Association of IL1A and IL1B loci with primary open angle glaucoma.

Abstract

Recent studies suggest that glaucoma is a neurodegenerative disease in which secondary degenerative losses occur after primary insult by raised Intraocular pressure (IOP) or by other associated factors. It has been reported that polymorphisms in the IL1A and IL1B genes are associated with Primary Open Angle Glaucoma (POAG). The purpose of our study was to investigate the role of these polymorphisms in eastern Indian POAG patients.
The study involved 315 unrelated POAG patients, consisting of 116 High Tension Glaucoma (HTG) patients with intra ocular pressure (IOP) > 21 mmHg and 199 non-HTG patients (presenting IOP < 20 mmHg), and 301 healthy controls from eastern India. Genotypes were determined by polymerase chain reaction and restriction digestion for three single nucleotide polymorphisms (SNPs): IL1A (-889C/T; rs1800587), IL1B (-511C/T; rs16944) and IL1B (3953C/T; rs1143634). Haplotype frequency was determined by Haploview 4.1 software. The association of individual SNPs and major haplotypes was evaluated using chi-square statistics. The p-value was corrected for multiple tests by Bonferroni method.
No significant difference was observed in the allele and genotype frequencies for IL1A and IL1B SNPs between total pool of POAG patients and controls. However, on segregating the patient pool to HTG and non-HTG groups, weak association was observed for IL1A polymorphism (-889C/T) where -889C allele was found to portray risk (OR = 1.380; 95% CI = 1.041-1.830; p = 0.025) for non-HTG patients. Similarly, 3953T allele of IL1B polymorphism (+3953C/T) was observed to confer risk to HTG group (OR = 1.561; 95% CI = 1.022-2.385; p = 0.039). On haplotype analysis it was observed that TTC was significantly underrepresented in non-HTG patients (OR = 0.538; 95% CI = 0.356- 0.815; p = 0.003) while TCT haplotype was overrepresented in HTG patients (OR = 1.784; 95% CI = 1.084- 2.937; p = 0.022) compared to control pool. However, after correction for multiple tests by Bonferroni method, an association of only TTC haplotype with non-HTG cases sustained (pcorrected = 0.015) and expected to confer protection.
The study suggests that the genomic region containing the IL1 gene cluster influences the POAG pathogenesis mostly in non-HTG patients in eastern India. A similar study in additional and larger cohorts of patients in other population groups is necessary to further substantiate the observation.

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Open Access
RESEARCH ARTICLE
Association of IL1A and IL1B loci with primary open
angle glaucoma
© 2010 Mookherjee et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Com-
mons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduc-
tion in any medium, provided the original work is properly cited.
Research article
Suddhasil Mookherjee1, Deblina Banerjee1, Subhadip Chakraborty1, Antara Banerjee1, Indranil Mukhopadhyay2,
Abhijit Sen3 and Kunal Ray*1
Abstract
Background: Recent studies suggest that glaucoma is a neurodegenerative disease in which secondary degenerative
losses occur after primary insult by raised Intraocular pressure (IOP) or by other associated factors. It has been reported
that polymorphisms in the IL1A and IL1B genes are associated with Primary Open Angle Glaucoma (POAG). The
purpose of our study was to investigate the role of these polymorphisms in eastern Indian POAG patients.
Methods: The study involved 315 unrelated POAG patients, consisting of 116 High Tension Glaucoma (HTG) patients
with intra ocular pressure (IOP) > 21 mmHg and 199 non-HTG patients (presenting IOP < 20 mmHg), and 301 healthy
controls from eastern India. Genotypes were determined by polymerase chain reaction and restriction digestion for
three single nucleotide polymorphisms (SNPs): IL1A (-889C/T; rs1800587), IL1B (-511C/T; rs16944) and IL1B (3953C/T;
rs1143634). Haplotype frequency was determined by Haploview 4.1 software. The association of individual SNPs and
major haplotypes was evaluated using chi-square statistics. The p-value was corrected for multiple tests by Bonferroni
method.
Results: No significant difference was observed in the allele and genotype frequencies for IL1A and IL1B SNPs between
total pool of POAG patients and controls. However, on segregating the patient pool to HTG and non-HTG groups, weak
association was observed for IL1A polymorphism (-889C/T) where -889C allele was found to portray risk (OR = 1.380;
95% CI = 1.041-1.830; p = 0.025) for non-HTG patients. Similarly, 3953T allele of IL1B polymorphism (+3953C/T) was
observed to confer risk to HTG group (OR = 1.561; 95% CI = 1.022-2.385; p = 0.039). On haplotype analysis it was
observed that TTC was significantly underrepresented in non-HTG patients (OR = 0.538; 95% CI = 0.356- 0.815; p =
0.003) while TCT haplotype was overrepresented in HTG patients (OR = 1.784; 95% CI = 1.084- 2.937; p = 0.022)
compared to control pool. However, after correction for multiple tests by Bonferroni method, an association of only TTC
haplotype with non-HTG cases sustained (pcorrected = 0.015) and expected to confer protection.
Conclusion: The study suggests that the genomic region containing the IL1 gene cluster influences the POAG
pathogenesis mostly in non-HTG patients in eastern India. A similar study in additional and larger cohorts of patients in
other population groups is necessary to further substantiate the observation.
Background
Glaucoma is a heterogeneous group of optic neuropathy,
characterized by typical visual field loss, often associated
with elevated intra ocular pressure. It affects over 60 mil-
lion people worldwide and is the second largest cause of
blindness after cataract [1]. Among different subtypes of
glaucoma, Primary Open Angle Glaucoma (POAG) is the
most frequently occurring subtype. Till date 25 loci have
been implicated in the pathogenesis of POAG with three
underlying genes, i.e. Myocilin [2], Optineurin [3] and
WDR36 [4]. Recent studies suggest that POAG is caused
mainly by genetic predisposition and interaction with
other risk factors. It is estimated that 72% of all POAG
cases represent the inherited and familial form of the dis-
ease that does not show a clear pattern of Mendelian
inheritance [5].
POAG results from progressive excavation of the optic
disc with corresponding loss of vision by raised Intraocu-
lar pressure (IOP) or by other associated factors. How-
* Correspondence: kunalray@gmail.com
1 Molecular & Human Genetics Division, Indian Institute of Chemical Biology,
Council of Scientific & Industrial Research, Kolkata, India
Full list of author information is available at the end of the article

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ever, a substantial number of POAG patients have normal
IOP - a condition referred to as normal tension glaucoma
(NTG). The main aqueous outflow pathway, controlling
IOP, consists of series of endothelial cell-lined channels in
the angle of the anterior chamber such as the trabecular
meshwork (TM), Schlemm's canal, the collector channels
and the episcleral venous system. Accumulation of extra-
neous materials or cells within the TM, alterations in the
cell structure, accelerated TM cell death and collapse of
trabecular beams as a result of oxidative stress, vascular
dysregulation and aging causes reduced outflow in open
angle glaucoma. The cells surviving the sublethal injury
tend to mount a protective response involving the expres-
sion of new genes [6].
It has been suggested that, in addition to other contrib-
uting factors, immune system in the body may have a role
in neuro-degeneration in glaucoma [7]. For example,
antibody levels against some of the self proteins [8,9] e.g.
anti-NSE antibody [10], antibodies to heat shock proteins
[11] and beta 2 glycoproteins [12] have been reported to
be significantly higher in glaucoma patients. It has been
reported that Interleukin-1 (IL1), an inflammatory
cytokine, is involved in ischemic and excitotoxic damage
in the retina [6]. Unlike the normal TM cells, glaucoma-
tous cells show endogenous expression of IL1. In vitro
experiments show that a stress response specific to the
aqueous outflow pathway is activated in the TM cells and
controlled by an Interleukin-1 (IL1) autocrine feedback
loop through transcription factor NF-kappaB [6]. It has
also been suggested that the chronic activation of the
stress response in the aqueous outflow pathways might
initiate and exacerbate high tension glaucoma (HTG) [6].
Shaftel et al have demonstrated that IL1 protein pro-
motes the development of β-amyloid deposits in
Alzheimer's patients [13]. Similar β-amyloid plaques are
found in the Retinal Ganglion Cells (RGC) of mouse with
experimental glaucoma. Drugs which work to prevent
this buildup of the β-amyloid protein in Alzheimer brains
can be used to treat glaucoma in animal models [14].
Interestingly, studies conducted on Japanese and German
population groups suggest a high frequency of POAG in
patients with Alzheimer's disease (AD) [15,16]. Both AD
and glaucoma show similar neuronal degeneration.
Moreover, patients with AD shows high level of axonal
degeneration of the optic nerve and retinal cell damage,
especially ganglion cells [17,18]. The ε4 allele of APOE
has been shown to be significantly over expressed in AD
and glaucoma patients, though the contribution remains
controversial [19,20]. Thus glaucoma may be viewed as a
neuro-degenerative disorder
Alzheimer's with common genetic risk factors, mecha-
nisms and pathways.
Several SNPs in the IL1 gene cluster (i.e. IL1A -889C/T;
IL1B -511 C/T & 3953C/T) have been reported to be
associated with Alzheimer's disease [21,22]. Recently
these SNPs have also been examined for association with
POAG through multiple studies in Chinese population
[23-27]. Two independent studies by Wang et al [25] and
Lin et al [24] have claimed association of POAG with
IL1A (-889 C/T) and IL1B (3953 C/T) SNPs, respectively.
However, a recent study by How et al [23] reported a lack
of association of these polymorphisms with the disease.
In the context of developing information on the poten-
tial involvement of the IL1 gene cluster in POAG, we
aimed to investigate the role of this genomic region using
the well-studied three SNPs (IL1A -889C/T, IL1B -511C/
T, IL1B +3953C/T) (Figure 1) in eastern Indian patient
cohort.
similar to that of
Methods
Selection of study subjects
The patients with POAG and control subjects were
recruited for the study from Dristipradip Eye Clinic, Kol-
kata. Individuals in both the cohorts are inhabitants of
Kolkata, West Bengal (eastern India), speak Bengali lan-
guage, and belong to the Indo-European linguistic group.
The patient cohort consisted of 315 POAG patients con-
sisting of 116 patients with presenting IOP > 21 mmHg,
Figure 1 Location of the SNPs in IL1 gene cluster IL1 alpha, IL1 beta, IL1 receptor antagonist (RN). The promoter regions and coding sequences
of the genes are shown in light and dark shaded boxes. The dbSNP refrence SNP ID for each SNP is provided. rs1800587 and rs16944 are located in
the promoter (p) region of IL1A and IL1B, respectively while rs1143634 represents a synonymous SNP (F105F) in the coding (c) sequence of IL1B gene.
The genomic distances between the SNPs are also shown.

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considered as high tension glaucoma (HTG) cases and
199 non-HTG patients with presenting IOP < 20 mmHg.
Diagnosis of POAG involved clinical, ocular and sys-
temic examinations. Intraocular pressure (IOP) was mea-
sured by Goldmann applanation tonometry (Haag-Streit
USA Inc., Mason, OH) followed by pachymetry (Ocuscan
A, Alcon, Texas, USA). A Goldman 3-mirror gonioscope
(Ocular Instrument, Bellevue, WA) was used to assess the
angles of the anterior chamber and the optic disc. The
optic disc was also evaluated with a +78D lens. Auto-
mated threshold field analysis was done using the Hum-
phrey Field Analyzer II (Carl Zeiss, Dublin, CA). The
retinal nerve fiber layer (RNFL) was investigated by Scan-
ning Laser Polarimetry (SLP) with variable corneal com-
pensation technique (GDx-Vcc, Carl Zeiss, Dublin, CA).
An increased intraocular pressure above 21 mmHg, sig-
nificant cupping of the optic disc (> 0.7) with or without
peripapillary changes and the presence of clinically open
angle (angle of the anterior chamber) on Gonioscopy
raised the suspicion of POAG, which was confirmed by
typical reproducible visual field changes, viz. arcuate,
Bjerrum, Seidel, paracentral and annular scotoma and
nasal steps and Scanning Laser Polarimetry for RNFL
analysis (Nerve fibre Indicator > 30). The pre-perimetric
cases were identified by RNFL analysis. These patients
were categorised as the HTG patients. There was also
another group of individuals, the non-HTG patients, who
had an IOP less than 20 mmHg on presentation but had
cupping of the optic disc, RNFL loss diagnosed by SLP
and visual field changes characteristic of POAG. In each
case the IOP has been corrected for central corneal thick-
ness (CCT). Thus, the patient pool consisted of 315 adult
onset open angle glaucoma cases. The age at diagnosis
ranged from 42 to 88 years, with a mean ± standard devi-
ation of 64 ± 10 years. However, individuals with any his-
tory of inflammation or ocular trauma (past and present)
and ocular hypertension were excluded from this study.
In this study, 301 controls were recruited following the
criteria which include: age > 40 years (mean age ± SD,
55.7 ± 10.7 years), without any family history of glaucoma
or ocular hypertension, IOP less than 20 mmHg in both
eyes in at least last two check ups, CCT greater than 500
μm in both eyes, no visual field defect, normal Scanning
Laser Polarimeter parameters (i.e. a good yellowish bow
type scan pattern, deviation map within normal limit, a
good double hump pattern in conduction map, TSNIT
parameters within normal limit, Nerve Fibre Indicator <
30 for both eyes), cup discs were physiological and similar
in both eyes, cup to disc ratio < 0.2, no defect in disc rim
or margin and no sphincter haemorrhage around the
disc. Individuals with high myopia, diabetes and hyper-
tension were excluded from the control group.
Collection of blood samples and genomic DNA preparation
Eight milliliters of peripheral blood was collected with
EDTA from the POAG patients and normal individuals
with their written consent. Genomic DNA was prepared
from fresh whole blood using the PAX gene blood DNA
isolation kit (Qiagen, Hilden, Germany). The DNA was
dissolved in TE (10 mM Tris-HCl, 1 mM EDTA, pH 8.0).
The study protocol adhered to the tenets of the Declara-
tion of Helsinki and was approved by the Institutional
Review Board.
Genotyping
Genotyping was done by polymerase chain reaction and
restriction digestion (PCR-RFLP). All the PCR reaction
was carried out in 20 μl reaction volume using 80 ng of
total genomic DNA with Ex Prime Taq Premix (GeNet
Bio, South Korea) with specific primers for IL1A (-889C/
T), IL1B (-511C/T) and IL1B (3953C/T) (Integrated DNA
Technologies, Coralville, Iowa, USA) as described previ-
ously [25,27]. The PCR conditions were as follows: an ini-
tial denaturation at 95°C for 4 mins followed by 30 cycles
of 30 secs of denaturation at 95°C, 30 secs of annealing at
58-60°C and 30 secs of extension at 72°C, with a final
extension at 72°C for 4 mins. All the PCR products were
detected on a 6% polyacrylamide gel with ethidium-bro-
mide staining. The PCR products for both IL1A and IL1B
were subjected to restriction digestion with appropriate
enzymes from NEB (New England Biolabs Inc. Beverly,
MA) for 3 hrs at optimum temperatures as described pre-
viously [25,27]. The digested products were analyzed on a
6% polyacrylamide gel and the alleles were scored as
described in earlier studies [25,27].
Bioinformatics and Statistical analysis
Haplotypes and their frequencies were determined for
comparison between patients and controls using Haplo-
view 4.1 software http://www.broad.mit.edu/mpg/haplo-
view[28]. The allele frequencies of the SNP and
haplotypes were compared between patients and controls
using chi square test. The p-values were corrected for
multiple testing by Bonferroni method.
Results
Lack of association between IL1A and IL1B polymorphisms
and POAG
On genotyping 315 patients for three selected SNPs, IL1A
(-889C/T), IL1B (-511C/T) and IL1B (3953C/T), no sig-
nificant association was observed with the POAG. How-
ever, on further sub-dividing the patients according to
their presenting IOP, i.e. IOP > 21 mmHg (HTG) and the
IOP < 20 mmHg (non-HTG), marginal associations were
observed in non-HTG patient group and HTG patient

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group with IL1A (-889C) allele (OR = 1.380; 95% CI =
1.041- 1.830; p = 0.025) and IL1B (+3953T) allele (OR =
1.561; 95% CI = 1.022- 2.385; p = 0.039), respectively.
However, the observed association was nullified after
Bonferroni correction for multiple tests (Table 1). There-
fore, our data did not unequivocally suggest having an
association with the three SNPs examined.
IL1 haplotype distribution among patient and control
group
Haplotype analysis revealed presence of total eight hap-
lotypes in both patients and controls. No significant LD
was observed between the three SNPs genotyped (Fig-
ure 2). Among the eight haplotypes, five (CTC, CCC,
TTC, TCT, TCC) represented the majority of chromo-
somes (~95%) in both POAG patients and controls
(Table 2). The remaining three haplotypes (TTT, CTT,
CCT) were in much lower frequency (i.e. up to max.
3.5%) and not considered for further analysis as pre-
dicted number of patients and controls carrying these
haplotypes were too small for carrying out any associa-
tion study. Thus for haplotypes five tests were consid-
ered for Bonferroni correction. Among the five major
haplotypes TTC haplotype was significantly under-rep-
Table 1: Allele frequencies of SNPs in IL-1 gene cluster in POAG patients and controls
SNP (Gene) AllelePatient Subgroup Patient (n) Control (n)p-value Adjusted p-Value*OR (95% CI)
-889C/T
rs1800587
(IL-1A)
POAG0.71 (448) 0.204--
C Non-HTG0.74 (296) 0.68 (408)0.0250.075 1.380(1.041-1.830)
HTG0.66 (152)0.534--
POAG 0.29 (182)0.204--
T Non-HTG0.26 (102) 0.32 (194) 0.0250.0750.725(0.547-0.961)
HTG0.34 (80) 0.534--
-511C/T
rs16944
(IL-1B)
POAG 0.44 (276)0.114--
C Non-HTG0.43 (173)0.39(237)0.197--
HTG0.44 (103)0.185--
POAG 0.56 (354)0.114--
T Non-HTG0.57 (225)0.61(365)0.197--
HTG 0.56 (129)0.185--
+3953C/T
rs1143634
(IL-1B)
POAG0.85 (538)0.102--
CNon-HTG0.87 (345) 0.89 (533)0.380--
HTG 0.83 (193)0.0390.117 0.641(0.419-0.979)
POAG 0.15 (92)0.102-
TNon-HTG0.13 (53)0.11(69)0.380--
HTG0.17 (39) 0.039 0.1171.561(1.022-2.385)
* P-values were adjusted by Bonferroni method.
The location of the SNP in the genomic region and corresponding dbSNP reference ID is furnished. None of the 3 SNPs was significantly
associated with the entire POAG patient cohort or when it is subdivided to two groups: HTG (IOP > 21 mmHg) and non-HTG (IOP < 20 mmHg).
The numbers in parenthesis next to 'Allele frequency' represent numbers of chromosomes in the group. The weak association of IL1A (-889C/T)
and IL1B (+3953C/T) polymorphisms were nullified after Bonferroni adjustments for multiple tests.

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resented in non-HTG patient group (OR = 0.538.; 95%
CI = 0.356-0.815; p = 0.003) as well as the entire POAG
(OR = 0.697; 95% CI = 0.498-0.976; p = 0.036) group
compared to the control group (Table 2). However after
correction for multiple tests by Bonferroni method, the
association for POAG group did not sustain. Thus, it
appears that the lower p-value for POAG is contributed
entirely due to the association with the non-HTG group
since no significant association of TTC haplotype was
observed with HTG patient group. On the other hand,
TCT haplotype was found to be over-represented in
HTG patient group (OR = 1.784; 95% CI = 1.084-2.937;
p = 0.022) but the association was nullified after correc-
tion for multiple tests (Table 2).
In an attempt to examine whether the effect of the
TTC haplotypes in non-HTG patients is related to age,
we divided these patients into two groups: (i) up to 50
years, and (ii) above 50 years. We observed that only
the patients above 50 years TTC haplotype appeared to
have a protective effect (OR = 0.477; 95% CI = 0.287-
0.792; p = 0.004) (Table 3) even after Bonferroni
correction.
Figure 2 LD structure between the three SNPs in POAG patients
and controls. No significant LD was observed between the three
SNPs. The LD value was calculated using Haploview 4.1 software. The
values in the boxes are the r-squared pairwise LD values given by the
software. The standard r-squared color scheme is followed here i.e the
extent of LD increases with darker shades.
Table 2: Haplotype distribution in IL1 gene cluster between POAG patients and controls
HaplotypesPatient SubgroupFrequency in PatientsFrequency in Controls p-valueAdjusted p-value*OR (95% CI)
CTCPOAG
Non-HTG
HTG
0.411(259)
0.423(168)
0.381(88)
0.423(255) 0.699
0.963
0.244--
CCC POAG
Non-HTG
HTG
0.264(166)
0.28(111)
0.245 (57)
0.234(141) 0.235
0.111
0.727
-
-
-
-
-
-
TTC
POAG
Non-HTG
HTG
0.108(68)
0.085(34)
0.15(35)
0.146(89)
0.036
0.003
0.912
0.18
0.015
-
0.697(0.498-0.976)
0.538(0.356-0.815)
-
TCTPOAG
Non-HTG
HTG
0.084(53)
0.059(23)
0.121(28)
0.072(43)0.406
0.395
0.022
-
-
0.11
-
1.784(1.084-2.937)
TCCPOAG
Non-HTG
HTG
0.071(45)
0.078(31)
0.055(13)
0.081(49)0.510
0.841
0.211
-
-
-
Haplotype was constructed using SNPs of IL1 gene cluster in the following order:-889 C/T (rs1800587), -511C/T (rs16944) and +3953C/T
(rs1143634). TTC haplotype have the potential to act as a protective haplotype in non-HTG patient group. The numbers in parenthesis in 3rd and
4th columns represent numbers of chromosomes in the group. * p-values were adjusted by Bonferroni method.

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Discussion
Glaucoma being a complex disease, understanding of the
molecular mechanism underlying its pathogenesis is lim-
ited. In addition to the identified genetic loci linked to
POAG, other genetic factors e.g. OPA1, Apolipoprotein E,
CYP1B1, E-cahderin, OPTC have been reported to ele-
vate the risk of retinal degeneration characteristic of glau-
coma [20,29-32]. However, the interplay of genetic factors
and environmental cues causing POAG pathogenesis
remains almost unexplored.
The association of the immune system with glaucoma
has seemingly conflicting aspects as neuroprotective or
neurodestructive. T-cell-mediated immune response may
initially be beneficial to limit neurodegeneration. How-
ever, a failure to properly control aberrant, stress-induced
immune response likely converts the protective immunity
to an autoimmune neurodegenerative process that can
facilitate the progression of neurodegeneration in some
glaucoma patients specially in Normal Tension Glaucoma
(NTG) cases [8]. Proliferating cytokines, such as IL1 has
been reported [6] to have a role in the immune response
in glaucoma patients. The IL1 induces the expression and
processing of β-amyloid expression protein (APP), result-
ing in the increased production of secreted APP and fur-
ther activation of microglia and overexpression of IL1
[33]. Studies show a higher expression of such microglial
cells in glaucomatous optic nerve heads, which may play a
role in neurodegeneration [8]. IL1 has been shown to
promote optic nerve damage by increasing the synthesis
of matrix mellanoproteinase-9 (MMP) in glaucoma
mouse model, which mimics some aspects of glaucoma
[34,35]. IL1 has also been reported to be involved in
increased generation of reactive oxygen species (ROS)
[36] and nitric oxide synthesis [37], implicated in retinal
ganglion cell damage leading to neurodegeneration.
Previous studies [6,24,25] suggest that single nucleotide
polymorphisms (SNPs) in IL1 gene cluster could influ-
ence glaucoma pathogenesis. Such studies have been car-
ried out so far exclusively in Chinese population groups.
For example, Wang et al [25] reported that the IL1A (-
889T) allele is significantly over-represented in HTG
patients with an IOP > 21 mmHg but not with NTG
cases. Lin et al [24] reported a possible association of the
IL1B (+3953T) polymorphism with POAG in a study car-
ried out in a smaller number of patients which could not
be further replicated in recent studies by Wang et al [27]
in NTG patients and by How et al in POAG [23] patients
(Table 4). Such variation might be due to overall genome
of the population groups under study. However, lack of
information about the ethnicity and interrelationship
between these populations [23-27] limits further analysis
of the data.
We observed weak association of IL1A (-889C) allele
and IL1B (+3953T) allele with non-HTG and HTG
patient groups respectively but these associations, if any,
did not sustain after Bonferroni correction for multiple
tests. However, haplotype analysis based on these 3 SNPs
examined, suggests one of the haplotype (TTC) provides
protection for non-HTG patients above 50 years of age.
This indicates that age also has an effect on this associa-
tion. It is possible that many patients in our cohort with
IOP < 20 mmHg represent NTG but we prefer to describe
those as non-HTG cases in the absence of rigorous moni-
toring of diurnal variations of IOP in all the patients. It
appears that apparent association of the IL1 haplotype
with entire POAG patient group is due to high attribut-
able protection of TTC haplotype only for non-HTG
cases.
It would be a worthwhile effort to examine association
of these haplotypes in other population groups of India. A
Table 3: Age based distribution of haplotypes in non-HTG patients and controls
Haplotype Frequency
in Patients
Frequency
in Controls
p-valueAdjusted
p-value*
OR(95%CI)
AGE GROUP ≤ 50
CTC0.294 (13) 0.420 (131)0.115--
CCC 0.342(15) 0.227 (71)0.100--
TTC0.160 (7)0.144 (45)0.794--
AGE GROUP > 50
CCC0.277 (98)0.253 (66)0.489--
CTC 0.435 (154)0.417 (109)0.637
TTC 0.080 (28) 0.152 (40)0.0040.02 0.477 (0.287-0.792)
*P-values were adjusted by Bonferroni method

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recent study carried out by the Indian Genome Variation
Consortium shows that the Indian population is divided
into 4 distinct linguistic groups - Indo-European, Austro
Asiatic, Tibeto-Burman and Dravidian, and there exists
high degree inter-population variance in allele frequen-
cies due to their genetic diversity [38]. Among the previ-
ous studies in Chinese population, How et al reported
lack of association of the haplotypes with POAG patients
[23]. Lack of required information in other pervious pub-
lications [23-27] prevented us from attempting a meta-
analysis of the data to examine the effect of these haplo-
types on glaucoma pathogenesis.
It has been reported that IL1A (-889T) allele upregu-
lates transcription of the gene and thus increases the level
of the gene product in Alzheimer's disease [39] as well as
the plasma level of IL1B [40]. IL1B (+3953T) allele is
known to increase the level of IL1B production at least by
four fold compared to IL1B (+3953C) allele. It is possible
that IL1A (-889C) allele and IL1B (+3953T) allele influ-
ence the risk of non-HTG and HTG respectively by alter-
ing the expression of the respective proteins [41].
However, since the genomic region represented by spe-
cific haplotype (and not just this SNP) is associated with
non-HTG cases, it appears likely that other yet unidenti-
fied variant might be influencing the course of pathogen-
esis. We plan to explore the question further by fine
mapping the genomic region with additional SNPs and
then finding the suspect variant (if any) by deep sequenc-
ing the minimum critical region. It is also certainly very
important that such studies are replicated in larger and
more importantly in additional glaucoma patient cohorts
to examine whether the IL1 locus has a major effect on
glaucoma pathogenesis. Interestingly, the IL1 gene cluster
(2q13) resides in close proximity to one of the POAG loci
(GLC1B at 2cen-2q13), also linked to NTG cases [42]. It
would be interesting to explore whether there is more
than one genomic region associated with POAG in this
segment of chromosome 2. Finally, understanding the
role of IL 1 in glaucoma pathogenesis and identification
of predisposing variants might pave the way for better
management of the disease through prevention and treat-
ment as appropriate.
Conclusion
Our data suggest that IL1A and IL1B SNPs studied are
not associated with POAG. However, haplotype con-
structed with these SNPs are associated with POAG, spe-
cifically in non-HTG patients above 50 years of age,
which might be due to additional variants located in the
genomic region examined. The observation needs to be
further vindicated by similar studies in larger and addi-
tional cohort of POAG patients.
Competing interests
The authors declare that they have no competing interests.
Table 4: Published reports of IL1 gene cluster polymorphisms in Chinese population group
SNPsPatient (n) Control (n) Results Reference
-889C/T
(IL1A)
156 POAG167 -889T allele is associated with POAGWang et al, Mol Vis, 2006
162 NTG 167 No associationWang et al, J of Glaucoma, 2007
194 POAG
(100 HTG, 94 NTG) and 125 PACG
79 No associationHow et al, IOVS, 2007
-511C/T
(IL1B)
58 POAG105 No associationLin et al, Ophthalmologica, 2003
231 NTG245No associationWang et al, Mol Vis, 2007
194 POAG
(100 HTG, 94 NTG) and 125 PACG
79 No associationHow et al, IOVS, 2007
+3953C/T
(IL1B)
58 POAG105+3953T allele is associated with POAGLin et al, Ophthalmologica, 2003
231 NTG245 No associationWang et al, Mol Vis, 2007
194 POAG
(100 HTG, 94 NTG) and 125 PACG
79 No association How et al, IOVS, 2007

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Authors' contributions
SM, DB, SC and AB carried out the wet lab experiments and also involved in
data analysis. SM, DB has prepared the manuscript. IM gave intellectual inputs
in statistical analysis of the data. AS led the group of ophthalmologist in Dristi
Pradip Eye clinic and selected the patient and control samples used in the
study. KR conceived the study, led the group in designing experimental strate-
gies and provided intellectual input for giving final shape of the manuscript. All
authors read and approved the final manuscript.
Acknowledgements
The authors are thankful to the patients who participated in this study. The
Council of Scientific and Industrial Research, Govt. of India supported the study
through funding grants (MLP-0016 and SIP-007) and pre-doctoral fellowships
to SM, DB, SC. AB is supported by a post-doctoral fellowship from Department
of Biotechnology, India. We are also thankful to the Mr. Kausik Bhattacharya,
who worked as summer trainee in our lab, for his excellent technical support.
Author Details
1Molecular & Human Genetics Division, Indian Institute of Chemical Biology,
Council of Scientific & Industrial Research, Kolkata, India, 2Human Genetics
Unit, Indian Statistical Institute, Kolkata, India and 3Dristi Pradip, Jodhpur Park,
Kolkata, India
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Received: 16 July 2009 Accepted: 19 June 2010
Published: 19 June 2010
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Cite this article as: Mookherjee et al., Association of IL1A and IL1B loci with
primary open angle glaucoma BMC Medical Genetics 2010, 11:99