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1.
FASEB J ; 37(2): e22776, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-36688817

RESUMO

AQP5 plays a crucial role in maintaining corneal transparency and the barrier function of the cornea. Here, we found that in the corneas of Aqp5-/- mice at older than 6 months, loss of AQP5 significantly increased corneal neovascularization, inflammatory cell infiltration, and corneal haze. The results of immunofluorescence staining showed that upregulation of K1, K10, and K14, and downregulation of K12 and Pax6 were detected in Aqp5-/- cornea and primary corneal epithelial cells. Loss of AQP5 aggravated wound-induced corneal neovascularization, inflammation, and haze. mRNA sequencing, western blotting, and qRT-PCR showed that Wnt2 and Wnt6 were significantly decreased in Aqp5-/- corneas and primary corneal epithelial cells, accompanied by decreased aggregation in the cytoplasm and nucleus of ß-catenin. IIIC3 significantly suppressed corneal neovascularization, inflammation, haze, and maintained corneal transparent epithelial in Aqp5-/- corneas. We also found that pre-stimulated Aqp5-/- primary corneal epithelial cells with IIIC3 caused the decreased expression of K1, K10, and K14, the increased expression of K12, Pax6, and increased aggregation in the cytoplasm and nucleus of ß-catenin. These findings revealed that AQP5 may regulate corneal epithelial homeostasis and function through the Wnt/ß-catenin signaling pathway. Together, we uncovered a possible role of AQP5 in determining corneal epithelial cell fate and providing a potential therapeutic target for corneal epithelial dysfunction.


Assuntos
Neovascularização da Córnea , Via de Sinalização Wnt , Camundongos , Animais , Aquaporina 5/genética , Neovascularização da Córnea/metabolismo , beta Catenina/metabolismo , Córnea/metabolismo , Células Epiteliais/metabolismo , Inflamação/metabolismo
2.
Mol Cell Biochem ; 2024 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-38556592

RESUMO

Ferroptosis is a newly discovered type of regulated cell death participated in multiple diseases. Different from other classical cell death programs such as necrosis and apoptosis, ferroptosis involving iron-catalyzed lipid peroxidation is characterized by Fe2+ accumulation and mitochondria alterations. The phenomenon of oxidative stress following organ ischemia-reperfusion (I/R) has recently garnered attention for its connection to the onset of ferroptosis and subsequent reperfusion injuries. This article provides a comprehensive overview underlying the mechanisms of ferroptosis, with a further focus on the latest research progress regarding interference with ferroptotic pathways in organ I/R injuries, such as intestine, lung, heart, kidney, liver, and brain. Understanding the links between ferroptosis and I/R injury may inform potential therapeutic strategies and targeted agents.

3.
Biochem Biophys Res Commun ; 680: 184-193, 2023 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-37742347

RESUMO

Cataract is lens opacity, which is a common blinding eye disease worldwide. Aquaporin 5 (AQP5) is expressed in the human and mouse lenses. This study aimed to investigate the underlying mechanisms of AQP5 in the senescence of lens epithelial cells (LECs). Primary LECs were isolated and cultured from Aqp5+/+ and Aqp5-/- mice. Western blot or immunofluorescence staining of p16, Ki67, MitoSOX, JC-1 and phalloidin was used in the experiments to evaluate the changes in the primary LECs. The primary Aqp5-/- LECs showed increased p16 expression and mitochondrial reactive oxygen species, decreased mitochondrial membrane potential and activity, and cytoskeletal disorders. When the cells were pretreated with Mito-TEMPO, the Aqp5-/- mice showed decreased p16 expression, reduced mitochondrial dysfunction and cytoskeletal disorders. Our results revealed that AQP5 deficiency promotes the senescence of primary LECs through mitochondrial dysfunction. This provides a new perspective for the treatment of cataracts by regulating AQP5 expression.

4.
Exp Eye Res ; 233: 109557, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37380095

RESUMO

As a water channel protein, aquaporin 5 (AQP5) is essential for the maintenance of the normal physiological functions of ocular tissues. This review provides an overview of the expression and function of AQP5 in the eye and discusses their role in related eye diseases. Although AQP5 plays a vital role in ocular functions, such as maintaining corneal and lens transparency, regulating water movement, and maintaining homeostasis, some of its functions in ocular tissues are still unclear. Based on the key role of AQP5 in eye function, this review suggests that in the future, eye diseases may be treated by regulating the expression of aquaporin.


Assuntos
Oftalmopatias , Cristalino , Humanos , Aquaporina 5 , Córnea/metabolismo , Cristalino/metabolismo , Oftalmopatias/metabolismo , Fenômenos Fisiológicos Oculares
5.
FASEB J ; 36(10): e22553, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-36111980

RESUMO

Mesenchymal stromal cells (MSCs) are attractive candidates for treating hepatic disorders given their potential to enhance liver regeneration and function. The paracrine paradigm may be involved in the mechanism of MSC-based therapy, and exosomes (Exo) play an important role in this paracrine activity. Hypoxia significantly improves the effectiveness of MSC transplantation. However, whether hypoxia preconditioned MSCs (Hp-MSCs) can enhance liver regeneration, and whether this enhancement is mediated by Exo, are unknown. In this study, mouse bone marrow-derived MSCs (BM-MSCs) and secreted Exo were injected through the tail vein. We report that Hp-MSCs promote liver regeneration after partial hepatectomy in mice through their secreted exosomes. Interestingly, MSC-Exo were concentrated in liver 6 h after administration and mainly taken up by macrophages, but not hepatocytes. Compared with normoxic MSC-Exo (N-Exo), hypoxic MSC-Exo (Hp-Exo) enhanced M2 macrophage polarization both in vivo and in vitro. Microarray analysis revealed significant enrichment of microRNA (miR)-182-5p in Hp-Exo compared with that in N-Exo. In addition, miR-182-5p knockdown partially abolished the beneficial effect of Hp-Exo. Finally, Hp-MSC-derived exosomal miR-182-5p inhibited theprotein expression of forkhead box transcription factor 1 (FOXO1) in macrophages, which inhibited toll-like receptor 4 (TLR4) expression and subsequently induced an anti-inflammatory response. These results highlight the therapeutic potential of Hp-Exo in liver regeneration and suggest that miR-182-5p from Hp-Exo facilitates macrophage polarization during liver regeneration by modulating the FOXO1/TLR4 signaling pathway.


Assuntos
Regeneração Hepática , Macrófagos , Células-Tronco Mesenquimais , MicroRNAs , Animais , Medula Óssea/metabolismo , Proteína Forkhead Box O1/genética , Proteína Forkhead Box O1/metabolismo , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Hipóxia/metabolismo , Regeneração Hepática/genética , Macrófagos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Camundongos , MicroRNAs/metabolismo , Receptor 4 Toll-Like/metabolismo
6.
Int J Mol Sci ; 24(4)2023 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-36834481

RESUMO

Acute liver injury (ALI) is a globally important public health issue that, when severe, rapidly progresses to acute liver failure, seriously compromising the life safety of patients. The pathogenesis of ALI is defined by massive cell death in the liver, which triggers a cascade of immune responses. Studies have shown that the aberrant activation of the nod-like receptor protein 3 (NLRP3) inflammasome plays an important role in various types of ALI and that the activation of the NLRP3 inflammasome causes various types of programmed cell death (PCD), and these cell death effectors can in turn regulate NLRP3 inflammasome activation. This indicates that NLRP3 inflammasome activation is inextricably linked to PCD. In this review, we summarize the role of NLRP3 inflammasome activation and PCD in various types of ALI (APAP, liver ischemia reperfusion, CCl4, alcohol, Con A, and LPS/D-GalN induced ALI) and analyze the underlying mechanisms to provide references for future relevant studies.


Assuntos
Lesão Pulmonar Aguda , Inflamassomos , Humanos , Lesão Pulmonar Aguda/metabolismo , Apoptose , Inflamassomos/metabolismo , Lipopolissacarídeos , Fígado/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteínas NLR
7.
Am J Pathol ; 191(11): 1974-1985, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34390680

RESUMO

Aquaporins (AQPs) are a highly conserved group of membrane proteins that play critical roles in water and small solute transport across epithelial and endothelial barriers. The aim of this study was to determine whether AQP5, a well-known water channel protein, also plays a role in corneal epithelial wound healing. AQP5 knockout (AQP5-/-) mice were generated using CRISPR/Cas9. A corneal wound healing model was established using epithelial debridement of corneas. The time to corneal epithelial and nerve regeneration was significantly delayed in the AQP5-/- mice. Reduction in Ki-67-positive cells and nerve growth factor (NGF) expression was confirmed in the AQP5-/- mice during healing. Epithelial and nerve regeneration rates were significantly increased in the AQP5-/- mice after treatment with NGF, which was accompanied by recovered levels of phosphorylated Akt. NGF treatment also improved the recovery of corneal nerve fiber density and sensitivity in the AQP5-/- mice. While the promotion of NGF induced corneal epithelial and nerve regeneration, Akt inhibitor reversed Akt reactivation. A significant impairment of corneal wound healing in the AQP5-/- mice resulted from distinct defects in corneal epithelial cell proliferation and nerve regeneration. These results provide evidence for the involvement of aquaporin in cell proliferation and suggest that AQP5 induction could be a potential therapy for accelerating the resurfacing of corneal defects.


Assuntos
Aquaporina 5/metabolismo , Epitélio Corneano/metabolismo , Regeneração Nervosa/fisiologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Cicatrização/fisiologia , Animais , Proliferação de Células/fisiologia , Córnea , Lesões da Córnea/metabolismo , Lesões da Córnea/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais/fisiologia
8.
Toxicol Appl Pharmacol ; 437: 115889, 2022 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-35065992

RESUMO

Quercetin (Que) exhibits excellent biological activity; however, its clinical development is hindered owing to the poor water solubility. In this study, Que. was loaded on polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer (PVCL-PVA-PEG, Soluplus) micelles through a thin-film hydration process, and their tumor angiogenesis inhibition ability was investigated. The particle size of Soluplus-Que micelles was 55.3 ± 1.8 nm, and the micelles stayed stability within 9 months. Soluplus-Que micelles can enhance the cell uptake of Que. and transport the micelles to intracellular lysosomes and mitochondria. The MTT assay results revealed that Soluplus-Que micelles enhanced the cytotoxicity of Que. on HUVEC cells. Furthermore, Soluplus-Que micelles inhibited migration and invasion of HUVEC cells, as well as inhibited the neovascularization of chick embryo allantoic membrane (CAM). The in vivo study revealed that Soluplus-Que micelles significantly inhibit the growth of H22 solid tumors, with low toxic side effects. Soluplus-Que inhibited the expression of CD31 (a marker of angiogenesis) and the PI3K/Akt/VEGF pathway in tumor tissues, indicating its potential to hold back tumor growth via the inhibition of angiogenesis. Our findings indicated that as a delivery system, Soluplus micelles demonstrate potential for the delivery of poorly soluble drugs for tumor treatment.


Assuntos
Micelas , Neovascularização Patológica/prevenção & controle , Fosfatidilinositol 3-Quinases/metabolismo , Polietilenoglicóis/química , Polímeros/química , Polivinil/química , Quercetina/farmacologia , Inibidores da Angiogênese , Animais , Movimento Celular/efeitos dos fármacos , Embrião de Galinha , Sistemas de Liberação de Medicamentos/métodos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana , Humanos , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Quercetina/química , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo
9.
J Pharmacol Sci ; 148(1): 19-30, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34924126

RESUMO

Hepatic ischemia/reperfusion (I/R) injury contributes to morbidity and mortality during liver resection or transplantation, with limited effective treatments available. Here, we investigated the potential benefits and underlying mechanisms of pterostilbene (Pt), a natural component of blueberries and grapes, in preventing hepatic I/R injury. Male C57BL/6 mice subjected to partial warm hepatic I/R and human hepatocyte cell line L02 cells exposed to anoxia/reoxygenation (A/R) were used as in vivo and in vitro models, respectively. Our findings showed that pretreatment with Pt ameliorated hepatic I/R injury by improving liver histology, decreasing hepatocyte apoptosis, and reducing plasma ALT and AST levels. Likewise, cell apoptosis, mitochondrial membrane dysfunction, and mitochondrial ROS overproduction in L02 cells triggered by the A/R challenge in vitro were reduced due to Pt administration. Mechanistically, Pt treatment efficiently enhanced mitophagy and upregulated PINK1, Parkin, and LC3B expression. Notably, the protective effect of Pt was largely abrogated after cells were transfected with PINK1 siRNA. Moreover, Pt pretreatment promoted hepatocyte proliferation and liver regeneration in the late phase of hepatic I/R. In conclusion, our findings provide evidence that Pt exerts hepatoprotective effects in hepatic I/R injury by upregulating PINK1-mediated mitophagy.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Infarto Hepático/genética , Infarto Hepático/prevenção & controle , Mitofagia/efeitos dos fármacos , Mitofagia/genética , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/prevenção & controle , Estilbenos/farmacologia , Estilbenos/uso terapêutico , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Hepatócitos/fisiologia , Humanos , Regeneração Hepática/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética
10.
Mol Vis ; 27: 679-690, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-35002213

RESUMO

Purpose: AQP5-/- mice spontaneously exhibit dry eye symptoms. The purpose of this study was to assess the endoplasmic reticulum (ER) stress-mediated inflammation generated by a deficiency of aquaporin 5 (AQP5) in the lacrimal gland. Methods: Hematoxylin and eosin (H&E) staining, Oil Red O staining, and transmission electron microscopy (TEM) analysis were performed to identify structural changes in lacrimal gland epithelial cells because of AQP5 deficiency. Corneal epithelial defects were assessed with sodium fluorescein staining. The expression profiles of mRNA and proteins were determined by quantitative real-time reverse transcription PCR (qRT-PCR) and western blot. Mice in the quercetin group were injected intraperitoneally with 40 mg/kg of quercetin, and the control group was injected with an equal volume of dimethyl sulfoxide (DMSO) for 4 weeks. Results: Aqueous tear secretion fell at about 50% in 1- and 6-month-old AQP5-/- mice compared with that of AQP5+/+ mice. TEM showed that the ER structure was damaged. ER stress was significantly increased in the lacrimal gland of AQP5-/- mice. Lipid droplets accumulated in the matrix and acinar cells, and changes occurred in the lipid metabolism and gene expression levels for PPARα, CPT1α, and CPT2 in the AQP5-/- mice. Immune cell infiltration and increases in the gene expression levels of the chemokines CXCL1, CXCL2, and CCL5 were found in the lacrimal gland of AQP5-/- mice. Quercetin partially reversed ER stress levels, inflammation, and lipid accumulation, and it inhibited tear secretion. Conclusions: The study data indicated that a deficiency of AQP5 induced pathophysiological changes and functional decompensation of the lacrimal gland. Quercetin may improve the inflammation in the lacrimal glands of AQP5-/- mice by regulating the ER stress levels.


Assuntos
Aquaporina 5 , Estresse do Retículo Endoplasmático , Aparelho Lacrimal , Animais , Aquaporina 5/genética , Homeostase , Inflamação/genética , Camundongos , Camundongos Knockout , Lágrimas
11.
Toxicol Appl Pharmacol ; 419: 115511, 2021 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-33819459

RESUMO

Hesperetin (HSP) has excellent biological activities with poor water solubility which limits its clinical development. In this study, we successfully prepared a novel, self-assembled micelle based on Rebaudioside A (RA) for oral delivery of HSP with improved bioavailability and therapeutic effects. We found that RA and HSP could be formylated into nanomicelles with particle sizes of 4.541 nm ± 0.048 nm. HSP was readily encapsulated into RA micelles and this improved its water solubility (to 12.74 mg/mL ± 0.28 mg/mL). The MTT results showed that RA-HSP enhanced the cytotoxicity, the clonal formation inhibitory activity, and cell migration inhibitory activity of HSP in human breast cancer MDA-MB-231 cells. The mechanism results showed that RA-HSP induced cell apoptosis by inducing the production of reactive oxygen species (ROS), destroying the mitochondrial membrane potential (MMP), and inhibiting the PI3K/Akt signaling pathway. Moreover, RA-HSP enhanced the anticancer activity, increased the oral bioavailability and tissue distribution of HSP in vivo. Moreover, the mechanism studies in vivo found that HSP inhibited PI3K/Akt signaling pathway with low side effects. These findings indicate that RA micelle formulations have great potential in oral drug delivery systems for the delivery of hydrophobic drugs.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Diterpenos do Tipo Caurano/química , Portadores de Fármacos , Hesperidina/farmacologia , Nanopartículas , Administração Oral , Animais , Antineoplásicos Fitogênicos/administração & dosagem , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacocinética , Apoptose/efeitos dos fármacos , Disponibilidade Biológica , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Composição de Medicamentos , Liberação Controlada de Fármacos , Feminino , Hesperidina/administração & dosagem , Hesperidina/química , Hesperidina/farmacocinética , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Micelas , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Solubilidade , Distribuição Tecidual , Carga Tumoral/efeitos dos fármacos
12.
Exp Eye Res ; 205: 108485, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33582182

RESUMO

The pathogenesis of congenital cataract (CC), a major disease associated with blindness in infants, is complex and diverse. Aquaporin 5 (AQP5) represents an essential membrane water channel. In the present study, whole exome sequencing revealed a novel heterozygous missense mutation of AQP5 (c.152 T > C, p. L51P) in the four generations of the autosomal dominant CC (adCC) family. By constructing a mouse model of AQP5 knockout (KO) using the CRISPR/Cas9 technology, we observed that the lens of AQP5-KO mice showed mild opacity at approximately six months of age. miR-124-3p.1 expression was identified to be downregulated in the lens of AQP5-KO mice as evidenced by qRT-PCR analysis. A dual luciferase reporter assay confirmed that vimentin was a target gene of miR-124-3p.1. Organ-cultured AQP5-KO mouse lenses were showed increased opacity compared to those of WT mice, and vimentin expression was upregulated as determined by RT-PCR, western blotting, and immunofluorescence staining. After miR-124-3p.1 agomir was added, the lens opacity in WT mice and AQP5-KO mice decreased, accompanied by the downregulation of vimentin. AQP5-L51P increased vimentin expression of in human lens epithelial cells. Therefore, a missense mutation in AQP5 (c.152 T > C, p. L51P) was associated with adCC, and AQP5 could participate in the maintenance of lens transparency by regulating vimentin expression via miR-124-3p.1.


Assuntos
Aquaporina 5/fisiologia , Catarata/prevenção & controle , Regulação da Expressão Gênica/fisiologia , Cristalino/metabolismo , MicroRNAs/genética , Vimentina/genética , Adulto , Idoso , Animais , Western Blotting , Catarata/genética , Catarata/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Linhagem , Reação em Cadeia da Polimerase em Tempo Real
13.
Biochem Biophys Res Commun ; 519(2): 396-401, 2019 11 05.
Artigo em Inglês | MEDLINE | ID: mdl-31519322

RESUMO

Annexin A1 (AnxA1) has been shown to exert potent anti-inflammatory and anti-fibrotic activities in a range of systemic inflammatory disorders. Corneal scarring is characterized by myofibroblast differentiation and disorganized extracellular matrix deposition. This study was aim to explore the potential therapeutic properties of Ac2-26, a mimetic peptide of AnnexinA1 (AnxA1), on TGF-ß induced human corneal myofibroblast differentiation and mechanical injury-induced mouse corneal haze. The results found that Ac2-26 treatment dose dependently reduced α-SMA level and other fibrogenic gene expressions in HTK cells stimulated by exogenous TGF-ß1. While this anti-fibrotic effect was abolished by an FPR2/ALX inhibitor WRW4. In mice, topical Ac2-26 application suppressed the development of corneal scarring, inhibited myofibroblast differentiation, while promoted the corneal epithelial wound healing. Moreover, Ac2-26 treatment inhibited Ly6G + neutrophil infiltration and reduced corneal inflammatory response. The results provided in vivo and in vitro supports the anti-fibrotic and anti-inflammatory effects of AnxA1 derived peptide Ac2-26, and suggest that AnxA1 mimetic agents might be a promising strategy for the treatment of corneal scarring.


Assuntos
Anexina A1/farmacologia , Lesões da Córnea/tratamento farmacológico , Inflamação/tratamento farmacológico , Peptídeos/farmacologia , Estresse Mecânico , Animais , Diferenciação Celular/efeitos dos fármacos , Lesões da Córnea/metabolismo , Lesões da Córnea/patologia , Inflamação/metabolismo , Inflamação/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL
14.
Mol Vis ; 24: 274-285, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29643724

RESUMO

Purpose: To investigate the effect and mechanism of proresolving lipid mediator resolvin D1 (RvD1) on the corneal epithelium and the restoration of mechanical sensation in diabetic mice. Methods: Type 1 diabetes was induced in mice with intraperitoneal streptozocin injections. The healthy and diabetic mice underwent removal of the central corneal epithelium, and then 100 ng/ml RvD1 or its formyl peptide receptor 2 (FPR2) antagonist WRW4 was used to treat the diabetic mice. Regeneration of the corneal epithelium and nerves was observed with sodium fluorescein staining and whole-mount anti-ß3-tubulin fluorescence staining. The inflammatory response level was measured with hematoxylin and eosin staining (inflammatory cell infiltration), enzyme-linked immunosorbent assay (tumor necrosis factor alpha and interleukin-1 beta content), myeloperoxidase activity, and fluorescence staining (macrophage content). The reactive oxygen species (ROS) and glutathione (GSH) levels were examined with incubation with fluorescent probes, and oxidative stress-related protein expression levels were evaluated with fluorescence staining and western blotting. Results: Topical application of RvD1 promoted regeneration of the corneal epithelium in diabetic mice, accompanied by the reactivation of signaling and inflammation resolution related to regeneration of the epithelium. Furthermore, RvD1 directly attenuated the accumulation of ROS and nicotinamide adenine dinucleotide phosphate oxidase 2/4 expression, while RvD1 enhanced GSH synthesis and reactivated the Nrf2-ARE signaling pathway that was impaired in the corneal epithelium in the diabetic mice. More interestingly, topical application of RvD1 promoted regeneration of corneal nerves and completely restored impaired mechanical sensitivity of the cornea in diabetic mice. In addition, the promotion of corneal epithelial wound healing by RvD1 in diabetic mice was abolished by its FPR2 antagonist WRW4. Conclusions: Topical application of RvD1 promotes corneal epithelial wound healing and the restoration of mechanical sensation in diabetic mice, which may be related to the lipid mediator's regulation of inflammation resolution, the reactivation of regenerative signaling in the epithelium, and the attenuation of oxidative stress.


Assuntos
Lesões da Córnea/tratamento farmacológico , Diabetes Mellitus Experimental/tratamento farmacológico , Ácidos Docosa-Hexaenoicos/farmacologia , Receptores de Formil Peptídeo/genética , Regeneração/efeitos dos fármacos , Percepção do Tato/efeitos dos fármacos , Cicatrização/efeitos dos fármacos , Administração Tópica , Animais , Hidrolases de Éster Carboxílico/genética , Hidrolases de Éster Carboxílico/metabolismo , Lesões da Córnea/complicações , Lesões da Córnea/genética , Lesões da Córnea/patologia , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patologia , Ácidos Docosa-Hexaenoicos/antagonistas & inibidores , Epitélio Corneano/efeitos dos fármacos , Epitélio Corneano/metabolismo , Epitélio Corneano/patologia , Regulação da Expressão Gênica , Glutationa/agonistas , Glutationa/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NADH NADPH Oxirredutases/genética , NADH NADPH Oxirredutases/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Oligopeptídeos/farmacologia , Nervo Óptico/efeitos dos fármacos , Nervo Óptico/metabolismo , Peroxidase/genética , Peroxidase/metabolismo , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Receptores de Formil Peptídeo/antagonistas & inibidores , Receptores de Formil Peptídeo/metabolismo , Estreptozocina , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
15.
Exp Eye Res ; 167: 122-127, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29246498

RESUMO

MicroRNA-204 (miR-204) is highly expressed in cornea, here we explored the role and mechanism of miR-204 in corneal neovascularization (CNV). Mouse CNV was induced by intrastromal placement of suture in BALB/c mice with the subconjunctival injection of miR-204 agomir or negative control. Human primary limbal epithelial cells (LECs) and immortalized microvascular endothelial cells (HMECs) were used to evaluate the expression changes and anti-angiogenic effects of miR-204 under biomechanical stress (BS). The expression and localization of miR-204, vascular endothelial growth factor (VEGF) and their receptors were detected by quantitative real-time PCR, in situ hybridization, immunohistochemistry and Western blot. The results showed that miR-204 expression was mainly localized in epithelium and down-expressed in vascularized cornea. Subconjunctival injection of miR-204 agomir inhibited CNV and reduced the expression of VEGF and VEGF receptor 2. Similarly, miR-204 overexpression attenuated the increased expression of VEGF by biomechanical stress in LECs, and suppressed the proliferation, migration, and tube formation of HMECs. These novel findings indicate that epithelium-derived miR-204 inhibits suture-induced CNV through regulating VEGF and VEGF receptor 2.


Assuntos
Neovascularização da Córnea/prevenção & controle , Modelos Animais de Doenças , Epitélio Corneano/metabolismo , MicroRNAs/fisiologia , Animais , Vasos Sanguíneos/efeitos dos fármacos , Western Blotting , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Neovascularização da Córnea/metabolismo , Neovascularização da Córnea/patologia , Células Endoteliais/metabolismo , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos BALB C , MicroRNAs/farmacologia , Reação em Cadeia da Polimerase em Tempo Real , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
16.
Stem Cells ; 33(5): 1566-76, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25546438

RESUMO

Ciliary neurotrophic factor (CNTF), a well-known neuroprotective cytokine, has been found to play an important role in neurogenesis and functional regulations of neural stem cells. As one of the most innervated tissue, however, the role of CNTF in cornea epithelium remains unclear. This study was to explore the roles and mechanisms of CNTF in the activation of corneal epithelial stem/progenitor cells and wound healing of both normal and diabetic mouse corneal epithelium. In mice subjecting to mechanical removal of corneal epithelium, the corneal epithelial stem/progenitor cell activation and wound healing were promoted by exogenous CNTF application, while delayed by CNTF neutralizing antibody. In cultured corneal epithelial stem/progenitor cells, CNTF enhanced the colony-forming efficiency, stimulated the mitogenic proliferation, and upregulated the expression levels of corneal epithelial stem/progenitor cell-associated transcription factors. Furthermore, the promotion of CNTF on the corneal epithelial stem/progenitor cell activation and wound healing was mediated by the activation of STAT3. Moreover, in diabetic mice, the content of CNTF in corneal epithelium decreased significantly when compared with that of normal mice, and the supplement of CNTF promoted the diabetic corneal epithelial wound healing, accompanied with the advanced activation of corneal epithelial stem/progenitor cells and the regeneration of corneal nerve fibers. Thus, the capability of expanding corneal epithelial stem/progenitor cells and promoting corneal epithelial wound healing and nerve regeneration indicates the potential application of CNTF in ameliorating limbal stem cell deficiency and treating diabetic keratopathy.


Assuntos
Fator Neurotrófico Ciliar/farmacologia , Epitélio Corneano/patologia , Células-Tronco/citologia , Cicatrização/efeitos dos fármacos , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Diabetes Mellitus Experimental/patologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Epitélio Corneano/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator de Transcrição STAT3/metabolismo , Células-Tronco/efeitos dos fármacos
17.
Exp Eye Res ; 123: 37-42, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24746620

RESUMO

γ-secretase inhibitor has been shown to promote intestinal goblet cell differentiation. We now demonstrated that the in vitro addition of γ-secretase inhibitor in the culture of human conjunctival epithelial cells significantly promoted the differentiation of conjunctival goblet cells with typical droplet-like phenotype, positive periodic acid-Schiff and goblet cell-specific Muc5Ac, cytokeratin 7 and Helix pomatia agglutinin lectin staining. Moreover, topical application of γ-secretase inhibitor promoted the differentiation of mouse conjunctival goblet cells in vivo. Furthermore, the expression of Notch target gene HES-1 was down-regulated during the differentiation of conjunctival goblet cells. In addition, we found that the recombinant conjunctival epithelium on amniotic membrane showed less goblet cell density and abnormal location when compared with normal conjunctival epithelium, which were improved by the addition of γ-secretase inhibitor in the final induction.


Assuntos
Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Diferenciação Celular/fisiologia , Túnica Conjuntiva/citologia , Dipeptídeos/farmacologia , Inibidores Enzimáticos/farmacologia , Células Caliciformes/citologia , Âmnio , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Western Blotting , Contagem de Células , Células Cultivadas , Túnica Conjuntiva/metabolismo , Técnica Indireta de Fluorescência para Anticorpo , Células Caliciformes/metabolismo , Proteínas de Homeodomínio/metabolismo , Humanos , Queratina-7/genética , Queratina-7/metabolismo , Lectinas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mucina-5AC/metabolismo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição HES-1
18.
Invest Ophthalmol Vis Sci ; 65(8): 25, 2024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-39017635

RESUMO

Purpose: Abnormalities in aquaporins are implicated in the pathological progression of dry eye syndrome. Retinoic acid (RA) regulates cellular proliferation, differentiation, and apoptosis in the cornea, thereby being associated with dry eye disease (DED). The objective of this study is to explore the underlying mechanisms responsible for RA metabolic abnormalities in corneas lacking aquaporin 5 (AQP5). Methods: Dry eye (DE) models were induced via subcutaneous scopolamine hydrobromide. Aqp5 knockout (Aqp5-/-) mice and DE mice were utilized to assess corneal epithelial alterations. Tear secretion, goblet cell counts, and corneal punctate defects were evaluated. The impact of Aqp5 on RA-related enzymes and receptors was investigated using pharmacological RA or SR (A JunB inhibitor), a transcription factor JunB inhibitor, treatment in mouse corneal epithelial cells (CECs), or human corneal epithelial cells (HCECs). The HCECs and NaCl-treated HCECs underwent quantitative real-time PCR (qRT-PCR), immunofluorescent, Western blot, and TUNEL assays. The regulation of transcription factor JunB on Aldh1a1 was explored via ChIP-PCR. Results: Aqp5 and Aldh1a1 were reduced in both CECs of DE mice and NaCl-induced HCECs. Aqp5-/- mice exhibited DE phenotype and reduced Aldh1a1. RA treatment reduced apoptosis, promoted proliferation, and improved the DE phenotype in Aqp5-/- mice. JunB enrichment in the Aldh1a1 promoter was identified by ChIP-PCR. SR significantly increased Aldh1a1 expression, Ki67, and ΔNp63-positive cells, and decreased TUNEL-positive cells in CECs and HCECs. Conclusions: Our findings demonstrated the downregulation of Aqp5 expression and aberrant RA metabolism in DE conditions. Knockout of Aqp5 resulted in reduced production of RA through activation of JunB, subsequently leading to the manifestation of DE symptoms.


Assuntos
Apoptose , Aquaporina 5 , Modelos Animais de Doenças , Síndromes do Olho Seco , Camundongos Knockout , Tretinoína , Animais , Aquaporina 5/genética , Aquaporina 5/biossíntese , Aquaporina 5/metabolismo , Síndromes do Olho Seco/metabolismo , Síndromes do Olho Seco/patologia , Síndromes do Olho Seco/genética , Camundongos , Tretinoína/farmacologia , Epitélio Corneano/metabolismo , Epitélio Corneano/patologia , Reação em Cadeia da Polimerase em Tempo Real , Camundongos Endogâmicos C57BL , Western Blotting , Humanos , Células Cultivadas , Lágrimas/metabolismo , Marcação In Situ das Extremidades Cortadas , Regulação da Expressão Gênica , Proliferação de Células
19.
Mol Med Rep ; 29(4)2024 04.
Artigo em Inglês | MEDLINE | ID: mdl-38391117

RESUMO

Acetaminophen (APAP) overdose is the primary cause of drug­induced acute liver failure in numerous Western countries. NLR family pyrin domain containing 3 (NLRP3) inflammasome activation serves a pivotal role in the pathogenesis of various forms of acute liver injury. However, the cellular source for NLRP3 induction and its involvement during APAP­induced hepatotoxicity have not been thoroughly investigated. In the present study, hematoxylin and eosin staining was performed to assess histopathological changes of liver tissue. Immunohistochemistry staining(NLRP3, Caspase­1, IL­1ß, GSDMD and Caspase­3), western blotting (NLRP3, Caspase­1, IL­1ß, GSDMD and Caspase­3) and RT­qPCR (NLRP3, Caspase­1 and IL­1ß) were performed to assess the expression of NLRP3/GSDMD signaling pathway. TUNEL staining was performed to assess apoptosis of liver tissue. The serum expression levels of inflammatory factors (IL­6, IL­18, IL­1ß and TNF­α) were assessed using ELISA and inflammation of liver tissue was assessed using immunohistochemistry (Ly6G and CD68) and RT­qPCR (TNF­α, Il­6, Mcp­1, Cxcl­1, Cxcl­2). A Cell Counting Kit­8 was performed to assess cell viability and apoptosis. Protein and gene expression were analyzed by western blotting (PCNA, CCND1) and RT­qPCR (CyclinA2, CyclinD1 and CyclinE1). Through investigation of an APAP­induced acute liver injury model (AILI), the present study demonstrated that APAP overdose induced activation of NLRP3 and cleavage of gasdermin D (GSDMD) in hepatocytes, both in vivo and in vitro. Additionally, mice with hepatocyte­specific knockout of Nlrp3 exhibited reduced liver injury and lower mortality following APAP intervention, accompanied by decreased infiltration of inflammatory cells and attenuated inflammatory response. Furthermore, pharmacological blockade of NLRP3/GSDMD signaling using MCC950 or disulfiram significantly ameliorated liver injury and reduced hepatocyte death. Notably, hepatocyte Nlrp3 deficiency promoted liver recovery by enhancing hepatocyte proliferation. Collectively, the present study demonstrated that inhibition of the NLRP3 inflammasome protects against APAP­induced acute liver injury by reducing hepatocyte pyroptosis and suggests that targeting NLRP3 may hold therapeutic potential for treating AILI.


Assuntos
Doença Hepática Crônica Induzida por Substâncias e Drogas , Proteína 3 que Contém Domínio de Pirina da Família NLR , Camundongos , Animais , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Inflamassomos/metabolismo , Acetaminofen/efeitos adversos , Piroptose , Caspase 3 , Fator de Necrose Tumoral alfa , Interleucina-6 , Hepatócitos/metabolismo
20.
Biochem Biophys Res Commun ; 430(2): 598-603, 2013 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-23219815

RESUMO

Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is a member of the immunoglobulin super family and has been observed to have two paradoxical functions: tumor suppression and the promotion of tumor invasion. In the present study, we discovered that CEACAM1 functions as an adhesion inhibitor and a migration promoter. The CEACAM1 transfected cells, either 293-CEACAM1 or LOVO/trans-CEACAM1, was proved to have lower adhesion rate. Furthermore, HT29/siRNA-CEACAM1 cells had a higher adhesion rate than HT29 cells. These results indicated that CEACAM1 was an inhibitor of cell-matrix adhesion. Additionally, 293-CEACAM1 LOVO/trans-CEACAM1 cells exhibited better motility in a trans-well migration assay. N-cadherin expression levels were positively correlated with CEACAM1 in 293-CEACAM1, LOVO/trans-CEACAM1 and HT29/siRNA-CEACAM1 cells. When blocked by a GC-4 antibody, the adhesive capacities of 293-CEACAM1 and LOVO/trans-CEACAM1 were recovered and the motilities of them were suppressed, which suggested that CEACAM1 functioned through N-cadherin.


Assuntos
Antígenos CD/metabolismo , Caderinas/biossíntese , Moléculas de Adesão Celular/metabolismo , Movimento Celular , Metástase Neoplásica/patologia , Antígenos CD/genética , Adesão Celular , Moléculas de Adesão Celular/genética , Células HEK293 , Células HT29 , Humanos , Modelos Biológicos
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