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Ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry(UPLC-Q/TOF-MS) was employed to examine the impact of Coptidis Rhizoma(CR) and its processed products on the metabolism in the rat model of oral ulcer due to excess heat and to compare the effectiveness of CR and its three products. Male SD rats were randomly allocated to the sham-operation(Sham), model(M, oral ulcer due to excess heat), CR, wine/Zingiberis Rhizoma Recens/Euodiae Fructus processed CR(wCR/zCR/eCR), and Huanglian Shangqing Tablets(HST) groups. Except the Sham group, the other groups were administrated with Codonopsis Radix-Astragali Radix decoction by gavage for two consecutive weeks. The anal temperature and water consumption of rats were monitored throughout the modeling period of excess heat. Following the completion of the modeling, oral ulcer was modeled with acetic acid. Hematoxylin-eosin(HE) staining was employed to observe the mucosal pathological changes in oral ulcer. A colorimetric assay was employed to determine the serum level of glutathione peroxidase(GSH-Px). Enzyme-linked immunosorbent assay(ELISA) was conducted to determine the levels of tumor necrosis factor-alpha(TNF-α), interleukin-6(IL-6), interleukin-1ß(IL-1ß), superoxide dismutase(SOD), and malondialdehyde(MDA) in the serum. The non-targeted metabolomics analysis based on UPLC-Q/TOF-MS was conducted on the serum samples. Metabolic profiles were then built, and the potential biomarkers were screened by principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA). The Mev software was used to establish a heat map and conduct cluster analysis on the quantitative results of the markers. The online databases including MBRole, KEGG, and MetaboAnalyst were used for pathway enrichment analysis and metabolic network building. The experimental results showed that the modeling led to pathological damage to the oral mucosa, elevated serum levels of TNF-α, IL-6, IL-1ß, and MDA, and lowered levels of SOD and GSH-Px in rats. The drug administration recovered all the indices to varying extents, and wCR exhibited the best performance. Non-targeted metabolomics identified 48 differential metabolites including 27 metabolites in the positive ion mode and 21 metabolites in the negative ion mode. Five enriched pathways were common, including glycerophospholipid metabolism, linoleic acid metabolism, and tyrosine metabolism. Conclusively, CR and its three processed products could alleviate the inflammation and oxidative stress injury in rats suffering from oral ulcers due to excess heat by regulating lipid and amino acid metabolism. Notably, wCR demonstrated the most significant therapeutic effect.
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Medicamentos Herbarios Chinos , Úlceras Bucales , Ratas , Masculino , Animales , Medicamentos Herbarios Chinos/farmacología , Úlceras Bucales/tratamiento farmacológico , Interleucina-6 , Calor , Factor de Necrosis Tumoral alfa , Ratas Sprague-Dawley , Metabolómica/métodos , Cromatografía Líquida de Alta Presión , Superóxido Dismutasa , BiomarcadoresRESUMEN
INTRODUCTION: Retinitis pigmentosa (RP) caused by the photoreceptor cell degeneration is currently incurable and leads to partial or complete blindness eventually. 3,5-dimethoxy-4-hydroxy myricanol (DM) is a novel compound isolated from the leaves of Micromelum integerrimum, with proliferative activities on NIH3T3 cells. This study was to investigate whether DM could mitigate retinal degeneration of rd10 mice, a well-characterized mouse model of RP. MATERIALS AND METHOD: Rd10 mice were treated with DM daily by intraperitoneal injection from postnatal day 12 (P12) to P26. Electroretinography (ERG) reflects the mass response of photoreceptor cells and was used to test the outer retinal function after DM treatment. Haematoxylin and Eosin staining was used to show the retinal morphology and evaluate the rod photoreceptor cell loss. TUNEL assay was used to detect the apoptosis-positive cells. Inflammatory factors were measured by ELISA to show the inflammatory response. Real-time PCR and western blot were applied to measure the gene and protein change to explore the underlying mechanisms. RESULTS: Results showed that DM significantly improved the retinal function by increasing the ERG amplitude, preserving the retinal morphology, reducing photoreceptor cell apoptosis, decreasing inflammatory response, and inhibiting endoplasmic reticulum stress in rd10 mice. CONCLUSION: This is the first time when the protective effects of DM against photoreceptor cell degeneration of rd10 mice have been demonstrated, providing scientific rationale to develop DM as a potential agent to treat RP.
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Antiinflamatorios/uso terapéutico , Diarilheptanoides/uso terapéutico , Células Fotorreceptoras de Vertebrados/efectos de los fármacos , Degeneración Retiniana/tratamiento farmacológico , Animales , Apoptosis/efectos de los fármacos , Ratones Endogámicos C57BL , Ratones MutantesRESUMEN
Exposure to reactive oxygen species (ROS) leads to the development and progression of retinal degenerative diseases. However, the exact mechanisms are not fully understood. In this article, the role of angiotensin II type 1 receptor (AT1R) signaling in H(2)O(2)-induced retinal damage was examined. Mouse photoreceptor-derived 661 W cells were treated with the AT1R blockers valsartan, losartan and candesartan before exposure to H(2)O(2). Cell viability, intracellular ROS level, mitochondrial membrane potential (MMP), cytochrome-c level, DNA fragmentation, caspase activity and gene expression were detected. Pre-treatment of 661 W cells with AT1R blockers significantly decreased H(2)O(2)-mediated toxicity and reduced the ROS level. In addition, apoptosis-related biochemical indicators showed that pre-incubation of AT1R blockers would elevate the MMP, decrease the release of cytochrome-c and formation of DNA fragmentation, and inhibit activities of caspase-3 and caspase-9 in exogenous H(2)O(2)-treated 661 W cells. Moreover, treatment with AT1R blockers suppressed the expression of Egr1, Fosl1 and Lox12. These results suggest that AT1R signaling mediates H(2)O(2)-induced apoptosis, at least partially through generating the ROS and increasing the levels of proapoptotic molecules in 661 W cells. AT1R blockade may provide a new therapeutic approach for preventing oxidative stress-induced retinal neural damage.
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Bloqueadores del Receptor Tipo 1 de Angiotensina II/farmacología , Peróxido de Hidrógeno/toxicidad , Células Fotorreceptoras de Vertebrados/efectos de los fármacos , Degeneración Retiniana/prevención & control , Animales , Apoptosis/efectos de los fármacos , Caspasas/metabolismo , Técnicas de Cultivo de Célula , Línea Celular , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Citocromos c/metabolismo , Expresión Génica/efectos de los fármacos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/genética , Células Fotorreceptoras de Vertebrados/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptor de Angiotensina Tipo 1/genética , Degeneración Retiniana/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Chloride channels (ClCs) have gained worldwide interest because of their molecular diversity, widespread distribution in mammalian tissues and organs, and their link to various human diseases. Nine different ClCs have been molecularly identified and functionally characterized in mammals. ClC-2 is one of nine mammalian members of the ClC family. It possesses unique biophysical characteristics, pharmacological properties, and molecular features that distinguish it from other ClC family members. ClC-2 has wide organ/tissue distribution and is ubiquitously expressed. Published studies consistently point to a high degree of conservation of ClC-2 function and regulation across various species from nematodes to humans over vast evolutionary time spans. ClC-2 has been intensively and extensively studied over the past two decades, leading to the accumulation of a plethora of information to advance our understanding of its pathophysiological functions; however, many controversies still exist. It is necessary to analyze the research findings, and integrate different views to have a better understanding of ClC-2. This review focuses on ClC-2 only, providing an analytical overview of the available literature. Nearly every aspect of ClC-2 is discussed in the review: molecular features, biophysical characteristics, pharmacological properties, cellular function, regulation of expression and function, and channelopathies.
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Canales de Cloruro/metabolismo , Cloruros/metabolismo , Animales , Canales de Cloruro CLC-2 , Canalopatías/fisiopatología , Canales de Cloruro/química , Canales de Cloruro/genética , Humanos , Miocitos Cardíacos/metabolismo , Regiones Promotoras Genéticas , Estructura Terciaria de ProteínaRESUMEN
BACKGROUND: Overhanging filtering bleb is a common complication after trabeculectomy and surgical repair is an effective treatment when the patient presents with apparent symptoms. Filtering bleb relevant infection including in the filtering bleb itself and even endophthalmitis in some severe cases has been reported. However, corneal fungal infection after filtering bleb repair is rarely reported. CASE SUMMARY: A 57-year-old Chinese man who had sensations of redness and foreign body sensations in the left eye 3 wk after repair of overhanging filtering bleb. 3 wk ago, due to sensations of a foreign body in the left eye for 3 years with worsening for 3 mo. The patient was diagnosed as overhanging filtering bleb and underwent a repair of overhanging filtering bleb. Postoperative, the filtering bleb formed well and the intraocular pressure is normal. But the patient gradually develop redness, pain and a grey infiltrate of the cornea in the eye. Finally it developed into fungal corneal ulcer. Through asking the medical history, we found the patient had irregularly self-medicated for years with glucocorticoid eye drops for years to relieve the foreign body sensation in the eye caused by filtering bleb overhanging. Because the glucocorticoid eye drops he used years ago had provide normal sensation to the eye. After 3 mo of anti-fungal treatment, the inflammation was controlled. CONCLUSION: In addition to avoiding the development of overhanging filtering bleb after trabeculectomy, the present case report also suggests that clinicians should pay more attention to the patient's ocular self-medication history. Particularly in patients with a history of glaucoma or eye surgery. Because these patients may be exposed to more types of eye drops than other individuals, they may select the wrong medications for long-term use, based on their previous experience.
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Three pairs of undescribed diarylpentanoid enantiomers (1-3) and five undescribed phenylpropanoids (4-8), along with seven known compounds, were isolated from the roots of Anthriscus sylvestris. The structures of compounds (1-8) were determined by analysis of their 1D and 2D NMR spectra, HRESIMS, and electronic circular dichroism. In addition, the inhibitory activities against hypoxia-stimulated pulmonary arterial smooth muscle cells abnormal proliferation were evaluated by MTT assay. The mRNA expression levels of Bcl-2, BAX, Caspase3, and IL-6 were detected by quantitative real-time PCR. The results showed that compounds (-)-1, (+)-1, (-)-2, (+)-3, 4, 8-10, 14, and 15 inhibited the abnormal proliferation of PASMCs by regulating the levels of apoptosis and inflammatory factors.
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Apiaceae , Extractos Vegetales , Extractos Vegetales/química , Arteria Pulmonar , Proliferación CelularRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Asthma is a chronic airway inflammatory disease. Current treatment of mainstream medications has significant side effects. There is growing evidence that the refractoriness of asthma is closely related to common changes in the lung and intestine. The lungs and intestines, as sites of frequent gas exchange in the body, are widely populated with gas signaling molecules NO and CO, which constitute NO-CO metabolism and may be relevant to the pathogenesis of asthma in the lung and intestine. The Chinese herbal formula Tingli Dazao Xiefei Decoction (TD) is commonly used in clinical practice to treat asthma with good efficacy, but there are few systematic evaluations of the efficacy of asthma on NO-CO metabolism, and the mode of action of its improving effect on the lung and intestine is unclear. AIM OF THE STUDY: To investigate the effect of TD on the lung and intestine of asthmatic rats based on NO-CO metabolism. MATERIALS AND METHODS: In vivo, we established a rat asthma model by intraperitoneal injection of sensitizing solution with OVA atomization, followed by intervention by gavage administration of TD. We simultaneously examined alterations in basal function, pathology, NO-CO metabolism, inflammation and immune cell homeostasis in the lungs and intestines of asthmatic rats, and detected changes in intestinal flora by macrogenome sequencing technology, with a view to multi-angle evaluation of the treatment effects of TD on asthmatic rats. In vitro, lung cells BEAS-2B and intestinal cells NCM-460 were used to establish a model of lung injury causing intestinal injury using LPS and co-culture chambers, and lung cells or intestinal cells TD-containing serum was administered to intervene. Changes in inflammatory, NO-CO metabolism-related, cell barrier-related and oxidative stress indicators were measured in lung cells and intestinal cells to evaluate TD on intestinal injury by way of amelioration and in-depth mechanism. RESULTS: In vivo, our results showed significant basal functional impairment in the lung and intestine of asthmatic rats, and an inflammatory response, immune cell imbalance and intestinal flora disturbance elicited by NO-CO metabolic disorders were observed (P < 0.05 or 0.01). The administration of TD was shown to deliver a multidimensional amelioration of the impairment induced by NO-CO metabolic disorders (P < 0.05 or 0.01). In vitro, the results showed that LPS-induced lung cells BEAS-2B injury could cause NO-CO metabolic disorder-induced inflammatory response, cell permeability damage and oxidative stress damage in intestinal cells NCM-460 (P < 0.01). The ameliorative effect on intestinal cells NCM-460 could only be exerted when TD-containing serum interfered with lung cells BEAS-2B (P < 0.01), suggesting that the intestinal ameliorative effect of TD may be exerted indirectly through the lung. CONCLUSION: TD can ameliorate NO-CO metabolism in the lung and thus achieve the indirectly amelioration of NO-CO metabolism in the intestine, ultimately achieving co-regulation of lung and intestinal inflammation, immune imbalance, cellular barrier damage, oxidative stress and intestinal bacterial disorders in asthma in vivo and in vitro. Targeting lung and intestinal NO-CO metabolic disorders in asthma may be a new therapeutic idea and strategy for asthma.
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Asma , Enfermedades Intestinales , Enfermedades Metabólicas , Ratas , Animales , Ratones , Lipopolisacáridos/farmacología , Pulmón , Intestinos/patología , Estrés Oxidativo , Inflamación/patología , Enfermedades Intestinales/patología , Enfermedades Metabólicas/metabolismo , Ovalbúmina/farmacología , Ratones Endogámicos BALB C , Modelos Animales de EnfermedadRESUMEN
Nuclear factor- (erythroid-derived 2-) like 2 (Nrf2) is a regulator of many processes of life, and it plays an important role in antioxidant, anti-inflammatory, and antifibrotic responses and in cancer. This review is focused on the potential mechanism of Nrf2 in the occurrence and development of ocular diseases. Also, several Nrf2 inducers, including noncoding RNAs and exogenous compounds, which control the expression of Nrf2 through different pathways, are discussed in ocular disease models and ocular cells, protecting them from dysfunctional changes. Therefore, Nrf2 might be a potential target of protecting ocular cells from various stresses and preventing ocular diseases.
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Oftalmopatías/genética , Oftalmopatías/terapia , Factor 2 Relacionado con NF-E2/uso terapéutico , Animales , Oftalmopatías/patología , Humanos , Ratones , Estrés OxidativoRESUMEN
Eyelid conditioning, including delay eyelid conditioning and trace eyelid conditioning, has been used extensively to study neural structures and mechanisms of learning and memory as a form of associative learning. In the present study, microcurrent electrical stimulation was used to stimulate the medial prefrontal cortex (mPFC) to induce delay eyelid conditioning in guinea pigs. The acquisition rate and relative latency of the conditioned eyelid response (CR) and the startle eyelid response (SR) were analyzed. The mPFC sites in the guinea pigs were examined under a light microscope following Nissl staining. In addition, the expression of Fos protein in neurons was detected using immunohistochemistry and western blot analysis. The results indicated that the acquisition rates of CR and SR were increased significantly (P<0.05), whilst the relative latencies of CR and SR were decreased significantly (P<0.05). Lesions were observed in the mPFC regions in the training group when compared with the control group. In addition, immunohistochemistry and western blot analysis revealed that Fos expression was significantly increased in the training group when compared with the sham group for the control and resident-intruder test guinea pigs (P<0.05). Therefore, the enhancement of delay eyelid conditioning by microcurrent electrical stimulation of the mPFC in guinea pigs was triggered by the expression of Fos protein. The observations of the present study further expand the understanding of conditioned reflexes and may aid future investigations into the formation of eyelid conditioning and the mechanisms underlying the circuit in various conditions.
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OBJECTIVES: The purpose of this study was to investigate the role of chloride channel protein 2 (ClC-2) in glutamate-induced apoptosis in the retinal ganglion cell line (RGC-5). MATERIALS AND METHODS: RGC-5 cells were treated with 1 mM glutamate for 24 hr. The expression of ClC-2, Bax, and Bcl-2 was detected by western blot analysis. Cell survival and apoptosis were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and flow cytometry assays, respectively. Caspase-3 and -9 activities were determined by a colorimetric assay. The roles of ClC-2 in glutamate-induced apoptosis were examined by using ClC-2 complementary deoxyribonucleic acid (cDNA) and small inference ribonucleic acid (RNA) transfection technology. RESULTS: Overexpression of ClC-2 in RGC-5 cells significantly decreased glutamate-induced apoptosis and increased cell viability, whereas silencing of ClC-2 with short hairpin (sh) RNA produced opposite effects. ClC-2 overexpression increased the expression of Bcl-2, decreased the expression of Bax, and decreased caspase-3 and -9 activation in RGC-5 cells treated with glutamate, but silencing of ClC-2 produced opposite effects. CONCLUSION: Our data suggest that ClC-2 chloride channels might play a protective role in glutamate-induced apoptosis in retinal ganglion cells via the mitochondria-dependent apoptosis pathway.
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We have cloned ClC-3B, a novel alternative splicing variant of ClC-3 (ClC-3A) that is expressed predominantly in epithelial cells. ClC-3B has a different, slightly longer C-terminal end than ClC-3A and contains a consensus motif for binding to the second PDZ (PSD95/Dlg/ZO-1) domain of the epithelium-specific scaffolding protein EBP50. Both in vitro and in vivo binding assays demonstrate interaction between ClC-3B and EBP50. C127 mouse mammary epithelial cells transfected with ClC-3B alone showed diffuse immunoreactivity for ClC-3B in the cytoplasmic region. In contrast, when EBP50 was cotransfected with ClC-3B, strong immunoreactivity for ClC-3B appeared at the leading edges of membrane ruffles. Patch-clamp experiments revealed that cotransfection of ClC-3B and EBP50 resulted in a remarkable increase in outwardly rectifying Cl- channel (ORCC) activities at the leading edges of membrane ruffles in C127 cells. The electrophysiological properties of the ClC-3B-induced ORCCs are similar to those of ORCCs described in native epithelial cells. When cystic fibrosis transmembrane conductance regulator (CFTR) was cotransfected with ClC-3B and EBP50, ClC-3B-dependent ORCCs were activated via the protein kinase A-dependent pathway. These findings indicate that ClC-3B is itself a CFTR-regulated ORCC molecule or its activator.
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Proteínas Portadoras/metabolismo , Canales de Cloruro/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Fosfoproteínas/metabolismo , Intercambiadores de Sodio-Hidrógeno , 1-Metil-3-Isobutilxantina/farmacología , Células 3T3 , Empalme Alternativo , Animales , Células CHO , Calcimicina/farmacología , Proteínas Portadoras/genética , Línea Celular , Canales de Cloruro/genética , Colforsina/farmacología , Cricetinae , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Expresión Génica , Humanos , Ionóforos/farmacología , Potenciales de la Membrana/efectos de los fármacos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Fosfoproteínas/genética , Unión Proteica , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , TransfecciónRESUMEN
Akita Prefecture has had the highest suicide rate in Japan for the past 9 years. To obtain further information on suicide attempts by self-immolation in Akita, we performed a statistical analysis of patients in this prefecture who attempted to burn themselves. Over the past 6 years, 541 patients suffering from burns were transferred to medical emergency units. Of these, 35 (6.5%) attempted self-immolation, most of whom were between 20 and 60 years of age. Women over 50 years of age outnumbered men in the same age group. All 35 patients sustained flame burns. The total burn surface area (TBSA), burn index (BI), rate of inhalation injury, and mortality rate were all significantly higher in the patients who attempted self-immolation than in those with nonsuicidal burns. Most (68.6%) of the self-immolation attempts were made indoors. Because the Japanese are not generally a very religious people, training to help them cultivate a philosophy of life and educating them in moral science to help them form a personal view of life and death may be necessary to prevent suicides.
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Quemaduras/epidemiología , Intento de Suicidio/estadística & datos numéricos , Adolescente , Adulto , Distribución por Edad , Anciano , Anciano de 80 o más Años , Quemaduras/patología , Quemaduras/psicología , Femenino , Humanos , Puntaje de Gravedad del Traumatismo , Japón/epidemiología , Masculino , Persona de Mediana Edad , Pronóstico , Estaciones del AñoRESUMEN
OBJECTIVE: To study the efficacy of electrolyzed oxidizing water (EOW) and hydrocolloid occlusive dressings in the acceleration of epithelialization in excised burn-wounds in rats. METHODS: Each of the anesthetized Sprague-Dawley rats (n=28) was subjected to a third-degree burn that covered approximately 10% of the total body surface area. Rats were assigned into four groups: Group I (no irrigation), Group II (irrigation with physiologic saline), Group III (irrigation with EOW) and Group IV (hydrocolloid occlusive dressing after EOW irrigation). Wounds were observed macroscopically until complete epithelialization was present, then the epithelialized wounds were examined microscopically. RESULTS: Healing of the burn wounds was the fastest in Group IV treated with hydrocolloid occlusive dressing together with EOW. Although extensive regenerative epidermis was seen in each Group, the proliferations of lymphocytes and macrophages associated with dense collagen deposition were more extensive in Group II, III and IV than in Group I. These findings were particularly evident in Group III and IV. CONCLUSIONS: Wound Healing may be accelerated by applying a hydrocolloid occlusive dressing on burn surfaces after they are cleaned with EOW.
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Quemaduras/terapia , Coloides/uso terapéutico , Apósitos Oclusivos , Agua/administración & dosificación , Animales , Masculino , Ratas , Ratas Sprague-Dawley , Cicatrización de HeridasRESUMEN
Normal mammalian terminal erythroid differentiation is a precisely regulated process during which the progenitor cells execute particular programs to form a mature erythrocytic phenotype. In the present study, it was found that RbAp48, a histone-binding protein associated with retinoblastoma protein, was upregulated during terminal erythroid maturation in vivo and in vitro. This indicated that RbAp48, at least in part, participated in the regulation of murine erythropoiesis. Following sodium butyrate (SB) induction, murine erythroleukemia (MEL) cells began to re-enter erythroid differentiation and the ratio of differentiated cells reached ~80% at 72 h. The erythroid maturation-related mRNA expression of α-globin, ß-globin and glycophorin A (GPA) was increased markedly, which indicated that SB induced MEL differentiation. During MEL differentiation, the RbAp48 level showed a 1.5-fold increase at 72 h, and the globin transcription factor (GATA)-1 level was also upregulated in the early stage of differentiation. By contrast, the c-Myc level was gradually downregulated in MEL differentiation. Using an immunofluorescence assay, the results of the study directly showed that the average fluorescence intensity of RbAp48 in each cell reached an almost 1.7-fold increase at 72 and 96 h. This was consistent with the western blot results of RbAp48 during MEL differentiation. In addition, reduced expression of RbAp48 by RNA inference decreased SB-induced MEL differentiation by ~20%, indicating that a high level of RbAp48 was essential for MEL differentiation. Taken together, these results established a functional link between RbAp48 and erythroid differentiation.
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The chloride channel protein 2 (CLC2) is important in maintaining the volume of trabecular meshwork cells by adjusting the outflow of aqueous solutions and maintaining the fluid balance. However, little is known concerning the functions of CLC2 in the cytoskeleton, specifically in human trabecular meshwork (HTM) cells. In the present study, two CLC-2 specific siRNAs (siRNA1 and siRNA2) that target CLC-2 mRNA were designed. The siRNAs were transfected into the HTM cells and the results showed that siRNA1 in particular decreased the expression of CLC2 by ~45%. Furthermore, an siRNA1mediated CLC-2 knockdown significantly reconstructed the actin cytoskeleton and formed crosslinked actin networks. In addition, the downregulation of the expression of CLC2 was associated with increased TGFß and Smad2 activities in the HTM cells following 24 h of transfection. In conclusion, these results suggest that CLC2 knockdown promotes trabecular meshwork cytoskeletal disorders and may activate the TGFß/Smad signaling pathway. Thus, CLC2 may be a promising and potential novel therapeutic strategy for combating primary openangle glaucoma.
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Canales de Cloruro/metabolismo , Fibras de Estrés/metabolismo , Malla Trabecular/patología , Actinas/metabolismo , Canales de Cloruro CLC-2 , Células Cultivadas , Canales de Cloruro/genética , Citoesqueleto/metabolismo , Expresión Génica , Técnicas de Silenciamiento del Gen , Glaucoma/patología , Humanos , ARN Interferente Pequeño/genética , Transducción de Señal , Proteínas Smad/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Vinculina/metabolismo , beta Catenina/metabolismoRESUMEN
Sex determination is a vital part of the medico-legal system but can be difficult in cases where the integrity of the body has been compromised. The purpose of this study was to develop a technique for sex assessment from measurements of the first lumber vertebrate. Twenty-nine linear measurements and five ratios were collected from 113 Chinese adult males and 97 Chinese adult females using digital three-dimensional anthropometry methods. By using discriminant analysis, we found that 23 linear measurements and two ratios identified sexual dimorphism (P<0.01), with predictive accuracy ranging from 57.1% to 86.6%. Using a stepwise method of discriminant function analysis, we found three dimensions predicted sex with 88.6% accuracy: (a) upper end-plate width (EPWu), (b) left pedicle height (PHl), and (c) middle end-plate depth (EPDm). This study shows that a single first lumber vertebra can be used for this purpose, and that the discriminant equation will help forensic determination of sex in the Chinese population.
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Vértebras Lumbares/anatomía & histología , Determinación del Sexo por el Esqueleto/métodos , Adulto , Pueblo Asiatico , China , Análisis Discriminante , Femenino , Antropología Forense , Humanos , Imagenología Tridimensional , Vértebras Lumbares/diagnóstico por imagen , Masculino , Tomografía Computarizada EspiralRESUMEN
AIM: To quantity and accurately evaluate pulmonary microvascular permeability. METHODS: To measure capillary fluid filtration coefficient (Kf) by using the gravimetric method in isolated perfused rabbit lungs. RESULTS: The amount of Kf is 4.78 +/- 0.73 mg x min(-1) x cmH2O(-1) x g(-1). CONCLUSION: The advantage of this method is exactly and directly. This method shows the important effect on realizing the pulmonary physiologic condition and evaluating the extent of acute lung edema and injury.
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Permeabilidad Capilar , Pulmón/irrigación sanguínea , Perfusión , Animales , Técnicas In Vitro , Masculino , Circulación Pulmonar , ConejosRESUMEN
Modulation of ion permeability during the cell cycle is one of the key events in cell cycle progression. We have compared the effects of K+ and Cl- channel blockers on the cell cycle in synchronous and asynchronous NIH3T3 cells. The Cl- channel blocker 5-N-2-(3-phenylpropylamino) benzoic acid (NPPB; 0.2 mM) inhibited entry into S phase in synchronous cells but not in asynchronous cells, while the K+ channel blocker 4-aminopyridine (4-AP) showed similar inhibitory effects in both conditions. In NIH3T3 cells synchronized by serum deprivation/replenishment, G0-to-G1 transition occurred within 8 h after serum addition, and the G1/S checkpoint at 10-14 h. NPPB applied only at 0-8 or 8-14 h after serum addition inhibited entry into S phase. Cl- permeability measured as 125I efflux increased at 4 and 10 h after serum addition. Ki-67-negative cells, which represent quiescent G0 phase cells, progressively decreased in number until 8 h after serum addition. The Cl- channel blockers (NPPB and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid [DIDS]) but not the K+ channel blocker (4-AP) significantly decreased the rate of reduction in number of Ki-67-negative cells. These data indicate that an increase in Cl- permeability plays an important role in reentry of quiescent cells into the proliferating phase, in addition to the known effects on passage through the G1/S checkpoint.
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Canales de Cloruro/antagonistas & inhibidores , Fibroblastos/citología , Fase de Descanso del Ciclo Celular/efectos de los fármacos , Células 3T3 , 4-Aminopiridina/farmacología , Inhibidores de la Angiogénesis/farmacología , Animales , Proteínas Sanguíneas/farmacología , División Celular/efectos de los fármacos , Fibroblastos/química , Fibroblastos/metabolismo , Fase G1/efectos de los fármacos , Radioisótopos de Yodo , Antígeno Ki-67/análisis , Ratones , Nitrobenzoatos/farmacología , Bloqueadores de los Canales de Potasio/farmacología , Fase S/efectos de los fármacosRESUMEN
Cl(-) channel activities vary during the cell cycle and are thought to play various roles including regulation of cell volume. We have shown previously that ClC-2 channels are directly phosphorylated and functionally regulated by the M phase-specific cyclin-dependent kinase p34(cdc2)/cyclin B. We investigate here to determine whether the expression levels of ClC-2 channel protein vary during the cell cycle. Immunoblot and immunocytochemical analyses of cells cycle-synchronized by serum depletion/replenishment reveal that ClC-2 channel protein is expressed predominantly at M phase in cells with two nuclei and a clear constriction ring, whereas RNA blot analysis shows that ClC-2 mRNA expression does not change during the cell cycle. Ubiquitin assays reveal that the ClC-2 channels are ubiquitinated at M phase, whereas the magnitude of ubiquitination is suppressed by incubation with olomoucine, an inhibitor of p34(cdc2)/cyclin B, and it is almost completely abolished in ClC-2 channels having an S632A mutation, which cannot be phosphorylated by p34(cdc2)/cyclin B, indicating that ubiquitination of ClC-2 channels requires phosphorylation by M phase-specific p34(cdc2)/cyclin B. Regulation at the post-transcriptional level, including phosphorylation-dependent ubiquitination, may contribute to M phase-specific expression of ClC-2 channels. Cell cycle-dependent regulation of expression at the protein level in addition to the regulation of function suggests that the ClC-2 channel plays a physiological role in the cell cycle.