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1.
Anticancer Res ; 39(7): 3543-3551, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31262878

RESUMO

BACKGROUND/AIM: Both bevacizumab (BEV) and soluble fms-like tyrosine kinase-1 (sFlt-1) have demonstrated anti-angiogenic effects, thereby causing hypertension and proteinuria. We hypothesized that anti-preeclamptic drugs that combat the action of sFlt-1 may reduce BEV's anti-tumor efficacy. MATERIALS AND METHODS: 3D co-cultured human mini-tumors consisting of endothelial cells, fibroblasts, and cancer cells were developed. The influence of anti-preeclamptic drugs and BEV on the invasion of mini-tumors embedded in collagen gel was evaluated. RESULTS: Mini-tumor spheroids that contained MDA-MB-231 cells showed higher invasion ability than spheroids with A549. Among the six anti-preeclamptic drugs investigated, only nicorandil enhanced the invasion of mini-tumors and inhibited the action of BEV. Glibenclamide, an ATP-sensitive potassium channel inhibitor, completely quenched the action of nicorandil on mini-tumors. CONCLUSION: In the human mini-tumor model, nicorandil aggravated the invasion of mini-tumors. These data raise the possibility that concomitant use of nicorandil counteracts the efficacy of BEV therapy.


Assuntos
Inibidores da Angiogênese/farmacologia , Anti-Hipertensivos/farmacologia , Bevacizumab/farmacologia , Esferoides Celulares/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Técnicas de Cocultura , Feminino , Fibroblastos/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Nicorandil/farmacologia , Pré-Eclâmpsia , Gravidez , Esferoides Celulares/fisiologia , Fator A de Crescimento do Endotélio Vascular/farmacologia
2.
Int J Nanomedicine ; 14: 2253-2263, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30992665

RESUMO

Background: Treatment of wounds with the help of nanoparticles (NPs) is more effective and superior in comparison to traditional wound healing methods as it protects and sustains active drug release at the wound site thus enhancing the safety of the drug and reducing the possibility of side effects. The advantages of this method are the possibility of allowing a reduction in administered dose, limiting toxicity levels to the minimum, and increasing safety of topical delivery of the drug. Materials and methods: We report the synthesis of a novel poly (lactic-co-glycolic acid) (PLGA) NP-based multicargo delivery system for growth factors and antimicrobial peptide. Growth factors vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) were entrapped in PLGA NPs by solvent diffusion method and an antimicrobial peptide (K4) was conjugated to the NP by carbodiimide chemistry. The developed multiple cargo delivery systems with growth factors (VEGF and bFGF) and an antimicrobial peptide (K4) were investigated and optimized for potential wound healing. Results: The system showed a sustained release of growth factors and was evaluated for cytotoxicity by MTT and live/dead assay, which revealed that the bioactivity of the growth factor-entrapped NPs was higher than that of free growth factors, and it also induced enhanced cell proliferation in vitro. Conclusion: The development of a system for the codelivery of growth factors (VEGF and bFGF) and an antimicrobial peptide (K4) was investigated for potential wound healing application. The entrapment of growth factors with very high efficiency is an advantage in this method along with its sustained release from the nanoparticulate system, which will enhance the angiogenesis. Our system also displayed broad-spectrum antimicrobial activity against both gram-positive and gram-negative bacteria.


Assuntos
Peptídeos Catiônicos Antimicrobianos/administração & dosagem , Peptídeos Catiônicos Antimicrobianos/farmacologia , Sistemas de Liberação de Medicamentos , Peptídeos e Proteínas de Sinalização Intercelular/administração & dosagem , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Nanopartículas/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Cicatrização/efeitos dos fármacos , Animais , Antibacterianos/administração & dosagem , Antibacterianos/farmacologia , Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/síntese química , Morte Celular/efeitos dos fármacos , Linhagem Celular , Ensaios de Migração Celular , Liberação Controlada de Fármacos , Difusão Dinâmica da Luz , Fator 2 de Crescimento de Fibroblastos/farmacologia , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Humanos , Cinética , Testes de Sensibilidade Microbiana , Nanopartículas/ultraestrutura , Tamanho da Partícula , Fator A de Crescimento do Endotélio Vascular/administração & dosagem , Fator A de Crescimento do Endotélio Vascular/farmacologia
3.
Int J Mol Med ; 43(5): 2230-2240, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30864673

RESUMO

Hair follicles (HFs) are a well­characterized niche for adult stem cells (SCs), and include epithelial and melanocytic SCs. HF cells are an accessible source of multipotent adult SCs for the generation of the interfollicular epidermis, HF structures and sebaceous glands in addition to the reconstitution of novel HFs in vivo. In the present study, it was demonstrated that HF cells are able to be induced to differentiate into cardiomyocyte­like cells in vitro under specific conditions. It was determined that HF cells cultured on OP9 feeder cells in KnockOut­Dulbecco's modified Eagle's medium/B27 in the presence of vascular endothelial growth factors differentiated into cardiomyocyte­like cells that express markers specific to cardiac lineage, but do not express non­cardiac lineage markers including neural stem/progenitor cell, HF bulge cells or undifferentiated spermatogonia markers. These cardiomyocyte­like cells exhibited a spindle­ and filament­shaped morphology similar to that presented by cardiac muscles and exhibited spontaneous beating that persisted for over 3 months. These results demonstrate that SC reprogramming and differentiation may be induced without resulting in any genetic modification, which is important for the clinical applications of SCs including tissue and organ regeneration.


Assuntos
Folículo Piloso/citologia , Miócitos Cardíacos/citologia , Animais , Biomarcadores/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem da Célula/efeitos dos fármacos , Células Cultivadas , Meios de Cultura/farmacologia , Células Alimentadoras/citologia , Células Alimentadoras/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Camundongos , Miócitos Cardíacos/efeitos dos fármacos , Fenótipo , Fator A de Crescimento do Endotélio Vascular/farmacologia
4.
Int J Pharm ; 561: 236-243, 2019 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-30853484

RESUMO

While vascular endothelial growth factor (VEGF) is an acknowledged potent pro-angiogenic agent there is a need to deliver it at an appropriate concentration for several days to achieve angiogenesis. The aim of this study was to produce microspheres of biodegradable polylactic-co-glycolic acid (PLGA) tailored to achieve sustained release of VEGF at an appropriate concentration over seven days, avoiding excessive unregulated release of VEGF that has been associated with the formation of leaky blood vessels. Several formulations were examined to produce microspheres loaded with both human serum albumin (HSA) and VEGF to achieve release of VEGF between 3 and 10 ng per ml for seven days to match the therapeutic window desired for angiogenesis. In vitro experiments showed an increase in endothelial cell proliferation in response to microspheres bearing VEGF. Similarly, when microspheres containing VEGF were added to the chorionic membrane of fertilised chicken eggs, there was an increase in the development of blood vessels over seven days in response, which was significant for microspheres bearing VEGF and HSA, but not VEGF alone. There was an increase in both blood vessel density and branching - both signs of proangiogenic activity. Further, there was clearly migration of cells to the VEGF loaded microspheres. In summary, we describe the development of an injectable delivery vehicle to achieve spatiotemporal release of physiologically relevant levels of VEGF for several days and demonstrate the angiogenic response to this. We propose that such a treatment vehicle would be suitable for the treatment of ischemic tissue or wounds.


Assuntos
Liberação Controlada de Fármacos , Microesferas , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Albumina Sérica/química , Fator A de Crescimento do Endotélio Vascular/química , Animais , Plásticos Biodegradáveis/química , Proliferação de Células/fisiologia , Galinhas , Córion/irrigação sanguínea , Preparações de Ação Retardada/química , Composição de Medicamentos/métodos , Células Endoteliais/fisiologia , Humanos , Neovascularização Fisiológica/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/farmacologia
5.
Bioanalysis ; 11(5): 381-392, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30892063

RESUMO

AIM: To evaluate the accuracy of the Quantikine Human VEGF Immunoassay (R&D Systems) in the presence of VEGF inhibitors. MATERIALS & METHODS: Quantikine VEGF ELISA (R&D), anti-VEGF165 mAb (R&D), VEGF165 and aflibercept (Regeneron), ranibizumab and bevacizumab (Genentech). RESULTS: Binding affinity of anti-VEGF165 mAb for VEGF was threefold weaker than aflibercept, but 33- and 40-fold stronger than ranibizumab or bevacizumab. Extended incubation of VEGF complexed with inhibitors led to VEGF dissociation from ranibizumab and bevacizumab, but not aflibercept, and subsequent binding by the immunoassay capture antibody. The immunoassay also detected VEGF:ranibizumab and VEGF:bevacizumab complexes but not VEGF:aflibercept complexes. CONCLUSION: The immunoassay cannot accurately quantitate VEGF in the presence of these VEGF inhibitors as they interfere with the capture and detection of free VEGF.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Ensaio de Imunoadsorção Enzimática/métodos , Fator A de Crescimento do Endotélio Vascular/uso terapêutico , Inibidores da Angiogênese/farmacologia , Humanos , Fator A de Crescimento do Endotélio Vascular/farmacologia
6.
Invest Ophthalmol Vis Sci ; 60(2): 605-614, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30726503

RESUMO

Purpose: Surgical techniques such as trabeculectomy aim to treat glaucoma by making an incision into the scleral tissue, to create an alternative drainage pathway for aqueous to flow into the sub-Tenon's/subconjunctival space. However, tissue fibrosis and wound healing occurring after the procedures can reduce the success rate. This study aims to investigate the synergistic effects of aqueous humor in combination with shear stress on the fibrosis response occurring in Tenon's capsule and conjunctival tissue (TCCT) after glaucoma surgery. Methods: Two-dimensional (2D) and 3D in vitro TCCT models were constructed by seeding porcine Tenon's capsule + conjunctival fibroblasts in collagen gel. These were used to investigate key growth factors (singular and natural form) with shear stress, which are believed to influence tissue fibrosis after glaucoma surgery. In addition to cell proliferation assessments, a nondestructive assay to quantify neocollagen synthesis in TCCT models, in response to these factors, has been applied up to 14 days. Results: TCCT fibroblast proliferation increased significantly with doses of TGF-ß, TNF-α, and VEGF, in comparison with the control. Furthermore, fibroblasts exposed to 50% aqueous humor had significantly increased proliferation and actin expression. Shear stress-induced mechanotransduction was also found to promote metabolic activity across experimental conditions. Neocollagen labeling cross validated the fibrosis process. Conclusions: Shear stress appeared to enhance the influence of key growth factors and further promoted fibrotic response within the model. These findings offer a useful insight for further study into the wound-healing response triggered by aqueous fluid outflow after glaucoma surgery.


Assuntos
Túnica Conjuntiva/patologia , Modelos Animais de Doenças , Fibroblastos/patologia , Glaucoma/patologia , Cápsula de Tenon/patologia , Actinas/metabolismo , Animais , Humor Aquoso/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Colágeno/biossíntese , Ensaio de Imunoadsorção Enzimática , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibrose , Imagem Tridimensional , Imuno-Histoquímica , Suínos , Trabeculectomia/métodos , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta/farmacologia , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Fator A de Crescimento do Endotélio Vascular/farmacologia , Vimentina/metabolismo
7.
Phytomedicine ; 57: 95-104, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30668328

RESUMO

BACKGROUND: Several components isolated from rhubarb, the root of Rheum undulatum L., including emodin, rhein, rhaponticin, and piceatannol, have been reported to induce cell death and inhibit metastasis in various types of cancer. Recently, piceatannol-3-O-ß-D-glucopyranoside (PG) isolated from rhubarb was demonstrated to improve vascular dysfunction by inhibiting arginase activity. PURPOSE: In this study, we examined the anti-cancer activities of PG, including effects on the proliferation, metastasis, and angiogenesis of endothelial and malignant cancer cells. RESULTS: We found that PG did not affect the proliferation of human fibrosarcoma (HT1080) and human umbilical vein endothelial cells (HUVECs) at treatments up to 100  µM. However, PG efficiently suppressed the metastatic ability of HT1080 cells, as determined by scratch wound migration, transwell migration/invasion assay, and three-dimensional (3D) spheroid invasion assay. PG significantly suppressed the phorbol 12-myristate 13-acetate (PMA)-induced increase of matrix metalloproteinase (MMP)-9 expression as well as gelatinolytic MMP-9 activity, which are essential for cancer metastasis. In addition, PG treatment reduced the production of proangiogenic factors in HT1080 cells under normoxic and hypoxic conditions and suppressed hypoxia-induced activation of the hypoxia-inducible factor (HIF)-1α pathway. We also found that HUVEC angiogenic activity, including migration and tubular structure formation, were significantly reduced by PG treatment. Moreover, in an in ovo chick chorioallantoic membrane assay, spontaneous and vascular endothelial growth factor (VEGF)-induced vessel formation were significantly inhibited by PG treatment. CONCLUSION: These results collectively indicate that PG has potent anti-metastatic and anti-angiogenic activities with no cytotoxicity. Thus, PG may be useful to limit the hyperplasia of malignant tumors and the spread of cancer to distant secondary organs.


Assuntos
Inibidores da Angiogênese/farmacologia , Fibrossarcoma/tratamento farmacológico , Glucosídeos/farmacologia , Estilbenos/farmacologia , Adulto , Animais , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Embrião de Galinha , Membrana Corioalantoide/irrigação sanguínea , Membrana Corioalantoide/efeitos dos fármacos , Fibrossarcoma/patologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Neovascularização Patológica/tratamento farmacológico , Acetato de Tetradecanoilforbol/farmacologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Fator A de Crescimento do Endotélio Vascular/farmacologia
8.
Vascul Pharmacol ; 115: 18-25, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30634049

RESUMO

Although didymin, a dietary flavonoid glycoside from citrus fruits, known to be a potent antioxidant with anti-cancer activities, its role in angiogenesis is not known. In this study, we examined the effect of didymin on VEGF-induced angiogenesis in vitro and in vivo models. Our results suggest that treatment of human umbilical vein endothelial cell (HUVECs) with didymin significantly prevented the VEGF-induced cell proliferation, migration, and invasion. Further, didymin significantly prevented the VEGF-induced endothelial tube formation in culture. Didymin also attenuated the VEGF-induced generation of ROS, activation of NF-κB and the expression of adhesion molecules such as VCAM-1, ICAM-1, and E-selectin in HUVECs. Further, didymin also prevented the VEGF-induced microvessel sprouting in ex vivo mouse aortic rings. Most importantly, didymin significantly prevented the invasion of endothelial cells and formation of blood capillary-like structures in Matrigel plug model of angiogenesis in mice. Thus, our results suggest a novel antiangiogenic efficacy of didymin in addition to its reported anti-cancer properties, which warrant further development of this agent for cancer therapy.


Assuntos
Indutores da Angiogênese/farmacologia , Inibidores da Angiogênese/farmacologia , Flavonoides/farmacologia , Glicosídeos/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , NF-kappa B/antagonistas & inibidores , Neovascularização Fisiológica/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/farmacologia , Animais , Moléculas de Adesão Celular/metabolismo , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Masculino , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos
9.
Yi Chuan ; 41(1): 76-84, 2019 Jan 20.
Artigo em Chinês | MEDLINE | ID: mdl-30686787

RESUMO

Vascular endothelial growth factor (VEGF) is a dimeric glycoprotein that induces proliferation and migration of vascular endothelial cells as well as regulation of capillary formation around hair follicles which affects the growth and development of hair follicles. cgVEGF164 is a major splice variant of the cashmere goat VEGF-A gene, but its regulation on hair follicles is rarely known. In order to investigate the role of cgVEGF164 on the growth of murine hair follicles, we produced keratin 14 promoter-driven cgVEGF164 transgenic mice via pronuclear microinjection. Firstly, the diameter and density of hair follicles of transgenic mice were compared with non-transgenic control mice in paraffin sections stained by hematoxylin-eosin (H&E). Then, protein expression levels and the phosphorylation of ERK1/2, AKT1 and LEF1 were examined by Western blot. There are five positive individuals among the neonatal mice (positive rate is 8.5%). Compared with non-transgenic control mice, the diameter and density of hair follicles in transgenic mice are both obviously increased. The expression levels of P-ERK1/2/ERK1/2, P-AKT1/AKT1 and P-LEF1/LEF1 are significantly higher in transgenic mice than those in non-transgenic control mice. Based on these results, we conclude that cgVEGF164 as a growth factor can improve the growth of hair follicles which might be mediated by increasing the levels of ERK1/2, AKT1, and LEF1 protein phosphorylation.


Assuntos
Folículo Piloso/crescimento & desenvolvimento , Isoformas de Proteínas/farmacologia , Fator A de Crescimento do Endotélio Vascular/farmacologia , Animais , Cabras , Camundongos , Camundongos Transgênicos , Fosforilação
10.
J Biotechnol ; 284: 84-90, 2018 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-30134149

RESUMO

VEGF165b has been shown to be an effective anti-cancer agent; however, its short half-life limits further application in the clinical field. The development of a mutant VEGF165b with a prolonged half-life is urgently needed for its future application. A mutant VEGF165b was generated by inactivation of its plasmin cleavage site. The mutant and native VEGF165b proteins without purification tags were expressed via the Pichia pastoris expression system followed by purification with a HiTrap heparin affinity chromatography column through optimization of the purification conditions. Furthermore, its binding affinity with VEGF Receptors and its functions in vitro and in vivo were examined. Results showed that the half-life of mutant VEGF165b increased to approximately 10 times (Intravenous), 9.1 times (Intraperitoneal) and 5.4 times (Subcutaneous) greater than that of VEGF165b, and the mutation did not cause significant alteration of VEGFR1 and VEGFR2 binding affinity. Mutant VEGF165b inhibited the proliferation and migration of HUVECs in vitro, similar to the native VEGF165b. In a mouse melanoma model, mutant VEGF165b exhibited stronger anti-tumor activity in comparison with its native counterpart. These results indicate that the mutant VEGF165b had a prolonged half-life and retained the anti-angiogenic activity of the native VEGF165b, suggesting that this novel mutant VEGF165b may be a stronger anti-cancer agent.


Assuntos
Antineoplásicos , Melanoma/tratamento farmacológico , Fator A de Crescimento do Endotélio Vascular , Animais , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Meia-Vida , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Masculino , Camundongos Endogâmicos C57BL , Mutação , Pichia/genética , Ratos Sprague-Dawley , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Fator A de Crescimento do Endotélio Vascular/farmacologia , Fator A de Crescimento do Endotélio Vascular/uso terapêutico , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
11.
Theriogenology ; 120: 147-156, 2018 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-30121547

RESUMO

Current research suggests that supplementing in vitro culture (IVC) media with vascular endothelial growth factor (VEGF) may have beneficial effects on the development of porcine embryos in vitro. However, the molecular signaling mechanisms underlying this effect are unclear. Therefore, we aimed to investigate the effects of VEGF on molecular signaling events during in vitro embryonic development of porcine embryos. Porcine oocytes matured in vitro were fertilized, and the resultant zygotes were cultured with 5 ng/mL of VEGF supplemented with or without fetal bovine serum from day 4 till day 7. Without VEGF and/or FBS served as the control group. Real-time quantitative PCR was used to detect expression patterns of apoptosis- and oxidative stress-related genes in day 7 blastocysts (BLs). Early-stage apoptosis was detected by annexin-V assays in day 2 and day 7 embryos. We found that the addition of VEGF throughout the culture period with or without FBS supplementation significantly improved embryo survival and development. Supplementation with VEGF in the IVC medium significantly increased early BL formation (p < 0.05), although addition of FBS on day 4 significantly increased hatched BL formation (p < 0.05) regardless of VEGF supplementation. However, supplementation of media with both VEGF and FBS increased the formation of expanded BLs synergistically. The average total cell numbers per BL were significantly (p < 0.05) higher in embryos supplemented with VEGF and FBS than in those supplemented with either VEGF or FBS alone. We also found that accumulation of reactive oxygen species in VEGF-treated embryos was significantly lower (p < 0.05) than that in untreated embryos. The mRNA levels of caspase-3 were significantly lower (p < 0.05), and those of Bcl-2 and Nrf-2 were significantly higher (p < 0.05) in embryos grown in VEGF-supplemented media than in embryos grown in non-supplemented media. Furthermore, on day 2, the numbers of viable embryos (44.06 ±â€¯3.94%) and blastomeres (67.18 ±â€¯3.60%) were significantly higher (p < 0.05), and the numbers of early apoptotic embryos (55.94 ±â€¯3.94) and blastomeres (23.23 ±â€¯4.22) were significantly lower (p < 0.05) in VEGF-treated BLs than in controls. Furthermore, the numbers of early apoptotic cells in BLs on day 7 were also significantly lower (p < 0.05) in VEGF-treated BLs than in controls. Overall, our results indicate that supplementing IVC media with VEGF during in vitro culture of porcine embryos increases their developmental potential.


Assuntos
Blastocisto/efeitos dos fármacos , Fertilização In Vitro/veterinária , Suínos/embriologia , Fator A de Crescimento do Endotélio Vascular/farmacologia , Animais , Apoptose , Blastocisto/fisiologia , Meios de Cultura , Técnicas de Cultura Embrionária/veterinária , Desenvolvimento Embrionário/efeitos dos fármacos , Estresse Oxidativo , Transdução de Sinais
12.
Zhonghua Kou Qiang Yi Xue Za Zhi ; 53(1): 42-48, 2018 Jan 09.
Artigo em Chinês | MEDLINE | ID: mdl-29972963

RESUMO

Objective: To evaluate the effect of vascular endothelial growth factor (VEGF)-loaded microspheres on dental pulp tissue regeneration and vascularization in vivo. Methods:In vitro release experiment and human umbilical vein endothelial cell migration experiment were conducted with VEGF loaded microspheres. The dental pulp stem cells (DPSC) were co-cultured with VEGF microspheres to observe the compatibility between the cells and the microspheres. DPSC and VEGF loaded microspheres were injected into the root lumen through the apical foramen, which were then transplanted subcutaneously into nude mice. Histological and immunohistochemical features were observed after nine weeks. Results: DPSCs attached and spread on the surface of the microspheres. HE staining showed that the regenerated pulp-like tissue fulfilled the whole apex and middle third of the root. Differentiated odontoblast-like cells aligned with the existing tubular root dentin. Conclusions: VEGF-loaded microspheres promoted the regeneration of pulp-like tissues and formation of blood vessels.


Assuntos
Polpa Dentária/efeitos dos fármacos , Microesferas , Regeneração/efeitos dos fármacos , Células-Tronco/fisiologia , Fator A de Crescimento do Endotélio Vascular/farmacologia , Animais , Diferenciação Celular , Movimento Celular/efeitos dos fármacos , Técnicas de Cocultura , Polpa Dentária/irrigação sanguínea , Polpa Dentária/citologia , Polpa Dentária/fisiologia , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Camundongos , Camundongos Nus , Neovascularização Fisiológica/efeitos dos fármacos , Odontoblastos , Regeneração/fisiologia , Transplante de Células-Tronco/métodos
13.
Int J Mol Sci ; 19(7)2018 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-29933613

RESUMO

Vascular endothelial growth factor (VEGF) is important for maintaining healthy endothelium, which is crucial for vascular integrity. In this paper, we show that VEGF stimulates the nuclear translocation of endothelial differentiation-related factor 1 (EDF1), a highly conserved intracellular protein implicated in molecular events that are pivotal to endothelial function. In the nucleus, EDF1 serves as a transcriptional coactivator of peroxisome proliferator-activated receptor gamma (PPARγ), which has a protective role in the vasculature. Indeed, silencing EDF1 prevents VEGF induction of PPARγ activity as detected by gene reporter assay. Accordingly, silencing EDF1 markedly inhibits the stimulatory effect of VEGF on the expression of FABP4, a PPARγ-inducible gene. As nitric oxide is a marker of endothelial function, it is noteworthy that we report a link between EDF1 silencing, decreased levels of FABP4, and nitric oxide production. We conclude that EDF1 is required for VEGF-induced activation of the transcriptional activity of PPARγ.


Assuntos
Proteínas de Ligação a Calmodulina/genética , Proteínas de Ligação a Ácido Graxo/genética , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , PPAR gama/genética , Fator A de Crescimento do Endotélio Vascular/farmacologia , Proteínas de Ligação a Calmodulina/antagonistas & inibidores , Proteínas de Ligação a Calmodulina/metabolismo , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Proteínas de Ligação a Ácido Graxo/metabolismo , Regulação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Óxido Nítrico/biossíntese , PPAR gama/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Transcrição Genética , Fator A de Crescimento do Endotélio Vascular/metabolismo
14.
Colloids Surf B Biointerfaces ; 167: 550-559, 2018 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-29730577

RESUMO

The purpose of this work was to investigate if the biomimetically conjugated VEGF and HUVECs co-culture could modulate the osteogenic and angiogenic differentiation of MSCs derived from rat and human bone marrow (rMSCs and hMSCs). After treated by ammonia plasma, Poly(lactic-co-glycolic acid) (PLGA) electrospun nanofibers were immobilized with VEGF through heparin to fulfil the sustained release. The proliferation capacity of rMSCs and hMSCs on neat PLGA nanofibers (NF) and VEGF immobilized NF (NF-VEGF) surfaces were assessed by CCK-8 and compared when MSCs were mono-cultured and co-cultured with HUVECs. The effect of VEGF and HUVECs co-culturing on osteogenic and angiogenic differentiation of rMSCs and hMSCs were investigated by calcium deposits and CD31 expression on NF and NF-VEGF surfaces. The results indicated that VEGF has been biomimetically immobilized onto PLGA nanofibers surface and kept sustained release successfully. The CD31 staining results showed that both VEGF and HUVECs co-culture could enhance the angiogenesis of rMSCs and hMSCs. However, the proliferation and osteogenic differentiation of MSCs when cultured with VEGF and HUVECs showed a species dependent response. Taken together, VEGF immobilization and co-culture with HUVECs promoted angiogenesis of MSCs, indicating a good strategy for vascularization in bone tissue engineering.


Assuntos
Células Endoteliais da Veia Umbilical Humana/fisiologia , Células-Tronco Mesenquimais/fisiologia , Neovascularização Fisiológica/fisiologia , Osteogênese/fisiologia , Fator A de Crescimento do Endotélio Vascular/química , Animais , Biomimética/métodos , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/fisiologia , Células Cultivadas , Técnicas de Cocultura , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Ácido Láctico/química , Células-Tronco Mesenquimais/efeitos dos fármacos , Nanofibras/química , Neovascularização Fisiológica/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Ratos Sprague-Dawley , Propriedades de Superfície , Engenharia Tecidual/métodos , Fator A de Crescimento do Endotélio Vascular/farmacologia
15.
Cell Biol Int ; 42(8): 1060-1068, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29745446

RESUMO

The improvement of fat graft viability might depend on the presence of multipotent resident adipose derived stem cells (ADSCs) which is the important component of stromal vascular fraction (SVF). Vascular endothelial growth factor (VEGF) and angiogenin-1 (Ang-1) are responsible for neovascularization. However, their half-life is too short to produce a biological effect. We thus investigated whether VEGF-ANG-1-polylactic acid (PLA) microspheres could enhance the angiogenic properties of ADSCs. PLA microspheres containing VEGF and ANG-1 were prepared by in vitro ultrasonic emulsification and characterized according to their encapsulation efficiency (EE), drug-loading rate (DL), particle size, and drug release. The systemic toxicity of empty loaded nanospheres (NPs) and the ability of these microspheres to promote the proliferation and differentiation of ADSCs were evaluated. The EE and DL were above 86 and 2.8%, respectively. The drug release was completed after 20 days. Systemic toxicity was verified in ADSCs that received the unloaded NPs. It was observed that ADSCs treated with VEGF-ANG-1-PLA microspheres had an increase in the proliferation and the number of CD31 positive cells. ADSCs proliferation and differentiation toward endothelial cells (ECs) could be enhanced by the addition of VEGF-ANG-1-PLA nano-sustained release microspheres.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Nanosferas/química , Poliésteres/farmacologia , Ribonuclease Pancreático/farmacologia , Fator A de Crescimento do Endotélio Vascular/farmacologia , Tecido Adiposo/citologia , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Células Cultivadas , Preparações de Ação Retardada , Liberação Controlada de Fármacos , Fator de Crescimento de Hepatócito/genética , Fator de Crescimento de Hepatócito/metabolismo , Humanos , Antígeno Ki-67/metabolismo , Microscopia Eletrônica de Transmissão , Molécula-1 de Adesão Celular Endotelial a Plaquetas/genética , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Poliésteres/química , Poliésteres/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ribonuclease Pancreático/química , Ribonuclease Pancreático/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismo , Fator A de Crescimento do Endotélio Vascular/química , Fator A de Crescimento do Endotélio Vascular/metabolismo
16.
J Appl Oral Sci ; 26: e20170437, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29791567

RESUMO

Tissue bioengineering has been applied to Endodontics to seek a more biological treatment. The presence of blood vessels is crucial for cell nutrition during tissue formation. Objective This study analysed the application of vascular endothelial growth factor (VEGF) in the angiogenesis of mature root canals. Material and methods Upper first molars of twelve 13-week old Wistar male rats were used. The root pulp of the mesiobuccal canal was removed and the root canal instrumented with K-files up to size #25. Periapical bleeding was induced into the root canal by introducing a #15 K-file beyond the apex. The teeth on the right side of the arch were filled up with blood clot (G1), whereas those on the left side were filled up with blood clot plus 50 ng/ml of VEGF (G2). Teeth were sealed with light-curing glass-ionomer cement and the animals were sacrificed after 60 days. The maxilla was dissected and fixed before obtaining serial sections for histological processing with haematoxylin-eosin (HE) and immunohistochemical factor-VIII. Immunohistochemical labelling was evaluated using scores for statistical analysis. Results Immunohistological analysis demonstrated the presence of angiogenesis in both groups, but with higher angiogenic maturation in G2 during the experimental period (p<0.05). HE staining showed connective tissue with absence of odontoblasts in all specimens. Conclusions It can be concluded that it is possible to obtain angiogenesis in mature root canals with or without the use of VEGF, although the latter tends to accelerate blood vessel formation.


Assuntos
Cavidade Pulpar/irrigação sanguínea , Neovascularização Fisiológica/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/farmacologia , Animais , Bioengenharia , Coagulação Sanguínea/fisiologia , Cavidade Pulpar/efeitos dos fármacos , Imuno-Histoquímica , Masculino , Ratos Wistar , Reprodutibilidade dos Testes , Tratamento do Canal Radicular/métodos , Fatores de Tempo , Resultado do Tratamento
17.
J Vasc Res ; 55(3): 125-135, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29779031

RESUMO

BACKGROUND: The development of models that incorporate intact microvascular networks enables the investigation of multicellular dynamics during angiogenesis. Our laboratory introduced the rat mesentery culture model as such a tool, which would be enhanced with mouse tissue. Since mouse mesentery is avascular, an alternative is mouse mesometrium, the connective tissue of uterine horns. The study's objective was to demonstrate that mouse mesometrium contains microvascular networks that can be cultured to investigate multicellular dynamics during angiogenesis. METHODS: Harvested mesometrium tissues from C57Bl/6 female mice were cultured in media with serum for up to 7 days. PECAM, NG2, αSMA, and LYVE-1 labeling identified endothelial cells, pericytes, smooth muscle cells, and lymphatic endothelial cells, respectively. RESULTS: These cells comprised microvascular networks with arterioles, venules, and capillaries. Compared to day 0, capillary sprouts per vascular length were increased by 3 and 5 days in culture (day 0, 0.08 ± 0.01; day 3, 3.19 ± 0.78; day 5, 2.49 ± 0.05 sprouts/mm; p < 0.05). Time-lapse imaging of cultured tissues from FlkEGFP mice showcases the use of the model for lineage studies. The impact is supported by the identification of endothelial cell jumping from one sprout to another. CONCLUSION: These results introduce a novel culture model for investigating multicellular dynamics during angiogenesis in real-time ex vivo microvascular networks.


Assuntos
Microvasos/fisiologia , Neovascularização Fisiológica , Útero/irrigação sanguínea , Actinas/metabolismo , Animais , Antígenos/metabolismo , Biomarcadores/metabolismo , Feminino , Glicoproteínas/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Microvasos/efeitos dos fármacos , Microvasos/metabolismo , Modelos Animais , Neovascularização Fisiológica/efeitos dos fármacos , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Proteoglicanas/metabolismo , Fatores de Tempo , Imagem com Lapso de Tempo , Técnicas de Cultura de Tecidos , Fator A de Crescimento do Endotélio Vascular/farmacologia
18.
Int J Mol Sci ; 19(5)2018 Apr 28.
Artigo em Inglês | MEDLINE | ID: mdl-29710813

RESUMO

Skeletal muscle tissue engineering (TE) aims to efficiently repair large congenital and acquired defects. Biological acellular scaffolds are considered a good tool for TE, as decellularization allows structural preservation of tissue extracellular matrix (ECM) and conservation of its unique cytokine reservoir and the ability to support angiogenesis, cell viability, and proliferation. This represents a major advantage compared to synthetic scaffolds, which can acquire these features only after modification and show limited biocompatibility. In this work, we describe the ability of a skeletal muscle acellular scaffold to promote vascularization both ex vivo and in vivo. Specifically, chicken chorioallantoic membrane assay and protein array confirmed the presence of pro-angiogenic molecules in the decellularized tissue such as HGF, VEGF, and SDF-1α. The acellular muscle was implanted in BL6/J mice both subcutaneously and ortotopically. In the first condition, the ECM-derived scaffold appeared vascularized 7 days post-implantation. When the decellularized diaphragm was ortotopically applied, newly formed blood vessels containing CD31⁺, αSMA⁺, and vWF⁺ cells were visible inside the scaffold. Systemic injection of Evans Blue proved function and perfusion of the new vessels, underlying a tissue-regenerative activation. On the contrary, the implantation of a synthetic matrix made of polytetrafluoroethylene used as control was only surrounded by vWF⁺ cells, with no cell migration inside the scaffold and clear foreign body reaction (giant cells were visible). The molecular profile and the analysis of macrophages confirmed the tendency of the synthetic scaffold to enhance inflammation instead of regeneration. In conclusion, we identified the angiogenic potential of a skeletal muscle-derived acellular scaffold and the pro-regenerative environment activated in vivo, showing clear evidence that the decellularized diaphragm is a suitable candidate for skeletal muscle tissue engineering and regeneration.


Assuntos
Diafragma/química , Espaço Extracelular/química , Neovascularização Fisiológica , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Células Cultivadas , Quimiocina CXCL12/análise , Quimiocina CXCL12/farmacologia , Embrião de Galinha , Diafragma/citologia , Feminino , Fator de Crescimento de Hepatócito/análise , Fator de Crescimento de Hepatócito/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fator A de Crescimento do Endotélio Vascular/análise , Fator A de Crescimento do Endotélio Vascular/farmacologia
19.
J Photochem Photobiol B ; 183: 385-390, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29772391

RESUMO

This investigation aimed to develop the silver (Ag) nanoparticles incorporated vascular endothelial growth factor (VEGF) for the improvement wound healing and reduce Aseptic necrosis in treatment of femoral fracture healing. The spherical shaped Ag nanoparticles with improved morphology have been effectively synthesized via microwave assisted method using ionic liquids 1-dodecyl-3-methylimidazolium chloride. The morphological structure and crystalline properties of Ag nanoparticles are analyzed by using UV, XRD and TEM-EDX analytical methods. The average grain size of the Ag nanoparticles is 20 nm, which was observed by defining the width of the (111) Bragg reflection with the Debye-Scherer formula and TEM results. The biological analyses confirmed that the Ag nanoparticles with VEGF molecules are promoted the cell adhesion and proliferation of Human mesenchymal stem cells (MSCs) cells. Ag NPs at appropriate concentrations have favorable biocompatibility to encourage cell activation properties like cell proliferation, cytokines release and chemotaxis. In the present study, our experimental results indicated that Ag NPs incorporated VEGF material are highly favorable to fracture healing and mainly as blood vessel repair. The surface morphology improved synthetic Ag NPs using ionic liquids has shown advantageous for cell activity and also improve the materials performances with VEGF for the regeneration of femoral fractures.


Assuntos
Portadores de Fármacos/química , Líquidos Iônicos/química , Nanopartículas Metálicas/química , Prata/química , Fator A de Crescimento do Endotélio Vascular/química , Adesão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Criança , Citocinas/metabolismo , Fraturas do Fêmur/patologia , Fraturas do Fêmur/cirurgia , Cabeça do Fêmur/patologia , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Tamanho da Partícula , Fator A de Crescimento do Endotélio Vascular/farmacologia , Fator A de Crescimento do Endotélio Vascular/uso terapêutico , Cicatrização/efeitos dos fármacos
20.
Cancer Sci ; 109(6): 1981-1994, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29664206

RESUMO

Infantile hemangioma (IH) is a benign tumor that is formed by aberrant angiogenesis and that undergoes spontaneous regression over time. Propranolol, the first-line therapy for IH, inhibits angiogenesis by downregulating activation of the vascular endothelial growth factor (VEGF) pathway, which is hyperactivated in IH. However, this treatment is reportedly ineffective for 10% of tumors, and 19% of patients relapse after propranolol treatment. Both pro-angiogenic and anti-angiogenic factors regulate angiogenesis, and pigment epithelium-derived factor (PEDF) is the most effective endogenous anti-angiogenic factor. PEDF/VEGF ratio controls many angiogenic processes, but its role in IH and the relationship between this ratio and propranolol remain unknown. Results of the present study showed that the PEDF/VEGF ratio increased during the involuting phase of IH compared with the proliferating phase. Similarly, in hemangioma-derived endothelial cells (HemEC), which were isolated with magnetic beads, increasing the PEDF/VEGF ratio inhibited proliferation, migration, and tube formation and promoted apoptosis. Mechanistically, the VEGF receptors (VEGFR1 and VEGFR2) and PEDF receptor (laminin receptor, LR) were highly expressed in both IH tissues and HemEC, and PEDF inhibited HemEC function by binding to LR. Interestingly, we found that propranolol increased the PEDF/VEGF ratio but did so by lowering VEGF expression rather than by upregulating PEDF as expected. Furthermore, the combination of PEDF and propranolol had a more suppressive effect on HemEC. Consequently, our results suggested that the PEDF/VEGF ratio played a pivotal role in the spontaneous regression of IH and that the combination of PEDF and propranolol might be a promising treatment strategy for propranolol-resistant IH.


Assuntos
Proteínas do Olho/metabolismo , Hemangioma/tratamento farmacológico , Fatores de Crescimento Neural/metabolismo , Propranolol/uso terapêutico , Serpinas/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Apoptose/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Proteínas do Olho/genética , Proteínas do Olho/farmacologia , Hemangioma/irrigação sanguínea , Hemangioma/metabolismo , Humanos , Lactente , Neovascularização Patológica/metabolismo , Neovascularização Patológica/prevenção & controle , Fatores de Crescimento Neural/genética , Fatores de Crescimento Neural/farmacologia , Receptores de Laminina/genética , Receptores de Laminina/metabolismo , Remissão Espontânea , Serpinas/genética , Serpinas/farmacologia , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/farmacologia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 1 de Fatores 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 , Vasodilatadores/uso terapêutico
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