Hipertensão arterial pulmonar. Fisiopatologia, aspectos genéticos e resposta ao uso crônico do sildenafil
Instituto do Coração do Hospital das Clínicas - FMUSP - São Paulo, SP
Nas últimas décadas, tem havido crescente interesse pelo entendimento dos mecanismos fisiopatológicos operantes na hipertensão pulmonar, sobretudo na hipertensão arterial pulmonar (HAP). Os progressos têm sido mais evidentes com relação à assim chamada HAP idiopática (esporádica e familial, anteriormente chamada de hipertensão pulmonar primária). Genes foram identificados cujas alterações podem estar relacionadas ao desencadeamento da doença (
A hipertensão pulmonar é uma síndrome clínica e hemodinâmica, que resulta no aumento da resistência vascular na pequena circulação, em geral por mecanismos mistos, envolvendo vasoconstrição e remodelagem da parede arterial (arteriolar)1. A classificação mais recente da hipertensão pulmonar2 inclui cinco itens, dos quais o primeiro recebe a designação de hipertensão arterial pulmonar (HAP). Nesta forma de apresentação estão incluídas entidades semelhantes sob o ponto de vista fisiopatológico, como por exemplo a HAP idiopática, a familial e a associada a cardiopatias congênitas.
A compreensão dos mecanismos etiopatogênicos e fisiopatológicos operantes na HAP progrediu significantemente nas últimas décadas e tem sido a razão do desenvolvimento de novos recursos terapêuticos. Assim sendo, o objetivo da presente atualização é apresentar os conceitos mais recentes acerca da fisiopatologia da HAP. Em seguida, são apresentadas duas famílias, com a intenção de ressaltar mecanismos ligados à gênese da doença e ainda outros, implicados com a resposta à intervenção terapêutica.
Há estudos que demonstram uma tendência à vasoconstrição na HAP. O aumento na produção de tromboxane A2, um vasoconstritor, bem como a produção deficiente de prostaciclina, vasodilatador, são achados em pacientes com a forma idiopática da HAP. Um outro exemplo desse desequilíbrio, relacionado à disfunção endotelial, é a demonstração de expressão reduzida da óxido nítrico sintase endotelial (eNOS), e expressão aumentada de endotelina-1 (ET-1), um potente vasoconstritor3,4,5.
A maioria dos eventos que levam à vasoconstrição na HAP, em última análise, fazem-no através do aumento de cálcio nas células musculares lisas vasculares. A forma mais exaustivamente estudada de aumento do tônus na pequena circulação é a vasoconstrição pulmonar por hipóxia. A redução na tensão alveolar de oxigênio promove uma elevação rápida e reversível da resistência vascular pulmonar, que se inicia com menos de um minuto, revertendo rapidamente com a normalização das condições de oxigenação6. O mecanismo que inicia a vasoconstrição é a inibição de vários canais de potássio, designados por Kv, seguida de despolarização da membrana celular e entrada de cálcio através dos canais tipo L. Essa é uma resposta única da vasculatura pulmonar, em contraste à resposta vasodilatadora sistêmica frente à hipóxia. Além disso, há evidências de que os canais de cálcio tipo L possam ser diretamente influenciados pela hipóxia7. A vasoconstrição pulmonar é, em condições normais, um mecanismo adaptativo com a finalidade de equilibrar a ventilação e a perfusão, restringindo o fluxo sangüíneo em áreas pouco ventiladas (como, por exemplo, nas atelectasias) em favor de áreas de melhor ventilação. O mecanismo preciso da vasoconstrição não está bem esclarecido, mas acredita-se no envolvimento de diversos fatores (provavelmente diferentes na hipóxia aguda e crônica) sempre resultando em despolarização da membrana e aumento do cálcio intracelular. Apesar de evidências confirmando esta hipótese, alguns autores crêem ser este apenas um mecanismo facilitador, e não o mediador principal da vasoconstrição7.
Uma teoria complementar para a vasoconstrição pulmonar por hipóxia sugere a participação de mecanismos "redox" na célula muscular lisa. Em condições normais, a geração de espécies reativas de oxigênio (ERO) pelas mitocôndrias manteria um "estado oxidado" dos canais de potássio, que permaneceriam abertos. Durante a hipóxia, o transporte de elétrons mitocondrial está reduzido, bem como a geração de ERO. O conseqüente "estado reduzido" do citosol resultaria na inibição dos canais de potássio e despolarização da membrana celular7,8. De forma alternativa, o sistema "sensor" de oxigênio poderia não estar nas mitocôndrias, mas sim junto à membrana plasmática, representado por NADPH oxidases ou enzimas similares. Outra possibilidade é que a hipóxia module as correntes de potássio nos Kv através da atividade do citocromo P450. Estudos que envolveram sua inibição demonstraram queda na corrente de potássio, causando despolarização da membrana celular8.
Em relação à hipóxia crônica, os mecanismos causadores da vasoconstrição pulmonar podem ser diversos. A atividade dos canais Kv pode estar limitada por redução na transcrição dos seus genes. Em ratos, a hipóxia crônica diminui a expressão de RNAm e proteína da subunidade-a dos canais de potássio em células musculares lisas, causando um decréscimo funcional dos mesmos8. Além disso, a deprivação de oxigênio aumenta a atividade funcional do fator de transcrição induzível por hipóxia (HIF-1). Em condições de tensão normal de oxigênio a subunidade-a deste transativador é degradada. Em hipóxia, porém, a degradação é inibida, havendo acúmulo de HIF-1a, dimerização com HIF-1b e formacão de HIF-1 funcional. Este é responsável pela ativação de genes críticos, como o fator de crescimento endotelial vascular (VEGF) e a ET-18.
Uma característica particular em pacientes com a forma idiopática da HAP é a redução no número de canais de potássio funcionantes. Alterações nos canais de potássio foram encontradas em células musculares lisas destes pacientes, mas não em outras formas de HAP. Comparando-se indivíduos normais com portadores de hipertensão arterial sistêmica, HAP associada a outras doenças e HAP idiopática, verificou-se que as células musculares lisas deste último grupo apresentaram expressão reduzida de subunidades dos canais Kv9. Isto acarretaria um decréscimo nas correntes de potássio, permitindo a entrada de cálcio e vasoconstrição pulmonar sustentada.
A seguir, passamos à discussão de alguns mecanismos relacionados à remodelagem vascular pulmonar. Esses mecanismos envolvem fatores de crescimento, matriz extracelular, receptores de membrana e atividades proteolíticas.
Dentre os fatores de crescimento e seus receptores, a superfamília TGF-b (
O VEGF tem sido amplamente estudado, mais precisamente na HAP induzida por hipóxia. Liga-se com alta afinidade a dois receptores do tipo tirosina-quinase: VEGFR-1 e VEGFR-2. Por ser mitógeno de células endoteliais, está envolvido no crescimento das mesmas, por exemplo, no interior das assim chamadas lesões plexiformes12. A hipóxia crônica, que acompanha algumas formas de HAP, cursa com aumento da expressão do VEGF e de seus receptores13. Outros fatores de crescimento também são influenciados pela hipóxia, como o fator de crescimento derivado de plaquetas (PDGF), de fibroblastos (FGF-2), insulina-símile (IGF-1) e o fator de crescimento epidérmico (EGF).
O óxido nítrico (NO), um vasodilatador derivado do endotélio, também tem ação antiproliferativa na musculatura lisa vascular14, possivelmente ligada à geração de GMP cíclico intracelular. Com efeito, tem sido cogitado que inibidores da enzima fosfodiesterase 5 (que degrada GMP cíclico) poderiam atuar sobre o remodelamento vascular pulmonar induzido por hipóxia15. Esta ação protetora do GMP cíclico estaria relacionada à via das MAPP quinases e à inibição do fator de transcrição AML1 (
A prostaciclina, semelhantemente ao NO, é um potente vasodilatador e inibidor da adesão plaquetária e crescimento celular. É o principal metabólito do ácido araquidônico no endotélio vascular, com ação na célula muscular lisa, através do aumento do AMP cíclico. Além disso, a prostaciclina inibe o efeito da endotelina-1 (ET-1). Pacientes com HAP apresentam síntese e excreção urinária diminuídas da prostaciclina4.
O papel da serotonina na HAP está relacionado à indução de hiperplasia e hipertrofia de células musculares lisas e efeito mitogênico também em células endoteliais e fibroblastos. A ação da serotonina na circulação pulmonar é ampla e envolve também a vasoconstrição, por exemplo, em resposta à hipóxia17. Níveis plasmáticos aumentados de serotonina são observados em algumas formas de HAP18. Além disso, a expressão aumentada de seu transportador poderia estar relacionada a acúmulo de serotonina intracelular19.
A ET-1 é um potente vasoconstritor produzido, predominantemente, pela célula endotelial, com ação mitogênica, inflamatória e de proliferação sobre a célula muscular lisa. Apesar de não existir estoques intracelulares de ET-1, estímulos específicos induzem sua síntese e secreção em minutos. Há dois tipos de receptores de endotelina: ETA (encontrados em células musculares lisas e miócitos cardíacos) e ETB (encontrados em células endoteliais e musculares lisas). A ligação da ET-1 aos seus receptores ativa a enzima fosfolipase C, aumentando os níveis intracelulares de cálcio, o que resulta em vasoconstrição prolongada. O diacil-glicerol, produzido a partir da fosfolipase C ativada, estimula a proteína-quinase C, enzima mediadora da ação mitogênica da ET-120. Semelhantemente à serotonina, a ação da ET-1 é potencializada na presença de outros fatores de crescimento21. Outros efeitos relevantes da ET-1 são a estimulação da produção de citocinas e fatores de crescimento22, a indução da formação de proteínas da matriz extracelular23 e a potencialização dos efeitos do TGF-b e PDGF21. Estudos em pulmões de ratos normais demonstraram níveis de RNAm da ET-1 cinco vezes maiores se comparados com qualquer outro órgão24. Em condições normais, os pulmões retiram ET-1 da circulação. Na HAP, os pulmões sintetizam ET-1 em quantidades aumentadas, o que resulta em aumento dos níveis circulantes, com implicações prognósticas3. Nessas condições, a circulação pulmonar passa a ser alvo desse peptídeo.
Além da atuação dos fatores de crescimento e outras moléculas sinalizadoras na gênese do remodelamento vascular pulmonar, eventos envolvendo componentes da matriz extracelular são de extrema relevância (
Os mecanismos descritos acima, de maneira resumida, constituem, na verdade, um modelo proposto de remodelamento vascular pulmonar a partir de vários estudos experimentais1. Compreende-se desta forma que o remodelamento vascular na HAP é um processo complexo que muito provavelmente envolve fatores séricos, indução de proteases, modificações da matriz extracelular, disponibilização de fatores de crescimento, ativação de seus receptores e, finalmente, mitose. Além disso, muitas alterações envolvendo matriz extracelular, integrinas e fatores de crescimento, contribuem para a alteração do fenótipo das células musculares lisas em vasos pulmonares, que ao perderem a característica contrátil, transformam-se em células secretoras e capazes de se movimentar em direção à camada íntima1.
Além da participação expressiva do endotélio nas alterações do tônus e no remodelamento vascular pulmonar presentes na HAP, seu envolvimento no processo vaso-oclusivo propriamente dito torna-se relevante, uma vez que há perda progressiva de suas características anticoagulantes, antitrombóticas e antiaderentes, e desencadeamento de mecanismos que facilitam a coagulação e a adesão celular e inibem a fibrinólise.
Entre as substâncias de origem endotelial, que têm se mostrado alteradas na HAP com implicações nos mecanismos descritos, encontram-se a trombomodulina, o ativador tecidual do plasminogênio (t-PA), seu inibidor (PAI-1), a selectina-P e o fator de Von Willebrand (FvW), além do óxido nítrico e da prostaciclina. Além disso, têm sido identificadas outras alterações pró-trombóticas, como a presença de anticorpos antifosfolípides e a hiperhomocisteinemia.
A trombomodulina é um proteoglicano de membrana plasmática. Sua porção extracelular é clivada em várias regiões, com conseqüente liberação de fragmentos na corrente sangüínea, cuja concentração pode ser facilmente determinada. Esses fragmentos são conhecidos como a fração solúvel. A trombomodulina tem propriedades de ligação à trombina, o que ativa a proteína C que, por sua vez, degrada os fatores V e VIII da coagulação26. Em pacientes com HAP, os níveis plasmáticos de trombomodulina estão diminuídos, provavelmente como resultado de síntese endotelial reduzida26,27. Além disso, há relatos que demonstram baixos níveis plasmáticos de proteína C, assim como seu co-fator, a proteína S28. Essas alterações podem refletir uma síntese deficiente ou mesmo consumo por coagulação intravascular.
Pacientes com a síndrome antifosfolípide, associada ou não ao
A homocisteína é um aminoácido que, em indivíduos normais, pode facilitar a coagulação e adesão celular. Sabe-se que a presença de níveis plasmáticos elevados de homocisteína está associada à disfunção endotelial e plaquetária, com conseqüente formação de trombos e vasoconstrição34. Sugere-se que a hiperhomocisteinemia possa exercer algum um papel na patogênese da HAP, pois há referência a aumento dos níveis deste aminoácido em pacientes com a doença34.
Uma forma particular de disfunção endotelial em pacientes com HAP é aquela associada à hipoxemia, dada a acentuada tendência à trombose. Isto ocorre na assim chamada síndrome de Eisenmenger, onde, em virtude da presença de defeitos intracardíacos com desvio de sangue da direita para a esquerda, desenvolve-se a hipoxemia crônica. Alterações na coagulação e fibrinólise nesses pacientes associam-se a um alto risco de eventos tromboembólicos, havendo relato de até 35% de ocorrência de trombose arterial pulmonar35. Contribui para isto o fato de a hipóxia promover exocitose dos corpúsculos de Weibel-Palade nas células endoteliais, resultando em liberação do FvW e da selectina-P. Além disso, sabe-se que a hipóxia induz a expressão do RNAm e da proteína do PAI-1, além de suprimir a expressão de trombomodulina. Portanto, na HAP associada à hipoxemia crônica é justificável imaginar que mecanismos como o recrutamento de leucócitos e plaquetas, a inibição da fibrinólise e a perda de controle sobre a coagulação, sejam operantes e contribuam para o evento vaso-oclusivo.
Além de fornecer estimativa do risco trombótico em indivíduos com HAP, os marcadores de disfunção endotelial parecem ter implicações prognósticas. Estudo em pacientes com HAP idiopática e secundária a cardiopatias congênitas correlacionou os níveis elevados do FvW com redução da sobrevida36. Pacientes com a forma idiopática da doença apresentaram níveis mais altos, associados a menor expectativa de vida37. Outros marcadores bioquímicos também estão sendo estudados, com finalidade prognóstica, como as catecolaminas, o ácido úrico e os peptídeos natriuréticos (ANP e BNP). Apenas o BNP, porém, parece ter correlação com a sobrevida, com níveis elevados em pacientes com HAP idiopática38.
A disfunção endotelial na HAP leva a um estado pró-trombótico, no qual a participação das plaquetas parece inequívoca. Estas têm sido alvo de estudos há vários anos, pois seu papel na HAP é mais que meramente trombogênico, havendo envolvimento também no processo de remodelamento vascular.
Estudos clínicos têm trazido evidências de ativação plaquetária endógena em pacientes com HAP, sugerindo que haja um estado crônico de consumo. Essa sugestão deve-se à observação da redução do número de plaquetas circulantes e níveis elevados de b-tromboglobulina39, uma substância liberada na fase de secreção. Sugere-se que esta ativação favoreça a formação de agregados celulares na circulação, envolvendo plaquetas entre si ou em associação com leucócitos e hemácias40, o que poderia nos vasos pulmonares, contribuir para a progressão da doença. A ativação plaquetária na HAP também promove a liberação de grânulos densos, contendo agentes mitogênicos e substâncias vasoconstritoras. Uma dessas substâncias é o tromboxane A2, potente vasoconstritor cuja produção está elevada em pacientes com HAP41. Também são liberados serotonina e fatores de crescimento, como PDGF, EGF e TGF-b, com efeito mitogenico em células musculares lisas vasculares, fibroblastos e células endoteliais42.
Em diferentes formas de HAP, o envolvimento (ativação) plaquetário tem sido considerado como evento secundário à disfunção endotelial, exposição do subendotélio e alterações das condições de fluxo. Exceção faz-se à situação identificada como doença de estoque das plaquetas (
A serotonina é o principal componente dos grânulos densos das plaquetas, sendo liberada a partir de sua ativação. A resposta normal da célula endotelial à serotonina é a liberação de NO, levando a um relaxamento da musculatura lisa e conseqüente vasodilatação44. Porém, em casos de disfunção endotelial, a serotonina tornar-se-ia incapaz de estimular a liberação de NO. Na circulação pulmonar, essa ocorrência favoreceria o desenvolvimento de HAP por mecanismos diversos. A serotonina, ou 5-hidroxitriptamina (5-HT), exerce sua função através da ligação ao seu receptor 5-HTR, e seu transportador 5-HTT. Ao ligar-se ao 5-HTR, a serotonina ativa a produção de IP3, induzindo a liberação de cálcio intracelular do retículo sarcoplasmático, com conseqüente vasoconstrição45. Por outro lado, a geração de diacilglicerol está implicada na indução da via mitogênica da proteína-quinase C. A ligação da serotonina ao 5-HTT inicia eventos intracelulares, envolvendo a formação de espécies reativas de oxigênio e a ativação da via das MAPP quinases, resultando na expressão de genes envolvidos em hipertrofia e proliferação celular46. Vários estudos sugerem que a serotonina tenha um papel na patogênese da HAP, inclusive na forma idiopática. Já foi demonstrada a presença de polimorfismo na região promotora do gene que codifica o 5-HTT, com expressão aumentada do mesmo19. Assim, esse gene é hoje considerado como "facilitador" no processo de remodelagem vascular pulmonar.
Uma importante condição clínica, ligada ao metabolismo da serotonina e potencialmente causadora de HAP, é o uso de anorexígenos orais, como a fenfluramina. No sistema nervoso central, o efeito seria exercido por inibição do 5-HTT e, conseqüentemente, da recaptura da serotonina em neurônios. Na periferia, também por interferir com o transportador, a fenfluramina é capaz de impedir a captura da serotonina nas plaquetas e em células endoteliais pulmonares. As conseqüências são o aumento da serotonina livre, de forma semelhante ao que ocorre na
Como citado anteriormente, mutações no gene que codifica o BMPR2 alteram sua sinalização que, em condições normais. suprimiria a proliferação e favoreceria a apoptose de células musculares lisas de artérias pulmonares. Sugere-se que a sinalização a partir do BMPR2 poderia antagonizar os efeitos da serotonina e que mutações nesse receptor permitiriam uma resposta celular exacerbada à mesma, principalmente nos pacientes com polimorfismo do 5-HTT47. Essa relação porém é, por enquanto, meramente especulativa.
A HAP é uma complicação comumente encontrada em várias condições inflamatórias sistêmicas, tais como a esclerodermia e o
Além dos achados inflamatórios, vale a pena enfatizar a associação da HAP com doenças ou condições de caráter imune: infecção pelo vírus da imunodeficiência humana48, síndrome POEMS (discrasia de célula plasmática com polineuropatia, organomegalia, endocrinopatia, proteína M e alterações de pele)50, presença de anticorpos auto-imunes51 e fenômeno de Raynaud52. Os principais aspectos imunogenéticos nessas doenças correspondem à presença de auto-anticorpos e de certos alelos do complexo de histocompatibilidade (MHC,
Na tentativa de identificar mecanismos que levam à condição de inflamação encontrada na HAP, observou-se aumento de vários mediadores. Já foi demonstrado que pacientes com HAP idiopática apresentam níveis séricos elevados de IL-1 e IL-656. Outro achado importante é o envolvimento de quimiocinas, que são citocinas quimiotáticas capazes de direcionar o recrutamento e a migração de leucócitos. Uma dessas quimiocinas, designada por RANTES (
A fractalcina é uma quimiocina que também parece ter papel importante na HAP: promove o recrutamento de leucócitos através do receptor transmembrana CX3CR1, com captura de forma rápida e adesão dependente de integrinas. Demonstrou-se em pacientes com HAP elevação dos níveis plasmáticos da fractalcina, aumento do seu RNAm, assim como aumento da expressão e função do receptor CX3CR1 em linfócitos59.
Outra quimiocina que pode estar envolvida no processo de inflamação da HAP idiopática é MCP-1 (
Apesar dos mecanismos inflamatórios estarem estabelecidos em HAP idiopática e associada a doenças do tecido conectivo, ainda há controvérsia sobre seu papel na HAP associada a cardiopatias congênitas. Em relato recente sobre a presença de células inflamatórias em artérias pulmonares periféricas de pacientes com HAP associada a cardiopatias congênitas, foi detectada predominância de macrófagos jovens nas camadas íntima e média, sugerindo reação inflamatória em curso62. Sendo os macrófagos responsáveis pela produção de várias citocinas, especula-se que possam ter participação relevante em eventos mediados por essas substâncias (por exemplo, replicação celular) em pacientes com HAP.
Tendo sido apresentada a visão atual acerca dos mecanismos fisiopatológicos mais relevantes que operam na HAP, passamos à apresentação de duas famílias com a doença.
O estudo genético foi realizado em cinco indivíduos desta família, conforme demonstrado no heredograma. O estudo consistiu na amplificação, pela reação da polimerase em cadeia (PCR), dos 13 exons do gene que codifica o receptor BMPR2. O produto da reação foi então submetido a sequenciamento por método automatizado. Conforme demonstrado na
O paciente DED, 15 anos de idade, encontrava-se em classe funcional IV na admissão, com pressão média na pulmonar de 62 mmHg, impossibilitado de percorrer qualquer distância no teste de 6 minutos de caminhada. Após início de tratamento com sildenafil por via oral, na dose de 225 mg ao dia, observou-se melhora importante na capacidade física, passando a caminhar 399 m e 471 m, respectivamente, aos 6 meses e 1 ano. Observou-se também incremento do fluxo pulmonar, estimado através do ecocardiograma, de 50% em 6 meses e 38% em 1 ano. Sua irmã (AKD), 18 anos de idade na admissão, teve manifestações (cansaço) de menor intensidade e início mais tardio. Iniciou o tratamento em classe funcional II, com pressão média pulmonar de 50 mmHg e com distância percorrida de 309 m no teste de 6 minutos de caminhada. Sob o uso do sildenafil (225 mg/dia), houve aumento na distância percorrida para 504 m aos 6 meses e 1 ano. Também foi observado incremento de 26% no fluxo pulmonar.
O paciente EMB, 33 anos de idade, encontrava-se em classe funcional IV na admissão, chegando a fazer uso de drogas vasoativas. A presença de estertores pulmonares, com desaparecimento mediante terapêutica diurética, fez suspeitar de doença veno-oclusiva, diagnóstico posteriormente confirmado por estudo hemodinâmico e biópsia pulmonar.
A avaliação deste paciente incluiu o teste agudo de vasodilatação com óxido nítrico (10 ppm) e sildenafil (75 mg por via oral). O teste demonstrou vasodilatação pulmonar significante com ambos os estímulos aplicados separadamente, havendo queda expressiva da resistência vascular (de 27,9 unidades Wood/m2, em condição basal, para 8,3 e 11,1 U/m2, respectivamente, com óxido nítrico e sildenafil). Entretanto, a pressão média capilar pulmonar estabilizou-se em nível mais seguro com sildenafil, comparado ao óxido nítrico, com valores de 15 e 29 mmHg, respectivamente. Os resultados deste estudo permitiram a orientação do uso crônico do sildenafil (225 mg/dia) associado a diuréticos, com melhora clínica evidente após um ano, sendo possível retirar o paciente da fila de espera para o transplante pulmonar. A distância percorrida no teste de caminhada passou de 112 m, na condição pré-tratamento, para 294 m e 408 m, em 6 meses e 1 ano, respectivamente. O ecocardiograma também mostrou incremento do fluxo pulmonar de 50% e 120%.
Estas observações ressaltam alguns aspectos importantes da HAP. Em primeiro lugar, o diagnóstico da forma familial, que na prática clínica possibilitaria a identificação da doença em estágios menos avançado, em indivíduos com sintomatologia discreta ou ausente. Em segundo lugar, a importância das mutações no gene BMPR2 no cenário fisiopatológico, com a possibilidade de se identificar tais alterações gênicas em pelo menos 50% das famílias acometidas. Por último, o papel fisiopatológico do óxido nítrico e das vias a ele relacionadas, como justificativa para a aplicação racional de recursos terapêuticos com capacidade de aumentar os níveis intracelulares de GMP cíclico em arteríolas pulmonares.
Em vista dos mecanismos discutidos, fica claro que a HAP é uma síndrome cujo início e progressão se processa de maneira diversa, dependendo do caso. Não é possível imaginar o mesmo mecanismo ou conjunto de mecanismos fisiopatológicos explicando a vasoconstrição e a evolução da remodelagem vascular em todas as formas de HAP. Além disso, mesmo se considerarmos uma única forma da doença, os mecanismos fisiopatológicos envolvidos podem variar de acordo com a fase evolutiva. Assim, percebe-se as dificuldades no desenvolvimento de recursos terapêuticos utilizáveis em humanos, uma vez que isso dependeria da clara definição dos mecanismos operantes em um caso particular ou ainda da identificação de alterações comuns a vários pacientes com diferentes formas da doença. Mesmo assim, as investigações em fisiopatologia da HAP continuam sendo relevantes. Sem elas, aquisições terapêuticas como os prostanóides, os inibidores de receptores de endotelina e de fosfodiesterases, hoje em uso, não teriam sido possíveis.
AML-1, leucemia mielógena aguda-1; AMP, monofosfato de adenina; ANP, peptídeo atrial natriurético; Apo A-1, apolipoproteína A1; BMP, proteína morfogenética óssea; BMPR2, receptor 2 do BMP; BNP, peptídeo cerebral natriurético; EGF, fator de crescimento epidérmico; ERO, espécies reativas de oxigênio; ERK, quinase com regulação extracelular; ET-1, endotelina-1; EVE, elastase vascular endógena; FGF-2, fator de crescimento de fibroblastos; FvW, fator de von Willebrand; GMP, monofosfato de guanina; HAP, hipertensão arterial pulmonar; HIF-1, fator induzível por hipóxia; 5-HT, 5-hidroxitriptamina; 5-HTR, receptor da 5-hidroxitriptamina; 5-HTT, transportador da 5-hidroxitriptamina; IGF-1, fator de crescimento insulina-símile; IL-1, interleucina-1; IL-6, interleucina-6; Kv, canal de potássio dependente de voltagem; MCP-1, proteína quimiotática de monócitos; MHC, complexo de histocompatibilidade principal; NO, óxido nítrico; NOS, óxido nítrico sintase; PAI-1, inibidor do ativador do plasminogênio; PDGF, fator de crescimento derivado de plaquetas; PKC, proteína-quinase C; POEMS (síndrome), discrasia de célula plasmática, polineuropatia, organomegalia,endocrinopatia, proteína M e alterações de pele; TGF-b, fator de crescimento transformador do tipo beta; TN-C, tenascina-C; t-PA, ativador tecidual do plasminogenio; VEGF, fator de crescimento endotelial vascular; VEGFR, receptor do fator de crescimento endotelial vascular.
1. Rabinovitch M. Pathobiology of pulmonary hypertension. Extracellular matrix. Clin Chest Med 2001; 22: 433-449.
2. Simonneau G, Galiè N, Rubin LJ et al. Clinical classification of pulmonary hypertension. J Am Coll Cardiol 2004, 43: 5S-12S.
3. Galiè N, Griggione F, Bacchi-Reggiani L et al. Relation of endothelin-1 to survival in primary pulmonary hypertension. Eur J Clin Invest 1996; 26: 273.
4. Tuder RM Cool CD, Geraci MW et al. Prostacyclin synthase expression is decreased in lungs from patients with severe pulmonary hypertension. Am J Respir Crit Care Med 1999; 159: 1925-32.
5. Giaid A, Saleh D. Reduced expression of endothelial nitric oxide synthase in the lungs of patients with pulmonary hypertension. N Engl J Med 1995; 333: 214-21.
6. Archer S, Wu X, Thébaud B et al. Preferential expression and function of voltage-gated, O2-sensitive K+ channels in resistance pulmonary arteries explains regional heterogeneity in hypoxic pulmonary vasoconstriction. Circ Res 2004; 95: 308-18.
7. Ward JPT, Snetkov VA, Aaronson PI. Calcium, mitochondria and oxygen sensing in the pulmonary circulation. Cell Calcium 2004; 36: 209-20.
8. Mandegar M, Remillard CV, Yuan JXJ. Ion channels in pulmonary arterial hypertension. Prog Cardiov Dis 2002; 45: 81-114.
9. Yuan X-J, Wang J, Juhaszova M et al. Attenuated K+ channel gene transcription in primary pulmonary hypertension. Lancet 1998; 351: 726-7.
10. Lane KB, Machado RD, Pauciulo MW et al. Heterozigous germline mutations in a TGF-b receptor, BMPR2, are the cause of familial primary pulmonary hypertension. The International PPH Consortium. Nat Genet 2000; 26: 81-4.
11. Atkinson C, Stewart S, Upton PD et al. Primary pulmonary hypertension is associated with reduced pulmonary vascular expression of type II bone morphogenetic protein receptor. Circulation 2002; 105: 1672-8.
12. Christman BW, McPherson CD, Newman JH et al. An imbalance between the excretion of thromboxane and prostacyclin metabolites in pulmonary hypertension. N Engl J Med 1992; 372: 70-5.
13. Hirose S, Hosoda Y, Furuya S et al. Expression of vascular endothelial growth factor and its receptor correlates closely with formation of the plexiform lesion in human pulmonary hypertension. Pathol Int 2000; 50: 472-9.
14. Gillespie MN, Rippetoe PE, Haven AC et al. Polyamines and epidermal growth factor in monocrotaline-induced pulmonary hypertension. Am Rev Respir Dis 1989; 140: 1463-6.
15. Walford G, Loscalzo J. Nitric oxide in vascular biology. J Thromb Haemost 2003; 1: 2112-18.
16. Rabinovitch M. EVE and beyond, retro and prospective insights. Am J Physiol 1999; 277: L5-L12.
17. Launay JM, Hervé P, Peoc'h K et al. Function of the serotonin 5-hydroxytryptamine 2B receptor in pulmonary hypertension. Nat Med 2002; 8: 1129-35.
18. Egermayer P, Town GI, Peacock AJ. Role of serotonin in the pathogenesis of acute and chronic pulmonary hypertension. Thorax 1999; 54: 161-8.
19. Eddahibi S, Humbert M, Fadel E et al. Hyperplasia of pulmonary artery smooth muscle cells is causally related to overexpression of the serotonin transporter in primary pulmonary hypertension. Chest 2002; 121: 97S-98S.
20. Assender JW, Irenius E, Fredholm BB. Endothelin-1 causes a prolonged protein kinase C activation and acts as a co-mitogen in vascular smooth muscle cells. Acta Physiol Scand 1996; 157: 451-60.
21. Weissberg PL, Witchel C, Davenport AD et al. The endothelin peptides ET-1, ET-2, ET-3 and sarafatoxin S6c are comitogenic with platelet derived growth factor for vascular smooth muscle cells. Atherosclerosis 1990; 83: 257-62.
22. Agui T, Xin X, Cai Y et al. Stimulation of interleukin-6 production by endothelin in rat bone marrow-derived stromal cells. Blood 1994; 84: 2531-8.
23. Peifley KA, Winkless JA. Angiotensin II and endothelin-1 increase fibroblast growth factor-2 mRNA expression in vascular smooth muscle cells. Biochem Biophys Res Commun 1998; 242: 202-08.
24. Matsumoto H, Suzuki N, Onda H et al. Abundance of endothelin-3 in rat intestine, pituitary gland and brain. Biochem Biophys Res Commun 1989; 164: 74-80.
25. Rabinovitch M. Elastase and the pathobiology of unexplained pulmonary hypertension. Chest 1998; 114: 213S-224S.
26. Cacoub P, Karmochkine M, Dorent R et al. Plasma levels of thrombomodulin in pulmonary hypertension. Am J Med 1996; 101: 160-4.
27. Sakamaki F, Kyotani S, Nagaya N et al. Increased plasma P-selectin and decreased thrombomodulin in pulmonary arterial hypertension were improved by continuous prostacyclin therapy. Circulation 2000; 102: 2720-5.
28. Maeda NY, Lopes AA, Da Rocha TR et al. Plasma anticoagulant system in patients with pulmonary hypertension. Braz J Med Biol Res 1990; 23: 251-4.
29. Esmon NL, Safa O, Smirnov MD et al. Antiphospholipid antibodies and the protein C pathway. J Autoimmun 2000; 15: 221-5.
30. Meroni PL, Riboldi P. Pathogenic mechanism mediating antiphospholipid syndrome. Curr Opin Rheumatol 2001; 13: 377-82.
31. Espinosa G, Cervera R, Font J et al. The lung in the antiphospholipid syndrome. Ann Rheum Dis 2002; 61: 195-8.
32. Moser KM, Auger WR, Fedullo PF. Chronic major-vessel thromboembolic pulmonary hypertension. Circulation 1990; 81: 1735-43.
33. Asherson RA, Higenbottan TW, Dinh Xuan AT et al. Pulmonary hypertension in a lupus clinic: experience with twenty-four patients. J Rheumatol 1990; 17: 1292-8.
34. Arroliga AC, Sandur S, Jacobsen D et al. Association between hyperhomocysteinemia and primary pulmonary hypertension. Respir Med 2003; 97: 825-9.
35. Caramuru LH, Maeda NY, Bydlowski SD et al. Age-dependent likelihood of
36. Lopes AA, Maeda NY, Gonçalves RC et al. Endothelial cell dysfunction correlates differentially with survival in primary and secondary pulmonary hypertension. Am Heart J 2000; 139: 618-23.
37. Lopes AA, Maeda NY. Circulating von Willebrand factor antigen as a predictor of short-term prognosis in pulmonary hypertension. Chest 1998; 114: 1276-82.
38. Nagaya N, Nishikimi T, Uematsu M et al. Plasma brain natriuretic peptide as a prognostic indicador in patients with primary pulmonary hipertensión. Circulation 2000;102: 865-70.
39. Caramuru LH, Soares RPS, Maeda NY et al. Hypoxia and altered platelet behavior influence Von Willebrand factor multimeric composition in secondary pulmonary hypertension. Clin Appl Thromb Hemost 2003; 9: 251-8.
40. Lopes AA, Maeda NY, Almeida A et al. Circulating platelets aggregates indicative of in vivo platelet activation in pulmonary hypertension. Angiology 1993; 44: 701-06.
41. Christman BW, McPherson CD, Newman JH et al. An imbalance between the excretion of thromboxane and prostacyclin metabolites in pulmonary hypertension. N Engl J Med 1992; 327: 70-5.
42. Pakala R, Willerson JT, Benedict CR. Mitogenic effect of serotonin on vascular endothelial cells. Circulation 1994; 90: 1919-26.
43. Herve P, Drouet L, Launay JM et al. Primary pulmonary hypertension in a patient with a familial platelet storage pool disease: rolo of serotonin. Am J Med 1990; 89: 117-20.
44. Flavahan NA, Shimokawa H, Vanhoutte PM. Pertussis toxin inhibits endothelium-dependent relaxations to certain agonists in porcine coronary arteries. J Physiol (Lond) 1989; 408: 549-60.
45. Boulanger C, Lorenz RR, Hendrickson H et al. Release of different relaxing factors by porcine endothelial cells in culture. FASEB 1998; 2: A314.
46. Eddahibi S, Fabre V, Boni C et al. Induction of serotonin transporter by hypoxia in pulmonary vascular smooth muscle – relationship with the mitogenic ction of serotonin. Circ Res 1999; 84: 329-36.
47. Rothman RB, Ayestas MA, Dersch CM et al. Aminorex, fenfluramine and chlorphentermine are serotonin transporter substrates. Implications for primary pulmonary hypertension. Circulation 1999; 100: 869-75.
48. Cool CD, Kennedy D, Voelkel NF et al. Pathogenesis and evolution of plexiform lesions in pulmonary hypertension associated with scleroderma and human immunodeficiency virus infection. Human Pathol 1997; 28: 434-42.
49. Tuder RM, Groves B, Badesch DB et al. Exuberant endothelial cell growth and elements of inflammation are present in plexiform lesions of pulmonary hypertension. Am J Pathol 1994; 144: 275-85.
50. Lesprit P, Godeau B, Authier FJ et al. Pulmonary hypertension in POEMS syndrome: a new feature mediated by cytokines. Am J Respir Crit Care Med 1998; 157: 907-11.
51. Isern RA, Yaneva M, Weiner E et al. Autoantibodies in patients with primary pulmonary hipertensión: association with anti-Ku. Am J Med 1992; 93: 307-12.
52. Wigley FM. Raynaud's phenomenon and other features of scleroderma, including pulmonary hypertension. Curr Opin Rheumatol 1996; 8: 561-8.
53. Langevitz PD, Buskila D, Gladman DD et al. HLA alleles in systemic sclerosis: association with pulmonary hypertension and outcome. Br J Rheumatol 1992; 31: 609-13.
54. Barst RJ, Flaster ER, Menon A et al. Evidence for the association of unexplained pulmonary hypertension in children with the major histocompatibility complex. Circulation 1992; 85: 249-58.
55. Morse JH, Barst RJ, Fotino M et al. Primary pulmonary hypertension, tissue plasminogen activator antibodies, and HLA-DQ7. Am J Respir Crit Care Med 1997; 155: 274-8.
56. Humbert M, Monti G, Breno TF et al. Increased interleukin-1 and interleukin-6 serum concentrations in severe primary pulmonary hypertension. Am J Respir Crit Care Med 1995; 151: 1628-31.
57. Molet S, Furukawa K, Maghazechi A et al. Chemokine and cytokine-induced expression of endothelin-1 and endothelin converting enzyme-1 in endothelial cells. J Allergy Clin Immunol 2000; 105: 333-8.
58. Dorfmüller P, Zarka V, Durand-Gasselin I et al. Chemokine RANTES in severe pulmonary arterial hypertension. Am J Respir Crit Care Med 2002; 165: 534-9.
59. Balabanian K, Foussat A, Dorfmüller P et al. CX3C chemokine fractalkine in pulmonary arterial hypertension. Am J Respir Crit Care Med 2002; 165: 1-8.
60. Sica A, Wang JM, Colotta F et al. Monocyte chemotatic and activating factor gene expression induced in endothelial cells by IL-1 and tumor necrosis factor. J Immunol 1990; 144: 3034-8.
61. Hashimoto K, Nakamura K, Fugio H et al. Epoprostenol therapy decreases elevated circulating levels of monocyte chemoattractant protein-1 in patients with primary pulmonary hypertension. Circ J 2004; 68: 227-31.
62. Pinto RFA, Higuchi ML, Aiello VD. Decreased numbers of T-lymphocytes and predominance of recently recruited macrophages in the walls of peripheral pulmonary arteries from 26 patients with pulmonary hypertension secondary to congenital cardiac shunts. Cardiovasc Pathol 2004, 13: 268-75.
Antonio Augusto Lopes InCor - Cardiologia Pediátrica e Cardiopatias Congênitas do Adulto Av. Dr. Eneas de Carvalho Aguiar, 44 05403-000 - São Paulo, SP E-mail:
Enviado em: 02/03/2005
Aceito em: 04/04/2005
Pulmonary arterial hypertension. Pathophysiology, genetic aspects and response to the chronic use of sildenafil
Instituto do Coração do Hospital das Clínicas - FMUSP - São Paulo, SP - Brazil
In the last decades there has been a growing interest for the understanding of pathophysiological mechanisms of pulmonary hypertension, especially in pulmonary arterial hypertension (PAH). Progresses have been more evident concerning the so-called idiopathic PAH (sporadic and familial, previously called primary pulmonary hypertension). Genes were identified whose changes may be related to the disease unleashing (bone morphogenetic protein type 2 receptor, BMPR2). Other genes have been identified as facilitators (serotonin transporter). Besides, from studies in animal models and cellular culture, a series of biological changes have been identified. Such changes involve endothelial cells, smooth muscle cells, fibroblasts, extracellular matrix, pericellular proteolysis, growth factors and their receptors, among others. The present review aims at identifying the main pathophysiological changes known at PAH so far. Besides, two PAH families are shown with the objective of emphasizing the importance of genetic diagnosis and draw the attention to signaling pathways (nitric oxide/cyclic GMP), currently used in therapeutic interventions (sildenafil, chronic use).
Pulmonary hypertension is a clinical and hemodynamic syndrome, which results in the increase of vascular resistance in lungs, generally through mixed mechanisms, involving vasoconstriction and remodeling of arterial (arteriolar)1 wall. The most recent classification of pulmonary hypertension2 includes five items, from which the first is called pulmonary arterial hypertension (PAH). In such way of presentation similar entities under the pathophysiological point of view are included, such as, for instance, idiopathic PAH, familial PAH and that associated with congenital heart diseases.
The understanding of pathophysiological mechanisms operating in PAH has significantly progressed in the last decades and has been the reason for the development of new therapeutic resources. So, the objective of the current updating is to show the most recent concepts on PAH pathophysiology. Afterwards two families are shown, with the aim of emphasizing mechanisms connected to disease's genesis and others, implied with response to therapeutic intervention.
There are studies demonstrating a trend to vasoconstriction in PAH. The increase in production of thromboxane A2, a vasoconstrictor, as well as the insufficient production of prostacyclin, a vasodilator, are found in patients with idiopathic PAH. Another example of that imbalance, related to endothelial dysfunction, is the demonstration of the reduced expression of endothelial nitric oxide synthase (eNOS), and the increased expression of endothelin-1 (ET-1), a powerful vasoconstrictor3-5.
Most of the events that lead to vasoconstriction in PAH, in last analysis, do it through the increase of calcium in vascular smooth muscle cells. The most studied model of tonus increase in the pulmonary circulation is the vasoconstriction due to hypoxia. The reduction in alveolar oxygen tension promotes a fast and reversible increase of pulmonary vascular resistance, which starts in less than one minute, quickly reversing with the normalization of oxygenation conditions6. The mechanism starting vasoconstriction is the inhibition of many potassium channels, designated by Kv, followed by cellular membrane depolarization and entry of calcium through L-type channels. That is a unique response of pulmonary vasculature, as opposed to systemic vasodilation against hypoxia7. Pulmonary vasoconstriction is, under normal conditions, an adaptative mechanism with the aim of balancing ventilation and perfusion, restricting blood flow in low ventilated areas (as, for example, in atelectasis) in favor of better ventilated areas. The accurate mechanism of vasoconstriction is not very clear, but it is believed to involve many factors (probably different in acute and chronic hypoxia), always resulting in depolarization of membrane and increase of intracellular calcium. Despite evidences confirming such hypothesis, some authors believe that it is only a facilitating mechanism7.
A complementary theory for pulmonary vasoconstriction due to hypoxia suggests the participation of redox mechanisms in smooth muscle cells. Under normal conditions, the generation of reactive oxygen species (ROS) in mitochondrias would keep an "oxidated status" of potassium channels, which would stay open. During hypoxia, mitochondrial electron transport is reduced, as well as the generation of ROS. The consequent "reduced status" of cytosol would result in inhibition of potassium channels and depolarization of cellular membrane7,8. Alternatively, the oxygen "sensor" system could not be in mitochondrias, but along the plasmatic membrane, represented by NADPH oxydases or similar enzymes. Another possibility is that hypoxia modulate potassium currents in Kv through cytochrome P450. Studies that involved its inhibition demonstrated a decrease in potassium currents, causing depolarization of cellular membrane8.
Regarding chronic hypoxia, mechanisms causing pulmonary vasoconstriction can be many. The activity of Kv channels may be limited due to reduction in their genes' transcription. In rats, chronic hypoxia diminishes mRNA expression and protein of a-subunit of potassium channels in smooth muscle cells, causing their functional decrease8. Besides, oxygen restriction increases the functional activity of hypoxia-induced transcription factor (HIF-1). Under conditions of normal oxygen tension, the a-subunit of that transactivator is degraded. In hypoxia, however, degradation is inhibited, with accumulation of HIF-1a, dimerization with HIF-1a and formation of functional HIF-1. That is responsible for activation of critical genes, as the vascular endothelial growth factor (VEGF) and ET-18.
A particular characteristic in patients with idiopathic PAH is the reduction of the number of functioning potassium channels. Changes in potassium channels were found in those patients' smooth muscle cells, but not in other PAH forms. By comparing normal individuals with carriers of systemic arterial hypertension, PAH associated to other diseases and idiopathic PAH, it was verified that smooth muscle cells of that last group showed a reduced expression of subunits of Kv channels9. That would result in a decrease in potassium currents, allowing for the entry of calcium and sustained pulmonary vasoconstriction.
Next, we start the discussion of some pulmonary vascular remodeling-related mechanisms. Such mechanisms involve growth factors, extracellular matrix, membrane receptors and proteolytic activities.
Among the growth factor and their receptors, the TGF-a (transforming growth factor) super-family is of special relevance in PAH genesis, as it has an important role in physiological processes of inflammation, immunity and cellular differentiation and proliferation. It consists of multifunctional mediators, including TGF-a and BMP (bone morphogenetic proteins). Studies have demonstrated that patients with idiopathic PAH have changes in BMP type 2 receptor (BMPR2), more precisely mutations in the gene that codifies it and that is located in chromosome 2q3310. BMPR2 binds cytokines as TGF-a, BMP, activin, inhibin and other growth and differentiation factors. It performs its signaling function through the formation of a heterodimeric complex on the cellular surface, along with type 1 receptor. When activated, that complex starts the signaling inside the cell through specific enzymes, known as Smad. That sign is translocated to the nucleus, where it will regulate the gene transcription. Mutations in BMPR2 would cause a deficient signaling, resulting in the loss of antiproliferative mechanisms, for example, in pulmonary circulation. It has been demonstrated that BMPR2 has its expression dramatically reduced in pulmonary circulation in idiopathic PAH patients, even in individuals who do not have mutations in this receptor11.
VEGF has been widely studied, more precisely in hypoxia-induced PAH. It highly connects with two tyrosine-kinase-type receptors: VEGFR-1 and VEGFR-2. For being an endothelial cell mitogen, it is involved in their growth, for example, inside the so-called plexiform lesions12. Chronic hypoxia, which follows some PAH forms, cruises with the increase of VEGF expression and its receptors13. Other growth factors are also influenced by hypoxia, as the platelet-derived growth factor (PDGF), fibroblast-derived growth factor (FGF-2), insulin-like growth factor (IGF-1) and the epidermal growth factor (EGF).
Nitric oxide (NO), an endothelium-derived vasodilator, has also an antiproliferative action over vascular smooth musculature14, possibly connected to intracellular cyclic GMP generation. In fact, it has been considered that phosphodiesterase 5 enzyme (which degrades cyclic GMP) inhibitors could act on hypoxia-induced pulmonary vascular remodeling15. Such protective action of cyclic GMP would be related to the pathway of MAPP kinases and to inhibition of transcription factor AML1 (acute myelogenous leukemia)16.
Prostacyclin, similar to NO, is a powerful vasodilator and inhibitor of platelet adhesion and cellular growth. It is the main metabolite of arachidonic acid in vascular endothelium, with action on smooth muscle cell, through the increase of cyclic AMP. Besides, prostacyclin inhibits the effect of endothelin-1 (ET-1). Patients with PAH show diminished prostacyclin urinary excretion4.
The role of serotonin in PAH is related to the induction of hyperplasia and hypertrophy of smooth muscle cells and mitogenic effect also in endothelial cells and fibroblasts. The action of serotonin in pulmonary circulation is wide and also includes vasoconstriction, for instance, in response to hypoxia17. Increased plasmatic levels of serotonin are observed in some forms of PAH18. Besides, the increased expression of its transporter could be related to accumulation of intracellular serotonin19.
ET-1 is a powerful vasoconstrictor predominantly produced by the endothelial cell, with mitogenic, inflammatory and proliferative action on the smooth muscle cell. Despite the absence of intracellular storages of ET-1, specific stimuli induce its synthesis and secretion in minutes. There are two types of endothelin receptors: ETA (found in smooth muscle cells and cardiac myocytes) and ETB (found in endothelial and smooth muscle cells). The binding of ET-1 to its receptors activates phospholipase C enzyme, increasing intracellular calcium levels, which results in a prolonged vasoconstriction. Diacylglycerol, produced from activated phospholipase C, stimulates protein-kinase C, a mediator enzyme of the mitogenic action of ET-120. Similar to serotonin, the action of ET-1 is potentized in the presence of other growth factors21. Other relevant effects of ET-1 are the stimulation for the production of cytokines and growth factors22, induction of the formation of extracellular matrix proteins23 and the potentiation of the effects of TGF-â and PDGF21. Studies in normal rat lungs demonstrated ET-1 mRNA levels five times higher if compared with any other organ24. Under normal conditions, the lungs remove ET-1 from the circulation. In PAH, the lungs synthesize ET-1 in increased amount, which results in an increase of circulating levels, with prognostic implications3. Under such conditions, pulmonary circulation becomes a target of this peptide.
Besides the action of growth factors and other signaling molecules in pulmonary vascular remodeling, events involving components of extracellular matrix are extremely relevant (
The mechanisms described above, in a summarized way, constitute in fact a proposed model of pulmonary vascular remodeling from many experimental studies1. It is understood that vascular remodeling in PAH is a complex process that most probably involves serum factors, protease induction, changes in extracellular matrix, availability of growth factors, activation of their receptors, and finally, mitosis. Besides, many changes involving extracellular matrix, integrins and growth factors, contribute to changes in the phenotype of smooth muscle cells from a contractile to a secretory one1.
In addition to expressive participation of the endothelium in changes of tonus and in pulmonary vascular remodeling, which are present in PAH, its involvement in the vaso-occlusive process, strictly speaking, becomes relevant as there is a progressive loss of its anticoagulant and antithrombotic properties, and an unleashing of mechanisms that facilitate coagulation and cellular adhesion, and inhibit fibrinolysis.
Among endothelial substances, which shown to be changed in PAH, with implications in the mechanisms described, there are thrombomodulin, tissue-type plasminogen activator (t-PA), its inhibitor (PAI-1), P-selectin and Von Willebrand factor (FvW), besides NO and prostacyclin. Besides, other prothrombotic changes have been identified, such as the presence of antiphospholipid antibodies and hyperhomocysteinemia.
Thrombomodulin is a plasma membrane proteoglycan. Its extracellular portion is cleaved in many regions, with a consequent release of fragments in blood flow, whose concentration can be easily determined. Those fragments are known as the soluble fraction. Thrombomodulin has thrombin-binding properties, which activates protein C that, for its turn, degrades coagulation factors V and VIII26. In PAH patients, thrombomodulin plasma levels are diminished, probably as a result of reduced endothelial synthesis26,27. Besides, there are reports that demonstrate low plasma levels of protein C, as well as its co-factor, protein S28. Such changes may reflect a defective synthesis or even consumption due to intravascular coagulation.
Patients with antiphospholipid syndrome, associated or not to systemic
Homocysteine is an amino acid that may facilitate coagulation and cellular adhesion in normal individuals. It is known that the presence of high plasmatic levels of homocysteine is associated to endothelial and platelet dysfunction, with a consequent formation of thrombi and vasoconstriction34. It is suggested that hyperhomocysteinemia may play some role in PAH pathogenesis, as there are studies showing increased levels of this amino acid in patients with the disease34.
A particular form of endothelial dysfunction in PAH patients is the one associated with hypoxemia, provided the stressed trend to thrombosis that takes place in the so-called Eisenmenger's syndrome, in which, due to the presence of intracardiac defects with blood deviation from right to left, chronic hypoxemia develops. Changes in coagulation and fibrinolysis in those patients are associated to a high risk of thromboembolic events, with reports of up of 35% of occurrence of pulmonary arterial thrombosis35. The fact that hypoxia promotes exocytosis of Weibel-Palade bodies in endothelial cells, which results in FvW and P-selectin release, contributes to that. Besides, it has been shown that hypoxia induces the expression of PAI-1 mRNA and protein in addition to the suppression of thrombomodulin. Therefore, in chronic hypoxemia-associated PAH it is justifiable to imagine that mechanisms as the recruitment of leucocytes and platelets, fibrinolysis inhibition and the loss of control over coagulation, are operative and contribute to the vaso-occlusive event.
In addition to supplying an estimate of thrombotic risk in PAH individuals, endothelial dysfunction markers seem to have prognostic implications. A study in patients with idiopathic PAH and secondary to congenital heart diseases correlated high FvW levels with decreased survival36. Patients with the idiopathic form of the disease showed higher levels, associated to a lower survival expectancy37. Other biochemical markers are also being studied, with prognostic aim, such as catecholamines, uric acid, and natriuretic peptides (ANP and BNP). However, only BNP seems to have correlation with survival, with high levels in idiopathic PAH patients38.
Endothelial dysfunction in PAH leads to a prothrombotic status, in which platelet participation seems unequivocal. They have been the target of studies for many years, as their role in PAH is more than merely thrombogenic, with an involvement also in the vascular remodeling process.
Clinical studies have been bringing about evidences of endogenous platelet activation in PAH patients, suggesting that there is a chronic consumption status. Such suggestion is due to the observation of the reduction of circulating platelet number and high levels of a-thromboglobulin39, a substance released at secretion stage. It is suggested that such activation favors the formation of cellular aggregates in circulation, involving platelets among themselves or in association with leucocytes and erythrocytes40, which could contribute to the disease progression in pulmonary vessels. Platelet activation in PAH also promotes the release of dense granules, containing mitogenic agents and vasoconstrictor substances. One of those substances is thromboxane A2, a powerful vasoconstrictor whose production is high in PAH patients41. Serotonin and growth factors, such as PDGF, EGF and TGF-a, are also released, with mitogenic effect on vascular smooth muscle cells, fibroblasts and endothelial cells42.
In different forms of PAH, the platelet involvement (activation) has been regarded as a secondary event to endothelial dysfunction, sub-endothelium exposure and changes in flow conditions. Exception is made to a situation identified as platelet storage pool disease, a rare, familial entity, in which there is a deficit in the ability of storing serotonin in dense granules, with extremely high levels of that substance in the plasma, causing pulmonary vasoconstriction43. With the report of PAH associated to that disease43, a series of studies was started in an attempt to demonstrate a causal association with serotonin.
Serotonin is the main component of platelets dense granules, being released from its activation. The normal response from the endothelial cell to serotonin is NO release, leading to vasorelaxation44. However, in endothelial dysfunction, serotonin is unable to stimulate NO release. In pulmonary circulation, such occurrence would favor PAH development through many mechanisms. Serotonin or 5-hydroxytryptamine (5-HT) performs its function through the connection to its receptor 5-HTR, and its transporter 5-HTT. When connected to 5-HTR, serotonin activates IP3 production, inducing the release of intracellular calcium from sarcoplasmatic reticulum, with consequent vasoconstriction45. On the other hand, diacylglycerol generation is implied in the induction of protein-kinase C mitogenic pathway. Serotonin connection to 5-HTT starts intracellular events, involving the formation of oxygen reactive species and the activation of MAPP kinase pathway, resulting in the expression of genes involved with hypertrophy and cellular proliferation46. Many studies suggest that serotonin has a role in PAH pathogenesis, including the idiopathic form. The presence of polymorphisms in the promoter region of the gene that encodes 5-HTT has already been demonstrated, with its expression increased19. So, that gene is today regarded as a "facilitator" in the pulmonary vascular remodeling process.
An important clinical condition, connected to serotonin metabolism and potentially PAH causer, is the use of oral anorexigenic, such as fenfluramine. In central nervous system, the effect would be performed through inhibition of 5-HTT and consequently serotonin reuptake in neurons. In the periphery, also for interfering with the transporter, fenfluramine is able to inhibits serotonin reuptake in platelets and pulmonary endothelial cells. The consequences are the increase of free serotonin, in a similar way to what happens in platelet storage pool disease, and pulmonary vasoconstriction19,46. Not every oral fenfluramine user develops PAH, which suggests that disorders in serotonin metabolism and in its transporter functioning are facilitating events that need predisposing factors for the development of the disease.
As previously mentioned, mutations in the gene that encodes BMPR2 change intracellular signaling which, in normal conditions, would suppress proliferation and favor the apoptosis of smooth muscle cells in pulmonary arteries. It is suggested that signaling from BMPR2 could antagonize the effects of serotonin and that mutations in that receptor would allow for an aggravated cellular response to it, especially in patients with polymorphisms of 5-HTT47. However, that relation is merely speculative for the time being.
Pulmonary arterial hypertension is complication usually found under many systemic inflammatory conditions, such as scleroderma and the systemic
In addition to inflammatory findings, it is important to emphasize the association of PAH with diseases or conditions of immune character: infection through human immunodeficiency virus48, POEMS (monoclonal plasma cell dyscrasia with a monoclonal protein present in plasma, polyneuropathy, organomegaly and skin changes) syndrome50, presence of autoimmune antibodies51 and Raynaud's phenomenon52. The main immunogenetic aspects in those diseases correspond to the presence of autoantibodies and certain alleles from class II major histocompatibility complex (MHC). Similarly, some patients with idiopathic PAH show high levels of antinuclear antibodies51 and a prevalence of certain alleles present in autoimmune diseases. So, an increased frequency of HLA-DR52 in patients with scleroderma and PAH53, of HLA-DR3 and HLA-DR52 in infants with idiopathic PAH54 has been observed. An increase in the frequency of HLA-DQ7 in infants and adults with idiopathic PAH has also been demonstrated. Such finding is important, as the allele is associated to the presence of lupus anticoagulant in patients with systemic
In an attempt to identify mechanisms that lead to inflammatory conditions found in PAH, an increase of many mediators has been observed. It has been already demonstrated that idiopathic PAH patients have high serum levels of IL-1 and IL-656. Another important finding is the involvement of chemokines, which are chemoattractant cytokines able to direct leucocyte recruiting and migration. One of those chemokines, designated by RANTES (regulated upon activation, normal T cell expressed and secreted), performs the recruitment of monocytes and T cells. Besides, RANTES chemokine is able to induce endothelin-1 converting enzyme, resulting in an increase of mitogenic activity and vasoconstriction57. In patients with idiopathic PAH, increased amounts of RANTES mRNA has been demonstrated, with endothelial cells as the main source58.
Fractalkine is a chemokine that also seems to have an important role in PAH: it promotes leucocyte recruitment through CX3CR1 transmembrane receptor, with fast capture and integrin-dependent adhesion. Increased fractalkine plasma levels and mRNA, as well as an increased of the expression and function of CX3CR1 receptor in lymphocites59 has been demonstrated in patients with PAH.
Another chemokine that may be involved in idiopathic PAH inflammatory process is MCP-1 (monocyte chemoattractant protein), which has a strong chemotaxis power in relation to mononuclear cells. It is produced in many cell types, including monocytes, endothelial and smooth muscle cells. It participates in monocyte activation and migration and also plays a role in proliferation of smooth muscle cells. It is also described that IL-1 and IL-6 cytokines may induce MCP-1 expression in smooth muscle and endothelial cells60. Serum concentration of MCP-1 is high in idiopathic PAH patients, a factor that may be related to the progression of the disease61.
Despite the inflammatory mechanisms established in idiopathic PAH and their association with connective tissue diseases, there is still controversy on their role in congenital cardiopathy-associated PAH. In a recent report on the presence of inflammatory cells in peripheral pulmonary arteries of congenital cardiopathy-associated PAH, a predominance of young macrophages in intimal and medial layers was detected, which suggests an inflammatory reaction in progress62. Being macrophages responsible for the production of many cytokines, it is speculated that they may have a relevant participation in events mediated by such substances (for example, cell replication) in patients with PAH.
After presenting the current view on the most relevant pathophysiological mechanisms operating in PAH, we are going to present two families with the disease.
The genetic study was carried out in five individuals of this family, as demonstrated in the pedigree. The study consisted of the amplification, through polymerase chain reaction (PCR), of 13 exons of the gene that codifies BMPR2 receptor. The product from the reaction was then submitted to sequencing through automatic method. As demonstrated in
Patient DED, 15 years old, was in functional class IV at admission, with mean pulmonary pressure of 62 mmHg, unable to walk any distance in the 6-minute walk test. After the beginning of treatment with sildenafil orally, in the dose of 225 mg a day, an important improvement in physical capacity was observed, as he started to walk 399m and 471m, respectively, at 6 months and 1 year. An increment of pulmonary flow, estimated through echocardiogram, of 50% at 6 months and 38% at 1 year was also noted. His sister (AKD), 18 years old at admission, had lower intensity manifestations and later beginning of the disease. The treatment was initiated in functional class II, with mean pulmonary pressure of 50 mmHg and a 309m walked distance. Under use of sildenafil (225 mg / day), there was an increase to 504m at 6 months and 1 year. An increment of 26% in pulmonary flow was also noted.
Patient EMB, 33 years old, was in functional class IV at admission, getting to make use of vasoactive drugs. The presence of pulmonary rales, with disappearance by means of diuretic therapy, raised the suspicion of veno-occlusive disease, a diagnosis later confirmed through hemodynamic study and pulmonary biopsy.
The assessment of this patient included acute vasodilatation test with nitric oxide (10ppm) and sildenafil (75 mg orally). The test demonstrated significant pulmonary vasodilatation with both stimuli separately applied, with an expressive decrease of vascular resistance (from 27.9 Wood units/m2, under basal condition, to 8.3 and 11.1U/m2, respectively, with nitric oxide and sildenafil). However, the pulmonary capillary pressure was stabilized on a safer level with sildenafil, compared to nitric oxide, with values of 15 and 29 mmHg, respectively. Results from this study allowed for the guidance of chronic use of sildenafil (225 mg/day) associated to diuretics, with a clear clinical improvement after a year, being possible to remove the patient from the waiting list for pulmonary transplantation. The six-minute walked distance was 112m at pretreatment condition and 294m and 408 m at 6 months and 1 year, respectively. Echocardiogram also showed an increase of pulmonary flow of 50% and 120%.
Such observations emphasize some important aspects of PAH. First, the familial form diagnosis, which in clinical practice would make possible the identification of the disease in less advanced stages, in individuals with discreet or absent symptomatology. Second, the importance of mutations in BMPR2 gene in pathophysiological scene, with the possibility of identifying such gene-related changes in at least 50% of ill-taken families. Finally, the pathophysiological role of NO and related pathways, as a rationale for the use of therapeutic resources with ability to increase cyclic GMP intracellular levels in pulmonary arterioles.
In consideration of the mechanisms discussed, it is clear that PAH is a syndrome whose beginning and progression processes in a diverse way, depending on the case. It is not possible to imagine the same pathophysiological mechanism or set of pathophysiological mechanisms explaining vasoconstriction and the evolution of vascular remodeling in all forms of PAH. Besides, even if we consider a single form of the disease, the pathophysiological mechanisms may vary depending on the stage. So, difficulties in the development of therapeutic resources usable in humans are noticed, as it would depend on the clear definition of operative mechanisms in a particular case or even on the identification of changes common to several patients with different forms of the disease. Even so, investigations in PAH pathophysiology have been relevant. Without them, therapeutic acquisitions as prostanoids, endothelin antagonists and phosphodiesterase inhibitors, currently in use, would not have been possible.
AML-1, acute myelogenic leukemia-1; AMP, adenine monophosphate; ANP, atrial natriuretic peptide; Apo A-1, apolipoprotein A1; BMP, bone morphogenetic protein; BMPR2, receptor 2 of BMP; BNP, brain natriuretic peptide; EGF, epidermal growth factor; ERO, oxygen reactive species; ERK, extracellular signal-related kinase; ET-1, endothelin-1; EVE, endogenous vascular elastase; FGF-2, fibroblast growth factor; FvW, von Willebrand factor; GMP, guanine monophosphate; PAH, pulmonary arterial hypertension; HIF-1, hypoxia-inducible factor; 5-HT, 5-hydroxytryptamine; 5-HTR, 5-hydroxytryptamine receptor; 5-HTT, 5-hydroxytryptamine transporter; IGF-1, insulin-like growth factor; IL-1, interleukin-1; IL-6, interleukin-6; Kv, voltage-dependent potassium channel; MCP-1, monocyte chemoattractant protein; MHC, main histocompatibility complex; NO, nitric oxide; NOS, nitric oxide synthase; PAI-1, plasminogen activator inhibitor; PDGF, platelet-derived growth factor; PKC, protein-kinase C; POEMS (syndrome), plasma cell dyscrasia with polyneuropathy, organomegaly, protein M and skin changes; TGF-b, beta type transformer growth factor; TN-C, tenascin-C; t-PA, plasminogen tissular activator; VEGF, vascular endothelial growth factor; VEGFR, vascular endothelial growth factor receptor.
1. Rabinovitch M. Pathobiology of pulmonary hypertension. Extracellular matrix. Clin Chest Med 2001; 22: 433-49.
2. Simonneau G, Galiè N, Rubin LJ, et al. Clinical classification of pulmonary hypertension. J Am Coll Cardiol 2004, 43: 5S-12S.
3. Galiè N, Griggione F, Bacchi-Reggiani L, et al. Relation of endothelin-1 to survival in primary pulmonary hypertension. Eur J Clin Invest 1996; 26: 273.
4. Tuder RM, Cool CD, Geraci MW, et al. Prostacyclin synthase expression is decreased in lungs from patients with severe pulmonary hypertension. Am J Respir Crit Care Med 1999; 159: 1925-32.
5. Giaid A, Saleh D. Reduced expression of endothelial nitric oxide synthase in the lungs of patients with pulmonary hypertension. N Engl J Med 1995; 333: 214-21.
6. Archer S, Wu X, Thébaud B, et al. Preferential expression and function of voltage-gated, O2-sensitive K+ channels in resistance pulmonary arteries explains regional heterogeneity in hypoxic pulmonary vasoconstriction. Circ Res 2004; 95: 308-18.
7. Ward JPT, Snetkov VA, Aaronson PI. Calcium, mitochondria and oxygen sensing in the pulmonary circulation. Cell Calcium 2004; 36: 209-20.
8. Mandegar M, Remillard CV, Yuan JXJ. Ion channels in pulmonary arterial hypertension. Prog Cardiov Dis 2002; 45: 81-114.
9. Yuan X-J, Wang J, Juhaszova M, et al. Attenuated K+ channel gene transcription in primary pulmonary hypertension. Lancet 1998; 351: 726-7.
10. Lane KB, Machado RD, Pauciulo MW, et al. Heterozigous germline mutations in a TGF-a receptor, BMPR2, are the cause of familial primary pulmonary hypertension. The International PPH Consortium. Nat Genet 2000; 26: 81-4.
11. Atkinson C, Stewart S, Upton PD, et al. Primary pulmonary hypertension is associated with reduced pulmonary vascular expression of type II bone morphogenetic protein receptor. Circulation 2002; 105: 1672-8.
12. Christman BW, McPherson CD, Newman JH, et al. An imbalance between the excretion of thromboxane and prostacyclin metabolites in pulmonary hypertension. N Engl J Med 1992; 372: 70-5.
13. Hirose S, Hosoda Y, Furuya S, et al. Expression of vascular endothelial growth factor and its receptor correlates closely with formation of the plexiform lesion in human pulmonary hypertension. Pathol Int 2000; 50: 472-9.
14. Gillespie MN, Rippetoe PE, Haven AC, et al. Polyamines and epidermal growth factor in monocrotaline-induced pulmonary hypertension. Am Rev Respir Dis 1989; 140: 1463-6.
15. Walford G, Loscalzo J. Nitric oxide in vascular biology. J Thromb Haemost 2003; 1: 2112-18.
16. Rabinovitch M. EVE and beyond, retro and prospective insights. Am J Physiol 1999; 277: L5-L12.
17. Launay JM, Hervé P, Peoc'h K, et al. Function of the serotonin 5-hydroxytryptamine 2B receptor in pulmonary hypertension. Nat Med 2002; 8: 1129-35.
18. Egermayer P, Town GI, Peacock AJ. Role of serotonin in the pathogenesis of acute and chronic pulmonary hypertension. Thorax 1999; 54: 161-8.
19. Eddahibi S, Humbert M, Fadel E, et al. Hyperplasia of pulmonary artery smooth muscle cells is causally related to overexpression of the serotonin transporter in primary pulmonary hypertension. Chest 2002; 121: 97S-98S.
20. Assender JW, Irenius E, Fredholm BB. Endothelin-1 causes a prolonged protein kinase C activation and acts as a co-mitogen in vascular smooth muscle cells. Acta Physiol Scand 1996; 157: 451-60.
21. Weissberg PL, Witchel C, Davenport AD, et al. The endothelin peptides ET-1, ET-2, ET-3 and sarafatoxin S6c are comitogenic with platelet derived growth factor for vascular smooth muscle cells. Atherosclerosis 1990; 83: 257-62.
22. Agui T, Xin X, Cai Y, et al. Stimulation of interleukin-6 production by endothelin in rat bone marrow-derived stromal cells. Blood 1994; 84: 2531-8.
23. Peifley KA, Winkless JA. Angiotensin II and endothelin-1 increase fibroblast growth factor-2 mRNA expression in vascular smooth muscle cells. Biochem Biophys Res Commun 1998; 242: 202-8.
24. Matsumoto H, Suzuki N, Onda H, et al. Abundance of endothelin-3 in rat intestine, pituitary gland and brain. Biochem Biophys Res Commun 1989; 164: 74-80.
25. Rabinovitch M. Elastase and the pathobiology of unexplained pulmonary hypertension. Chest 1998; 114: 213S-224S.
26. Cacoub P, Karmochkine M, Dorent R, et al. Plasma levels of thrombomodulin in pulmonary hypertension. Am J Med 1996; 101: 160-4.
27. Sakamaki F, Kyotani S, Nagaya N, et al. Increased plasma P-selectin and decreased thrombomodulin in pulmonary arterial hypertension were improved by continuous prostacyclin therapy. Circulation 2000; 102: 2720-5.
28. Maeda NY, Lopes AA, Da Rocha TR, et al. Plasma anticoagulant system in patients with pulmonary hypertension. Braz J Med Biol Res 1990; 23: 251-4.
29. Esmon NL, Safa O, Smirnov MD, et al. Antiphospholipid antibodies and the protein C pathway. J Autoimmun 2000; 15: 221-5.
30. Meroni PL, Riboldi P. Pathogenic mechanism mediating antiphospholipid syndrome. Curr Opin Rheumatol 2001; 13: 377-82.
31. Espinosa G, Cervera R, Font J, et al. The lung in the antiphospholipid syndrome. Ann Rheum Dis 2002; 61: 195-8.
32. Moser KM, Auger WR, Fedullo PF. Chronic major-vessel thromboembolic pulmonary hypertension. Circulation 1990; 81: 1735-43.
33. Asherson RA, Higenbottan TW, Dinh Xuan AT, et al. Pulmonary hypertension in a lupus clinic: experience with twenty-four patients. J Rheumatol 1990; 17: 1292-8.
34. Arroliga AC, Sandur S, Jacobsen D, et al. Association between hyperhomocysteinemia and primary pulmonary hypertension. Respir Med 2003; 97: 825-9.
35. Caramuru LH, Maeda NY, Bydlowski SD, et al. Age-dependent likelihood of
36. Lopes AA, Maeda NY, Gonçalves RC, et al. Endothelial cell dysfunction correlates differentially with survival in primary and secondary pulmonary hypertension. Am Heart J 2000 139: 618-23.
37. Lopes AA, Maeda NY. Circulating von Willebrand factor antigen as a predictor of short-term prognosis in pulmonary hypertension. Chest 1998; 114: 1276-82.
38. Nagaya N, Nishikimi T, Uematsu M, et al. Plasma brain natriuretic peptide as a prognostic indicador in patients with primary pulmonary hipertensión. Circulation 2000 102: 865-70.
39. Caramuru LH, Soares RPS, Maeda NY, et al. Hypoxia and altered platelet behavior influence Von Willebrand factor multimeric composition in secondary pulmonary hypertension. Clin Appl Thromb Hemost 2003; 9: 251-8.
40. Lopes AA, Maeda NY, Almeida A, et al. Circulating platelets aggregates indicative of in vivo platelet activation in pulmonary hypertension. Angiology 1993; 44: 701-06.
41. Christman BW, McPherson CD, Newman JH, et al. An imbalance between the excretion of thromboxane and prostacyclin metabolites in pulmonary hypertension. N Engl J Med 1992; 327: 70-5.
42. Pakala R, Willerson JT, Benedict CR. Mitogenic effect of serotonin on vascular endothelial cells. Circulation 1994; 90: 1919-26.
43. Herve P, Drouet L, Launay JM, et al. Primary pulmonary hypertension in a patient with a familial platelet storage pool disease: rolo of serotonin. Am J Med 1990; 89: 117-20.
44. Flavahan NA, Shimokawa H, Vanhoutte PM. Pertussis toxin inhibits endothelium-dependent relaxations to certain agonists in porcine coronary arteries. J Physiol (Lond) 1989; 408: 549-60.
45. Boulanger C, Lorenz RR, Hendrickson H, et al. Release of different relaxing factors by porcine endothelial cells in culture. FASEB 1998; 2: A314.
46. Eddahibi S, Fabre V, Boni C, et al. Induction of serotonin transporter by hypoxia in pulmonary vascular smooth muscle – relationship with the mitogenic ction of serotonin. Circ Res 1999; 84: 329-36.
47. Rothman RB, Ayestas MA, Dersch CM, et al. Aminorex, fenfluramine and chlorphentermine are serotonin transporter substrates. Implications for primary pulmonary hypertension. Circulation 1999; 100: 869-75.
48. Cool CD, Kennedy D, Voelkel NF, et al. Pathogenesis and evolution of plexiform lesions in pulmonary hypertension associated with scleroderma and human immunodeficiency virus infection. Human Pathol 1997; 28: 434-42.
49. Tuder RM, Groves B, Badesch DB, et al. Exuberant endothelial cell growth and elements of inflammation are present in plexiform lesions of pulmonary hypertension. Am J Pathol 1994; 144: 275-85.
50. Lesprit P, Godeau B, Authier FJ, et al. Pulmonary hypertension in POEMS syndrome: a new feature mediated by cytokines. Am J Respir Crit Care Med 1998; 157: 907-11.
51. Isern RA, Yaneva M, Weiner E, et al. Autoantibodies in patients with primary pulmonary hipertensión: association with anti-Ku. Am J Med 1992; 93: 307-12.
52. Wigley FM. Raynaud's phenomenon and other features of scleroderma, including pulmonary hypertension. Curr Opin Rheumatol 1996; 8: 561-8.
53. Langevitz PD, Buskila D, Gladman DD, et al. HLA alleles in systemic sclerosis: association with pulmonary hypertension and outcome. Br J Rheumatol 1992; 31: 609-13.
54. Barst RJ, Flaster ER, Menon A, et al. Evidence for the association of unexplained pulmonary hypertension in children with the major histocompatibility complex. Circulation 1992; 85: 249-58.
55. Morse JH, Barst RJ, Fotino M, et al. Primary pulmonary hypertension, tissue plasminogen activator antibodies, and HLA-DQ7. Am J Respir Crit Care Med 1997; 155: 274-8.
56. Humbert M, Monti G, Breno TF, et al. Increased interleukin-1 and interleukin-6 serum concentrations in severe primary pulmonary hypertension. Am J Respir Crit Care Med 1995; 151: 1628-31.
57. Molet S, Furukawa K, Maghazechi A, et al. Chemokine and cytokine-induced expression of endothelin-1 and endothelin converting enzyme-1 in endothelial cells. J Allergy Clin Immunol 2000; 105: 333-8.
58. Dorfmüller P, Zarka V, Durand-Gasselin I, et al. Chemokine RANTES in severe pulmonary arterial hypertension. Am J Respir Crit Care Med 2002; 165: 534-9.
59. Balabanian K, Foussat A, Dorfmüller P, et al. CX3C chemokine fractalkine in pulmonary arterial hypertension. Am J Respir Crit Care Med 2002; 165: 1-8.
60. Sica A, Wang JM, Colotta F, et al. Monocyte chemotatic and activating factor gene expression induced in endothelial cells by IL-1 and tumor necrosis factor. J Immunol 1990; 144: 3034-8.
61. Hashimoto K, Nakamura K, Fugio H, et al. Epoprostenol therapy decreases elevated circulating levels of monocyte chemoattractant protein-1 in patients with primary pulmonary hypertension. Circ J 2004; 68: 227-31.
62. Pinto RFA, Higuchi ML, Aiello VD. Decreased numbers of T-lymphocytes and predominance of recently recruited macrophages in the walls of peripheral pulmonary arteries from 26 patients with pulmonary hypertension secondary to congenital cardiac shunts. Cardiovasc Pathol 2004, 13: 268-75.
Antonio Augusto Lopes InCor Cardiologia Pediátrica e Cardiopatias Congênitas do Adulto Av. Dr. Eneas de Carvalho Aguiar, 44 05403-000 - São Paulo, SP E-mail:
Sent for publishing on 03/02/2005
Accepted on 04/04/2005