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1.
J Neurosurg ; : 1-12, 2021 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-34740197

RESUMO

OBJECTIVE: Hereditary hemorrhagic telangiectasia is the only condition associated with multiple inherited brain arteriovenous malformations (AVMs). Therefore, a mouse model was developed with a genetics-based approach that conditionally deleted the causative activin receptor-like kinase 1 (Acvrl1 or Alk1) gene. Radiographic and histopathological findings were correlated, and AVM stability and hemorrhagic behavior over time were examined. METHODS: Alk1-floxed mice were crossed with deleter mice to generate offspring in which both copies of the Alk1 gene were deleted by Tagln-Cre to form brain AVMs in the mice. AVMs were characterized using MRI, MRA, and DSA. Brain AVMs were characterized histopathologically with latex dye perfusion, immunofluorescence, and Prussian blue staining. RESULTS: Brains of 55 Tagln-Cre+;Alk12f/2f mutant mice were categorized into three groups: no detectable vascular lesions (group 1; 23 of 55, 42%), arteriovenous fistulas (AVFs) with no nidus (group 2; 10 of 55, 18%), and nidal AVMs (group 3; 22 of 55, 40%). Microhemorrhage was observed on MRI or MRA in 11 AVMs (50%). AVMs had the angiographic hallmarks of early nidus opacification, a tangle of arteries and dilated draining veins, and rapid shunting of blood flow. Latex dye perfusion confirmed arteriovenous shunting in all AVMs and AVFs. Microhemorrhages were detected adjacent to AVFs and AVMs, visualized by iron deposition, Prussian blue staining, and macrophage infiltration using CD68 immunostaining. Brain AVMs were stable on serial MRI and MRA in group 3 mice (mean age at initial imaging 2.9 months; mean age at last imaging 9.5 months). CONCLUSIONS: Approximately 40% of transgenic mice satisfied the requirements of a stable experimental AVM model by replicating nidal anatomy, arteriovenous hemodynamics, and microhemorrhagic behavior. Transgenic mice with AVFs had a recognizable phenotype of hereditary hemorrhagic telangiectasia but were less suitable for experimental modeling. AVM pathogenesis can be understood as the combination of conditional Alk1 gene deletion during embryogenesis and angiogenesis that is hyperactive in developing and newborn mice, which translates to a congenital origin in most patients but an acquired condition in patients with a confluence of genetic and angiogenic events later in life. This study offers a novel experimental brain AVM model for future studies of AVM pathophysiology, growth, rupture, and therapeutic regression.

2.
Transl Stroke Res ; 2021 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-34674144

RESUMO

We have previously demonstrated that deletion of activin receptor-like kinase 1 (Alk1) or endoglin in a fraction of endothelial cells (ECs) induces brain arteriovenous malformations (bAVMs) in adult mice upon angiogenic stimulation. Here, we addressed three related questions: (1) could Alk1- mutant bone marrow (BM)-derived ECs (BMDECs) cause bAVMs? (2) is Alk1- ECs clonally expended during bAVM development? and (3) is the number of mutant ECs correlates to bAVM severity? For the first question, we transplanted BM from PdgfbiCreER;Alk12f/2f mice (EC-specific tamoxifen-inducible Cre with Alk1-floxed alleles) into wild-type mice, and then induced bAVMs by intra-brain injection of an adeno-associated viral vector expressing vascular endothelial growth factor and intra-peritoneal injection of tamoxifen. For the second question, clonal expansion was analyzed using PdgfbiCreER;Alk12f/2f;confetti+/- mice. For the third question, we titrated tamoxifen to limit Alk1 deletion and compared the severity of bAVM in mice treated with low and high tamoxifen doses. We found that wild-type mice with PdgfbiCreER;Alk12f/2f BM developed bAVMs upon VEGF stimulation and Alk1 gene deletion in BMDECs. We also observed clusters of ECs expressing the same confetti color within bAVMs and significant proliferation of Alk1- ECs at early stage of bAVM development, suggesting that Alk1- ECs clonally expanded by local proliferation. Tamoxifen dose titration revealed a direct correlation between the number of Alk1- ECs and the burden of dysplastic vessels in bAVMs. These results provide novel insights for the understanding of the mechanism by which a small fraction of Alk1 or endoglin mutant ECs contribute to development of bAVMs.

3.
Circ Res ; 127(9): 1122-1137, 2020 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-32762495

RESUMO

RATIONALE: Hereditary hemorrhagic telangiectasia (HHT) is a genetic disease caused by mutations in ENG, ALK1, or SMAD4. Since proteins from all 3 HHT genes are components of signal transduction of TGF-ß (transforming growth factor ß) family members, it has been hypothesized that HHT is a disease caused by defects in the ENG-ALK1-SMAD4 linear signaling. However, in vivo evidence supporting this hypothesis is scarce. OBJECTIVE: We tested this hypothesis and investigated the therapeutic effects and potential risks of induced-ALK1 or -ENG overexpression (OE) for HHT. METHODS AND RESULTS: We generated a novel mouse allele (ROSA26Alk1) in which HA (human influenza hemagglutinin)-tagged ALK1 and bicistronic eGFP expression are induced by Cre activity. We examined whether ALK1-OE using the ROSA26Alk1 allele could suppress the development of arteriovenous malformations (AVMs) in wounded adult skin and developing retinas of Alk1- and Eng-inducible knockout (iKO) mice. We also used a similar approach to investigate whether ENG-OE could rescue AVMs. Biochemical and immunofluorescence analyses confirmed the Cre-dependent OE of the ALK1-HA transgene. We could not detect any pathological signs in ALK1-OE mice up to 3 months after induction. ALK1-OE prevented the development of retinal AVMs and wound-induced skin AVMs in Eng-iKO as well as Alk1-iKO mice. ALK1-OE normalized expression of SMAD and NOTCH target genes in ENG-deficient endothelial cells (ECs) and restored the effect of BMP9 (bone morphogenetic protein 9) on suppression of phosphor-AKT levels in these endothelial cells. On the other hand, ENG-OE could not inhibit the AVM development in Alk1-iKO models. CONCLUSIONS: These data support the notion that ENG and ALK1 form a linear signaling pathway for the formation of a proper arteriovenous network during angiogenesis. We suggest that ALK1 OE or activation can be an effective therapeutic strategy for HHT. Further research is required to study whether this therapy could be translated into treatment for humans.


Assuntos
Receptores de Activinas Tipo II/metabolismo , Malformações Arteriovenosas/prevenção & controle , Células Endoteliais/metabolismo , Telangiectasia Hemorrágica Hereditária/metabolismo , Receptores de Activinas Tipo II/deficiência , Receptores de Activinas Tipo II/genética , Alelos , Animais , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Malformações Arteriovenosas/genética , Modelos Animais de Doenças , Endoglina/deficiência , Endoglina/genética , Endoglina/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Fator 2 de Diferenciação de Crescimento/metabolismo , Camundongos , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , RNA não Traduzido , Receptores Notch/genética , Receptores Notch/metabolismo , Vasos Retinianos/anormalidades , Transdução de Sinais , Pele/irrigação sanguínea , Pele/lesões , Proteína Smad4/genética , Proteína Smad4/metabolismo , Telangiectasia Hemorrágica Hereditária/genética , Fator de Crescimento Transformador beta
4.
Angiogenesis ; 23(3): 339-355, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32112176

RESUMO

BACKGROUND: TMEM100 is identified as a downstream gene of bone morphogenetic protein 9 (BMP9) signaling via activin receptor-like kinase 1 (ALK1), which is known to participate in lymphangiogenesis as well as angiogenesis. TMEM100 has been shown to be important for blood vessel formation and maintenance, but its role in the development of lymphatic vasculature remains unknown. The objective is to investigate the role of TMEM100 in development of the lymphatic system. METHODS AND RESULTS: Global Tmem100 gene deletion was induced by tamoxifen on 10.5 days post-coitus. Tmem100-inducible knockout (iKO) embryos in embryonic days (E)14.5-16.5 exhibited edema and blood-filled enlarged lymphatics with misconnections between veins and lymphatic vessels. For a reciprocal approach, we have generated a novel mouse line in which TMEM100 overexpression (OE) can be induced in endothelial cells by intercrossing with Tie2-Cre driver. TMEM100-OE embryos at E12.5-14.5 exhibited edema with small size and number of lymphatic vessels, the exact opposite phenotypes of Tmem100-iKOs. In Tmem100-iKO embryos, the number of progenitors of lymphatic endothelial cells (LECs) in the cardinal vein was increased, while it was decreased in TMEM100-OE embryos. The activity of NOTCH signaling, which limits the number of progenitors of LECs in the cardinal vein, was decreased in Tmem100-iKO embryos, whereas it was increased in TMEM100-OE embryos. CONCLUSION: TMEM100 plays an important role in the specification of LECs in the cardinal veins, at least in part, by regulating the NOTCH signaling.


Assuntos
Células Endoteliais/metabolismo , Células Progenitoras Endoteliais/metabolismo , Vasos Linfáticos/metabolismo , Proteínas de Membrana/metabolismo , Animais , Feminino , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout
5.
J Clin Invest ; 130(2): 942-957, 2020 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-31689244

RESUMO

Hereditary hemorrhagic telangiectasia (HHT), a genetic bleeding disorder leading to systemic arteriovenous malformations (AVMs), is caused by loss-of-function mutations in the ALK1/ENG/Smad1/5/8 pathway. Evidence suggests that HHT pathogenesis strongly relies on overactivated PI3K/Akt/mTOR and VEGFR2 pathways in endothelial cells (ECs). In the BMP9/10-immunoblocked (BMP9/10ib) neonatal mouse model of HHT, we report here that the mTOR inhibitor, sirolimus, and the receptor tyrosine kinase inhibitor, nintedanib, could synergistically fully block, but also reversed, retinal AVMs to avert retinal bleeding and anemia. Sirolimus plus nintedanib prevented vascular pathology in the oral mucosa, lungs, and liver of the BMP9/10ib mice, as well as significantly reduced gastrointestinal bleeding and anemia in inducible ALK1-deficient adult mice. Mechanistically, in vivo in BMP9/10ib mouse ECs, sirolimus and nintedanib blocked the overactivation of mTOR and VEGFR2, respectively. Furthermore, we found that sirolimus activated ALK2-mediated Smad1/5/8 signaling in primary ECs - including in HHT patient blood outgrowth ECs - and partially rescued Smad1/5/8 activity in vivo in BMP9/10ib mouse ECs. These data demonstrate that the combined correction of endothelial Smad1/5/8, mTOR, and VEGFR2 pathways opposes HHT pathogenesis. Repurposing of sirolimus plus nintedanib might provide therapeutic benefit in patients with HHT.


Assuntos
Células Endoteliais , Indóis/farmacologia , Sirolimo/farmacologia , Proteína Smad1 , Proteína Smad5 , Proteína Smad8 , Serina-Treonina Quinases TOR , Telangiectasia Hemorrágica Hereditária , Receptor 2 de Fatores de Crescimento do Endotélio Vascular , Receptores de Activinas Tipo II/genética , Receptores de Activinas Tipo II/metabolismo , Animais , Proteínas Morfogenéticas Ósseas/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Fator 2 de Diferenciação de Crescimento/genética , Fator 2 de Diferenciação de Crescimento/metabolismo , Camundongos , Camundongos Knockout , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Proteína Smad1/genética , Proteína Smad1/metabolismo , Proteína Smad5/genética , Proteína Smad5/metabolismo , Proteína Smad8/genética , Proteína Smad8/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Telangiectasia Hemorrágica Hereditária/tratamento farmacológico , Telangiectasia Hemorrágica Hereditária/genética , Telangiectasia Hemorrágica Hereditária/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
6.
Int J Mol Sci ; 20(21)2019 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-31731545

RESUMO

Arteriovenous malformations (AVMs) are abnormal connections of vessels that shunt blood directly from arteries into veins. Rupture of brain AVMs (bAVMs) can cause life-threatening intracranial bleeding. Even though the majority of bAVM cases are sporadic without a family history, some cases are familial. Most of the familial cases of bAVMs are associated with a genetic disorder called hereditary hemorrhagic telangiectasia (HHT). The mechanism of bAVM formation is not fully understood. The most important advances in bAVM basic science research is the identification of somatic mutations of genes in RAS-MAPK pathways. However, the mechanisms by which mutations of these genes lead to AVM formation are largely unknown. In this review, we summarized the latest advance in bAVM studies and discussed some pathways that play important roles in bAVM pathogenesis. We also discussed the therapeutic implications of these pathways.


Assuntos
Malformações Arteriovenosas Intracranianas , Hemorragias Intracranianas , Sistema de Sinalização das MAP Quinases/genética , Mutação , Telangiectasia Hemorrágica Hereditária , Feminino , Humanos , Malformações Arteriovenosas Intracranianas/genética , Malformações Arteriovenosas Intracranianas/metabolismo , Malformações Arteriovenosas Intracranianas/patologia , Malformações Arteriovenosas Intracranianas/terapia , Hemorragias Intracranianas/genética , Hemorragias Intracranianas/metabolismo , Hemorragias Intracranianas/patologia , Hemorragias Intracranianas/terapia , Masculino , Telangiectasia Hemorrágica Hereditária/genética , Telangiectasia Hemorrágica Hereditária/metabolismo , Telangiectasia Hemorrágica Hereditária/patologia , Telangiectasia Hemorrágica Hereditária/terapia
8.
Angiogenesis ; 22(1): 145-155, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30191360

RESUMO

Pazopanib (Votrient) is an orally administered tyrosine kinase inhibitor that blocks VEGF receptors potentially serving as anti-angiogenic treatment for hereditary hemorrhagic telangiectasia (HHT). We report a prospective, multi-center, open-label, dose-escalating study [50 mg, 100 mg, 200 mg, and 400 mg], designed as a proof-of-concept study to demonstrate efficacy of pazopanib on HHT-related bleeding, and to measure safety. Patients, recruited at 5 HHT Centers, required ≥ 2 Curacao criteria AND [anemia OR severe epistaxis with iron deficiency]. Co-primary outcomes, hemoglobin (Hgb) and epistaxis severity, were measured during and after treatment, and compared to baseline. Safety monitoring occurred every 1.5 weeks. Seven patients were treated with 50 mg pazopanib daily. Six/seven showed at least 50% decrease in epistaxis duration relative to baseline at some point during study; 3 showed at least 50% decrease in duration during Weeks 11 and 12. Six patients showed a decrease in ESS of > 0.71 (MID) relative to baseline at some point during study; 3/6 showed a sustained improvement. Four patients showed > 2 gm improvement in Hgb relative to baseline at one or more points during study. Health-related QOL scores improved on all SF-36 domains at Week 6 and/or Week 12, except general health (unchanged). There were 19 adverse events (AE) including one severe AE (elevated LFTs, withdrawn from dosing at 43 days); with no serious AE. In conclusion, we observed an improvement in Hgb and/or epistaxis in all treated patients. This occurred at a dose much lower than typically used for oncologic indications, with no serious AE. Further studies of pazopanib efficacy are warranted.


Assuntos
Hemorragia , Pirimidinas , Sulfonamidas , Telangiectasia Hemorrágica Hereditária , Adulto , Feminino , Hemorragia/sangue , Hemorragia/tratamento farmacológico , Humanos , Masculino , Pessoa de Meia-Idade , Pirimidinas/administração & dosagem , Pirimidinas/farmacocinética , Sulfonamidas/administração & dosagem , Sulfonamidas/farmacocinética , Telangiectasia Hemorrágica Hereditária/sangue , Telangiectasia Hemorrágica Hereditária/tratamento farmacológico
9.
J Am Heart Assoc ; 7(21): e009514, 2018 11 06.
Artigo em Inglês | MEDLINE | ID: mdl-30571376

RESUMO

Background Hereditary hemorrhagic telangiectasia ( HHT ) is a rare genetic vascular disorder caused by mutations in endoglin ( ENG ), activin receptor-like kinase 1 ( ACVRL 1; ALK 1), or SMAD 4. Major clinical symptoms of HHT are arteriovenous malformations ( AVM s) found in the brain, lungs, visceral organs, and mucosal surface. Animal models harboring mutations in Eng or Alk1 recapitulate all of these HHT clinical symptoms and have been useful resources for studying mechanisms and testing potential drugs. However, animal models representing SMAD 4 mutations have been lacking. The goal of this study is to evaluate Smad4-inducible knockout ( iKO ) mice as an animal model of HHT and compare the phenotypes with other established HHT animal models. Methods and Results Global Smad4 deletion was induced at neonatal and adult stages, and hemoglobin levels, gastrointestinal hemorrhage, and presence of aberrant arteriovenous connections were examined. Neonatal Smad4- iKO mice exhibited signs of gastrointestinal bleeding and AVM s in the brain, intestine, nose, and retina. The radial expansion was decreased, and AVM s were detected on both distal and proximal retinal vasculature of Smad4- iKO s. Aberrant smooth muscle actin staining was observed in the initial stage AVM s and their connecting veins, indicating abnormal arterial flow to veins. In adult mice, Smad4 deficiency caused gastrointestinal bleeding and AVM s along the gastrointestinal tract and wounded skin. HHT -related phenotypes of Smad4- iKO s appeared to be comparable with those found in Alk1- iKO and Eng- iKO mice. Conclusions These data further confirm that SMAD signaling is crucial for normal arteriovenous network formation, and Smad4- iKO will be an alternative animal model of AVM research associated with HHT .


Assuntos
Malformações Arteriovenosas/genética , Modelos Animais de Doenças , Camundongos , Proteína Smad4/deficiência , Proteína Smad4/genética , Telangiectasia Hemorrágica Hereditária/genética , Fatores Etários , Animais , Animais Recém-Nascidos , Camundongos Knockout , Fenótipo
10.
Cardiovasc Res ; 113(13): 1677-1687, 2017 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-29016745

RESUMO

Aims: Chemokine CXCL12 (stromal derived factor 1: SDF1) has been shown to play important roles in various processes of cardiovascular development. In recent avian studies, CXCL12 signalling has been implicated in guidance of cardiac neural crest cells for their participation in the development of outflow tract and cardiac septum. The goal of this study is to investigate the extent to which CXCL12 signalling contribute to the development of aortic arch and pulmonary arteries in mammals. Methods and results: Novel Cxcl12-LacZ reporter and conditional alleles were generated. Using whole mount X-gal staining with the reporter allele and vascular casting techniques, we show that the domain branching pattern of pulmonary arteries in Cxcl12-null mice is completely disrupted and discordant with that of pulmonary veins and airways. Cxcl12-null mice also displayed abnormal and superfluous arterial branches from the aortic arch. The early steps of pharyngeal arch remodelling in Cxcl12-null mice appeared to be unaffected, but vertebral arteries were often missing and prominent aberrant arteries were present parallel to carotid arteries or trachea, similar to aberrant vertebral artery or thyroid ima artery, respectively. Analysis with computed tomography not only confirmed the results from vascular casting studies but also identified abnormal systemic arterial supply to lungs in the Cxcl12-null mice. Tie2-Cre mediated Cxcr4 deletion phenocopied the Cxcl12-null phenotypes, indicating that CXCR4 is the primary receptor for arterial patterning, whereas Cxcl12 or Cxcr4 deletion by Wnt1-Cre did not affect aortic arch patterning. Conclusion: CXCL12-CXCR4 signalling is essential for the correct patterning of aortic arches and pulmonary arteries during development. Superfluous arteries in Cxcl12-null lungs and the aortic arch infer a role of CXCL12 in protecting arteries from uncontrolled sprouting during development of the arterial system.


Assuntos
Aorta Torácica/metabolismo , Padronização Corporal , Quimiocina CXCL12/metabolismo , Artéria Pulmonar/metabolismo , Receptores CXCR4/metabolismo , Malformações Vasculares/metabolismo , Animais , Aorta Torácica/anormalidades , Aorta Torácica/diagnóstico por imagem , Aortografia/métodos , Quimiocina CXCL12/deficiência , Quimiocina CXCL12/genética , Angiografia por Tomografia Computadorizada , Regulação da Expressão Gênica no Desenvolvimento , Genótipo , Idade Gestacional , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neovascularização Fisiológica , Fenótipo , Artéria Pulmonar/anormalidades , Artéria Pulmonar/diagnóstico por imagem , Receptores CXCR4/deficiência , Receptores CXCR4/genética , Transdução de Sinais , Malformações Vasculares/diagnóstico por imagem , Malformações Vasculares/genética
11.
Cardiovasc Pathol ; 31: 26-33, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28820968

RESUMO

INTRODUCTION: Activin receptor-like kinase 1 (ALK1) mediates signaling via the transforming growth factor beta-1 (TGFß1), a pro-fibrogenic cytokine. No studies have defined a role for ALK1 in heart failure. HYPOTHESIS: We tested the hypothesis that reduced ALK1 expression promotes maladaptive cardiac remodeling in heart failure. METHODS AND RESULTS: In patients with advanced heart failure referred for left ventricular (LV) assist device implantation, LV Alk1 mRNA and protein levels were lower than control LV obtained from patients without heart failure. To investigate the role of ALK1 in heart failure, Alk1 haploinsufficient (Alk1+/-) and wild-type (WT) mice were studied 2 weeks after severe transverse aortic constriction (TAC). LV and lung weights were higher in Alk1+/- mice after TAC. Cardiomyocyte area and LV mRNA levels of brain natriuretic peptide and ß-myosin heavy chain were increased similarly in Alk1+/- and WT mice after TAC. Alk-1 mice exhibited reduced Smad 1 phosphorylation and signaling compared to WT mice after TAC. Compared to WT, LV fibrosis and Type 1 collagen mRNA and protein levels were higher in Alk1+/- mice. LV fractional shortening was lower in Alk1+/- mice after TAC. CONCLUSIONS: Reduced expression of ALK1 promotes cardiac fibrosis and impaired LV function in a murine model of heart failure. Further studies examining the role of ALK1 and ALK1 inhibitors on cardiac remodeling are required.


Assuntos
Receptores de Activinas Tipo II/metabolismo , Insuficiência Cardíaca/metabolismo , Miocárdio/patologia , Remodelação Ventricular/fisiologia , Receptores de Ativinas Tipo I/metabolismo , Adulto , Animais , Feminino , Fibrose/metabolismo , Insuficiência Cardíaca/patologia , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Miocárdio/metabolismo
12.
Heart Vessels ; 32(5): 628-636, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28213819

RESUMO

Activin like kinase-1 (AlK-1) mediates signaling via the transforming growth factor beta (TGFß) family of ligands. AlK-1 activity promotes endothelial proliferation and migration. Reduced AlK-1 activity is associated with arteriovenous malformations. No studies have examined the effect of global AlK-1 deletion on indices of cardiac remodeling. We hypothesized that reduced levels of AlK-1 promote maladaptive cardiac remodeling. To test this hypothesis, we employed AlK-1 conditional knockout mice (cKO) harboring the ROSA26-CreER knock-in allele, whereby a single dose of intraperitoneal tamoxifen triggered ubiquitous Cre recombinase-mediated excision of floxed AlK-1 alleles. Tamoxifen treated wild-type (WT-TAM; n = 5) and vehicle treated AlK-1-cKO mice (cKO-CON; n = 5) served as controls for tamoxifen treated AlK-1-cKO mice (cKO-TAM; n = 15). AlK-1 cKO-TAM mice demonstrated reduced 14-day survival compared to cKO-CON controls (13 vs 100%, respectively, p < 0.01). Seven days after treatment, cKO-TAM mice exhibited reduced left ventricular (LV) fractional shortening, progressive LV dilation, and gastrointestinal bleeding. After 14 days total body mass was reduced, but LV and lung mass increased in cKO-TAM not cKO-CON mice. Peak LV systolic pressure, contractility, and arterial elastance were reduced, but LV end-diastolic pressure and stroke volume were increased in cKO-TAM, not cKO-CON mice. LV AlK-1 mRNA levels were reduced in cKO-TAM, not cKO-CON mice. LV levels of other TGFß-family ligands and receptors (AlK5, TBRII, BMPRII, Endoglin, BMP7, BMP9, and TGFß1) were unchanged between groups. Cardiomyocyte area and LV levels of BNP were increased in cKO-TAM mice, but LV levels of ß-MHC and SERCA were unchanged. No increase in markers of cardiac fibrosis, Type I collagen, CTGF, or PAI-1, were observed between groups. No differences were observed for any variable studied between cKO-CON and WT-TAM mice. Global deletion of AlK-1 is associated with the development of high output heart failure without maladaptive remodeling. Future studies exploring the functional role of AlK-1 in cardiac remodeling independent of systemic AVMs are required.


Assuntos
Receptores de Ativinas Tipo I/genética , Regulação da Expressão Gênica , Insuficiência Cardíaca/genética , RNA/genética , Função Ventricular Esquerda/fisiologia , Remodelação Ventricular/fisiologia , Receptores de Ativinas Tipo I/biossíntese , Receptores de Activinas Tipo II , Alelos , Animais , Modelos Animais de Doenças , Progressão da Doença , Insuficiência Cardíaca/metabolismo , Insuficiência Cardíaca/fisiopatologia , Camundongos Knockout , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais
13.
Nat Commun ; 8: 14477, 2017 02 16.
Artigo em Inglês | MEDLINE | ID: mdl-28205519

RESUMO

Non-alcoholic fatty liver disease (NAFLD) is a common metabolic disorder in obese individuals. Adenine nucleotide translocase (ANT) exchanges ADP/ATP through the mitochondrial inner membrane, and Ant2 is the predominant isoform expressed in the liver. Here we demonstrate that targeted disruption of Ant2 in mouse liver enhances uncoupled respiration without damaging mitochondrial integrity and liver functions. Interestingly, liver specific Ant2 knockout mice are leaner and resistant to hepatic steatosis, obesity and insulin resistance under a lipogenic diet. Protection against fatty liver is partially recapitulated by the systemic administration of low-dose carboxyatractyloside, a specific inhibitor of ANT. Targeted manipulation of hepatic mitochondrial metabolism, particularly through inhibition of ANT, may represent an alternative approach in NAFLD and obesity treatment.


Assuntos
Translocador 2 do Nucleotídeo Adenina/metabolismo , Trifosfato de Adenosina/metabolismo , Fígado Gorduroso/metabolismo , Resistência à Insulina , Mitocôndrias Hepáticas/metabolismo , Substâncias Protetoras/metabolismo , Translocador 2 do Nucleotídeo Adenina/genética , Animais , Atractilosídeo/análogos & derivados , Dieta Hiperlipídica , Modelos Animais de Doenças , Fígado Gorduroso/terapia , Feminino , Técnica Clamp de Glucose , Hiperinsulinismo , Metabolismo dos Lipídeos , Lipogênese , Fígado/metabolismo , Fígado/patologia , Masculino , Camundongos , Camundongos Knockout , Membranas Mitocondriais/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Hepatopatia Gordurosa não Alcoólica/terapia , Obesidade/metabolismo , Obesidade/terapia , Ácido Pirúvico/metabolismo
15.
Sci Rep ; 6: 35444, 2016 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-27739498

RESUMO

Transforming growth factor (TGF)-ß signaling disorder has emerged as a common molecular signature for aortic aneurysm development. The timing of postnatal maturation plays a key role in dictating the biological outcome of TGF-ß signaling disorders in the aortic wall. In this study, we investigated the impact of deficiency of TGFß receptors on the structural homeostasis of mature aortas. We used an inducible Cre-loxP system driven by a Myh11 promoter to delete Tgfbr1, Tgfbr2, or both in smooth muscle cells (SMCs) of adult mice. TGFBR1 deficiency resulted in rapid and severe aneurysmal degeneration, with 100% penetrance of ascending thoracic aortas, whereas TGFBR2 deletion only caused mild aortic pathology with low (26%) lesion prevalence. Removal of TGFBR2 attenuated the aortic pathology caused by TGFBR1 deletion and correlated with a reduction of early ERK phosphorylation. In addition, the production of angiotensin (Ang)-converting enzyme was upregulated in TGFBR1 deficient aortas at the early stage of aneurysmal degeneration. Inhibition of ERK phosphorylation or blockade of AngII type I receptor AT1R prevented aneurysmal degeneration of TGFBR1 deficient aortas. In conclusion, loss of SMC-Tgfbr1 triggers multiple deleterious pathways, including abnormal TGFBR2, ERK, and AngII/AT1R signals that disrupt aortic wall homeostasis to cause aortic aneurysm formation.


Assuntos
Aneurisma Aórtico/genética , Sistema de Sinalização das MAP Quinases , Miócitos de Músculo Liso/metabolismo , Proteínas Serina-Treonina Quinases/genética , Receptores de Fatores de Crescimento Transformadores beta/genética , Angiotensina II/metabolismo , Animais , Aorta/metabolismo , Aorta/patologia , Aneurisma Aórtico/metabolismo , Deleção de Genes , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Peptidil Dipeptidase A/metabolismo , Proteínas Serina-Treonina Quinases/deficiência , Proteínas Serina-Treonina Quinases/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptor do Fator de Crescimento Transformador beta Tipo II , Receptores de Angiotensina/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/deficiência , Receptores de Fatores de Crescimento Transformadores beta/metabolismo
16.
JAMA ; 316(9): 943-51, 2016 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-27599329

RESUMO

IMPORTANCE: Epistaxis is a major factor negatively affecting quality of life in patients with hereditary hemorrhagic telangiectasia (HHT; also known as Osler-Weber-Rendu disease). Optimal treatment for HHT-related epistaxis is uncertain. OBJECTIVE: To determine whether topical therapy with any of 3 drugs with differing mechanisms of action is effective in reducing HHT-related epistaxis. DESIGN, SETTING, AND PARTICIPANTS: The North American Study of Epistaxis in HHT was a double-blind, placebo-controlled randomized clinical trial performed at 6 HHT centers of excellence. From August 2011 through March 2014, there were 121 adult patients who met the clinical criteria for HHT and had experienced HHT-related epistaxis with an Epistaxis Severity Score of at least 3.0. Follow-up was completed in September 2014. INTERVENTIONS: Patients received twice-daily nose sprays for 12 weeks with either bevacizumab 1% (4 mg/d), estriol 0.1% (0.4 mg/d), tranexamic acid 10% (40 mg/d), or placebo (0.9% saline). MAIN OUTCOMES AND MEASURES: The primary outcome was median weekly epistaxis frequency during weeks 5 through 12. Secondary outcomes included median duration of epistaxis during weeks 5 through 12, Epistaxis Severity Score, level of hemoglobin, level of ferritin, need for transfusion, emergency department visits, and treatment failure. RESULTS: Among the 121 patients who were randomized (mean age, 52.8 years [SD, 12.9 years]; 44% women with a median of 7.0 weekly episodes of epistaxis [interquartile range {IQR}, 3.0-14.0]), 106 patients completed the study duration for the primary outcome measure (43 were women [41%]). Drug therapy did not significantly reduce epistaxis frequency (P = .97). After 12 weeks of treatment, the median weekly number of bleeding episodes was 7.0 (IQR, 4.5-10.5) for patients in the bevacizumab group, 8.0 (IQR, 4.0-12.0) for the estriol group, 7.5 (IQR, 3.0-11.0) for the tranexamic acid group, and 8.0 (IQR, 3.0-14.0) for the placebo group. No drug treatment was significantly different from placebo for epistaxis duration. All groups had a significant improvement in Epistaxis Severity Score at weeks 12 and 24. There were no significant differences between groups for hemoglobin level, ferritin level, treatment failure, need for transfusion, or emergency department visits. CONCLUSIONS AND RELEVANCE: Among patients with HHT, there were no significant between-group differences in the use of topical intranasal treatment with bevacizumab vs estriol vs tranexamic acid vs placebo and epistaxis frequency. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT01408030.


Assuntos
Inibidores da Angiogênese/administração & dosagem , Bevacizumab/administração & dosagem , Epistaxe/tratamento farmacológico , Telangiectasia Hemorrágica Hereditária/complicações , Administração Intranasal , Administração Tópica , Adulto , Idoso , Antifibrinolíticos/administração & dosagem , Transfusão de Sangue , Método Duplo-Cego , Epistaxe/etiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Qualidade de Vida , Índice de Gravidade de Doença , Ácido Tranexâmico/administração & dosagem , Resultado do Tratamento
17.
Oncotarget ; 7(35): 55957-55969, 2016 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-27517154

RESUMO

Age-related macular degeneration (AMD) is the leading cause of blindness in aging populations of industrialized countries. The drawbacks of inhibitors of vascular endothelial growth factor (VEGFs) currently used for the treatment of AMD, which include resistance and potential serious side-effects, require the identification of new therapeutic targets to modulate angiogenesis. BMP9 signaling through the endothelial Alk1 serine-threonine kinase receptor modulates the response of endothelial cells to VEGF and promotes vessel quiescence and maturation during development. Here, we show that BMP9/Alk1 signaling inhibits neovessel formation in mouse models of pathological ocular angiogenesis relevant to AMD. Activating Alk1 signaling in laser-induced choroidal neovascularization (CNV) and oxygen-induced retinopathy (OIR) inhibited neovascularization and reduced the volume of vascular lesions. Alk1 signaling was also found to interfere with VEGF signaling in endothelial cells whereas BMP9 potentiated the inhibitory effects of VEGFR2 signaling blockade, both in OIR and laser-induced CNV. Together, our data show that targeting BMP9/Alk1 efficiently prevents the growth of neovessels in AMD models and introduce a new approach to improve conventional anti-VEGF therapies.


Assuntos
Receptores de Ativinas Tipo I/metabolismo , Envelhecimento/patologia , Fator 2 de Diferenciação de Crescimento/metabolismo , Degeneração Macular/patologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptores de Ativinas Tipo I/genética , Receptores de Activinas Tipo II , Adenoviridae/genética , Animais , Neovascularização de Coroide/patologia , Modelos Animais de Doenças , Células Endoteliais/patologia , Ensaio de Imunoadsorção Enzimática , Vetores Genéticos/genética , Fator 2 de Diferenciação de Crescimento/genética , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Doenças Retinianas/patologia , Transdução de Sinais , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores
18.
Sci Rep ; 6: 27932, 2016 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-27305836

RESUMO

The specification of left-right asymmetry of the visceral organs is precisely regulated. The earliest breakage of left-right symmetry occurs as the result of leftward flow generated by asymmetric beating of nodal cilia, which eventually induces asymmetric Nodal/Lefty/Pitx2 expression on the left side of the lateral plate mesoderm. PIERCE1 has been identified as a p53 target gene involved in the DNA damage response. In this study, we found that Pierce1-null mice exhibit severe laterality defects, including situs inversus totalis and heterotaxy with randomized situs and left and right isomerisms. The spectrum of laterality defects was closely correlated with randomized expression of Nodal and its downstream genes, Lefty1/2 and Pitx2. The phenotype of Pierce1-null mice most closely resembled that of mutant mice with impaired ciliogenesis and/or ciliary motility of the node. We also found the loss of asymmetric expression of Cerl2, the earliest flow-responding gene in the node of Pierce1-null embryos. The results suggest that Pierce1-null embryos have defects in generating a symmetry breaking signal including leftward nodal flow. This is the first report implicating a role for PIERCE1 in the symmetry-breaking step of left-right asymmetry specification.


Assuntos
Padronização Corporal , Proteínas de Ciclo Celular/metabolismo , Animais , Proteínas de Ciclo Celular/deficiência , Regulação da Expressão Gênica no Desenvolvimento , Mesoderma , Camundongos , Camundongos Knockout
19.
Angiogenesis ; 19(4): 451-461, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27325285

RESUMO

An abnormally high number of macrophages are present in human brain arteriovenous malformations (bAVM) with or without evidence of prior hemorrhage, causing unresolved inflammation that may enhance abnormal vascular remodeling and exacerbate the bAVM phenotype. The reasons for macrophage accumulation at the bAVM sites are not known. We tested the hypothesis that persistent infiltration and pro-inflammatory differentiation of monocytes in angiogenic tissues increase the macrophage burden in bAVM using two mouse models and human monocytes. Mouse bAVM was induced through deletion of AVM causative genes, Endoglin (Eng) globally or Alk1 focally, plus brain focal angiogenic stimulation. An endothelial cell and vascular smooth muscle cell co-culture system was used to analyze monocyte differentiation in the angiogenic niche. After angiogenic stimulation, the Eng-deleted mice had fewer CD68(+) cells at 2 weeks (P = 0.02), similar numbers at 4 weeks (P = 0.97), and more at 8 weeks (P = 0.01) in the brain angiogenic region compared with wild-type (WT) mice. Alk1-deficient mice also had a trend toward more macrophages/microglia 8 weeks (P = 0.064) after angiogenic stimulation and more RFP(+) bone marrow-derived macrophages than WT mice (P = 0.01). More CD34(+) cells isolated from peripheral blood of patients with ENG or ALK1 gene mutation differentiated into macrophages than those from healthy controls (P < 0.001). These data indicate that persistent infiltration and pro-inflammatory differentiation of monocytes might contribute to macrophage accumulation in bAVM. Blocking macrophage homing to bAVM lesions should be tested as a strategy to reduce the severity of bAVM.


Assuntos
Malformações Arteriovenosas Intracranianas/patologia , Monócitos/patologia , Receptores de Ativinas Tipo I/deficiência , Receptores de Ativinas Tipo I/genética , Receptores de Activinas Tipo II , Animais , Diferenciação Celular , Técnicas de Cocultura , Modelos Animais de Doenças , Endoglina/deficiência , Endoglina/genética , Células Endoteliais/patologia , Humanos , Malformações Arteriovenosas Intracranianas/genética , Malformações Arteriovenosas Intracranianas/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Miócitos de Músculo Liso/patologia , Neovascularização Patológica/genética
20.
Arterioscler Thromb Vasc Biol ; 36(4): 707-17, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26821948

RESUMO

OBJECTIVE: To determine the role of Gja5 that encodes for the gap junction protein connexin40 in the generation of arteriovenous malformations in the hereditary hemorrhagic telangiectasia type 2 (HHT2) mouse model. APPROACH AND RESULTS: We identified GJA5 as a target gene of the bone morphogenetic protein-9/activin receptor-like kinase 1 signaling pathway in human aortic endothelial cells and importantly found that connexin40 levels were particularly low in a small group of patients with HHT2. We next took advantage of the Acvrl1(+/-) mutant mice that develop lesions similar to those in patients with HHT2 and generated Acvrl1(+/-); Gja5(EGFP/+) mice. Gja5 haploinsufficiency led to vasodilation of the arteries and rarefaction of the capillary bed in Acvrl1(+/-) mice. At the molecular level, we found that reduced Gja5 in Acvrl1(+/-) mice stimulated the production of reactive oxygen species, an important mediator of vessel remodeling. To normalize the altered hemodynamic forces in Acvrl1(+/-); Gja5(EGFP/+) mice, capillaries formed transient arteriovenous shunts that could develop into large malformations when exposed to environmental insults. CONCLUSIONS: We identified GJA5 as a potential modifier gene for HHT2. Our findings demonstrate that Acvrl1 haploinsufficiency combined with the effects of modifier genes that regulate vessel caliber is responsible for the heterogeneity and severity of the disease. The mouse models of HHT have led to the proposal that 3 events-heterozygosity, loss of heterozygosity, and angiogenic stimulation-are necessary for arteriovenous malformation formation. Here, we present a novel 3-step model in which pathological vessel caliber and consequent altered blood flow are necessary events for arteriovenous malformation development.


Assuntos
Receptores de Activinas Tipo II/metabolismo , Receptores de Ativinas Tipo I/metabolismo , Malformações Arteriovenosas/enzimologia , Conexinas/metabolismo , Células Endoteliais/enzimologia , Vasos Retinianos/enzimologia , Telangiectasia Hemorrágica Hereditária/enzimologia , Receptores de Ativinas Tipo I/genética , Receptores de Activinas Tipo II/genética , Animais , Malformações Arteriovenosas/genética , Malformações Arteriovenosas/patologia , Células Cultivadas , Conexinas/genética , Modelos Animais de Doenças , Predisposição Genética para Doença , Haploinsuficiência , Humanos , Camundongos Mutantes , Camundongos Transgênicos , Neovascularização Patológica , Fenótipo , Interferência de RNA , Espécies Reativas de Oxigênio/metabolismo , Vasos Retinianos/patologia , Transdução de Sinais , Telangiectasia Hemorrágica Hereditária/genética , Telangiectasia Hemorrágica Hereditária/patologia , Transfecção , Remodelação Vascular
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