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1.
Nucleic Acids Res ; 52(5): 2212-2230, 2024 Mar 21.
Artículo en Inglés | MEDLINE | ID: mdl-38364871

RESUMEN

Nonreference sequences (NRSs) are DNA sequences present in global populations but absent in the current human reference genome. However, the extent and functional significance of NRSs in the human genomes and populations remains unclear. Here, we de novo assembled 539 genomes from five genetically divergent human populations using long-read sequencing technology, resulting in the identification of 5.1 million NRSs. These were merged into 45284 unique NRSs, with 29.7% being novel discoveries. Among these NRSs, 38.7% were common across the five populations, and 35.6% were population specific. The use of a graph-based pangenome approach allowed for the detection of 565 transcript expression quantitative trait loci on NRSs, with 426 of these being novel findings. Moreover, 26 NRS candidates displayed evidence of adaptive selection within human populations. Genes situated in close proximity to or intersecting with these candidates may be associated with metabolism and type 2 diabetes. Genome-wide association studies revealed 14 NRSs to be significantly associated with eight phenotypes. Additionally, 154 NRSs were found to be in strong linkage disequilibrium with 258 phenotype-associated SNPs in the GWAS catalogue. Our work expands the understanding of human NRSs and provides novel insights into their functions, facilitating evolutionary and biomedical researches.


Asunto(s)
Genoma Humano , Estudio de Asociación del Genoma Completo , Grupos de Población , Humanos , Diabetes Mellitus Tipo 2/genética , Desequilibrio de Ligamiento , Fenotipo , Polimorfismo de Nucleótido Simple , Genética de Población , Grupos de Población/genética
2.
Proc Natl Acad Sci U S A ; 119(15): e2109934119, 2022 04 12.
Artículo en Inglés | MEDLINE | ID: mdl-35394880

RESUMEN

Photoreceptor connecting cilium (CC) is structurally analogous to the transition zone (TZ) of primary cilia and gates the molecular trafficking between the inner and the outer segment (OS). Retinal dystrophies with underlying CC defects are manifested in a broad array of syndromic conditions known as ciliopathies as well as nonsyndromic retinal degenerations. Despite extensive studies, many questions remain in the mechanism of protein trafficking across the photoreceptor CC. Here, we genetically inactivated mouse Tmem138, a gene encoding a putative transmembrane protein localized to the ciliary TZ and linked to ciliopathies. Germline deletion of Tmem138 abolished OS morphogenesis, followed by rapid photoreceptor degeneration. Tmem138 was found localized to the photoreceptor CC and was required for localization of Ahi1 to the distal subdomain of the CC. Among the examined set of OS proteins, rhodopsin was mislocalized throughout the mutant cell body prior to OS morphogenesis. Ablation of Tmem138 in mature rods recapitulated the molecular changes in the germline mutants, causing failure of disc renewal and disintegration of the OS. Furthermore, Tmem138 interacts reciprocally with rhodopsin and a related protein Tmem231, and the ciliary localization of the latter was also altered in the mutant photoreceptors. Taken together, these results suggest a crucial role of Tmem138 in the functional organization of the CC, which is essential for rhodopsin localization and OS biogenesis.


Asunto(s)
Ciliopatías , Degeneración Retiniana , Cilios/metabolismo , Ciliopatías/metabolismo , Humanos , Proteínas de la Membrana , Cilio Conector de los Fotorreceptores , Degeneración Retiniana/metabolismo , Rodopsina/genética , Rodopsina/metabolismo
3.
Exp Eye Res ; 243: 109906, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38657786

RESUMEN

Pediatric cataract, including congenital and developmental cataract, is a kind of pediatric vision-threatening disease with extensive phenotypic heterogeneity and multiple mechanisms. We aimed to investigate the metabolite profile of aqueous humor (AH) in patients with pediatric cataracts, and identify underlying mutual correlations between differential metabolites. Metabolomic profiles of AH were analyzed and compared between pediatric cataract patients (n = 33) and age-related cataract patients without metabolic diseases (n = 29), using global untargeted metabolomics with ultra-high-performance liquid chromatography tandem mass spectrometry. Principal component analysis, partial least squares discriminant analysis and heat map were applied. Enriched pathway analysis was conducted using Kyoto Encyclopedia of Genes and Genomes. Receiver-operating characteristic (ROC) analyses were employed to select potential biomarkers. A total of 318 metabolites were identified, of which 54 differential metabolites (25 upregulated and 29 downregulated) were detected in pediatric cataract group compared with controls (variable importance of projection >1.0, fold change ≥1.5 or ≤ 0.667 and P < 0.05). A significant accumulation of N-Acetyl-Dl-glutamic acid was observed in pediatric cataract group. The differential metabolites were mainly enriched in histidine metabolism (increased L-Histidine and decreased 1-Methylhistamine) and the tryptophan metabolism (increased N-Formylkynurenine and L-Kynurenine). 5-Aminosalicylic acid showed strong positive mutual inter-correlation with L-Tyrosinemethylester and N,N-Diethylethanolamine, both of which were down-regulated in pediatric cataract group. The ROC analysis implied 11 metabolites served as potential biomarkers for pediatric cataract patients (all area under the ROC curve ≥0.900). These results illustrated novel potential metabolites and metabolic pathways in pediatric cataract, which provides new insights into the pathophysiology of pediatric cataract.


Asunto(s)
Humor Acuoso , Biomarcadores , Catarata , Metabolómica , Humanos , Humor Acuoso/metabolismo , Catarata/metabolismo , Metabolómica/métodos , Masculino , Femenino , Preescolar , Cromatografía Líquida de Alta Presión , Niño , Biomarcadores/metabolismo , Curva ROC , Espectrometría de Masas en Tándem , Metaboloma/fisiología , Lactante
4.
Exp Eye Res ; 243: 109899, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38636802

RESUMEN

Virus-like particles (VLP) are a promising tool for intracellular gene delivery, yet their potential in ocular gene therapy remains underexplored. In this study, we bridged this knowledge gap by demonstrating the successful generation and application of vesicular stomatitis virus glycoprotein (VSVG)-pseudotyped mouse PEG10 (MmPEG10)-VLP for intraocular mRNA delivery. Our findings revealed that PEG10-VLP can efficiently deliver GFP mRNA to adult retinal pigment epithelial cell line-19 (ARPE-19) cells, leading to transient expression. Moreover, we showed that MmPEG10-VLP can transfer SMAD7 to inhibit epithelial-mesenchymal transition (EMT) in RPE cells effectively. In vivo experiments further substantiated the potential of these vectors, as subretinal delivery into adult mice resulted in efficient transduction of retinal pigment epithelial (RPE) cells and GFP reporter gene expression without significant immune response. However, intravitreal injection did not yield efficient ocular expression. We also evaluated the transduction characteristics of MmPEG10-VLP following intracameral delivery, revealing transient GFP protein expression in corneal endothelial cells without significant immunotoxicities. In summary, our study established that VSVG pseudotyped MmPEG10-based VLP can transduce mitotically inactive RPE cells and corneal endothelial cells in vivo without triggering an inflammatory response, underscoring their potential utility in ocular gene therapy.


Asunto(s)
Técnicas de Transferencia de Gen , ARN Mensajero , Epitelio Pigmentado de la Retina , Animales , Ratones , Epitelio Pigmentado de la Retina/metabolismo , ARN Mensajero/genética , Terapia Genética/métodos , Vectores Genéticos , Ratones Endogámicos C57BL , Humanos , Proteínas Fluorescentes Verdes/genética , Transición Epitelial-Mesenquimal , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo
5.
Graefes Arch Clin Exp Ophthalmol ; 262(6): 1829-1838, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38197993

RESUMEN

PURPOSE: To investigate the effect of posterior keratometry (PK) on the accuracy of 10 intraocular lens (IOL) power calculation formulas using standard keratometry (K) and total keratometry (TK). METHODS: This is a retrospective consecutive case-series study. The IOL power was calculated using K and TK measured by IOLMaster 700 in 6 new-generation formulas (Barrett Universal II, Emmetropia Verifying Optical (EVO) 2.0, RBF Calculator 3.0, Hoffer QST, Kane, and Ladas Super Formula) and 4 traditional formulas (Haigis, Hoffer Q, Holladay 1, and SRK/T). The arithmetic prediction error (PE) and mean absolute PE (MAE) were evaluated. The locally-weighted scatterplot smoothing was performed to assess the relationship between PE and PK. RESULTS: A total of 576 patients (576 eyes) who underwent cataract surgery were included. Compared with using K, all formulas using TK showed a hyperopic shift in the whole group. Specifically, for eyes with PK exceeding -5.90 D, all formulas using TK exhibited a hyperopic shift (all P < 0.001), while eyes with PK less than -5.90 D showed a myopic shift (all P < 0.001). The MAE of new-generation formulas calculated with TK and K showed no statistical differences, while the MAE of traditional formulas with TK was larger (TK: 0.34 ~ 0.43 D; K: 0.33 ~ 0.42 D, all P < 0.05). CONCLUSIONS: The prediction bias of formulas with TK increased as PK deviated from -5.90 D. TK did not improve the prediction accuracy of new-generation formulas, and even performed worse in traditional formulas.


Asunto(s)
Biometría , Córnea , Lentes Intraoculares , Óptica y Fotónica , Refracción Ocular , Humanos , Estudios Retrospectivos , Refracción Ocular/fisiología , Femenino , Masculino , Biometría/métodos , Anciano , Córnea/diagnóstico por imagen , Reproducibilidad de los Resultados , Persona de Mediana Edad , Agudeza Visual/fisiología , Facoemulsificación/métodos , Anciano de 80 o más Años , Estudios de Seguimiento , Implantación de Lentes Intraoculares/métodos
6.
Proc Natl Acad Sci U S A ; 118(11)2021 03 16.
Artículo en Inglés | MEDLINE | ID: mdl-33836603

RESUMEN

Retinal neovascularization is a leading cause of severe visual loss in humans, and molecular mechanisms of microglial activation-driven angiogenesis remain unknown. Using single-cell RNA sequencing, we identified a subpopulation of microglia named sMG2, which highly expressed necroptosis-related genes Rip3 and Mlkl. Genetic and pharmacological loss of function demonstrated that hypoxia-induced microglial activation committed to necroptosis through the RIP1/RIP3-mediated pathway. Specific deletion of Rip3 gene in microglia markedly decreased retinal neovascularization. Furthermore, hypoxia induced explosive release of abundant FGF2 in microglia through RIP3-mediated necroptosis. Importantly, blocking signaling components of the microglia necropotosis-FGF2 axis largely ablated retinal angiogenesis and combination therapy with simultaneously blocking VEGF produced synergistic antiangiogenic effects. Together, our data demonstrate that targeting the microglia necroptosis axis is an antiangiogenesis therapy for retinal neovascular diseases.


Asunto(s)
Microglía/patología , Necroptosis , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Enfermedades de la Retina/patología , Animales , Quimioterapia Combinada , Factor 2 de Crecimiento de Fibroblastos/antagonistas & inhibidores , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Hipoxia/patología , Ratones , Microglía/metabolismo , Necroptosis/efectos de los fármacos , Neovascularización Patológica , Proteínas Quinasas/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/antagonistas & inhibidores , Enfermedades de la Retina/tratamiento farmacológico , Enfermedades de la Retina/metabolismo , Vasos Retinianos/efectos de los fármacos , Vasos Retinianos/metabolismo , Vasos Retinianos/patología , Transducción de Señal/efectos de los fármacos , Factor A de Crecimiento Endotelial Vascular/antagonistas & inhibidores , Factor A de Crecimiento Endotelial Vascular/metabolismo
7.
Hum Genet ; 142(4): 577-593, 2023 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-36964785

RESUMEN

CLCN2 encodes a two-pore homodimeric chloride channel protein (CLC-2) that is widely expressed in human tissues. The association between Clcn2 and the retina is well-established in mice, as loss-of-function of CLC-2 can cause retinopathy in mice; however, the ocular phenotypes caused by CLCN2 mutations in humans and the underlying mechanisms remain unclear. The present study aimed to define the ocular features and reveal the pathogenic mechanisms of CLCN2 variants associated with retinal degeneration in humans using an in vitro overexpression system, as well as patient-induced pluripotent stem cell (iPSC)-derived retinal pigment epithelium (RPE) cells and retinal organoids (ROs). A patient carrying the homozygous c.2257C > T (p.R753X) nonsense CLCN2 mutation was followed up for > 6 years. Ocular features were comprehensively characterized with multimodality imaging and functional examination. The patient presented with severe bilateral retinal degeneration with loss of photoreceptor and RPE. In vitro, mutant CLC-2 maintained the correct subcellular localization, but with reduced channel function compared to wild-type CLC-2 in HEK293T cells. Additionally, patient iPSC-derived RPE cells carrying the CLCN2 mutation exhibited dysfunctional ClC-2 chloride channels and outer segment phagocytosis. Notably, these functions were rescued following the repair of the CLCN2 mutation using the CRISPR-Cas9 system. However, this variant did not cause significant photoreceptor degeneration in patient-derived ROs, indicating that dysfunctional RPE is likely the primary cause of biallelic CLCN2 variant-mediated retinopathy. This study is the first to establish the confirmatory ocular features of human CLCN2-related retinal degeneration, and reveal a pathogenic mechanism associated with biallelic CLCN2 variants, providing new insights into the cause of inherited retinal dystrophies.


Asunto(s)
Células Madre Pluripotentes Inducidas , Distrofias Retinianas , Animales , Humanos , Ratones , Canales de Cloruro/genética , Codón sin Sentido , Células HEK293 , Mutación , Fagocitosis/genética , Especies Reactivas de Oxígeno/metabolismo , Distrofias Retinianas/metabolismo , Epitelio Pigmentado de la Retina/metabolismo , Epitelio Pigmentado de la Retina/patología
8.
Development ; 147(22)2020 11 30.
Artículo en Inglés | MEDLINE | ID: mdl-33144400

RESUMEN

In terrestrial animals, the lacrimal drainage apparatus evolved to serve as conduits for tear flow; however, little is known about the ontogenesis of this system. Here, we define the anatomy of the fully formed tear duct in mice, characterize crucial morphogenetic events for the development of tear duct components and identify the site for primordial tear duct (PTD) initiation. We report that the PTD originates from the orbital lacrimal lamina, a junction formed by the epithelia of the maxillary and lateral nasal processes. We demonstrate that Prickle1, a key component of planar cell polarity signaling, is expressed in progenitors of the PTD and throughout tear duct morphogenesis. Disruption of Prickle1 stalls tear duct elongation; in particular, the loss of basement membrane deposition and aberrant cytoplasmic accumulation of laminin are salient. Altered cell adhesion, cytoskeletal transport systems, vesicular transport systems and cell axis orientation in Prickle1 mutants support the role of Prickle1 in planar cell polarity. Taken together, our results highlight a crucial role of Prickle1-mediated polarized basement membrane secretion and deposition in PTD elongation.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Membrana Basal/embriología , Polaridad Celular/fisiología , Proteínas con Dominio LIM/metabolismo , Conducto Nasolagrimal/embriología , Organogénesis/fisiología , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Membrana Basal/citología , Adhesión Celular/fisiología , Citoesqueleto/genética , Citoesqueleto/metabolismo , Proteínas con Dominio LIM/genética , Ratones , Conducto Nasolagrimal/citología
9.
J Neuroinflammation ; 20(1): 119, 2023 May 22.
Artículo en Inglés | MEDLINE | ID: mdl-37217935

RESUMEN

BACKGROUND: Chronic inflammation significantly contributes to photoreceptor death in blinding retinal diseases such as age-related macular degeneration (AMD) and retinitis pigmentosa (RP). Bromodomain and extraterminal domain (BET) proteins are epigenetic readers that act as key proinflammatory factors. We recently found the first-generation BET inhibitor JQ1 alleviated sodium iodate-induced retinal degeneration by suppressing cGAS-STING innate immunity. Here, we investigated the effects and mechanism of dBET6, a proteolysis­targeting chimera (PROTAC) small molecule that selectively degrades BET by the ubiquitin‒proteasome system, in light-induced retinal degeneration. METHODS: Mice were exposed to bright light to induce retinal degeneration, and the activation of cGAS-STING was determined by RNA-sequencing and molecular biology. Retinal function, morphology, photoreceptor viability and retinal inflammation were examined in the presence and absence of dBET6 treatment. RESULTS: Intraperitoneal injection of dBET6 led to the rapid degradation of BET protein in the retina without detectable toxicity. dBET6 improved retinal responsiveness and visual acuity after light damage (LD). dBET6 also repressed LD-induced retinal macrophages/microglia activation, Müller cell gliosis, photoreceptor death and retinal degeneration. Analysis of single-cell RNA-sequencing results revealed cGAS-STING components were expressed in retinal microglia. LD led to dramatic activation of the cGAS-STING pathway, whereas dBET6 suppressed LD-induced STING expression in reactive macrophages/microglia and the related inflammatory response. CONCLUSIONS: This study indicates targeted degradation of BET by dBET6 exerts neuroprotective effects by inhibiting cGAS-STING in reactive retinal macrophages/microglia, and is expected to become a new strategy for treatment of retinal degeneration.


Asunto(s)
Degeneración Retiniana , Ratones , Animales , Degeneración Retiniana/etiología , Degeneración Retiniana/prevención & control , Degeneración Retiniana/metabolismo , Inflamación/metabolismo , Nucleotidiltransferasas , ARN
10.
Cell Commun Signal ; 21(1): 134, 2023 06 14.
Artículo en Inglés | MEDLINE | ID: mdl-37316948

RESUMEN

BACKGROUND: Age-related macular degeneration (AMD), characterized by the degeneration of retinal pigment epithelium (RPE) and photoreceptors, is the leading cause of irreversible vision impairment among the elderly. RPE senescence is an important contributor to AMD and has become a potential target for AMD therapy. HTRA1 is one of the most significant susceptibility genes in AMD, however, the correlation between HTRA1 and RPE senescence hasn't been investigated in the pathogenesis of AMD. METHODS: Western blotting and immunohistochemistry were used to detect HTRA1 expression in WT and transgenic mice overexpressing human HTRA1 (hHTRA1-Tg mice). RT-qPCR was used to detect the SASP in hHTRA1-Tg mice and ARPE-19 cells infected with HTRA1. TEM, SA-ß-gal was used to detect the mitochondria and senescence in RPE. Retinal degeneration of mice was investigated by fundus photography, FFA, SD-OCT and ERG. The RNA-Seq dataset of ARPE-19 cells treated with adv-HTRA1 versus adv-NC were analyzed. Mitochondrial respiration and glycolytic capacity in ARPE-19 cells were measured using OCR and ECAR. Hypoxia of ARPE-19 cells was detected using EF5 Hypoxia Detection Kit. KC7F2 was used to reduce the HIF1α expression both in vitro and in vivo. RESULTS: In our study, we found that RPE senescence was facilitated in hHTRA1-Tg mice. And hHTRA1-Tg mice became more susceptible to NaIO3 in the development of oxidative stress-induced retinal degeneration. Similarly, overexpression of HTRA1 in ARPE-19 cells accelerated cellular senescence. Our RNA-seq revealed an overlap between HTRA1-induced differentially expressed genes associated with aging and those involved in mitochondrial function and hypoxia response in ARPE-19 cells. HTRA1 overexpression in ARPE-19 cells impaired mitochondrial function and augmented glycolytic capacity. Importantly, upregulation of HTRA1 remarkably activated HIF-1 signaling, shown as promoting HIF1α expression which mainly located in the nucleus. HIF1α translation inhibitor KC7F2 significantly prevented HTRA1-induced cellular senescence in ARPE-19 cells, as well as improved the visual function in hHTRA1-Tg mice treated with NaIO3. CONCLUSIONS: Our study showed elevated HTRA1 contributes to the pathogenesis of AMD by promoting cellular senescence in RPE through damaging mitochondrial function and activating HIF-1 signaling. It also pointed out that inhibition of HIF-1 signaling might serve as a potential therapeutic strategy for AMD. Video Abstract.


Asunto(s)
Degeneración Retiniana , Anciano , Humanos , Animales , Ratones , Epitelio Pigmentado de la Retina , Transducción de Señal , Mitocondrias , Núcleo Celular
11.
Int J Syst Evol Microbiol ; 73(11)2023 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-37917540

RESUMEN

Two Gram-stain negative, aerobic and rod-shaped bacterial strains, DHOD12T and 7GSK02T, were isolated from forest soil of Dinghushan Biosphere Reserve, Guangdong Province, PR China. Strain DHOD12T grew at 4-42 °C (optimum, 28-33 °C), pH 4.0-8.5 (optimum, pH 5.5-6.5) and in the presence of 0-1.5 % (w/v; optimum, 0-0.5 %)NaCl; while strain 7GSK02T grew at 12-42 °C (optimum, 28-33 °C), pH 4.0-8.5 (optimum, pH 5.0-6.0) and in the presence of 0-0.5 % (w/v; optimum, 0 %) NaCl. Strains DHOD12T and 7GSK02T had the highest 16S rRNA sequence similarities of 98.0 and 98.3 % with the same species Trinickia mobilis DHG64T, respectively, and 98.4 % between themselves. In the 16S rRNA phylogeny, they formed a clade that was sister to a major cluster consisting of all described Trinickia species. Phylogenomic analyses with the UBCG and PhyloPhlAn methods consistently showed that strains DHOD12T and 7GSK02T formed a clade with T. mobilis DHG64T that was a sister of a cluster containing the remainder of the Trinickia species. The DNA G+C contents of strains DHOD12T and 7GSK02T were 63.1 and 64.6 mol%, respectively. Digital DNA-DNA hybridization and average nucleotide identity values of strains DHOD12T, 7GSK02T and their closely related strains were in the ranges of 21.6-31.4 % and 77.1-86.9 %, respectively. These two strains had the same major respiratory quinone, ubiquinone-8, and both had C16 : 0, C17 : 0 cyclo and summed feature 8 (C18 : 1 ω7c/C18 : 1 ω6c) as their major fatty acids. Their major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol. Genomic analysis indicated that the two strains could have the potential to degrade aromatic compounds like other Trinickia species. On the basis of phenotypic and phylogenetic results, strains DHOD12T and 7GSK02T represent two novel species of the genus Trinickia, for which the names Trinickia violacea sp. nov. (type strain DHOD12T=LMG 30258T=CGMCC 1.15436T) and Trinickia terrae sp. nov. (type strain 7GSK02T=CGMCC 1.15432T=KCTC 62468T) are proposed.


Asunto(s)
Burkholderiaceae , Ácidos Grasos , Ácidos Grasos/química , Fosfolípidos/química , ARN Ribosómico 16S/genética , Filogenia , Cloruro de Sodio , ADN Bacteriano/genética , Análisis de Secuencia de ADN , Composición de Base , Técnicas de Tipificación Bacteriana , Bosques
12.
Mol Cell ; 59(6): 931-40, 2015 Sep 17.
Artículo en Inglés | MEDLINE | ID: mdl-26365380

RESUMEN

Glaucoma, a blinding neurodegenerative disease, whose risk factors include elevated intraocular pressure (IOP), age, and genetics, is characterized by accelerated and progressive retinal ganglion cell (RGC) death. Despite decades of research, the mechanism of RGC death in glaucoma is still unknown. Here, we demonstrate that the genetic effect of the SIX6 risk variant (rs33912345, His141Asn) is enhanced by another major POAG risk gene, p16INK4a (cyclin-dependent kinase inhibitor 2A, isoform INK4a). We further show that the upregulation of homozygous SIX6 risk alleles (CC) leads to an increase in p16INK4a expression, with subsequent cellular senescence, as evidenced in a mouse model of elevated IOP and in human POAG eyes. Our data indicate that SIX6 and/or IOP promotes POAG by directly increasing p16INK4a expression, leading to RGC senescence in adult human retinas. Our study provides important insights linking genetic susceptibility to the underlying mechanism of RGC death and provides a unified theory of glaucoma pathogenesis.


Asunto(s)
Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Glaucoma de Ángulo Abierto/metabolismo , Proteínas de Homeodominio/fisiología , Células Ganglionares de la Retina/fisiología , Transactivadores/fisiología , Secuencia de Aminoácidos , Animales , Estudios de Casos y Controles , Muerte Celular , Línea Celular , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , Glaucoma de Ángulo Abierto/genética , Glaucoma de Ángulo Abierto/patología , Humanos , Ratones Endogámicos C57BL , Ratones Noqueados , Datos de Secuencia Molecular , Mutación Missense , Regulación hacia Arriba
13.
Vascular ; 31(5): 892-901, 2023 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-35466838

RESUMEN

OBJECTIVES: The aim of this study is to identify the peri-procedural risk factors and outcomes of hemodynamic instability (HI) after carotid artery stenting (CAS). METHODS: A single-center, retrospective study was performed in 168 patients who underwent CAS procedure between September 2017 and September 2020. The presence of HI, as defined by hypertension (systolic blood pressure >160 mmHg), hypotension (systolic blood pressure <90 mmHg), and/or bradycardia (heart rate <60 bpm), was recorded. Long-period HI was defined as persistent HI lasting more than 24 h. Patient demographics, comorbidities, peri-procedural variables, and risk factors were recorded. Clinical outcomes including cerebral hyperperfusion syndrome, hemorrhage, transient ischemic attack (TIA), stroke, myocardial infarction, and mortality within 30 days of the procedure were evaluated. Logistic regression was used to analyze the independent risk factors of long-period HI following CAS. RESULTS: Among 168 patients (mean age, 68.2 ± 8.3 years; 81.5% male), the frequency of post-procedural long-period HI was noted in 42 patients (25.0%). Male was prone to experience HI (odds ratio, 9.156, p = 0.021). Aggressive inflation pressure (>7 atm) and 5 mm balloon for pre-dilatation were risk factors of long-period HI (OR, 7.372, p = 0.035; OR, 3.527, p = 0.023). Intraoperative peak blood pressure and larger-sized stents remained independent predictors for the development of HI (OR, 1.043, p = 0.027, and OR, 1.973, p = 0.015). Patients with prolonged HI were more likely to suffer TIA and stroke compared to other patients and significant difference was found in the occurrence of TIA (p < 0.05). Non-significance was found in mortality rate and other outcomes. CONCLUSIONS: CAS-induced HI occurs in a considerable percentage while several peri-procedural variables are determined as independent predictors to develop long-period HI. Patients with prolonged HI are associated with increased risk of neurologic events and thus standardized intervention as well as management of long-period HI are of critical importance during clinical process.


Asunto(s)
Estenosis Carotídea , Ataque Isquémico Transitorio , Accidente Cerebrovascular , Humanos , Masculino , Persona de Mediana Edad , Anciano , Femenino , Estenosis Carotídea/complicaciones , Estenosis Carotídea/diagnóstico por imagen , Estenosis Carotídea/terapia , Ataque Isquémico Transitorio/etiología , Estudios Retrospectivos , Stents/efectos adversos , Angioplastia/efectos adversos , Arteria Carótida Común , Presión Sanguínea , Accidente Cerebrovascular/etiología , Factores de Riesgo , Resultado del Tratamiento
14.
BMC Med Educ ; 23(1): 912, 2023 Nov 30.
Artículo en Inglés | MEDLINE | ID: mdl-38037047

RESUMEN

BACKGROUND: Traditional problem-based learning (PBL) relying on tutored learning in small groups is very resource-intensive. Little is known about the benefits of PBL in a large classroom setting. This paper introduced a PBL case into the traditional didactic biochemistry course and investigated the acceptability of total online or partial online PBL in a large classroom setting introduced during the coronavirus pandemic. METHODS: The students were allocated into either total online Group 1, partial online Group 2, or partial online and with poorer academic performance Group 3. A questionnaire comprising of 8 closed-ended questions and 2 open-ended questions and final exam performances were used to evaluate the acceptability of total online or partial online PBL in a large classroom setting. The 8 closed-ended questions were analysed by the Kruskal-Wallis test or chi-square tests. The word cloud analysis of the 2 open-ended questions were conducted by Wenjuanxing. Students' performances in the final examination were analysed by One-way Anova. RESULTS: Both total online and partial online PBL were rated highly by the students. Overall, there were no significant differences in the effectiveness evaluation of PBL between Group 2 and Group 3. There were no significant differences in final exam performances between Group 1 and Group 2. However, Group 1 rated the effectiveness of PBL much higher than Group 2 and 3. Word cloud analysis of the 2 open-ended questions showed students' positive perspectives of PBL. In biochemistry teaching, from the perspective of the students, the expected optimal number of useful PBL cases might be 2. CONCLUSIONS: Both total online and partial online PBL in a large classroom setting were widely accepted as a beneficial supplement to traditional biochemistry classes.


Asunto(s)
Aprendizaje Basado en Problemas , Estudiantes , Humanos , Evaluación Educacional , Bioquímica/educación , Encuestas y Cuestionarios
15.
Nature ; 531(7594): 323-8, 2016 Mar 17.
Artículo en Inglés | MEDLINE | ID: mdl-26958831

RESUMEN

The repair and regeneration of tissues using endogenous stem cells represents an ultimate goal in regenerative medicine. To our knowledge, human lens regeneration has not yet been demonstrated. Currently, the only treatment for cataracts, the leading cause of blindness worldwide, is to extract the cataractous lens and implant an artificial intraocular lens. However, this procedure poses notable risks of complications. Here we isolate lens epithelial stem/progenitor cells (LECs) in mammals and show that Pax6 and Bmi1 are required for LEC renewal. We design a surgical method of cataract removal that preserves endogenous LECs and achieves functional lens regeneration in rabbits and macaques, as well as in human infants with cataracts. Our method differs conceptually from current practice, as it preserves endogenous LECs and their natural environment maximally, and regenerates lenses with visual function. Our approach demonstrates a novel treatment strategy for cataracts and provides a new paradigm for tissue regeneration using endogenous stem cells.


Asunto(s)
Catarata/terapia , Cristalino/citología , Cristalino/fisiología , Recuperación de la Función , Regeneración/fisiología , Células Madre/citología , Visión Ocular/fisiología , Animales , Catarata/congénito , Catarata/patología , Catarata/fisiopatología , Extracción de Catarata , Células Epiteliales/citología , Células Epiteliales/metabolismo , Proteínas del Ojo/metabolismo , Proteínas de Homeodominio/metabolismo , Homeostasis , Humanos , Macaca , Factor de Transcripción PAX6 , Factores de Transcripción Paired Box/metabolismo , Complejo Represivo Polycomb 1/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Células Madre/metabolismo
16.
Dev Dyn ; 250(7): 974-985, 2021 07.
Artículo en Inglés | MEDLINE | ID: mdl-33449399

RESUMEN

BACKGROUND: Genetic programs underlying preimplantation development and early lineage segregation are highly conserved across mammals. It has been suggested that nonhuman primates would be better model organisms for human embryogenesis, but a limited number of studies have investigated the monkey preimplantation development. In this study, we collect single cells from cynomolgus monkey preimplantation embryos for transcriptome profiling and compare with single-cell RNA-seq data derived from human and mouse embryos. RESULTS: By weighted gene-coexpression network analysis, we found that cynomolgus gene networks have greater conservation with human embryos including a greater number of conserved hub genes than that of mouse embryos. Consistently, we found that early ICM/TE lineage-segregating genes in monkeys exhibit greater similarity with human when compared to mouse, so are the genes in signaling pathways such as LRP1 and TCF7 involving in WNT pathway. Last, we tested the role of one conserved pre-EGA hub gene, SIN3A, using a morpholino knockdown of maternal RNA transcripts in monkey embryos followed by single-cell RNA-seq. We found that SIN3A knockdown disrupts the gene-silencing program during the embryonic genome activation transition and results in developmental delay of cynomolgus embryos. CONCLUSION: Taken together, our study provided new insight into evolutionarily conserved and divergent transcriptome dynamics during mammalian preimplantation development.


Asunto(s)
Blastómeros/metabolismo , Desarrollo Embrionario/genética , Macaca fascicularis/embriología , Adulto , Animales , Blastocisto , Blastómeros/citología , Linaje de la Célula/genética , Células Cultivadas , Embrión de Mamíferos , Desarrollo Embrionario/fisiología , Femenino , Perfilación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Redes Reguladoras de Genes/fisiología , Humanos , Macaca fascicularis/genética , Macaca mulatta , Masculino , Ratones , Embarazo , Complejo Correpresor Histona Desacetilasa y Sin3/genética , Complejo Correpresor Histona Desacetilasa y Sin3/fisiología , Análisis de la Célula Individual/veterinaria , Transcriptoma/genética
17.
Am J Physiol Lung Cell Mol Physiol ; 320(1): L84-L98, 2021 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-33146564

RESUMEN

Coronavirus disease 2019 (COVID-19), driven by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was declared a global pandemic in March 2020. Pathogenic T cells and inflammatory monocytes are regarded as the central drivers of the cytokine storm associated with the severity of COVID-19. In this study, we explored the characteristic peripheral cellular profiles of patients with COVID-19 in both acute and convalescent phases by single-cell mass cytometry (CyTOF). Using a combination of algorithm-guided data analyses, we identified peripheral immune cell subsets in COVID-19 and revealed CD4+ T-cell depletion, T-cell differentiation, plasma cell expansion, and the reduced antigen presentation capacity of innate immunity. Notably, COVID-19 induces a dysregulation in the balance of monocyte populations by the expansion of the monocyte subsets. Collectively, our results represent a high-dimensional, single-cell profile of the peripheral immune response to SARS-CoV-2 infection.


Asunto(s)
Linfocitos T CD4-Positivos/inmunología , COVID-19/inmunología , Leucocitos Mononucleares/inmunología , Monocitos/inmunología , SARS-CoV-2/inmunología , Adulto , Anciano , Presentación de Antígeno/inmunología , Linfocitos T CD4-Positivos/citología , COVID-19/patología , Síndrome de Liberación de Citoquinas/inmunología , Síndrome de Liberación de Citoquinas/patología , Citocinas/inmunología , Células Dendríticas/citología , Células Dendríticas/inmunología , Femenino , Humanos , Células Asesinas Naturales/citología , Células Asesinas Naturales/inmunología , Leucocitos Mononucleares/citología , Depleción Linfocítica , Masculino , Persona de Mediana Edad , Monocitos/citología , Células Plasmáticas/citología , Análisis de la Célula Individual
18.
Liver Int ; 41(8): 1956-1968, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-33786995

RESUMEN

BACKGROUND: Liver fibrosis is pathologically important in the liver cirrhosis progression. The epithelial-mesenchymal transition (EMT) is crucial for organ fibrosis. Macrophage-stimulating protein (MSP) and its receptor tyrosine kinase, RON, promote cellular EMT. However, their role in liver fibrosis is unclear. Here, we clarify the biological profile, potential mechanisms and therapeutic targets of the MSP-RON pathway in liver fibrosis. MATERIALS AND METHODS: Macrophage-stimulating protein expression and its correlation with clinicopathological characteristics of cirrhosis were evaluated in 57 clinical cases and a control group. The effect of MSP-RON pathway in liver fibrosis was determined in vitro and in vivo. The therapeutic effects of MSP or RON inhibition on liver fibrosis were evaluated in a mouse liver fibrosis model. RESULTS: Macrophage-stimulating protein is upregulated in liver cirrhosis, which was associated with poor patient prognosis. The MSP-RON pathway promoted hepatocytes EMT. MSP-RON-induced EMT depends on the transforming growth factor beta (TGF-ß) pathway and is regulated by TGF-ß inhibitors. In animal models, an MSP blocking antibody and a small molecule inhibitor of RON, BMS-777607, both inhibited liver fibrosis progression. CONCLUSION: Our study revealed that MSP is an important biomarker in liver cirrhosis progression and can be used to prognose patients. The MSP-RON pathway promotes the EMT of hepatocytes and the progress of fibrosis via a TGF-ß related pathway. Consequently, we identified a new treatment strategy for liver cirrhosis through targeted inhibition of MSP/RON. This research increases the understanding of EMT-modulated liver fibrosis and provides new insights into biomarkers and therapeutic targets of liver fibrosis.


Asunto(s)
Transición Epitelial-Mesenquimal , Cirrosis Hepática , Factor de Crecimiento Transformador beta , Animales , Factor de Crecimiento de Hepatocito , Humanos , Ratones , Proteínas Tirosina Quinasas Receptoras
19.
J Pineal Res ; 71(1): e12716, 2021 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-33426650

RESUMEN

Retinopathy of prematurity is a vision-threatening disease associated with retinal hypoxia-ischemia, leading to the death of retinal neurons and chronic neuronal degeneration. During this study, we used the oxygen-induced retinopathy mice model to mimic retinal hypoxia-ischemia phenotypes to investigate further the neuroprotective effect of melatonin on neonatal retinal neurons. Melatonin helped maintain relatively normal inner retinal architecture and thickness and preserve inner retinal neuron populations in avascular areas by rescuing retinal ganglion and bipolar cells, and horizontal and amacrine neurons, from apoptosis. Meanwhile, melatonin recovered visual dysfunction, as reflected by the improved amplitudes and implicit times of a-wave, b-wave, and oscillatory potentials. Additionally, elevated cleaved caspase-3 and Bax protein levels and reduced Bcl-2 protein levels in response to hypoxia-ischemia were diminished after melatonin treatment. Moreover, melatonin increased BDNF and downstream phospho-TrkB/Akt/ERK/CREB levels. ANA-12, a TrkB receptor antagonist, antagonized these melatonin actions and reduced melatonin-induced neuroprotection. Furthermore, melatonin rescued the reduction in melatonin receptor expression. This study suggests that melatonin exerted anti-apoptotic and neuroprotective effects in inner retinal neurons after hypoxia-ischemia, at least partly due to modulation of the BDNF-TrkB pathway.


Asunto(s)
Melatonina/farmacología , Fármacos Neuroprotectores/farmacología , Neuronas Retinianas/efectos de los fármacos , Neuronas Retinianas/patología , Retinopatía de la Prematuridad , Animales , Animales Recién Nacidos , Hipoxia/etiología , Hipoxia/patología , Isquemia/etiología , Isquemia/patología , Ratones , Ratones Endogámicos C57BL , Retinopatía de la Prematuridad/complicaciones , Retinopatía de la Prematuridad/patología
20.
Nature ; 523(7562): 607-11, 2015 Jul 30.
Artículo en Inglés | MEDLINE | ID: mdl-26200341

RESUMEN

The human lens is comprised largely of crystallin proteins assembled into a highly ordered, interactive macro-structure essential for lens transparency and refractive index. Any disruption of intra- or inter-protein interactions will alter this delicate structure, exposing hydrophobic surfaces, with consequent protein aggregation and cataract formation. Cataracts are the most common cause of blindness worldwide, affecting tens of millions of people, and currently the only treatment is surgical removal of cataractous lenses. The precise mechanisms by which lens proteins both prevent aggregation and maintain lens transparency are largely unknown. Lanosterol is an amphipathic molecule enriched in the lens. It is synthesized by lanosterol synthase (LSS) in a key cyclization reaction of a cholesterol synthesis pathway. Here we identify two distinct homozygous LSS missense mutations (W581R and G588S) in two families with extensive congenital cataracts. Both of these mutations affect highly conserved amino acid residues and impair key catalytic functions of LSS. Engineered expression of wild-type, but not mutant, LSS prevents intracellular protein aggregation of various cataract-causing mutant crystallins. Treatment by lanosterol, but not cholesterol, significantly decreased preformed protein aggregates both in vitro and in cell-transfection experiments. We further show that lanosterol treatment could reduce cataract severity and increase transparency in dissected rabbit cataractous lenses in vitro and cataract severity in vivo in dogs. Our study identifies lanosterol as a key molecule in the prevention of lens protein aggregation and points to a novel strategy for cataract prevention and treatment.


Asunto(s)
Catarata/tratamiento farmacológico , Catarata/metabolismo , Lanosterol/farmacología , Lanosterol/uso terapéutico , Agregado de Proteínas/efectos de los fármacos , Agregación Patológica de Proteínas/tratamiento farmacológico , Adulto , Secuencia de Aminoácidos , Amiloide/química , Amiloide/efectos de los fármacos , Amiloide/metabolismo , Amiloide/ultraestructura , Animales , Secuencia de Bases , Catarata/congénito , Catarata/genética , Catarata/patología , Línea Celular , Niño , Cristalinas/química , Cristalinas/genética , Cristalinas/metabolismo , Cristalinas/ultraestructura , Perros , Femenino , Humanos , Lanosterol/administración & dosificación , Cristalino/efectos de los fármacos , Cristalino/metabolismo , Cristalino/patología , Masculino , Modelos Moleculares , Datos de Secuencia Molecular , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteínas Mutantes/ultraestructura , Linaje , Agregación Patológica de Proteínas/patología
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