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1.
Eur Heart J ; 40(37): 3081-3094, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31114854

RESUMO

AIMS: The Brugada syndrome (BrS) is an inherited cardiac disorder predisposing to ventricular arrhythmias. Despite considerable efforts, its genetic basis and cellular mechanisms remain largely unknown. The objective of this study was to identify a new susceptibility gene for BrS through familial investigation. METHODS AND RESULTS: Whole-exome sequencing performed in a three-generation pedigree with five affected members allowed the identification of one rare non-synonymous substitution (p.R211H) in RRAD, the gene encoding the RAD GTPase, carried by all affected members of the family. Three additional rare missense variants were found in 3/186 unrelated index cases. We detected higher levels of RRAD transcripts in subepicardium than in subendocardium in human heart, and in the right ventricle outflow tract compared to the other cardiac compartments in mice. The p.R211H variant was then subjected to electrophysiological and structural investigations in human cardiomyocytes derived from induced pluripotent stem cells (iPSC-CMs). Cardiomyocytes derived from induced pluripotent stem cells from two affected family members exhibited reduced action potential upstroke velocity, prolonged action potentials and increased incidence of early afterdepolarizations, with decreased Na+ peak current amplitude and increased Na+ persistent current amplitude, as well as abnormal distribution of actin and less focal adhesions, compared with intra-familial control iPSC-CMs Insertion of p.R211H-RRAD variant in control iPSCs by genome editing confirmed these results. In addition, iPSC-CMs from affected patients exhibited a decreased L-type Ca2+ current amplitude. CONCLUSION: This study identified a potential new BrS-susceptibility gene, RRAD. Cardiomyocytes derived from induced pluripotent stem cells expressing RRAD variant recapitulated single-cell electrophysiological features of BrS, including altered Na+ current, as well as cytoskeleton disturbances.

2.
Am J Hum Genet ; 102(1): 133-141, 2018 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-29304371

RESUMO

Intracranial aneurysms (IAs) are acquired cerebrovascular abnormalities characterized by localized dilation and wall thinning in intracranial arteries, possibly leading to subarachnoid hemorrhage and severe outcome in case of rupture. Here, we identified one rare nonsense variant (c.1378A>T) in the last exon of ANGPTL6 (Angiopoietin-Like 6)-which encodes a circulating pro-angiogenic factor mainly secreted from the liver-shared by the four tested affected members of a large pedigree with multiple IA-affected case subjects. We showed a 50% reduction of ANGPTL6 serum concentration in individuals heterozygous for the c.1378A>T allele (p.Lys460Ter) compared to relatives homozygous for the normal allele, probably due to the non-secretion of the truncated protein produced by the c.1378A>T transcripts. Sequencing ANGPTL6 in a series of 94 additional index case subjects with familial IA identified three other rare coding variants in five case subjects. Overall, we detected a significant enrichment (p = 0.023) in rare coding variants within this gene among the 95 index case subjects with familial IA, compared to a reference population of 404 individuals with French ancestry. Among the 6 recruited families, 12 out of 13 (92%) individuals carrying IA also carry such variants in ANGPTL6, versus 15 out of 41 (37%) unaffected ones. We observed a higher rate of individuals with a history of high blood pressure among affected versus healthy individuals carrying ANGPTL6 variants, suggesting that ANGPTL6 could trigger cerebrovascular lesions when combined with other risk factors such as hypertension. Altogether, our results indicate that rare coding variants in ANGPTL6 are causally related to familial forms of IA.

3.
J Am Coll Cardiol ; 70(3): 358-370, 2017 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-28705318

RESUMO

BACKGROUND: Inherited cardiac conduction disease is a rare bradyarrhythmia associated with mutations in various genes that affect action potential propagation. It is often characterized by isolated conduction disturbance of the His-Purkinje system, but it is rarely described as a syndromic form. OBJECTIVES: The authors sought to identify the genetic defect in families with a novel bradyarrhythmia syndrome associated with bone malformation. METHODS: The authors genetically screened 15 European cases with genotype-negative de novo atrioventricular (AV) block and their parents by trio whole-exome sequencing, plus 31 Japanese cases with genotype-negative familial AV block or sick sinus syndrome by targeted exon sequencing of 457 susceptibility genes. Functional consequences of the mutation were evaluated using an in vitro cell expression system and in vivo knockout mice. RESULTS: The authors identified a connexin-45 (Cx45) mutation (p.R75H) in 2 unrelated families (a de novo French case and a 3-generation Japanese family) who presented with progressive AV block, which resulted in atrial standstill without ventricular conduction abnormalities. Affected individuals shared a common extracardiac phenotype: a brachyfacial pattern, finger deformity, and dental dysplasia. Mutant Cx45 expressed in Neuro-2a cells showed normal hemichannel assembly and plaque formation. However, Lucifer yellow dye transfer and gap junction conductance between cell pairs were severely impaired, which suggested that mutant Cx45 impedes gap junction communication in a dominant-negative manner. Tamoxifen-induced, cardiac-specific Cx45 knockout mice showed sinus node dysfunction and atrial arrhythmia, recapitulating the intra-atrial disturbance. CONCLUSIONS: Altogether, the authors showed that Cx45 mutant p.R75H is responsible for a novel disease entity of progressive atrial conduction system defects associated with craniofacial and dentodigital malformation.


Assuntos
Bloqueio Atrioventricular/etiologia , Conexinas/genética , DNA/genética , Deformidades Dentofaciais/complicações , Mutação , Adolescente , Adulto , Animais , Bloqueio Atrioventricular/genética , Bloqueio Atrioventricular/fisiopatologia , Criança , Pré-Escolar , Conexinas/metabolismo , Análise Mutacional de DNA , Deformidades Dentofaciais/genética , Deformidades Dentofaciais/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Eletrocardiografia , Feminino , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Linhagem , Fenótipo , Adulto Jovem
4.
Cell Signal ; 33: 10-21, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28179144

RESUMO

The p38 MAPK signaling pathway is essential in the cellular response to stress stimuli, in particular in the endothelial cells that are major target of external stress. The importance of the bioactive sphingolipid ceramide generated by acid sphingomyelinase is also firmly established in stress-induced endothelial apoptotic cell death. Despite a suggested link between the p38 MAPK and ceramide pathways, the exact molecular events of this connection remain elusive. In the present study, by using two different activators of p38 MAPK, namely anisomycin and ionizing radiation, we depicted how ceramide generated by acid sphingomyelinase was involved in p38 MAPK-dependent apoptosis of endothelial cells. We first proved that both anisomycin and ionizing radiation conducted to apoptosis through activation of p38 MAPK in human microvascular endothelial cells HMEC-1. We then found that both treatments induced activation of acid sphingomyelinase and the generation of ceramide. This step was required for p38 MAPK activation and apoptosis. We finally showed that irradiation, as well as treatment with exogenous C16-ceramide or bacterial sphingomyelinase, induced in endothelial cells a deep reorganization of the plasma membrane with formation of large lipid platforms at the cell surface, leading to p38 MAPK activation and apoptosis in endothelial cells. Altogether, our results proved that the plasma membrane reorganization leading to ceramide production is essential for stress-induced activation of p38 MAPK and apoptosis in endothelial cells and established the link between the acid sphingomyelinase/ceramide and p38 MAPK pathways.


Assuntos
Apoptose , Membrana Celular/metabolismo , Ceramidas/metabolismo , Sistema de Sinalização das MAP Quinases , Esfingomielina Fosfodiesterase/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Células Endoteliais/metabolismo , Ativação Enzimática , Humanos , Microdomínios da Membrana/metabolismo , Modelos Biológicos , Estresse Fisiológico
5.
J Am Heart Assoc ; 5(6)2016 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-27287695

RESUMO

BACKGROUND: The Brugada syndrome is an inherited cardiac arrhythmia associated with high risk of sudden death. Although 20% of patients with Brugada syndrome carry mutations in SCN5A, the molecular mechanisms underlying this condition are still largely unknown. METHODS AND RESULTS: We combined whole-exome sequencing and linkage analysis to identify the genetic variant likely causing Brugada syndrome in a pedigree for which SCN5A mutations had been excluded. This approach identified 6 genetic variants cosegregating with the Brugada electrocardiographic pattern within the pedigree. In silico gene prioritization pointed to 1 variant residing in KCNAB2, which encodes the voltage-gated K(+) channel ß2-subunit (Kvß2-R12Q). Kvß2 is widely expressed in the human heart and has been shown to interact with the fast transient outward K(+) channel subunit Kv4.3, increasing its current density. By targeted sequencing of the KCNAB2 gene in 167 unrelated patients with Brugada syndrome, we found 2 additional rare missense variants (L13F and V114I). We then investigated the physiological effects of the 3 KCNAB2 variants by using cellular electrophysiology and biochemistry. Patch-clamp experiments performed in COS-7 cells expressing both Kv4.3 and Kvß2 revealed a significant increase in the current density in presence of the R12Q and L13F Kvß2 mutants. Although biotinylation assays showed no differences in the expression of Kv4.3, the total and submembrane expression of Kvß2-R12Q were significantly increased in comparison with wild-type Kvß2. CONCLUSIONS: Altogether, our results indicate that Kvß2 dysfunction can contribute to the Brugada electrocardiographic pattern.


Assuntos
Síndrome de Brugada/genética , Mutação com Ganho de Função/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Eletrocardiografia , Feminino , Predisposição Genética para Doença/genética , Genótipo , Humanos , Masculino , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Linhagem , Polimorfismo de Nucleotídeo Único/genética , Canais de Potássio Shal/genética , Sequenciamento Completo do Exoma
6.
Int J Cardiol ; 207: 349-58, 2016 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-26820365

RESUMO

BACKGROUND: Progressive cardiac conduction disease (PCCD) is one of the most common cardiac conduction disturbances. It has been causally related to rare mutations in several genes including SCN5A, SCN1B, TRPM4, LMNA and GJA5. METHODS AND RESULTS: In this study, by applying targeted next-generation sequencing (NGS) in 95 unrelated patients with PCCD, we have identified 13 rare variants in the TRPM4 gene, two of which are currently absent from public databases. This gene encodes a cardiac calcium-activated cationic channel which precise role and importance in cardiac conduction and disease is still debated. One novel variant, TRPM4-p.I376T, is carried by the proband of a large French 4-generation pedigree. Systematic familial screening showed that a total of 13 family members carry the mutation, including 10 out of the 11 tested affected individuals versus only 1 out of the 21 unaffected ones. Functional and biochemical analyses were performed using HEK293 cells, in whole-cell patch-clamp configuration and Western blotting. TRPM4-p.I376T results in an increased current density concomitant to an augmented TRPM4 channel expression at the cell surface. CONCLUSIONS: This study is the first extensive NGS-based screening of TRPM4 coding variants in patients with PCCD. It reports the third largest pedigree diagnosed with isolated Progressive Familial Heart Block type I and confirms that this subtype of PCCD is caused by mutation-induced gain-of-expression and function of the TRPM4 ion channel.


Assuntos
Progressão da Doença , Marcação de Genes/métodos , Predisposição Genética para Doença/genética , Bloqueio Cardíaco/genética , Análise de Sequência de DNA/métodos , Canais de Cátion TRPM/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Doença do Sistema de Condução Cardíaco , Criança , Feminino , Variação Genética/genética , Células HEK293 , Bloqueio Cardíaco/diagnóstico , Humanos , Masculino , Pessoa de Meia-Idade , Linhagem
7.
Hum Mol Genet ; 24(10): 2757-63, 2015 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-25650408

RESUMO

The Brugada syndrome (BrS) is a rare heritable cardiac arrhythmia disorder associated with ventricular fibrillation and sudden cardiac death. Mutations in the SCN5A gene have been causally related to BrS in 20-30% of cases. Twenty other genes have been described as involved in BrS, but their overall contribution to disease prevalence is still unclear. This study aims to estimate the burden of rare coding variation in arrhythmia-susceptibility genes among a large group of patients with BrS. We have developed a custom kit to capture and sequence the coding regions of 45 previously reported arrhythmia-susceptibility genes and applied this kit to 167 index cases presenting with a Brugada pattern on the electrocardiogram as well as 167 individuals aged over 65-year old and showing no history of cardiac arrhythmia. By applying burden tests, a significant enrichment in rare coding variation (with a minor allele frequency below 0.1%) was observed only for SCN5A, with rare coding variants carried by 20.4% of cases with BrS versus 2.4% of control individuals (P = 1.4 × 10(-7)). No significant enrichment was observed for any other arrhythmia-susceptibility gene, including SCN10A and CACNA1C. These results indicate that, except for SCN5A, rare coding variation in previously reported arrhythmia-susceptibility genes do not contribute significantly to the occurrence of BrS in a population with European ancestry. Extreme caution should thus be taken when interpreting genetic variation in molecular diagnostic setting, since rare coding variants were observed in a similar extent among cases versus controls, for most previously reported BrS-susceptibility genes.


Assuntos
Síndrome de Brugada/genética , Predisposição Genética para Doença , Mutação , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Adulto , Arritmias Cardíacas/genética , Síndrome de Brugada/diagnóstico , Grupo com Ancestrais do Continente Europeu , Feminino , Frequência do Gene , Genes , Estudos de Associação Genética , Humanos , Masculino , Pessoa de Meia-Idade , Análise de Sequência de DNA
8.
Biochem Biophys Res Commun ; 414(4): 750-5, 2011 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-22001926

RESUMO

Endothelial cells of the microvasculature are major target of ionizing radiation, responsible of the radiation-induced vascular early dysfunctions. Molecular signaling pathways involved in endothelial responses to ionizing radiation, despite being increasingly investigated, still need precise characterization. Small GTPase RhoA and its effector ROCK are crucial signaling molecules involved in many endothelial cellular functions. Recent studies identified implication of RhoA/ROCK in radiation-induced increase in endothelial permeability but other endothelial functions altered by radiation might also require RhoA proteins. Human microvascular endothelial cells HMEC-1, either treated with Y-27632 (inhibitor of ROCK) or invalidated for RhoA by RNA interference were exposed to 15Gy. We showed a rapid radiation-induced activation of RhoA, leading to a deep reorganisation of actin cytoskeleton with rapid formation of stress fibers. Endothelial early apoptosis induced by ionizing radiation was not affected by Y-27632 pre-treatment or RhoA depletion. Endothelial adhesion to fibronectin and formation of focal adhesions increased in response to radiation in a RhoA/ROCK-dependent manner. Consistent with its pro-adhesive role, ionizing radiation also decreased endothelial cells migration and RhoA was required for this inhibition. These results highlight the role of RhoA GTPase in ionizing radiation-induced deregulation of essential endothelial functions linked to actin cytoskeleton.


Assuntos
Movimento Celular/efeitos da radiação , Endotélio Vascular/efeitos da radiação , Proteína rhoA de Ligação ao GTP/metabolismo , Actinas/metabolismo , Amidas/farmacologia , Apoptose/efeitos da radiação , Adesão Celular/efeitos da radiação , Linhagem Celular , Citoesqueleto/metabolismo , Endotélio Vascular/enzimologia , Endotélio Vascular/fisiologia , Fibronectinas/metabolismo , Humanos , Piridinas/farmacologia , Quinases Associadas a rho/antagonistas & inibidores , Quinases Associadas a rho/genética , Quinases Associadas a rho/metabolismo , Proteína rhoA de Ligação ao GTP/antagonistas & inibidores , Proteína rhoA de Ligação ao GTP/genética
9.
Clin Cancer Res ; 17(18): 5973-81, 2011 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-21821698

RESUMO

PURPOSE: Mantle cell lymphoma (MCL) is considered to be incurable. ABT-737 is a BH3 mimetic that targets Bcl-2, which is overexpressed in MCL and implicated in drug resistance. The present work investigated the antitumor effect of ABT-737. EXPERIMENTAL DESIGN: Six MCL cell lines and primary MCL cells (n = 13) were used. Sensitivity to ABT-737 was assessed, and expression levels of Bcl-2 and Mcl-1 were analyzed. Finally, ABT-737 was combined with other cytotoxic agents to promote tailored therapy. RESULTS: MINO and GRANTA-519 cell lines were highly sensitive to ABT-737 [the median lethal dose (LD50) = 20 and 80 nmol/L, respectively], whereas other cell lines were resistant. In primary MCL cells, 46% of patients' samples were sensitive to ABT-737. The analysis of protein expression levels revealed that both sensitive cell lines and primary MCL cells could be characterized by a Bcl-2(high)/Mcl-1(low) profile, whereas resistant MCL cells contained high levels of Mcl-1. ABT-737 induced a rapid disruption of both Bcl-2/Bax and Bcl-2/Bik complexes. In addition, silencing of Mcl-1 by siRNA sensitized MCL cell lines to ABT-737. Similarly, flavopiridol, which induces Mcl-1 downregulation, in combination with ABT-737 led to a synergistic anti-MCL effect in ABT-737-resistant cell lines. This synergy was also observed when ABT-737 was combined with either bortezomib or cytarabine. CONCLUSIONS: The present work shows that ABT-737 induces strong apoptosis in MCL cells expressing a Bcl-2(high)/Mcl-1(low) profile. In ABT-737-resistant MCL cells, downregulation of Mcl-1 overcomes Mcl-1-induced resistance and synergizes ABT-737 effects. Our results strongly support the use of ABT-737 according to the Bcl-2/Mcl-1 tumor cell profiles in the treatment of MCL.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Compostos de Bifenilo/farmacologia , Linfoma de Célula do Manto/metabolismo , Nitrofenóis/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Sulfonamidas/farmacologia , Proteínas Reguladoras de Apoptose/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Sinergismo Farmacológico , Humanos , Proteínas de Membrana/metabolismo , Complexos Multiproteicos/metabolismo , Proteína de Sequência 1 de Leucemia de Células Mieloides , Piperazinas/farmacologia , Proteína X Associada a bcl-2/metabolismo
10.
Cancer Res ; 70(23): 9905-15, 2010 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-21118968

RESUMO

A previous in vitro study showed that sphingosine-1-phosphate (S1P), a ceramide antagonist, preserved endothelial cells in culture from radiation-induced apoptosis. We proposed to validate the role of S1P in tissue radioprotection by inhibiting acute gastrointestinal (GI) syndrome induced by endothelial cell apoptosis after high dose of radiation. Retro-orbital S1P was injected in mice exposed to 15 Gy, a dose-inducing GI syndrome within 10 days. Overall survival and apoptosis on intestines sections were studied. Intestinal cell type targeted by S1P and early molecular survival pathways were researched using irradiated in vitro cell models and in vivo mouse models. We showed that retro-orbital S1P injection before irradiation prevented GI syndrome by inhibiting endothelium collapse. We defined endothelium as a specific therapeutic target because only these cells and not intestinal epithelial cells, or B and T lymphocytes, were protected. Pharmacologic approaches using AKT inhibitor and pertussis toxin established that S1P affords endothelial cell protection in vitro and in vivo through a mechanism involving AKT and 7-pass transmembrane receptors coupled to Gi proteins. Our results provide strong pharmacologic and mechanistic proofs that S1P protects endothelial cells against acute radiation enteropathy.


Assuntos
Apoptose/efeitos dos fármacos , Intestino Delgado/efeitos dos fármacos , Lisofosfolipídeos/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Esfingosina/análogos & derivados , Animais , Apoptose/efeitos da radiação , Western Blotting , Medula Óssea/efeitos dos fármacos , Medula Óssea/patologia , Medula Óssea/efeitos da radiação , Linhagem Celular , Células Cultivadas , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais/efeitos da radiação , Trato Gastrointestinal/efeitos dos fármacos , Trato Gastrointestinal/patologia , Trato Gastrointestinal/efeitos da radiação , Humanos , Imuno-Histoquímica , Intestino Delgado/citologia , Intestino Delgado/efeitos da radiação , Tecido Linfoide/efeitos dos fármacos , Tecido Linfoide/patologia , Tecido Linfoide/efeitos da radiação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação/efeitos dos fármacos , Fosforilação/efeitos da radiação , Lesões Experimentais por Radiação/tratamento farmacológico , Esfingosina/farmacologia , Síndrome
11.
Radiat Res ; 167(2): 185-93, 2007 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17390726

RESUMO

The response of endothelial cells (EC) to high radiation doses leads to damage of normal tissue or tumor. The precise mechanisms of the endothelial-tissue linkage are still largely unknown. We investigated the possible involvement of a bystander effect, secondary to endothelial damage, in tissue response to radiation. Proliferating human intestinal epithelial T84 cells were grown in a non-contact co-culture with confluent primary human microvascular EC (HMVEC-L). The bystander response in unirradiated T84 cells co-cultured with irradiated EC was studied by evaluating cell growth, cell death and epithelial morphology. Twenty-four hours after exposure of EC to 15 Gy, unirradiated T84 cells showed a decreased cell number (29%) and percentage in mitosis (66%) as well as increased apoptosis (1.5-fold) and cell surface area (1.5-fold), highlighting the involvement of bystander effects on T84 cells after irradiation of EC. Furthermore, the responses of T84 cells were amplified when EC and T84 cells were irradiated together, indicating that the bystander response in T84 cells adds further to direct radiation damage. As opposed to direct irradiation, the T84 cell bystander response did not involve the cell cycle-related protein p21(Waf1) (CDKN1A) and pro-apoptosis protein BAX. The bystander effect was specific to EC since the irradiation of human colon fibroblasts did not induce bystander responses in unirradiated T84 cells. These results strengthen previous in vivo evidence of the role of EC in tissue damage by radiation. In addition, this study provides a suitable and useful model to identify soluble factors involved in bystander effects secondary to endothelial damage. Modulating such factors may have important clinical implications.


Assuntos
Endotélio Vascular/efeitos da radiação , Células Epiteliais/metabolismo , Células Epiteliais/efeitos da radiação , Mucosa Intestinal/metabolismo , Intestinos/efeitos da radiação , Apoptose , Efeito Espectador , Ciclo Celular , Técnicas de Cocultura , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Citocinas/metabolismo , Endotélio Vascular/citologia , Humanos , Microcirculação , Mitose , RNA Mensageiro/metabolismo , Radiação , Proteína X Associada a bcl-2/metabolismo
12.
Cancer Res ; 67(4): 1803-11, 2007 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-17308123

RESUMO

Because of the central role of the endothelium in tissue homeostasis, protecting the vasculature from radiation-induced death is a major concern in tissue radioprotection. Premitotic apoptosis and mitotic death are two prevalent cell death pathways induced by ionizing radiation. Endothelial cells undergo apoptosis after radiation through generation of the sphingolipid ceramide. However, if mitotic death is known as the established radiation-induced death pathway for cycling eukaryotic cells, direct involvement of mitotic death in proliferating endothelial radiosensitivity has not been clearly shown. In this study, we proved that proliferating human microvascular endothelial cells (HMEC-1) undergo two waves of death after exposure to 15 Gy radiation: an early premitotic apoptosis dependent on ceramide generation and a delayed DNA damage-induced mitotic death. The fact that sphingosine-1-phosphate (S1P), a ceramide antagonist, protects HMEC-1 only from membrane-dependent apoptosis but not from DNA damage-induced mitotic death proves the independence of the two pathways. Furthermore, adding nocodazole, a mitotic inhibitor, to S1P affected both cell death mechanisms and fully prevented radiation-induced death. If our results fit with the standard model in which S1P signaling inhibits ceramide-mediated apoptosis induced by antitumor treatments, such as radiotherapy, they exclude, for the first time, a significant role of S1P-induced molecular survival pathway against mitotic death. Discrimination between ceramide-mediated apoptosis and DNA damage-induced mitotic death may give the opportunity to define a new class of radioprotectors for normal tissues in which quiescent endothelium represents the most sensitive target, while excluding malignant tumor containing pro-proliferating angiogenic endothelial cells that are sensitive to mitotic death.


Assuntos
Apoptose/efeitos dos fármacos , Ceramidas/antagonistas & inibidores , Dano ao DNA , Células Endoteliais/efeitos dos fármacos , Lisofosfolipídeos/farmacologia , Esfingosina/análogos & derivados , Apoptose/efeitos da radiação , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/fisiologia , Processos de Crescimento Celular , Ceramidas/biossíntese , Desipramina/farmacologia , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Células Endoteliais/efeitos da radiação , Humanos , Mitose/efeitos dos fármacos , Mitose/fisiologia , Nocodazol/farmacologia , Esfingosina/farmacologia
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