Lack of Association Between Polymorphism −592A/C in the Promoter Region of the IL10 Gene and Tourette's Syndrome in a Family-Based Association Study in the Chinese Han Population

Abstract

We evaluated the genetic contribution of the IL10−592A/C polymorphism in 108 trios with Tourette's syndrome (TS) including all their parents in a Chinese Han population by using the transmission disequilibrium test and haplotype relative risk design; no evidence for association or linkage disequilibrium was found between IL10−592A/C polymorphism and TS. These results suggested that IL10 may not play a major role in the development of TS in the Chinese Han population.

Introduction

Tourette's syndrome (TS) is an inherited and complex neuro-developmental disorder of childhood that is often associated with various psychiatric morbidities (Jimenez-Shahed, 2009). With rapid development of science and technology and changes of life style, the prevalence and epidemiology of this disorder are more complex than was once thought (Robertson, 2000). Although the exact disease etiology and susceptibility gene remain poorly understood following decades of extensive studies, some evidence suggested that cytokines may play an important role in the pathogenesis of TS. As an important immunoregulatory cytokine, IL10 is involved in vivo in restricting inflammatory responses. In addition, IL10 promotes B-cell proliferation and antibody production (Fuss et al., 2002). As functional polymorphism in the promoter of the IL10 gene, −592A/C was found to be related to IL10 production (Turner et al., 1997) and has been reported to be associated with several diseases, such as schizophrenia, early carotid atherosclerosis, and systemic lupus erythematosus (Chong et al., 2004; Ozbey et al., 2009; Xie et al., 2010).

To test the hypothesis that functional variants of IL10−592A/C may influence disposition to TS development, the present study was conducted on the possible genetic association between this polymorphism and TS in a Chinese Han population.

Material and Methods

Study population

A total of 108 trios in the study were recruited from The Child Healthcare Department of the Affiliated Hospital of Qingdao University Medical College and Linyi People's Hospital. TS cases comprised 25 female and 83 male outpatients, aged between 5 and 18 years. All probands were diagnosed independently by two experienced psychiatrists according to the DSM-IV criteria and the TS Classification Study Group (Freeman et al., 1995). The protocol was approved by the Ethics Committees of Qingdao University Medical College Hospital. Patients gave informed consent. If the subject were a child or disabled person, the informed consent would be issued by the parents or guardians. Subjects were excluded if they presented with unclear diagnosis and incomplete medical record data.

DNA extraction

Five milliliters of blood was drawn from each subject into a vacutainer containing sodium citrate. The genomic DNA was extracted using a commercially available kit (TIANamp Blood DNA kit; Beijing, China) according to standard methods.

Genotyping

The polymorphism at −592A/C in the promoter region of the IL10 gene was amplified by polymerase chain reaction; the polymerase chain reaction product was digested with the restriction enzyme RsaI at 37°C overnight and was electrophoresed in 3% agarose gels, according to Xie et al. (2010). To confirm genotype, several subjects were selected for DNA sequencing techniques.

Statistical analysis

All data analyses were carried out using the Statistical Package for Social Sciences (Version12.0 for Windows; SPSS, Inc., Chicago, IL). For all data of 108 TS trios, the Hardy-Weinberg equilibrium of the genotype distribution was tested using the homogeneity chi-square test. A family-based study was performed to assess genetic association by means of haplotype relative risk (HRR) and transmission disequilibrium test (TDT) statistics. To increase the efficiency of the test, we performed analysis by haplotype-based HRR.

Results

The results showed that allelic and genotypic distribution of IL10−592A/C was not significantly different from the distribution expected according to Hardy-Weinberg equilibrium in TS patients (χ²=2.64 with 1 df and p=0.104) and their parents (χ²=3.27 with 1 df and p=0.07) (Table 1). As we found no excess of transmission of one allele from the heterozygous parents to their affected children using TDT and HRR (TDT=1.301, df=1, p=0.468; HRR=0.722; 95% CI: 0.327—1.594), no evidence for association or linkage disequilibrium was found between IL10−592A/C polymorphism and TS (Tables 2 and 3). To increase the efficiency of the test, we performed analysis by haplotype-based HRR, and still no association was found (χ²=0.189, p>0.05).

Table 1.

Genotype Distribution and Relative Allele Frequencies of IL10−592A/C Gene in Chinese with Tourette's Syndrome Patients (n=108) and Parents

	Genotype frequency (%)			Allele frequency (%)		Hardy-Weinberg
	AA	AC	CC	A	C	χ²	P
Patients	22 (0.20)	44 (0.41)	42 (0.39)	88 (0.41)	128 (0.59)	2.64	0.104
Parents	47 (0.22)	93 (0.43)	76 (0.35)	187 (0.43)	245 (0.57)	3.27	0.07

Table 2.

The Results of Transmission Disequilibrium Test Analysis

	Untransmitted allele
Transmitted allele	A	C	Total
A	a=47	b=41	W=88
C	c=52	d=17	X=128
Total	Y=99	Z=117	2N=432

Transmission disequilibrium test=(b - c)²/(b+c)=1.301, p=0.254.

Table 3.

The Results of Haplotype Relative Risk Analysis

	Genotype of control
Genotype of case	AA, AC	CC	Total
AA, AC	A=33	B=21	W=54
CC	C=37	D=17	X=54
Total	Y=70	Z=38	N=108

χ²=0.649, p=0.420, haplotype relative risk=WZ/XY=0.722, 95% CI: 0.327-1.594.

Discussion

Previous investigation indicated that an infectious or postinfectious (especially streptococcus) process may be an important environmental factor in the pathogenesis of TS patients (Riedel et al., 1998), which suggested poststreptococcal autoimmune abnormalities were implied in this disease. IL10 has important immunosuppressive and antiinflammatory effects on immune responses to both foreign and self-antigens that are primarily mediated by its inhibitory activities on the function of APCs (Moore et al., 2001). Jochen et al. have investigated interleukin 10 receptor 1 variants S138G and G330R in patients and 250 healthy controls for association with TS, but no significant association was found (Kindler et al., 2008). This is in agreement with our study. Our results revealed no significant association between the polymorphisms −592A/C within IL10 with TS, which suggested that IL10 may not play a major role in the development of TS in the Chinese Han population. To our knowledge, this is the first report of investigating an SNP in the IL10 gene in relation to TS by TDT, in which we concluded that the −592A/C was not associated with susceptibility to TS in the Chinese Han population. However, ethnic differences in genetic polymorphisms may determine the functions in different populations. Thus, the relationship between genetic polymorphism and TS will be needed more thorough investigation.

Footnotes

Acknowledgments

We thank all the probands for their participation. This work was supported by the National Basic Research Program of China (2007CB511905), the National Infrastructure Program of Chinese Genetic Resources (2006DKA21300).

Disclosure Statement

No competing financial interests exist.

References

Chong

, Ip

, Wong

et al. 2004. Association of interleukin-10 promoter polymorphisms with systemic lupus erythematosus. Genes Immun, 5:484-492.

Freeman

, Fast

, Kent

. 1995. DSM-IV criteria for Tourette's. J Am Acad Child Adolesc Psychiatry, 34:400-401.

Fuss

, Boirivant

, Lacy

et al. 2002. The interrelated roles of TGF-beta and IL10 in the regulation of experimental colitis. J Immunol, 168:900-908.

Jimenez-Shahed

. 2009. Tourette syndrome. Neurol Clin, 27:737-755.

Kindler

, Schosser

, Stamenkovic

et al. 2008. Tourette's syndrome is not associated with interleukin-10 receptor 1 variants on chromosome 11q23.3. Psychiatry Res, 157:235-239.

Moore

, de Waal Malefyt

, Coffman

et al. 2001. Interleukin-10 and the interleukin-10 receptor. Annu Rev Immunol, 19:683-765.

Ozbey

, Tug

, Namli

. 2009. Interleukin-10 gene promoter polymorphism in patients with schizophrenia in a region of East Turkey. World J Biol Psychiatry, 10:461-468.

Riedel

, Straube

, Schwarz

et al. 1998. Disease presenting as Tourette's disorder. Lancet, 351:418-419.

Robertson

. 2000. Tourette syndrome, associated conditions and the complexities of treatment. Brain, 123:425-462.

10.

Turner

, Williams

, Sankaran

et al. 1997. An investigation of polymorphism in the interleukin-10 gene promoter. Eur J Immunogenet, 24:1-8.

11.

Xie

, Myint

, Zhao

et al. 2010. Relationship between-592A/C polymorphism of interleukin-10 (IL-10) gene and risk of early carotid atherosclerosis. Int J Cardiol, 143:102-104.