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1.
Cardiovasc Res ; 120(7): 745-755, 2024 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-38507654

RESUMO

AIMS: In hypoxia, endothelial cells (ECs) proliferate, migrate, and form new vasculature in a process called angiogenesis. Recent studies have suggested that ECs rely on glycolysis to meet metabolic needs for angiogenesis in ischaemic tissues, and several studies have investigated the molecular mechanisms integrating angiogenesis and endothelial metabolism. Here, we investigated the role of stem cell factor (SCF) and its receptor, cKIT, in regulating endothelial glycolysis during hypoxia-driven angiogenesis. METHODS AND RESULTS: SCF and cKIT signalling increased the glucose uptake, lactate production, and glycolysis in human ECs under hypoxia. Mechanistically, SCF and cKIT signalling enhanced the expression of genes encoding glucose transporter 1 (GLUT1) and glycolytic enzymes via Akt- and ERK1/2-dependent increased translation of hypoxia inducible factor 1A (HIF1A). In hypoxic conditions, reduction of glycolysis and HIF-1α expression using chemical inhibitors significantly reduced the SCF-induced in vitro angiogenesis in human ECs. Compared with normal mice, mice with oxygen-induced retinopathy (OIR), characterized by ischaemia-driven pathological retinal neovascularization, displayed increased levels of SCF, cKIT, HIF-1α, GLUT1, and glycolytic enzymes in the retina. Moreover, cKIT-positive neovessels in the retina of mice with OIR showed elevated expression of GLUT1 and glycolytic enzymes. Further, blocking SCF and cKIT signalling using anti-SCF neutralizing IgG and cKIT mutant mice significantly reduced the expression of HIF-1α, GLUT1, and glycolytic enzymes and decreased the pathological neovascularization in the retina of mice with OIR. CONCLUSION: We demonstrated that SCF and cKIT signalling regulate angiogenesis by controlling endothelial glycolysis in hypoxia and elucidated the SCF/cKIT/HIF-1α axis as a novel metabolic regulation pathway during hypoxia-driven pathological angiogenesis.


Assuntos
Hipóxia Celular , Transportador de Glucose Tipo 1 , Glicólise , Subunidade alfa do Fator 1 Induzível por Hipóxia , Proteínas Proto-Oncogênicas c-kit , Transdução de Sinais , Fator de Células-Tronco , Animais , Humanos , Fator de Células-Tronco/metabolismo , Fator de Células-Tronco/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Transportador de Glucose Tipo 1/metabolismo , Transportador de Glucose Tipo 1/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Proteínas Proto-Oncogênicas c-kit/genética , Células Endoteliais da Veia Umbilical Humana/metabolismo , Camundongos Endogâmicos C57BL , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Neovascularização Retiniana/metabolismo , Neovascularização Retiniana/patologia , Neovascularização Retiniana/genética , Camundongos , Neovascularização Fisiológica , Células Cultivadas , Modelos Animais de Doenças , Glucose/metabolismo
2.
BMB Rep ; 55(11): 565-570, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36016502

RESUMO

Pulmonary arterial hypertension (PAH) is a progressive and devastating disease whose pathogenesis is associated with a phenotypic switch of pulmonary arterial vascular smooth muscle cells (PASMCs). Bone morphogenetic protein (BMP) signaling and potassium two pore domain channel subfamily K member 3 (KCNK3) play crucial roles in PAH pathogenesis. However, the relationship between BMP signaling and KCNK3 expression in the PASMC phenotypic switching process has not been studied. In this study, we explored the effect of BMPs on KCNK3 expression and the role of KCNK3 in the BMP-mediated PASMC phenotypic switch. Expression levels of BMP receptor 2 (BMPR2) and KCNK3 were downregulated in PASMCs of rats with PAH compared to those in normal controls, implying a possible association between BMP/BMPR2 signaling and KCNK3 expression in the pulmonary vasculature. Treatment with BMP2, BMP4, and BMP7 significantly increased KCNK3 expression in primary human PASMCs (HPASMCs). BMPR2 knockdown and treatment with Smad1/5 signaling inhibitor substantially abrogated the BMP-induced increase in KCNK3 expression, suggesting that KCNK3 expression in HPASMCs is regulated by the canonical BMP-BMPR2-Smad1/5 signaling pathway. Furthermore, KCNK3 knockdown and treatment with a KCNK3 channel blocker completely blocked BMP-mediated anti-proliferation and expression of contractile marker genes in HPAMSCs, suggesting that the expression and functional activity of KCNK3 are required for BMP-mediated acquisition of the quiescent PASMC phenotype. Overall, our findings show a crosstalk between BMP signaling and KCNK3 in regulating the PASMC phenotype, wherein BMPs upregulate KCNK3 expression and KCNK3 then mediates BMP-induced phenotypic switching of PASMCs. Our results indicate that the dysfunction and/or downregulation of BMPR2 and KCNK3 observed in PAH work together to induce aberrant changes in the PASMC phenotype, providing insights into the complex molecular pathogenesis of PAH. [BMB Reports 2022; 55(11): 565-570].


Assuntos
Hipertensão Pulmonar , Músculo Liso Vascular , Proteínas do Tecido Nervoso , Canais de Potássio de Domínios Poros em Tandem , Animais , Humanos , Ratos , Receptores de Proteínas Morfogenéticas Ósseas Tipo II/genética , Receptores de Proteínas Morfogenéticas Ósseas Tipo II/metabolismo , Proteínas Morfogenéticas Ósseas/genética , Proliferação de Células , Células Cultivadas , Hipertensão Pulmonar/metabolismo , Hipertensão Pulmonar/patologia , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Fenótipo , Transdução de Sinais , Canais de Potássio de Domínios Poros em Tandem/genética , Proteínas do Tecido Nervoso/genética
3.
Int J Mol Sci ; 23(16)2022 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-36012732

RESUMO

The fibroblast growth factor (FGF) family has various biological functions, including cell growth, tissue regeneration, embryonic development, metabolism, and angiogenesis. In the case of hair growth, several members of the FGF family, such as FGF1 and FGF2, are involved in hair growth, while FGF5 has the opposite effect. In this study, the regulation of the hair growth cycle by FGF12 was investigated. To observe its effect, the expression of FGF12 was downregulated in mice and outer root sheath (ORS) by siRNA transfection, while FGF12 overexpression was carried out using FGF12 adenovirus. For the results, FGF12 was primarily expressed in ORS cells with a high expression during the anagen phase of hair follicles. Knockdown of FGF12 delayed telogen-to-anagen transition in mice and decreased the hair length in vibrissae hair follicles. It also inhibited the proliferation and migration of ORS cells. On the contrary, FGF12 overexpression increased the migration of ORS cells. FGF12-overexpressed ORS cells induced the telogen-to-anagen transition in the animal model. In addition, FGF12 overexpression regulated the expression of PDGF-CC, MDK, and HB-EGF, and treatment of these factors exhibited hair growth promotion. Altogether, FGF12 promoted hair growth by inducing the anagen phase of hair follicles, suggesting the potential for hair loss therapy.


Assuntos
Fatores de Crescimento de Fibroblastos , Folículo Piloso , Cabelo , Animais , Ciclo Celular , Fatores de Crescimento de Fibroblastos/genética , Fatores de Crescimento de Fibroblastos/metabolismo , Cabelo/crescimento & desenvolvimento , Folículo Piloso/metabolismo , Camundongos , Vibrissas
4.
Pharmaceutics ; 13(10)2021 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-34683841

RESUMO

The wet type of age-related macular degeneration (AMD) accompanies the subfoveal choroidal neovascularization (CNV) caused by the abnormal extension or remodeling of blood vessels to the macula and retinal pigment epithelium (RPE). Vascular endothelial growth factor (VEGF) is known to play a crucial role in the pathogenesis of the disease. In this study, we tried to repurpose an investigational anticancer drug, rivoceranib, which is a selective inhibitor of VEGF receptor-2 (VEGFR2), and evaluate the therapeutic potential of the drug for the treatment of wet-type AMD in a laser-induced CNV mouse model using microsphere-based sustained drug release formulations. The PLGA-based rivoceranib microsphere can carry out a sustained delivery of rivoceranib for 50 days. When administered intravitreally, the sustained microsphere formulation of rivoceranib effectively inhibited the formation of subfoveal neovascular lesions in mice.

5.
Pharmaceutics ; 13(8)2021 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-34452268

RESUMO

Stem cell factor (SCF) and its receptor, cKIT, are novel regulators of pathological neovascularization in the eye, which suggests that inhibition of SCF/cKIT signaling may be a novel pharmacological strategy for treating neovascular age-related macular degeneration (AMD). This study evaluated the therapeutic potential of a newly developed fully human monoclonal antibody targeting cKIT, NN2101, in a murine model of neovascular AMD. In hypoxic human endothelial cells, NN2101 substantially inhibited the SCF-induced increase in angiogenesis and activation of the cKIT signaling pathway. In a murine model of neovascular AMD, intravitreal injection of NN2101 substantially inhibited the SCF/cKIT-mediated choroidal neovascularization (CNV), with efficacy comparable to aflibercept, a vascular endothelial growth factor inhibitor. A combined intravitreal injection of NN2101 and aflibercept resulted in an additive therapeutic effect on CNV. NN2101 neither caused ocular toxicity nor interfered with the early retinal vascular development in mice. Ocular pharmacokinetic analysis in rabbits indicated that NN2101 demonstrated a pharmacokinetic profile suitable for intravitreal injection. These findings provide the first evidence of the potential use of the anti-cKIT blocking antibody, NN2101, as an alternative or additive therapeutic for the treatment of neovascular AMD.

6.
J Med Chem ; 64(9): 5535-5550, 2021 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-33902285

RESUMO

Diabetic retinopathy is the leading cause of blindness which is associated with excessive angiogenesis. Using the structure of wondonin marine natural products, we previously created a scaffold to develop a novel type of antiangiogenesis agent that possesses minimized cytotoxicity. To overcome its poor pharmaceutical properties, we further modified the structure. A new scaffold was derived in which the stereogenic carbon was changed to nitrogen and the 1,2,3-triazole ring was replaced by an alkyl chain. By comparing the bioactivity versus cytotoxicity, compound 31 was selected, which has improved aqueous solubility and an enhanced selectivity index. Mechanistically, 31 suppressed angiopoietin-2 (ANGPT2) expression induced by high glucose in retinal cells and exhibited in vivo antiangiogenic activity in choroidal neovascularization and oxygen-induced retinopathy mouse models. These results suggest the potential of 31 as a lead to develop antiangiogenic small-molecule drugs to treat diabetic retinopathy and as a chemical tool to elucidate new mechanisms of angiogenesis.


Assuntos
Inibidores da Angiogênese/farmacologia , Desenho de Fármacos , Neovascularização Fisiológica/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Inibidores da Angiogênese/química , Inibidores da Angiogênese/metabolismo , Inibidores da Angiogênese/uso terapêutico , Angiopoietina-2/genética , Angiopoietina-2/metabolismo , Animais , Sobrevivência Celular/efeitos dos fármacos , Retinopatia Diabética/tratamento farmacológico , Retinopatia Diabética/patologia , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Estabilidade de Medicamentos , Glucose/farmacologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/uso terapêutico , Relação Estrutura-Atividade , Triazóis/química , Triazóis/metabolismo , Triazóis/farmacologia , Triazóis/uso terapêutico
7.
Hypertension ; 76(6): 1778-1786, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33100045

RESUMO

Loss of BMP (bone morphogenic protein) signaling induces a phenotype switch of pulmonary arterial smooth muscle cells (PASMCs), which is the pathological basis of pulmonary vascular remodeling in pulmonary arterial hypertension (PAH). Here, we identified FGF12 (fibroblast growth factor 12) as a novel regulator of the BMP-induced phenotype change in PASMCs and elucidated its role in pulmonary vascular remodeling during PAH development. Using murine models of PAH and lung specimens of patients with PAH, we observed that FGF12 expression was significantly reduced in PASMCs. In human PASMCs, FGF12 expression was increased by canonical BMP signaling. FGF12 knockdown blocked the antiproliferative and prodifferentiation effect of BMP on human PASMCs, suggesting that FGF12 is required for the BMP-mediated acquisition of the quiescent and differentiated PASMC phenotype. Mechanistically, FGF12 regulated the BMP-induced phenotype change by inducing MEF2a (myocyte enhancer factor 2a) phosphorylation via p38MAPK signaling, thereby modulating the expression of MEF2a target genes involved in cell proliferation and differentiation. Furthermore, we observed that TG (transgenic) mice with smooth muscle cell-specific FGF12 overexpression were protected from chronic hypoxia-induced PAH development, pulmonary vascular remodeling, and right ventricular hypertrophy. Consistent with the in vitro data using human PASMCs, FGF12 TG mice showed increased MEF2a phosphorylation and a substantial change in MEF2a target gene expression, compared with the WT (wild type) controls. Overall, our findings demonstrate a novel BMP/FGF12/MEF2a pathway regulating the PASMC phenotype switch and suggest FGF12 as a potential target for the development of therapeutics for ameliorating pulmonary vascular remodeling in PAH.


Assuntos
Fatores de Crescimento de Fibroblastos/genética , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Hipertensão Arterial Pulmonar/genética , Remodelação Vascular/genética , Animais , Proteínas Morfogenéticas Ósseas/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Proliferação de Células/genética , Células Cultivadas , Fatores de Crescimento de Fibroblastos/metabolismo , Humanos , Sistema de Sinalização das MAP Quinases/genética , Fatores de Transcrição MEF2/genética , Fatores de Transcrição MEF2/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Músculo Liso Vascular/citologia , Hipertensão Arterial Pulmonar/fisiopatologia , Artéria Pulmonar/metabolismo , Artéria Pulmonar/fisiopatologia , Ratos Sprague-Dawley
8.
Exp Mol Med ; 52(10): 1744-1753, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-33051573

RESUMO

Dipeptidyl peptidase-4 (DPP-4) inhibitors are used for the treatment of type 2 diabetes mellitus (DM). Recent studies have shown that beyond their effect in lowing glucose, DPP-4 inhibitors mitigate DM-related microvascular complications, such as diabetic retinopathy. However, the mechanism by which pathological retinal neovascularization, a major clinical manifestation of diabetic retinopathy, is inhibited is unclear. This study sought to examine the effects of evogliptin, a potent DPP-4 inhibitor, on pathological retinal neovascularization in mice and elucidate the mechanism by which evogliptin inhibits angiogenesis mediated by vascular endothelial growth factor (VEGF), a key factor in the vascular pathogenesis of proliferative diabetic retinopathy (PDR). In a murine model of PDR, an intravitreal injection of evogliptin significantly suppressed aberrant retinal neovascularization. In human endothelial cells, evogliptin reduced VEGF-induced angiogenesis. Western blot analysis showed that evogliptin inhibited the phosphorylation of signaling molecules associated with VEGF-induced cell adhesion and migration. Moreover, evogliptin substantially inhibited the VEGF-induced activation of adenosine 5'-diphosphate ribosylation factor 6 (Arf6), a small guanosine 5'-triphosphatase (GTPase) that regulates VEGF receptor 2 signal transduction. Direct activation of Arf6 using a chemical inhibitor of Arf-directed GTPase-activating protein completely abrogated the inhibitory effect of evogliptin on VEGF-induced activation of the angiogenic signaling pathway, which suggests that evogliptin suppresses VEGF-induced angiogenesis by blocking Arf6 activation. Our results provide insights into the molecular mechanism of the direct inhibitory effect of the DPP-4 inhibitor evogliptin on pathological retinal neovascularization. In addition to its glucose-lowering effect, the antiangiogenic effect of evogliptin could also render it beneficial for individuals with PDR.


Assuntos
Fatores de Ribosilação do ADP/metabolismo , Inibidores da Dipeptidil Peptidase IV/farmacologia , Piperazinas/farmacologia , Neovascularização Retiniana/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Fator 6 de Ribosilação do ADP , Fatores de Ribosilação do ADP/genética , Inibidores da Angiogênese/farmacologia , Animais , Modelos Animais de Doenças , Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Masculino , Camundongos , Fosforilação , Neovascularização Retiniana/tratamento farmacológico , Neovascularização Retiniana/etiologia , Neovascularização Retiniana/patologia , Transdução de Sinais/efeitos dos fármacos
9.
Biomol Ther (Seoul) ; 28(1): 98-103, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31284709

RESUMO

Marfan syndrome (MFS), a connective tissue disorder caused by mutations in the fibrillin-1 (Fbn1) gene, has vascular manifestations including aortic aneurysm, dissection, and rupture. Its vascular pathogenesis is assumed to be attributed to increased transforming growth factor ß (TGFß) signaling and blockade of excessive TGFß signaling has been thought to prevent dissection and aneurysm formation. Here, we investigated whether galunisertib, a potent small-molecule inhibitor of TGFß receptor I (TßRI), attenuates aneurysmal disease in a murine model of MFS (Fbn1C1039G/+) and compared the impact of galuninsertib on the MFS-related vascular pathogenesis with that of losartan, a prophylactic agent routinely used for patients with MFS. Fbn1C1039G/+ mice were administered galunisertib or losartan for 8 weeks, and their ascending aortas were assessed for histopathological changes and phosphorylation of Smad2 and extracellular signal-regulated kinase 1/2 (Erk1/2). Mice treated with galunisertib or losartan barely exhibited phosphorylated Smad2, suggesting that both drugs effectively blocked overactivated canonical TGFß signaling in Fbn1C1039G/+ mice. However, galunisertib treatment did not attenuate disrupted medial wall architecture and only partially decreased Erk1/2 phosphorylation, whereas losartan significantly inhibited MFS-associated aortopathy and markedly decreased Erk1/2 phosphorylation in Fbn1C1039G/+ mice. These data unexpectedly revealed that galunisertib, a TßRI inhibitor, showed no benefits in aneurysmal disease in MFS mice although it completely blocked Smad2 phosphorylation. The significant losartaninduced inhibition of both aortic vascular pathogenesis and Smad2 phosphorylation implied that canonical TGFß signaling might not prominently drive aneurysmal diseases in MFS mice.

10.
Arterioscler Thromb Vasc Biol ; 39(10): 2120-2131, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31434494

RESUMO

OBJECTIVE: Aberrant neovascularization is a leading cause of blindness in several eye diseases, including age-related macular degeneration and proliferative diabetic retinopathy. The identification of key regulators of pathological ocular neovascularization has been a subject of extensive research and great therapeutic interest. Here, we explored the previously unrecognized role of cKIT and its ligand, SCF (stem cell factor), in the pathological ocular neovascularization process. Approach and Results: Compared with normoxia, hypoxia, a crucial driver of neovascularization, caused cKIT to be highly upregulated in endothelial cells, which significantly enhanced the angiogenic response of endothelial cells to SCF. In murine models of pathological ocular neovascularization, such as oxygen-induced retinopathy and laser-induced choroidal neovascularization models, cKIT and SCF expression was significantly increased in ocular tissues, and blockade of cKIT and SCF using cKit mutant mice and anti-SCF neutralizing IgG substantially suppressed pathological ocular neovascularization. Mechanistically, SCF/cKIT signaling induced neovascularization through phosphorylation of glycogen synthase kinase-3ß and enhancement of the nuclear translocation of ß-catenin and the transcription of ß-catenin target genes related to angiogenesis. Inhibition of ß-catenin-mediated transcription using chemical inhibitors blocked SCF-induced in vitro angiogenesis in hypoxia, and injection of a ß-catenin agonist into cKit mutant mice with oxygen-induced retinopathy significantly enhanced pathological neovascularization in the retina. Conclusions; Our data reveal that SCF and cKIT are promising novel therapeutic targets for treating vision-threatening ocular neovascular diseases.


Assuntos
Regulação da Expressão Gênica , Doenças Retinianas/genética , Doenças Retinianas/metabolismo , Neovascularização Retiniana/genética , Fator de Células-Tronco/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Análise de Variância , Inibidores da Angiogênese/farmacologia , Animais , Células Cultivadas , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Hipóxia/complicações , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-kit/genética , Doenças Retinianas/patologia , Doenças Retinianas/fisiopatologia , Transdução de Sinais/genética
11.
Mol Med Rep ; 19(4): 3061-3070, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30816523

RESUMO

The Src kinase family (SKF) includes non­receptor tyrosine kinases that interact with many cellular cytosolic, nuclear and membrane proteins, and is involved in the progression of cellular transformation and oncogenic activity. However, there is little to no evidence on the effect of SKF or its inhibitors on melanogenesis. Therefore, the present study investigated whether C­terminal Src kinase inhibition can induce melanogenesis and examined the associated signaling pathways and mRNA expression of melanogenic proteins. First, whether stimulators of melanogenesis, such as ultraviolet B and α­melanocyte­stimulating hormone, can dephosphorylate Src protein was evaluated, and the results revealed that SU6656 and PP2 inhibited the phosphorylation of Src in G361 cells. Src inhibition by these chemical inhibitors induced melanogenesis in G361 cells and upregulated the mRNA expression levels of melanogenesis­associated genes encoding microphthalmia­associated transcription factor, tyrosinase­related protein 1 (TRP1), TRP2, and tyrosinase. In addition, Src inhibition by small interfering RNA induced melanogenesis and upregulated the mRNA expression levels of melanogenesis­associated genes. As the p38 mitogen­activated protein kinase (MAPK) and cyclic adenosine monophosphate response element binding (CREB) pathways serve key roles in melanogenesis, the present study further examined whether Src mediates melanogenesis via these pathways. As expected, Src inhibition via SU6656 or PP2 administration induced the phosphorylation of p38 or CREB, as determined by western blotting analysis, and increased the levels of phosphorylated p38 or CREB, as determined by immunofluorescence staining. In addition, the induced pigmentation and melanin content of G361 cells by Src inhibitors was significantly inhibited by p38 or CREB inhibitors. Taken together, these data indicate that Src is associated with melanogenesis, and Src inhibition induces melanogenesis via the MAPK and CREB pathways in G361 cells.


Assuntos
Transformação Celular Neoplásica/metabolismo , Melanoma/etiologia , Melanoma/metabolismo , Quinases da Família src/antagonistas & inibidores , Biomarcadores , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Humanos , Indóis/farmacologia , Melaninas/biossíntese , Melanoma/patologia , Fosforilação , Pirimidinas/farmacologia , RNA Interferente Pequeno/genética , Sulfonamidas/farmacologia , Raios Ultravioleta , alfa-MSH/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Quinases da Família src/genética , Quinases da Família src/metabolismo
12.
Sci Rep ; 9(1): 1656, 2019 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-30733541

RESUMO

Peroxisome proliferator-activated receptor (PPAR)-α/γ dual agonists have been developed to treat metabolic diseases; however, most of them exhibit side effects such as body weight gain and oedema. Therefore, we developed a novel PPARα/γ dual agonist that modulates glucose and lipid metabolism without adverse effects. We synthesised novel compounds composed of coumarine and chalcone, determined their crystal structures, and then examined their binding affinity toward PPARα/γ. We investigated the expression of PPARα and PPARγ target genes by chemicals in HepG2, differentiated 3T3-L1, and C2C12 cells. We examined the effect of chemicals on glucose and lipid metabolism in db/db mice. Only MD001 functions as a PPARα/γ dual agonist in vitro. MD001 increased the transcriptional activity of PPARα and PPARγ, resulting in enhanced expression of genes related to ß-oxidation and fatty acid and glucose uptake. MD001 significantly improved blood metabolic parameters, including triglycerides, free fatty acids, and glucose, in db/db mice. In addition, MD001 ameliorated hepatic steatosis by stimulating ß-oxidation in vitro and in vivo. Our results demonstrated the beneficial effects of the novel compound MD001 on glucose and lipid metabolism as a PPARα/γ dual agonist. Consequently, MD001 may show potential as a novel drug candidate for the treatment of metabolic disorders.


Assuntos
Chalconas/farmacologia , Cumarínicos/farmacologia , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Glucose/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , PPAR alfa/agonistas , PPAR gama/agonistas , Animais , Chalconas/química , Cumarínicos/química , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/patologia , Células HEK293 , Células Hep G2 , Humanos , Hipoglicemiantes/química , Hipoglicemiantes/farmacologia , Resistência à Insulina , Masculino , Camundongos , Camundongos Endogâmicos C57BL
13.
Biomol Ther (Seoul) ; 26(3): 306-312, 2018 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-29129045

RESUMO

In a previous study, we have demonstrated that S-methylmethionine sulfonium (SMMS) confers wound-healing and photoprotective effects on the skin, suggesting that SMMS can be used as a cosmetic raw material. However, it has an unpleasant odor. Therefore, in the present study, we synthesized odor-free SMMS derivatives by eliminating dimethyl sulfide, which is the cause of the unpleasant odor and identified two derivatives that exhibited skin-protective effects: one derivative comprised (2S,4S)- and (2R,4S)-2-phenylthiazolidine-4-carboxylic acid and the other comprised (2S,4R)-, (2S,4S)-, (2R,4R)-, and (2R,4S)-2-phenyl-1,3-thiazinane-4-carboxylic acid. We performed in vitro proliferation assays using human dermal fibroblasts (hDFs) and an immortalized human keratinocyte cell line (HaCaT). The two SMMS derivatives were shown to increase hDF and HaCaT cell proliferation as well as improve their survival by protecting against ultraviolet exposure. Moreover, the derivatives regulated the expression of collagen type I and MMP mRNAs against ultraviolet exposure in hDFs, suggesting that these derivatives can be developed as cosmetic raw materials.

14.
Mol Vis ; 23: 823-831, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29225458

RESUMO

Purpose: Vascular endothelial growth factor (VEGF) is a principal mediator of pathological ocular neovascularization, which is the leading cause of blindness in various ocular diseases. As Src, a non-receptor tyrosine kinase, has been implicated as one of the major signaling molecules in VEGF-mediated neovascularization, the present study aimed to investigate whether dasatinib, a potent Src kinase inhibitor, could suppress pathological ocular neovascularization in murine models of oxygen-induced retinopathy (OIR) and choroidal neovascularization (CNV). Methods: Tube formation, scratch wounding migration, and cell proliferation assays were performed to measure the inhibitory effect of dasatinib on VEGF-induced angiogenesis in human retinal microvascular endothelial cells. Murine models of OIR and laser-induced CNV were used to assess the preventive effect of an intravitreal injection of dasatinib on pathological neovascularization in the retina and choroid. Neovascularization and Src phosphorylation were evaluated with immunofluorescence staining. Results: Dasatinib efficiently inhibited VEGF-induced endothelial proliferation, wounding migration, and tube formation. In mice with OIR and laser injury-induced CNV, eyes treated with a single intravitreal injection of dasatinib exhibited significant decreases in pathological neovascularization compared with that of controls injected with vehicle. The dasatinib-treated OIR mice also showed a decrease in Src phosphorylation in the periretinal tufts. The intravitreal injection of dasatinib did not cause ocular toxicity at the treatment dose administered. Conclusions: These results demonstrated that dasatinib suppressed pathological neovascularization in the mouse retina and choroid. Therefore, dasatinib may be indicated for the treatment of ischemia-induced proliferative retinopathy and neovascular age-related macular degeneration.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Neovascularização de Coroide/tratamento farmacológico , Dasatinibe/uso terapêutico , Inibidores de Proteínas Quinases/uso terapêutico , Neovascularização Retiniana/tratamento farmacológico , Animais , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Neovascularização de Coroide/etiologia , Neovascularização de Coroide/metabolismo , Modelos Animais de Doenças , Endotélio Vascular/efeitos dos fármacos , Feminino , Angiofluoresceinografia , Humanos , Marcação In Situ das Extremidades Cortadas , Injeções Intravítreas , Fotocoagulação a Laser/efeitos adversos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Oxigênio/toxicidade , Fosforilação , Neovascularização Retiniana/induzido quimicamente , Neovascularização Retiniana/metabolismo , Vasos Retinianos/citologia , Fator A de Crescimento do Endotélio Vascular/efeitos adversos
15.
Oncotarget ; 8(55): 94805-94818, 2017 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-29212268

RESUMO

Adipose tissue is considered to be an endocrine organ, and adipocyte size correlates with insulin resistance and metabolic parameters in obesity. There is little data on the effects of angiopoietin-1 in adipose tissue and kidney in streptozotocin (STZ)-induced diabetes. In this study, we investigated the protective effect of COMP-angiopoietin-1 (COMP-Ang1), a potent variant of angiopoietin-1, on vascular endothelial cells in epididymal adipose tissue and its regulatory effect on other metabolic parameters, such as lipid droplet diameter, macrophage infiltration, and renal inflammation in STZ-treated mice. Our data showed that COMP-Ang1 increased the density of platelet endothelial cell adhesion molecule-1 (PECAM-1)-1-positive vascular endothelial cells in adipose tissue, which were significantly decreased by treatment with STZ. COMP-Ang1 ameliorated the STZ-induced decrease in lipid droplet diameter and increase in macrophage infiltration in adipose tissue. Serum free fatty acid and triglyceride levels were decreased after administration of COMP-Ang1. There was a beneficial effect on serum insulin levels after treatment with COMP-Ang1 in STZ-induced diabetic mice. Fasting blood glucose levels in COMP-Ang1-treated mice were significantly lower than those of LacZ-treated mice. Cotreatment with COMP-Ang1 and STZ also had similar effects on the above parameters. Administration of soluble Tie2, an inhibitor of angiopoietin-1, reversed the effects of COMP-Ang1. COMP-Ang1 was found to ameliorate the up-regulation of proinflammatory molecules and F4/80-positive macrophage infiltration in the kidneys of STZ-treated mice. COMP-Ang1 increased the phosphorylation of Akt in epididymal adipose tissue and kidneys of STZ-induced diabetic mice. These data indicate that COMP-Ang1 regulates lipogenic effects in adipose tissue and renal inflammation in STZ-induced diabetic mice.

16.
Int J Nanomedicine ; 12: 4813-4822, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28740387

RESUMO

Pathological angiogenesis is one of the major symptoms of severe ocular diseases, including corneal neovascularization. The blockade of vascular endothelial growth factor (VEGF) action has been recognized as an efficient strategy for treating corneal neovascularization. In this study, we aimed to investigate whether nanoparticle-based delivery of apatinib, a novel and selective inhibitor of VEGF receptor 2, inhibits VEGF-mediated angiogenesis and suppresses experimental corneal neovascularization. Water-insoluble apatinib was encapsulated in nanoparticles composed of human serum albumin (HSA)-conjugated polyethylene glycol (PEG). In vitro angiogenesis assays showed that apatinib-loaded HSA-PEG (Apa-HSA-PEG) nanoparticles potently inhibited VEGF-induced tube formation, scratch wounding migration, and proliferation of human endothelial cells. In a rat model of alkali burn injury-induced corneal neovascularization, a subconjunctival injection of Apa-HSA-PEG nanoparticles induced a significant decrease in neovascularization compared to that observed with an injection of free apatinib solution or phosphate-buffered saline. An in vivo distribution study using HSA-PEG nanoparticles loaded with fluorescent hydrophobic model drugs revealed the presence of a substantial number of nanoparticles in the corneal stroma within 24 h after injection. These in vitro and in vivo results demonstrate that apatinib-loaded nanoparticles may be promising for the prevention and treatment of corneal neovascularization-related ocular disorders.


Assuntos
Indutores da Angiogênese/administração & dosagem , Neovascularização da Córnea/tratamento farmacológico , Nanopartículas/administração & dosagem , Neovascularização Patológica/tratamento farmacológico , Piridinas/administração & dosagem , Indutores da Angiogênese/farmacologia , Animais , Modelos Animais de Doenças , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Nanopartículas/química , Polietilenoglicóis/química , Piridinas/farmacologia , Ratos Sprague-Dawley , Albumina Sérica/química , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores
17.
Invest Ophthalmol Vis Sci ; 58(9): 3592-3599, 2017 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-28715845

RESUMO

Purpose: Vascular endothelial growth factor (VEGF) signaling via VEGF receptor 2 (VEGFR2) plays a crucial role in pathologic ocular neovascularization. In this study, we investigated the antiangiogenic effect of apatinib, a pharmacologic inhibitor of VEGFR2 tyrosine kinase, against oxygen-induced retinopathy (OIR) and laser-induced choroidal neovascularization (CNV) in mice. Methods: Western blotting and in vitro angiogenesis assays were performed using human retinal microvascular endothelial cells (HRMECs). OIR was induced in neonatal mice by exposure to 75% oxygen from postnatal day (P) 7 to P12 and to room air from P12 to P17. Experimental CNV was induced in mice using laser photocoagulation. Apatinib was intravitreally and orally administered to mice. Neovascularization and phosphorylation of VEGFR2 were evaluated by immunofluorescence staining. Results: Apatinib inhibited VEGF-mediated activation of VEGFR2 signaling and substantially reduced VEGF-induced proliferation, migration, and cord formation in HRMECs. A single intravitreal injection of apatinib significantly attenuated retinal or choroidal neovascularization in mice with OIR or laser injury-induced CNV, respectively. Retinal or choroidal tissues of the eyes treated with apatinib exhibited substantially lower phosphorylation of VEGFR2 than those of controls injected with vehicle. Intravitreal injection of apatinib did not cause noticeable ocular toxicity. Moreover, oral administration of apatinib significantly reduced laser-induced CNV in mice. Conclusions: Our study demonstrates that apatinib inhibits pathologic ocular neovascularization in mice with OIR or laser-induced CNV. Apatinib may, therefore, be a promising drug for the prevention and treatment of ischemia-induced proliferative retinopathy and neovascular age-related macular degeneration.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Neovascularização de Coroide/tratamento farmacológico , Modelos Animais de Doenças , Piridinas/uso terapêutico , Neovascularização Retiniana/tratamento farmacológico , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Administração Oral , Animais , Animais Recém-Nascidos , Western Blotting , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Neovascularização de Coroide/metabolismo , Neovascularização de Coroide/patologia , Células Endoteliais/efeitos dos fármacos , Feminino , Angiofluoresceinografia , Técnica Indireta de Fluorescência para Anticorpo , Injeções Intravítreas , Fotocoagulação a Laser , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Oxigênio/toxicidade , Fosforilação , Gravidez , Neovascularização Retiniana/metabolismo , Neovascularização Retiniana/patologia , Vasos Retinianos/citologia , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
18.
Arch Pharm Res ; 40(2): 197-203, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-27988882

RESUMO

Src kinase signaling is important in the regulation of microvascular barrier function and endothelial hyperpermeability. This study was designed to evaluate the protective effect of dasatinib, a potent Src inhibitor used clinically for the treatment of cancer, against the breakdown of the blood-retinal barrier (BRB) and the retinal vascular leakage caused by vascular endothelial growth factor (VEGF) and diabetes. We examined the effects of dasatinib on VEGF-induced endothelial hyperpermeability and the loss of vascular endothelial (VE)-cadherin, an endothelial junctional protein. Dasatinib inhibited VEGF-induced phosphorylation of Src in human retinal microvascular endothelial cells (HRMECs). In vitro and in vivo vascular permeability assays showed that dasatinib blocked the VEGF-enhanced hyperpermeability of HRMECs and decreased VEGF-mediated retinal vascular leakage in mice. Immunofluorescent staining of VE-cadherin showed that dasatinib abolished the junctional disappearance of VE-cadherin in VEGF-treated HRMECs and murine retinal vasculature. In addition, we examined the protective effect of dasatinib against diabetes-induced retinal vascular leakage in streptozotocin-induced diabetic rats. An intravitreal injection of dasatinib substantially inhibited the development of hyperpermeable retinal vasculature. Our results indicate that dasatinib is a promising agent for the prevention and treatment of diabetes-induced retinal vascular leakage.


Assuntos
Barreira Hematorretiniana/efeitos dos fármacos , Permeabilidade Capilar/efeitos dos fármacos , Dasatinibe/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Quinases da Família src/antagonistas & inibidores , Animais , Antígenos CD/metabolismo , Caderinas/metabolismo , Dasatinibe/administração & dosagem , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/complicações , Retinopatia Diabética/tratamento farmacológico , Retinopatia Diabética/metabolismo , Células Endoteliais/efeitos dos fármacos , Humanos , Injeções Intravítreas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação , Inibidores de Proteínas Quinases/administração & dosagem , Ratos , Ratos Sprague-Dawley , Retina/metabolismo , Vasos Retinianos/efeitos dos fármacos , Transdução de Sinais , Estreptozocina/toxicidade , Quinases da Família src/metabolismo
19.
Arch Pharm Res ; 40(1): 1-12, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27921262

RESUMO

In 2006, Shinya Yamanaka first reported that in vitro reprogramming of somatic cells toward pluripotency was achieved by simple induction of specific transcription factors. Induced pluripotent stem cell (iPSC) technology has since revolutionized the ways in which we explore the mechanisms of human diseases and develop therapeutics. Here, I describe the recent advances in human iPSC-based disease modeling and drug discovery and discuss the current challenges. Additionally, I outline potential future applications of human iPSCs in classifying patients based on their response to drugs in clinical trials and elucidating optimal patient-specific therapeutic strategies, which will contribute to reduced attrition rates and the development of precision medicine.


Assuntos
Descoberta de Drogas/tendências , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/patologia , Modelos Biológicos , Preparações Farmacêuticas/metabolismo , Medicina de Precisão/tendências , Animais , Reprogramação Celular/efeitos dos fármacos , Reprogramação Celular/fisiologia , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Preparações Farmacêuticas/administração & dosagem , Medicina de Precisão/métodos
20.
Mol Med Rep ; 14(5): 4706-4714, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27748854

RESUMO

It has previously been demonstrated that hypoxia has diverse stimulatory effects on adipose­derived stem cells (ASCs), however, metabolic responses under hypoxia remain to be elucidated. Thus, the present study aimed to investigate the glucose uptake and metabolism of ASCs under hypoxic conditions, and to identify the underlying molecular mechanisms. ASCs were cultured in 1% oxygen, and experiments were conducted in vitro. As determined by proteomic analysis and western blotting, GAPDH and enolase 1 (ENO1) expression were upregulated under hypoxia. In addition, lactate production was significantly increased, and mRNA levels of glycolytic enzymes, including GAPDH, ENO1, hexokinase 2 (HK2), and lactate dehydrogenase α (LDHα) were upregulated. Hypoxia­inducible factor 1­α (HIF­1α) expression was increased as demonstrated by western blotting, and a pharmacological inhibitor of HIF­1α significantly attenuated hypoxia­induced lactate production and expression of glycolytic enzymes. It was also observed that hypoxia significantly increased glucose uptake in ASCs, and glucose transporter (GLUT)1 and GLUT3 expression were upregulated under hypoxia. Pharmacological inhibition of the HIF­1α signaling pathways also attenuated hypoxia­induced GLUT1 and GLUT3 expression. These results collectively indicate that hypoxia increases glucose uptake via GLUT1 and GLUT3 upregulation, and induces lactate production of ASCs via GAPDH, ENO1, HK2, and LDHα. Furthermore, HIF­1α is involved in glucose uptake and metabolism of ASCs.


Assuntos
Tecido Adiposo/citologia , Tecido Adiposo/metabolismo , Glucose/metabolismo , Hipóxia/metabolismo , Células-Tronco/metabolismo , Feminino , Regulação Enzimológica da Expressão Gênica , Proteínas Facilitadoras de Transporte de Glucose/genética , Glicólise , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Ácido Láctico/biossíntese , Proteoma , Proteômica/métodos
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