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1.
Am J Physiol Cell Physiol ; 326(5): C1293-C1307, 2024 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-38525543

RESUMEN

Given the widespread application of glucocorticoids in ophthalmology, the associated elevation of intraocular pressure (IOP) has long been a vexing concern for clinicians, yet the underlying mechanisms remain inconclusive. Much of the discussion focuses on the extracellular matrix (ECM) of trabecular meshwork (TM). It is widely agreed that glucocorticoids impact the expression of matrix metalloproteinases (MMPs), leading to ECM deposition. Since Zn2+ is vital for MMPs, we explored its role in ECM alterations induced by dexamethasone (DEX). Our study revealed that in human TM cells treated with DEX, the level of intracellular Zn2+ significantly decreased, accompanied by impaired extracellular Zn2+ uptake. This correlated with changes in several Zrt-, Irt-related proteins (ZIPs) and metallothionein. ZIP8 knockdown impaired extracellular Zn2+ uptake, but Zn2+ chelation did not affect ZIP8 expression. Resembling DEX's effects, chelation of Zn2+ decreased MMP2 expression, increased the deposition of ECM proteins, and induced structural disarray of ECM. Conversely, supplementation of exogenous Zn2+ in DEX-treated cells ameliorated these outcomes. Notably, dietary zinc supplementation in mice significantly reduced DEX-induced IOP elevation and collagen content in TM, thereby rescuing the visual function of the mice. These findings underscore zinc's pivotal role in ECM regulation, providing a novel perspective on the pathogenesis of glaucoma.NEW & NOTEWORTHY Our study explores zinc's pivotal role in mitigating extracellular matrix dysregulation in the trabecular meshwork and glucocorticoid-induced ocular hypertension. We found that in human trabecular meshwork cells treated with dexamethasone, intracellular Zn2+ significantly decreased, accompanied by impaired extracellular Zn2+ uptake. Zinc supplementation rescues visual function by modulating extracellular matrix proteins and lowering intraocular pressure, offering a direction for further exploration in glaucoma management.


Asunto(s)
Glaucoma , Malla Trabecular , Ratones , Humanos , Animales , Malla Trabecular/metabolismo , Dexametasona/farmacología , Glucocorticoides/farmacología , Glaucoma/patología , Presión Intraocular , Proteínas de la Matriz Extracelular/metabolismo , Matriz Extracelular/metabolismo , Metaloproteinasas de la Matriz/metabolismo , Zinc/metabolismo , Células Cultivadas
2.
J Neuroinflammation ; 20(1): 91, 2023 Apr 07.
Artículo en Inglés | MEDLINE | ID: mdl-37029422

RESUMEN

BACKGROUND: Retinal ischemia-reperfusion (RIR) injury refers to an obstruction in the retinal blood supply followed by reperfusion. Although the molecular mechanism underlying the ischemic pathological cascade is not fully understood, neuroinflammation plays a crucial part in the mortality of retinal ganglion cells. METHODS: Single-cell RNA sequencing (scRNA-seq), molecular docking, and transfection assay were used to explore the effectiveness and pathogenesis of N,N-dimethyl-3ß-hydroxycholenamide (DMHCA)-treated mice with RIR injury and DMHCA-treated microglia after oxygen and glucose deprivation/reoxygenation (OGD/R). RESULTS: DMHCA could suppress inflammatory gene expression and attenuate neuronal lesions, restoring the retinal structure in vivo. Using scRNA-seq on the retina of DMHCA-treated mice, we provided novel insights into RIR immunity and demonstrated nerve injury-induced protein 1 (Ninjurin1/Ninj 1) as a promising treatment target for RIR. Moreover, the expression of Ninj1, which was increased in RIR injury and OGD/R-treated microglia, was downregulated in the DMHCA-treated group. DMHCA suppressed the activation of the nuclear factor kappa B (NF-κB) pathways induced by OGD/R, which was undermined by the NF-κB pathway agonist betulinic acid. Overexpressed Ninj1 reversed the anti-inflammatory and anti-apoptotic function of DMHCA. Molecular docking indicated that for Ninj1, DMHCA had a low binding energy of - 6.6 kcal/mol, suggesting highly stable binding. CONCLUSION: Ninj1 may play a pivotal role in microglia-mediated inflammation, while DMHCA could be a potential treatment strategy against RIR injury.


Asunto(s)
FN-kappa B , Daño por Reperfusión , Ratones , Animales , FN-kappa B/metabolismo , Transducción de Señal , Simulación del Acoplamiento Molecular , Oxígeno , Células Ganglionares de la Retina/patología , Daño por Reperfusión/metabolismo , Inflamación/tratamiento farmacológico , Factores de Crecimiento Nervioso , Moléculas de Adhesión Celular Neuronal
3.
Free Radic Biol Med ; 212: 415-432, 2024 02 20.
Artículo en Inglés | MEDLINE | ID: mdl-38134974

RESUMEN

The molecular mechanism of how reduced mobile zinc (Zn2+) affected retinal ganglion cell (RGC) survival and optic nerve regeneration after optic nerve crush (ONC) injury remains unclear. Here, we used conditionally knocked out ZnT-3 in the amacrine cells (ACs) of mice (CKO) in order to explore the role of reactive oxygen species (ROS), nuclear factor erythroid 2-related factor 2 (NFE2L2, Nrf2) and autophagy in the protection of RGCs and axon regeneration after ONC injury. We found that reduced Zn2+ can promote RGC survival and axonal regeneration by decreasing ROS, activating Nrf2, and inhibiting autophagy. Additionally, autophagy after ONC is regulated by ROS and Nrf2. Visual function in mice after ONC injury was partially recovered through the reduction of Zn2+, achieved by using a Zn2+ specific chelator N,N,N',N'-tetrakis-(2-Pyridylmethyl) ethylenediamine (TPEN) or through CKO mice. Overall, our data reveal the crosstalk between Zn2+, ROS, Nrf2 and autophagy following ONC injury. This study verified that TPEN or knocking out ZnT-3 in ACs is a promising therapeutic option for the treatment of optic nerve damage and elucidated the postsynaptic molecular mechanism of Zn2+-triggered damage to RGCs after ONC injury.


Asunto(s)
Etilenodiaminas , Traumatismos del Nervio Óptico , Células Ganglionares de la Retina , Ratones , Animales , Factor 2 Relacionado con NF-E2/genética , Especies Reactivas de Oxígeno , Axones/fisiología , Regeneración Nerviosa , Traumatismos del Nervio Óptico/tratamiento farmacológico , Traumatismos del Nervio Óptico/genética , Zinc , Modelos Animales de Enfermedad
4.
Sci Adv ; 10(31): eado0866, 2024 Aug 02.
Artículo en Inglés | MEDLINE | ID: mdl-39093964

RESUMEN

As part of the central nervous system, the optic nerve, composed of axons from retinal ganglion cells (RGCs), generally fails to regenerate on its own when injured in adult mammals. An innovative approach to promoting optic nerve regeneration involves manipulating the interactions between amacrine cells (ACs) and RGCs. Here, we identified a unique AC subtype, dopaminergic ACs (DACs), that responded early after optic nerve crush by down-regulating neuronal activity and reducing retinal dopamine (DA) release. Activating DACs or augmenting DA release with levodopa demonstrated neuroprotective effects and modestly enhanced axon regeneration. Within this context, we pinpointed the DA receptor D1 (DRD1) as a critical mediator of DAC-derived DA and showed that RGC-specific Drd1 overexpression effectively overcame subtype-specific barriers to regeneration. This strategy markedly boosted RGC survival and axon regeneration after crush and preserved vision in a glaucoma model. This study unveils the crucial role of DAC-derived DA signaling in optic nerve regeneration, holding promise for therapeutic insights into neural repair.


Asunto(s)
Células Amacrinas , Dopamina , Regeneración Nerviosa , Nervio Óptico , Células Ganglionares de la Retina , Transducción de Señal , Animales , Células Amacrinas/metabolismo , Dopamina/metabolismo , Regeneración Nerviosa/efectos de los fármacos , Nervio Óptico/metabolismo , Células Ganglionares de la Retina/metabolismo , Células Ganglionares de la Retina/efectos de los fármacos , Traumatismos del Nervio Óptico/metabolismo , Traumatismos del Nervio Óptico/tratamiento farmacológico , Traumatismos del Nervio Óptico/patología , Ratones , Axones/metabolismo , Axones/fisiología , Receptores de Dopamina D1/metabolismo , Visión Ocular/fisiología , Modelos Animales de Enfermedad
5.
Transl Vis Sci Technol ; 12(11): 21, 2023 11 01.
Artículo en Inglés | MEDLINE | ID: mdl-37975842

RESUMEN

Purpose: Trabecular meshwork (TM) fibrosis is a crucial pathophysiological process in the development of primary open-angle glaucoma. Pirfenidone (PFD) is a new, broad-spectrum antifibrotic agent approved for the treatment of idiopathic pulmonary fibrosis. This study investigated the inhibitory effect of PFD on TM fibrosis and evaluated its efficacy in lowering intraocular pressure (IOP). Methods: Human TM cells were isolated, cultured, and characterized. Cell Counting Kit-8 was used to evaluate the proliferation and toxicity of different concentrations of PFD on normal or fibrotic TM cells. TM cells were treated with transforming growth factor beta-2 (TGF-ß2) in the absence or presence of PFD. Western blotting and immunofluorescence analyses were used to analyze changes in the TM cell cytoskeleton and extracellular matrix (ECM) proteins, including alpha-smooth muscle actin (α-SMA), F-actin, collagen IV (COL IV), and fibronectin (FN). An ocular hypertension (OHT) mouse model was induced with Ad-TGF-ß2C226/228S and then treated with PFD or latanoprost (LT) eye drops to confirm the efficacy of PFD in lowering IOP. Results: PFD inhibited the proliferation of fibrotic TM cells in a dose-dependent manner and inhibited TGF-ß2-induced overexpression of α-SMA, COL IV, and FN in TM cells. PFD stabilized F-actin. In vivo, PFD eye drops reduced the IOP of the OHT models and showed no significant difference compared with LT eye drops. Conclusions: PFD inhibited TGF-ß2-induced TM cell fibrosis by rearranging the disordered cytoskeleton and decreasing ECM deposition, thereby enhancing the aqueous outflow from the TM outflow pathway and lowering IOP, which provides a potential new approach to treating glaucoma. Translational Relevance: Our work with pirfenidone provides a new approach to treat glaucoma.


Asunto(s)
Glaucoma de Ángulo Abierto , Glaucoma , Hipertensión Ocular , Animales , Humanos , Ratones , Actinas/metabolismo , Células Cultivadas , Fibrosis , Glaucoma de Ángulo Abierto/tratamiento farmacológico , Soluciones Oftálmicas/farmacología , Malla Trabecular/metabolismo , Malla Trabecular/patología , Factor de Crecimiento Transformador beta2/farmacología
6.
Neural Regen Res ; 18(12): 2773-2780, 2023 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-37449644

RESUMEN

Vision depends on accurate signal conduction from the retina to the brain through the optic nerve, an important part of the central nervous system that consists of bundles of axons originating from retinal ganglion cells. The mammalian optic nerve, an important part of the central nervous system, cannot regenerate once it is injured, leading to permanent vision loss. To date, there is no clinical treatment that can regenerate the optic nerve and restore vision. Our previous study found that the mobile zinc (Zn2+) level increased rapidly after optic nerve injury in the retina, specifically in the vesicles of the inner plexiform layer. Furthermore, chelating Zn2+ significantly promoted axonal regeneration with a long-term effect. In this study, we conditionally knocked out zinc transporter 3 (ZnT3) in amacrine cells or retinal ganglion cells to construct two transgenic mouse lines (VGATCreZnT3fl/fl and VGLUT2CreZnT3fl/fl, respectively). We obtained direct evidence that the rapidly increased mobile Zn2+ in response to injury was from amacrine cells. We also found that selective deletion of ZnT3 in amacrine cells promoted retinal ganglion cell survival and axonal regeneration after optic nerve crush injury, improved retinal ganglion cell function, and promoted vision recovery. Sequencing analysis of reginal ganglion cells revealed that inhibiting the release of presynaptic Zn2+ affected the transcription of key genes related to the survival of retinal ganglion cells in postsynaptic neurons, regulated the synaptic connection between amacrine cells and retinal ganglion cells, and affected the fate of retinal ganglion cells. These results suggest that amacrine cells release Zn2+ to trigger transcriptomic changes related to neuronal growth and survival in reginal ganglion cells, thereby influencing the synaptic plasticity of retinal networks. These results make the theory of zinc-dependent retinal ganglion cell death more accurate and complete and provide new insights into the complex interactions between retinal cell networks.

7.
Antioxidants (Basel) ; 11(10)2022 Oct 10.
Artículo en Inglés | MEDLINE | ID: mdl-36290724

RESUMEN

Retinal ganglion cells (RGCs), the projection neurons of the eye, are irreversibly lost once the optic nerve is injured, which is a critical mechanism of glaucoma. Mobile zinc (Zn2+) levels rapidly increase in retinal interneuron amacrine cells and Zn2+ is then transferred to RGCs via the Zn2+ transporter protein ZnT-3, triggering RGC loss in optic nerve injury. Zn2+ chelation and ZnT-3 deletion promote long-term RGC survival. However, the downstream signaling pathways of Zn2+ in RGCs remains unknown. Here, we show that increased levels of Zn2+ upregulate the expression and activity of mitochondrial zinc metallopeptidase OMA1 in the retina, leading to the cleavage of DELE1 and activation of cytosolic eIF2α kinase PKR, triggering the integrated stress response (ISR) in RGCs. Our study identified OMA1 and ISR as the downstream molecular mechanisms of retinal Zn2+ and potential targets for preventing the progression of Zn2+-associated neuronal damage.

8.
Drug Deliv ; 28(1): 634-641, 2021 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-33779455

RESUMEN

To increase the amount of pirfenidone (PFD) loaded in polyvinyl alcohol (PVA) film embedded soft contact lens (SCL), and evaluate its function of sustaining delivery of drug in vitro and in vivo. Drug loading efficiency within PVA film and SCLs, drug release from SCLs in vitro, and the effects of parameters of SCLs and external environment on drug release in vitro were evaluated by ultraviolet-visible spectrophotometer at 312 nm. Safety of SCLs was evaluated in vitro by transformed human corneal epithelial cell. Safety in vivo was determined by optical coherence tomography and histology of anterior segment of rabbits. Drug release study in tear fluid and aqueous humor were measured by ultra-performance liquid chromatography. SCLs had smooth surface and were fit for experimental rabbits. Amount of PFD in PVA film and SCLs were 153.515 µg ± 12.508 and 127.438 µg ± 19.674, respectively, PFD in PVA film was significantly higher than SCLs (p=.006) and closed to 150 µg (targeting amount of PFD to be loaded). Thickness of SCLs, molecular weight of PVA, and amount of PVA used in SCLs affected drug release in vitro significantly. Thickness of PVA film and amount of drug in SCLs had no effect on drug release rate in vitro. SCLs were safe in vitro and in vivo, PFD released from SCLs could be detected around 12 hours in tears and aqueous humor, and the concentration of drug was higher than eye drop at all detected time points while amount of PFD in SCLs was lower than eye drop. Drug loaded PVA film embedded SCLs may be a promising ocular drug delivery system.


Asunto(s)
Antiinflamatorios no Esteroideos/administración & dosificación , Lentes de Contacto Hidrofílicos , Sistemas de Liberación de Medicamentos/métodos , Alcohol Polivinílico/química , Piridonas/administración & dosificación , Animales , Antiinflamatorios no Esteroideos/farmacología , Química Farmacéutica , Preparaciones de Acción Retardada , Liberación de Fármacos , Células Epiteliales , Humanos , Hidrogeles/química , Piridonas/farmacología , Conejos , Lágrimas/química
9.
J Ocul Pharmacol Ther ; 37(2): 75-83, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33297836

RESUMEN

Purpose: The aim of this study was to fabricate pirfenidone (PFD)-loaded soft contact lenses (SCLs), explore their characteristics, and evaluate their efficiency on extended delivery of PFD in vitro and in vivo. Methods: PFD-loaded SCLs were fabricated by embedding an insert of PFD and polyvinyl alcohol (PVA) into 2 layers of silicone elastomer. The optical transparency, water content, and protein deposition were measured. Transformed human corneal epithelial cells were used to test the cytotoxicity of SCLs. The release rate of PFD by SCLs in vitro was evaluated by an ultraviolet-visible spectrophotometer. Toxicity of SCLs was assessed by inspection of ocular surface irritation in rabbits before and after contact lens wear. The concentrations of PFD in tears and aqueous humor of rabbits' eyes as a function of time were determined by high-performance liquid chromatography for SCLs and 30 µL of 0.5% PFD eye drops. Results: SCLs possessed good light transmittance. Blank SCLs had poor water content (0.548% ± 0.330), and an improved water content was found in PVA film-loaded SCLs (11.022% ± 1.508, P = 0.010). No lysozyme and human serum albumin were found in SCLs. There was no significant toxicity of SCLs in vitro and in vivo. SCLs prolonged the residence time of PFD in tears and aqueous humor of rabbit eyes by 5 times compared with the eye drop instillation while around 1/10 of the eye drop dosage was loaded in SCLs. Conclusions: PFD-loaded SCLs can significantly prolong the residence time of PFD and may be a promising ocular drug delivery system.


Asunto(s)
Lentes de Contacto Hidrofílicos , Sistemas de Liberación de Medicamentos , Soluciones Oftálmicas/química , Piridonas/química , Animales , Femenino , Humanos , Conejos
10.
Exp Ther Med ; 15(4): 3385-3391, 2018 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-29545859

RESUMEN

Hedyotis diffusa Willd (HDW) is a constituent of several Chinese medicines used clinically to treat inflammatory diseases, including airway inflammation. The aim of the present study was to investigate whether HDW serves a protective role in suppressing chronic airway inflammation and its underlying mechanisms. A mouse model of chronic smoking was induced via exposure to cigarette smoke (CS) for 30 days, increasing the exposure time for up to 5 min per day and the administration of lipopolysaccharide (LPS). Mice were gavaged with HDW (50 or 100 mg/kg body weight), dexamethasone (1 mg/kg body weight) or normal saline (NS, 0.9%) 1 h prior to CS challenge. Compared with CS and LPS (SL)-induced mice, the levels of interleukin (IL)-1ß, tumor necrosis factor-α and transforming growth factor-ß in bronchoalveolar lavage fluid from HDW+SL mice were significantly decreased and IL-10 was markedly reduced. Histological examination of the lung tissues revealed that HDW treatment alleviates airway inflammation. In addition, the administration of HDW to human bronchial epithelial BEAS-2B cells suppressed the activity of the nuclear factor (NF)-κB signaling pathway. The results of the present study demonstrate that HDW has a therapeutic effect in COPD and the underlying mechanism may be attributed to inhibition of the NF-κB pathway.

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