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AIM: To investigate the difference in the initial surgical results between a new monofocal intraocular lens (IOL) with enhanced intermediate vision and the standard monofocal IOL in patients with retinal disease. METHODS: We retrospectively reviewed the medical records of patients with retinal disease who underwent cataract surgery due to accompanying cataracts. Types of retinal diseases were investigated and best-corrected distant visual acuity, distant uncorrected visual acuity (UCVA), intermediate UCVA, near UCVA, and spherical equivalent were recorded at each visit. The surgical results were investigated at 1 day, 1 week, and 1 month after surgery. RESULTS: Seventeen eyes treated with a new monofocal IOL enhanced for intermediate vision (ICB00 group) and 18 eyes treated with the standard monofocal IOL (AAB00 group) were included in this study. There were no significant differences in the baseline characteristics, including the type of underlying retinal disease, between the groups. There were no significant differences between the groups in terms of distant, intermediate, or near UCVA at day 1 and week 1 after surgery. However, at 1 month after surgery, the ICB00 group showed a significantly better intermediate vision improvement than the AAB00 group (p = 0.001). CONCLUSION: Even in patients with cataract accompanied by retinal disease, the use of the ICB00 IOL showed significant improvement in intermediate vision compared to the use of the AAB00 (standard monofocal) IOL. The ICB00 IOL might be a good option for patients with cataract and retinal disease in the era of increased intermediate vision needs in daily life.
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Catarata , Lentes Intraoculares , Facoemulsificación , Enfermedades de la Retina , Humanos , Implantación de Lentes Intraoculares/métodos , Estudios Retrospectivos , Catarata/complicaciones , Enfermedades de la Retina/complicaciones , Enfermedades de la Retina/cirugía , Diseño de Prótesis , Facoemulsificación/métodos , Satisfacción del PacienteRESUMEN
PURPOSE: To evaluate the clinical efficacy and safety of combination treatment with intense pulsed light (IPL), LipiFlow thermal pulsation, and manual meibomian gland expression (MGX) for patients with refractory meibomian gland dysfunction (MGD). METHODS: A total of 23 eyes from 23 patients who underwent combination treatment with IPL, LipiFlow, and MGX for refractory MGD were included in the study. All patients were followed up for 12 months after the first treatment. All patients were examined prior to treatment and underwent three sessions of IPL, LipiFlow, and MGX, a subsequent three sessions of IPL and MGX, and six subsequent sessions of MGX. Patients were examined 10 weeks, 6 months, and 12 months after the first treatment. The ophthalmologic examination included ocular surface disease index (OSDI), non-invasive tearfilm break-up time (NIBUT), lipid layer thickness (LLT), meibomian gland expressibility (MGE), meibomian gland morphology (meiboscore), and tear meniscus height (TMH). RESULTS: OSDI, NIBUT, LLT, and MGE significantly improved with treatment, which was maintained during the follow-up period. TMH and meiboscore remained unchanged throughout follow-up. No serious adverse events occurred during follow-up. CONCLUSIONS: Combination therapy with IPL, LipiFlow, and MGX was clinically effective and safe for the treatment of refractory MGD. Combination treatment could be considered for patients with insufficient improvement despite conventional MGD treatments, including IPL treatment with MGX.
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Síndromes de Ojo Seco , Disfunción de la Glándula de Meibomio , Humanos , Glándulas Tarsales/metabolismo , Disfunción de la Glándula de Meibomio/terapia , Fototerapia , Resultado del Tratamiento , Lípidos , Lágrimas/metabolismo , Síndromes de Ojo Seco/metabolismoRESUMEN
The purpose of this study was to investigate whether and how topical nerve growth factor (NGF) attenuates streptozotocin (STZ)-induced diabetic cataracts in vivo. Rats were randomly divided into three groups, including the normal control rat group, STZ-induced diabetic cataract rat group (DM group), and STZ-induced diabetic cataract rat group treated with 200 µg/mL recombinant rat ß-NGF (DM + NGF group). Cataract formation was evaluated by portable slit lamp biomicroscopy following pupil dilation at 8 weeks. The expression levels of NGF, aldose reductase (AR), and Na+/K+-ATPase in the lens epithelial cells (LECs) of the three groups were measured in the presence or absence of topical NGF. TUNEL-positive LECs were quantified to determine if hyperglycemia caused LEC apoptosis. At 8 weeks, the mean cataract score in the control group was significantly lower than that in DM and DM + NGF groups, and the score in the DM + NGF group was significantly lower than that in the DM group. At the equatorial zone and anterior central zone of lens, NGF and Na+/K+-ATPase expression levels were significantly decreased in the DM group; however, they were partially restored in the DM + NGF group. At the equatorial zone and anterior central zone of lens, AR expression and TUNEL-positive apoptotic LECs were significantly increased in the DM group compared with the control group, however, they were significantly decreased in the DM + NGF group. In conclusion, topical NGF could delay the progression of diabetic cataracts by attenuating polyol pathway activation and increasing Na+/K+-ATPase protein levels.
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Catarata/prevención & control , Diabetes Mellitus Experimental/prevención & control , Factor de Crecimiento Nervioso/uso terapéutico , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Administración Oftálmica , Animales , Apoptosis , Catarata/inducido químicamente , Catarata/enzimología , Catarata/patología , Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Experimental/enzimología , Diabetes Mellitus Experimental/patología , Hiperglucemia/metabolismo , Etiquetado Corte-Fin in Situ , Masculino , Soluciones Oftálmicas , Polímeros , Ratas , Ratas Sprague-Dawley , Estreptozocina/toxicidad , Regulación hacia ArribaRESUMEN
Two-photon microscopy (TPM) is a three dimensional (3D) microscopic technique based on nonlinear two-photon fluorescence, which has been tested as an alternative to reflectance confocal microscopy (RCM) for detecting fungal keratitis via optical imaging. Although TPM provided images with better contrast than RCM for fungal keratitis, its imaging speed was relatively low because of weak intrinsic signal. Moxifloxacin, a Food and Drug Administration (FDA)-approved antibiotic, was recently used as a cell-labeling agent for TPM. In this study, moxifloxacin was used to label fungal cells for TPM imaging of fungal keratitis models. Fungal cell suspensions and ex vivo fungal keratitis-affected rabbit corneas were prepared using two types of fungal pathogens, Aspergillus fumigatus and Candida albicans, and TPM imaging was performed both with and without moxifloxacin treatment. Fungal cells with enhanced fluorescence were clearly visible by TPM of moxifloxacin-treated fungal cell suspensions. TPM of moxifloxacin-treated fungal keratitis rabbit corneas revealed both the infecting fungal cells and corneal cells similar to those observed in TPM without moxifloxacin treatment, albeit with approximately 10-times enhanced fluorescence. Fungal cells were distinguished from corneal cells on the basis of their distinct morphologies. Thus, TPM with moxifloxacin labeling might be useful for the detection of fungal keratitis at the improved imaging speed.
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Aspergilosis/diagnóstico por imagen , Candidiasis/diagnóstico por imagen , Técnicas de Diagnóstico Oftalmológico , Infecciones Fúngicas del Ojo/diagnóstico por imagen , Queratitis/diagnóstico por imagen , Microscopía Confocal/métodos , Microscopía Fluorescente/métodos , Moxifloxacino/administración & dosificación , Coloración y Etiquetado/métodos , Animales , Modelos Animales de Enfermedad , Infecciones Fúngicas del Ojo/microbiología , Imagenología Tridimensional/métodos , Queratitis/microbiología , ConejosRESUMEN
BACKGROUND: Epidural blood patch (EBP) has been shown to be an effective treatment option for spontaneous intracranial hypotension (SIH). We investigated whether response to the EBP was related to the distance of the injection site from the leakage site in patients with SIH. METHODS: We reviewed patients with SIH who underwent EBP at a single hospital. Patients were assigned to group R (response after EBP) or group N (no response after EBP). We then analyzed the demographics, clinical characteristics, leakage site, leakage length, EBP injection level and distance from leakage site, and injected EBP volume. RESULTS: Sixty-two patients were included in the analysis. The overall response rate to EBP was 59.7% (37 patients). The leakage length and injection distance from the leakage site did not differ between the two groups. Age, gender, body mass index, leakage site, and EBP volume did not differ significantly between the two groups. CONCLUSION: The clinical effect of EBP in SIH was not affected by leakage length or injection distance to leakage site. Further large studies must be conducted to investigate the efficacy of targeted EBP for SIH.
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Parche de Sangre Epidural/métodos , Hipotensión Intracraneal/cirugía , Adulto , Anciano , Anciano de 80 o más Años , Índice de Masa Corporal , Pérdida de Líquido Cefalorraquídeo/etiología , Femenino , Humanos , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Estudios RetrospectivosRESUMEN
Chinese hamster ovary (CHO) cells are the most widely used host cell lines for the commercial production of therapeutic proteins including Fc-fusion proteins. During the culture of recombinant CHO (rCHO) cells, host cell proteins (HCPs), secreted from viable cells and released from dead cells, accumulate extracellularly, potentially impairing product quality. In this study, the HCPs that accumulated extracellularly in batch and fed-batch cultures of Fc-fusion protein-producing rCHO cell lines (DG-Fc and DUKX-Fc) were identified and quantified using nanoflow liquid chromatography-tandem mass spectrometry (LC-MS/MS), followed by gene ontology and functional analysis. When the proteome database of Cricetulus griseus was used as a reference to identify the HCPs, more HCPs were identified for DG-Fc (1632 HCPs in batch culture and 1733 HCPs in fed-batch culture) than for DUKX-Fc (1114 HCPs in batch culture and 1002 HCPs in fed-batch culture). Clustering analysis of HCPs, which were classified into four clusters according to their concentration profiles during culture, showed that the concentration profiles of HCPs affecting the quality of Fc-fusion proteins correlated with changes in Fc-fusion protein quality. Taken together, the dataset of HCPs obtained in this study using the two different rCHO cell lines provides insights into the determination of appropriate target proteins to be removed during the culture and purification steps so as to ensure good Fc-fusion protein quality. Biotechnol. Bioeng. 2017;114: 2267-2278. © 2017 Wiley Periodicals, Inc.
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Técnicas de Cultivo Celular por Lotes/métodos , Células CHO/fisiología , Perfilación de la Expresión Génica/métodos , Fragmentos Fc de Inmunoglobulinas/metabolismo , Proteoma/metabolismo , Proteínas Recombinantes de Fusión/biosíntesis , Animales , Células CHO/clasificación , Cricetulus , Medios de Cultivo , Ingeniería Genética/métodos , Fragmentos Fc de Inmunoglobulinas/química , Mapeo Peptídico/métodos , Proteoma/química , Proteínas Recombinantes de Fusión/químicaRESUMEN
Chinese hamster ovary (CHO) cell cultivation for production of therapeutic proteins is accompanied by production of metabolic wastes, mostly ammonia and lactate. To reduce ammonia production, the glutamine synthetase (GS) system was used to develop therapeutic monoclonal antibody (mAb)-producing CHO cells (SM-0.025). Additionally, the lactate dehydrogenase-A (LDH-A) was downregulated with shRNA to reduce lactate production in SM-0.025. The resulting mAb-producing cell lines (#2, #46, and #52) produced less ammonia than the host cell line during the exponential phase due to GS protein overexpression. LDH-A downregulation in SM-0.025 not only reduced lactate production but also further reduced ammonia production. Among the three LDH-A-downregulated clones, clone #2 had the highest mAb production along with significantly reduced specific lactate and ammonia production rates compared to those in SM-0.025. Waste reduction increased the galactosylation level of N-glycosylation, which improved mAb quality. LDH-A downregulation was also successfully applied to the host cell lines (CHO K1 and GS knockout CHO-K1). However, LDH-A downregulated host cells could not survive the pool-selection process wherein glutamine was excluded and methionine sulfoximine was added to the media. Taken together, LDH-A downregulation in the mAb-producing cell line generated with the GS system successfully reduced both ammonia and lactate levels, improving mAb galactosylation. However, LDH-A downregulation could not be applied to host cell lines because it hampered the selection process of the GS system.
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Amoníaco/metabolismo , Glutamato-Amoníaco Ligasa/metabolismo , L-Lactato Deshidrogenasa/genética , L-Lactato Deshidrogenasa/metabolismo , Ácido Láctico/metabolismo , Animales , Anticuerpos Monoclonales/biosíntesis , Anticuerpos Monoclonales/uso terapéutico , Células CHO , Supervivencia Celular , Cricetulus , Medios de Cultivo/química , Regulación hacia Abajo , Isoenzimas/genética , Isoenzimas/metabolismo , Lactato Deshidrogenasa 5 , ARN Interferente Pequeño , TransfecciónRESUMEN
This paper presents a thermal convection-based sensor fabricated using simple microelectromechanical systems (MEMS)-based processes. This sensor can be applied to both acceleration and inclination measurements without modifying the structure. Because the operating mechanism of the accelerometer is the thermal convection of a gas medium, a simple model is proposed and developed in which the performance of the thermal convection-based accelerometer is closely associated with the Grashof number, Gr and the Prandtl number, Pr. This paper discusses the experiments that were performed by varying several parameters such as the heating power, cavity size, gas media, and air pressure. The experimental results demonstrate that an increase in the heating power, pressure, and cavity size leads to an increase in the accelerometer sensitivity. However, an increase in the pressure and/or cavity size results in a decrease in the frequency bandwidth. This paper also discusses the fact that a working-gas medium with a large thermal diffusivity and small kinematic viscosity can widen the frequency bandwidth and increase the sensitivity, respectively.
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In response to the escalating concern over the effect of environmental factors on ocular health, this study aimed to investigate the impact of air pollution-associated particulate matter (PM) on ocular allergy and inflammation. C57BL/6 mice were sensitized with ovalbumin (OVA) topically and aluminum hydroxide via intraperitoneal injection. Two weeks later, the mice were challenged with OVA and exposed to PM. Three groups-naive, OVA, and OVA-sensitized with PM exposure (OVA + PM) groups-were induced to an Allergic Eye disease (AED) model. Parameters including clinical signs, histological changes, inflammatory cell infiltration, serum OVA-specific immunoglobulins E (IgE) levels, mast cells degranulation, cellular apoptosis and T-cell cytokines were studied. The results demonstrate that exposure with PM significantly exacerbates ocular allergy, evidenced by increased eye-lid edema, mast cell degranulation, inflammatory cytokines (IL-4, IL-5 and TNF-α), cell proliferation (Ki67), and serum IgE, polymorphonuclear leukocytes (PMN), and apoptosis and reduced goblet cells. These findings elucidate the detrimental impact of PM exposure on exacerbating the severity of AED. Noticeably, diminished goblet cells highlight disruptions in ocular surface integrity, while increased PMN infiltration with an elevated production of IgE signifies a systemic allergic response with inflammation. In conclusion, this study not only scientifically substantiates the association between air pollution, specifically PM, and ocular health, but also underscores the urgency for further exploration and targeted interventions to mitigate the detrimental effects of environmental pollutants on ocular surfaces.
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Dry eye disease (DED) is a growing health concern that impacts millions of individuals every year, and is associated with corneal injury, excessive oxidative stress and inflammation. Current therapeutic strategies, including artificial tears and anti-inflammatory agents, are unable to achieve a permanent clinical cure due to their temporary nature or adverse side effects. Therefore, here, we investigated the effectiveness of the topical administration of programmed death-ligand 1 (PD-L1) in the mouse model of DED. The model was generated in C57BL/6 mice by excising the extra orbital lacrimal gland and causing desiccation stress with scopolamine injections. Subsequently, either phosphate-buffered saline (3 µL/eye) or PD-L1 (0.5 µg/mL) was topically administered for 10 days. Tear volume was evaluated with phenol red thread, and corneal fluorescein staining was observed to quantify the corneal epithelial defect. Corneas were collected for histological analysis, and the expression levels of inflammatory signaling proteins such as CD4, CD3e, IL-17, IL-1ß, pIkB-α, pNF-kB and pERK1/2 were assessed through immunofluorescence and Western blot techniques. Our results demonstrate that desiccating stress-induced corneal epithelial defect and tear secretion were significantly improved by topical PD-L1 and could reduce corneal CD4+ T cell infiltration, inflammation and apoptosis in a DED mouse model by downregulating IL-17 production and ERK1/2-NFkB pathways.
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Síndromes de Ojo Seco , Epitelio Corneal , Animales , Ratones , Ratones Endogámicos C57BL , Antígeno B7-H1 , Interleucina-17 , Síndromes de Ojo Seco/tratamiento farmacológico , Modelos Animales de Enfermedad , InflamaciónRESUMEN
Genetic engineering approaches to inhibit cell death in Chinese hamster ovary (CHO) cell cultures have been limited primarily to anti-apoptosis engineering. Recently, autophagy has received attention as a new anti-cell death engineering target in addition to apoptosis. In order to achieve a more efficient protection of cells from the stressful culture conditions, the simultaneous targeting of anti-apoptosis and pro-autophagy in CHO cells (DG44) was attempted by co-overexpressing an anti-apoptotic protein, Bcl-2, and a key regulator of autophagy pathway, Beclin-1, respectively. Co-overexpression of Bcl-2 and Beclin-1 exhibited a longer culture period as well as higher viability during serum-free suspension culture, compared with the control (without co-overexpression of Bcl-2 and Beclin-1) and Bcl-2 overexpression only. In addition to the efficient inhibition of apoptosis by Bcl-2 overexpression, Beclin-1 overexpression successfully induced the increase in the autophagic marker protein, LC3-II, and autophagosome formation with the decrease in mTOR activity. Co-immunoprecipitation and qRT-PCR experiments revealed that the enforced expression of Beclin-1 increased Ulk1 expression and level of free-Beclin-1 that did not bind to the Bcl-2 despite the Bcl-2 overexpression. Under other stressful culture conditions such as treatment with sodium butyrate and hyperosmolality, co-overexpression of Bcl-2 and Beclin-1 also protected the cells from cell death more efficiently than Bcl-2 overexpression only, implying the potential of autophagy induction. Taken together, the data obtained here provide the evidence that pro-autophagy engineering together with anti-apoptosis engineering yields a synergistic effect and successfully enhances the anti-cell death engineering of CHO cells.
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Proteínas Reguladoras de la Apoptosis/biosíntesis , Apoptosis , Autofagia , Biotecnología/métodos , Proteínas Proto-Oncogénicas c-bcl-2/biosíntesis , Animales , Proteínas Reguladoras de la Apoptosis/genética , Células CHO , Técnicas de Cultivo de Célula/métodos , Supervivencia Celular , Cricetulus , Proteínas Proto-Oncogénicas c-bcl-2/genéticaRESUMEN
Intubation with videolaryngoscopy has become popular in various clinical settings. However, despite the use of a videolaryngoscope, difficult intubation still exists and intubation failure has been reported. This retrospective study assessed the efficacy of the 2 maneuvers in improving the glottic view during videolaryngoscopic intubation. The medical records of patients who underwent videolaryngoscopic intubation and whose glottal images were stored in electronic medical charts were reviewed. The videolaryngoscopic images were divided into 3 categories according to the applied optimization techniques as follows: conventional method, with the blade tip located in the vallecular; backward-upward-rightward pressure (BURP) maneuver; and epiglottis lifting maneuver. Four independent anesthesiologists scored the visualization of the vocal folds using the percentage of glottic opening (POGO, 0-100%) scoring system. A total of 128 patients with 3 laryngeal images were analyzed. The glottic view was the most improved in the epiglottis lifting maneuver among all the techniques. The median POGO scores were 11.3, 36.9, and 63.1 in the conventional method, BURP, and epiglottis lifting maneuver, respectively (Pâ <â .001). There were significant differences in the distribution of POGO grades according to the application of BURP and epiglottis lifting maneuvers. In the POGO grades 3 and 4 subgroups, the epiglottis lifting maneuver was more effective than the BURP maneuver in improving the POGO score Inadequate visualization of the vocal folds occurred even when intubation was performed using a videolaryngoscope. The application of optimization maneuvers, such as BURP and epiglottis lifting by the blade tip, could improve the glottic view.
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Glotis , Pliegues Vocales , Humanos , Estudios Retrospectivos , Registros Médicos , Intubación IntratraquealRESUMEN
AIM: To characterize the anti-inflammatory and anti-apoptotic effects of N-acetylcysteine (NAC) in streptozotocin (STZ)-induced diabetic rat corneal epithelium and human corneal epithelial cells (HCECs) exposed to a high-glucose environment. METHODS: HCECs were incubated in 0, 5, 50 mmol/L glucose medium, or 50 mmol/L glucose medium with NAC for 24h. Diabetes was induced in rats by intraperitoneal injection of 65 mg/kg STZ and some of these rats were topically administered NAC to corneas with 3 mice per group. We characterized receptor for advanced glycation end-products (RAGE) expression using immunofluorescence, and interleukin (IL)-1ß and cleaved caspase-3 (CCAP-3) expression using immunohistochemistry. Circulating tumor necrosis factor (TNF)-α concentration was measured by ELISA and cleaved poly-ADP ribose polymerase (PARP) concentration was quantified by Western blotting. Apoptotic cells were detected using TUNEL assay and annexin V and propidium iodide staining. RESULTS: Diabetic rats had higher expression of RAGE (2.46±0.13 fold), IL-1ß, and CCAP-3 in apoptotic cells of their corneas than control rats. The expression of RAGE (1.83±0.11 fold), IL-1ß, and CCAP-3, and the number of apoptotic cells, were reduced by topical NAC treatment. HCECs incubated in 50 mmol/L glucose medium showed high concentrations of TNF-α (310±2.00 pg/mL) and cleaved PARP (7.43±0.56 fold), and more extensive apoptosis than cells in 50 mmol/L glucose medium. However, the addition of NAC reduced the concentrations of TNF-α (153.67±2.31 pg/mL) and cleaved PARP (5.55±0.31 fold) and the number of apoptotic cells. CONCLUSION: NAC inhibits inflammation and apoptosis in the corneas of diabetic rats and HCECs maintained in a high-glucose environment.
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PURPOSE: To evaluate the clinical efficacy and safety of immediate manual meibomian gland expression (MGX) after LipiFlow thermal pulsation (TearScience Inc, Morrisville, NC) for obstructive meibomian gland dysfunction and to compare the LipiFlow only and MGX after LipiFlow. METHODS: Patients who underwent immediate manual MGX after LipiFlow or who received only LipiFlow treatment were included. Thirty eyes from 15 patients were enrolled in each group. All patients underwent 3 treatments at monthly intervals. All patients were followed up for 6 months after treatment. All patients were examined before and at 3 and 6 months after treatment. Examinations included the Ocular Surface Disease Index score, noninvasive tear film breakup time (NIBUT), lipid layer thickness (LLT), corneal and conjunctival staining, and tear meniscus height. RESULTS: The Ocular Surface Disease Index scores improved in both groups during the follow-up periods (P = 0.001 and P = 0.001). In the LipiFlow-only group, the NIBUT and LLT significantly improved at 3 months (P < 0.001 and P = 0.006) but deteriorated at 6 months. In the MGX after LipiFlow group, the NIBUT and LLT improved at 3 months (P < 0.001 and P < 0.001), and this improvement was maintained at 6 months. The improvement of NIBUT at 3 months was greater in the MGX after LipiFlow group (3.24 ± 1.16 to 9.25 ± 1.36 s) than in the LipiFlow-only group (3.78 ± 1.75 to 7.18 ± 2.70 s), and the improvements of the LLT at 6 months were greater in the MGX after LipiFlow group (30.27 ± 10.74 to 46.93 ± 20.81 µm) than in the LipiFlow-only group (34.70 ± 10.79 to 38.73 ± 14.70 µm). CONCLUSIONS: Both LipiFlow only and MGX after LipiFlow were clinically effective for obstructive meibomian gland dysfunction. However, the efficacy and persistence of treatment were greater in patients who received MGX after LipiFlow.
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Hipertermia Inducida/métodos , Disfunción de la Glándula de Meibomio/terapia , Glándulas Tarsales/metabolismo , Lágrimas/metabolismo , Adulto , Femenino , Humanos , Masculino , Disfunción de la Glándula de Meibomio/metabolismo , Persona de Mediana Edad , Estudios Retrospectivos , Resultado del Tratamiento , Adulto JovenRESUMEN
AIM: To evaluate clinical outcomes of unilateral implantation of a diffractive multifocal intraocular lens (IOL) in patients with contralateral monofocal IOL. METHODS: Twenty-two patients who already had implantation of a monofocal IOL in unilateral eye underwent implantation of a diffractive multifocal IOL in contralateral eye were enrolled. After 1, 6, and 12mo, uncorrected and distant corrected distant visual acuity (UCDVA and DCDVA), uncorrected and distant corrected intermediate-visual acuity (UCIVA and DCIVA), uncorrected and distant corrected near visual acuity (UCNVA and DCNVA), and contrast sensitivity were obtained. Halo/glare symptoms, spectacle dependence, and patient satisfaction were also evaluated. RESULTS: The mean age was 67.86±7.25y and the average interval between two IOL implantations was 645.82±878.44d. At 1mo, binocular UCDVA was lower than 0.20 logMAR in 76% of patients (mean 0.12±0.13 logMAR), which increased to 90% by 6 and 12mo. The binocular UCDVA was significantly better than the monocular results (P<0.05) at 1, 6, and 12mo. Additionally, UCNVA was lower than 0.40 logMAR in 82% of patients, increasing to 90% by 6 and 12mo. Mean UCNVA in the multifocal IOL implanted eye was statistically significantly better than that in the monofocal IOL implanted eye (P<0.05) at 1, 6, and 12mo. About 5% of patients at 1 and 6mo, reported "severe glare or halo". Patient satisfaction rates were 95% and 91% at 6 and 12mo, respectively. CONCLUSION: Unilateral implantation of multifocal IOL in patients with a contralateral, monofocal IOL implantation results in high patient satisfaction rate, with low severe glare or halo rate during follow-up. It can represent a good option for patients who have previously had a monofocal IOL implantation regardless of two year interval duration between two IOL implantations.
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PURPOSE: To compare the protective effects of cyclosporine A emulsion (Restasis: 0.05% cyclosporine A) (CsAE) and cyclosporine A cationic emulsion (Ikervis: 0.1% cyclosporine A) (CsACE) on cellular inflammation, apoptosis, proliferation, and survival in an in vitro dry eye model. METHODS: The concentration of CsA in CsAE and CsACE was verified using a liquid chromatography tandem mass spectrometry system. Human corneal epithelial cells were subjected to desiccation stress. Human corneal epithelial cells were incubated with or without 3 groups of cyclosporine A medium (CsAE 1:50, CsACE 1:50, and CsACE 1:100). p-NF-κB p65, p-IκBα, Bax, Bcl-xL, p-Erk1/2, and p-Akt levels were determined using Western blots, and TNF-α levels were quantified using an enzyme-linked immunosorbent assay. RESULTS: The CsA concentration of CsACE 1:100 was nearly the same as that of CsAE 1:50. Compared with CsAE 1:50 (0.78 ± 0.19 fold), the p-NF-κB p65 level was further reduced in CsACE 1:50 (0.38 ± 1.20 fold) and 1:100 (0.29 ± 0.11 fold) as well as p-IκBα. Levels of TNF-α were also lower in CsACE 1:50 and 1:100 than in CsAE 1:50. Induction of the apoptotic protein Bax was significantly decreased in CsACE 1:50 and 1:100 compared with CsAE 1:50, whereas that of the antiapoptotic protein Bcl-xL was increased in CsACE 1:50 and 1:100. p-ERK1/2 and p-Akt levels were higher in CsACE 1:50 and 1:100 than in CsAE 1:50. CONCLUSIONS: CsACE had more potent anti-inflammatory and antiapoptotic effects than CsAE in a transwell desiccation stress model. CsACE also enhanced proliferation and survival factors under desiccation stress compared with CsAE in this in vitro dry eye model.
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Apoptosis/efectos de los fármacos , Ciclosporina/uso terapéutico , Desecación/métodos , Síndromes de Ojo Seco/tratamiento farmacológico , Epitelio Corneal/patología , Estrés Mecánico , Lágrimas/metabolismo , Línea Celular , Síndromes de Ojo Seco/patología , Emulsiones , Epitelio Corneal/efectos de los fármacos , Humanos , Inmunosupresores/uso terapéuticoRESUMEN
The protein kinase Snf1 is a major component of the glucose derepression pathway in yeast and a regulator of gene expression for the cell wall degrading enzyme (CWDE) in some plant pathogenic fungi. To address the molecular function of Snf1 in Magnaporthe oryzae, which causes the rice blast disease, MoSNF1 was cloned and functionally characterized using gene knock-out strategies. MoSNF1 functionally complemented the growth defect of the yeast snf1 mutant on a non-fermenting carbon source. However, the growth rate of the Deltamosnf1 mutant on various carbon sources was reduced independent of glucose, and the expression of the CWDE genes in the mutant was induced during derepressing condition like the wild type. The pre-penetration stage including conidial germination and appressorium formation of the Deltamosnf1 was largely impaired, and the pathogenicity of the Deltamosnf1 was significantly reduced. Most strikingly, the Deltamosnf1 mutant produced only a few conidia and had a high frequency of abnormally shaped conidia compared to the wild type. Our results suggest that MoSNF1 is a functional homolog of yeast Snf1, but its contribution to sporulation, vegetative growth and pathogenicity is critical in M. oryzae.
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Proteínas Fúngicas/genética , Magnaporthe/genética , Oryza/microbiología , Enfermedades de las Plantas/microbiología , Esporas Fúngicas/genética , Proteínas Fúngicas/fisiología , Prueba de Complementación Genética , Magnaporthe/patogenicidad , Magnaporthe/fisiología , Mutación , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/fisiología , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crecimiento & desarrollo , Esporas Fúngicas/fisiología , Virulencia/genéticaRESUMEN
OBJECTIVE: We compared the effects of two anesthetics, isoflurane and propofol, on the nuclear or cytosolic localization of nuclear factor erythroid 2-related factor 2 (Nrf2), mRNA expression levels of excitatory amino acid carrier 1 (EAAC1), and glutathione (GSH) protein levels in the rat hippocampus. METHODS: Fifty-two adult male Sprague-Dawley rats were randomly divided into three groups: a control group, a group that received propofol for 240 minutes (P240), and a group that received isoflurane for 240 minutes (I240). We compared GSH protein and EAAC1 mRNA expression levels in the rat hippocampus and evaluated Nrf2 content in cytosolic and nuclear fractions in the three groups. RESULTS: GSH protein and EAAC1 mRNA expression levels were significantly higher in the I240 and P240 groups compared with the control group. The I240 and P240 groups showed lower Nrf2 protein levels in the cytosolic fractions, but higher levels in the nuclear fractions compared with the control group. CONCLUSION: Treatment with isoflurane or propofol may enhance GSH production by facilitating translocation of Nrf2 into the nucleus and increasing EAAC1mRNA expression in the rat hippocampus. Isoflurane and propofol show similar profiles in EAAC1 expression-associated GSH production.
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Transportador 3 de Aminoácidos Excitadores/genética , Glutatión/metabolismo , Hipocampo/metabolismo , Isoflurano/farmacología , Propofol/farmacología , Anestesia General , Animales , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Citosol/efectos de los fármacos , Citosol/metabolismo , Transportador 3 de Aminoácidos Excitadores/metabolismo , Hipocampo/efectos de los fármacos , Masculino , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Neuronas/efectos de los fármacos , Neuronas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas Sprague-DawleyRESUMEN
Purpose: To evaluate the effect of diquafosol on corneal epithelium in a dry eye model using Transwell culture and a scopolamine-induced dry eye rat model. Methods: Desiccation stress induced in an in vitro dry eye model using human corneal epithelial cells was used, and the cells were incubated with or without diquafosol media diluted at 1:100. Reactive oxygen species (ROS) generation was measured using 2',7'-dichlorofluorescein diacetate (DCFH-DA). Apoptosis was analyzed, and levels of phosphorylated Erk1/2, phosphorylated p90RSK, phosphorylated Akt, IκB-α, and NF-κB-p65 were determined. Levels of IL-1ß, TNF-α, IL-6, IL-8, and GM-CSF were quantified. To investigate the in vivo effects of diquafosol, we induced dry eye in Wistar rats using scopolamine hydrobromide. The rats were divided into three groups: control, dry eye, and dry eye diquafosol; topical DIQUAS was applied four times daily for 28 days. We used immunohistochemistry to detect the levels of phosphorylated Erk1/2, phosphorylated p90RSK, and IL-1ß, and used the TUNEL assay in corneal tissue. Results: The distribution of highly fluorescent dichlorofluorescein and the proportion of annexin V- and PI-positive cells decreased in the diquafosol medium. Diquafosol increased the levels of phospho-Erk1/2, phospho-90RSK, phospho-Akt, and IκB-α, whereas it significantly decreased the levels of NF-κB-p65, IL-1ß, and TNF-α. In vivo, apoptosis was enhanced in dry eye group. This response was markedly reduced and the level of phosphorylated p90RSK and phosphorylated ERK1/2 were upregulated and IL-1ß was downregulated by DIQUAS. Conclusions: Diquafosol treatment reduced intracellular ROS levels, apoptosis, and inflammation, all of which were increased in the dry eye model through desiccation.
Asunto(s)
Apoptosis/efectos de los fármacos , Síndromes de Ojo Seco/tratamiento farmacológico , Queratitis/prevención & control , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Polifosfatos/uso terapéutico , Proteínas Quinasas S6 Ribosómicas 90-kDa/metabolismo , Nucleótidos de Uracilo/uso terapéutico , Animales , Western Blotting , Citocinas/metabolismo , Modelos Animales de Enfermedad , Síndromes de Ojo Seco/metabolismo , Epitelio Corneal/efectos de los fármacos , Femenino , Inflamación/metabolismo , Inflamación/prevención & control , Queratitis/metabolismo , Soluciones Oftálmicas/uso terapéutico , Fosforilación , Agonistas del Receptor Purinérgico P2Y/uso terapéutico , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Generation of recombinant Chinese hamster ovary (rCHO) cell lines is critical for the production of therapeutic proteins. However, the high degree of phenotypic heterogeneity among generated clones, referred to as clonal variation, makes the rCHO cell line development process inefficient and unpredictable. Here, we investigated the major genomic causes of clonal variation. We found the following: (1) consistent with previous studies, a strong variation in rCHO clones in response to hypothermia (33 vs 37 °C) after random transgene integration; (2) altered DNA sequence of randomly integrated cassettes, which occurred during the integration process, affecting the transgene expression level in response to hypothermia; (3) contrary to random integration, targeted integration of the same expression cassette, without any DNA alteration, into three identified integration sites showed the similar response of transgene expression in response to hypothermia, irrespective of integration site; (4) switching the promoter from CMV to EF1α eliminated the hypothermia response; and (5) deleting the enhancer part of the CMV promoter altered the hypothermia response. Thus, we have revealed the effects of integration methods and cassette design on transgene expression levels, implying that rCHO cell line generation can be standardized through detailed genomic understanding. Further elucidation of such understanding is likely to have a broad impact on diverse fields that use transgene integration, from gene therapy to generation of production cell lines.