Transcript ASSOCIATION OF CYTOKINE GENE POLYMORPHISMS WITH …

Single nucleotide polymorphisms in
genes for cytokines interleukin (IL)-2,
IL-6 and TNFalpha influence severity of
osteolysis after total hip arthroplasty
F. Mrazek1, J. Gallo2, A. Arakelyan1, Z. Kubistova1,
M. Petrek1
1 Laboratory
of Immunogenomics, Dept. of Immunology
2 Department of Orthopaedics
Palacky University and University Hospital Olomouc,
Czech Republic
Supported by Czech Govt. Funding IGA MZ NR9490 and MSM 6198959205.
BACKGROUND
Periprosthetic osteolysis (OL) is dominant long-term
complication of the total hip arthroplasty (THA) which can
result in prosthetic failure and re-operation.
Pathogenesis of OL is complex – both biological and
mechanical factors play an important role.
Polyethylene wear particles liberated through the prosthetic
surfaces stimulate an inflammatory response leading to
osteolysis.
Substantial interindividual variability in the severity of OL
suggests implication of genetic factors.
CYTOKINES & OSTEOLYSIS
 cytokines participate in bone resorption pathway
(proliferation, differentiation and function of osteoclasts)
- tumor necrosis factor (TNF)
- interleukin (IL)-1
- macrophage-colony stimulating factor (M-CSF)
- receptor activator of nuclear factor kappa beta ligand
(RANKL)
 production and regulation of cytokines is affected by
particular variants of cytokine genes
 cytokine / cytokine receptor polymorphisms – candidates
for participation in genetic susceptibility to OL.
HYPOTHESIS & AIM
Cytokines
Cytokine gene polymorphisms
???
Periprosthetic Osteolysis
Genetic component
Individual susceptibility
Is there any association between the variants of cytokine
genes and susceptibility to periprosthetic OL/ premature
failure after total hip arthroplasty?
SUBJECTS & DESIGN
205 patients after total hip arthroplasty (THA)
Severe osteolysis*
Mild osteolysis*
Screening Phase
N=55
N=34
N=21
Replication Phase
N=150
N=82
N=68
Healthy subjects
Combined analysis
N=205
N=116
N=89
N=150
*Severe (types III-V) and mild (I+II) osteolysis according to Saleh et al classification,
J Bone Joint Surg Am, 83: 1040, 2001.
METHODS
1. Genotyping of cytokine gene polymorphisms
- 22 selected cytokine gene polymorphisms (Table 1)
- Polymerase Chain Reaction with Sequence Specific
Primers (PCR-SSP)
- „The Cytokine Typing Tray kit“, University of Heidelberg
2. Statistics
- conformity of genotype distribution to the Hardy-Weinberg
equilibrium: Chi-square test
- differences between allelic, genotype and phenotype
(„carriage rate“) frequencies: Chi-square test
- risk assessment: odds ratio, population attributable risk (PAR)
- survival analysis – Kaplan-Meier curves, log-rang test
Table 1: Overview of investigated cytokine
single nucleotide polymorphisms
Cytokine
Gene location
Gene polymorphisms (SNP)
IL-1α
2q14
-889 C/T
IL-1β
2q14
-511 C/T, +3962 C/T
IL-1R
2q12
pstI 1970 C/T
IL-1RA
2q14
Mspa1 11100 T/C
IL-4Ra
16p12
+1902 A/G
IL-12
3q25
-1188 A/C
gIFN
12q14
UTR 5644 A/T
TGFβ
19q13
Codon 10 T/C, Codon 25 G/C
TNFα
6p21
-308 G/A, -238 G/A
IL-2
4q26
-330 T/G, +166 G/T
IL-4
5q31
-1098 T/G, -590 C/T, -33 C/T
IL-6
7p21
-174 G/C, nt 565 G/A
IL-10
1q31
-1082 A/G, -819 C/T, -592 C/A
RESULTS I
Associations of cytokine gene polymorphisms with severity
of osteolysis
The proportion of TNF-238*A and IL-6-174*G allele carriers was
higher in patients with severe OL than in those with mild OL in
both study phases and in combined analysis (Fig. 1, 2).
By contrast, combined analysis revealed that the allele IL-2330*G was protective from severe osteolysis (Fig. 3).
RESULTS II
Combined analysis of THA survival and risk of revision with
cytokine gene variants
Carriage of TNF-238*A allele was associated with worse longterm THA survival (Fig. 4). On the other hand, THA survived
better in patients carrying IL-2-330*G allele (Fig. 5).
Similarly, a risk for THA revision was higher in TNF-238*A allele
carriers (p=0.017) and decreased in patients carrying
IL-2-330*G allele (p=0.02).
Figure 1: Comparison of proportion of TNF-238*A allele
carriers in patients with severe / mild OL and healthy control
subjects (Controls) - combined analysis
0.14
0.13
Allele carriage
0.12
0.10
0.08
0.08
0.06
0.04
0.02
0.02
0.00
Severe OL
Mild OL
Controls
Severe OL versus mild OL: odds ratio = 6.6, p=0.005, PAR%=5.2*
* PAR% - population attributable risk percentage – percentage of the incidence of a disease
in the population that is due to a certain exposure
Figure 2: Comparison of proportion of IL-6-174*G allele
carriers in patients with severe / mild OL and healthy control
subjects (Controls) - combined analysis
0.90
0.85
Allele carriage
0.80
0.83
0.70
0.70
0.60
0.50
0.40
0.30
0.20
0.10
0.00
Severe OL
Mild OL
Controls
Severe OL versus mild OL: odds ratio = 2.5, p=0.007, PAR%=31.5
Figure 3: Comparison of proportion of IL-2-330*G allele
carriers in patients with severe / mild OL and healthy control
subjects (Controls) - combined analysis
0.64
0.70
Allele carriage
0.60
0.56
0.50
0.50
0.40
0.30
0.20
0.10
0.00
Severe OL
Mild OL
Controls
Severe OL versus mild OL: odds ratio = 0.55, p=0.043
Figure 4: Comparison of long-term THA survival between the
carriers (blue curve) and non-carriers (green curve) of the
TNF-238*A allele (log rank test: p=0.022)
Mean survival:
TNF-238*A carriers: 6.7 years
TNF-238*A non carriers: 8.0 years
Figure 5: Comparison of long-term THA survival between the
carriers (blue curve) and non-carriers (green curve) of
IL-2-330*G allele (log rank test: p=0.018)
Mean survival:
IL-2-330*G carriers: 7.8 years
IL-2-330*G non carriers: 6.7 years
CONCLUSIONS
The alleles TNF-238*A and IL-6-174*G may predispose
patients after total hip arthroplasty (THA) to the development
of severe periprosthetic osteolysis (OL).
By contrast, carriage of IL-2-330*G allele appeared to be
protective from severe OL in our group of THA patients.
Furthermore, certain variants of cytokine genes were
associated with worse (TNF-238*A) or better (IL-2-330*G)
overall THA outcome (long-term THA survival and risk for
THA revision).