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Differential Endothelial VCAM1 Expression and Implications for Sickle Cell Anemia Vasculopathy

2021-06-15Conference object Open access
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dc.contributor.authorSilva, Marisa
dc.contributor.authorCoelho, Andreia
dc.contributor.authorVargas, Sofia
dc.contributor.authorFaustino, Paula
dc.date.accessioned2022-02-22T16:42:28Z
dc.date.available2022-02-22T16:42:28Z
dc.date.issued2021-06-15
dc.descriptionComunicação oral por convitept_PT
dc.description.abstractBackground: Vascular disease is systemic in sickle cell anemia (SCA), with profound effects in organs like the brain,where stroke is the most severe end of the cerebral vasculopathy spectrum. Endothelial dysfunction is an important pathobiological mechanism in SCA systemic vasculopathy, with upregulation of adhesion molecules (e.g., VCAM-1), lower nitric oxide bioavailability, and increased oxidative stress. In previous association studies, we found positive associations between the presence of three specific VCAM1 gene promoter haplotypes and i) high blood flow velocities in the median cerebral artery, and ii) a chronic hemolysis biochemical marker. Aims: The aims of our work were: a) to investigate the functional role of those VCAM1 promoter haplotypes in endothelial cell response following endothelial activation through TNF-α stimulation; b) to assess the modulation role of proinflammatory and/or pro-oxidative stimuli on endothelial VCAM1 expression; and, finally, to evaluate how hydroxyurea (HU) treatment would affect that expression. Methods: After molecular cloning of three VCAM1 promoter haplotype constructs, using pGL4 promoterless vectors, haplotype sequence was confirmed, by Sanger sequencing, prior to transfection. Transfection experiments for each construct were performed, with or without TNF-α stimulation, using EAhy926, and HBEC as macrovascular and microvascular endothelial cell models, respectively. Differences in promoter activity were assessed by luciferase reporter assay. RNA was extracted from non-transfected EAhy926 and HBEC cell cultures stimulated or not with TNF-a and/or hemin, and with or without HU treatment. RT-qPCR was performed to analyze VCAM1 expression. HMOX1 and NOS3 were also analyzed for comparison purposes. Results: Our results showed that two VCAM1 promoter haplotypes, previously associated with pediatric cerebral vasculopathy and hemolysis in SCA, increased promoter activity in transfected and TNF-α-stimulated EA.hy926 and HBEC cells, consistent with a higher VCAM1 expression in macro and microvascular settings. In non-transfected cells, we also observed TNF-a-induced VCAM1 overexpression as well as heme-induced overexpression of HMOX1 in both cell models. Heme did not affect VCAM1 nor NOS3 expression and the latter was also not affected by TNF-a stimulus. Hydroxyurea treatment lowered TNF-a-induced VCAM1 and NOS3 expression but did not affect heme-induced HMOX1 expression. Summary/Conclusion: These data further indicate that VCAM1 haplotypes we previously associated with pediatric cerebral vasculopathy and hemolysis in SCA, induce higher VCAM1 expression potentially affecting both cerebral and systemic vasculopathy risk. The differential endothelial expression of VCAM1, NOS3, and HMOX1 after proinflammatory and/or pro-oxidative stimuli also reinforces their genetic modulation role in SCA systemic vasculopathy.pt_PT
dc.description.versionN/Apt_PT
dc.identifier.urihttp://hdl.handle.net/10400.18/7965
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.subjectSickle Cell Anemiapt_PT
dc.subjectCerebrovascular Diseasept_PT
dc.subjectHemept_PT
dc.subjectVCAM-1pt_PT
dc.subjectModificadores Genéticospt_PT
dc.subjectAnemia das Células Falciformespt_PT
dc.subjectDrepanocitosept_PT
dc.subjectDoenças Genéticaspt_PT
dc.titleDifferential Endothelial VCAM1 Expression and Implications for Sickle Cell Anemia Vasculopathypt_PT
dc.typeconference object
dspace.entity.typePublication
oaire.citation.conferencePlace(online)pt_PT
oaire.citation.title26th EHA-European Hematology Association, 9-17 June 2021pt_PT
person.familyNameFaustino
person.givenNamePaula
person.identifier.ciencia-idF01A-353A-433E
person.identifier.orcid0000-0002-6269-4867
person.identifier.ridM-3519-2014
person.identifier.scopus-author-id8158641100
rcaap.rightsopenAccesspt_PT
rcaap.typeconferenceObjectpt_PT
relation.isAuthorOfPublication94303e78-8b7d-4e24-811d-3af3b1a4e330
relation.isAuthorOfPublication.latestForDiscovery94303e78-8b7d-4e24-811d-3af3b1a4e330

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