Genetic Markers Associated to Dyslipidemia in HIV-Infected

8112019 Genetic Markers Associated to Dyslipidemia in HIV-Infected

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Hindawi Publishing CorporationTe Scienti1047297c World JournalVolume 983090983088983089983091 Article ID 983094983088983096983092983089983093 983089983088 pageshttpdxdoiorg983089983088983089983089983093983093983090983088983089983091983094983088983096983092983089983093

Research ArticleGenetic Markers Associated to Dyslipidemia in HIV-InfectedIndividuals on HAART

Rosmeri K Lazzaretti1 Aline S Gasparotto2 Marina G de M Sassi2 Cariacutesi A Polanczyk13

Regina Kuhmmer1 Jussara M Silveira4 Rossana P Basso4 Cezar A T Pinheiro5

Mariacircngela F Silveira5 Eduardo Sprinz6 and Vanessa S Mattevi2

983089 Servico de Cardiologia Hospital de Cl ınicas de Porto Alegre Universidade Federal do Rio Grande do Sul983097983088983088983091983093-983097983088983091 Porto Alegre RS Brazil

983090 Universidade Federal de Ciencias da Saude de Porto Alegre Rua Sarmento Leite 983090983092983093 Sala 983091983088983097 983097983088983088983093983088-983089983095983088 Porto Alegre RS Brazil 983091 Servico de Cardiologia Hospital de Cl ınicas de Porto Alegre Universidade Federal do Rio Grande do Sul

Instituto de Avaliacao de ecnologias em Saude 983097983088983088983091983093-983097983088983091 Porto Alegre RS Brazil 983092 Hospital Universit ario Dr Miguel Riet Correa Jr 983097983094983090983088983088-983088983088983088 Rio Grande RS Brazil 983093 Servico de Assist encia Especializada em HIVAIDS Universidade Federal de Pelotas 983097983094983088983091983088-983088983088983089 Pelotas RS Brazil 983094 Servico de Medicina Interna Hospital de Cl ınicas de Porto Alegre Universidade Federal do Rio Grande do Sul

983097983088983088983091983093-983097983088983091 Porto Alegre RS Brazil

Correspondence should be addressed to Vanessa S Mattevi vmatteviucspaedubr

Received 983091983089 July 983090983088983089983091 Accepted 983090983096 August 983090983088983089983091

Academic Editors S C Fuchs M B Moreira and B Oyeledun

Copyright copy 983090983088983089983091 Rosmeri K Lazzaretti et al Tis is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

Tisstudy evaluatedthe impact o 983097 single nucleotide polymorphisms (SNPs) in 983094 candidate genes ( APOB APOA983093 APOE APOC983091SCAP and LDLR) over dyslipidemia in HIV-inected patients on stable antiretroviral therapy (AR) with undetectable viral loadsBlood samples were collected rom 983094983089983092 patients at reerence services in the cities o Porto Alegre Pelotas and Rio Grande in BrazilTe SNPs were genotyped by conventional polymerase chain reaction (PCR) and real-time PCR Te prevalence o dyslipidemiawas particularly high among the protease inhibitors-treated patients (983095983097) APOE (rs983092983090983097983091983093983096 and rs983095983092983089983090) genotypes and APOA983093minus983089983089983091983089gtC (rs983094983094983090983095983097983097) were associated with plasma triglycerides (G) and low-density-lipoprotein cholesterol levels (LDL-C) Te APOA983093 minus983089983089983091983089gtC (rs983094983094983090983095983097983097) and SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094) polymorphisms were signi1047297cantly associated with high-density-lipoprotein cholesterol levels Te mean values o the total cholesterol and LDL-C levels were associated with both the APOB SPInsDel (rs983089983095983090983092983088983092983092983089) and APOB XbaI (rs983094983097983091) polymorphisms In conclusion our data support the importance o genetic actors inthe determination o lipid levels in HIV-inected individuals Due to the relatively high number o carriers o these risk variantsstudies to veriy treatment implications o genotyping beore HAAR initiation may be advisable to guide the selection o anappropriate antiretroviral therapy regimen

1 Introduction

Te use o antiretroviral therapy (AR) as a standard o care has changed the prognosis o human immunode1047297ciency

virus (HIV) inection by decreasing mortality and improvingquality o lie [983089 983090] Despite the clinical bene1047297ts long-termAR is associated with a complex spectrum o unwantedmetabolic effects including dyslipidemia that eventually

might lead to increased risk o cardiovascular diseases[983091]

Nevertheless these side effects are not universal to allindividuals on AR and even vary in individuals with com-parable AR demographic immunologic and virologicalcharacteristics Tis variability suggests that host geneticactors and inherited predispositions may have a signi1047297cantin1047298uence on the appearance o metabolic alterations [983092]



983090 Te Scienti1047297c World Journal

Te exact mechanism o dyslipidemia is not ully under-stood but is most likely multiactorial In the general pop-ulation genetic variation accounts or approximately 983092983091ndash983096983091 o the variability in lipid plasma levels [983093] Recentcandidate gene studies [983094ndash983089983089] as well as genome-wide-basedassociation studies have identi1047297ed certain single nucleotide

polymorphisms (SNPs) that could account or a signi1047297cantportion o the variation in blood lipid levels [983089983090ndash983089983092]In HIV inection genetic predisposition may help to

explain the variability among patients with respect to theeffectso protease inhibitors (PIs) on lipid metabolism [983089983088 983089983089]We have hypothesized that this variation is attributable tothe joint effect o HIV inection and AR together with theunderlying genetic predisposition present in these individu-als Te aim o this study was to investigate the requencieso 983097 SNPs in 983094 candidate genes and to identiy associationsbetween these SNPs and the plasma lipid levels o patients onstable AR with undetectable viral loads

2 Methods

983090983089 Subjects We conducted a cross-sectional study with 983094983089983092patients who were diagnosed with HIV-983089 inection accordingto the criteria o the Centers or Disease Control andPrevention [983089983093] All subjects were more than 983089983095 years old hadregularly used AR or at least 983089983090 months had a viral loadbelow the detection limit o the test (983093983088 copiesmL VersantHIV-983089 RNA 983091983088 Assay (bDNA) Siemens Germany) andwere recruited rom three reerral centers in southern Brazil(HIVAIDS Ambulatory Unit o Hospital de Clınicas romPorto AlegreRS HIV Ambulatory Care o Hospital Univer-sitario Dr Miguel Riet Correa Jr rom Rio GrandeRS and

HIVAIDS Specialized Assistance Service rom PelotasRS)rom March 983090983088983088983094 to November 983090983088983088983096 Pregnant women andthose with neurological disease that prevented understandingand proper consent were not included in the study Te study protocol was approved by the Research Ethics Committees o the three centers and o the Universidade Federal de Cienciasda Saude de Porto Alegre and all participants signed aninormed consent statement beore they were included in thestudy (protocol numbers 983088983093983090983097983093 983095983089983096983088983096 983089983093983092983088983095 and 983089983092983089983088983094resp)

983090983090 Study Protocol Te routine evaluation consisted o visitsevery 983092 months in each center or an evaluation by the

patientsrsquo attending physicians as well as laboratory evalua-tions that included measurements o CD983092 cell counts viralload and lipid levels Te patients were invited to participatein the study and had their inormation and a blood sampleor DNA extraction collected during one o these visits

An interview was perormed at enrollment to obtaindemographic and liestyle inormation Details o HIVinection (time rom diagnosis as well as current andprior antiretroviral medications) lipid-lowering interven-tion and relevant clinical variables were obtained rommedical records Te interviewer phenotypically de1047297ned thepatientsrsquo ethnicities because there might be a strong culturaltendency to claim European ancestry in Brazil [983089983094] Patients

were classi1047297ed as Euro- or Aro-descendants because theAmerindian contribution is very low in the Brazilian SouthRegion [983089983095]

983090983091 Laboratory Analysis Blood samples were collected andsent to theMolecular Biology Laboratory orDNA extraction

Lipid pro1047297les included determinations o total cholesterol(C) high-density lipoprotein (HDL-C) triglycerides (G)and when possible low-density lipoprotein (LDL-C) aferasting or 983089983090 hours LDL-C was calculated using the Friede-wald ormula LDL-C = C minus HDL-C minus G983093 i triglyceridelevels were below 983092983088983088 mgdL

Dyslipidemia was de1047297ned by asting triglyceridesplasma levels ge983089983093983088 mgdL andor asting total cholesterolge983090983088983088 mgdL andor LDL-C ge983089983091983088 mgdL andor HDL-Clt983092983088 mgdL Participants were instructed not to perorm any

vigorous physical activity or ingest alcohol in the 983090983092 hoursprior to the blood collection [983089983096]

Genomic DNA was obtained rom peripheral leukocytes

by a standard salting-out technique [983089983097] Te genotypes o apolipoprotein B gene ( APOB) polymorphisms were deter-mined using polymerase chain reaction (PCR) based pro-cedures in 983092983089983088 individuals rom the outpatient clinic o theHospital de Clınicas de Porto Alegre Te polymorphism o insertiondeletion o signal peptide SP InsDel (rs983089983095983090983092983088983092983092983089)was ampli1047297ed by PCR using primers as previously described[983090983088] and directly analyzed by electrophoresis in 983096 polyacry-lamide gels Te XbaI 983095983094983095983091Cgt (rs983094983097983091) polymorphism wasampli1047297ed by PCR using the primers described by Pan et al[983090983089] and genotypes were determined by digestion with XbaIrestriction endonuclease and electrophoresis in 983090 agarosegels

Te SNPs o apolipoprotein A-V ( APOA983093) minus983089983089983091983089gtC(rs983094983094983090983095983097983097) and S983089983097W (983093983094CgtG rs983091983089983091983093983093983088983094) apolipoproteinE( APOE) 983091983091983092gtC (E983092 rs983092983090983097983091983093983096) and 983092983095983090Cgt (E983090 rs983095983092983089983090)apolipoprotein C-III ( APOC983091) 983091983090983091983096CgtG (rs983093983089983090983096) sterol reg-ulatory element-binding actor cleavage-activating protein(SCAP ) 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094) and low-density lipoproteinreceptor (LDLR) intron 983089983097Ggt (rs983094983093983089983089983095983090983088) were genotypedby real-time PCR using the aqMan methodology and arelisted in able 983089 Candidate SNPs were selected via review o PubMed reports o SNP associations with dyslipidemia in thegeneral population or among HIV-inected individuals [983094ndash983089983089]

983090983092 Statistical Analyses Te results are expressed as themean plusmn standard deviation (SD) or continuous variablesand as proportions or categorical variables Variables thatdid not have a normal distribution (triglycerides) weretransormed into natural logarithms beore the statisticaltests were applied Allele requencies were estimated by gene

counting 2 analysis that was used to test or deviations ingenotype requencies rom the Hardy-Weinberg equilibrium(HWE)

For the APOC983091 983091983090983091983096CgtG (rs983093983089983090983096) APOA983093 minus983089983089983091983089gtC(rs983094983094983090983095983097983097) and S983089983097W (983093983094CgtG) (rs983091983089983091983093983093983088983094) SCAP 983090983091983096983094AgtG(rs983089983090983092983096983095983095983091983094) and LDLR intron 983089983097Ggt (rs983094983093983089983089983095983090983088) polymor-phisms the association analyses were perormed according



Te Scienti1047297c World Journal 983091

983137983138983148983141 983089 Genotypic and allelic requencies o polymorphisms analyzed

Polymorphisms Genotypic requency Allelic requency

APOA983093 minus983089983089983091983089gtC (rs983094983094983090983095983097983097)a C

983093983089983093 (983096983092983095) 983088983097983090 983088983088983096C 983097983088 (983089983092983096)

CC 983091 (983088983093)

otal 983094983088983096

APOA983093 S983089983097W (983093983094CgtG rs983091983089983091983093983093983088983094)a C G

CC 983093983088983088 (983096983090) 983088983097983088 983088983089983088

CG 983089983088983091 (983089983094983097)

GG 983095 (983089983089)

otal 983094983089983088

APOB SP InsDel (rs983089983095983090983092983088983092983092983089)a Ins Del

Insins 983089983097983091 (983092983095983091) 983088983095983088 983088983091983088

Insdel 983089983095983094 (983092983091983089)

Deldel 983091983097 (983097983094)otal 983092983088983096

APOB XbaI 983095983094983095983091Cgt (rs983094983097983091)a C

CC 983089983090983090 (983091983088) 983088983093983093 983088983092983093

C 983090983088983091 (983092983097983097)

983096983090 (983090983088983089)

otal 983092983088983095

APOC983091 983091983090983091983096CgtG (rs983093983089983090983096)b C G

CC 983092983095983096 (983095983096983096) 983088983096983097 983088983089983089

CG 983089983090983095 (983090983088983097)

GG 983090 (983088983091)

otal 983094983088983095

APOEa E983090 E983091 E983092E983090E983090 983093 (983088983096) 983088983088983095 983088983095983097 983088983089983092

E983090E983091 983094983095 (983089983089983090)

E983090E983092 983097 (983089983093)

E983091E983091 983091983095983097 (983094983091983094)

E983091E983092 983089983090983088 (983090983088983089)

E983092E983092 983089983094 (983090983095)

otal 983093983097983094

LDLR intron 983089983097Ggt (rs983094983093983089983089983095983090983088)a G

GG 983092983096983095 (983096983088983090) 983088983097983088 983088983089983088

G 983089983089983091 (983089983096983094)

983095 (983089983090)

otal 983094983088983095

SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094)a A G

AA 983089983094983096 (983090983095983095) 983088983093983090 983088983092983096

AG 983090983097983094 (983092983096983097)

GG 983089983092983090 (983090983091983092)

otal 983094983088983094

Te difference in the number o individuals among single nucleotide polymorphisms (SNPs) is due to ailure in genotyping some SNPs in the whole sampleGenotypic requencies presented as number o patients ()a2 test or Hardy-Weinberg equilibrium 1038389 gt 005b

2 test or Hardy-Weinberg equilibrium 1038389 = 0032




983137983138983148983141 983090 Characteristics o the study participants

All study participants1038389 = 614

PI-sparingAR

1038389 = 311

PI-basedAR

1038389 = 303

1103925 value

Demographic

Age years 430 plusmn 10 42 plusmn 10 43 plusmn 9

0114sect

Male sex 1038389 () 983091983092983089 (983093983093983093) 983089983095983097 (983093983096) 983089983094983090 (983093983092) 983088983091983092983096lowast

Ethnicity 1038389 ()

Euro-Brazilians 983091983092983097 (983093983095) 983089983094983094 (983092983096) 983089983096983091 (983094983088) 983088983088983097983092lowast

Aro-Brazilians 983090983094983093 (983092983091) 983089983092983093 (983092983095) 983089983090983088 (983092983088)

Physical activity 1038389 () 983089983093983094 (983090983093) 983095983092 (983090983092) 983096983090 (983090983095) 983088983091983096983097lowast

Cigarette smoking 1038389 () 983089983096983094 (983091983088) 983089983088983091 (983091983091) 983096983091 (983090983096) 983088983089983093983091lowast

Clinical

CD983092 cellsuL 533 plusmn 266 516 plusmn 255 552 plusmn 277 0095sect

Terapy time (months) 68 plusmn 41 57 plusmn 38 76 plusmn 41 lt9830889830889830881sect983205

Lipid-lowering drugs 1038389 () 983089983088983093 (983089983095) 983092983090 (983089983092) 983094983091 (983090983089) 983088983088983090983089lowast

Metabolic

riglycerides mgdL 196 plusmn 171 170 plusmn 131 223 plusmn 202 lt9830889830889830881para983205

otal cholesterol mgdL 192 plusmn 48 189 plusmn 44 196 plusmn 52 0056sect

HDL-C mgdL 48 plusmn 16 51 plusmn 17 46 plusmn 14 lt9830889830889830881sect

LDL-C mgdL 107 plusmn 37 105 plusmn 34 109 plusmn 40 0175sect

Dyslipidemia 1038389 () 983092983093983093 (983095983092) 983090983089983093 (983094983097) 983090983092983088 (983095983097) 983088983088983088983094lowast

Hypertriglyceridemia 983090983097983091 (983092983097) 983089983090983090 (983092983088) 983089983095983089 (983093983097) lt983088983088983088983089lowast

Hypercholesterolemia 983090983092983093 (983092983089) 983089983089983096 (983091983097) 983089983090983095 (983092983092) 983088983090983094983095lowast

Low HDL-C 983089983096983093 (983091983089) 983095983092 (983090983092) 983089983089983089 (983091983096) lt983088983088983088983089lowast

Data presented as mean plusmn SD or number o patients ()PI protease inhibitors AR antiretroviral therapy HDL-C high-density lipoprotein LDL-C low-density lipoprotein9832051038389 value expressed with tests perormed with ln-transormed variablesectStudent 1103925-test or independent samples

lowast2-est with Yates correctionparaStudent 1103925-test or independent samples with ln-transormed variable

to the dominant model due to the low number o individ-uals who were homozygous or the minor allele and werepooled with subjects with the heterozygous genotype As or

APOE SNPs 983091983091983092gtC (rs983092983090983097983091983093983096) and 983092983095983090Cgt (rs983095983092983089983090) whichtogether de1047297ne the APOE E983090 E983091 and E983092 alleles the subjectswere analyzed in 983091 genotype categories E983090E983091 homozygotesor the E983091 allele and E983091E983092 Subjects with the rare genotypesE983090E983090 (1038389 = 5) E983090E983092 (1038389 = 9) and E983092E983092 (1038389 = 1 6) wereexcluded rom the statistical analyses

Te esting Haplotype Effects in Association Studies Pro-gram (version 983091983089 HESIAS Paris France) was used or theanalysis o linkage disequilibrium between polymorphismswithin the same gene [983090983090]

General linear model analyses were used to test or theinteraction between SNPs and variables and to adjust thelipid pro1047297le or covariables Te ollowing variables wereincluded in the models and underwent stepwise removalaccording to the greatest 1103925 values ethnic group genderage physical activity cigarette smoking use o lipid-loweringagents PI use body mass index (BMI) and the presenceo polymorphism Only those variables that were signi1047297cantpredictors were included in the 1047297nal model Correction or

multiple testing was perormed using the Bonerroni methodTe data were analyzed with the Statistical Package or SocialSciences (version 983090983088983088 SPSS Chicago Illinois) Te valueindicating statistical signi1047297cance was 1103925 lt 005

3 Results

983091983089 Study Participants Te main demographic clinical and

metabolic characteristics o the individuals enrolled in thestudy are shown in able 983090 O the 983094983089983092 patients 983093983093983093 weremales 983093983095 were characterized as Euro-descendants and themean patient age was 43 plusmn 10 years Te mean duration o AR was 68 plusmn 41 months Regarding metabolic parameters983090983092983093 (983092983089) patients had hypercholesterolemia 983090983097983091 (983092983097) hadhypertriglyceridemia and 983089983096983093 (983091983089) had low HDL-C levels

Te total patient population was strati1047297ed according tocurrent PI use or nonuse Te prevalence o dyslipidemia wasparticularly high among the PI-treated patients (983095983097 versus983094983097 in nonusers 1103925 = 0006) Although the mean levelso C and LDL-C did not differ signi1047297cantly between thePI-treated and non-PI-treated subjects the latter had higher




983137983138983148983141 983091 Mean metabolic variables according to polymorphisms analyzed

Polymorphisms

otal cholesterol

(mgdL)

LDL-C

(mgdL)

HDL-C

(mgdL)

riglycerides

(mgdL)

907317 M plusmn SD 907317 M plusmn SD 907317 M plusmn SD 907317 M plusmn SD

APOA983093 minus983089983089983091983089gtC (rs983094983094983090983095983097983097)

983092983097983097 191 plusmn 80 983092983093983096 106 plusmn 72 983092983097983096 48 plusmn 88 983092983097983097 187 plusmn 157

C + CC 983097983090 196 plusmn 78 983096983091 107 plusmn 12 983097983090 45 plusmn 16 983097983090 243 plusmn 230

P value 983088983093983092983097 983088983097983093983091 983088983088983092983095983076 983088983088983088983091lowast983203

P value corrected lowastlowast983089983088983088983088 983089983088983088983088 983088983091983095983094 983088983088983090983092

APOA983093 S983089983097W (983093983094CgtG rs983091983089983091983093983093983088983094)

CC 983092983096983094 192 plusmn 49 983092983092983091 106 plusmn 37 983092983096983093 48 plusmn 15 983092983096983094 199 plusmn 177

CG + GG 983089983088983094 197 plusmn 43 983097983097 112 plusmn 35 983089983088983095 50 plusmn 17 983089983088983095 183 plusmn 144

P value 983088983091983091983088 983088983088983092983095

983075

983088983089983091983088 983088983091983097983096

lowast


APOB SP InsDel (rs983089983095983090983092983088983092983092983089)

Insins 983089983097983090 189 plusmn 42a 983089983095983094 99 plusmn 31c 983089983097983090 52 plusmn 15 983089983097983091 195 plusmn 167

Insdel 983089983095983093 193 plusmn 51ab 983089983093983096 106 plusmn 36cd 983089983095983093 50 plusmn 14 983089983095983094 195 plusmn 193

Deldel 983091983097 210 plusmn 45b 983091983095 120 plusmn 41d 983091983097 54 plusmn 13 983091983097 187 plusmn 103

P value 0036sect 0006sect 983088983090983092983093 983088983097983097983094lowast


APOB XbaI 983095983094983095983091Cgt (rs983094983097983091)

CC 983089983090983090 183 plusmn 42e 983089983089983090 95 plusmn 35g 983089983090983090 52 plusmn 15 983089983090983090 187 plusmn 126

C 983090983088983090 194 plusmn 42e 983089983096983093 107 plusmn 31h 983090983088983089 50 plusmn 14 983090983088983091 193 plusmn 163

983096983089 203 plusmn 59

983095983091 110 plusmn 41ih

983096983090 51 plusmn 12 983096983090 212 plusmn 248P value 0007sect 0002sect 983088983095983088983093 983088983095983097983089lowast


APOC983091 983091983090983091983096CgtG (rs983093983089983090983096)



P value 983088983095983089983096 983088983096983091983091 983088983090983097983093 983088983097983088983093lowast


APOE genotype

E983090E983091 983094983092 188 plusmn 60 983093983094 93 plusmn 34 j 983094983091 46 plusmn 14 983094983092 263 plusmn 350l

E983091E983091 983091983094983093 193 plusmn 47 983091983091983091 109 plusmn 37k 983091983094983094 49 plusmn 16 983091983094983094 186 plusmn 128m


P value 983088983090983088983097 0002para 983088983091983095983089 0033lowastsect


LDLR intron 983089983097Ggt (rs983094983093983089983089983095983090983088)

GG 983092983095983093 193 plusmn 48 983092983091983088 107 plusmn 36 983092983095983094 48 plusmn 16 983092983095983091 198 plusmn 184

G + 983089983089983092 191 plusmn 51 983089983088983095 105 plusmn 41 983089983089983091 50 plusmn 16 983089983089983092 183 plusmn 109

P value 983088983094983089983090 983088983092983090983095 983088983089983090983088 983088983097983095983089lowast





983137983138983148983141 983091 Continued

Polymorphisms

otal cholesterol(mgdL)

LDL-C(mgdL)

HDL-C(mgdL)

riglycerides(mgdL)


SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094)

AA 983089983094983092 195 plusmn 49 983089983092983094 107 plusmn 39 983089983094983092 50 plusmn 17n 983089983094983092 201 plusmn 162

AG 983090983096983095 194 plusmn 48 983090983094983091 107 plusmn 35 983090983096983095 49 plusmn 15no 983090983096983095 197 plusmn 197

GG 983089983091983095 188 plusmn 47 983089983090983097 107 plusmn 41 983089983091983095 45 plusmn 14o 983089983091983096 188 plusmn 124

P value 983088983092983092983091 983088983097983095983088 983088983088983091983090983205 983088983097983095983089lowast


HDL-C high-density lipoprotein LDL-C low density lipoproteinlowast1038389 value expressed with tests perormed with logn-transormed variablelowastlowast1038389 value afer B onerroni correction or multiple testingsectAdjusted or gender age and lipid-lowering agents useparaAdjusted or age and lipid-lowering agents use983076Adjusted or gender cigarette smoking and PI use983203Adjusted or gender age lipid-lowering agents use and PI983075Adjusted or gender age ethnic group BMI and lipid-lowering agents use983205Adjusted or gender cigarette smoking ethnic group and lipid-lowering agents useabukey test InsIns versus InsDel 1038389 = 0619 InsIns versus DelDel 1038389 = 0027 InsDel versus DelDel 1038389 = 0145cdukey test InsIns versus InsDel 1038389 = 0231 InsIns versus DelDel 1038389 = 0006 InsDel versus DelDel 1038389 = 0071e ukey test CC versus C 1038389 = 0053 CC versus 1038389 = 0007 C versus 1038389 = 0598ghiukey test CC versus C 1038389 = 0006 CC versus 1038389 = 0008 C versus 1038389 = 0830

jk ukey test E983090E983091 versus E983091E983091 1038389 = 0011 E983090E983091 versus E983091E983092 1038389 = 0005 E983091E983091 versus E983091E983092 1038389 = 0628lmukey test E983090E983091 versus E983091E983091 1038389 = 0003 E983090E983091 versus E983091E983092 1038389 = 0029 E983091E983091 versus E983091E983092 1038389 = 0875noukey test AA versus AG 1038389 = 0594 AA versus GG 1038389 = 0014 AG versus GG 1038389 = 0071

plasma HDL-C levels (51 plusmn 17 mgdL versus 46 plusmn 14 mgdL1103925 lt 0001) and lower triglycerides levels (170 plusmn 131 mgdL

versus 223 plusmn 202 mgdL 1103925 lt 0001) than the PI-treatedsubjects Tereore PI use was tested as a covariate in allstatistical analyses and was included when signi1047297cant

Te genotype and allele requencies o the analyzed SNPsare shown in able 983089 For all studied polymorphisms therewas no departure rom Hardy-Weinberg equilibrium exceptor APOC983091 983091983090983091983096CgtG (rs983093983089983090983096) able 983091 summarizes the asso-ciation analysis o the SNPs with serum lipid concentrations

983091983090 riglycerides wo SNPs contributed signi1047297cantly tothe modi1047297cation o G levels Te plasma G levels weredifferent among APOE genotypes (1103925 = 0033) the E983090 allelebeing associated with increased G levels (ukey test E983090E983091

versus E983091E983091 1103925 = 0003 E983090E983091 versus E983091E983092 1103925 = 0029)

However this result was no longer signi1047297cant afer correctionor multiple testing (1103925corrected = 0132)Te effect o the APOA983093 minus983089983089983091983089gtC (rs983094983094983090983095983097983097) polymor-

phism on plasma lipids was observed in C-allele carrierswho presented higher triglyceride levels than those withthe genotype (243 plusmn 230 mgdL and 187 plusmn 157 mgdLrespectively 1103925 = 0003) Tis nominal 1103925-value remained sta-tistically signi1047297cant afer Bonerroni correction or multiplecomparisons (1103925corrected = 0024)

983091983091 HDL Cholesterol Te APOA983093 minus983089983089983091983089gtC (rs983094983094983090983095983097983097)polymorphism was also signi1047297cantly associated with HDL-C levels Te homozygotes presented higher HDL-C

concentrationsthan C-carriers (1103925 = 0047) afer the same sta-tistical approach and adjustment or covariates Furthermorea statistically signi1047297cant association o the SCAP 983090983091983096983094AgtG(rs983089983090983092983096983095983095983091983094) variant was observed with HDL-C levels (1103925 =0032) As shown in able 983091 the 983090983091983096983094GG homozygotes hadlower HDL-C levels while the 983090983091983096983094AA homozygotes showedan increase o 5 plusmn 2 mgdL when compared to GG homozy-gotes (ukey test AA versus GG 1103925 = 0014) However bothassociations were no longer signi1047297cant afer the Bonerronicorrection or multiple comparisons

983091983092 otal Cholesterol and LDL Cholesterol Initially theaverage C levels were associated with both the APOB SPInsDel (rs983089983095983090983092983088983092983092983089 able 983091 1103925 = 0036) and APOB XbaI983095983094983095983091Cgt (rs983094983097983091 1103925 = 0007) polymorphisms Te post hoc testshowed that there were differences between the homozygotes

or both polymorphisms (ukey test InsIns versus DelDel 1103925 = 0027 and CC versus 1103925 = 0007 resp)Neither results were signi1047297cant afer Bonerroni correctionwas applied (1103925corrected = 0320 and 1103925corrected = 0056 resp)

Te mean LDL-C levels were also different betweenthe genotypes (1103925 = 0007 and 1103925 = 0002 resp) orboth polymorphisms and were statistically signi1047297cant aferBonerroni correction was applied (1103925corrected = 0048 and1103925corrected = 0016 resp) Te ukey test showed that inthe polymorphism o the signal peptide differences wereound between the homozygotes (InsIns versus DelDel 1103925 =0006) and in the polymorphism in exon 983090983094 o the gene thehomozygous group with regard to the C allele differed rom




983137983138983148983141 983092 Effect o APOB Del haplotype in total cholesterol and LDL-C

Risk haplotype C (mgdL) LDL-C (mgdL)

907317 Mean plusmn SD 1103925 value 907317 Mean plusmn SD 1103925-value

Noncarriers 983090983089983091 188 plusmn 42a 983088983088983091983094 983089983097983094 100 plusmn 32b 983088983088983089983089

Heterozygous Del 983089983094983091 195 plusmn 51 983089983092983095 108 plusmn 36

Homozygotes Del 983090983095 212 plusmn 48a

983090983093 118 plusmn 43b

C total cholesterol LDL-C low density lipoproteinResults rom analysis o varianceaukey test noncarriers versus homozygous or Del 1038389 = 0036bukey test noncarriers versus homozygous or Del 1038389 = 0038

the other groups (CC versus C 1103925 = 0006 CC versus 1103925 = 0008)

A signi1047297cant linkage disequilibrium ( = 072 1103925 lt0001) was detected between polymorphisms associatingalleles Del and as well as Ins and C Based on these resultswe analyzed the effect o the haplotype combining alleles Del and on the levels o these lipid parameters Te haplotypeanalysis showed that homozygotes or the risk haplotypeshowed signi1047297cantly higher mean levels o C and LDL-C(able 983092) Te post hoc test showed that the differences wereound between noncarriers and homozygotes or the Del haplotype (ukey test 1103925 = 0036 and 1103925 = 0038 resp)

Te APOE genotype was associated with elevated plasmaLDL-C levels (1103925 = 0002 1103925corrected = 0008 able 983091) Incontrast to what was observed regarding the G levels theE983090 allele was associated with a decrease in the LDL-C levelswhile the E983092 allele was associated with an increase as thedifferences were ound between the E983090 and E983092 allele carriers(ukey test E983090E983091 versus E983091E983091 1103925 = 0011 E983090E983091 versusE983091E983092 1103925 = 0005)

Regarding APOA983093 S983089983097W (983093983094CgtG) (rs983091983089983091983093983093983088983094) there wasa marginal association with LDL-C levels Carriers o theG allele and CC genotype showed a mean and an SD o 112plusmn35 mgdL and 106plusmn37 mgdL respectively (1103925 = 0047)that was adjusted or gender age BMI and use o lipid-lowering agents However it did not remain signi1047297cant aferBonerroni correction

No signi1047297cant contribution to plasma C G HDL-C or LDL-C was identi1047297ed in the present dataset withregard to APOC983091 983091983090983091983096CgtG (rs983093983089983090983096) and LDLR intron 983089983097Ggt(rs983094983093983089983089983095983090983088)

Analyses were also perormed separately or users and

nonusers o IPs (data not shown) and similar effects wereobserved in both groups although in some cases these effectswere not signi1047297cant

4 Discussion

Te contributions o several polymorphisms to dyslipidemiain 983094983089983092 HIV-983089-inected patients on HAAR were addressedin the present multicenter study Nine polymorphisms in983094 candidate genes were analyzed or their association withdyslipidemia As expected according to the literature thisadverse effect was more prevalent in patients receiving PIs

([983090983091] and reerences therein) However we ound no statis-tically signi1047297cant difference in C and LDL-C levels betweenPI-treated and non-PI-treated patients probably associatedwith the more prevalent use o lipid-lowering agents by thePI group (983090983089 versus 983089983092 1103925 = 0021)

Te allelic requencies o the APOB APOA983093 APOESCAP and LDLR genotypes were similar to those previously described in populations rom the same geographical regionor ethnicity [983096ndash983089983089] Te APOC983091 983091983090983091983096CgtG (rs983093983089983090983096) genotypicrequencies were not distributed according to what wasexpected under Hardy-Weinberg equilibrium (1103925 = 0032able 983089) Tis small deviation was most likely due to thelowernumber o homozygotes (983090 patients 983088983091) or the rare alleleobserved in our study 983095 were expected according to Hardy-Weinbergrsquos proportions Nevertheless the allele requencies( = 011) were very close to those ound in previous studieso individuals not inected with HIV who live in SouthernBrazil ( = 010) as shown by Fiegenbaum et al [983096] and toparticipants in the Swiss HIV Cohort Study (SHCS) whichwere = 010 and = 009 as shown by Rotger et al [983089983089]and arr et al [983090983092] respectively

Our study demonstrated a strong association betweenthe E983091E983092 genotype and elevated plasma LDL-C which is inagreementwith the results o previous studies [983090983093 983090983094]TeE983092

variant is typically associated with increased levels o LDL-Candlow plasmaG levels whereas the E983090 variant is associatedwith lower LDL-C levels in the general population [983090983093 983090983094]In a large meta-analysis the E983092 variant was associated witha 983092983090 increase in cardiovascular disease risk [983090983095] AlthoughE983090 isoorms bind to LDL receptors much more weakly thanE983091 or E983092 isoorms the catabolism o the particle containingisoorm E983090 is slower this may result in a lower rate o LDL-

C ormation Moreover according to the conclusion o themeta-analysis o Bennet et al [983090983096] there is an approximately linear relationship between the APOE genotypes and bothLDL-C levels and to the risk o cardiovascular disease

Carriers o non-E983091E983091 genotypes o APOE appear to beat risk o ritonavir-associated hypertriglyceridemia and thisrisk appears to be enhanced by the association with APOC983091

variants [983090983097] According to the SHCSthe interaction between APOE and APOC983091 is associated with an extreme risk o developing hypertriglyceridemia in individuals treated withritonavir [983090983092] Our data did not allow us to perorm thesame analyses as the vast majority o our PI-users wereon ritonavir Nevertheless our 1047297ndings are in agreement




with these data reinorcing the association o E983090 allele andincreased G levels With respect to the E983091E983091 genotypeaccording to our data as well as to those o [983091983088] HIV-positivepatients have a lower risk o developing high triglyceridelevels afer starting HAAR than non-E983091E983091 genotypes

APOA983093 also plays an important role in the modulation

o blood lipid metabolism it is predominantly synthesizedin the liver and is secreted into the plasma where it playsa central role in regulating G metabolism [983091983089 983091983090] wopolymorphisms in the APOA983093 gene minus983089983089983091983089gtC and S983089983097W(983093983094CgtG) have been shown to be associated with elevatedtriglyceride levels in different populations [983091983089 983091983091] In ourstudy or APOA983093 minus983089983089983091983089gtC individuals with at least oneC allele had higher G levels and lower levels o HDL-C However only the association with G levels remainedsigni1047297cant afer the Bonerroni correction was applied Teseresults are consistent with previous study in HIV-inectedpatients indicated that minus983089983089983091983089C carriers experienced markedincreases in triglyceride levels during a 983091-year ollow-upwhile no change was recorded in patients carrying the

minus983089983089983091983089

normal allele [983091983092] A similar effect but o lesser magnitudewas demonstrated regarding changes in cholesterol [983091983092]

o our knowledge this is the 1047297rst study conducted inan HIV-positive setting to assess the association o SCAP 983090983091983096983094AgtG rs983089983090983092983096983095983095983091983094 with dyslipidemia Te SCAP pathway controls cellular cholesterol homeostasis Initially in ouranalysis we ound that AA homozygous patients showedhigher HDL-C levels however this result was no longersigni1047297cant afer Bonerroni correction was applied Carrierso the 983090983091983096983094G allele rom non-HIV-inected Brazilians treatedwith simvastatin exhibit reduced C and G levels as shownby Fiegenbaum et al [983095] but no association was ound withHDL-C levels

Te mean C and LDL-C levels showed statistically sig-ni1047297cant differences or genotypes o both polymorphisms inthe APOB gene SP InsDel (rs983089983095983090983092983088983092983092983089) and XbaI 983095983094983095983091Cgt(rs983094983097983091) Te homozygotes or the alleles and Del had higherlevels o these lipids than individuals heterozygous or bothalleles who had intermediate levels this result is compatiblewith a co-dominant effect o these polymorphisms Teseresults are in agreement with published data showing thatthe Del and alleles are associated with increased levelso C andor LDL-C in different populations with distinctdiseases [983097 983091983093ndash983091983095] However Xu et al and Ye et al ound noassociation o the polymorphism in the signal peptide o thegene with C and LDL-C levels in the Finnish and Chinese

populations respectively [983091983096 983091983097]Te signi1047297cant linkage disequilibrium observed between

polymorphisms as reported in other studies [983090983088 983091983097] led toan analysis o risk haplotype (patients with a combination o alleles Del and ) Te patients who were homozygous orthe risk haplotype showed higher total cholesterol and LDL-C while an intermediate effect was observed in patients withthe heterozygous haplotype Rios et al [983097] also observed theassociation o the haplotype with LDL-C levels in a Brazilianpopulation in a comparison o patients with and withoutcoronary artery disease

Tere are two possible causes or these changes in lipidmetabolism related to both polymorphisms in the APOB

gene Te 1047297rst cause is that three amino acids leu-ala-leu thatare included in the allele Ins anddeleted in the allele Del couldalter the hydrophobicity o the signal peptide o the proteinand thereby alter the rate o translocation o new APOBpeptides synthesized in the cytoplasm to the endoplasmicreticulum [983091983096] Te second hypothesis is based on the act

that there is no amino acid change in the unctional proteinin both polymorphisms Tese could then be in linkagedisequilibrium with an unknown change in the DNA thatmay cause lipid abnormalities [983090983088]

wo urther SNPs that were proposed in the litera-ture ( APOC983091 983091983090983091983096CgtG (rs983093983089983090983096) and LDLR intron 983089983097Ggt(rs983094983093983089983089983095983090983088) did not contribute to the plasma lipid levels inthe present dataset which may re1047298ect a limited effect o theseSNPs in HIV-inected patients

All analyses were also perormed by stratiying thepatients according to PI use (data not shown) however sim-ilar effects were observed in both groups Furthermore the

variables were not statistically signi1047297cant in some analysesand this was most likely attributable to the reduction in thenumber o individuals when they are analyzed separatelyTis 1047297nding suggests that in our sample the lipid-geneinteraction is independent o the type o HAAR used andthat the variation observed is similar to that ound in thegeneral population

We acknowledge some limitations o the present studyTe cross-sectional design and the inclusion o patientsreceiving several different antiretroviral regimens which isa difficulty inherent to any study involving current HAARas by de1047297nition should include at least three different drugsTis act might hinder the accomplishment o any pharma-cogenomics study regarding this therapy Furthermore weound a deviation rom the HWE observed in the currentpopulation or the APOC983091 983091983090983091983096CgtG (rs983093983089983090983096) SNP Althoughthis deviation may indicate genotyping error laboratory procedures were in place to detect errors including blindedno-template controls and DNA sample replicates Anotherlimitation that we might acknowledge is the restricted num-ber o SNPs analyzed We cannot rule out the possibility thatother SNPs in the genes investigated here are associated withthe phenotypes studied

In conclusion our data support the importance o geneticactors on o lipid levels in HIV-inected individuals rom apreviously uninvestigated region o the world We ound noevidence o interaction o these genetic variants with the useo non-nucleoside transcriptase reverse inhibitors or protease

inhibitors Due to the relatively high number o carriers o these risk variants (eg APOA983093 minus983089983089983091983089gtC = 983089983093 APOB risk haplotype = 983092983095 APOE = 983090983088983089) studies to veriy treatmentimplications o genotyping are desirable

Authorsrsquo Contribution

Eduardo Sprinz and Vanessa S Mattevi were joint seniorauthors on this work Aline S Gasparotto and Marina Gde M Sassi genotyped the patients and perormed theexperiments Rosmeri K Lazzaretti Eduardo Sprinz ReginaKuhmmer Jussara M Silveira Rossana P Basso Cezar




A Pinheiro and Mariangela F Silveira were responsibleor recruiting and clinical evaluation o patients VanessaS Mattevi conceived and designed the experiments CarısiA Polanczyk reviewed the paper Rosmeri K Lazzarettiand Vanessa S Mattevi perormed the statistical analysesRosmeri K Lazzaretti Vanessa S Mattevi and Eduardo

Sprinz wrote the paper

Disclosure

Te number o contributors is justi1047297ed by the perormance o sample collections and patientsrsquo evaluations in three differentreerence centers

Conflict of Interests

Te authors declare that there is no con1047298ict o interests

Acknowledgments

A special acknowledgement in memoriam o Pro Jorge PintoRibeiro an example o scientist and educator He will besadly missed but his enduring and substantial legacy willbe intensely alive Te authors thank Grasiela Agnes orhelping with qPCR experiments Tis work was supported by Departamento de DS AIDS e Hepatites Virais Ministerioda Saude Brazil Conselho Nacional de DesenvolvimentoCientı1047297co e ecnologico (CNPq) Fundacao de Amparo aPesquisa do Estado do Rio Grande do Sul (FAPERGS) andPRONEX-FAPERGSCNPq

References[983089] B O aiwo X Li F Palella et al ldquoHigher risk o AIDS or death

in patients with lower CD983092 cell counts afer virally suppressiveHAARrdquo HIV Medicine vol 983089983088 no 983089983088 pp 983094983093983095ndash983094983094983088 983090983088983088983097

[983090] J R P Marins L F Jamal S Y Chen et al ldquoDramaticimprovement in survival among adult Brazilian AIDS patientsrdquo AIDS vol 983089983095 no 983089983089 pp 983089983094983095983093ndash983089983094983096983090 983090983088983088983091

[983091] M P Dube J H Stein J A Aberg et al ldquoGuidelines or theevaluation and management o dyslipidemia in human immun-ode1047297ciency virus (HIV)-inected adults receiving antiretroviraltherapy recommendations o the HIV Medicine Association o the Inectious Disease Society o America and the Adult AIDSClinical rials Grouprdquo Clinical Infectious Diseases vol 983091983095 no 983093pp 983094983089983091ndash983094983090983095 983090983088983088983091

[983092] J Fox M Boffito and A Winston ldquoTe clinical implicationso antiretroviral pharmacogenomicsrdquo Pharmacogenomics vol983095 no 983092 pp 983093983096983095ndash983093983097983094 983090983088983088983094

[983093] M-H Chang A Yesupriya R M Ned P W Mueller and N FDowling ldquoGenetic variants associated with asting blood lipidsin the US population third National Health and NutritionExamination Surveyrdquo BMC Medical Genetics vol 983089983089 no 983089article 983094983090 983090983088983089983088

[983094] F M de Andrade S W Malu JB Schuch et al ldquoTein1047298uenceo the S983089983097W SNP o the APOA983093 gene on triglyceride levelsin southern Brazil interactions with the APOE gene sex andmenopause statusrdquo Nutrition Metabolism and Cardiovascular Diseases vol 983090983089 no 983096 pp 983093983096983092ndash983093983097983088 983090983088983089983089

[983095] M Fiegenbaum F R Silveira C R van der Sand et alldquoDeterminants o variable response to simvastatin treatmentthe role o common variants o SCAP SREBF-983089a and SREBF-983090 genesrdquo Pharmacogenomics Journal vol 983093 no 983094 pp 983091983093983097ndash983091983094983092983090983088983088983093

[983096] M Fiegenbaum F M de Andrade and M H Hutz ldquoAssociationbetween plasma lipid parameters and APOC983091 genotypes inBrazilian subjects effect o gender smoking and APOE geno-typesrdquo Clinica Chimica Acta vol 983091983096983088 no 983089-983090 pp 983089983095983093ndash983089983096983089 983090983088983088983095

[983097] D L S Rios A F Vargas M R orres A J Zago SM Callegari-Jacques and M H Hutz ldquoInteraction betweenSREBP-983089a and APOB polymorphisms in1047298uences total and low-density lipoprotein cholesterol levels in patients with coronary artery diseaserdquo Clinical Genetics vol 983094983091 no 983093 pp 983091983096983088ndash983091983096983093983090983088983088983091

[983089983088] M Arnedo P aff e R Sahli et al ldquoContribution o 983090983088single nucleotide polymorphisms o 983089983091 genes to dyslipidemiaassociated with antiretroviral therapyrdquo Pharmacogenetics and Genomics vol 983089983095 no 983097 pp 983095983093983093ndash983095983094983092 983090983088983088983095

[983089983089] M Rotger C Bayard P aff e et al ldquoContribution o genome-wide signi1047297cant single-nucleotide polymorphisms andantiretroviral therapy to dyslipidemia in HIV-inected individ-uals a longitudinal studyrdquo Circulation vol 983090 no 983094 pp 983094983090983089ndash983094983090983096983090983088983088983097

[983089983090] S Kathiresan C J Willer G M Peloso et al ldquoCommon variants at 983091983088 loci contribute to polygenic dyslipidemiardquo NatureGenetics vol 983092983089 no 983089 pp 983093983094ndash983094983093 983090983088983088983097

[983089983091] M S Sandhu D M Waterworth S L Debenham et al ldquoLDL-cholesterol concentrations a genome-wide association studyrdquoTe Lancet vol 983091983095983089 no 983097983094983089983089 pp 983092983096983091ndash983092983097983089 983090983088983088983096

[983089983092] C Wallace S J Newhouse P Braund et al ldquoGenome-wideassociation study identi1047297es genes or biomarkers o cardiovas-cular disease serum urate and dyslipidemiardquo American Journal of Human Genetics vol 983096983090 no 983089 pp 983089983091983097ndash983089983092983097 983090983088983088983096

[983089983093] E Schneider S Whitmore K M Glynn K DominguezA Mitsch and M McKenna ldquoRevised surveillance casede1047297nitions or HIV inection among adults adolescents andchildren aged lt983089983096 months and or HIV inection and AIDSamong children aged 983089983096 months to lt983089983091 yearsndashUnited States983090983088983088983096rdquo Morbidity and Mortality Weekly Report vol 983093983095 no 983089983088pp 983089ndash983089983090 983090983088983088983096

[983089983094] V M Zembrzuski S M Callegari-Jacques and M H HutzldquoApplication o an Arican ancestry index as a genomic controlapproach in a Brazilian populationrdquo Annals of Human Genetics vol 983095983088 no 983094 pp 983096983090983090ndash983096983090983096 983090983088983088983094

[983089983095] N P C Santos E M Ribeiro-Rodrigues A K C Ribeiro-dos-Santos et al ldquoAssessing individual interethnic admixture andpopulation substructure using a 983092983096-insertion-deletion (INSEL)

ancestry-inormative marker (AIM) panelrdquo Human Mutation vol 983091983089 no 983090 pp 983089983096983092ndash983089983097983088 983090983088983089983088

[983089983096] Expert Panel on Detection Evaluation and reatment o HighBlood Cholesterol in Adults ldquoExecutive summary o the thirdreport o the National Cholesterol Education Program (NCEP)expert panel on detection evaluation and treatment o highblood cholesterol in adults (adult treatment panel III)rdquo Journal of the American Medical Association vol 983090983096983093 no 983089983097 pp 983090983092983096983094ndash983090983092983097983095 983090983088983088983089

[983089983097] D K Lahiri and J L Nurnberger Jr ldquoA rapid non-enzymaticmethodor thepreparation oHMW DNA rom bloodor RFLPstudiesrdquo Nucleic Acids Research vol 983089983097 no 983089983097 p 983093983092983092983092 983089983097983097983089

[983090983088] P E PajukantaL M ValstaA Aro P Pietinen HelioandMJ ikkanen ldquoTe effects o the apolipoprotein B signal peptide




(insdel) and XbaI polymorphisms on plasma lipid responses todietary changerdquo Atherosclerosis vol 983089983090983090 no 983089 pp 983089ndash983089983088 983089983097983097983094

[983090983089] J-P Pan A-N Chiang J J ai S-P Wang and M-S ChangldquoRestriction ragment length polymorphisms o apolipoproteinB gene in Chinese population with coronary heart diseaserdquoClinical Chemistry vol 983092983089 no 983091 pp 983092983090983092ndash983092983090983097 983089983097983097983093

[983090983090] D A regouet and V Garelle ldquoA new JAVA interace imple-mentation o HESIAS testing haplotype effects in associationstudiesrdquo Bioinformatics vol 983090983091 no 983096 pp 983089983088983091983096ndash983089983088983091983097 983090983088983088983095

[983090983091] E R Feeney andP W G Mallon ldquoHIV and HAAR-associateddyslipidemiardquo Te Open Cardiovascular Medicine Journal vol983093 pp 983092983097ndash983094983091 983090983088983089983089

[983090983092] P E arr P affe G Bleiber et al ldquoModeling the in1047298uenceo APOC983091 APOE and NF polymorphisms on the risk o antiretroviral therapymdashassociated lipid disordersrdquo Journal of Infectious Diseases vol 983089983097983089 pp 983089983092983089983097ndash983089983092983090983094 983090983088983088983093

[983090983093] J M Ordovas J Lopez-Miranda F Perez-Jimenez et al ldquoEffecto apolipoprotein E and A-IV phenotypes on the low density lipoprotein responseto HMG CoA reductaseinhibitor therapyrdquo Atherosclerosis vol 983089983089983091 no 983090 pp 983089983093983095ndash983089983094983094 983089983097983097983093

[983090983094] J Pedro-Botet E J Schaeer R G Bakker-Arkema et alldquoApolipoprotein E genotype affects plasma lipid response toatorvastatin in a gender speci1047297c mannerrdquo Atherosclerosis vol983089983093983096 no 983089 pp 983089983096983091ndash983089983097983091 983090983088983088983089

[983090983095] R W Mahley and S C Rall Jr ldquoApolipoprotein E ar morethan a lipid transport proteinrdquo Annual Review of Genomics and Human Genetics vol 983089 no 983090983088983088983088 pp 983093983088983095ndash983093983091983095 983090983088983088983088

[983090983096] A M Bennet E di Angelantonio Z Ye et al ldquoAssociation o apolipoprotein e genotypes with lipid levels and coronary riskrdquo Journal of the American Medical Association vol 983090983097983096 no 983089983089 pp983089983091983088983088ndash983089983091983089983089 983090983088983088983095

[983090983097] J Fauvel E Bonnet J-B Ruidavets et al ldquoAn interactionbetween apo C-III variants and protease inhibitors contributesto high triglyceridelow HDL levels in treated HIV patientsrdquo

AIDS vol 983089983093 no 983089983096 pp 983090983091983097983095ndash983090983092983088983094 983090983088983088983089

[983091983088] A Marzocchetti J Schwarz S di Giambenedetto et al ldquoTeeffect o polymorphisms in candidate genes on the long-termrisk o lipodystrophy and dyslipidemia in HIV-inected whitepatients starting antiretroviral therapyrdquo AIDS Research and Human Retroviruses vol 983090983095 no 983089983090 pp 983089983090983097983097ndash983089983091983088983097 983090983088983089983089

[983091983089] O Seda and L Sedova ldquoNew Apolipoprotein A-V comparativegenomics meets metabolismrdquo Physiological Research vol983093983090pp983089983092983089ndash983089983092983094 983090983088983088983091

[983091983090] E S ai andJ M Ordovas ldquoClinical signi1047297cance o apolipopro-tein A983093rdquo Current Opinion in Lipidology vol 983089983097 no 983092 pp 983091983092983097ndash983091983093983092 983090983088983088983096

[983091983091] C-Q Lai E-S ai C E an et al ldquoTeAPOA983093 locus is a strong

determinant o plasma triglyceride concentrations across ethnicgroups in Singaporerdquo Journal of Lipid Research vol 983092983092 no 983089983090pp 983090983091983094983093ndash983090983091983095983091 983090983088983088983091

[983091983092] M Guardiola R Ferre J Salazar et al ldquoProtease inhibitor-associated dyslipidemia in HIV-inected patients is strongly in1047298uenced by the APOA983093-983089983089983091983089 rarr C gene variationrdquo Clinical Chemistry vol 983093983090 no 983089983088 pp 983089983097983089983092ndash983089983097983089983097 983090983088983088983094

[983091983093] A Kay W Marz M M Hoffmann et al ldquoCoronary artery disease and dyslipidemia within Europe genetic variants inlipid transport gene loci in German subjects with prematurecoronary artery diseaserdquo Atherosclerosis Supplements vol 983091 no983089 pp 983090983095ndash983091983091 983090983088983088983090

[983091983094] P R urner P J almud S Visvikis C Ehnholm and Liret ldquoDNA polymorphisms o the apoprotein B gene are

associated with altered plasma lipoprotein concentrations butnot with perceived risk o cardiovascular disease europeanAtherosclerosis Research Studyrdquo Atherosclerosis vol 983089983089983094 no 983090pp 983090983090983089ndash983090983091983092 983089983097983097983093

[983091983095] M Benn B G Nordestgaard J S Jensen P Grande H Sillesenand A ybjaeligrg-Hansen ldquoPolymorphism in APOB associatedwith increased low-density lipoprotein levels in both genders inthe general populationrdquo Journal of Clinical Endocrinology and Metabolism vol 983097983088 no 983089983088 pp 983093983095983097983095ndash983093983096983088983091 983090983088983088983093

[983091983096] C-F Xu M J ikkanen J K Huttunen et al ldquoApolipoproteinB signal peptide insertiondeletion polymorphism is associatedwith Ag epitopes and involved in the determination o serumtriglyceride levelsrdquo Journal of Lipid Research vol 983091983089 no 983095 pp983089983090983093983093ndash983089983090983094983089 983089983097983097983088

[983091983097] P Ye B Chen and S Wang ldquoAssociation o polymorphisms o the apolipoprotein B gene with coronary heart disease in HanChineserdquo Atherosclerosis vol 983089983089983095 no 983089 pp 983092983091ndash983093983088 983089983097983097983093



Submit your manuscripts at

httpwwwhindawicom




Te exact mechanism o dyslipidemia is not ully under-stood but is most likely multiactorial In the general pop-ulation genetic variation accounts or approximately 983092983091ndash983096983091 o the variability in lipid plasma levels [983093] Recentcandidate gene studies [983094ndash983089983089] as well as genome-wide-basedassociation studies have identi1047297ed certain single nucleotide

polymorphisms (SNPs) that could account or a signi1047297cantportion o the variation in blood lipid levels [983089983090ndash983089983092]In HIV inection genetic predisposition may help to

explain the variability among patients with respect to theeffectso protease inhibitors (PIs) on lipid metabolism [983089983088 983089983089]We have hypothesized that this variation is attributable tothe joint effect o HIV inection and AR together with theunderlying genetic predisposition present in these individu-als Te aim o this study was to investigate the requencieso 983097 SNPs in 983094 candidate genes and to identiy associationsbetween these SNPs and the plasma lipid levels o patients onstable AR with undetectable viral loads

2 Methods

983090983089 Subjects We conducted a cross-sectional study with 983094983089983092patients who were diagnosed with HIV-983089 inection accordingto the criteria o the Centers or Disease Control andPrevention [983089983093] All subjects were more than 983089983095 years old hadregularly used AR or at least 983089983090 months had a viral loadbelow the detection limit o the test (983093983088 copiesmL VersantHIV-983089 RNA 983091983088 Assay (bDNA) Siemens Germany) andwere recruited rom three reerral centers in southern Brazil(HIVAIDS Ambulatory Unit o Hospital de Clınicas romPorto AlegreRS HIV Ambulatory Care o Hospital Univer-sitario Dr Miguel Riet Correa Jr rom Rio GrandeRS and

HIVAIDS Specialized Assistance Service rom PelotasRS)rom March 983090983088983088983094 to November 983090983088983088983096 Pregnant women andthose with neurological disease that prevented understandingand proper consent were not included in the study Te study protocol was approved by the Research Ethics Committees o the three centers and o the Universidade Federal de Cienciasda Saude de Porto Alegre and all participants signed aninormed consent statement beore they were included in thestudy (protocol numbers 983088983093983090983097983093 983095983089983096983088983096 983089983093983092983088983095 and 983089983092983089983088983094resp)

983090983090 Study Protocol Te routine evaluation consisted o visitsevery 983092 months in each center or an evaluation by the

patientsrsquo attending physicians as well as laboratory evalua-tions that included measurements o CD983092 cell counts viralload and lipid levels Te patients were invited to participatein the study and had their inormation and a blood sampleor DNA extraction collected during one o these visits

An interview was perormed at enrollment to obtaindemographic and liestyle inormation Details o HIVinection (time rom diagnosis as well as current andprior antiretroviral medications) lipid-lowering interven-tion and relevant clinical variables were obtained rommedical records Te interviewer phenotypically de1047297ned thepatientsrsquo ethnicities because there might be a strong culturaltendency to claim European ancestry in Brazil [983089983094] Patients

were classi1047297ed as Euro- or Aro-descendants because theAmerindian contribution is very low in the Brazilian SouthRegion [983089983095]

983090983091 Laboratory Analysis Blood samples were collected andsent to theMolecular Biology Laboratory orDNA extraction

Lipid pro1047297les included determinations o total cholesterol(C) high-density lipoprotein (HDL-C) triglycerides (G)and when possible low-density lipoprotein (LDL-C) aferasting or 983089983090 hours LDL-C was calculated using the Friede-wald ormula LDL-C = C minus HDL-C minus G983093 i triglyceridelevels were below 983092983088983088 mgdL

Dyslipidemia was de1047297ned by asting triglyceridesplasma levels ge983089983093983088 mgdL andor asting total cholesterolge983090983088983088 mgdL andor LDL-C ge983089983091983088 mgdL andor HDL-Clt983092983088 mgdL Participants were instructed not to perorm any

vigorous physical activity or ingest alcohol in the 983090983092 hoursprior to the blood collection [983089983096]

Genomic DNA was obtained rom peripheral leukocytes

by a standard salting-out technique [983089983097] Te genotypes o apolipoprotein B gene ( APOB) polymorphisms were deter-mined using polymerase chain reaction (PCR) based pro-cedures in 983092983089983088 individuals rom the outpatient clinic o theHospital de Clınicas de Porto Alegre Te polymorphism o insertiondeletion o signal peptide SP InsDel (rs983089983095983090983092983088983092983092983089)was ampli1047297ed by PCR using primers as previously described[983090983088] and directly analyzed by electrophoresis in 983096 polyacry-lamide gels Te XbaI 983095983094983095983091Cgt (rs983094983097983091) polymorphism wasampli1047297ed by PCR using the primers described by Pan et al[983090983089] and genotypes were determined by digestion with XbaIrestriction endonuclease and electrophoresis in 983090 agarosegels

Te SNPs o apolipoprotein A-V ( APOA983093) minus983089983089983091983089gtC(rs983094983094983090983095983097983097) and S983089983097W (983093983094CgtG rs983091983089983091983093983093983088983094) apolipoproteinE( APOE) 983091983091983092gtC (E983092 rs983092983090983097983091983093983096) and 983092983095983090Cgt (E983090 rs983095983092983089983090)apolipoprotein C-III ( APOC983091) 983091983090983091983096CgtG (rs983093983089983090983096) sterol reg-ulatory element-binding actor cleavage-activating protein(SCAP ) 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094) and low-density lipoproteinreceptor (LDLR) intron 983089983097Ggt (rs983094983093983089983089983095983090983088) were genotypedby real-time PCR using the aqMan methodology and arelisted in able 983089 Candidate SNPs were selected via review o PubMed reports o SNP associations with dyslipidemia in thegeneral population or among HIV-inected individuals [983094ndash983089983089]

983090983092 Statistical Analyses Te results are expressed as themean plusmn standard deviation (SD) or continuous variablesand as proportions or categorical variables Variables thatdid not have a normal distribution (triglycerides) weretransormed into natural logarithms beore the statisticaltests were applied Allele requencies were estimated by gene

counting 2 analysis that was used to test or deviations ingenotype requencies rom the Hardy-Weinberg equilibrium(HWE)

For the APOC983091 983091983090983091983096CgtG (rs983093983089983090983096) APOA983093 minus983089983089983091983089gtC(rs983094983094983090983095983097983097) and S983089983097W (983093983094CgtG) (rs983091983089983091983093983093983088983094) SCAP 983090983091983096983094AgtG(rs983089983090983092983096983095983095983091983094) and LDLR intron 983089983097Ggt (rs983094983093983089983089983095983090983088) polymor-phisms the association analyses were perormed according







983093983089983093 (983096983092983095) 983088983097983090 983088983088983096C 983097983088 (983089983092983096)

CC 983091 (983088983093)

otal 983094983088983096


CC 983093983088983088 (983096983090) 983088983097983088 983088983089983088

CG 983089983088983091 (983089983094983097)

GG 983095 (983089983089)

otal 983094983089983088


Insins 983089983097983091 (983092983095983091) 983088983095983088 983088983091983088

Insdel 983089983095983094 (983092983091983089)

Deldel 983091983097 (983097983094)otal 983092983088983096


CC 983089983090983090 (983091983088) 983088983093983093 983088983092983093

C 983090983088983091 (983092983097983097)

983096983090 (983090983088983089)

otal 983092983088983095

APOC983091 983091983090983091983096CgtG (rs983093983089983090983096)b C G

CC 983092983095983096 (983095983096983096) 983088983096983097 983088983089983089

CG 983089983090983095 (983090983088983097)

GG 983090 (983088983091)

otal 983094983088983095

APOEa E983090 E983091 E983092E983090E983090 983093 (983088983096) 983088983088983095 983088983095983097 983088983089983092

E983090E983091 983094983095 (983089983089983090)

E983090E983092 983097 (983089983093)

E983091E983091 983091983095983097 (983094983091983094)

E983091E983092 983089983090983088 (983090983088983089)

E983092E983092 983089983094 (983090983095)

otal 983093983097983094


GG 983092983096983095 (983096983088983090) 983088983097983088 983088983089983088

G 983089983089983091 (983089983096983094)

983095 (983089983090)

otal 983094983088983095

SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094)a A G

AA 983089983094983096 (983090983095983095) 983088983093983090 983088983092983096

AG 983090983097983094 (983092983096983097)

GG 983089983092983090 (983090983091983092)

otal 983094983088983094








PI-sparingAR

1038389 = 311

PI-basedAR

1038389 = 303

1103925 value

Demographic


0114sect

Male sex 1038389 () 983091983092983089 (983093983093983093) 983089983095983097 (983093983096) 983089983094983090 (983093983092) 983088983091983092983096lowast



Aro-Brazilians 983090983094983093 (983092983091) 983089983092983093 (983092983095) 983089983090983088 (983092983088)



Clinical




Metabolic
















3 Results








Polymorphisms

otal cholesterol

(mgdL)

LDL-C

(mgdL)

HDL-C

(mgdL)

riglycerides

(mgdL)


APOA983093 minus983089983089983091983089gtC (rs983094983094983090983095983097983097)



P value 983088983093983092983097 983088983097983093983091 983088983088983092983095983076 983088983088983088983091lowast983203


APOA983093 S983089983097W (983093983094CgtG rs983091983089983091983093983093983088983094)



P value 983088983091983091983088 983088983088983092983095

983075

983088983089983091983088 983088983091983097983096

lowast


APOB SP InsDel (rs983089983095983090983092983088983092983092983089)






APOB XbaI 983095983094983095983091Cgt (rs983094983097983091)



983096983089 203 plusmn 59

983095983091 110 plusmn 41ih



APOC983091 983091983090983091983096CgtG (rs983093983089983090983096)



P value 983088983095983089983096 983088983096983091983091 983088983090983097983093 983088983097983088983093lowast


APOE genotype









P value 983088983094983089983090 983088983092983090983095 983088983089983090983088 983088983097983095983089lowast





983137983138983148983141 983091 Continued

Polymorphisms


LDL-C(mgdL)

HDL-C(mgdL)

riglycerides(mgdL)


SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094)




P value 983088983092983092983091 983088983097983095983088 983088983088983091983090983205 983088983097983095983089lowast

























983090983093 118 plusmn 43b









4 Discussion















minus983089983089983091983089






















Disclosure




Acknowledgments





















































httpwwwhindawicom







983093983089983093 (983096983092983095) 983088983097983090 983088983088983096C 983097983088 (983089983092983096)

CC 983091 (983088983093)

otal 983094983088983096


CC 983093983088983088 (983096983090) 983088983097983088 983088983089983088

CG 983089983088983091 (983089983094983097)

GG 983095 (983089983089)

otal 983094983089983088


Insins 983089983097983091 (983092983095983091) 983088983095983088 983088983091983088

Insdel 983089983095983094 (983092983091983089)

Deldel 983091983097 (983097983094)otal 983092983088983096


CC 983089983090983090 (983091983088) 983088983093983093 983088983092983093

C 983090983088983091 (983092983097983097)

983096983090 (983090983088983089)

otal 983092983088983095

APOC983091 983091983090983091983096CgtG (rs983093983089983090983096)b C G

CC 983092983095983096 (983095983096983096) 983088983096983097 983088983089983089

CG 983089983090983095 (983090983088983097)

GG 983090 (983088983091)

otal 983094983088983095

APOEa E983090 E983091 E983092E983090E983090 983093 (983088983096) 983088983088983095 983088983095983097 983088983089983092

E983090E983091 983094983095 (983089983089983090)

E983090E983092 983097 (983089983093)

E983091E983091 983091983095983097 (983094983091983094)

E983091E983092 983089983090983088 (983090983088983089)

E983092E983092 983089983094 (983090983095)

otal 983093983097983094


GG 983092983096983095 (983096983088983090) 983088983097983088 983088983089983088

G 983089983089983091 (983089983096983094)

983095 (983089983090)

otal 983094983088983095

SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094)a A G

AA 983089983094983096 (983090983095983095) 983088983093983090 983088983092983096

AG 983090983097983094 (983092983096983097)

GG 983089983092983090 (983090983091983092)

otal 983094983088983094








PI-sparingAR

1038389 = 311

PI-basedAR

1038389 = 303

1103925 value

Demographic


0114sect

Male sex 1038389 () 983091983092983089 (983093983093983093) 983089983095983097 (983093983096) 983089983094983090 (983093983092) 983088983091983092983096lowast



Aro-Brazilians 983090983094983093 (983092983091) 983089983092983093 (983092983095) 983089983090983088 (983092983088)



Clinical




Metabolic
















3 Results








Polymorphisms

otal cholesterol

(mgdL)

LDL-C

(mgdL)

HDL-C

(mgdL)

riglycerides

(mgdL)


APOA983093 minus983089983089983091983089gtC (rs983094983094983090983095983097983097)



P value 983088983093983092983097 983088983097983093983091 983088983088983092983095983076 983088983088983088983091lowast983203


APOA983093 S983089983097W (983093983094CgtG rs983091983089983091983093983093983088983094)



P value 983088983091983091983088 983088983088983092983095

983075

983088983089983091983088 983088983091983097983096

lowast


APOB SP InsDel (rs983089983095983090983092983088983092983092983089)






APOB XbaI 983095983094983095983091Cgt (rs983094983097983091)



983096983089 203 plusmn 59

983095983091 110 plusmn 41ih



APOC983091 983091983090983091983096CgtG (rs983093983089983090983096)



P value 983088983095983089983096 983088983096983091983091 983088983090983097983093 983088983097983088983093lowast


APOE genotype









P value 983088983094983089983090 983088983092983090983095 983088983089983090983088 983088983097983095983089lowast





983137983138983148983141 983091 Continued

Polymorphisms


LDL-C(mgdL)

HDL-C(mgdL)

riglycerides(mgdL)


SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094)




P value 983088983092983092983091 983088983097983095983088 983088983088983091983090983205 983088983097983095983089lowast

























983090983093 118 plusmn 43b









4 Discussion















minus983089983089983091983089






















Disclosure




Acknowledgments





















































httpwwwhindawicom






PI-sparingAR

1038389 = 311

PI-basedAR

1038389 = 303

1103925 value

Demographic


0114sect

Male sex 1038389 () 983091983092983089 (983093983093983093) 983089983095983097 (983093983096) 983089983094983090 (983093983092) 983088983091983092983096lowast



Aro-Brazilians 983090983094983093 (983092983091) 983089983092983093 (983092983095) 983089983090983088 (983092983088)



Clinical




Metabolic
















3 Results








Polymorphisms

otal cholesterol

(mgdL)

LDL-C

(mgdL)

HDL-C

(mgdL)

riglycerides

(mgdL)


APOA983093 minus983089983089983091983089gtC (rs983094983094983090983095983097983097)



P value 983088983093983092983097 983088983097983093983091 983088983088983092983095983076 983088983088983088983091lowast983203


APOA983093 S983089983097W (983093983094CgtG rs983091983089983091983093983093983088983094)



P value 983088983091983091983088 983088983088983092983095

983075

983088983089983091983088 983088983091983097983096

lowast


APOB SP InsDel (rs983089983095983090983092983088983092983092983089)






APOB XbaI 983095983094983095983091Cgt (rs983094983097983091)



983096983089 203 plusmn 59

983095983091 110 plusmn 41ih



APOC983091 983091983090983091983096CgtG (rs983093983089983090983096)



P value 983088983095983089983096 983088983096983091983091 983088983090983097983093 983088983097983088983093lowast


APOE genotype









P value 983088983094983089983090 983088983092983090983095 983088983089983090983088 983088983097983095983089lowast





983137983138983148983141 983091 Continued

Polymorphisms


LDL-C(mgdL)

HDL-C(mgdL)

riglycerides(mgdL)


SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094)




P value 983088983092983092983091 983088983097983095983088 983088983088983091983090983205 983088983097983095983089lowast

























983090983093 118 plusmn 43b









4 Discussion















minus983089983089983091983089






















Disclosure




Acknowledgments





















































httpwwwhindawicom





Polymorphisms

otal cholesterol

(mgdL)

LDL-C

(mgdL)

HDL-C

(mgdL)

riglycerides

(mgdL)


APOA983093 minus983089983089983091983089gtC (rs983094983094983090983095983097983097)



P value 983088983093983092983097 983088983097983093983091 983088983088983092983095983076 983088983088983088983091lowast983203


APOA983093 S983089983097W (983093983094CgtG rs983091983089983091983093983093983088983094)



P value 983088983091983091983088 983088983088983092983095

983075

983088983089983091983088 983088983091983097983096

lowast


APOB SP InsDel (rs983089983095983090983092983088983092983092983089)






APOB XbaI 983095983094983095983091Cgt (rs983094983097983091)



983096983089 203 plusmn 59

983095983091 110 plusmn 41ih



APOC983091 983091983090983091983096CgtG (rs983093983089983090983096)



P value 983088983095983089983096 983088983096983091983091 983088983090983097983093 983088983097983088983093lowast


APOE genotype









P value 983088983094983089983090 983088983092983090983095 983088983089983090983088 983088983097983095983089lowast





983137983138983148983141 983091 Continued

Polymorphisms


LDL-C(mgdL)

HDL-C(mgdL)

riglycerides(mgdL)


SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094)




P value 983088983092983092983091 983088983097983095983088 983088983088983091983090983205 983088983097983095983089lowast

























983090983093 118 plusmn 43b









4 Discussion















minus983089983089983091983089






















Disclosure




Acknowledgments





















































httpwwwhindawicom




983137983138983148983141 983091 Continued

Polymorphisms


LDL-C(mgdL)

HDL-C(mgdL)

riglycerides(mgdL)


SCAP 983090983091983096983094AgtG (rs983089983090983092983096983095983095983091983094)




P value 983088983092983092983091 983088983097983095983088 983088983088983091983090983205 983088983097983095983089lowast

























983090983093 118 plusmn 43b









4 Discussion















minus983089983089983091983089






















Disclosure




Acknowledgments





















































httpwwwhindawicom










983090983093 118 plusmn 43b









4 Discussion















minus983089983089983091983089






















Disclosure




Acknowledgments





















































httpwwwhindawicom







minus983089983089983091983089






















Disclosure




Acknowledgments





















































httpwwwhindawicom






Disclosure




Acknowledgments





















































httpwwwhindawicom






























httpwwwhindawicom




httpwwwhindawicom

Documents

Genetic Markers Associated to Dyslipidemia in HIV-Infected