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1.
Aging (Albany NY) ; 13(7): 10275-10288, 2021 04 04.
Artigo em Inglês | MEDLINE | ID: mdl-33819188

RESUMO

Coupling between osteogenesis and angiogenesis determines bone morphology. A decrease in the osteogenic ability of bone marrow mesenchymal stem cells (BMSCs) is one of the underlying causes of senile osteoporosis (OP). Here, we investigated the involvement of circular RNAs (circRNAs) in the osteogenic differentiation of BMSCs and the pathogenesis of senile OP. We sequenced RNA and found decreases expression of hsa_circ_0006215 in BMSCs from patients with OP. We further assessed the role of hsa_circ_0006215 in the osteogenic differentiation of BMSCs using lentivirus-mediated hsa_circ_0006215 overexpression and knockdown. Overexpression of hsa_circ_0006215 promoted the osteogenic differentiation of BMSCs. Luciferase reporter and RNA pull-down assays revealed that hsa_circ_0006215 bound to miRNA-942-5p and thus regulated RUNX2 and vascular endothelial growth factor (VEGF) expression in BMSCs. We assessed osteogenesis and vascular coupling in co-cultured cells, and the role of hsa_circ_0006215 in bone formation in vivo using a cortical bone defect model. We found that hsa_circ_0006215 promoted bone defect repair. Overall, our results showed that hsa_circ_0006215 has an important function in osteogenesis and could be a novel target for treating senile OP.


Assuntos
Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/metabolismo , Osteogênese/fisiologia , RNA Circular/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Diferenciação Celular/fisiologia , Regulação da Expressão Gênica/fisiologia , Humanos , Neovascularização Fisiológica/fisiologia , Osteoporose/metabolismo
2.
Aging (Albany NY) ; 13(6): 7781-7799, 2021 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-33764901

RESUMO

Angiogenesis - the formation of new blood capillaries- is impaired in aging animals and contributes to the pathogenesis of age-related diseases. A transcription factor, Twist1, contributes to the pathogenesis of age- and angiogenesis-related diseases such as pulmonary fibrosis and atherosclerosis. However, the mechanism by which Twist1 controls age-dependent decline in angiogenesis remains unclear. In this report, we have demonstrated that the levels of Twist1 are higher, while the expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) that stimulates angiogenesis, is lower in endothelial cells (ECs) isolated from aged human adipose tissues and mouse lungs compared to those from young tissues. Knockdown of Twist1 in aged human ECs increases the levels of PGC1α and angiogenic factor receptor, vascular endothelial growth factor receptor (VEGFR2), and restores EC proliferation and migration, while inhibition of PGC1α suppresses these effects. Knockdown of Twist1 in supplemented aged ECs also restores vascular networks in the subcutaneously implanted gel, while these effects are abrogated by knockdown of PGC1α. Age-dependent inhibition of post-pneumonectomy (PNX) lung growth is suppressed in Tie2-specific Twist1 conditional knockout mouse lungs, in which VEGFR2 expression increases after PNX. These results suggest that upregulation of endothelial Twist1 mediates age-dependent decline in angiogenesis and regenerative lung growth.


Assuntos
Envelhecimento/metabolismo , Pulmão/metabolismo , Neovascularização Fisiológica/fisiologia , Transdução de Sinais/fisiologia , Proteína 1 Relacionada a Twist/metabolismo , Adulto , Fatores Etários , Idoso , Animais , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Receptor TIE-2/genética , Receptor TIE-2/metabolismo , Regeneração/fisiologia , Proteína 1 Relacionada a Twist/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
3.
Yakugaku Zasshi ; 141(3): 335-341, 2021.
Artigo em Japonês | MEDLINE | ID: mdl-33642501

RESUMO

Blood vessels including arteries, veins, and capillaries, are densely spread throughout the body. One round of systemic blood circulation through these blood vessels occurs approximately every minute, and blood sent by the heart transports oxygen, nutrients, and fluid to cells throughout the body. This nourishes cells, tissues, and organs and maintains homeostasis. The relatively simple structure of blood vessels consists of endothelial cells surrounded by a basal lamina and pericytes covering the outer layer. However, blood vessels patterning markedly varies among tissues. The diversity and plasticity of vascular networks are considered vital for this system to facilitate distinct functions for each tissue. Recent studies revealed that blood vessels create a tissue-specific niche, thus attracting attention as biologically active sites for tissue development. This vascular niche establishes specialized microenvironments through both direct physical contact and secreted-soluble factors. Here, we review advances in our understanding of how the vascular niche is utilized by neural stem and progenitor cells during neocortical development, and describe future perspectives regarding new treatment strategies for neural diseases utilizing this vascular niche.


Assuntos
Neocórtex/citologia , Neocórtex/embriologia , Neovascularização Fisiológica/fisiologia , Células-Tronco Neurais/fisiologia , Células-Tronco/fisiologia , Animais , Diferenciação Celular , Humanos , Camundongos , Neocórtex/fisiologia , Nicho de Células-Tronco/fisiologia
4.
J Biosci Bioeng ; 131(6): 686-695, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33775542

RESUMO

Angiogenesis is a pressing issue in tissue engineering associated with restoration of blood supply to ischemic tissues and promotion of rapid vascularization of tissue-engineered grafts. Fibroblast growth factor-2 (FGF-2) plays a vital role in processes such as angiogenesis and is an attractive candidate for tissue engineering. While skeletal muscle tissue engineering is established, the role of FGF-2 in endothelial function to promote angiogenesis after transplantation is unclear. Here, a culture system comprising a five-layered sheet of human skeletal muscle cells co-incubated on green fluorescent protein-expressing human umbilical vein endothelial cells (GFP-HUVECs) mimicking in vivo angiogenesis was used to investigate the role of FGF-2 in vascularization of engineered tissues. The basal level of FGF-2 in cultured media of skeletal muscle cell sheets was undetectable. Therefore, cell sheets co-incubated with GFP-HUVECs were exogenously treated with 10 ng/mL FGF-2, and endothelial network formation was evaluated. After prolonged culture, the endothelial network length and connectivity increased following treatment with FGF-2 as compared with control treatment. The numbers of medium and long endothelial networks significantly increased inside the sheet longer than 0.2 and 0.4 cm, respectively, after FGF-2 treatment. Time-lapse microscopy monitoring dynamic endothelial behavior revealed that FGF-2-mediated maintenance of endothelial connection and retardation of endothelial network disconnection after 72 h. The present study suggests the precise role of FGF-2 in maintaining endothelial connection and the extent of the endothelial network in skeletal muscle cell sheets. This understanding can be applied to design in vitro pre-vascularized tissue and graft integration prospects.


Assuntos
Comunicação Celular/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Músculo Esquelético/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Meios de Cultura/química , Meios de Cultura/farmacologia , Fator 2 de Crescimento de Fibroblastos/metabolismo , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Músculo Esquelético/citologia , Músculo Esquelético/fisiologia , Neovascularização Fisiológica/efeitos dos fármacos , Neovascularização Fisiológica/fisiologia , Organoides/citologia , Organoides/efeitos dos fármacos , Organoides/fisiologia , Engenharia Tecidual/métodos
5.
Int J Mol Sci ; 22(4)2021 Feb 23.
Artigo em Inglês | MEDLINE | ID: mdl-33672103

RESUMO

Hydrogen sulfide (H2S) has long been considered as a toxic gas, but as research progressed, the idea has been updated and it has now been shown to have potent protective effects at reasonable concentrations. H2S is an endogenous gas signaling molecule in mammals and is produced by specific enzymes in different cell types. An increasing number of studies indicate that H2S plays an important role in cardiovascular homeostasis, and in most cases, H2S has been reported to be downregulated in cardiovascular diseases (CVDs). Similarly, in preclinical studies, H2S has been shown to prevent CVDs and improve heart function after heart failure. Recently, many H2S donors have been synthesized and tested in cellular and animal models. Moreover, numerous molecular mechanisms have been proposed to demonstrate the effects of these donors. In this review, we will provide an update on the role of H2S in cardiovascular activities and its involvement in pathological states, with a special focus on the roles of exogenous H2S in cardiac protection.


Assuntos
Fármacos Cardiovasculares/farmacologia , Doenças Cardiovasculares/tratamento farmacológico , Sulfeto de Hidrogênio/administração & dosagem , Sulfeto de Hidrogênio/metabolismo , Administração por Inalação , Animais , Doenças Cardiovasculares/metabolismo , Movimento Celular , Humanos , Neovascularização Fisiológica/fisiologia , Estresse Oxidativo/fisiologia
6.
Life Sci ; 272: 119234, 2021 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-33607158

RESUMO

Stroke still ranks as a most lethal disease worldwide. Angiogenesis during the chronic phase of ischemic stroke can alleviate ischemic injury and attenuate neurological deficit. XQ-1H is a new compound derived from the structure modification of ginkgolide B, which exerts anti-inflammation and neuroprotection against cerebral ischemic injury during the acute or subacute phase. However, whether XQ-1H facilitates angiogenesis and neural functional recovery during the chronic phase remains unclear. This research was designed to explore whether XQ-1H promotes angiogenesis after ischemic stroke and to preliminarily elucidate the mechanism. In vitro, XQ-1H was found to facilitate proliferation, migration and tube formation in bEnd.3 cells. In vivo, XQ-1H raised the CD31 positive microvessel number and increased focal cerebral blood flow in mice exposed to cerebral ischemic injury, and improved the neurological function. Mechanism studies revealed that XQ-1H exerted angiogenesis promoting effect via the PI3K/Akt/GSK3ß/ß-catenin/VEGF signal pathway, which was reversed by LY294002 (the specific inhibitor of PI3K/Akt). In conclusion, XQ-1H exerts angiogenetic effect both in vivo and in vitro, which is a potential agent against ischemic stroke during chronic phase.


Assuntos
Isquemia Encefálica/tratamento farmacológico , Ginkgolídeos/metabolismo , Ginkgolídeos/farmacologia , Lactonas/metabolismo , Lactonas/farmacologia , Animais , Encéfalo/metabolismo , Isquemia Encefálica/metabolismo , Isquemia Encefálica/fisiopatologia , Circulação Cerebrovascular/efeitos dos fármacos , China , Glicogênio Sintase Quinase 3 beta/metabolismo , Infarto da Artéria Cerebral Média/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microvasos/metabolismo , Neovascularização Fisiológica/efeitos dos fármacos , Neovascularização Fisiológica/fisiologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Recuperação de Função Fisiológica/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Acidente Vascular Cerebral/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , beta Catenina/metabolismo
7.
Am J Med Sci ; 361(6): 765-775, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33582157

RESUMO

BACKGROUND: Stem cell therapy is among the novel approaches for the treatment of post-myocardial infarction cardiomyopathy. This study aims to compare the effect of stromal-derived factor 1 α (SDF1α), mesenchymal stem cells (MSCs) in combination with the lentiviral production of vascular endothelial growth factor (VEGF) on infarct area, vascularization and eventually cardiac function in a rat model of myocardial infarction (MI). METHODS: The influence of SDf1α on MSCs survival was investigated. MSCs were transduced via a lentiviral vector containing VEGF. After that, the effect of mesenchymal stem cell transfection of VEGF-A165 and SDf1α preconditioning on cardiac function and scar size was investigated in five groups of MI rat models. The MSC survival, cardiac function, scar size, angiogenesis, and lymphocyte count were assessed 72 hours and 6 weeks after cell transplantation. RESULTS: SDF1α decreased the lactate dehydrogenase release in MSCs significantly. Also, the number of viable cells in the SDF1α-pretreated group was meaningfully more than the control. The left ventricular systolic function significantly enhanced in groups with p240MSC, SDF1αMSC, and VEGF-A165MSC in comparison to the control group. CONCLUSIONS: These findings suggest that SDF1α pretreatment and overexpressing VEGF in MSCs could augment the MSCs' survival in the infarcted myocardium, reduce the scar size, and improve the cardiac systolic function.


Assuntos
Quimiocina CXCL12/administração & dosagem , Células-Tronco Mesenquimais/efeitos dos fármacos , Infarto do Miocárdio/terapia , Neovascularização Fisiológica/efeitos dos fármacos , Animais , Células Cultivadas , Masculino , Células-Tronco Mesenquimais/fisiologia , Infarto do Miocárdio/diagnóstico por imagem , Infarto do Miocárdio/fisiopatologia , Neovascularização Fisiológica/fisiologia , Ratos , Ratos Wistar , Proteínas Recombinantes/administração & dosagem
8.
Nat Neurosci ; 24(4): 478-488, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33510480

RESUMO

Neural-derived signals are crucial regulators of CNS vascularization. However, whether the vasculature responds to these signals by means of elongating and branching or in addition by building a feedback response to modulate neurodevelopmental processes remains unknown. In this study, we identified bidirectional crosstalk between the neural and the vascular compartment of the developing CNS required for oligodendrocyte precursor cell specification. Mechanistically, we show that neural progenitor cells (NPCs) express angiopoietin-1 (Ang1) and that this expression is regulated by Sonic hedgehog. We demonstrate that NPC-derived Ang1 signals to its receptor, Tie2, on endothelial cells to induce the production of transforming growth factor beta 1 (TGFß1). Endothelial-derived TGFß1, in turn, acts as an angiocrine molecule and signals back to NPCs to induce their commitment toward oligodendrocyte precursor cells. This work demonstrates a true bidirectional collaboration between NPCs and the vasculature as a critical regulator of oligodendrogenesis.


Assuntos
Diferenciação Celular/fisiologia , Células Endoteliais/metabolismo , Neovascularização Fisiológica/fisiologia , Neurogênese/fisiologia , Células Precursoras de Oligodendrócitos/citologia , Animais , Camundongos Endogâmicos C57BL , Células-Tronco Neurais/citologia , Células Precursoras de Oligodendrócitos/metabolismo
9.
Nat Biomed Eng ; 5(1): 89-102, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33483713

RESUMO

Using endogenous mesenchymal stem cells for treating myocardial infarction and other cardiovascular conditions typically results in poor efficacy, in part owing to the heterogeneity of the harvested cells and of the patient responses. Here, by means of high-throughput screening of the combinatorial space of mechanical-strain level and of the presence of particular kinase inhibitors, we show that human mesenchymal stem cells can be mechanically and pharmacologically conditioned to enhance vascular regeneration in vivo. Mesenchymal stem cells conditioned to increase the activation of signalling pathways mediated by Smad2/3 (mothers against decapentaplegic homolog 2/3) and YAP (Yes-associated protein) expressed markers that are associated with pericytes and endothelial cells, displayed increased angiogenic activity in vitro, and enhanced the formation of vasculature in mice after subcutaneous implantation and after implantation in ischaemic hindlimbs. These effects were mediated by the crosstalk of endothelial-growth-factor receptors, transforming-growth-factor-beta receptor type 1 and vascular-endothelial-growth-factor receptor 2. Mechanical and pharmacological conditioning can significantly enhance the regenerative properties of mesenchymal stem cells.


Assuntos
Fenômenos Biomecânicos/fisiologia , Células-Tronco Mesenquimais/fisiologia , Neovascularização Fisiológica/fisiologia , Regeneração/fisiologia , Adulto , Animais , Feminino , Humanos , Isquemia , Masculino , Transplante de Células-Tronco Mesenquimais , Camundongos , Neovascularização Fisiológica/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Receptores de Fatores de Crescimento/metabolismo , Regeneração/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Adulto Jovem
10.
Commun Biol ; 4(1): 82, 2021 01 19.
Artigo em Inglês | MEDLINE | ID: mdl-33469143

RESUMO

Bone has a remarkable potential for self-healing and repair, yet several injury types are non-healing even after surgical or non-surgical treatment. Regenerative therapies that induce bone repair or improve the rate of recovery are being intensely investigated. Here, we probed the potential of bone marrow stem cells (BMSCs) engineered with chemically modified mRNAs (modRNA) encoding the hBMP-2 and VEGF-A gene to therapeutically heal bone. Induction of osteogenesis from modRNA-treated BMSCs was confirmed by expression profiles of osteogenic related markers and the presence of mineralization deposits. To test for therapeutic efficacy, a collagen scaffold inoculated with modRNA-treated BMSCs was explored in an in vivo skull defect model. We show that hBMP-2 and VEGF-A modRNAs synergistically drive osteogenic and angiogenic programs resulting in superior healing properties. This study exploits chemically modified mRNAs, together with biomaterials, as a potential approach for the clinical treatment of bone injury and defects.


Assuntos
Proteína Morfogenética Óssea 2/metabolismo , Osso e Ossos/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Materiais Biocompatíveis , Células da Medula Óssea/metabolismo , Regeneração Óssea/fisiologia , Diferenciação Celular , Células Cultivadas , China , Colágeno/metabolismo , Masculino , Células-Tronco Mesenquimais/metabolismo , Neovascularização Fisiológica/fisiologia , Osteogênese/fisiologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Engenharia Tecidual
11.
PLoS Comput Biol ; 17(1): e1008055, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33411727

RESUMO

We introduce a hybrid two-dimensional multiscale model of angiogenesis, the process by which endothelial cells (ECs) migrate from a pre-existing vascular bed in response to local environmental cues and cell-cell interactions, to create a new vascular network. Recent experimental studies have highlighted a central role of cell rearrangements in the formation of angiogenic networks. Our model accounts for this phenomenon via the heterogeneous response of ECs to their microenvironment. These cell rearrangements, in turn, dynamically remodel the local environment. The model reproduces characteristic features of angiogenic sprouting that include branching, chemotactic sensitivity, the brush border effect, and cell mixing. These properties, rather than being hardwired into the model, emerge naturally from the gene expression patterns of individual cells. After calibrating and validating our model against experimental data, we use it to predict how the structure of the vascular network changes as the baseline gene expression levels of the VEGF-Delta-Notch pathway, and the composition of the extracellular environment, vary. In order to investigate the impact of cell rearrangements on the vascular network structure, we introduce the mixing measure, a scalar metric that quantifies cell mixing as the vascular network grows. We calculate the mixing measure for the simulated vascular networks generated by ECs of different lineages (wild type cells and mutant cells with impaired expression of a specific receptor). Our results show that the time evolution of the mixing measure is directly correlated to the generic features of the vascular branching pattern, thus, supporting the hypothesis that cell rearrangements play an essential role in sprouting angiogenesis. Furthermore, we predict that lower cell rearrangement leads to an imbalance between branching and sprout elongation. Since the computation of this statistic requires only individual cell trajectories, it can be computed for networks generated in biological experiments, making it a potential biomarker for pathological angiogenesis.


Assuntos
Células Endoteliais , Modelos Biológicos , Neovascularização Fisiológica/fisiologia , Animais , Diferenciação Celular/fisiologia , Linhagem Celular , Movimento Celular/fisiologia , Células Cultivadas , Células Endoteliais/citologia , Células Endoteliais/fisiologia , Camundongos , Neovascularização Patológica/fisiopatologia , Transdução de Sinais/fisiologia , Transcriptoma/fisiologia
12.
Am J Physiol Cell Physiol ; 320(2): C240-C249, 2021 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-33406025

RESUMO

Chronic kidney disease (CKD) is associated with a substantial increased risk of cardiovascular disease. There is growing evidence that uremic metabolites, which accumulate in the blood with CKD, have detrimental impacts on endothelial cell health and function. However, the molecular mechanisms by which uremic metabolites negatively impact endothelial cell biology are not fully understood. In this study, activation of the aryl hydrocarbon receptor (AHR) via indoxyl sulfate, a known uremic metabolite, was found to impair endothelial cell tube formation and proliferation but not migratory function. Moreover, aortic ring cultures treated with indoxyl sulfate also exhibited decreased sprouting and high AHR activation. Next, genetic knockdown of the AHR using shRNA was found to rescue endothelial cell tube formation, proliferation, and aortic ring sprouting. Similarly, pharmacological AHR antagonism using resveratrol and CH223191 were also found to rescue angiogenesis in cell and aortic ring cultures. Finally, a constitutively active AHR (CAAHR) vector was generated and used to confirm AHR-specific effects. Expression of the CAAHR recapitulated the impaired tube formation and proliferation in cultured endothelial cells and decreased sprouting in aortic ring cultures. Taken together, these data define the impact of AHR activation on angiogenesis and highlight the potential for therapeutic AHR antagonists, which may improve angiogenesis in the context of CKD and cardiovascular disease.


Assuntos
Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Indicã/farmacologia , Neovascularização Fisiológica/efeitos dos fármacos , Neovascularização Fisiológica/fisiologia , Receptores de Hidrocarboneto Arílico/metabolismo , Animais , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Técnicas de Cultura de Órgãos
13.
Transplant Proc ; 53(1): 417-426, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32958221

RESUMO

BACKGROUND: Transplantation of living allogeneic bone segments may permit reconstruction of large defects, particularly if viability is maintained without immunosuppression. Development of a new autogenous osseous blood supply accomplishes this goal in rodent experimental models. This study evaluates potential systemic and local inflammatory responses to this angiogenesis in a large-animal model. METHODS: Vascularized allogeneic tibia segments were transplanted orthotopically into matched tibial defects in Yucatan minipigs. Microvascular anastomoses of bone nutrient artery and vein were supplemented by intramedullary placement of an autogenous arteriovenous (AV) bundle in group 1. Group 2 served as a no-angiogenesis control. A 3-drug immunosuppression regimen was withdrawn after 2 weeks. During the 20-week survival period, periodic leukocyte counts and inflammatory cytokine levels were measured. Thereafter, osteocyte survival was quantified and transplant rejection graded by histologic examination and quantitative real-time polymerase chain reaction of immunologic markers. RESULTS: Both groups developed an initial systemic response, which resolved after 4 to 6 weeks. No differences were seen in blood cytokine levels. Interleukin 2 expression was diminished in group 1 tibiae. As expected, nutrient pedicles had thrombosed without sustained immunosuppression, occluded by intimal hyperplasia. In group 1, angiogenesis from the autogenous AV bundle resulted in significantly less osteonecrosis (P = .04) and fibrosis (P = .02) than group 2 allotransplants. CONCLUSIONS: Systemic immune responses to large-bone allotransplants were not increased by generation of an autogenous osseous blood supply within porcine tibial bone allotransplants. Implanted AV bundles diminished inflammation and fibrosis and improved bone viability when compared to no-angiogenesis controls.


Assuntos
Artérias/transplante , Transplante Ósseo/métodos , Transplante Autólogo/métodos , Transplante Homólogo/métodos , Veias/transplante , Aloenxertos/imunologia , Anastomose Cirúrgica , Animais , Autoenxertos/imunologia , Osso e Ossos/irrigação sanguínea , Rejeição de Enxerto , Neovascularização Fisiológica/fisiologia , Suínos , Porco Miniatura
14.
Methods Mol Biol ; 2206: 67-88, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32754812

RESUMO

Several studies are available addressing the mechanisms of vascular morphogenesis in order to unravel how cooperative cell behavior can follow from the underlying, genetically regulated behavior of endothelial cells and from cell-to-cell and cell-to-extracellular matrix interactions. From the morphological standpoint several aspects of the process are of interest. They include the way the pattern of vessels fills the available tissue space and how the network grows during the angiogenic process, namely how a main trunk divides into smaller branches, and how branching occurs at different distances from the root point of a vascular tree. A third morphological aspect of interest concerns the spatial relationship between vessels and tissue cells able to secrete factors modulating endothelial cells self-organization, thus influencing vascular rearrangement.In the present chapter image analysis methods allowing for a quantitative characterization of these morphological aspects will be detailed and discussed. They are almost based on concepts derived from the theoretical framework represented by spatial statistics.


Assuntos
Células Endoteliais/fisiologia , Processamento de Imagem Assistida por Computador/métodos , Morfogênese/fisiologia , Neovascularização Fisiológica/fisiologia , Animais , Vasos Sanguíneos/fisiologia , Comunicação Celular/fisiologia , Embrião de Galinha , Galinhas , Matriz Extracelular/fisiologia , Microvasos/fisiologia
15.
Methods Mol Biol ; 2206: 103-127, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32754814

RESUMO

For more than 2000 years, the avian embryo has helped scientists understand questions of developmental and cell biology. As early as 350 BC Aristotle described embryonic development inside a chicken egg (Aristotle, Generation of animals. Loeb Classical Library (translated), vol. 8, 1943). In the seventeenth century, Marcello Malpighi, referred to as the father of embryology, first diagramed the microscopic morphogenesis of the chick embryo, including extensive characterization of the cardiovascular system (Pearce Eur Neurol 58(4):253-255, 2007; West, Am J Physiol Lung Cell Mol Physiol 304(6):L383-L390, 2016). The ease of accessibility to the embryo and similarity to mammalian development have made avians a powerful system among model organisms. Currently, a unique combination of classical and modern techniques is employed for investigation of the vascular system in the avian embryo. Here, we will introduce the essential techniques of embryonic manipulation for experimental study in vascular biology.


Assuntos
Galinhas/fisiologia , Neovascularização Fisiológica/fisiologia , Codorniz/fisiologia , Animais , Embrião de Galinha , Embrião de Mamíferos/fisiologia , Desenvolvimento Embrionário/fisiologia , Modelos Animais
16.
Methods Mol Biol ; 2206: 143-150, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32754816

RESUMO

Pericytes are integral part of neurovascular unit and play a role in the maintenance of blood-brain barrier integrity, angiogenesis, and cerebral blood flow regulation. Despite their important functional roles, a univocal phenotypic identification is still emerging also for the lack of a "pan-pericyte" marker. In the present study, we describe in detail the method for performing fluorescence immunohistochemistry on thick free-floating sections from human fetal brain in high resolution laser confocal microscopy. This method enables to obtain three-dimensional images of pericytes and provides insights about their distribution and localization in the microvessels of human developing brain.


Assuntos
Encéfalo/irrigação sanguínea , Microscopia Confocal/métodos , Microvasos/citologia , Pericitos/citologia , Barreira Hematoencefálica/citologia , Circulação Cerebrovascular/fisiologia , Humanos , Imageamento Tridimensional/métodos , Imuno-Histoquímica/métodos , Neovascularização Fisiológica/fisiologia
17.
Methods Mol Biol ; 2206: 193-203, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32754819

RESUMO

The capability of forming functional blood vessel networks is critical for the characterization of endothelial cells. In this chapter, we will review a modified in vivo vascular network forming assay by replacing traditional mouse tumor-derived Matrigel with a well-defined collagen-fibrin hydrogel. The assay is reliable and does not require special equipment, surgical procedure, or a skilled person to perform. Moreover, investigators can modify this method on-demand for testing different cell sources, perturbation of gene functions, growth factors, and pharmaceutical molecules, and for the development and investigation of strategies to enhance neovascularization of engineered human tissues and organs.


Assuntos
Bioensaio/métodos , Vasos Sanguíneos/citologia , Microvasos/citologia , Neovascularização Fisiológica/fisiologia , Animais , Colágeno/metabolismo , Combinação de Medicamentos , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Fibrina/metabolismo , Humanos , Hidrogéis/metabolismo , Laminina/metabolismo , Camundongos , Camundongos Nus , Proteoglicanas/metabolismo , Engenharia Tecidual/métodos
18.
Methods Mol Biol ; 2206: 205-222, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32754820

RESUMO

The zebrafish has emerged as a valuable and important model organism for studying vascular development and vascular biology. Here, we discuss some of the approaches used to study vessels in fish, including loss-of-function tools such as morpholinos and genetic mutants, along with methods and considerations for assessing vascular phenotypes. We also provide detailed protocols for methods used for vital imaging of the zebrafish vasculature, including microangiography and long-term time-lapse imaging. The methods we describe, and the considerations we suggest using for assessing phenotypes observed using these methods, will help ensure reliable, valid conclusions when assessing vascular phenotypes following genetic or experimental manipulation of zebrafish.


Assuntos
Angiografia/métodos , Vasos Sanguíneos/fisiologia , Peixe-Zebra/fisiologia , Animais , Vasos Sanguíneos/metabolismo , Morfolinos/metabolismo , Neovascularização Fisiológica/fisiologia , Fenótipo , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/metabolismo
19.
Spine (Phila Pa 1976) ; 46(15): E802-E809, 2021 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-33337674

RESUMO

STUDY DESIGN: In vivo studies of the vascular system in ossification of the posterior longitudinal ligament (OPLL) model mice. OBJECTIVE: The aim of this study was to investigate blood coagulability, vascular morphology, and vasculogenesis capability, known as venous thromboembolism (VTE) risk factors in the ossification model, tiptoe walking (ttw) mice. SUMMARY OF BACKGROUND DATA: Patients with OPLL are more likely to develop VTE after spinal cord injury. Capillary mesh invasion of spinal ligaments precedes spinal ligament ossification in ttw mice. Investigation on vascular systems of ttw mice may contribute to clarifying its pathology. METHODS: Coagulability of blood samples from ttw and C57BL/6 (WT) mice were evaluated at 8, 16, and 24 weeks of age. Vascular morphology was assessed from a Hematoxylin-Eosin stained section by measuring vessel area. A tube formation assay was performed with endothelial cells isolated from the aorta to assess vasculogenesis. RESULTS: Prothrombin time was significantly shorter in ttw mice than in WT at 8 and 16 weeks. Fibrinogen had a greater increase in ttw mice than in WT at 16 weeks. The vascular area and vascular wall area were significantly smaller in ttw mice than in WT at all timepoints. The ratio of vascular wall area to vascular area was significantly smaller in ttw mice than in WT at 24 weeks. The endothelial cells from ttw mice formed significantly higher numbers of total branching points than WT cells. CONCLUSION: Ossification model mice had impaired blood coagulation and vascular morphology and high capacity for vasculogenesis. With regard to the pathogenesis of VTE, ttw mice harbor an environment that promotes the development of VTE.Level of Evidence: N/A.


Assuntos
Modelos Animais de Doenças , Ossificação do Ligamento Longitudinal Posterior , Animais , Coagulação Sanguínea/fisiologia , Camundongos , Neovascularização Fisiológica/fisiologia
20.
Methods Mol Biol ; 2147: 163-173, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32840819

RESUMO

Functional vasculature is crucial for the maintenance of living tissues via the transport of oxygen, nutrients, and metabolic waste products. As a result, insufficient vascularization in thick engineered tissues will lead to cell death and necrosis due to mass transport and diffusional constraints. To circumvent these limitations, we describe the development of a microscale continuous optical bioprinting (µCOB) platform for 3D printing complex vascularized tissues with superior resolution and speed. By using the µCOB system, endothelial cells and other supportive cells can be printed directly into hydrogels with precisely controlled distribution and subsequent formation of lumen-like structures in vitro.


Assuntos
Materiais Biocompatíveis/síntese química , Bioimpressão/métodos , Vasos Sanguíneos/citologia , Regeneração Tecidual Guiada/instrumentação , Engenharia Tecidual/instrumentação , Tecidos Suporte/química , Animais , Órgãos Bioartificiais , Materiais Biocompatíveis/química , Circulação Sanguínea/fisiologia , Células Cultivadas , Células Endoteliais/citologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Hidrogéis/síntese química , Hidrogéis/química , Camundongos Endogâmicos C3H , Neovascularização Fisiológica/fisiologia , Impressão Tridimensional
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