Estudo Português de Hipercolesterolemia

7/14/2019 Estudo Portugus de Hipercolesterolemia

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Estudo Portugus de HipercolesterolemiaFamiliar: Apresentao do Estudo e

Resultados Preliminares [76]MAFALDA BOURBON, QUITRIA RATO, pelos Investigadores do Estudo Portugus de Hipercolesterolemia Familiar

Unidade de Investigao Cardiovascular, Centro de Biopatologia, Instituto Nacional de Sade Dr. Ricardo Jorge, Lisboa, PortugalServio de Cardiologia, Centro Hospitalar de Setbal, E.P.E., Hospital de So Bernardo, Setbal, Portugal

Rev Port Cardiol 2006; 25 (11): 999-1013

999

RESUMO

Introduo: A hipercolesterolemia familiar(FH) uma patologia gentica, autossmica

dominante, causada na maioria dos casos pelaausncia total ou parcial de receptores daslipoprotenas de baixa densidade (LDLR)

funcionais. Mutaes no gene LDLR levam aoaumento dos nveis de colesterol plasmtico, o

que provoca a sua deposio nas artrias econsequente incremento do risco de doena

coronria prematura. A forma homozigtica daFH rara, mas a heterozigtica comum

embora sub-diagnosticada em muitaspopulaes, nomeadamente na portuguesa.

Em 1999 iniciou-se no Instituto Nacional deSade Dr. Ricardo Jorge o Estudo Portugusde Hipercolesterolemia Familiar.

Objectivos: O objectivo do Estudo Portugusde Hipercolesterolemia Familiar a

realizao de um estudo epidemiolgico paraa determinao da prevalncia e distribuioda Hipercolesterolemia Familiar em Portugale a melhor compreenso da fisiopatologia da

doena cardiovascular. O objectivo dopresente trabalho apresentar os critrios e a

organizao do estudo assim como osresultados preliminares.

Material e Mtodos: A populao do estudoso indivduos de ambos os sexos, sem

restries etrias, com o diagnstico clnicode FH, sendo efectuada a caracterizao

bioqumica e molecular. Os critrios clnicosutilizados para o diagnstico de FH so

adaptados de Simon Broome HeartResearch Trust. Este estudo engloba cinco

fases: 1- seleco de indivduos com odiagnstico clnico de FH, 2- preenchimento

de um questionrio clnico e obteno do

ABSTRACT

Portuguese Familial Hypercholeste-rolemia Study: Presentation of the Study

and Preliminary ResultsIntroduction: Familial hypercholesterolemia(FH) is an autosomal dominant geneticdisorder caused, in the majority of cases, by apartial or total lack of functional low densitylipoprotein receptors (LDLR). Mutations inthe LDLR gene lead to increased plasmacholesterol levels, resulting in cholesteroldeposition in the arteries, thereby increasingthe risk of premature coronary heart disease.

The homozygous form of FH is rare butheterozygous FH is common, althoughunderdiagnosed in many populations,including the Portuguese. In 1999 thePortuguese Familial HypercholesterolemiaStudy was begun at the National Institute ofHealth.Objectives: The aim of the PortugueseFamilial Hypercholesterolemia Study is toperform an epidemiological study todetermine the prevalence and distribution ofFH in Portugal and to better understand thepathophysiology of coronary heart disease inthese patients. The aim of the present work isto present the study's criteria and organizationas well as its preliminary results.Methods: The study population consists ofindividuals of both sexes and all ages with aclinical diagnosis of FH, with biochemicaland molecular characterization beingperformed. The clinical criteria used for thediagnosis of FH were adapted from those ofthe Simon Broome Heart Research Trust. Thestudy is organized in five stages: 1. selection

Recebido para publicao: Setembro de 2005 Aceite para publicao: Agosto de 2006

Received for publication: September 2005 Accepted for publication: August 2006


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1000

Rev Port CardiolVol. 25 Novembro 06 / November 06

consentimento informado, 3- colheita desangue, 4- caracterizao bioqumica 5-

estudo molecular englobando a pesquisa demutaes em trs genes associados ao

fentipo de FH:LDLR, apolipoprotena B(APOB) eProprotein convertase

subtilisin/kexin type 9 (PCSK9).Resultados: Entre 1999 e Junho 2006estudaram-se os genes LDLR e ApoB de 141casos ndex (38 crianas e 103 adultos). Em

78 destes casos ndex (76 heterozigotos e doishomozigotos) foram encontradas 50 mutaes

diferentes no gene LDLR e em doisindivduos foi detectada a mutao ApoB3500.

Foi tambm realizado o estudo do genePCSK9 dos indivduos nos quais no foi

detectada uma mutao nos genes LDLR e

APOB, levando identificao de 2indivduos com mutao neste gene. O estudode 62 famlias levou identificao adicional

de 117 indivduos com FH, 90 adultos e 27crianas (86 adultos e 27

crianas/adolescentes com mutaes no geneLDLR, dois com a mutao ApoB3500 e dois

com mutao no gene PCSK9).Concluso: O diagnstico molecular da FH

permite a correcta identificao da patologia efundamenta a instituio de teraputica

farmacolgica mais agressiva e/ou precoce,com a consequente reduo do riscocardiovascular nos indivduos afectados.

Actualmente o Estudo Portugus deHipercolesterolemia Familiar tem a

colaborao de treze clnicos, mas deextrema importncia a colaborao de mais

mdicos em todo o pas, para que aprevalncia e distribuio da FH na

populao portuguesa seja caracterizada e ummaior nmero de indivduos possa usufruir

dos benefcios da teraputica adequada e/ouprecoce.

Palavras-ChaveHipercolesterolemia familiar; Diagnstico gentico;

Receptor das lipoprotenas de baixa densidade; Mutao;Doena coronria; Caso-ndex

of individuals with a clinical diagnosis of FH;2. completion of a clinical questionnaire anddeclaration of informed consent; 3. collectionof blood samples; 4. biochemicalcharacterization; 5. molecular study of threegenes associated with the FH phenotype:

LDLR, apolipoprotein B (APOB) andproprotein convertase subtilisin/kexin type 9(PCSK9).Results: Between 1999 and June 2006 theLDLR gene and theAPOB gene of 141 indexcases (38 children and 103 adults) werestudied. In 78 of these index cases (76heterozygotes and two homozygotes) 50different mutations in theLDLR gene wereidentified, and two unrelated individuals werefound to have the ApoB3500 mutation. The

PCSK9 gene was also studied in individualsin whom a mutation in theLDLR orAPOBgenes was not found, which identified twoindex cases with a mutation in this gene. Thestudy of 62 families led to the identification ofan additional 117 individuals with FH, 90adults and 27 children (86 adults and 27children with mutations in theLDLR gene,two adults with the ApoB3500 mutation, andtwo adults with a mutation in thePCSK9gene).

Conclusions: Genetic diagnosis enablescorrect identification of the disease andprovides the basis for more aggressivepharmacologic therapeutic interventions toreduce cardiovascular risk in affectedindividuals. At present thirteen clinicians arecollaborating in the Portuguese FH Study, butit is extremely important to obtain thecollaboration of more physicians throughoutthe country, so that the prevalence anddistribution of FH in the Portuguesepopulation can be characterized and a greaternumber of individuals can benefit fromappropriate and early therapy.

Key wordsFamilial hypercholesterolemia; Genetic diagnosis;Low-density lipoprotein receptor; Mutation;Coronary heart disease; Index case


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INTRODUO

Hipercolesterolemia Familiar

Ahipercolesterolemia familiar (FH) umapatologia gentica, autossmica dominante,com uma prevalncia heterozigtica de 1:500 na

maioria das populaes europeias, o que a tornauma das patologias genticas mais comuns (1).Num rastreio aleatrio a metade dos utentes doficheiro de um mdico de famlia da Beira Litoralinterior (752 utentes), 0,7% dos mesmos tinhamum colesterol total superior a 350 mg/dL etrigliceridos inferiores a 200 mg/dl, o que fazsuspeitar de hipercolesterolemia familiarnesta populao especfica (2). No encontrmosna reviso da literatura estudos de FH realizadosem amostras representativas da populao

portuguesa, ainda que s por critrios clnicos,que permitam determinar a sua prevalncia nonosso pas. Assim, e com base na prevalnciaencontrada na maioria das populaes europeias,calcula-se que existam cerca de 20.000 casos deFH em Portugal. A FH homozigtica rara,estimando-se que afecte uma pessoa num milho.

A base molecular da hipercolesterolemiafamiliar foi descoberta nos anos 80 pelosinvestigadores Goldstein e Brown, que em 1985

ganharam o prmio Nobel da Medicina pelosseus estudos sobre a regulao do metabolismodo colesterol, incluindo a descoberta que a FHera causada pela ausncia total ou parcial dereceptores das lipoprotenas de baixa densidadefuncionais (3). Logo de seguida o gene LDLR foiisolado (4) e as primeiras mutaes neste geneforam descritas (5). Desde ento, esta patologia temsido alvo de vrios estudos, que tm contribudopara uma melhor compreenso da sua basemolecular, assim como para o desenvolvimentodo diagnstico gentico. Ao longo dos anos outrosgenes associados ao fentipo de FH foramdescobertos, como o gene da apolipoprotena B(APOB) e mais recentemente o gene proproteinconvertase subtilisin/kexin type 9 (PCSK9).

Embora a FH seja uma das patologiasgenticas mais comuns, no havia nenhum estudoepidemiolgico ou registo desta patologia nonosso Pas, pelo que em 1999 no Centro deBiopatologia do Instituto Nacional de SadeDr. Ricardo Jorge (INSA) se decidiu iniciar o

presente estudo.

INTRODUCTION

Familial hypercholesterolemia

Familial hypercholesterolemia (FH) is anautosomal dominant genetic disorder, withthe heterozygous form having a prevalence of

1:500 in most European populations, and so it isamong the more common genetic disorders (1). In arandom screening of half the patients on a familyphysicians register in the Beira Litoral region(752 patients), 0.7% had total cholesterolabove 350 mg/dl and triglycerides below 200mg/dl, which suggests the presence of familialhypercholesterolemia in this population (2). Wefound no FH studies on representative samples ofthe Portuguese population in the literature, evenmeasured by clinical criteria alone, that would

enable its prevalence in Portugal to bedetermined. Based on the prevalence found inmost other European populations, it is estimatedthat there are around 20,000 cases of FH inPortugal. Homozygous FH is rare, estimated atone per million.

The molecular basis of familialhypercholesterolemia was discovered in the1980s by Goldstein and Brown, who won theNobel Prize for Medicine in 1985 for theirresearch on the regulation of cholesterol

metabolism, including the discovery that FH wascaused by a partial or total lack of functionallow-density lipoprotein receptors (LDLR) (3). TheLDLR gene was then isolated (4) and the firstmutations described (5). Since then, the disorderhas been the subject of various studies that havecontributed to a better understanding of itsmolecular basis and to the development ofgenetic diagnosis. Over the years, other genesassociated with the FH phenotype have beendiscovered, such as apolipoprotein B (APOB)and more recently proprotein convertasesubtilisin/kexin type 9 (PCSK9).

Although FH is among the most commongenetic disorders, there had been noepidemiological study or registry of the pathologyin Portugal, and so the Biopathology Center ofthe National Institute of Health (INSA) decidedto set up the present study in 1999.

Clinical diagnosis of familialhypercholesterolemia

The homozygous form of familial

hypercholesterolemia is rare but heterozygous 1001

MAFALDA BOURBON, QUITRIA RATORev Port Cardiol 2006; 25: 999-1013


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Diagnstico clnico da HipercolesterolemiaFamiliar

A forma homozigtica da hipercolesterolemiafamiliar rara, mas a heterozigtica comum,se bem que sub-diagnosticada (6), apesar depreencher os critrios da Organizao Mundial

de Sade para programas de rastreio em largaescala (7). A FH, causada por defeitos no gene doLDLR, e a Deficincia Familiar em ApoB (FDB),causada por defeitos no gene APOB (com umafrequncia entre 1:200 a 1:1250 dependendo daspopulaes), so clinicamente indistinguveis,sendo actualmente englobadas no diagnsticode hipercolesterolemia familiar(8,9,10). No entanto,em geral, as manifestaes clnicas da FDB somais leves que as causadas por defeitos no genedo LDLR (11,12).

A FH caracterizada por nveis elevados decolesterol total e das LDL, acima do percentil95% para o sexo e a idade, com valores decolesterol das lipoprotenas de alta densidade ede trigliceridos dentro da normalidade e umahistria familiar de hipercolesterolemia e dedoena cardiovascular prematura. Clinicamente aforma homozigtica da FH tem um fentipomais severo do que a heterozigtica (10). Ocolesterol total na forma heterozigtica varia entre

290 e 500 mg/dL e na forma homozigticahabitualmente encontra-se entre os 600 mg/dL eos 1000 mg/dL (13). O nvel elevado de colesterolno plasma frequentemente resulta em depsitosde colesterol nos tecidos extravasculares, quepor vezes podem ser facilmente identificados:xantomas, xantelasmas, arco corneano emindivduos ainda jovens (abaixo dos 45 anos) (13). Apresena de xantomas tendinosos geralmenteconsiderada patognomnica da FH, pois emboraeles possam estar presentes na hiperli-poproteinemia tipo III, na fitoesterolemia e naxantomatose cerebrotendinosa estas patologiasso muito raras (14). Os xantomas tendinosos somais facilmente reconhecidos no tendo deAquiles, onde causam irregularidades e o seuespessamento. Outra localizao frequente nos tendes extensores dos dedos das mos. Aecografia do Tendo de Aquiles em casossuspeitos pode ter interesse, j que a medio dasua espessura apresenta uma boa sensibilidade eespecificidade (15). Na populao portuguesa apresena de xantomas tendinosos parece ser

semelhante descrita na populao espanhola

FH is common, albeit underdiagnosed (6), despitemeeting the World Health Organization criteriafor large-scale screening programs (7). FH, causedby defects in the LDLR gene, and familialdefective ApoB (FDB), which is caused bydefects in the APOB gene and has a prevalence

of between 1:200 and 1:1250 depending onthe population, are clinically indistinguishableand are both currently included under thediagnosis of familial hypercholesterolemia (8, 9, 10).However, the clinical manifestations of FDB aregenerally milder than those caused by defects intheLDLR gene (11, 12).

FH is characterized by high levels of total andLDL cholesterol, above the 95th percentile forsex and age, with normal levels of high densitylipoprotein (HDL) cholesterol and triglycerides,

and a family history of hypercholesterolemiaand premature cardiovascular disease. Clinically,the homozygous form of FH has a more severephenotype than the heterozygous form (10). Totalcholesterol in heterozygous FH ranges between290 and 500 mg/dl, while in the homozygousform it is usually between 600 mg/dl and1000 mg/dl (13). Elevated plasma cholesterollevels frequently result in cholesterol depositionin extravascular tissues, which can be easilydetected in some cases, resulting in xanthoma,

xanthelasma, and corneal arcus in youngadults (under 45) (13). The presence of tendonxanthomas is generally considered diagnosticof FH, as although they can be due totype III hyperlipoproteinemia, phytosterolemiaor cerebrotendinous xanthomatosis, thesepathologies are extremely rare (14). Tendonxanthomas are most easily detected on theAchilles tendon, which appears irregular andthickened. Another common location is on thefinger tendons. Achilles tendon ultrasonographycan be useful in suspected cases, sinceassessment of the tendon's thickness has highsensitivity and specificity (15). The prevalenceof tendon xanthomas in the Portuguese populationappears to be similar to that found in Spain butis much lower than that reported in UnitedKingdom (16). Xanthelasmas may be found inpeople with a normal lipid profile; they are notuncommon in the elderly, and arcus senilis isoften found among those aged over 60 (13).

Differential diagnosis of FH from other causesof hypercholesterolemia, particularly common or

polygenic hypercholesterolemia, is important1002



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mas bastante menos frequente do que a referidana populao inglesa (16). Os xantelasmas podemencontrar-se em pessoas normolipidmicas eno so incomuns nos idosos, o arco senilencontra-se frequentemente em indivduosacima dos 60 anos (13).

A diferenciao da FH de outras causas dehipercolesterolemia, nomeadamente da hiperco-lesterolemia comum ou polignica, assume omaior interesse por a FH se associar a um elevadorisco de doena coronria prematura, no fatale fatal, j que o indivduo est exposto aelevados nveis de colesterol plasmtico desde onascimento (17-20). O aumento do colesterolplasmtico leva ao incremento da deposio departculas lipdicas nas artrias, dando origem a

aterosclerose precoce(21)

. O territrio coronrio omais atingido, mas o territrio arterial perifricotambm afectado (22). A associao entre FH e oaumento de risco para acidente vascular cerebralainda pouco clara (22). Nos indivduos com FHhomozigtica o ateroma da crossa da aorta e davlvula artica a manifestao cardaca maisfrequente, podendo levar a um estreitamento daaorta ascendente e arco artico, bem como aestenose dos stios das artrias coronrias (20). Emcrianas com a forma homozigtica da patologia

pode encontrar-se doena aterosclertica massivadesde os primeiros meses de vida, e a doenacardiovascular comea na infncia podendolevar morte na primeira ou segunda dcadasde vida, particularmente se o tratamento no foriniciado nos primeiros meses/anos de vida (20).Estudos recentes indicam que crianas comFH heterozigtica, a partir dos 12 anos deidade, j apresentam um aumento significativoda espessura da ntima-mdia das artriascarotdeas (17,23).

Como a FH uma patologia gentica,autossmica dominante, os descendentes de umindivduo afectado tm uma probabilidade de50% de ter a patologia. estimado que 5-10%dos doentes que sofrem um enfarte do miocrdioantes dos 55 anos de idade possam ter FH (24,25), oque demonstra a importncia do estudo destapatologia para a preveno cardiovascular. Deuma forma generalizada, considera-se que amelhor abordagem para a descoberta de novoscasos a que tem como base a investigao de

familiares de casos conhecidos (26). O estudo de

since FH is associated with a high risk ofpremature coronary heart disease, which can befatal, since affected individuals have beenexposed to high plasma cholesterol levelssince birth (17, 18, 19, 20). Elevated plasma cholesterolleads to increased deposition of lipid particles in

arteries, resulting in premature atherosclerosis (21).The coronary territory is the most affected butperipheral arteries may also be involved (22). It isstill unclear whether there is an associationbetween FH and increased risk of stroke (22). Themost common cardiac manifestation inindividuals with homozygous FH is atheroma inthe aortic arch or aortic valve, which can result innarrowing of the ascending aorta and aortic arch,as well as stenosis of the coronary artery ostia (20).Children with the homozygous form of the

disorder may develop widespread atheroscleroticdisease in the first months of life, andcardiovascular disease can begin in childhoodand lead to death in the first or second decade oflife, particularly if left untreated in the firstmonths or years (20). Recent studies indicate thatchildren with heterozygous FH present asignificant increase in carotid artery intima-media thickness from the age of twelve (17, 23).

Since FH is an autosomal dominant geneticdisorder, descendents of an affected individual

will have a 50% probability of developing thedisease. It is estimated that 5-10% of patientswho suffer myocardial infarction before the ageof 55 have FH (24, 25), which demonstrates theimportance for cardiovascular prevention ofscreening for the disease. It is generally believedthat the best way to detect new cases is toinvestigate relatives of known cases (26). Studyingpatients with premature cardiovascular diseaseand the children of patients who have died orsuffer from premature cardiovascular disease, aswell as screening 16-year-olds, have also beensuggested as cost-effective strategies for earlyidentification of these individuals (26, 27). Genetictesting for the LDL receptor is seen as the mosteffective. However, diagnosis is generally basedon a range of clinical and biochemical criteriaand family history. There is no internationalagreement on the clinical criteria for a diagnosisof FH, with three systems currently in use: in theUnited States, the MEDPED (Make EarlyDiagnoses to Prevent Early Deaths in MEDicalPEDigrees) system is used (28); in The

Netherlands, a scoring system classifying FH as 1003



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doentes com doena cardiovascular prematura, osfilhos de doentes que morrem ou sofremprematuramente de doena cardiovascular ou orastreio de jovens de 16 anos, tm sido tambmapontados como a estratgia mais custo-efectivapara a identificao precoce destes indiv-

duos (26, 27). O teste gentico para o receptor dasLDL visto como o mais eficaz. O diagnstico no entanto geralmente baseado num conjuntode critrios clnicos, bioqumicos e na histriafamiliar. No h acordo internacional nadefinio de critrios clnicos para o diagnsticode FH, sendo utilizados habitualmente trssistemas. Nos Estados Unidos da Amrica usa-seo sistema MEDPED (make early diagnosis,prevent early death on medical pedigrees) (28). NaHolanda um sistema de score definindo FH

em confirmada, provvel e possvel (Dutchfamilial hypercholesterolaemia diagnosticsystem) (19). No Reino Unido so utilizados oscritrios do registo de FH de Simon Broome queestabelecem o diagnstico de FH emconfirmada ou possvel (29,30) (Tabela I). Osvalores preditivo positivo e negativo destesdiferentes sistemas no tm sido largamentecomparados e dependem em parte dos critriosde seleco usados para o recrutamento dosindivduos a rastrear(8).

Diagnstico molecular deHipercolesterolemia Familiar

A FH maioritariamente causada por defeitosno gene do receptor das lipoprotenas de baixadensidade (LDLR), cuja funo a remoodo colesterol LDL do plasma, entregando-o aofgado para ser processado. Existe tambmoutra forma de hipercolesterolemia familiardenominada Deficincia Familiar em Apo B(FDB) que causada por defeitos no gene APOB.A ApoB o nico ligando da partcula LDLatravs do qual esta partcula se liga ao receptordas LDL (1). Quando o gene que codifica para esteligando se encontra mutado d origem a umaApoB deficiente, qual o receptor das LDL nose consegue ligar. Mutaes neste gene levamtambm acumulao de colesterol no plasma,tendo as mesmas consequncias clnicas que aFH. Enquanto que j foram descritas mais de 900mutaes no gene LDLR nas mais diversaspopulaes (base de dados www.fh.ucl.uk/fh) amaioria dos doentes com FDB apresentam

normalmente uma das quatro mutaes mais

confirmed, probable or possible (familialhypercholesterolemia diagnostic criteria of theDutch Lipid Clinic Network) is employed (19); andthe United Kingdom uses the criteria defined bythe Simon Broome Register, which establish adiagnosis of FH as confirmed or possible (29, 30)

(Table I). The positive and negative predictivevalues of these different systems have not beenthoroughly compared and they depend partly onthe selection criteria used to recruit theindividuals to be screened (8).

Genetic diagnosis of familialhypercholesterolemia

FH is caused in most cases by defects in theLDLR gene, whose function is to remove LDLcholesterol from plasma and deliver it to the liver

for processing. There is another form of familialhypercholesterolemia called familial defectiveapolipoprotein B (FDB), which is caused bydefects in the APOB gene. ApoB is the onlyligand of the LDL particle that can bind to theLDL receptor(1). If the gene that codes for thisligand is mutated, it leads to defective ApoB,which cannot bind to the LDL receptor. Mutationsin this gene also result in accumulation ofcholesterol in plasma, which has the sameclinical consequences as FH. Although more

than 900 mutations have been described for theLDLR gene in a wide variety of populations(database at www.fh.ucl.uk/fh), most patients withFDB present one of the four most frequentAPOBmutations, ApoB3500 being the most common

(31).Another gene associated with the FH phenotypewas discovered in 2004, proprotein convertasesubtilisin/kexin type 9 (32). This gene codes forthe protein with the same name, whose functionhas yet to be determined; it is known only that itis strongly expressed in the liver, where it appearsto perform some role in cholesterol homeostasis.So far seven different mutations have beendescribed in this gene (32, 33, 34) in patients in whomno mutations in LDLR or APOB had beendetected, and which may explain theirhypercholesterolemia. Thus, genetic diagnosisof FH is currently based on the search formutations in these three genes:LDLR, APOB andPCSK9.

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comuns no gene da APOB, sendo a mutaoApoB3500 a mais comum

(31). Em 2004 foidescoberto outro gene associado ao fentipo dehipercolesterolemia familiar, o gene PCSK9

(proprotein convertase subtilisin/kexin type

OBJECTIVES

The Portuguese Familial Hypercho-lesterolemia Study is an epidemiological study

aimed at determining the prevalence and 1005


Tabela I

Critrios de FH adaptados de Simon Broome Heart Research Trust

Hipercolesterolemia Familiar confirmada definida como:

(a) Caso-ndex: Criana menor de 16 anos com colesterol total acima de 200 mg/dL(5.20 mmol/L) ou colesterol LDL acima de 120 mg/dL (3.10 mmol/L);

Caso-ndex: Adulto com colesterol total acima de 290 mg/dL (7.5 mmol/L) oucolesterol LDL acima de 190mg/dL (4.9 mmol/L), e

(b) Xantomas tendinosos no caso-ndex ou familiar (pais, filhos, avs, irmos, tios) ou

(c) Evidncia gentica de mutao no gene receptor de LDL ou ApoB

Hipercolesterolemia Familiar possivel definida como:

(a) Caso-ndex: Criana menor de 16 anos com colesterol total acima de 200 mg/dL(5.20 mmol/L) ou colesterol LDL acima de 120 mg/dL (3.10 mmol/L);

Caso-ndex: Adulto com colesterol total acima de 290 mg/dL (7.5 mmol/L) oucolesterol LDL acima de 190mg/dL (4.9 mmol/L), e

(d) Histria familiar de enfarte do miocrdio antes dos 50 anos em avs e tios ouantes dos 60 anos nos pais, irmos e filhos e/ou histria familiar de nveiselevados de colesterol ( > 290 mg/dL) nos pais, irmos ou filhos; ou colesteroltotal acima de 290 mg/dL (7,5 mmol/L) nos avs e/ou tios.

Table I

FH criteria adapted from those of the Simon Broome Heart Research Trust

Confirmed familial hypercholesterolemia is defined as:

(a) Index case: Child under 16 with total cholesterol over 200 mg/dl (5.20 mmol/l)or LDL cholesterol over 120 mg/dl (3.10 mmol/l);

Index case: Adult with total cholesterol over 290 mg/dl (7.5 mmol/l) or LDLcholesterol over 190 mg/dl (4.9 mmol/l), and

(b) Tendon xanthoma in the index case or relative (parents, children, grandparents,siblings, aunts or uncles), or

(c) Genetic evidence of a mutation in the LDL receptor gene or APOB

Possible familial hypercholesterolemia is defined as:

(a) Index case: Child under 16 with total cholesterol over 200 mg/dl (5.20 mmol/l)or LDL cholesterol over 120 mg/dl (3.10 mmol/l);

Index case: Adult with total cholesterol over 290 mg/dl (7.5 mmol/l) or LDLcholesterol over 190 mg/dl (4.9 mmol/l), and

(d) Family history of myocardial infarction before the age of 50 in grandparents oraunts or uncles, or before the age of 60 in parents, siblings or children, and/orfamily history of elevated cholesterol levels (>290 mg/dl) in parents, siblings orchildren; or total cholesterol over 290 mg/dl (7.5 mmol/l) in grandparents and/oraunts or uncles.


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9) (32). Este gene codifica para uma protenacom o mesmo nome e cuja funo aindadesconhecida, sabendo-se somente que esta uma protena com alta expresso no fgado,onde se pensa que desempenhar algumafuno na homeostase do colesterol. Cerca de

sete mutaes diferentes j foram descritasneste gene (32-34) em doentes nos quais no tinhamsido detectadas mutaes nos genes LDLR eAPOB e que possivelmente explicam a suahipercolesterolemia. Assim, actualmente odiagnstico molecular da FH realiza-se pelapesquisa de mutaes nestes trs genes: LDLR,APOB ePCSK9.

OBJECTIVOS DO ESTUDO

O objectivo do Estudo Portugus deHipercolesterolemia Familiar a realizaode um estudo epidemiolgico para adeterminao da prevalncia e distribuioda Hipercolesterolemia Familiar em Portugale melhorar a compreenso da fisiopatologiada doena cardiovascular. O objectivo dopresente trabalho apresentar os critrios e aorganizao do estudo assim como os resultadospreliminares.

MATERIAIS E MTODOS

A populao do estudo so indivduos deambos os sexos, sem restries etrias, com odiagnstico clnico de FH, sendo efectuada acaracterizao bioqumica e molecular. Ametodologia que se passa a indicar mantem-seem curso, j que o estudo continua a decorrer.

Organizao do estudo1. Seleco de indivduos com o diagnstico

clnico de FH de acordo com os critriosapresentados na tabela I. Estes critrios foramadaptados de Simon Broome Heart ResearchTrust (29).

2. Preenchimento de um questionrio clnicopelo mdico assistente para cada caso-ndex ecada familiar. O mdico assistente aps anecessria explicao, obtm dos indivduos querecruta para o estudo (caso-ndex e familiares) oseu consentimento informado, sendo assinada a

respectiva declarao (no caso de menores de

distribution of FH in Portugal and improvingunderstanding of the pathophysiology of coronaryheart disease. The aim of the present work is topresent the studys criteria and organization aswell as its preliminary results.

METHODS

The study population consists of individualsof both sexes and all ages with a clinicaldiagnosis of FH, with biochemical and molecularcharacterization being performed. Themethodology described below is still being usedas the study continues.

Study organization

1. Selection of individuals with a clinicaldiagnosis of FH according to the criteria given inTable I, which were adapted from those of theSimon Broome Heart Research Trust (29).

2. Completion of a clinical questionnaire bythe attending physician for each index case andeach relative. After explaining the study, thephysician obtains informed consent from thoserecruited (index cases and relatives) by means ofa signed declaration (in the case of minors, thedeclaration is signed by the parents). A

confidential number is allocated to all samples.Steps are being taken to ensure that the studyscomputerized database conforms to legalrequirements.

3. Collection of blood samples performed asdescribed in Table II. The maximum time intervalbetween collection of samples and analysis is twodays, which means the sample must be sent foranalysis immediately after collection. Thiscondition is of the utmost importance for thestudy. The samples are sent by express post to theNational Institute of Health, for the attention ofthe Portuguese Familial HypercholesterolemiaStudy, appropriately packaged in a paddedenvelope, or alternatively, sample collection canbe performed at the Institute by priorappointment. Whenever possible, samples fromrelatives with or without suspected FH arecollected at the same time for subsequent familystudies.

4. Biochemical characterization performed bythe Clinical Chemistry Laboratory (CBP, INSA).This includes measurement of total and HDL

cholesterol, triglycerides and LDL cholesterol.1006



9/15

idade a declarao assinada pelos pais). A todasas amostras atribudo um nmero confidencial.Encontra-se em fase de legalizao a base dedados informatizada referente a este estudo.

3. Colheita de sangue realizada de acordo coma Tabela II. O tempo mximo entre a colheita das

amostras e a sua anlise no dever ultrapassaros dois dias, o que implica que a amostra sejaenviada logo aps a colheita. Esta condio deextrema importncia para o estudo. As amostrasso enviadas em correio azul para o InstitutoNacional de Sade Dr. Ricardo Jorge, a/c EstudoPortugus de Hipercolesterolemia Familiar,em envelope almofadado e bem acondicio-nadas ou, opcionalmente, a colheita realizadano Instituto aps marcao prvia. Sempre quepossvel feita a colheita simultnea a

familiares com e sem suspeita de FH paraposterior realizao dos estudos familiares.

Whenever possible, levels of apolipoprotein AI(ApoAI), ApoB and lipoprotein(a) are alsodetermined.

The results are sent to the attending physicianwithin two to five days of sample receipt orcollection.

5. Genetic study performed by theCardiovascular Research Unit (CBP, INSA). Thisconsists of the search for mutations in the LDLRgene and of the four most common mutations intheAPOB gene. ThePCSK9 gene has also beenstudied since 2005. Access to samples fromrelatives with or without suspected FH isessential in confirming the results of geneticstudy in the index case, which is why it isadvisable to collect samples from relatives at thesame time as the index case. Only after such

confirmation are the results of genetic study sentto the attending physician, within one to four

1007


Tabela II

Condies da colheita das amostras

1.1.1.1 Caso-ndex (as colheitas devem ser realizadas aps jejum de 12 horas)

Adultos:4 mL de sangue em tubo seco - separar e enviar s o soro10 mL (2x5mL) de sangue em tubos de EDTA

Crianas:2 mL de sangue em tubo seco - separar e enviar s o soro6 mL (2x3mL) de sangue em tubos de EDTA

1.1.1.2 Familiares do caso-ndex (as colheitas devem ser realizadas aps jejumde 12 horas)

4 mL de sangue em tubo seco - separar e enviar s o soro5 mL de sangue em tubos de EDTA

Table II

Conditions for collection of samples

Index case (samples must be collected after 12 hours fasting)

Adult:4 ml blood in a dry tube - separate and send serum only10 ml (2x5ml) blood in tubes with EDTA

Child:2 ml blood in a dry tube - separate and send serum only6 ml (2x3ml) blood in tubes with EDTA

Relatives of index case (samples must be collected after 12 hours fasting)

4 ml blood in a dry tube - separate and send the serum only

5 ml blood in a tube with EDTA


10/15

4. Caracterizao bioqumica realizada noLaboratrio de Qumica Clnica (CBP, INSA).A caracterizao bioqumica realizada atravsda determinao dos valores de colesterol total,colesterol das lipoprotenas de alta densidade(HDL), trigliceridos e colesterol das LDL. Sempre

que possvel os valores da apolipoprotenaAI (ApoAI), ApoB e lipoprotena (a) (Lp (a)) sotambm determinados.

Os resultados so enviados ao mdicoassistente no prazo mximo de dois a cinco dias,aps a recepo das amostras ou efectivao dacolheita.

5. Estudo molecular - realizado na Unidade deInvestigao Cardiovascular (CBP, INSA). Esteestudo efectuado atravs da pesquisa demutaes do gene LDLR e tambm pela pesquisa

das quatro mutaes mais comuns no gene daAPOB. A partir de 2005 o estudo do gene PCSK9tambm passou a ser efectuado. A existnciade amostras de familiares com e sem suspeitade FH um factor essencial para confirmaodo resultado molecular no caso-ndex, razopela qual aconselhvel a colheita de amostrasde familiares juntamente com o caso-ndex. Saps esta confirmao que o resultado doestudo enviado para o mdico assistente noprazo de 1 a 4 meses, podendo variar conforme

a existncia de financiamento externo. Este um estudo de investigao, gratuito para oscasos-ndex e familiares, financiado em partepelo prprio Instituto, mas que, como todosos estudos de investigao, depende definanciamento externo.

RESULTADOS

At Junho de 2006 estudaram-se os genesLDLR e APOB de 141 indivduos (38 crianase 103 adultos) (Tabela III) com diagnsticoclnico de FH. Dos 141 indivduos estudados78 apresentam uma mutao no gene LDLR(20 crianas e 58 adultos), 2 adultosapresentam a mutao APOB3500 no gene daAPOB, e outros 2 adultos apresentam umamutao no gene PCSK9 (Tabela III). No geneda LDLR foram encontradas 50 mutaesdiferentes, 19 destas mutaes no foramdescritas anteriormente, sendo actualmenteexclusivas da nossa populao. Identificou-se

dois casos ndex homozigotos, embora um seja

months, depending on availability of externalfunding. This is a research project, which is freeof charge to index cases and their relatives and ispartly financed by the Institute, but, as with allresearch studies, depends on external funding.

RESULTS

Up until June 2006 the LDLR and APOBgenes had been studied in 141 individuals (38children and 103 adults) (Table III) with a clinicaldiagnosis of FH. Of these 141, 78 (20 childrenand 58 adults) had a mutation in theLDLR gene,two adults had the APOB3500 mutation, andanother two adults had a mutation in the PCSK9gene (Table III). Fifty different mutations were

identified in the LDLR gene, 19 of which hadnot been previously described and are at presentexclusive to our study population. Two indexcases of homozygous FH were identified,although one of them was a compoundheterozygote, with two different mutations indifferent alleles of the LDLR gene. The studyof 62 families led to the identification of anadditional 117 individuals with FH (90 adultsand 27 children) (Table IV). Of these 117relatives, 113 had mutations in the LDLR gene,

two adults had the ApoB3500 mutation and twoadults a mutation in thePCSK9 gene (Table IV).

Table III

III Distribution of index cases recruited for the study,by age-group and mutations in the genes studied

IC: index case; 18: adults

Table IV

Distribution of all individuals recruited for the study(index cases and relatives), by age-group andmutations in the genes studied

IC: index case; R: relative; 18: adults1008


Age-group18

No. ofICs

studied38103

No. of ICswithLDLRmutation

2058

No. ICswithAPOBmutation

02

No. ICswithPCSK9

mutation02

Age-group18

No. ofICs+Rs38+65

103+284

No. ofICs+Rs with

LDLRmutation20+2758+86

No. ofICs+Rs with

APOBmutation

02+2

No. ofICs+Rs with

PCSK9mutation

02+2


11/15

um heterozigoto composto, ou seja, com duasmutaes diferentes em alelos diferentes do geneLDLR. Estudos familiares de 62 famlias levaram identificao adicional de 117 indivduos comFH, (90 adultos e 27 crianas) (Tabela IV). Destes117 familiares, 113 apresentavam mutaes no

gene LDLR, dois a mutao ApoB3500 e dois umamutao no gene PCSK9 (Tabela IV).

Tabela III

Distribuio dos casos ndex recebidos para o estudopor grupo etrio e mutaes encontradas nosdiferentes genes estudados

CI, caso ndex; < 18, crianas e adolescentes; >18, adultos

Tabela IV

Distribuio de todos os indivduos recebidos para oestudo (casos ndex e familiares) por grupo etrio emutaes encontradas nos diferentes genes estudados

CI, caso ndex; F, familiares; < 18, crianas e adolescentes; >18, adultos

DISCUSSO E CONCLUSO

Os critrios clnicos de hipercolesterolemiafamiliar usados neste estudo foram adaptados deSimon Broome Heart Research Trust uma vezque este trabalho foi iniciado em colaboraocom o laboratrio do Medical Reserach Council,Lipoprotein Group, Hammersmith Hospital,Londres, UK, que usa aqueles critrios (16). Almdisso os outros dois sistemas descritoscomparativamente no tm demonstrado bene-fcios significativos, pelo que se decidiu manteros critrios escolhidos inicialmente. Os mtodospara a anlise molecular da FH estoestabelecidos em Portugal e apenas o teste para apesquisa de grandes rearranjos no gene LDLRainda efectuado no laboratrio de Londres, mas

brevemente tambm passar a ser efectuado no

DISCUSSION AND CONCLUSION

The clinical criteria for familialhypercholesterolemia used in this study wereadapted from those of the Simon Broome HeartResearch Trust, as the project was begun in

collaboration with the Medical Research CouncilLaboratory, Lipoprotein Group, HammersmithHospital, London, which uses these criteria (16).Comparison of the other two systems referred toabove has not shown them to have significantadvantages, and so it was decided to keep to thecriteria initially chosen. Molecular analysismethods for FH are available in Portugal andonly the test for large rearrangements in theLDLR gene are still performed by the Londonlaboratory, but this too will soon be performed in

the Cardiovascular Research Unit laboratory(CBP, INSA).Four hundred and ninety individuals,

including index cases and relatives with orwithout a clinical diagnosis of FH, arecurrently enrolled in the Portuguese FamilialHypercholesterolemia Study. Genetic study hasidentified a total of 199 individuals with FH (82index cases and 117 relatives). It was onlypossible to perform a family study for two-thirdsof the subjects with a clinical diagnosis of FH,

and in most cases only close relatives. Thecooperation of family members can be difficult toobtain, and this is one of the problems of thestudy. Nevertheless, genetic study of relativesof affected individuals is undoubtedly the mostcost-effective method of detecting new casesof familial hypercholesterolemia and severalcountries have already begun cascade screeningprograms (35, 36, 37).

Analyzing the results by age-group, a total of47 children and adolescents (under 18 years ofage) were diagnosed with heterozygous FH, and152 adults were diagnosed with FH, of whom150 had the heterozygous form and two thehomozygous form. The two cases withhomozygous FH are the first to be reported amongthe Portuguese population. Interestingly, theseindividuals do not present the typical phenotypeof patients with homozygous FH, but are similarto that of individuals with heterozygous FH,which seems to indicate a positive effect ofenvironmental or genetic factors. The two indexcases with the APOB3500 mutation are the first to

be reported in the Portuguese literature. The 1009


Grupoetrio18

N deCI

estudados38

103

N CI commutaoLDLR

20

58

N CI commutaoAPOB

0

2

N CI commutaoPCSK9

0

2

Grupoetrio18

NCI+F

recebidos38+65

103+284

N CI+Fcom

mutaoLDLR20+2758+86

N CI+Fcom

mutaoAPOB

02+2

N CI+Fcom

mutaoPCSK9

02+2


12/15

Laboratrio da Unidade de InvestigaoCardiovascular, CBP, INSA .

Quatrocentos e noventa indivduos, entrecasos-ndex e familiares, com e sem diagnsticoclnico de FH, esto inscritos no Estudo

Portugus de Hipercolesterolemia Familiar. Oestudo molecular levou identificao de umtotal de 199 indivduos com FH (82 casos--ndex e 117 familiares). S foi possvel realizaro estudo familiar de 2/3 dos indivduos comdiagnstico clnico de FH , e na maioria dos casosapenas da famlia nuclear. A colaborao dosfamiliares parece ser difcil de obter, o que umadas dificuldades do estudo. No obstante, oestudos dos familiares de indivduos afectados sem dvida o mtodo mais custo-efectivo para a

deteco de novos casos de hipercolesterolemiafamiliar e vrios pases comearam j a realizarprogramas de cascade screening(35-37).

Analisando os resultados por gruposetrios, no total diagnosticaram-se 47 crianase adolescentes (menos de 18 anos de idade)com FH heterozigtica e 152 adultos com FH,sendo que 150 tm FH heterozigtica e 2 tmFH homozigtica. Os dois casos-ndex deFH homozigtica so os primeiros descritosna populao portuguesa. Curiosamente

estes indivduos no apresentam o fentipocaracterstico de doentes com FH homozi-gtica, assemelhando-se o seu fentipo ao deindivduos com FH heterozigtica, o que pareceindicar uma modulao positiva por factoresambientais ou genticos. Os dois caso-ndex coma mutao APOB3500 so os primeiros descritosna literatura portuguesa. A mutao encontradano gene PCSK9 de dois indivduos com odiagnstico clnico de FH uma mutao aindano descrita na literatura, sendo actualmenteexclusiva da nossa populao. Os 59 indivduosno quais no foi possvel detectar uma mutaonos trs genes estudados podero ter umamutao noutro gene ainda no descrito oupodem ter hipercolesterolemia polignica oumesmo dislipidemia familiar combinada, j quea histria familiar da maioria destes indivduossugere a presena de uma doena hereditria. Ofacto de no ter sido detectada uma mutaoexistente num dos trs genes estudados umahiptese que no pode ser ignorada, mas osmtodos laboratoriais so cada vez mais fiveis e

a evoluo da tcnica limita bastante estes erros.

mutation found in thePCSK9 gene of two patientswith a clinical diagnosis of FH has not beenpreviously described in the literature, and is todate exclusive to our study population. The 59individuals in whom no mutation was detected inthe three genes studied may have mutations in

other genes that have not yet been identifiedor they may have polygenic hypercholesterolemiaor familial compound hyperlipidemia, since thefamily history of most of these individuals wouldsuggest the presence of inherited disease. Thepossibility that a mutation in one of the threegenes studied was not in fact detected cannotbe discounted, but laboratory methods areincreasingly reliable and advances in thetechnique used mean such errors are veryuncommon.

The results presented here are preliminaryfindings from a wider epidemiological study stillin progress. At present thirteen cliniciansare collaborating in the study, but it is hopedthat all interested physicians will participate sothat a greater number of individuals can benefitfrom genetic diagnosis and the prevalence anddistribution of FH in the Portuguese populationcan be better characterized. The aim of thiswork is to highlight the clinical importanceof distinguishing between hereditary and

non-hereditary hypercholesterolemia, sinceindividuals with the inherited form are exposedto high cholesterol levels from birth andtherefore have an increased risk of prematurecardiovascular disease. Genetic study providesan accurate diagnosis as well as early detectionof FH in relatives, thus enabling appropriatetherapy to be instituted. Prompt andaccurate diagnosis is extremely important incardiovascular disease prevention as it providesthe basis for earlier and/or more aggressivetherapeutic measures, which have been shown tobe effective in reducing cardiovascular morbidityand mortality in adults and children (17, 18, 19).

The study of familial hypercholesterolemiais also extremely important in socioeconomicterms for families and society in general.Early diagnosis and appropriate treatment cansignificantly reduce the number of cardio-vascular events and prevent premature death,which have a heavy impact on families andsociety. Health authorities increasingly stress tothe medical community and the general

population the costs of medical interventions,1010



13/15

Os resultados preliminares agora apresen-tados so os primeiros a ser divulgados de umestudo epidemiolgico mais alargado ainda emcurso. Este estudo at agora contou com acolaborao de 13 mdicos, mas pretende-se quetodos os clnicos interessados participem no

mesmo, para que um maior nmero de indivduospossa usufruir dos benefcios do diagnsticomolecular e a prevalncia e distribuio daFH na populao portuguesa seja melhorcaracterizada. Pretende-se com este trabalhochamar a ateno para a importncia clnicada distino entre uma hipercolesterolemiahereditria e uma hipercolesterolemia nohereditria, uma vez que os indivduos comhipercolesterolemia hereditria esto expostos aconcentraes altas de colesterol desde o

nascimento, havendo por isso um risco acrescidode doena cardiovascular prematura. Odiagnstico molecular da FH permite odiagnstico correcto da mesma e possibilitatambm a identificao precoce de familiarescom FH, permitindo a sua adequada orientaoteraputica. O diagnstico correcto e atempadotem elevada importncia na preveno dadoena cardiovascular, pois fundamenta aintroduo de medidas teraputicas maisprecoces e/ou agressivas, que se tm mostrado

efectivas na reduo da morbilidade emortalidade cardiovascular em adultos ecrianas (17-19).

O estudo da hipercolesterolemia familiartem tambm grande importncia no contextoscio econmico das famlias e da sociedade.O diagnstico atempado e a teraputicaadequada podem reduzir significativamente onmero de eventos cardiovasculares e evitarmortes prematuras, o que tem custos familiarese sociais muito importantes. As autoridades desade chamam cada vez mais a ateno dacomunidade mdica e da populao em geralpara os custos das intervenes em sade, sejamelas farmacolgicas ou outras, j que os recursosso limitados e h que optimiz-los. Um estudode viabilidade econmica efectuado pelo grupoespanhol para o estudo da hipercolesterolemiafamiliar (www.colesterolfamiliar.com) demonstrouser mais custo-efectivo identificar e tratar osindivduos com hipercolesterolemia familiar,durante vrios anos e com teraputica adequadapara baixar os nveis de colesterol, do que no

os tratar, pois mais cedo ou mais tarde estes

pharmacological or otherwise, since resources arelimited and must be optimized. An economicviability study carried out by the Spanishfamilial hypercholesterolemia study group(www.colesterolfamiliar.com) demonstrated thatit is more cost-effective to identify and treat FH

with appropriate cholesterol-lowering therapyover several years than to leave it untreated, sincethese individuals will very probably requirecoronary revascularization sooner or later, as wellas having an increased risk of premature death,thus depriving society of potentially usefulmembers.

The Portuguese Familial Hypercholesterolemia Study is run by

the Cardiovascular Research Unit of the Center for Biopathology of

the Portuguese National Institute of Health, with Dr. Mafalda

Bourbon as coordinator and Dr. Quitria Rato as clinical consultant.Genetic Study: Dr. Mafalda Bourbon (head researcher), Dr. Ana

Catarina Alves (research assistant), Dr. Ana Margarida Medeiros

and Dr. Snia Silva (research trainees) - Cardiovascular Research

Unit, CBP, INSA, Lisbon. Clinical Researchers: Prof. Antnio

Guerra, Serv. de Pediatria, Hosp. de S. Joo, E.P.E., Porto; Dr.

Antnio Furtado, Serv. de Med. Interna, Hosp. Pedro Hispano,

Matosinhos; Prof. Jos Manuel Silva, Serv. de Med. Interna, HUC,

Coimbra; Prof. Graa Morais, Dept. de Bioqumica, Fac. de Cincias

Mdicas, UNL, Lisbon; Prof. Helosa Santos, Dr. Isabel Mendes

Gaspar, Serv. de Gentica, Hosp. de Santa Maria, E.P.E., Lisbon; Dr.

Ana Gaspar, Serv. de Pediatria, Hosp. de Santa Maria, E.P.E.,Lisbon; Dr. Pedro Marques da Silva, Serv. de Med. Interna, Hosp. de

Santa Marta, E.P.E, Lisbon; Dr. Joo Sequeira Duarte, Serv. de

Endocrinologia, Hosp. de Egas Moniz, Lisbon; Dr. Slvia Sequeira,

Serv. Doenas Metablicas, Hosp. D. Estefnia, Lisbon; Dr. Quitria

Rato, Serv. de Cardiologia, Centro Hospitalar de Setbal, E.P.E.,

Hospital de So Bernardo, Setbal; Dr. Mrio Amaro, Serv.

Medicina Interna, Hospital Garcia da Orta, Almada; Dr. Isabel

Azevedo, Serv. de Medicina Interna I, Hosp. dos Marmeleiros,

Funchal.

1011



14/15

indivduos so srios candidatos a revas-cularizao coronria, alm do risco acrescido demorte prematura em indivduos potencialmenteactivos para a sociedade.

O Estudo Portugus de Hipercolesterolemia Familiar

promovido pela Unidade de Investigao Cardiovascular do Centrode Biopatologia do Instituto Nacional de Sade Dr. Ricardo Jorge,

que tem como coordenadora a Doutora Mafalda Bourbon e

consultora clnica a Dra. Quitria Rato. Investigao Molecular:

Doutora Mafalda Bourbon (investigadora responsvel), Dra Ana

Catarina Alves (assistente de investigao), Dra Ana Margarida

Medeiros e Dra Snia Silva (estagirias de investigao) - Unidade

de Investigao Cardiovascular, CBP, INSA Lisboa. Investigao

Clnica: Prof. Doutor Antnio Guerra, Serv. de Pediatria, Hosp. de

S. Joo, E.P.E., Porto; Dr. Antnio Furtado, Serv. de Med. Interna,

Hosp. Pedro Hispano, Matosinhos; Prof. Doutor Jos Manuel Silva,

Serv. de Med. Interna, HUC, Coimbra; Prof Doutora Graa Morais,Dept. de Bioqumica, Fac. de Cincias Mdicas, UNL, Lisboa;

Profa. Helosa Santos, Dra Isabel Mendes Gaspar, Serv. de Gentica,

Hosp. de Santa Maria, E.P.E., Lisboa; Dra. Ana Gaspar Serv. de

Pediatria, Hosp. de Santa Maria, E.P.E., Lisboa; Dr. Pedro Marques

da Silva, Serv. de Med. Interna, Hosp. de Santa Marta, E.P.E,

Lisboa; Dr. Joo Sequeira Duarte, Serv. de Endocrinologia, Hosp. de

Egas Moniz, Lisboa; Dra. Slvia Sequeira, Serv. Doenas

Metablicas, Hosp. D. Estefnia, Lisboa; Dra. Quitria Rato, Serv.

de Cardiologia, Centro Hospitalar de Setbal, E.P.E., Hospital de

So Bernardo, Setbal; Dr. Mrio Amaro, Serv. Medicina Interna,

Hospital Garcia da Orta, Almada; Dra. Isabel Azevedo, Serv. deMedicina Interna I, Hosp. dos Marmeleiros, Funchal.

1012


Pedidos de separatas para:

Address for reprints:

MAFALDA BOURBON

Unidade de Investigao Cardiovascular

Centro de Biopatologia

Instituto Nacional de Sade Dr. Ricardo Jorge

Av. Padre Cruz, 1649-016 LISBOAE-mail: [email protected]

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Estudo Português de Hipercolesterolemia