Association Between Human Leukocyte Antigen Class-I and Hepatitis C: The First Report in Azeri Patients
It has been suggested that host genetic diversity may be associated
with Hepatitis C (HC). However, available data are tremendously heterogeneous
due to the influence of ethnic and geographical differences. This study aimed
to investigate possible association between certain Human Leukocyte Antigen
(HLA) class-I alleles with HC in a group of Azeri patients for the first time
in the literature. In a case-control study, 50 patients with confirmed HC (cases)
and 50 healthy age- and sex-matched counterparts (controls) were evaluated in
Tabriz Sina and Imam Reza Hospitals in a 2-year period of time (2011-2013).
The investigated HLA alleles in the present study were: A2, A3,
B35, B38, BW4, CW4 and CW7.
The A2-positive cases were significantly more frequent in the case
than in the control group (58 vs. 32%, p = 0.01, Odds Ratio (OR) = 2.9). Similar
trend was documented for A3 (62 vs. 26%, p<0.001, OR = 4.6), B35
(24 vs. 2%, p = 0.001, OR = 15.5) and BW4 (78 vs. 46%, p = 0.001,
OR = 4.2). In contrast, the rate of B38-positive (34 vs. 8%, p =
0.001, OR = 0.2) and CW7-positive (38 vs. 14%, p = 0.01, OR = 0.3)
cases was significantly higher in the hepatitis-C-negative subjects. There was
no significant difference in terms of the rate of CW4-positivity
between the two groups (20% in the cases vs. 34% in the controls, p = 0.12,
OR = 0.5). This study showed that there are significant associations between
certain HLA-I alleles with hepatitis C in Azeri patients. While some alleles
make the host prone to the disease, others may have a protective role in this
Received: March 15, 2013;
Accepted: March 20, 2013;
Published: April 25, 2014
Hepatitis C (HC) is major health problem worldwide, particularly in developing
countries in Asia and Africa (Hejazi et al., 2007;
Bidgoli et al., 2007; Hemeida
et al., 2011; Sohail et al., 2011;
Kilic et al., 2012; Barakat
et al., 2012; Abo Elmagd et al., 2011).
The disease is caused by an enveloped blood-borne virus, which is known as the
cause of post-transfusion non-A, non-B hepatitis. It is estimated the there
are approximately 200 million individuals with chronic form of HC in the world.
Although the average prevalence of the infection is rather high, it varies geographically
from 0.1% in industrial countries to about 18% in Africa. The main route of
transmission of HC virus is blood transfusion (Barth et
al., 2006; Zeisel et al., 2009).
The extent of destruction induced by HC virus is determined by two factors;
virus-related parameters such as heterogeneity, viral load and replicative potency
and host-related parameters such as the efficacy of immune response (Chang,
2003). However, different clinical courses in similar patients with a single
source of infection propose a possible role of host genetic factors (Sheehan
et al., 1997). Human Leukocyte Antigen (HLA) is central to the host
immune response and thus, is the nominee for investigating possible role of
genetics in HC (Tripathy et al., 2009). Although
there are numerous reports in this regard in the literature, the results are
widely incongruous, presumably due to ethnic and geographical differences between
studied populations (Wang et al., 2009; De
Almeida et al., 2011). The objective of the present study was to
investigate possible association between certain HLA class-I alleles and hepatitis
C in a group of Azeri patients for the first time.
MATERIALS AND METHODS
In this case-control setting, 50 patients with hepatitis-C (the case group)
and 50 healthy subjects (the control group) were recruited in a study during
a 2-year period of time (2011-2013) in Tabriz Imam Reza and Sina Teaching Hospitals.
The diagnosis of hepatitis-C infection was made by appropriate molecular and
This study was approved by the ethics committee of Tabriz University of the
Medical Sciences. Written consent was obtained from the participants.
The hepatitis-C infection was diagnosed by detection of anti Hepatitis C Virus
(HCV) antibody using Enzyme Linked Immunosorbent Assay (ELISA) technique (Ortho
Clinical diagnostics, Inc. 3rd generation, New Jersey, USA) and confirmed by
immunoblot assay. The same method was employed for rolling out infection in
the control group.
The control group was consisted of 50 age and sex-matched healthy individuals,
who were selected from organ donating volunteers.
Subjects positive for hepatitis-B (positive HBsAg) or Human Immunodeficiency
Virus (HIV) infections were not included. Both patients and controls had normal
liver function tests at the time of enrollment.
The Human Leukocyte Antigens (HLA) evaluated in the present work were as follows;
A2, A3, B35, B38, BW4,
CW4 and CW7. For HLA typing, 10 mL peripheral blood was
taken from the participants and its fibrinogen content was extracted. About
4 mL of this fibrinogen-free blood sample was mixed with 4 mL of Hanks
solution and the new product was mixed gently with ficoll-hypaque solution (density
= 1.077, ratio: 3/5 mL). After being centrifuged, lymphocytes were extracted
and washed with Hanks solution three more times. After being centrifuged
for three times the washed lymphocytes were placed into wells on a commercial
72-well class I typing trays (Biotest, Germany), previously coated with anti-HLA
Class-I antibodies. After 30 min incubating on a shaker, exogenous (rabbit)
complement was added into the plates. After 90 min shaking, eosin and formalin
were added and after 24 h plates were examined by using a standard reverse microscope.
After interpretation of the pattern of reactivity, the HLA typing was finalized
(Ray, 1980). The rate of HLA positivity was compared
between the case and control groups.
Statistical analysis: Data were shown as Mean±standard deviation
or number (%). The SPSS software for Windows (ver.16) was used. Independent
samples t test (for age) and the Chi-square test (for sex and the status of
HLA) were employed for analyzing. The p≤0.05 was considered statistically
Fifty hepatitis-C positive patients, including 35 males (70%) and 15 females
(30%) were compared with fifty hepatitis-C negative individuals, including 33
males (66%) and 17 females (34%). The two groups were comparable in terms of
participants gender (p = 0.69).
The mean age of the patients in the hepatitis-C positive group was 44.7±12.1
(range: 16-66) years vs. the mean age of 43.2±11.2 (range: 18-69) years
in the healthy group.
||Age distributions of the participants in hepatitis-C positive
and negative groups
||Positive human leukocyte antigen class I (HLAI)
in hepatitis-C positive and negative subjects
|Data are shown as frequency (%), p-value = 0.05 is statistically
significant, CI: Confidence interval
Age distribution of the participants in the two studied group is shown in
Fig. 1. The two groups were comparable in terms of their participants
age (p = 0.47).
The rate of HLAI positivity in the two studied groups is summarized
and compared in Table 1. Accordingly, the rate of A2-positive
cases was significantly higher in the patients than in the controls (58 vs.
32%, p = 0.01, Odds Ratio (OR) = 2.9).
Similar finding was documented in terms of A3 (62 vs. 26%, p<0.001,
OR = 4.6), B35 (24 vs. 2%, p = 0.001, OR = 15.5) and BW4 (78 vs.
46%, p = 0.001, OR = 4.2).
In contrast, the rate of B38-positive (34 vs. 8%, p = 0.001, OR
= 0.2) and CW7-positive (38 vs. 14%, p = 0.01, OR = 0.3) cases was
significantly higher in the hepatitis-C-negative group.
There was no significant difference as for the rate of CW4-positivity
between the two studied groups (20% in the cases vs. 34% in the controls, p
= 0.12, OR = 0.5).
It has been suggested that various HLA alleles might be not only associated
with susceptibility of infection with HCV, but also with its course, severity
and extra-hepatic manifestations (Sebastiani et al.,
2005; Hong et al., 2005; Farag
et al., 2013).
In this study, for the first time, a possible association between various types
of HLA-I and HCV infection was investigated in a group of Azeri patients and
normal counterparts. Based on the findings, HLA B35, A3, BW4
and A2 alleles were significantly higher in the patients than in
the controls (OR = 15.5, 4.6, 4.2 and 2.9, respectively). On the other hand,
HLA Cw7 and B37 were against HCV infection (OR = 0.3 and
0.2, respectively). There was no significant difference between the two groups
in term of HLA Cw4.
A possible association between host genetic diversity and HCV infection has
been examined in a number of previous studies. For example, in a study by Tripathy
et al., 2009) among 43 HCV-positive patients and 67 healthy counterparts,
there was a significant association between HLA-I alleles including A3,
A32, B15, B55, Cw16 and Cw18
with higher possibility of HCV infection.
Among the studied alleles in this study, only A3 was also a significant
risk factor for HCV infection in our series. This high rate of heterogeneity
indicates a strong effect of ethnicity in this regard. In addition, it has been
claimed that besides ethnicity, geographical differences also play an important
role in this regard (Wang et al., 2009; De
Almeida et al., 2011).
In another study by Paladino et al. (2007)
in Argentina, it was shown that the presence of HLA-I BW4 is a potent
risk factor for HCV infection. This allele was also considered as a portent
of a worse progression of the infection.
The result of this study is also in conformity with our findings, in which
BW4 was significantly associated with HCV infection.
In a Tunisian study by Ksiaa et al. (2007)
on 99 patients with chronic HCV infection (n = 75) or clearance (n = 24), they
found out that among HLA-I alleles, B35 was more frequently detected
in those with viral clearance than in those with chronic infection (21.7 vs.
In our series, as mentioned before, B35 was the most potent indicator
of HCV infection (OR = 15.5). It is not clear whether this allele may also play
a prognostic role in Azeri patients, as well or not. This should be clarified
in future studies.
In a series in Pakistan by Anis et al. (2007),
there was no significant difference between the patients with HCV infection
and normal controls in terms of HLA-I alleles.
Again, ethnic and geographical differences may justify the contradicting results
of this study comparing with ours.
In a Korean series by Yoon et al. (2005), 137
patients with chronic HCV infection were compared with 206 normal counterparts
for HLA class I molecules. According to their finding, frequency of A3
and B35 was significantly higher in the patients than in the controls.
Surprisingly, both the mentioned alleles were also significantly associated
with HCV infection in our series.
In a study in Ireland, McKiernan et al. (2004)
examined 141 patients with chronic infection and 86 with viral clearance. According
to their findings, HLA-I A3 was associated with clearance (39.5 vs.
The frequency of HLA A3 allele was 62% in our patients, which is
rather higher than both 39.5% and 19.1% in the mentioned study. In addition,
this allele was significantly associated with HCV infection in our series. However,
the prognostic role of this allele was not evaluated in the present work. This
high gap between the rates of expression between the two studies could be attributed
to ethnicity, as mentioned earlier.
In another study in Thailand, Vejbaesya et al.
(2000) did not report a significant difference between A and B alleles of
HLA-I between the patients with HCV infection and the healthy group.
This is in contrast with our findings, emphasizing on different ethnicity and
geographical area of the patients in two separate studies.
In this brief discussion, the reports of various studies regarding a possible
association between HLA and HCV infection are summarized. As underlined earlier,
because of a clear heterogeneity among these studies due to ethnic and geographical
differences, it is essential that every single ethnicity in every geographical
area reports its unique status in this regard. The present work is the first
report from Northwestern Iran with Azeri inhabitants in majority. According
to this report, different HLA alleles may be considered as a foible point, or
in contrast, as a forte for the host against HCV infection. Further studies,
especially with regard to possible association between HLA and the course/outcome
of infection are recommended.
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