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1.
Nat Commun ; 11(1): 4102, 2020 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-32796823

RESUMO

Emerging evidence suggests that intestinal stromal cells (IntSCs) play essential roles in maintaining intestinal homeostasis. However, the extent of heterogeneity within the villi stromal compartment and how IntSCs regulate the structure and function of specialized intestinal lymphatic capillary called lacteal remain elusive. Here we show that selective hyperactivation or depletion of YAP/TAZ in PDGFRß+ IntSCs leads to lacteal sprouting or regression with junctional disintegration and impaired dietary fat uptake. Indeed, mechanical or osmotic stress regulates IntSC secretion of VEGF-C mediated by YAP/TAZ. Single-cell RNA sequencing delineated novel subtypes of villi fibroblasts that upregulate Vegfc upon YAP/TAZ activation. These populations of fibroblasts were distributed in proximity to lacteal, suggesting that they constitute a peri-lacteal microenvironment. Our findings demonstrate the heterogeneity of IntSCs and reveal that distinct subsets of villi fibroblasts regulate lacteal integrity through YAP/TAZ-induced VEGF-C secretion, providing new insights into the dynamic regulatory mechanisms behind lymphangiogenesis and lymphatic remodeling.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/metabolismo , Fibroblastos/metabolismo , Mucosa Intestinal/metabolismo , Fatores de Transcrição/metabolismo , Fator C de Crescimento do Endotélio Vascular/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Proteínas de Ciclo Celular/genética , Células Cultivadas , Análise por Conglomerados , Ensaio de Imunoadsorção Enzimática , Fibroblastos/ultraestrutura , Citometria de Fluxo , Imunofluorescência , Hibridização in Situ Fluorescente , Mucosa Intestinal/ultraestrutura , Linfangiogênese/genética , Linfangiogênese/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Eletrônica , Neovascularização Fisiológica/genética , Neovascularização Fisiológica/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição/genética , Fator C de Crescimento do Endotélio Vascular/genética
2.
Cell Prolif ; 53(8): e12830, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32608556

RESUMO

OBJECTIVES: Skin serves as the major interface between the external environment and body which is liable to many kinds of injuries. Mesenchymal stem cell (MSC) therapy has been widely used and became a promising strategy. Pre-treatment with chemical agents, hypoxia or gene modifications can partially protect MSCs against injury, and the pre-treated MSCs show the improved differentiation, homing capacity, survival and paracrine effects regard to attenuating injury. The aim of this study was to investigate whether the exosomes from the educated MSCs contribute to accelerate wound healing process. MATERIALS AND METHODS: We extracted the exosomes from the two educated MSCs and utilized them in the cutaneous wound healing model. The pro-angiogenetic effect of exosomes on endothelial cells was also investigated. RESULTS: We firstly found that MSCs pre-treated by exosomes from neonatal serum significantly improved their biological functions and the effect of therapy. Moreover, we extracted the exosomes from the educated MSCs and utilized them to treat the cutaneous wound model directly. We found that the released exosomes from MSCs which educated by neonatal serum before had the more outstanding performance in therapeutic effect. Mechanistically, we revealed that the recipient endothelial cells (ECs) were targeted and the exosomes promoted their functions to enhance angiogenesis via regulating AKT/eNOS pathway. CONCLUSIONS: Our findings unravelled the positive effect of the upgraded exosomes from the educated MSCs as a promising cell-free therapeutic strategy for cutaneous wound healing.


Assuntos
Exossomos/metabolismo , Células-Tronco Mesenquimais/citologia , Neovascularização Fisiológica/fisiologia , Cicatrização/fisiologia , Animais , Diferenciação Celular/fisiologia , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Fibroblastos/metabolismo , Células Endoteliais da Veia Umbilical Humana/citologia , Humanos , Camundongos Endogâmicos C57BL , Pele/citologia
3.
Life Sci ; 254: 117813, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32428597

RESUMO

AIMS: This study aimed to investigate the effect of lymphocytes in wound healing and the underlying mechanisms, in diabetic and non-diabetic mice, using Balb/c recombination activating gene (Rag)-2 and interleukin 2 receptor gamma (IL-2Rγ) double knockout (KO) (RAG2-/- IL-2Rγ-/-) mice. MAIN METHODS: Wound healing in vivo was performed in control and STZ-induced diabetic mice, in both KO and WT mice. Inflammation and ROS production were evaluated by immunofluorescence microscopy analysis, antioxidant enzymes and angiogenesis were evaluated by quantitative PCR and immunofluorescence microscopy analysis, and wound closure kinetics evolution was evaluated by measurement of acetate tracing of the wound area. KEY FINDINGS: Wound closure was significantly delayed in KO mice, where the M1/M2 macrophage ratio and basal ROS levels were significantly increased, while antioxidant defenses and angiogenesis were significantly decreased. Moreover, the expected increase in matrix metallopeptidase (MMP)-9 protein levels in diabetic conditions was not observed in KO mice, suggesting that the mechanisms leading to the increase in MMP-9 observed in diabetic wounds may in part be lymphocyte-dependent. SIGNIFICANCE: Our results indicate that lack of lymphocytes compromises wound healing independent of diabetes. The lack of these cells, even in non-diabetic mice, mimics the phenotype observed in wounds under diabetic conditions. Moreover, the combination of diabetes and the lack of lymphocytes, further impair the wound healing conditions, indicating that when the innate regulatory function is lost in these KO mice, excessive M1 polarization, poor angiogenesis and impaired wound healing are worsen.


Assuntos
Proteínas de Ligação a DNA/fisiologia , Diabetes Mellitus Experimental/fisiopatologia , Subunidade gama Comum de Receptores de Interleucina/fisiologia , Linfócitos/fisiologia , Neovascularização Fisiológica/fisiologia , Cicatrização/fisiologia , Animais , Proteínas de Ligação a DNA/genética , Inflamação/metabolismo , Subunidade gama Comum de Receptores de Interleucina/genética , Ativação de Macrófagos/fisiologia , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Knockout , Espécies Reativas de Oxigênio/metabolismo , Pele/irrigação sanguínea , Pele/metabolismo
4.
Am J Pathol ; 190(8): 1763-1773, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32450152

RESUMO

Within the human lung, mast cells typically reside adjacent to the conducting airway and assume a mucosal phenotype (MCT). In rare pathologic conditions, connective tissue phenotype mast cells (MCTCs) can be found in the lung parenchyma. MCTCs accumulate in the lungs of infants with severe bronchopulmonary dysplasia, a chronic lung disease associated with preterm birth, which is characterized by pulmonary vascular dysmorphia. The human mast cell line (LUVA) was used to model MCTCs or MCTs. The ability of MCTCs to affect vascular organization during fetal lung development was tested in mouse lung explant cultures. The effect of MCTCs on in vitro tube formation and barrier function was studied using primary fetal human pulmonary microvascular endothelial cells. The mechanistic role of MCTC proteases was tested using inhibitors. MCTCLUVA but not MCTLUVA was associated with vascular dysmorphia in lung explants. In vitro, the addition of MCTCLUVA potentiated fetal human pulmonary microvascular endothelial cell interactions, inhibited tube stability, and disrupted endothelial cell junctions. Protease inhibitors ameliorated the ability of MCTCLUVA to alter endothelial cell angiogenic activities in vitro and ex vivo. These data indicate that MCTCs may directly contribute to disrupted angiogenesis in bronchopulmonary dysplasia. A better understanding of factors that regulate mast cell subtype and their different effector functions is essential.


Assuntos
Displasia Broncopulmonar/patologia , Células Endoteliais/patologia , Pulmão/patologia , Mastócitos/patologia , Neovascularização Fisiológica/fisiologia , Animais , Linhagem Celular , Células Cultivadas , Humanos , Camundongos
5.
Life Sci ; 255: 117763, 2020 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-32389831

RESUMO

AIMS: To explored the potential of human umbilical cord mesenchymal stem cells (hUCMSCs) as seed cells for dental pulp regeneration and the possibility of cotransplantation hUCMSCs and endothelial cells (ECs) for angiogenesis and pulp regeneration in vivo. MATERIALS AND METHODS: hUCMSCs and human umbilical vein endothelial cells (HUVECs) were cocultured for matrigel angiogenesis assay in vitro and Matrigel plug assay in vivo. Next, we used the transwell coculture system to coculture hUCMSCs and HUVECs in vitro for RNA- sequencing (RNA-seq). Last, encapsulated hUCMSCs and HUVECs in scaffolds were injected into the root segments, and transplanted into immunodeficient mice for dental pulp regeneration. KEY FINDINGS: In vitro Matrigel angiogenesis assay and in vivo Matrigel plug assay indicated that cocultured hUCMSCs and HUVECs promote vascular formation of HUVECs, especially in 1:5 (hUCMSCs:HUVECs) coculture group. The RNA-seq result indicated that cocultured HUVECs exhibited high Hif-1 signaling pathway activity. We performed the cell transfection assay to knock down HIF1A-AS2 in HUVECs and then coculture with hUCMSCs, and the expression of VEGFA, HIF1A and PECAM1 were reduced. In pulp regeneration assay, Cotransplantation of hUCMSCs and HUVECs (1,5) group showed pulp-like tissue regeneration. SIGNIFICANCE: Cocultured hUCMSCs and HUVECs can promote vascular formation of HUVECs, and the optimal coculture ration is 1:5 (hUCMSCs:HUVECs). hUCMSCs promote angiogenesis of HUVECs through the long noncoding RNA HIF1A-AS2-activation of the Hif-1 signaling pathway. Cotransplantation of hUCMSCs and HUVECs can regenerate dental pulp-like tissue in vivo.


Assuntos
Polpa Dentária/metabolismo , Células Endoteliais da Veia Umbilical Humana/transplante , Transplante de Células-Tronco Mesenquimais/métodos , Neovascularização Fisiológica/fisiologia , Animais , Técnicas de Cocultura , Colágeno/metabolismo , Polpa Dentária/citologia , Combinação de Medicamentos , Feminino , Técnicas de Silenciamento de Genes , Humanos , Fator 1 Induzível por Hipóxia/metabolismo , Laminina/metabolismo , Camundongos , Proteoglicanas/metabolismo , RNA Longo não Codificante/genética , Regeneração/fisiologia , Cordão Umbilical/citologia
7.
Yakugaku Zasshi ; 140(4): 513-519, 2020.
Artigo em Japonês | MEDLINE | ID: mdl-32238634

RESUMO

Repair of injured tissues requires angiogenesis, the growth of new blood vessels from pre-existing ones. Cutaneous wound healing is a complex and dynamic process by which skin tissue repairs itself after injury; however, how endothelial cells and pericytes form new blood vessels during cutaneous wound angiogenesis remains unclear. We recently developed a fluorescence-based live imaging system to analyze cutaneous wound angiogenesis in adult zebrafish. Employing this system, we found that endothelial cells and pericytes remain in a quiescent state in normal skin tissue, whereas cutaneous injury immediately activates both types of cells to induce angiogenesis. At 2 days post-injury (dpi), the injured vessels elongated, and some uninjured vessels became tortuous and began to sprout new branches. Then, vessel sprouting, elongation, bifurcation, and anastomosis progressively occurred to form the tortuous and disorganized vascular networks observed at 6 dpi. Thereafter, blood vessel tortuosity gradually decreased through the regression of excessive vessels, thereby leading to the formation of well-organized vessel networks at 42 dpi. Pericytes are thought to detach from the vessel wall to promote endothelial cell sprouting upon the induction of angiogenesis. However, not only endothelial cells but also pericytes proliferated to form pericyte-covered tortuous blood vessels in response to cutaneous injury, revealing an unexpected role of pericytes in cutaneous wound angiogenesis. Therefore, this live-imaging system for adult zebrafish is anticipated to make a valuable contribution to research advancements in understanding the angiogenesis that occurs during tissue repair.


Assuntos
Neovascularização Fisiológica/fisiologia , Imagem Óptica/métodos , Fenômenos Fisiológicos da Pele , Cicatrização/fisiologia , Animais , Células Endoteliais/fisiologia , Humanos , Modelos Animais , Pericitos/fisiologia , Peixe-Zebra
8.
Endocrinology ; 161(4)2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32141513

RESUMO

Follicle-stimulating hormone (FSH)-induced growth of ovarian follicles is independent of follicular vascularization. Recent evidence has indicated that follicular vascularization is critical to ovarian follicle development and survival. FSH, a gonadotropin that induces follicular growth and development, also acts as the major survival factor for antral follicles. FSH has been reported to stimulate angiogenesis in the theca layers mediated in part by the vascular endothelial growth factor A (VEGFA) and the transcription factor hypoxia inducible factor 1α (HIF-1α). However, it remains largely undetermined whether FSH-dependent growth and survival of antral follicles relies on FSH-induced vascularization. Here, we first demonstrated that induction of angiogenesis through the FSH-HIF-1α-VEGFA axis is not required for FSH-stimulated follicular growth in mouse ovary. FSH increased the total number of blood vessels in mouse ovarian follicles, which was correlated with elevated expression of VEGFA and HIF-1α in granulosa cells. In contrast, blocking of follicular angiogenesis using inhibitors against the HIF-1α-VEGFA pathway repressed vasculature formation in follicles despite FSH administration. Interestingly, by measuring follicular size and ovarian weight, we found that the suppression of angiogenesis via HIF-1α-VEGFA pathway did not influence FSH-mediated follicular growth. However, inhibition of FSH-induced follicular vascularization by PX-478, a small-molecule inhibitor that suppresses HIF-1α activity, blocked ovulation and triggered atresia in large follicles. On the other hand, PX-478 injection reduced oocyte quality via impairing the meiotic apparatus, showing a prominently defective spindle assembly and actin dynamics. Collectively, our findings unveiled a vascularization-independent effect of FSH on follicular growth, whereas follicular survival, ovulation, and oocyte development relies on FSH-mediated angiogenesis in the follicles.


Assuntos
Hormônio Foliculoestimulante/farmacologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Oócitos/crescimento & desenvolvimento , Folículo Ovariano/crescimento & desenvolvimento , Ovulação/metabolismo , Transdução de Sinais/fisiologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Feminino , Camundongos , Neovascularização Fisiológica/efeitos dos fármacos , Neovascularização Fisiológica/fisiologia , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Ovulação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
9.
Ann N Y Acad Sci ; 1470(1): 31-43, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32166759

RESUMO

Loss of vascular elasticity results from progressive degeneration of the extracellular matrix of elastic arteries under the effect of aging and certain diseases, including atherosclerosis. To investigate the influence of vessel wall stiffening on endothelial cell (EC) function, we seeded human umbilical vein ECs onto variably compliant polydimethylsiloxane substrates. When plated on the more compliant substrate, ECs assembled into capillary-like structures. By contrast, they failed to form a network on stiff substrates, even in the presence of vascular endothelial growth factor (VEGF). Cell proliferation and migration increased with stiffness, while ECs released more nitric oxide (NO) on the soft substrate. Treatment with VEGF increased migration and NO release in a stiffness-dependent manner. Atomic force microscopy measurement of cell elasticity along with actin fiber analysis revealed that ECs plated on the more compliant surface were mechanically softer, with mostly diffuse actin arrangement. Our results demonstrate that matrix stiffening induces actin reorganizations, reflected by cortical stiffening in ECs, which may lead to a decrease in their angiogenic capacity and NO release. Hence, the mechanical properties of ECs display a prognostic and therapeutic potential and might serve as a reliable biomarker of vascular function.


Assuntos
Células Endoteliais da Veia Umbilical Humana/metabolismo , Neovascularização Fisiológica/fisiologia , Óxido Nítrico/metabolismo , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Neovascularização Fisiológica/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/farmacologia
10.
Nat Commun ; 11(1): 615, 2020 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-32001693

RESUMO

Angiogenesis induction into damaged sites has long been an unresolved issue. Local treatment with pro-angiogenic molecules has been the most common approach. However, this approach has critical side effects including inflammatory coupling, tumorous vascular activation, and off-target circulation. Here, the concept that a structure can guide desirable biological function is applied to physically engineer three-dimensional channel networks in implant sites, without any therapeutic treatment. Microchannel networks are generated in a gelatin hydrogel to overcome the diffusion limit of nutrients and oxygen three-dimensionally. Hydrogel implantation in mouse and porcine models of hindlimb ischemia rescues severely damaged tissues by the ingrowth of neighboring host vessels with microchannel perfusion. This effect is guided by microchannel size-specific regenerative macrophage polarization with the consequent functional recovery of endothelial cells. Multiple-site implantation reveals hypoxia and neighboring vessels as major causative factors of the beneficial function. This technique may contribute to the development of therapeutics for hypoxia/inflammatory-related diseases.


Assuntos
Indutores da Angiogênese/efeitos adversos , Gelatina/química , Gelatina/farmacologia , Hidrogéis/química , Hidrogéis/farmacologia , Isquemia/terapia , Animais , Modelos Animais de Doenças , Células Endoteliais/patologia , Desenho de Equipamento , Feminino , Membro Posterior/irrigação sanguínea , Membro Posterior/diagnóstico por imagem , Membro Posterior/patologia , Hidrogéis/uso terapêutico , Hipóxia , Isquemia/diagnóstico por imagem , Isquemia/patologia , Macrófagos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Neovascularização Fisiológica/fisiologia , Doenças Vasculares Periféricas/patologia , Doenças Vasculares Periféricas/terapia , Próteses e Implantes , Suínos , Cicatrização
11.
Proc Natl Acad Sci U S A ; 117(9): 4792-4801, 2020 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-32075915

RESUMO

Angiogenesis, the formation of new blood vessels, is tightly regulated by gene transcriptional programs. Yin Ying 1 (YY1) is a ubiquitously distributed transcription factor with diverse and complex biological functions; however, little is known about the cell-type-specific role of YY1 in vascular development and angiogenesis. Here we report that endothelial cell (EC)-specific YY1 deletion in mice led to embryonic lethality as a result of abnormal angiogenesis and vascular defects. Tamoxifen-inducible EC-specific YY1 knockout (YY1 iΔEC ) mice exhibited a scarcity of retinal sprouting angiogenesis with fewer endothelial tip cells. YY1 iΔEC mice also displayed severe impairment of retinal vessel maturation. In an ex vivo mouse aortic ring assay and a human EC culture system, YY1 depletion impaired endothelial sprouting and migration. Mechanistically, YY1 functions as a repressor protein of Notch signaling that controls EC tip-stalk fate determination. YY1 deficiency enhanced Notch-dependent gene expression and reduced tip cell formation. Specifically, YY1 bound to the N-terminal domain of RBPJ (recombination signal binding protein for Ig Kappa J region) and competed with the Notch coactivator MAML1 (mastermind-like protein 1) for binding to RBPJ, thereby impairing the NICD (intracellular domain of the Notch protein)/MAML1/RBPJ complex formation. Our study reveals an essential role of endothelial YY1 in controlling sprouting angiogenesis through directly interacting with RBPJ and forming a YY1-RBPJ nuclear repression complex.


Assuntos
Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/metabolismo , Morfogênese/fisiologia , Neovascularização Patológica/metabolismo , Fator de Transcrição YY1/metabolismo , Animais , Proteínas de Transporte/metabolismo , Diferenciação Celular , Células Endoteliais/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Masculino , Camundongos/embriologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neovascularização Patológica/genética , Neovascularização Patológica/patologia , Neovascularização Fisiológica/genética , Neovascularização Fisiológica/fisiologia , Proteínas Nucleares , Ligação Proteica , Receptores Notch/metabolismo , Vasos Retinianos/metabolismo , Transdução de Sinais , Fatores de Transcrição , Fator de Transcrição YY1/genética
12.
Stroke ; 51(4): 1279-1289, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32075549

RESUMO

Background and Purpose- Bone marrow mononuclear cells (BM-MNCs) are a rich source of hematopoietic stem cells and have been widely used in experimental therapies for patients with ischemic diseases. Activation of angiogenesis is believed to be one of major BM-MNC mode of actions, but the essential mechanism by which BM-MNCs activate angiogenesis have hitherto been elusive. The objective of this study is to reveal the mechanism how BM-MNCs activate angiogenesis. Methods- We have evaluated the effect of direct cell-cell interaction between BM-MNC and endothelial cell on uptake of VEGF (vascular endothelial growth factor) into endothelial cells in vitro. Cerebral ischemia model was used to evaluate the effects of direct cell-cell interaction with transplanted BM-MNC on endothelial cell at ischemic tissue. Results- The uptake of VEGF into endothelial cells was increased by BM-MNC, while being inhibited by blockading the gap junction. Low-molecular-weight substance was transferred from BM-MNC into endothelial cells via gap junctions in vivo, followed by increased expression of hypoxia-inducible factor-1α and suppression of autophagy in endothelial cells. The concentration of glucose in BM-MNC cytoplasm was significantly higher than in endothelial cells, and transfer of glucose homologue from BM-MNC to endothelial cells was observed. Conclusions- Our findings demonstrated cell-cell interaction via gap junction is the prominent pathway for activation of angiogenesis at endothelial cells after ischemia and provided novel paradigm that energy source supply by stem cell to injured cell is one of the therapeutic mechanisms of cell-based therapy. Visual Overview- An online visual overview is available for this article.


Assuntos
Transplante de Medula Óssea/métodos , Comunicação Celular/fisiologia , Junções Comunicantes/fisiologia , Neovascularização Fisiológica/fisiologia , Acidente Vascular Cerebral/terapia , Animais , Células da Medula Óssea/fisiologia , Células Endoteliais da Veia Umbilical Humana/transplante , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Acidente Vascular Cerebral/patologia
13.
PLoS Comput Biol ; 16(1): e1006919, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31986145

RESUMO

During angiogenesis, new blood vessels sprout and grow from existing ones. This process plays a crucial role in organ development and repair, in wound healing and in numerous pathological processes such as cancer progression or diabetes. Here, we present a mathematical model of early stage angiogenesis that permits exploration of the relative importance of mechanical, chemical and cellular cues. Endothelial cells proliferate and move over an extracellular matrix by following external gradients of Vessel Endothelial Growth Factor, adhesion and stiffness, which are incorporated to a Cellular Potts model with a finite element description of elasticity. The dynamics of Notch signaling involving Delta-4 and Jagged-1 ligands determines tip cell selection and vessel branching. Through their production rates, competing Jagged-Notch and Delta-Notch dynamics determine the influence of lateral inhibition and lateral induction on the selection of cellular phenotypes, branching of blood vessels, anastomosis (fusion of blood vessels) and angiogenesis velocity. Anastomosis may be favored or impeded depending on the mechanical configuration of strain vectors in the ECM near tip cells. Numerical simulations demonstrate that increasing Jagged production results in pathological vasculatures with thinner and more abundant vessels, which can be compensated by augmenting the production of Delta ligands.


Assuntos
Modelos Biológicos , Neovascularização Patológica/fisiopatologia , Receptores Notch/metabolismo , Transdução de Sinais/fisiologia , Resposta Táctica/fisiologia , Algoritmos , Animais , Biologia Computacional , Simulação por Computador , Neovascularização Fisiológica/fisiologia
14.
Plast Reconstr Surg ; 145(2): 420-431, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31985635

RESUMO

BACKGROUND: Secondary lymphedema is a refractory disease, for which adipose-derived stem cells have shown some therapeutic potential. However, the mechanism of this action remains poorly understood. METHODS: The authors identified podoplanin-expressing adipose-derived stem cells, which allowed them to divide adipose-derived stem cells into podoplanin-positive and podoplanin-negative groups that they characterized in vitro. The authors then used a mouse hindlimb model for lymphedema to trace the fate of podoplanin-positive, podoplanin-negative, and unsorted adipose-derived stem cells in vivo. RESULTS: When induced in culture, podoplanin-positive cells were noted to up-regulate the expression of lymphatic endothelial cell markers, including LYVE-1, and assumed a cobblestone morphology. In addition, a substantial increase in lymphangiogenic cytokines was detected in the podoplanin-positive supernatant. The above findings were largely absent from the podoplanin-negative and unsorted groups. In the mouse model, the implanted cells relieved the limb lymphedema by promoting lymphangiogenesis, with the podoplanin-positive group showing the most significant effect. Immunocolocalization further revealed that the podoplanin-positive cells incorporated into lymphatic vessels were positive for LYVE-1. CONCLUSIONS: These data demonstrated that actions by means of both paracrine and differentiation pathways were involved in the adipose-derived stem cell-mediated therapeutic effects. The podoplanin-positive cells possessed lymphatic paracrine and differentiation abilities and may represent lymphatic endothelial cell precursor cells. The podoplanin-negative cells, which constitute a considerable proportion of the adipose-derived stem cells, may play an important paracrine role by secreting mesenchymal stem cell-related factors.


Assuntos
Linfangiogênese/fisiologia , Vasos Linfáticos/fisiologia , Glicoproteínas de Membrana/metabolismo , Células-Tronco Mesenquimais/fisiologia , Animais , Biomarcadores/metabolismo , Células Cultivadas , Citocinas/metabolismo , Modelos Animais de Doenças , Células Endoteliais/fisiologia , Feminino , Proteínas de Fluorescência Verde , Linfedema/fisiopatologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neovascularização Fisiológica/fisiologia , Fenótipo
15.
Nat Commun ; 11(1): 460, 2020 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-31974363

RESUMO

Recent interest in the control of bone metabolism has focused on a specialized subset of CD31hiendomucinhi vessels, which are reported to couple angiogenesis with osteogenesis. However, the underlying mechanisms that link these processes together remain largely undefined. Here we show that the zinc-finger transcription factor ZEB1 is predominantly expressed in CD31hiendomucinhi endothelium in human and mouse bone. Endothelial cell-specific deletion of ZEB1 in mice impairs CD31hiendomucinhi vessel formation in the bone, resulting in reduced osteogenesis. Mechanistically, ZEB1 deletion reduces histone acetylation on Dll4 and Notch1 promoters, thereby epigenetically suppressing Notch signaling, a critical pathway that controls bone angiogenesis and osteogenesis. ZEB1 expression in skeletal endothelium declines in osteoporotic mice and humans. Administration of Zeb1-packaged liposomes in osteoporotic mice restores impaired Notch activity in skeletal endothelium, thereby promoting angiogenesis-dependent osteogenesis and ameliorating bone loss. Pharmacological reversal of the low ZEB1/Notch signaling may exert therapeutic benefit in osteoporotic patients by promoting angiogenesis-dependent bone formation.


Assuntos
Neovascularização Fisiológica/fisiologia , Osteogênese/fisiologia , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/farmacologia , Idoso , Animais , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/farmacologia , Células Endoteliais/metabolismo , Epigênese Genética , Feminino , Humanos , Camundongos Knockout , Camundongos Transgênicos , Pessoa de Meia-Idade , Neovascularização Fisiológica/genética , Osteogênese/efeitos dos fármacos , Osteogênese/genética , Osteoporose/terapia , Ovariectomia , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Receptor Notch1/genética , Receptor Notch1/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética
16.
Cell Prolif ; 53(2): e12757, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31916327

RESUMO

OBJECTIVES: To testify that endothelial cells (ECs) induce astrocyte maturation by leukaemia inhibitory factor (LIF) secretion. MATERIALS AND METHODS: In vivo experiments, mice bearing floxed alleles of YAP were crossed with mice expressing a Cre recombinase driven by the endothelial Tek promoter (Tek-Cre) to finally obtain the following three genotypes: YAPf/f , Tek-Cre; YAPf/w , Tek-Cre; and YAPf/f . Retinal vascularization and astrocyte network were evaluated by whole-mount fluorescence and Western blotting. In vitro, experiments were performed in an astrocyte and human microvascular endothelial cell (HMEC-1) coculture model to analyse the mechanisms underlying the effect of endothelial YAP on astrocytes. RESULTS: In vivo, YAPf/f ;Tek-Cre mice showed delayed angiogenesis, sparse vessels and decreased glial fibrillary acidic protein (GFAP)+ astrocytes but aberrant growth of endothelial networks and immature astrocytes (platelet-derived growth factor A, PDGFRA+ astrocytes) overgrowth. In vitro, Yap deletion attenuated the LIF release that delayed the maturation of retinal astrocyte which was consistent with the results of HMEC-1-astrocyte coculture. The effect of YAP overexpression on LIF-LIFR axis in HMEC-1 interferes the GFAP expression of astrocyte. In contrast, LIF protein rescues the astrocytic GFAP expression when EC YAP was inhibited by siRNAs. CONCLUSIONS: We show that EC yes-associated protein (YAP) is not only a critical coactivator of Hippo signalling in retinal vessel development but also plays an essential role in retinal astrocyte maturation by regulating LIF production.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Astrócitos/metabolismo , Fator Inibidor de Leucemia/metabolismo , Retina/metabolismo , Vasos Retinianos/metabolismo , Fatores de Transcrição/metabolismo , Animais , Astrócitos/fisiologia , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Técnicas de Cocultura/métodos , Células Endoteliais/metabolismo , Células Endoteliais/fisiologia , Feminino , Proteína Glial Fibrilar Ácida/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Neovascularização Fisiológica/fisiologia , Neurogênese/fisiologia , Retina/fisiologia , Vasos Retinianos/fisiologia
17.
PLoS One ; 15(1): e0227286, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31910228

RESUMO

In order to unravel rapid mechano-chemical feedback mechanisms in sprouting angiogenesis, we combine selective plane illumination microscopy (SPIM) and tailored image registration algorithms - further referred to as SPIM-based displacement microscopy - with an in vitro model of angiogenesis. SPIM successfully tackles the problem of imaging large volumes while upholding the spatial resolution required for the analysis of matrix displacements at a subcellular level. Applied to in vitro angiogenic sprouts, this unique methodological combination relates subcellular activity - minute to second time scale growing and retracting of protrusions - of a multicellular systems to the surrounding matrix deformations with an exceptional temporal resolution of 1 minute for a stack with multiple sprouts simultaneously or every 4 seconds for a single sprout, which is 20 times faster than with a conventional confocal setup. Our study reveals collective but non-synchronised, non-continuous activity of adjacent sprouting cells along with correlations between matrix deformations and protrusion dynamics.


Assuntos
Imageamento Tridimensional/métodos , Microscopia Intravital/métodos , Neovascularização Fisiológica/fisiologia , Imagem com Lapso de Tempo , Algoritmos , Técnicas de Cultura de Células/métodos , Colágeno Tipo I , Marcadores Fiduciais , Células Endoteliais da Veia Umbilical Humana , Humanos , Hidrogéis , Microscopia de Fluorescência/métodos , Microesferas
18.
Plast Reconstr Surg ; 145(2): 348e-359e, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31985636

RESUMO

BACKGROUND: The authors developed a noncontact low-frequency ultrasound device that delivers high-intensity mechanical force based on phased-array technology. It may aid wound healing because it is likely to be associated with lower risks of infection and heat-induced pain compared with conventional ultrasound methods. The authors hypothesized that the microdeformation it induces accelerates wound epithelialization. Its effects on key wound-healing processes (angiogenesis, collagen accumulation, and angiogenesis-related gene transcription) were also examined. METHODS: Immediately after wounding, bilateral acute wounds in C57BL/6J mice were noncontact low-frequency ultrasound- and sham-stimulated for 1 hour/day for 3 consecutive days (10 Hz/90.6 Pa). Wound closure (epithelialization) was recorded every 2 days as the percentage change in wound area relative to baseline. Wound tissue was procured on days 2, 5, 7, and 14 (five to six per time point) and subjected to histopathology with hematoxylin and eosin and Masson trichrome staining, CD31 immunohistochemistry, and quantitative polymerase-chain reaction analysis. RESULTS: Compared to sham-treated wounds, ultrasound/phased-array-treated wounds exhibited significantly accelerated epithelialization (65 ± 27 percent versus 30 ± 33 percent closure), angiogenesis (4.6 ± 1.7 percent versus 2.2 ± 1.0 percent CD31 area), and collagen deposition (44 ± 14 percent versus 28 ± 13 percent collagen density) on days 5, 2, and 5, respectively (all p < 0.05). The expression of Notch ligand delta-like 1 protein (Dll1) and Notch1, which participate in angiogenesis, was transiently enhanced by treatment on days 2 and 5, respectively. CONCLUSIONS: The authors' noncontact low-frequency ultrasound phased-array device improved the wound-healing rate. It was associated with increased early neovascularization that was followed by high levels of collagen-matrix production and epithelialization. The device may expand the mechanotherapeutic proangiogenesis field, thereby helping stimulate a revolution in infected wound care.


Assuntos
Pele/lesões , Terapia por Ultrassom/métodos , Cicatrização/fisiologia , Ferimentos e Lesões/terapia , Animais , Colágeno/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neovascularização Fisiológica/fisiologia , Pele/metabolismo , Ferimentos e Lesões/metabolismo , Ferimentos e Lesões/patologia
19.
J Surg Res ; 249: 168-179, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31986359

RESUMO

BACKGROUND: Development of collateral vasculature is key in compensating for arterial occlusions in patients with peripheral artery disease (PAD). We aimed to examine the development of collateral pathways after ligation of native vessels in a porcine model of PAD. METHODS: Right hindlimb ischemia was induced in domestic swine (n = 11) using two versions of arterial ligation. Version 1 (n = 6) consisted of ligation with division of the right external iliac, profunda femoral, and superficial femoral arteries. Version 2 (n = 5) consisted of the ligation of version 1 with additional ligation with division of the right internal iliac artery. Development of collateral pathways was evaluated with standard angiography before arterial ligation and at termination (30 days later). Relative luminal diameter of the arteries supplying the ischemic right hind limb were determined by two-dimensional angiography. RESULTS: The dominant collateral pathway that developed after version 1 ligation connected the right internal iliac artery to the right profunda femoral and then to the right superficial femoral and popliteal artery. Mean luminal diameter of the right internal iliac artery at termination increased by 38% compared with baseline. Two codominant collateral pathways developed in version 2 ligation: (i) from the left profunda femoral artery to the reconstituted right profunda femoral artery and (ii) from the common internal iliac trunk and the left internal iliac artery to the reconstituted right internal iliac artery, which then supplied the right profunda femoral and then the right superficial femoral and popliteal artery. The mean diameter of the left profunda and the left internal iliac artery increased at termination by 26% and 21%, respectively (P < 0.05). CONCLUSIONS: Two versions of hindlimb ischemia induction (right ilio-femoral artery ligation with and without right internal iliac artery ligation) in swine produced differing collateral pathways, along with changes to the diameter of the inflow vessels (i.e., arteriogenesis).


Assuntos
Circulação Colateral/fisiologia , Isquemia/fisiopatologia , Neovascularização Fisiológica/fisiologia , Doença Arterial Periférica/fisiopatologia , Angiografia , Animais , Modelos Animais de Doenças , Artéria Femoral/diagnóstico por imagem , Artéria Femoral/cirurgia , Membro Posterior/irrigação sanguínea , Humanos , Artéria Ilíaca/diagnóstico por imagem , Artéria Ilíaca/cirurgia , Isquemia/diagnóstico por imagem , Isquemia/etiologia , Ligadura/efeitos adversos , Masculino , Doença Arterial Periférica/diagnóstico por imagem , Doença Arterial Periférica/etiologia , Fluxo Sanguíneo Regional/fisiologia , Sus scrofa
20.
J Vasc Res ; 57(2): 76-85, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31968349

RESUMO

Diabetes mellitus (DM)-induced impairment of collateral formation has been demonstrated in subjects with coronary artery disease, which contributes to unfavorable prognosis among diabetic individuals. In our previous studies, thioredoxin1 (Trx1) activity was shown to be decreased in diabetic cardiac tissues, but the reason of Trx1 inactivation and whether it mediates the impaired angiogenesis in ischemic myocardium is still to be identified. As thioredoxin-interacting protein (TXNIP), an endogenous inhibitor of Trx, is overexpressed in DM due to carbohydrate response element within its promoter, we hypothesized that inhibition of Trx1 by enhanced TXNIP expression in endothelial cells may play a role in hyperglycemia-induced impairment of angiogenesis. In the present study, we found that high glucose-mediated increase of TXNIP expression and TXNIP-Trx1 interaction induced the impairment in endothelial cell function and survival, since these detrimental effects are rescued by silencing TXNIP with small interfering RNA. In diabetic mice, TXNIP knockdown or recombinant human Trx1 treatment counteracted the impairment of angiogenesis, alleviated myocardial ischemic injury, and improved survival rate. All these data implicate that TXNIP upregulation and subsequently the increased formation of TXNIP-Trx1 complex is a novel pathologic pathway by which DM induces insufficient angiogenesis and thereby exacerbates myocardial ischemia injury.


Assuntos
Diabetes Mellitus Experimental/complicações , Isquemia Miocárdica/fisiopatologia , Neovascularização Fisiológica/fisiologia , Tiorredoxinas/fisiologia , Animais , Proteínas de Transporte/fisiologia , Diabetes Mellitus Experimental/fisiopatologia , Células Endoteliais/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL
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