Research Article Genetic Variations of Cytokines and

Research ArticleGenetic Variations of Cytokines and Cytokine Receptors inPsoriasis Patients from China

Xiao-Lan Li1 Chun-Feng Wu2 and Gui-Sheng Wu2

1 Department of Dermatology and Rheumatology Affiliated Yanrsquoan Hospital of Kunming Medical CollegeKunming Yunnan 650051 China

2 State Key Laboratory of Phytochemistry and Plant Resources in West China Kunming Institute of BotanyThe Chinese Academy of Sciences Kunming Yunnan 650201 China

Correspondence should be addressed to Xiao-Lan Li lixiaolan96hotmailcom and Gui-Sheng Wu wuguishengmailkibaccn

Received 20 March 2014 Accepted 11 May 2014 Published 25 May 2014

Academic Editor Ji-Fu Wei

Copyright copy 2014 Xiao-Lan Li et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

Psoriasis is a chronic inflammatory and hyperproliferative skin disease affected by both genetic and environmental factors Theaim of the present study was to investigate polymorphisms in a candidate gene family of interleukin (IL) in unrelated Chinesepatients with psoriasis and control subjects without psoriasis In this case-control study 200 unrelated Chinese psoriasis patientsand 298 age- and sex-matched control subjects were enrolled Genomic DNA was prepared from peripheral blood obtained fromall psoriasis patients and control subjects We genotyped seven single-nucleotide polymorphisms (SNPs) in candidate genes ofsix ILs IL4 IL10 IL12B IL13 IL15 and IL23R which have been shown in the literature to be associated with psoriasis in otherethnic groups Among the seven SNPs in the six IL genes studied only the rs3212227 in the IL12B gene was found to be associatedwith psoriasis at genotypic level in the studied population The CC genotype in the IL12B gene is a protective factor of psoriasis(119875 = 00218 OR = 051 95 CI 027ndash096) in Chinese Furthermore the studied Chinese population has extremely low minorallele frequency for IL23R Together the data reveal unique genetic patterns in Chinese that may be in part responsible for thelower risk for psoriasis in this population

1 Introduction

Psoriasis is an immunologically mediated chronic inflam-matory and hyperproliferative skin disease affected by bothgenetic and environmental factors The prevalence of psori-asis varied among populations with different genetic back-grounds and habitats from 3 inNorthern Europe and 2 inNorth America and the UK to 01ndash03 in American Indiansand East Asia [1 2] Psoriasis is proposed to be associatedwith other immune diseases such as arthritis and Crohnrsquosdisease [3] Initial causative research has identified strongassociation between psoriasis and the interleukin genes (IL4IL10 IL12B IL13 and IL23R) in the northern European fromUS and UK [4ndash7] Furthermore the SNP rs56245420 in theIL15 gene has been found to be associatedwith psoriasis in theChinese Han population but not in any of the UK Germanor US Caucasian populations investigated [8ndash10] since theminor allele frequency for this SNP and others across IL15

differs quite strikingly between the populations suggestingheterogeneity in the genetic susceptibility to psoriasis

In this study we aimed to determine if psoriasis isassociatedwith six IL genes that have been strongly associatedwith psoriasis in Europeans but not well studied in ChineseThe seven included SNPs are rs2243250 in the IL4 geners1800872 in the IL10 gene rs3212227 in the IL12B geners1800925 and rs20541 in the IL13 gene rs56245420 in the IL15gene and rs11209026 in the IL23R gene

2 Materials and Methods

21 Study Population A total of 200 psoriasis patients and298 healthy controls were recruited in this study (see Table 2)All participants did not suffer from any other diseases andbelonged to Han nationality in Yunnan Province China Thestudy was performed according to the Helsinki Declaration

Hindawi Publishing CorporationInternational Journal of GenomicsVolume 2014 Article ID 870597 5 pageshttpdxdoiorg1011552014870597

2 International Journal of Genomics

Table1Prim

ersequ

encesPC

Rprod

uctlengthsrestrictio

nendo

nucle

asesand

restric

tionpatte

rnsfor

theP

CR-RFL

Panalyzed

SNPs

SNP

Gene

Positionof

SNP

ingeno

mic

sequ

ence

Forw

ardprim

era

Reversep

rimer

Ann

eal

temperature

(∘C)

Prod

uct

leng

th(bp)

Restr

ictio

nendo

nucle

ases

Fragmentsof

frequ

entallele

geno

type

(bp)

Fragmentsof

heterozygous

geno

type

(bp)

Fragmentsof

rare

allele

geno

type

(bp)

Reference

rs2243250

IL4

132009154CT

TAAAC

TTGGGAG

AAC

ATGGT

TGGGGAAAG

ATAG

AGTA

ATA

49195

AvaII

195

22+173+195

22+173

[1]

rs1800872

IL10

206946

407AC

AGGTG

ATGTA

ATAT

CTCT

GT

TAAAT

ATCC

TCAAAG

TTCC

57303

RsaI

65+238

65+238+303

303

[2]

rs3212227

IL12B

158742950AC

TTCT

ATCT

GAT

TTGCT

TTA

TGAAAC

ATTC

CATA

CATC

C51

233

TaqI

233

68+165+233

68+165

[3]

rs1800

925

IL13

131992809CT

GTC

GCC

TTTT

CCTG

CTCT

TCCC

GC

GGAAT

CCAG

CATG

CCTT

GTG

AGG

65247

Bsh1236I

23+224

23+224+247

247

[4]

rs20541

IL13

13199596

4CT

TAGGCT

GAAG

ACGGGCA

GCA

AAG

AAAC

TTTT

TCGCG

AGGGC C

63199

MspI

22+177

22+177+199

199

[5]

rs56245420

IL15

142873720AT

TTTC

TGTT

ATTA

ACAAAC

ATCA

CTCT

GCA

ACAC

TTGTA

CATA

TTTT

TATT

CAAT A

T54

274

SspI

27+247

27+247+274

274

Thisstu

dya M

ismatch

isshow

nin

bold

andun

derlinedfont

International Journal of Genomics 3

Table 2 Genotyping of seven studied SNPs in psoriasis patients (119899 = 200) and controls (119899 = 298)

SNP Gene Population Genotype () Minor allele ()

rs2243250 IL4TT TC CC C

Controls 189 (634) 98 (329) 11 (37) 120 (201)Psoriasis 127 (635) 61 (305) 12 (60) 85 (213)

rs1800872 IL10AA AC CC C


rs3212227 IL12BAA AC CC C

Controls 119 (399) 128 (430) 51 (171) 230 (386)Psoriasis 77 (385) 104 (520) 19 (95)lowasta 142 (355)

rs1800925 IL13CC CT TT T




rs56245420 IL15AA AT TT T


rs11209026 IL23RGG GA AA A

Controls 298 (100) 0 0 0Psoriasis 200 (100) 0 0 0

lowasta119875 = 00218

with approval of the institutional review boards of the Affil-iated Yanrsquoan Hospital of Kunming Medical College and theKunming Institute of Botany Informed consent was obtainedfrom each participant before inclusion in this study

22 Determination of Genotype Genomic DNAs were iso-lated from whole blood using regular phenolchloroformmethod The SNP rs11209026 in the IL23R gene was geno-typed by the TaqMan allelic discrimination method (AppliedBiosystems) New PCR-RFLP methods were generated togenotype the SNP rs56245420 in the IL15 gene Primers 51015840-TTTCTGTTATTAACAAACATCACTCTG-31015840 and 51015840-CAACAC TTG TAC ATA TTT TTA TTC AAt AT-31015840 (mismatchis shown in bold lower case) were used for rs56245420 Otherfive SNPs were genotyped by PCR-RFLP methods describedpreviously with slight modification [11ndash15] PCR reactionwas carried out in a total volume of 20 120583L containing 20 ngof genomic DNA 1 times PCR buffer 15mMMgCl

2 200120583M

of each dNTP 30 ng of each primer and 1 unit of TaqDNA polymerase (TakaRa) Samples were denatured at 95∘Cfor 2min followed by 30 cycles of 94∘C for 45 sec 61∘C(rs2395029) or 54∘C (rs56245420) for 45 sec and 72∘C for45 sec and endedwith a final extension for 7min at 72∘C PCRproducts were digested with 4U of appropriate restrictionendonuclease and electrophoresed on 3 agarose gels andstained with ethidium bromide The restriction endonucle-ases PCR product lengths and restriction patterns are shownin Table 1

23 Data Analysis Statistics analysis was performed by SPSSsoftware for windows (SPSS Inc) The frequencies of geno-types and alleles for all the six studied loci were determinedassuming codominant inheritance The Hardy-Weinbergequilibrium (HWE) for six loci in psoriasis patients andcontrols was tested by means of chi-square tests The statis-tical significance of the genotype and allele frequency vari-ables between the psoriasis patients and control group wasevaluated by chi-square test with Yates correction for smallnumbers Relative risk associated with the significant geno-type was estimated by the odds ratio (OR) OR with 95confidence intervals (95 CI) was tested using a chi-squaredistribution and the null hypothesis being tested is OR = 1 119875values lt005 were considered as statistically significant

3 Results

Only one of the fourteen Hardy-Weinberg tests (seven poly-morphic loci each in the psoriasis patient and control groups)had 119875 values smaller than 005 (119875 = 001 at rs3212227in controls) All nine remaining genotype frequencies fitHardy-Weinberg expectations according to chi-square testsin psoriasis patients and controls (119875 gt 005) Therefore thereis no meaningful deviation from WHE and our populationis derived from random mating

Polymorphism (minor allele frequency gt 1) has beenfound for all studied SNPs except for rs11209026 in the IL23Rgene (Table 2) Table 2 shows that rs3212227 in the IL12B gene


Table 3 Correlation of psoriasis symptoms with SNPs in the IL genes

SNP Genotype Psoriasis Controls OR (95 CI) Allele Psoriasis Controls OR (95 CI)

rs2243250

TT 127 189 T 315 476TC 61 98 093 (063ndash137) C 85 120 107 (078ndash146)CC 12 11 162 (069ndash378)

TC-CC 73 109 100 (069ndash145)

rs1800872

AA 93 138 A 272 399AC 86 123 104 (071ndash152) C 128 197 095 (072ndash125)CC 21 37 084 (046ndash153)

AC-CC 107 160 099 (069ndash142)

rs3212227

CC 19 51 C 142 230AC 104 128 218 (121ndash392) A 258 366 114 (088ndash148)AA 77 119 171 (096ndash317)

AC-AA 181 247 197 (112ndash345)

rs1800925

CC 140 222 C 332 516CT 52 72 115 (076ndash174) T 68 80 132 (093ndash186)TT 8 4 317 (094ndash1072)

CT-CC 60 76 125 (084ndash186)

rs20541


CT-TT 100 152 096 (067ndash137)

rs56245420

AA 78 139 A 253 413AT 97 135 128 (087ndash187) T 147 183 128 (100ndash171)TT 25 24 185 (099ndash346)

AT-TT 122 159 136 (094ndash196)

(119875 = 00218) was associated with psoriasis at genotypic levelin the studied population Other SNPs examined were notassociated with psoriasis considered from single locus Asshown in Table 3 while the AC genotype (OR = 148 95CI 095ndash230) and the alleles (OR = 084 95 CI 063ndash113)at rs3212227 in the IL12B were not a risk factor of psoriasisthe CC genotype was a protective factor of psoriasis (OR =051 95 CI 027ndash096)

4 Discussion

The etiology of psoriasis is a complex interaction of envi-ronmental and biological factors Genetic factors may playa significant role in the risk of psoriasis in Chinese [10 16]Recent genetic studies indicate that the location of these genesvaries considerably among populations and families We areinterested to know if psoriasis is associatedwith the genes thathave been strongly associated with psoriasis in Europeans butnot well studied in Chinese

Our results showed that the IL12B gene was associatedwith psoriasis in Chinese at genotypic level (119875 lt 005) whichis in line with the findings from European studies [17] TheCC genotype for rs3212227 in IL12B is a protective factor(OR = 051) from psoriasis Similar result from studying SNPrs6887695 in IL12B showed that the minor allele C was aprotective factor from psoriasis [5]

The nonsynonymous SNP in IL23R rs11209026 widelythought to be the primary psoriasis-associated SNP in IL23Rin Europeans was found not to be polymorphic in Chinesewhich is in agreementwith the findings of others [18]The lowfrequencies of variant in IL23R are accordingly of low risk forpsoriasis in ChineseWith single SNP analysis no associationis found between the psoriasis and the IL4 IL10 IL13 andIL15 genes

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

Thisworkwas supported byGrants from theNationalNaturalScience Foundation of China (81360457) and the NaturalScience Foundation of Yunnan Province (2012FB099)

References

[1] J E Gudjonsson and J T Elder ldquoPsoriasis epidemiologyrdquoClinics in Dermatology vol 25 no 6 pp 535ndash546 2007

[2] R Parisi D P Symmons C E Griffiths and D M AshcroftldquoGlobal epidemiology of psoriasis a systematic review of


incidence and prevalencerdquo Journal of Investigative Dermatologyvol 133 no 2 pp 377ndash385 2013

[3] R P Nair T Henseler S Jenisch et al ldquoEvidence for twopsoriasis susceptibility loci (HLA and 17q) and two novelcandidate regions (16q and 20p) by genome-wide scanrdquoHumanMolecular Genetics vol 6 no 8 pp 1349ndash1356 1997

[4] K C Duffin andG G Krueger ldquoGenetic variations in cytokinesand cytokine receptors associated with psoriasis found bygenome-wide associationrdquo Journal of Investigative Dermatologyvol 129 no 4 pp 827ndash833 2009

[5] Y Liu C Helms W Liao et al ldquoA genome-wide associationstudy of psoriasis and psoriatic arthritis identifies new diseaselocirdquo PLoS Genetics vol 4 no 3 2008

[6] R P Nair K C Duffin C Helms et al ldquoGenome-wide scanreveals association of psoriasiswith IL-23 andNF-120581BpathwaysrdquoNature Genetics vol 41 no 2 pp 199ndash204 2009

[7] F Capon M J Bijlmakers N Wolf et al ldquoIdentificationof ZNF313RNF114 as a novel psoriasis susceptibility generdquoHuman Molecular Genetics vol 17 no 13 pp 1938ndash1945 2008

[8] R L Smith S Eyre R BWarrenH S Young C EMGriffithsand J Worthington ldquoNo association between polymorphismsin the interleukin-15 gene and early-onset psoriasis in a UKcohort suggests heterogeneity for this susceptibility locus iden-tified in Chinese psoriasis patientsrdquo Journal of InvestigativeDermatology vol 128 no 12 pp 2904ndash2905 2008

[9] W Weger A Hofer P Wolf et al ldquoRole of the interleukin15 96516AgtT and IL15 96330CgtA gene polymorphisms incaucasian patients with chronic plaque psoriasisrdquo Journal ofDermatological Science vol 51 no 2 pp 147ndash149 2008

[10] X J Zhang K L Yan Z M Wang et al ldquoPolymorphismsin interleukin-15 gene on chromosome 4q312 are associatedwith psoriasis vulgaris in Chinese populationrdquo Journal ofInvestigative Dermatology vol 127 no 11 pp 2544ndash2551 2007

[11] E Tarazona-Santos and S A Tishkoff ldquoDivergent patterns oflinkage disequilibrium and haplotype structure across globalpopulations at the interleukin-13 (IL13) locusrdquo Genes andImmunity vol 6 no 1 pp 53ndash65 2005

[12] P E Graves M Kabesch M Halonen et al ldquoA cluster of seventightly linked polymorphisms in the IL-13 gene is associatedwith total serum IgE levels in three populations of whitechildrenrdquo Journal of Allergy and Clinical Immunology vol 105no 3 pp 506ndash513 2000

[13] T Kawashima E Noguchi T Arinami et al ldquoLinkage andassociation of an interleukin 4 gene polymorphism with atopicdermatitis in Japanese familiesrdquo Journal ofMedical Genetics vol35 no 6 pp 502ndash504 1998

[14] C C Mok J S Lanchbury D W Chan and C S LauldquoInterleukin-10 promoter polymorphisms in Southern Chinesepatients with systemic lupus erythematosusrdquo Arthritis andRheumatology vol 41 no 6 pp 1090ndash1095 1998

[15] A Alvarado-NavarroMMontoya-Buelna J FMunoz-Valle RI Lopez-Roa C Guillen-Vargas and M Fafutis-Morris ldquoThe31015840UTR 1188 AC polymorphism in the interleukin-12p40 gene(IL-12B) is associated with lepromatous leprosy in the West ofMexicordquo Immunology Letters vol 118 no 2 pp 148ndash151 2008

[16] F Long C Sun D Deng X Zhou X P Li and Y P ZhangldquoTNF-238A is associated with juvenile onset psoriasis inpatients of Han population in Southwest Chinardquo Journal ofDermatological Science vol 36 no 2 pp 109ndash111 2004

[17] F Capon S Semprini B Dallapiccola and G Novelli ldquoEvi-dence for interaction between psoriasis-susceptibility loci on

chromosomes 6p21 and 1q21rdquo The American Journal of HumanGenetics vol 65 no 6 pp 1798ndash1800 1999

[18] S I Davidson X Wu Y Liu et al ldquoAssociation of ERAP1but not IL23R with ankylosing spondylitis in a Han ChinesepopulationrdquoArthritis and Rheumatism vol 60 no 11 pp 3263ndash3268 2009

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Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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International Journal of

Volume 2014

Zoology


Molecular Biology International

GenomicsInternational Journal of


The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014


BioinformaticsAdvances in

Marine BiologyJournal of



Signal TransductionJournal of


BioMed Research International

Evolutionary BiologyInternational Journal of



Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014


Genetics Research International


Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational



Enzyme Research



Microbiology


Table1Prim

ersequ

encesPC

Rprod

uctlengthsrestrictio

nendo

nucle

asesand

restric

tionpatte

rnsfor

theP

CR-RFL

Panalyzed

SNPs

SNP

Gene

Positionof

SNP

ingeno

mic

sequ

ence

Forw

ardprim

era

Reversep

rimer

Ann

eal

temperature

(∘C)

Prod

uct

leng

th(bp)

Restr

ictio

nendo

nucle

ases

Fragmentsof

frequ

entallele

geno

type

(bp)

Fragmentsof

heterozygous

geno

type

(bp)

Fragmentsof

rare

allele

geno

type

(bp)

Reference

rs2243250

IL4

132009154CT

TAAAC

TTGGGAG

AAC

ATGGT

TGGGGAAAG

ATAG

AGTA

ATA

49195

AvaII

195

22+173+195

22+173

[1]

rs1800872

IL10

206946

407AC

AGGTG

ATGTA

ATAT

CTCT

GT

TAAAT

ATCC

TCAAAG

TTCC

57303

RsaI

65+238

65+238+303

303

[2]

rs3212227

IL12B

158742950AC

TTCT

ATCT

GAT

TTGCT

TTA

TGAAAC

ATTC

CATA

CATC

C51

233

TaqI

233

68+165+233

68+165

[3]

rs1800

925

IL13

131992809CT

GTC

GCC

TTTT

CCTG

CTCT

TCCC

GC

GGAAT

CCAG

CATG

CCTT

GTG

AGG

65247

Bsh1236I

23+224

23+224+247

247

[4]

rs20541

IL13

13199596

4CT

TAGGCT

GAAG

ACGGGCA

GCA

AAG

AAAC

TTTT

TCGCG

AGGGC C

63199

MspI

22+177

22+177+199

199

[5]

rs56245420

IL15

142873720AT

TTTC

TGTT

ATTA

ACAAAC

ATCA

CTCT

GCA

ACAC

TTGTA

CATA

TTTT

TATT

CAAT A

T54

274

SspI

27+247

27+247+274

274

Thisstu

dya M

ismatch

isshow

nin

bold

andun

derlinedfont


















lowasta119875 = 00218



2 200120583M



3 Results






rs2243250


TC-CC 73 109 100 (069ndash145)

rs1800872


AC-CC 107 160 099 (069ndash142)

rs3212227


AC-AA 181 247 197 (112ndash345)

rs1800925


CT-CC 60 76 125 (084ndash186)

rs20541


CT-TT 100 152 096 (067ndash137)

rs56245420


AT-TT 122 159 136 (094ndash196)


4 Discussion






Acknowledgments


References





























Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology


















lowasta119875 = 00218



2 200120583M



3 Results






rs2243250


TC-CC 73 109 100 (069ndash145)

rs1800872


AC-CC 107 160 099 (069ndash142)

rs3212227


AC-AA 181 247 197 (112ndash345)

rs1800925


CT-CC 60 76 125 (084ndash186)

rs20541


CT-TT 100 152 096 (067ndash137)

rs56245420


AT-TT 122 159 136 (094ndash196)


4 Discussion






Acknowledgments


References





























Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology




rs2243250


TC-CC 73 109 100 (069ndash145)

rs1800872


AC-CC 107 160 099 (069ndash142)

rs3212227


AC-AA 181 247 197 (112ndash345)

rs1800925


CT-CC 60 76 125 (084ndash186)

rs20541


CT-TT 100 152 096 (067ndash137)

rs56245420


AT-TT 122 159 136 (094ndash196)


4 Discussion






Acknowledgments


References





























Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology



























Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology








Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology

Documents

Research Article Genetic Variations of Cytokines and