Get 20M+ Full-Text Papers For Less Than $1.50/day. Start a 14-Day Trial for You or Your Team.

Learn More →

Does the A9285g Polymorphism in Collagen Type XII α1 Gene Associate with the Risk of Anterior Cruciate Ligament Ruptures?

Does the A9285g Polymorphism in Collagen Type XII α1 Gene Associate with the Risk of Anterior... One of the most severe injuries sustained by athletesisruptureoftheanteriorcruciateligament (ACL).RecentinvestigationssuggestthatapredispositionforACLrupturemaybetheresultofspecific geneticsequencevariants.Inlightofthis,wedecided toinvestigatewhethertheCOL12A1A9285GpolymorphismwasassociatedwithACLrupturesinPolish footballplayers. We compared genotypic and allelic frequencies of the COL12A1A9285Gpolymorphismintwo groupsofathletes:91malefootballplayers(23± 3 years)withsurgicallydiagnosedprimaryACLruptureswhoqualifiedforligamentreconstruction(cases)and143apparentlyhealthy,malefootballplayers ofthesameethnicity,asimilaragecategory,anda comparable level of exposure to ACL injury, who were without any self-reported history of ligament ortendoninjury(controls).DNAsamplesextracted fromtheoralepithelialcellsweregenotypedbyus GalenOrthopaedics,Bieru,Poland DepartmentofPhysicalCultureandHealthPromotion,University ofSzczecin,Poland 3 DepartmentofClinicalandMolecularBiochemistry,Pomeranian MedicalUniversity,Szczecin,Poland 4 DepartmentofGenetics,UniversityofSzczecin,Szczecin,Poland 5 BoniFratresCatoviensis,Katowice,Poland 6 DepartmentofSportEducation,AcademyofPhysicalEducationand Sport,Gdask,Poland 7 DepartmentofPhysiology,InstituteofSport,Warsaw,Poland ing a real-time polymerase chain reaction (Ri-TiPCR)method. Thegenotypedistributioninthecaseswerenot different from those in controls (p=0.70).ThefrequencyoftheGallelewaslowerinthecases(18.1%) butnotstatisticallysignificant(p=0.40)whencomparedwithcontrols(21.3%). Ourresultsareincontradictiontothehypothesis that the COL12A1A9285GpolymorphismisassociatedwithapredispositionforACLinjury.However, theseconclusionsshouldbesupportedwithmoreexperimental studies on COL12A1polymorphisms. Keywords: Anterior cruciate ligament (ACL) rupture;Collagen; COL12A1gene;Polymorphism. INTRODUCTION One of the most severe injuries sustained by athletesisruptureoftheanteriorcruciateligament (ACL) [1]. The exact etiology ofACL ruptures is poorly understood, but recently conducted investigationsindicatethataround70.0%ofACLruptures are the consequence of forces applied to the knee atthetimeofinjury,whichresultfromtheathlete's own movements and do not involve contact with another athlete or object [2].That seems to be the reasonthattheriskofACLruptureissignificantly higher in sports requiring change in direction and rapiddecelerationduringcutting,pivotingandlanding [3]. Therefore, it is not surprising that one of thegroupsofathleteswiththehighestfrequencyof ACLrupturearefootballplayers[4]. AnalyzingindividualcasesofACLinjuriesin football players raises the question as to why one memberofateam(withthesameloadandmovement character) is exposed to ACL injury, while fellow team mates are not. Recent investigations suggestthattheriskofACLruptureresultsfromfamilialpredispositionsandspecificgeneticsequence variants[5].Atthispointintime,onlyafewstudieshavegivenevidenceoftheconnectionbetween ACLinjuryandspecificgeneticriskfactors[6-10], mostly among sequence variants within the COL5A1 and COL1A1 genes. Anterior cruciate ligaments are collagenous structuresconsistingofwaterandfibro-cartilaginous specific proteins, which build collagen fibrils [11]. Themainstructuralcomponentsofthisligamentare collagenstypeI,III-VI,XIIandXIV,butalsoproteoglycans such as decorin, lumican and versican, and glycoproteins such as elastin, tenascin C and cartilageoligomericmatrixprotein(COMP)[12]. Inthepresentstateofknowledge,anyproteins (and what is more, any genes that encode these proteins)thatarefunctionallyassociatedwithligamentscouldbepotentialcandidates[13-15].Hence, anysuchgenesandproteinsthathavealreadybeen implicatedwithACLinjuryshouldbedesignatedas candidatesofpriority[7].Inourstudy,wedecided to investigate COL12A1, which is one of the less frequentlystudiedgenesinthecontextofpredispositiontoACLinjury. The COL12A1 gene(121kb;mappedtochromosome 6q12-q13) encodes the 1 chains of the variouslong(XIIA)andshort(XIIB)homotrimeric isoforms of type XII collagen [16,17]. According to the database hosted by the National Center for BiotechnologyandInformation(NCBI),fivesingle nucleotide polymorphisms (SNPs) are shown in COL12A1 exons.Onlytwoofthem(rs240736and rs970547)wereidentifiedasnonsynonymousSNPs (i.e.SNPsthatchangetheaminoacidsequencein thegeneproduct)[13]. Inoneofthepreviousstudies,itwassuggested thatespeciallytheA/Gtransitionatposition162of exon65(9285A/G,S3058G,rs970547)mayalter the biomechanical properties of the collagen fibril andthusmayincreasetheriskofACLruptures[18]. In light of the facts mentioned above, we decided toinvestigatewhethertheCOL12A1A9285GpolymorphismwasassociatedwithACLrupturesinPolishfootballplayers. MATERIALS AND METHODS Study Subjects. A total of 91 male football players(23±3years)withsurgicallydiagnosedprimaryACLruptureswhoqualifiedforligamentreconstruction,wererecruitedforthisstudythrough Galen Orthopaedics, Bieru, Poland. The control groupcomprised143apparentlyhealthy,malefootballplayers(25±2.6years),ofthesameethnicity, a similar age category, and a comparable level of exposuretoACLinjury,whowerewithoutanyselfreportedhistoryofligamentortendoninjury. Ethics Committee.The study was conducted inaccordancewiththeethicalstandardsasdescribed byKruk[19].Additionally,thePomeranianMedicalUniversity(Szczecin,Poland)EthicsCommittee approvedthedetailsofthisstudyandallrelatedinformationalandconsentdocumentationbeforeany datacollection.InaccordancewiththePomeranian MedicalUniversityEthicsCommittee'sguidelines, theinvestigatorinformedallthesubjectsastothe benefits and possible risks associated with participationinthestudy,andallsubjectssignedawritten informedconsentdocumentindicatingtheirvoluntaryparticipation. Genotyping. Genomic DNA was extracted fromtheoralepithelialcellsusingGenEluteMammalianGenomicDNAMiniprepKit(Sigma-Aldrich Chemie, Steinheim, Germany) according to manufacturer's protocol. Allelic discrimination of the A9285G COL12A1 (rs970547) polymorphic site wasperformedusingaTaqManPre-DesignedSNP Genotyping Assays (Applied Biosystems, Foster City,CA,USA),includingprimersandfluorescently labelled(FAMandVIC)MGBprobesforthedetection of the alleles.All samples were genotyped on a Rotor-Gene real-time polymerase chain reaction (Re-Ti-PCR)instrument(CorbettResearch,Sydney, NSW,Australia).Thermalcyclerconditionswereas follows:aninitialstepat95°Cfor5min.,followed by45cyclesofdenaturationat94°C for 15 seconds andannealing/extensionat60°Cfor1min. Statistical Analysis.Anydifferencesingenotype and allele frequency were analyzed using 2 tests(orFisherexacttests).Oddsratios(OR)with 95% confidence intervals (95% CI) were calculated.AllcalculationswereperformedusingStatistica (StatSoft Inc., Tulsa, OK, USA; 2011). STATISTICA (data analysis software system, version 10, BALKAN JOURNAL OF MEDICAL GENETICS Ficek K, Stepien-Slodkowska M, Kaczmarczyk M, Maciejewska-Karlowska A, Sawczuk M, Cholewinski J, Leonska-Duniec A, Zarebska A, Cieszczyk P, Zmijewski P www.statsoft.com) was used for computing statistics,exceptHardy-Weinbergequilibriumwhichwas testedwiththeprogramminglanguageandenvironment R (http://www.r-project.org) and the test for lineartrend,whichwasperformedusingtheSTATCALC module in Epi Info (http://wwwn.cdc.gov/ epiinfo).Thep<0.05valueswereconsideredtobe statisticallysignificant. RESULTS Genotype distributions met Hardy-Weinberg proportionsinthecontrolgroup(p=0.81)andin the cases (p=1.0).ThedistributionsoftheA9285G COL12A1genotypesandallelesaregiveninTable 1.Thegenotypedistributioninthecaseswerenot different from those in controls (p=0.70).Underrepresentation of the GG genotype in the ACL rupturegroupwasnotstatisticallysignificant(p = 0.744, Fisher's exact test, recessive mode: GG vs. GA+AA).ThefrequencyoftheGallelewaslower inthecases(18.1%),butnotstatisticallysignificant (p = 0.40) when compared with controls (21.3%). GiventheGallele,thelikelihoodofACLinjurywas 0.82timeshigher(95%CI:0.50-1.34;p<0.00001; pad<0.0001)thaninthecontrolgroup. DISCUSSION Theroleofgeneticsinsportresearchincreases witheverypassingyear[20,21].Knowledgeofthe roleofindividualgenesintheprocessesoccurring inthehumanbodycanalsobeusedinsportrehabilitationandinjuryprevention[22].Precisedetermination of genotypes at risk for acute or chronic diseases related to sport will probably enable adjustments in individual training plans to greatly minimizetheriskofinjury. Approximately two-thirds ofACL tissue consistsofwater.Therestismadeupoftightlypacked parallelcollagenfibrilbundlesconsistingpredominately of type I collagen fibrils (60.0-80.0% dry massofligament)[11]. CollagenXIIbelongstothesubfamilyoffibrilassociatedcollagenswithinterruptedtriplehelices (FACIT) [23,24] that are believed to form interfibrillar connections and mediate fibril interaction withotherextracellularandcellsurfacemolecules withintendonsandothertissues[24,25].Onthebasis of this fact, we suspect that collagen XII may beconsideredaninfluentialcomponentinligament andtendonstrengthandflexibility. This assumption seems to be indirectly confirmedbythefactthatthekeyelementsoftendons' stretch-responsivenesshavebeenidentifiedinregulatory regions of the COL12A1 gene.Additionally, earlierstudiesindicatedthatcollagenXIIsignificantlypromotesthecontractionofcollagengels(suchas tenascinC)andconsequentlymodulatesthecellular responseoftissuetomechanicalstress[26-28].Furthermore, September et al. [8] suggested collagen XIImaybeinvolvedinsimilarbiologicalprocesses asbothtenascinCandtypeVcollagen,i.e.,regulationoftheassemblyoffibrils(fibrillogenesis). Table 1. GenotypeandallelefrequenciesoftheA9285GCOL12A1gene. Subjects Cases (n=91) Controls (n = 143) HWE 1.0 Genotype n (%) AA:61(67.0) AG:27(29.7) GG:3(3.3) AA:89(62.2) AG:47(32.9) GG:7(4.9) p Values 0.701a pD0.487b pR0.744b Allele (%) A(81.9) G(18.1) A(78.7) G(21.3) OR (95% CI) 0.82 (0.50-1.34) Allele p Value 0.400a HWE:Hardy-Weinbergequilibrium. a b 2:pvalue. pD and pRaretwo-sidedFisher'sexacttestprobabilitieswithdominant(GG+AGvs.AA)andrecessive (GG vs.AG+AA)modesofinheritanceofminoralleles(G),respectively. TheA9285G COL12A1 polymorphism within exon65,isanonsynonymouscodingvariant,which changestheaminoacidatposition3058fromaserinetoaglycine.Althoughthewildtypeserineaminoacidisaneutralpolaraminoacidwithalarger sidechainthanthesubstitutednonpolarneutralglycineaminoacid,someinvestigatorsspeculatethat this change in amino acid sequence may alter the biomechanicalpropertiesofthecollagenfibril[29]. Ontheotherhand,itisnotproventhatthisSNPhas anyeffectonproteinexpressionorfunction. The first report concerning the possible importance of COL12A1variants for achilles tendon injuries did not identify a statistically significant difference in the genotype or allele distribution in theA9285GCOL12A1polymorphism[29].Onthe otherhand,thesameinvestigatorsshowedthatthe A9285G COL12A1polymorphismisassociatedwith ACLrupturesinfemales[29].Theobtainedresults suggestedthatfemaleswithanAAgenotypeareat increased risk forACL ruptures (AA vs. GT+GG; OR=2.4;95%CI1.0-5.5;p<0.05).Additionally, Septemberet al.[13]observedatrendfortheAA genotype to be overrepresented (AA vs. GA+GG; p=0.08)infemaleparticipantswithafamilyhistoryofligamentinjury.Theambiguityoftheresults obtained by September et al. [13] and Posthumus et al. [29] and a lack of other studies concerning theroleA9285G COL12A1 inACLinjuresmaybe considered as one of the most important reasons to conductrepeatedinvestigationsinordertoidentify thegeneticbackgroundofindividualspredisposed totendonandligamentinjury. Ourresultswerecontrarytothehypothesisthat theA9285G COL12A1 polymorphismisassociated withACL injuries. We did not find any statistical difference in theA9285G genotype and allele frequencies in male football players with surgically confirmedprimaryACLrupturescomparedtoinjury-freeathletes.Tosummarize,consideringweonly investigated male subjects, we reached a similar conclusionasSeptemberet al.[13]. In our investigation, the ACL injury group closely resembled the control group of injury-free athletesinnumerousaspects,beingofsimilarages andethnicitiesandidenticalathleticdisciplines.The lastoftheseisakeycomponentduetothevariability of inciting events among different disciplines. Participantsfromthecontrolgroupinourstudyhad thesimilarinternalandexternalriskfactorsrelating to the examined phenotype, overcoming a known limitationofcase-controlstudies. WeinvestigatedmalefootballplayerswithsurgicallyconfirmedprimaryACLrupture,whowere qualifiedforanACLreconstructionprocedure.The controlgroupcomprisedofonlymalesofthesame ethnicity, similar in age, participating in the same sport,theirkneejointsbeingexposedtocomparable forcesandmovements,controllingthemanyinternalandexternalriskfactors.Thankstothissolution inourinvestigationscasesandcontrolsaresimilar in variables that may be related to the phenotype that is under examination, as well as the inciting eventsisverydifficult,aknownlimitationofcasecontrolstudies. The homogeneity of the investigated groups seemstobeastrengthofthisstudy,however,this factmaybeconsideredasalimitation,becausewe did not manage to confirm or deny the important roleoftheinvestigatedpolymorphismwithregard to the risk ofACL injury in women.We included only male participants, but it should be noted that womenare2-3timesmorelikelytosustainanACL injury than men [18]. A number of intrinsic risk factorsclassifiedasanatomical,hormonal,orneuromuscular,havebeenlinkedtothisobservedphenomenon.Inthecaseoffemalepatients,eachofthe aforementionedfactorsmaybemorelikelytoaffect thegeneenvironmentalinteraction,causingsignificantlyhigherincidenceofthispolymorphism'sassociationwithACLinjuries[30,31],arelationship thatisnotseenasclearlyinmen. Inconclusion,thisstudyfoundthatthereisno association between the A9285G COL12A1 polymorphismandACLrupturesinmen.Ontheother hand,thelackofstatisticalsignificanceingenotype andalleledistributionofA9285GCOL12A1 shown inourinvestigation,doesnotnecessarilymeanthat theinvestigatedpolymorphismhasnoeffectonACL injuries.Tendonandligamentinjuriesarecomplex, multifactorialconditions,causedbyinteractionsof anumberofdifferentproteins,encodedbydifferent genes on different chromosomes (gene-gene interactions),andtheinteractionsofthesegeneticcomponentswithdifferentenvironmentalfactors(geneenvironment interactions) [13]. Thus, our findings shouldbesupportedwithmoreexperimentalstudies on COL12A1polymorphismsincludingtheirinter- BALKAN JOURNAL OF MEDICAL GENETICS Ficek K, Stepien-Slodkowska M, Kaczmarczyk M, Maciejewska-Karlowska A, Sawczuk M, Cholewinski J, Leonska-Duniec A, Zarebska A, Cieszczyk P, Zmijewski P action with other genes. Additionally, our results needtobeconfirmedinalargersampleofsubjects. Lastly, it should be noted that genetic association studiesmustalwaysbeinterpretedwithcaution. Declaration of Interest. The authors report no conflictsofinterest.Theauthorsaloneareresponsibleforthecontentandwritingofthisarticle. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Balkan Journal of Medical Genetics de Gruyter

Does the A9285g Polymorphism in Collagen Type XII α1 Gene Associate with the Risk of Anterior Cruciate Ligament Ruptures?

Download PDF
6 pages

Loading next page...
 
/lp/de-gruyter/does-the-a9285g-polymorphism-in-collagen-type-xii-1-gene-associate-U10CiaI65j
Publisher
de Gruyter
Copyright
Copyright © 2014 by the
ISSN
1311-0160
eISSN
1311-0160
DOI
10.2478/bjmg-2014-0022
pmid
25741214
Publisher site
See Article on Publisher Site

Abstract

One of the most severe injuries sustained by athletesisruptureoftheanteriorcruciateligament (ACL).RecentinvestigationssuggestthatapredispositionforACLrupturemaybetheresultofspecific geneticsequencevariants.Inlightofthis,wedecided toinvestigatewhethertheCOL12A1A9285GpolymorphismwasassociatedwithACLrupturesinPolish footballplayers. We compared genotypic and allelic frequencies of the COL12A1A9285Gpolymorphismintwo groupsofathletes:91malefootballplayers(23± 3 years)withsurgicallydiagnosedprimaryACLruptureswhoqualifiedforligamentreconstruction(cases)and143apparentlyhealthy,malefootballplayers ofthesameethnicity,asimilaragecategory,anda comparable level of exposure to ACL injury, who were without any self-reported history of ligament ortendoninjury(controls).DNAsamplesextracted fromtheoralepithelialcellsweregenotypedbyus GalenOrthopaedics,Bieru,Poland DepartmentofPhysicalCultureandHealthPromotion,University ofSzczecin,Poland 3 DepartmentofClinicalandMolecularBiochemistry,Pomeranian MedicalUniversity,Szczecin,Poland 4 DepartmentofGenetics,UniversityofSzczecin,Szczecin,Poland 5 BoniFratresCatoviensis,Katowice,Poland 6 DepartmentofSportEducation,AcademyofPhysicalEducationand Sport,Gdask,Poland 7 DepartmentofPhysiology,InstituteofSport,Warsaw,Poland ing a real-time polymerase chain reaction (Ri-TiPCR)method. Thegenotypedistributioninthecaseswerenot different from those in controls (p=0.70).ThefrequencyoftheGallelewaslowerinthecases(18.1%) butnotstatisticallysignificant(p=0.40)whencomparedwithcontrols(21.3%). Ourresultsareincontradictiontothehypothesis that the COL12A1A9285GpolymorphismisassociatedwithapredispositionforACLinjury.However, theseconclusionsshouldbesupportedwithmoreexperimental studies on COL12A1polymorphisms. Keywords: Anterior cruciate ligament (ACL) rupture;Collagen; COL12A1gene;Polymorphism. INTRODUCTION One of the most severe injuries sustained by athletesisruptureoftheanteriorcruciateligament (ACL) [1]. The exact etiology ofACL ruptures is poorly understood, but recently conducted investigationsindicatethataround70.0%ofACLruptures are the consequence of forces applied to the knee atthetimeofinjury,whichresultfromtheathlete's own movements and do not involve contact with another athlete or object [2].That seems to be the reasonthattheriskofACLruptureissignificantly higher in sports requiring change in direction and rapiddecelerationduringcutting,pivotingandlanding [3]. Therefore, it is not surprising that one of thegroupsofathleteswiththehighestfrequencyof ACLrupturearefootballplayers[4]. AnalyzingindividualcasesofACLinjuriesin football players raises the question as to why one memberofateam(withthesameloadandmovement character) is exposed to ACL injury, while fellow team mates are not. Recent investigations suggestthattheriskofACLruptureresultsfromfamilialpredispositionsandspecificgeneticsequence variants[5].Atthispointintime,onlyafewstudieshavegivenevidenceoftheconnectionbetween ACLinjuryandspecificgeneticriskfactors[6-10], mostly among sequence variants within the COL5A1 and COL1A1 genes. Anterior cruciate ligaments are collagenous structuresconsistingofwaterandfibro-cartilaginous specific proteins, which build collagen fibrils [11]. Themainstructuralcomponentsofthisligamentare collagenstypeI,III-VI,XIIandXIV,butalsoproteoglycans such as decorin, lumican and versican, and glycoproteins such as elastin, tenascin C and cartilageoligomericmatrixprotein(COMP)[12]. Inthepresentstateofknowledge,anyproteins (and what is more, any genes that encode these proteins)thatarefunctionallyassociatedwithligamentscouldbepotentialcandidates[13-15].Hence, anysuchgenesandproteinsthathavealreadybeen implicatedwithACLinjuryshouldbedesignatedas candidatesofpriority[7].Inourstudy,wedecided to investigate COL12A1, which is one of the less frequentlystudiedgenesinthecontextofpredispositiontoACLinjury. The COL12A1 gene(121kb;mappedtochromosome 6q12-q13) encodes the 1 chains of the variouslong(XIIA)andshort(XIIB)homotrimeric isoforms of type XII collagen [16,17]. According to the database hosted by the National Center for BiotechnologyandInformation(NCBI),fivesingle nucleotide polymorphisms (SNPs) are shown in COL12A1 exons.Onlytwoofthem(rs240736and rs970547)wereidentifiedasnonsynonymousSNPs (i.e.SNPsthatchangetheaminoacidsequencein thegeneproduct)[13]. Inoneofthepreviousstudies,itwassuggested thatespeciallytheA/Gtransitionatposition162of exon65(9285A/G,S3058G,rs970547)mayalter the biomechanical properties of the collagen fibril andthusmayincreasetheriskofACLruptures[18]. In light of the facts mentioned above, we decided toinvestigatewhethertheCOL12A1A9285GpolymorphismwasassociatedwithACLrupturesinPolishfootballplayers. MATERIALS AND METHODS Study Subjects. A total of 91 male football players(23±3years)withsurgicallydiagnosedprimaryACLruptureswhoqualifiedforligamentreconstruction,wererecruitedforthisstudythrough Galen Orthopaedics, Bieru, Poland. The control groupcomprised143apparentlyhealthy,malefootballplayers(25±2.6years),ofthesameethnicity, a similar age category, and a comparable level of exposuretoACLinjury,whowerewithoutanyselfreportedhistoryofligamentortendoninjury. Ethics Committee.The study was conducted inaccordancewiththeethicalstandardsasdescribed byKruk[19].Additionally,thePomeranianMedicalUniversity(Szczecin,Poland)EthicsCommittee approvedthedetailsofthisstudyandallrelatedinformationalandconsentdocumentationbeforeany datacollection.InaccordancewiththePomeranian MedicalUniversityEthicsCommittee'sguidelines, theinvestigatorinformedallthesubjectsastothe benefits and possible risks associated with participationinthestudy,andallsubjectssignedawritten informedconsentdocumentindicatingtheirvoluntaryparticipation. Genotyping. Genomic DNA was extracted fromtheoralepithelialcellsusingGenEluteMammalianGenomicDNAMiniprepKit(Sigma-Aldrich Chemie, Steinheim, Germany) according to manufacturer's protocol. Allelic discrimination of the A9285G COL12A1 (rs970547) polymorphic site wasperformedusingaTaqManPre-DesignedSNP Genotyping Assays (Applied Biosystems, Foster City,CA,USA),includingprimersandfluorescently labelled(FAMandVIC)MGBprobesforthedetection of the alleles.All samples were genotyped on a Rotor-Gene real-time polymerase chain reaction (Re-Ti-PCR)instrument(CorbettResearch,Sydney, NSW,Australia).Thermalcyclerconditionswereas follows:aninitialstepat95°Cfor5min.,followed by45cyclesofdenaturationat94°C for 15 seconds andannealing/extensionat60°Cfor1min. Statistical Analysis.Anydifferencesingenotype and allele frequency were analyzed using 2 tests(orFisherexacttests).Oddsratios(OR)with 95% confidence intervals (95% CI) were calculated.AllcalculationswereperformedusingStatistica (StatSoft Inc., Tulsa, OK, USA; 2011). STATISTICA (data analysis software system, version 10, BALKAN JOURNAL OF MEDICAL GENETICS Ficek K, Stepien-Slodkowska M, Kaczmarczyk M, Maciejewska-Karlowska A, Sawczuk M, Cholewinski J, Leonska-Duniec A, Zarebska A, Cieszczyk P, Zmijewski P www.statsoft.com) was used for computing statistics,exceptHardy-Weinbergequilibriumwhichwas testedwiththeprogramminglanguageandenvironment R (http://www.r-project.org) and the test for lineartrend,whichwasperformedusingtheSTATCALC module in Epi Info (http://wwwn.cdc.gov/ epiinfo).Thep<0.05valueswereconsideredtobe statisticallysignificant. RESULTS Genotype distributions met Hardy-Weinberg proportionsinthecontrolgroup(p=0.81)andin the cases (p=1.0).ThedistributionsoftheA9285G COL12A1genotypesandallelesaregiveninTable 1.Thegenotypedistributioninthecaseswerenot different from those in controls (p=0.70).Underrepresentation of the GG genotype in the ACL rupturegroupwasnotstatisticallysignificant(p = 0.744, Fisher's exact test, recessive mode: GG vs. GA+AA).ThefrequencyoftheGallelewaslower inthecases(18.1%),butnotstatisticallysignificant (p = 0.40) when compared with controls (21.3%). GiventheGallele,thelikelihoodofACLinjurywas 0.82timeshigher(95%CI:0.50-1.34;p<0.00001; pad<0.0001)thaninthecontrolgroup. DISCUSSION Theroleofgeneticsinsportresearchincreases witheverypassingyear[20,21].Knowledgeofthe roleofindividualgenesintheprocessesoccurring inthehumanbodycanalsobeusedinsportrehabilitationandinjuryprevention[22].Precisedetermination of genotypes at risk for acute or chronic diseases related to sport will probably enable adjustments in individual training plans to greatly minimizetheriskofinjury. Approximately two-thirds ofACL tissue consistsofwater.Therestismadeupoftightlypacked parallelcollagenfibrilbundlesconsistingpredominately of type I collagen fibrils (60.0-80.0% dry massofligament)[11]. CollagenXIIbelongstothesubfamilyoffibrilassociatedcollagenswithinterruptedtriplehelices (FACIT) [23,24] that are believed to form interfibrillar connections and mediate fibril interaction withotherextracellularandcellsurfacemolecules withintendonsandothertissues[24,25].Onthebasis of this fact, we suspect that collagen XII may beconsideredaninfluentialcomponentinligament andtendonstrengthandflexibility. This assumption seems to be indirectly confirmedbythefactthatthekeyelementsoftendons' stretch-responsivenesshavebeenidentifiedinregulatory regions of the COL12A1 gene.Additionally, earlierstudiesindicatedthatcollagenXIIsignificantlypromotesthecontractionofcollagengels(suchas tenascinC)andconsequentlymodulatesthecellular responseoftissuetomechanicalstress[26-28].Furthermore, September et al. [8] suggested collagen XIImaybeinvolvedinsimilarbiologicalprocesses asbothtenascinCandtypeVcollagen,i.e.,regulationoftheassemblyoffibrils(fibrillogenesis). Table 1. GenotypeandallelefrequenciesoftheA9285GCOL12A1gene. Subjects Cases (n=91) Controls (n = 143) HWE 1.0 Genotype n (%) AA:61(67.0) AG:27(29.7) GG:3(3.3) AA:89(62.2) AG:47(32.9) GG:7(4.9) p Values 0.701a pD0.487b pR0.744b Allele (%) A(81.9) G(18.1) A(78.7) G(21.3) OR (95% CI) 0.82 (0.50-1.34) Allele p Value 0.400a HWE:Hardy-Weinbergequilibrium. a b 2:pvalue. pD and pRaretwo-sidedFisher'sexacttestprobabilitieswithdominant(GG+AGvs.AA)andrecessive (GG vs.AG+AA)modesofinheritanceofminoralleles(G),respectively. TheA9285G COL12A1 polymorphism within exon65,isanonsynonymouscodingvariant,which changestheaminoacidatposition3058fromaserinetoaglycine.Althoughthewildtypeserineaminoacidisaneutralpolaraminoacidwithalarger sidechainthanthesubstitutednonpolarneutralglycineaminoacid,someinvestigatorsspeculatethat this change in amino acid sequence may alter the biomechanicalpropertiesofthecollagenfibril[29]. Ontheotherhand,itisnotproventhatthisSNPhas anyeffectonproteinexpressionorfunction. The first report concerning the possible importance of COL12A1variants for achilles tendon injuries did not identify a statistically significant difference in the genotype or allele distribution in theA9285GCOL12A1polymorphism[29].Onthe otherhand,thesameinvestigatorsshowedthatthe A9285G COL12A1polymorphismisassociatedwith ACLrupturesinfemales[29].Theobtainedresults suggestedthatfemaleswithanAAgenotypeareat increased risk forACL ruptures (AA vs. GT+GG; OR=2.4;95%CI1.0-5.5;p<0.05).Additionally, Septemberet al.[13]observedatrendfortheAA genotype to be overrepresented (AA vs. GA+GG; p=0.08)infemaleparticipantswithafamilyhistoryofligamentinjury.Theambiguityoftheresults obtained by September et al. [13] and Posthumus et al. [29] and a lack of other studies concerning theroleA9285G COL12A1 inACLinjuresmaybe considered as one of the most important reasons to conductrepeatedinvestigationsinordertoidentify thegeneticbackgroundofindividualspredisposed totendonandligamentinjury. Ourresultswerecontrarytothehypothesisthat theA9285G COL12A1 polymorphismisassociated withACL injuries. We did not find any statistical difference in theA9285G genotype and allele frequencies in male football players with surgically confirmedprimaryACLrupturescomparedtoinjury-freeathletes.Tosummarize,consideringweonly investigated male subjects, we reached a similar conclusionasSeptemberet al.[13]. In our investigation, the ACL injury group closely resembled the control group of injury-free athletesinnumerousaspects,beingofsimilarages andethnicitiesandidenticalathleticdisciplines.The lastoftheseisakeycomponentduetothevariability of inciting events among different disciplines. Participantsfromthecontrolgroupinourstudyhad thesimilarinternalandexternalriskfactorsrelating to the examined phenotype, overcoming a known limitationofcase-controlstudies. WeinvestigatedmalefootballplayerswithsurgicallyconfirmedprimaryACLrupture,whowere qualifiedforanACLreconstructionprocedure.The controlgroupcomprisedofonlymalesofthesame ethnicity, similar in age, participating in the same sport,theirkneejointsbeingexposedtocomparable forcesandmovements,controllingthemanyinternalandexternalriskfactors.Thankstothissolution inourinvestigationscasesandcontrolsaresimilar in variables that may be related to the phenotype that is under examination, as well as the inciting eventsisverydifficult,aknownlimitationofcasecontrolstudies. The homogeneity of the investigated groups seemstobeastrengthofthisstudy,however,this factmaybeconsideredasalimitation,becausewe did not manage to confirm or deny the important roleoftheinvestigatedpolymorphismwithregard to the risk ofACL injury in women.We included only male participants, but it should be noted that womenare2-3timesmorelikelytosustainanACL injury than men [18]. A number of intrinsic risk factorsclassifiedasanatomical,hormonal,orneuromuscular,havebeenlinkedtothisobservedphenomenon.Inthecaseoffemalepatients,eachofthe aforementionedfactorsmaybemorelikelytoaffect thegeneenvironmentalinteraction,causingsignificantlyhigherincidenceofthispolymorphism'sassociationwithACLinjuries[30,31],arelationship thatisnotseenasclearlyinmen. Inconclusion,thisstudyfoundthatthereisno association between the A9285G COL12A1 polymorphismandACLrupturesinmen.Ontheother hand,thelackofstatisticalsignificanceingenotype andalleledistributionofA9285GCOL12A1 shown inourinvestigation,doesnotnecessarilymeanthat theinvestigatedpolymorphismhasnoeffectonACL injuries.Tendonandligamentinjuriesarecomplex, multifactorialconditions,causedbyinteractionsof anumberofdifferentproteins,encodedbydifferent genes on different chromosomes (gene-gene interactions),andtheinteractionsofthesegeneticcomponentswithdifferentenvironmentalfactors(geneenvironment interactions) [13]. Thus, our findings shouldbesupportedwithmoreexperimentalstudies on COL12A1polymorphismsincludingtheirinter- BALKAN JOURNAL OF MEDICAL GENETICS Ficek K, Stepien-Slodkowska M, Kaczmarczyk M, Maciejewska-Karlowska A, Sawczuk M, Cholewinski J, Leonska-Duniec A, Zarebska A, Cieszczyk P, Zmijewski P action with other genes. Additionally, our results needtobeconfirmedinalargersampleofsubjects. Lastly, it should be noted that genetic association studiesmustalwaysbeinterpretedwithcaution. Declaration of Interest. The authors report no conflictsofinterest.Theauthorsaloneareresponsibleforthecontentandwritingofthisarticle.

Journal

Balkan Journal of Medical Genetics – de Gruyter

Published: Jun 1, 2014

There are no references for this article.