gene may
lead to defects in gene activation or to changes in the
protein structure of the VDR, both of which could affect
the cellular functions of 1,25-dihydroxyvitamin D3.
Various VDR polymorphisms could also be linked to
each other or to unidentified genes that are important
determinants of disease risk.
In the 3’ end of the VDR gene, several polymorphisms (BsmI, ApaI, and TaqI) with strong linkage disequilibrium (LD) have been studied. Although these
nucleotide changes in the VDR gene are predicted to be
“silent” and to have no effect on the structure of the
expressed VDR protein, they may be involved in regulating VDR gene expression,11 or they could potentially be
linked with other truly functional nucleotide sequences
in the VDR gene.11,25 It has been suggested that the mRNA
coded from the TaqI t allele of the VDR gene is more
stable than the mRNA from the T allele of the VDR
gene.13
A non-silent VDR gene polymorphism is the FokI
polymorphism, found in exon 2, which is at a translation
initiation start site and is predicted to change the structure of the coded protein. A thymine-to-cytosine (T→C)
change found in the F allele leads to an alternative transNutrition Reviews® Vol. 67(5):289–293
lational start site and a VDR protein that is three amino
acids shorter than that of the f variant. Although the
difference between the two proteins is only three amino
acids, it has been suggested that the more commonly
observed shorter VDR protein is functionally more
active.11,25,26
Several studies have examined TaqI and FokI VDR
gene polymorphisms and their association with susceptibility to tuberculosis in different populations with inconclusive findings.11 An early study of this question by
Bellamy et al.10 found that the TaqI tt VDR genotype was
associated with decreased risk of TB in The Gambia
compared with the TT and Tt genotypes combined
(OR = 0.53, 95% CI 0.31–0.88). In contrast, a study in
India found that the tt TaqI VDR genotype was associated
with increased TB susceptibility, at least in women.27 A
recent study among native Paraguayans found that the
TaqI t VDR allele protects against active disease, but not
infection, while the FokI F VDR genotype was associated
with decreased risk of TB (assessed by PPD status).14
Other studies found no significant association between
the TaqI and FokI polymorphisms and risk of TB, but
probably had limited statistical power due to small
sample sizes.11
VITAMIN D RECEPTOR GENE POLYMORPHISMS AND
RESPONSE TO TREATMENT OF PULMONARY
TUBERCULOSIS
VDR polymorphisms may affect not only host susceptibility to tuberculosis, but also the response to treatment.
Two studies have focused on the association of polymorphisms in the FokI and TaqI VDR genes12 and TB
susceptibility; one of them also evaluated the association
with the ApaI VDR gene polymorphism.9
Roth et al.9 studied the association between FokI and
TaqI VDR gene polymorphisms and susceptibility to TB
and sputum conversion time following treatment among
inhabitants of an area in the outskirts of Lima, Peru,
where the incidence of TB is very high.12 A case-control
study design was used to evaluate the association between
VDR gene polymorphisms and susceptibility to TB. Study
cases were 103 persons, aged 15–45 years, with confirmed
TB (excluding HIV-positive patients and pregnant
women) who were receiving directly observed therapy, as
recommended by the WHO.28 Two age- and sex-matched
controls were chosen for each case, one with a positive
tuberculin test (PPD) and the other with a negative test.
The prevalence of the TaqI t allele in the VDR gene was
lower among the PPD-positive controls than among the
PPD-negative controls and even lower among the TB
cases, suggesting that the t allele was somehow protective
against TB susceptibility. However, there was no significant difference in TB susceptibility between the FokI VDR
Nutrition Reviews® Vol. 67(5):289–293
genotypes ff versus FF (OR = 0.84, 95% CI 0.34–2.86) or
Ff versus FF (OR = 0.64, 95% CI 0.25–1.62), or the TaqI
VDR genotype Tt versus TT (OR = 0.61, 95% CI 0.28–
1.29).
Although the FokI VDR genotype had no apparent
effect on susceptibility to TB, quite different effects were
seen regarding the response to TB treatment. To evaluate
this effect, the researchers1 conducted a cohort study
among 78 patients with confirmed pulmonary TB (positive sputum test and TB symptoms). Kaplan-Meier survival analysis for the total treatment-time follow-up
period showed that patients with the VDR FokI FF genotype had significantly faster conversion times in the
sputum cultures and in the auramine staining tests compared to those with the VDR FokI ff genotype, thus supporting the notion that the F allele, which results in the
expression of a shorter VDR protein, somehow enhances
the treatment response compared to the FokI f allele,
which codes for a slightly longer VDR protein. With
respect to the TaqI polymorphisms, no study participants
had the tt genotype. Among TB patients, the conversion
of culture tests – but not the auramine staining tests – was
significantly faster for those with the TT VDR genotype as
compared to the Tt genotype. In conclusion, there was a
beneficial effect on the host response to TB among participants with the TT or FF VDR genotypes, especially for
culture conversion, as compared to the Tt and the ff VDR
genotypes.12
Recently, Baab et al.12 conducted a similar study
among a population of women and men, aged 18–65
years, from Western Cape, South Africa, where the incidence of TB in 2003 was high (919/100,000). Study cases
were 249 persons with newly diagnosed TB (excluding
pregnant women and persons with HIV or other chronic
diseases); controls were 352 persons with no previous
history or symptoms of TB selected from clinics, households, and workplaces in the same geographical area. A
cohort study was conducted among the cases of TB, starting from the day of diagnosis up to 12 months. Patients
were required to provide sputum samples for ZiehlNielsen staining and M. tuberculosis culture on the day of
diagnosis, the following 2 days, then weekly up to month
2, and then monthly until month 6. After that, two subsequent samples were collected at months 9 and 12. Conversion time (from positive to negative M. tuberculosis in
sputum) was calculated in days from the day of diagnosis
to an average between the dates of the last date of a
positive result and the first of two successive negative
results if less than 92 days had elapsed between the positive and negative results. Otherwise, the last positive day
was used as the conversion day. Participants were categorized as “fast respondents” if their conversion time was
before day 55 after diagnosis, and “slow respondents”
otherwise.
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For the case-control study, no significant associations were observed between the individual VDR genotypes and the presence or absence of pulmonary TB.
Moreover, after adjusting for age and gender, no statistically significant differences were observed between the
diplotype frequencies in cases and controls. However, for
the response-to-treatment cohort study, Babb et al.9
found a faster smear conversion time for the VDR genotypes ApaI AA and TaqI TT and Tt as compared to the
ApaI aa and TaqI tt VDR genotypes, respectively. No statistically significant difference in the culture conversion
time was observed between genotypes. For the categorization between “fast respondents” and “slow respondents”, there was a significant trend to a faster smear
conversion in those with a VDR FokI f allele and for a
faster culture conversion in patients with the ApaI A
allele. In summary, Babb et al. found no significant association between the ApaI, TaqI and FokI polymorphisms
and susceptibility to TB. However, ApaI AA and TaqI Tt
and TT genotypes may contribute to a faster response to
treatment.9
The association between VDR polymorphisms and
response to TB treatment remains inconclusive, and larger
studies are needed. In the study among Peruvians, the FokI
F and TaqI T alleles were associated with a faster response
to TB treatment.9 In the South African study,there was also
a faster response to TB treatment among people with the
TaqI T and the ApaI A alleles, whereas no definitive association was observed for the FokI genotypes.12
Selection of the optimal method to evaluate response
to treatment (auramine staining or culture of sputum) is
of substantial importance given the differences found
among the studies. Roth et al.12 found significant associations for the TaqI genotypes among the TB cultures – not
the stained samples – suggesting that VDR genotypes are
potentially associated with mycobacterial viability, rather
than with the quantity of expectorated microorganisms.9
A low number of viable bacteria can give positive results
in the culture, but can be negative in the auramine staining test.12
CONCLUSION
Although the immune system responds quickly to the
presence of M. tuberculosis bacilli, this pathogen has
developed several ways to avoid being killed by macrophages and, instead, to reproduce inside them. Vitamin D
plays an important role in the immune response to M.
tuberculosis by promoting both formation of the phagolysosome and also production of the antimicrobial peptide
LL-37, which has direct bactericidal activity and an
immune-regulating function.
The available evidence does not allow for a solid
conclusion concerning the relationship between various
292
VDR polymorphisms and TB susceptibility or response
to treatment. However, these findings are intriguing and
support additional inquiry on this research question in
order to target individuals at risk of developing TB or to
devise new treatment modalities. Additional studies with
sufficient statistical power to investigate the effects of
various polymorphisms in the VDR gene or other relevant genes on TB are needed. These studies should recognize that the prevalence of certain genotypes varies
among different ethnic groups, which influences how
suitable different ethnicities may be for exploring particular gene-disease interactions. For example, in the study
among South Africans, 41 participants (6.8% of the total
population) had the tt VDR genotype3,4,22,24 compared to
none in the study among Peruvians.9 Also, the VDR FokI
f allele is known to occur less frequently in Africans than
in Caucasians and Asians,12 but the studies reviewed
above found that Peruvians and native Paraguayans have
the lowest prevalence of the FokI ff genotype reported so
far.25 Nevertheless, it should be possible to generalize the
functional effects associated with certain polymorphisms
to other populations because the physiological role and
underlying molecular mechanisms of vitamin D action
likely remain the same.14 The inconclusiveness of available studies highlights the critical need to better assess the
vitamin D status of these study populations. Future
studies should investigate possible interactions between
vitamin D status, genetic polymorphisms in the VDR
gene and other genes involved with vitamin D metabolism and function, and TB susceptibility and treatment
response.
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