CAROLINA GONCALVES FERNANDES
Índice h a partir de 2011
4
Projetos de Pesquisa
Unidades Organizacionais
LIM/64, Hospital das Clínicas, Faculdade de Medicina
6 resultados
Resultados de Busca
Agora exibindo 1 - 6 de 6
- Golgi-independent routes support protein disulfide isomerase externalization in vascular smooth muscle cells(2017) ARAUJO, Thais L. S.; FERNANDES, Carolina G.; LAURINDO, Francisco R. M.Extracellular pools of intracellular molecular chaperones are increasingly evident. The peri/epicellular(pec) pool of the endoplasmic reticulum redox chaperone protein disulfide isomerase-A1(PDI) is involved in thrombosis and vascular remodeling, while PDI externalization routes remain elusive. In endothelial cells, vesicular-type PDI secretion involves classical and unconventional pathways, while in platelets PDI exocytosis involves actin cytoskeleton. However, little is known about pecPDI in vascular smooth muscle cells(VSMC). Here, we showed that VSMC display a robust cell-surface(cs) PDI pool, which binds to cs independently of electrostatic forces. However, contrarily to other cells, soluble secreted PDI pool was undetectable in VSMC. Calcium ionophore A23187 and TNF alpha enhanced VSMC csPDI. Furthermore, VSMC PDI externalization occurred via Golgi-bypass unconventional route, which was independent of cytoskeleton or lysosomes. Secreted PDI was absent in ex vivo wild-type mice aortas but markedly enhanced in PDI-overexpressing mice. Such characterization of VSMC pecPDI reinforces cell-type and context specific routes of PDI externalization.
- Conserved Gene Microsynteny Unveils Functional Interaction Between Protein Disulfide Isomerase and Rho Guanine-Dissociation Inhibitor Families(2017) MORETTI, Ana I. S.; PAVANELLI, Jessyca C.; NOLASCO, Patricia; LEISEGANG, Matthias S.; TANAKA, Leonardo Y.; FERNANDES, Carolina G.; WOSNIAK JR., Joao; KAJIHARA, Daniela; DIAS, Matheus H.; FERNANDES, Denise C.; JO, Hanjoong; Ngoc-Vinh Tran; EBERSBERGER, Ingo; BRANDES, Ralf P.; BONATTO, Diego; LAURINDO, Francisco R. M.Protein disulfide isomerases (PDIs) support endoplasmic reticulum redox protein folding and cell-surface thiol-redox control of thrombosis and vascular remodeling. The family prototype PDIA1 regulates NADPH oxidase signaling and cytoskeleton organization, however the related underlying mechanisms are unclear. Here we show that genes encoding human PDIA1 and its two paralogs PDIA8 and PDIA2 are each flanked by genes encoding Rho guanine-dissociation inhibitors (GDI), known regulators of RhoGTPases/cytoskeleton. Evolutionary histories of these three microsyntenic regions reveal their emergence by two successive duplication events of a primordial gene pair in the last common vertebrate ancestor. The arrangement, however, is substantially older, detectable in echinoderms, nematodes, and cnidarians. Thus, PDI/RhoGDI pairing in the same transcription orientation emerged early in animal evolution and has been largely maintained. PDI/RhoGDI pairs are embedded into conserved genomic regions displaying common cis-regulatory elements. Analysis of gene expression datasets supports evidence for PDI/RhoGDI coexpression in developmental/inflammatory contexts. PDIA1/RhoGDIa were co-induced in endothelial cells upon CRISP-R-promoted transcription activation of each pair component, and also in mouse arterial intima during flow-induced remodeling. We provide evidence for physical interaction between both proteins. These data support strong functional links between PDI and RhoGDI families, which likely maintained PDI/RhoGDI microsynteny along > 800-million years of evolution.
- PDIA1 acts as master organizer of NOX1/NOX4 balance and phenotype response in vascular smooth muscle(2021) FERNANDES, Denise C.; JR, Joao Wosniak; GONCALVES, Renata C.; TANAKA, Leonardo Y.; FERNANDES, Carolina G.; ZANATTA, Daniela B.; MATTOS, Ana Barbosa M. de; STRAUSS, Bryan E.; LAURINDO, Francisco R. M.Changes in vascular smooth muscle cell (VSMC) phenotype underlie disease pathophysiology and are strongly regulated by NOX NADPH oxidases, with NOX1 favoring synthetic proliferative phenotype and NOX4 supporting differentiation. Growth factor-triggered NOX1 expression/activity strictly depends on the chaperone oxidoreductase protein disulfide isomerase-A1 (PDIA1). Intracellular PDIA1 is required for VSMC migration and cytoskeleton organization, while extracellular PDIA1 fine-tunes cytoskeletal mechanoadaptation and vascular remodeling. We hypothesized that PDIA1 orchestrates NOX1/NOX4 balance and VSMC phenotype. Using an inducible PDIA1 overexpression model in VSMC, we showed that early PDIA1 overexpression (for 24-48 h) increased NOX1 expression, hydrogen peroxide steady-state levels and spontaneous VSMC migration distances. Sustained PDIA1 overexpression for 72 h and 96 h supported high NOX1 levels while also increasing NOX4 expression and, remarkably, switched VSMC phenotype to differentiation. Differentiation was preceded by increased nuclear myocardin and serum response factor-response element activation, with no change in cell viability. Both NOX1 and hydrogen peroxide were necessary for later PDIA1-induced VSMC differentiation. In primary VSMC, PDIA1 knockdown decreased nuclear myocardin and increased the proliferating cell nuclear antigen expression. Newly-developed PDIA1 -overexpressing mice (TgPDIA1) exhibited normal general and cardiovascular baseline phenotypes. However, in TgPDIA1 carotids, NOX1 was decreased while NOX4 and calponin expressions were enhanced, indicating overdifferentiation vs. normal carotids. Moreover, in a rabbit overdistension injury model during late vascular repair, PDIA1 silencing impaired VSMC redifferentiation and NOX1/NOX4 balance. Our results suggest a model in which PDIA1 acts as an upstream organizer of NOX1/NOX4 balance and related VSMC phenotype, accounting for baseline differentiation setpoint.
conferenceObject Phenotype Switch and Altered Mechanosignaling in Vascular Smooth Muscle Cells From Marfan Syndrome Mice(2017) NOLASCO, Patricia; FERNANDES, Carolina G.; FERRAZ, Mariana S.; SOARES, Lazaro R.; ALENCAR, Adriano M.; LAURINDO, Francisco R.conferenceObject Protein Disulfide Isomerase-A1 Overexpression Enhances Vascular Calcification in Mice(2017) PESCATORE, Luciana A.; NOLASCO, Patricia; FARIAS-SILVA, Elisngela; JENSEN, Leonardo; FERNANDES, Carolina G.; ALMEIDA, Youri E.; CARMO, Luciana S.; LAURINDO, Laurindo R.; LIBERMAN, Marcel- Impaired vascular smooth muscle cell force-generating capacity and phenotypic deregulation in Marfan Syndrome mice(2020) NOLASCO, Patricia; FERNANDES, Carolina Goncalves; RIBEIRO-SILVA, Joao Carlos; OLIVEIRA, Percillia V. S.; SACRINI, Mariana; BRITO, Isis Vasconcelos de; BESSA, Tiphany Coralie De; PEREIRA, Lygia V.; TANAKA, Leonardo Y.; ALENCAR, Adriano; LAURINDO, Francisco Rafael MartinsMechanisms whereby fibrillin-1 mutations determine thoracic aorta aneurysms/dissections (TAAD) in Marfan Syndrome (MFS) are unclear. Most aortic aneurysms evolve from mechanosignaling deregulation, converging to impaired vascular smooth muscle cell (VSMC) force-generating capacity accompanied by synthetic phenotype switch. However, little is known on VSMC mechanoresponses in MFS pathophysiology. Here, we investigated traction force-generating capacity in aortic VSMC cultured from 3-month old mg Delta(lpn) MFS mice, together with morpho-functional and proteomic data. Cultured MFS-VSMC depicted marked phenotype changes vs. wild-type (WT) VSMC, with overexpressed cell proliferation markers but either lower (calponin-1) or higher (SM alphaactin and SM22) differentiation marker expression. In parallel, the increased cell area and its complex non-fusiform shape suggested possible transition towards a mesenchymal-like phenotype, confirmed through several markers (e.g. N-cadherin, Slug). MFS-VSMC proteomic profile diverged from that of WT-VSMC particularly regarding lower expression of actin cytoskeleton-regulatory proteins. Accordingly, MFS-VSMC displayed lower traction force-generating capacity and impaired contractile moment at physiological substrate stiffness, and markedly attenuated traction force responses to enhanced substrate rigidity. Such impaired mechanoresponses correlated with decreased number, altered morphology and delocalization of focal adhesions, as well as dis-organized actin stress fiber network vs. WT-VSMC. In VSMC cultured from 6-month-old mice, phenotype changes were attenuated and both WT-VSMC and MFS-VSMC generated less traction force, presumably involving VSMC aging, but without evident senescence. In summary, MFS-VSMC display impaired force-generating capacity accompanying a mesenchymal-like phenotype switch connected to impaired cytoskeleton/focal adhesion organization. Thus, MFS-associated TAAD involves mechanoresponse impairment common to other TAAD types, but through distinct mechanisms.