RESUMEN

Background: Extracellular cold-inducible RNA-binding protein (eCIRP) plays a vital role in the inflammatory response during cerebral ischaemia. However, the potential role and regulatory mechanism of eCIRP in traumatic brain injury (TBI) remain unclear. Here, we explored the effect of eCIRP on the development of TBI using a neural-specific CIRP knockout (KO) mouse model to determine the contribution of eCIRP to TBI-induced neuronal injury and to discover novel therapeutic targets for TBI. Methods: TBI animal models were generated in mice using the fluid percussion injury method. Microglia or neuron lines were subjected to different drug interventions. Histological and functional changes were observed by immunofluorescence and neurobehavioural testing. Apoptosis was examined by a TdT-mediated dUTP nick end labelling assay in vivo or by an annexin-V assay in vitro. Ultrastructural alterations in the cells were examined via electron microscopy. Tissue acetylation alterations were identified by non-labelled quantitative acetylation via proteomics. Protein or mRNA expression in cells and tissues was determined by western blot analysis or real-time quantitative polymerase chain reaction. The levels of inflammatory cytokines and mediators in the serum and supernatants were measured via enzyme-linked immunoassay. Results: There were closely positive correlations between eCIRP and inflammatory mediators, and between eCIRP and TBI markers in human and mouse serum. Neural-specific eCIRP KO decreased hemispheric volume loss and neuronal apoptosis and alleviated glial cell activation and neurological function damage after TBI. In contrast, eCIRP treatment resulted in endoplasmic reticulum disruption and ER stress (ERS)-related death of neurons and enhanced inflammatory mediators by glial cells. Mechanistically, we noted that eCIRP-induced neural apoptosis was associated with the activation of the protein kinase RNA-like ER kinase-activating transcription factor 4 (ATF4)-C/EBP homologous protein signalling pathway, and that eCIRP-induced microglial inflammation was associated with histone H3 acetylation and the α7 nicotinic acetylcholine receptor. Conclusions: These results suggest that TBI obviously enhances the secretion of eCIRP, thereby resulting in neural damage and inflammation in TBI. eCIRP may be a biomarker of TBI that can mediate the apoptosis of neuronal cells through the ERS apoptotic pathway and regulate the inflammatory response of microglia via histone modification.

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OBJECTIVE: MicroRNAs have been reported to be involved in the regulation of tumor progression. This study investigated the role of miR-152-3p in bladder cancer development. METHODS: A total of 67 bladder cancer cases were enrolled. miR-152-3p expression in bladder cancer tissues and cells were detected using quantitative reverse transcriptase polymerase chain reaction. Bladder cancer cells were transfected by miR-152-3p mimic and inhibitor to up-regulate and down-regulate miR-152-3p expression. After transfection, cell counting kit-8 assay, flow cytometry, Brdu staining assay, transwell experiment and wound healing assay were conducted to research the effect of miR-152-3p up-regulation/down-regulation on bladder cancer cell viability, apoptosis, proliferation, invasion and migration abilities. The expression of high-mobility group protein A2 (HMGA2) and autophagy-related proteins was researched using Western blot. The interaction between miR-152-3p and HMGA2 was explored by dual luciferase reporter gene assay. RESULTS: Low miR-152-3p expression in tumor tissues bladder cancer patients was associated with poor prognosis. miR-152-3p expression was abnormally down-regulated in bladder cancer cells. miR-152-3p up-regulation inhibited viability, proliferation, invasion, migration but promoted apoptosis of bladder cancer cells. miR-152-3p down-regulation showed the opposite effects. miR-152-3p up-regulation suppressed the expression of Beclin 1 and LC3II/LC3I proteins in bladder cancer cells, but miR-152-3p down-regulation increased them. HMGA2 was target of miR-152-3p, which could be directly inhibited by miR-152-3p. HMGA2 up-regulation reversed the inhibitory effect of miR-152-3p on bladder cancer cell malignant phenotype. CONCLUSION: miR-152-3p inhibited malignant phenotype of bladder cancer cell lines via suppressing HMGA2 expression.

Asunto(s)

MicroARNs , Neoplasias de la Vejiga Urinaria , Humanos , Neoplasias de la Vejiga Urinaria/genética , Autofagia/genética , Vejiga Urinaria , Línea Celular , Proteínas del Grupo de Alta Movilidad , Proliferación Celular/genética , MicroARNs/genética

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To investigate if a magnetic resonance imaging (MRI)-based model reduced postoperative biochemical failure (BF) incidence in patients with prostate cancer (PCa). From June 2018 to January 2020, we retrospectively analyzed 967 patients who underwent prostate bi-parametric MRI and radical prostatectomy (RP). After inclusion criteria were applied, 446 patients were randomized into research (n = 335) and validation cohorts (n = 111) at a 3:1 ratio. In addition to clinical variables, MRI models also included MRI parameters. The area under the curve (AUC) of receiver operating characteristic and decision curves were analyzed. The risk of postoperative BF, defined as persistently high or re-elevated prostate serum antigen (PSA) levels in patients with PCa with no clinical recurrence. In the research (age 69 [63-74] years) and validation cohorts (age 69 [64-74] years), the postoperative BF incidence was 22.39% and 27.02%, respectively. In the research cohort, the AUC of baseline and MRI models was 0.780 and 0.857, respectively, with a significant difference (P < 0.05). Validation cohort results were consistent (0.753 vs. 0.865, P < 0.05). At a 20% risk threshold, the false positive rate in the MRI model was lower when compared with the baseline model (31% [95% confidence interval (CI): 9-39%] vs. 44% [95% CI: 15-64%]), with the true positive rate only decreasing by a little (83% [95% CI: 63-94%] vs. 87% [95% CI: 75-100%]). 32 of 100 RPs can been performed, with no raise in quantity of patients with missed BF. We developed and verified a MRI-based model to predict BF incidence in patients after RP using preoperative clinical and MRI-related variables. This model could be used in clinical settings.

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Próstata , Neoplasias de la Próstata , Anciano , Humanos , Masculino , Imagen por Resonancia Magnética/métodos , Próstata/patología , Antígeno Prostático Específico , Prostatectomía , Neoplasias de la Próstata/diagnóstico por imagen , Neoplasias de la Próstata/cirugía , Estudios Retrospectivos , Persona de Mediana Edad

RESUMEN

BACKGROUND: Cold inducible RNA-binding protein (CIRP) is a key protein in the hypothermic therapy. Highly expressed CIRP exerts a neuroprotective effect on neurons. The aim of this study is to provide the evidence of the protective effects of CIRP on the glial cells and explore the downstream pathway of CIRP. RESULTS: The results of this study demonstrated that the cell viability of the glial cells with CIRP overexpression was increased significantly compared to the control. With CIRP overexpression, the epidermal growth factor (EGF) mRNA expression was found increasing significantly and the mRNA expressions of derived neurotrophic factor (BDNF), bcl-2, vascular endothelial growth factor (VEGF) and nerve growth factor (NGF) were not upregulated compared to the control. EGF and CIRP co-expression was demonstrated on the glial cells. With CIRP expression, EGF expression on the glial cells was increased statistically compared to the control. CONCLUSION: CIRP overexpression increases the cell viability of the glial cells, exerting a neuroprotective effect. EGF expression is activated on the glial cells with CIRP overexpression, implying a pathway of CIRP neuroprotection via EGF activation.

Asunto(s)

Factor de Crecimiento Epidérmico , Fármacos Neuroprotectores , Supervivencia Celular , Factor de Crecimiento Epidérmico/metabolismo , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Factor A de Crecimiento Endotelial Vascular , Neuroglía/metabolismo , ARN Mensajero/genética

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Novel reactive dyes with mono- and bi-acyl fluoride reactive groups have been designed and synthesized, which are obtained by using 2-amino-8-naphthol-6-sulfonic acid or 1-amino-8-naphthol-3,6-disulfonicacid as the coupling component and 4-aminobenzoyl fluoride (PABF) as the diazo component. Their structures have been defined by nuclear magnetic resonance spectroscopy and ultraviolet-visible spectra (UV-Vis). The novel reactive dyes were evaluated on cotton by using the exhaust dyeing method. The properties were examined in detail, and the results showed that the dye concentration of 4% (o.w.f), pH = 9, and salt-free was the most effective condition. The fixation of the novel reactive dyes on cotton was 60.27% and 64.13%, respectively. The micro-fluorine-containing reactive dyes have favorable dyeing properties owing to the covalent bond formed between the reactive group of dyes and the functional group of cotton fibers, which can achieve salt-free dyeing of cotton.

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Colorantes , Fluoruros , Colorantes/química , Fibra de Algodón , Naftoles , Textiles

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Large area requirements and huge energy consumption restrict the applications of microalgae in wastewater treatment. In this study, in-situ nutrient removal was tested using a floating permeable nutrients uptake system with pore sizes of 1, 5, 10, and 40 µm, and Chlorella sorokiniana and Scenedesmus acuminatus. Results showed that N transfer rate across FPNUS varied with membrane pore size and N-type. Average transfer rate of NH4+-N, NO3--N, and NO2--N across 1 µm membrane was 2.6, 14.6, and 2.3 mg m-2h-1, respectively, sufficient to support microalgal growth. The NH4+-N and NO3--N removal rate in shrimp wastewater reached 1.32 and 1.88 mg L-1d-1, comparable to some BNR processes used in RAS. According to the developed area ratio prediction model, FPNUS to pond area ratio of 21% is sufficient to balance N loading of 0.05 mg L-1d-1. These results indicate extraordinary potential of in-situ nutrient removal from wastewaters using FPNUS.

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Chlorella , Microalgas , Acuicultura , Biomasa , Nitrógeno/análisis , Nutrientes , Fósforo , Aguas Residuales

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Background: Clear cell renal cell carcinoma (ccRCC) is the main subtype of renal cell carcinoma and has different prognoses, especially in patients with metastasis. Here, we aimed to establish a novel model to predict overall survival (OS) and surgical benefit of ccRCC patients with distant metastasis. Methods: Using data from the Surveillance, Epidemiology, and End Results (SEER) databases, we identified 2185 ccRCC patients with distant metastasis diagnosed from 2010 to 2015. Univariate and multivariate Cox analysis were used to identify significant prognostic clinicopathological variables. By integrating these variables, a prognostic nomogram was constructed and evaluated using C-indexes and calibration curves. The discriminative ability of the nomogram was measured by analyses of receiver operating characteristic (ROC) curve. A risk stratification model was built according to each patient's total scores. Kaplan-Meier curves were performed in the low-, intermediate- and high-risk groups to evaluate the survival benefit of surgery. Results: Eight clinicopathological variables were included as independent prognostic factors in the nomogram: grade, marital status, T stage, N stage, bone metastasis, brain metastasis, liver metastasis, and lung metastasis. The nomogram had a better discriminative ability for predicting OS than Tumor-Node-Metastasis (TNM) stage. The C-index was 0.71 (95% CI 0.68-0.74) in the training cohort. The calibration plots demonstrated that the nomogram-based predictive outcomes had good consistency with the actual prognosis results. Total nephrectomy improved prognosis in both the low-risk and intermediate-risk groups, but partial nephrectomy could only benefit the low-risk group. Conclusions: We constructed a predictive nomogram and risk stratification model to evaluate prognosis in ccRCC patients with distant metastasis, which was valuable for prognostic stratification and making therapeutic decisions.

RESUMEN

Exosomal microRNAs (miRNAs) are suggested to reflect molecular changes occurring in their cells of origin and are potential indicators in the early detection of cancers. This study aimed to determine whether certain exosomal miRNAs from tumor tissue can be used as noninvasive biomarkers for clear cell renal cell carcinoma (ccRCC). Based on ccRCC miRNA expression profiles and the literature, we selected six miRNAs (miR-210, miR-224, miR-452, miR-155, miR-21, and miR-34a) and analyzed their expression in tissues, sera, and serum exosomes through quantitative real-time polymerase chain reaction in hypoxia-induced (with CoCl2 ) renal cell lines. miR-210, miR-224, miR-452, miR-155, and miR-21 were upregulated in tumor tissues compared with normal tissues. Serum miR-210 and miR-155 levels were higher in patients with ccRCC than in healthy controls (HCs). Furthermore, only exosomal miR-210 was significantly upregulated in patients with ccRCC than in HCs. Moreover, receiver operating characteristic (ROC) curve analysis revealed an area under the ROC curve of 0.8779 (95% confidence interval, 0.7987-0.9571) and a sensitivity and specificity of 82.5% and 80.0%, respectively. Moreover, exosomal miR-210 was upregulated at an advanced stage, and Fuhrman grade and metastasis decreased significantly one month after surgery. Acute hypoxia exposure activates miR-210 and release of exosomes with upregulated miR-210 in both normal and tumor RCC cell lines and interferes with vacuole membrane protein 1 mRNA expression, especially in the metastatic ccRCC cell line. In conclusion, Serum exosomal miR-210 originating from tumor tissue has potential as a novel noninvasive biomarker for the detection and prognosis of ccRCC.

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The cyanobacterial genera Raphidiopsis and Cylindrospermopsis morphologically resembled each other and are difficult to distinguish, especially when heterocytes do not form in Cylindrospermopsis species. The phylogeny based on multiple gene loci sequences in previous studies cannot divide the strains of these two genera into separate clusters. In the present study, four gene loci 16S rRNA, ITS-L, cpcBA-IGS and rpoC1, sequenced from many Chinese strains (44 Cylindrospermopsis strains and 16 Raphidiopsis strains), were analyzed to infer the phylogenetic relationship between the two genera. According to the 16S rRNA, cpcBA-IGS and rpoC1 gene phylogeny, Cylindrospermopsis and Raphidiopsis strains are intermixed and divide into two or three groups. By contrast, the ITS-L analysis implied that they respectively form two separate and unique clusters. A fragment of 7bp (RAGAAACT) in ITS-L was conserved for Raphidiopsis strains, which could be distinguished from Cylindrospermopsis strains. A pair of primers targeting the ITS-L fragment specific to Raphidiopsis strains were designed and further verified.

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Hyperglycemia may be associated with worse functional outcomes in patients with primary intracerebral hemorrhage. We performed a systematic review and meta-analysis to investigate the relationship between hyperglycemia and mortality risk in patients with primary intracerebral hemorrhage. We searched PubMed and Embase databases for studies investigating the association between hyperglycemia and mortality risk in patients with primary intracerebral hemorrhage. We estimated the pooled relative risk (RR) with its 95% confidence interval (95% CI) to assess the impact of hyperglycemia on mortality risk. Seventeen studies with a total of 6527 primary intracerebral hemorrhage patients were included. Meta-analysis of those studies showed that hyperglycemia significantly increased risk of mortality in patients with primary intracerebral hemorrhage (RR = 2.36, 95% CI 1.79-3.12). Subgroup analysis by time of follow-up showed that hyperglycemia significantly increased risk of short-term mortality (RR = 3.97, 95% CI 2.13-7.43) and long-term mortality (RR = 1.53, 95% CI 1.14-2.05). The RR of mortality for per 1-mmol/L increment in glucose level was 1.14 (95% CI 1.06-1.22). In patients with primary intracerebral hemorrhage, hyperglycemia significantly increases risk of both short-term mortality and long-term mortality.

Asunto(s)

Hemorragia Cerebral/complicaciones , Hiperglucemia/complicaciones , Hiperglucemia/mortalidad , Glucosa/metabolismo , Humanos , Sesgo de Publicación , Riesgo , Factores de Riesgo

RESUMEN

BACKGROUND: Pituitary adenoma is a common intracranial tumor in neurosurgery. Some pituitary adenomas have the characteristics of invasive growth make them unable to be removed completely by surgery leading to easy relapse. Discoidin domain receptor l (DDR1) is a new kind of tyrosine kinase receptor on the cell surface. DDR1 can be activated by tumor microenvironment signal in tumorigenesis, increasing MMP-2/9 expression and promoting the invasive ability of tumor cells. Anoxia can promote tumor growth and metastasis. This study investigated the impact of anoxic environment DDR1 expression in pituitary adenoma. MATERIAL AND METHODS: A primary hypoxia pituitary adenoma cell model was established and treated with DDR1 inhibitor nilotinib. Real-time PCR and Western blot were used to detect DDR1 mRNA and protein expression. ELISA was used to detect MMP-2/9 changes. MTT method was used to detect pituitary adenoma cell proliferation. We used a transwell chamber to determine pituitary adenoma cell invasion ability. RESULTS: DDR1 mRNA and protein were significantly overexpressed under hypoxia (P<0.05). MMP-2 and MMP-9 expression was obviously increased in supernatant (P<0.05). Pituitary adenoma cell proliferation and invasive ability improved markedly under hypoxia (P<0.05). Nilotinib could reduce DDR1 expression, decrease MMP-2 and MMP-9 expression, and inhibit pituitary adenoma cells proliferation and invasion. CONCLUSIONS: Hypoxia can increase DDR1 expression in pituitary adenoma cells, leading to improved MMP-2 and MMP-9 secretion, and promoting pituitary adenoma cell proliferation and invasion.

Asunto(s)

Adenoma/genética , Adenoma/metabolismo , Hipoxia de la Célula/genética , Hipoxia de la Célula/fisiología , Neoplasias Hipofisarias/genética , Neoplasias Hipofisarias/metabolismo , Proteínas Tirosina Quinasas Receptoras/genética , Proteínas Tirosina Quinasas Receptoras/metabolismo , Adenoma/patología , Adulto , Antineoplásicos/farmacología , Proliferación Celular/efectos de los fármacos , Receptor con Dominio Discoidina 1 , Femenino , Expresión Génica/efectos de los fármacos , Humanos , Masculino , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Persona de Mediana Edad , Invasividad Neoplásica , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias Hipofisarias/patología , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Pirimidinas/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Neoplásico/genética , ARN Neoplásico/metabolismo , Células Tumorales Cultivadas , Microambiente Tumoral

RESUMEN

Cold-inducible RNA-binding protein (CIRP) is induced by mild hypothermia in several mammals, but the precise mechanism by which CIRP mediates hypothermia-induced neuroprotection remains unknown. We aimed to investigate the molecular mechanisms by which CIRP protects the nervous system during mild hypothermia. Rat cortical neurons were isolated and cultured in vitro under mild hypothermia (32°C). Apoptosis was measured by annexin V and propidium iodide staining, visualized by flow cytometry. Neuron ultrastructure was visualized by transmission electron microscopy. CIRP overexpression and knockdown were achieved via infection with pL/IRES/GFP-CIRP and pL/shRNA/F-CIRP-A lentivirus. RT(2) Profiler PCR Array Pathway Analysis and western blotting were used to evaluate the effects of CIRP overexpresion/knockdown on the neurons׳ transcriptome. Neuron late apoptosis was significantly reduced at day 7 of culture by 12h hypothermia, but neuron ultrastructure remained relatively intact. RT(2) Profiler PCR Array Pathway Analysis of 84 apoptosis pathway-associated factors revealed that mild hypothermia and CIRP overexpression induce similar gene expression profiles, specifically alterations of genes implicated in the mitochondrial apoptosis pathway. Mild hypothermia-treated neurons up-regulated 12 and down-regulated 38 apoptosis pathway-associated genes. CIRP-overexpressing neurons up-regulated 15 and down-regulated 46 genes. CIRP-knocked-down hypothermia-treated cells up-regulated 9 and down-regulated 40 genes. Similar results were obtained at the protein level. In conclusion, CIRP may inhibit neuron apoptosis through the suppression of the mitochondria apoptosis pathway during mild hypothermia.

Asunto(s)

Apoptosis/fisiología , Proteínas y Péptidos de Choque por Frío/metabolismo , Mitocondrias/fisiología , Neuronas/fisiología , Proteínas de Unión al ARN/metabolismo , Animales , Western Blotting , Células Cultivadas , Corteza Cerebral/fisiología , Corteza Cerebral/ultraestructura , Proteínas y Péptidos de Choque por Frío/genética , Citometría de Flujo , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Vectores Genéticos , Hipotermia/genética , Hipotermia/patología , Hipotermia/fisiopatología , Lentivirus/genética , Análisis por Micromatrices , Microscopía Electrónica de Transmisión , Mitocondrias/ultraestructura , Neuronas/ultraestructura , Reacción en Cadena de la Polimerasa , ARN Interferente Pequeño , Proteínas de Unión al ARN/genética , Ratas Wistar , Regulación hacia Arriba/genética

RESUMEN

OBJECTIVE: To evaluate the effect of auscultation, partial pressure of carbon dioxide in end-expiration (P(ET)CO2), transillumination technique to judge whether the endotracheal tube is misplaced into the esophagus. METHODS: A blinded randomized controlled trial was conducted. Sixty patients with American Society of Anesthesiology (ASA) grade I - II undergoing endotracheal intubation in Fengxian Central Hospital admitted from September 2014 to February 2015 were enrolled. Two endotracheal tubes with the same size were respectively inserted into the trachea and esophagus for the same depth after general anesthesia by the same person. Two blinded anesthetists with different experience checked the tube position using three methods including auscultation, P(ET)CO2, and transillumination technique, respectively. The order of the tubes tested (trachea or esophagus) and the method used were randomized according to randomise numbers table. The experienced anesthetists conducted the test first, followed by an inexperienced anesthetist conducting the same methods. The numbers of right and wrong determinations with different methods by different anesthetists were recorded. RESULTS: Sixty patients underwent the procedures for 180 times, with intratracheal intubation for 90 times, and esophageal intubation for 90 times. It was shown that the results were not different in two groups [96.7% (174/180) vs. 92.2% (166/180), χ2 = 3.500, P = 0.057]. By using auscultation, the correct rate of experienced anesthetist was higher than that of inexperienced (95.0% vs. 78.3%, χ2 = 5.786, P = 0.013). Using P(ET)CO2, both anesthetists were correct in all cases, and the accuracy was 100%. Using transillumination, the experienced anesthetist was mistaken in 3 cases (accuracy was 95.0%), while the inexperienced mistook in 1 case (accuracy was 98.3%), and no significant difference was found between two groups χ2 = 0.500, P = 0.250). The correct rate of using transillumination was significantly higher than that of using auscultation χ2 = 7.563, P = 0.004). The sensitivity and specificity of the auscultation was 70.0% and 80.0%, that of transillumination technique was 96.7% and 93.3%, and P(ET)CO2 was 100%, respectively, for two groups. CONCLUSIONS: P(ET)CO2 is the most reliable method for determining tube position, and it is superior to auscultation and transillumination. Transillumination technique is superior to auscultation, irrespective of anesthetists' experience, while the accuracy of auscultation showed an obvious relationship with the anesthetists' experience.

Asunto(s)

Auscultación , Dióxido de Carbono/análisis , Intubación Intratraqueal/métodos , Transiluminación , Anestesia General , Esófago , Humanos , Variaciones Dependientes del Observador , Presión Parcial , Sensibilidad y Especificidad , Tráquea

RESUMEN

BACKGROUND: Cold-inducible RNA-binding protein (CIRP) is induced in response to hypothermia, where it exerts neuroprotective effects. Our preliminary studies revealed that it inhibits H2O2-induced apoptosis in rat neurons. In the current study, we report effective expression and purification approaches for the synthesis of CIRP, and assess its potential protective effects against oxidative stress. METHODS: CIRP-encoding was expressed using the prokaryotic expression system pGEX-4T-1, and SP-Sepharose and Sephacryl S-200 columns were used to purify rCIRP. To mimic ischemia/reperfusion injury-associated oxidative stress, neuro2a cells (N2a) were pre-treated with rCIRP for 2h, followed by hydrogen peroxide (H2O2 60 µmol/ml) for 24h. Cell viability was then quantified using an MTT assay. In addition, western blotting was performed to measure the cell cycle related signal transduction pathways. RESULTS: N2a cells exhibited decreased viability following H2O2 treatment, whereas rCIRP significantly improved viability following H2O2 treatment. CIRP also accelerated cell cycle progression from S to G2/M phase in cultured mouse neuroblastoma cells. In addition, CIRP increased levels of p-ERK and p-Akt, and also re-activated the cell cycle-related protein cyclin D1 and c-Myc. These results suggest that CIRP activated the Akt and ERK signal transduction pathways in N2a cells. CONCLUSIONS: Our findings suggest that CIRP could exert protective effects against oxidative stress, and that it might be a novel neuroprotective agent.

Asunto(s)

Fármacos Neuroprotectores/administración & dosificación , Proteínas de Unión al ARN/administración & dosificación , Proteínas de Unión al ARN/metabolismo , Animales , Ciclo Celular/efectos de los fármacos , Ciclo Celular/fisiología , Hipoxia de la Célula/efectos de los fármacos , Hipoxia de la Célula/fisiología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Clonación Molecular , Ciclina D1/metabolismo , Escherichia coli , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Expresión Génica , Peróxido de Hidrógeno , Ratones , Fármacos Neuroprotectores/aislamiento & purificación , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiología , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-myc/metabolismo , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/aislamiento & purificación , Proteínas Recombinantes/administración & dosificación , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo

RESUMEN

Hypoxia-ischemia (HI) encephalopathy is a frequent cause of disability and mortality with limited therapeutic options. Here, we collected peripheral blood mononuclear cells (PB-MNCs) from healthy donors and labeled them with CM-DiI before implanting these cells by tail-vein injection into rats at day 3 after hypoxia-ischemia (HI). For immune-suppression the animals received daily injections of cyclosporine throughout the experiment, commencing 24h before cell transplantation. Then we observed the PB-MNCs by fluorescent microscopy, examined motor function of rats by rotarod and cylinder tests, measured the lesion volume using image-pro plus software, and analyzed the apoptosis of neural cells in HI rats by tunnel assay. The results showed PB-MNCs could survive in the brain of hosts, migrate to the damage area and express neural marker. In addition, The HI rats that received PB-MNCs showed a reduction in motor function impairment, lesion volume and neural cell apoptosis. To better understand the mechanism of cell migration, PB-MNCs were also injected into normal rats via tail-vein. The expression of stromal cell-derived factor-1 (SDF-1) in the brain of normal and HI rats was measured by RT- PCR and western-blot, while the response of PB-MNCs in vitro to HI or normal brain extracts were measured by cell migration assay. Collectively these data suggest that the migration of PB-MNCs is directed to the damaged brain through an SDF-1-dependent pathway. Our results suggest that intravenous transplantation of PB-MNCs may be a feasible candidate for HI therapy.

Asunto(s)

Hipoxia-Isquemia Encefálica/terapia , Leucocitos Mononucleares/trasplante , Animales , Apoptosis , Encéfalo/metabolismo , Encéfalo/patología , Movimiento Celular , Supervivencia Celular , Quimiocina CXCL12/metabolismo , Factor Estimulante de Colonias de Granulocitos/farmacología , Movilización de Célula Madre Hematopoyética , Humanos , Hipoxia-Isquemia Encefálica/metabolismo , Hipoxia-Isquemia Encefálica/patología , Leucocitos Mononucleares/metabolismo , Actividad Motora , Ratas , Ratas Sprague-Dawley

RESUMEN

BACKGROUND: The RNA-binding motif protein 3 (RBM3), which is transcriptionally induced by low temperature and hypoxia, has recently been found to be upregulated in human tumors. However, its expression status in human astrocytoma is not well defned. This article focuses on the differential expression of RBM3 in human astrocytomas of different grades and normal brain tissues. METHODS: RBM3 was detected in astrocytomas and normal brain tissues by quantitative real-time PCR, immunohistochemistry, and Western blotting. Analysis of variance was performed on the data from quantitative real-time PCR. The Fisher's exact test was used to analyze the immunohistochemistry results. A P-value of less than 0.05 indicates a statistically significant difference. RESULTS: On one hand, the mRNA expression levels of three X-chromosome-related RBM genes (RBMX, RBM3, and RBM10) were detected by quantitative real-time PCR. The results showed that there were no significant differences in RBMX and RBM10 mRNA expression levels in human astrocytomas of different grades and normal brain tissues. However, RBM3 mRNA expression levels were elevated in high-grade (World Health Organization (WHO) Grade III-IV) astrocytomas versus low-grade (WHO Grade I-II) astrocytomas (5.06 ± 0.66 vs. 1.60 ± 0.58; P < 0.05) or normal controls (5.06 ± 0.66 vs. 1.03 ± 0.22; P < 0.05) as determined by quantitative real-time PCR analysis. On the other hand, immunohistochemistry showed an increased RBM3 labeling index in astrocytomas of different grades and normal brain tissues (positive staining rate: astrocytoma Grade IV, 92.9%; astrocytoma Grade III, 81.8%; astrocytoma Grade I-II, 50%; normal brain tissues, 37.5%; high-grade astrocytoma versus normal brain tissues, P < 0.05; high-grade astrocytoma versus low-grade astrocytoma, P < 0.05). The higher protein levels of RBM3 were also validated in high-grade astrocytomas and low-grade astrocytomas compared with normal brain tissues by Western blotting. CONCLUSIONS: These data suggest that the overexpression of RBM3 may serve as an important molecular mechanism underlying astrocytic carcinogenesis. Moreover, RBM3 may have proliferative and/or proto-oncogenic functions in human astrocytomas.

Asunto(s)

Astrocitoma/metabolismo , Proteínas de Unión al ARN/metabolismo , Astrocitoma/genética , Western Blotting , Humanos , Inmunohistoquímica , Técnicas In Vitro , Proteínas de Unión al ARN/genética , Reacción en Cadena en Tiempo Real de la Polimerasa

RESUMEN

OBJECTIVE: The expression cold-inducible RNA-binding protein (CIRP) is significantly enhanced in neurons under hypothermia, but its roles remain unclear. This study aims to investigate whether the cerebral protection under hypothermia is mediated by the CIRP-mediated inhibition of neuronal apoptosis. METHODS: Primary rat cortical neurons were isolated, cultured, and transduced with lentiviral CIRP-RNAi. Apoptosis of the transduced neurons was induced with 100 µmol/L H2O2, the treated cells were divided into two groups, and cultured in 37 °C or 32 °C incubator respectively. Cell viability was detected by MTT colorimetric assay. Neuronal apoptosis was detected by flow cytometry after labeling the cells with Hoechst 33342 and Annexin V-FITC/PI. The protein expressions of CIRP, activated caspase-3, and thioredoxin (TRX) were detected by Western blot. RESULTS: Under 32 °C, CIRP protein is significantly induced in cortical neurons; the expression of activated caspase-3 decreases, while the TRX expression increases. The rate of neuronal apoptosis is 4.5±0.8%. Under 37 °C, CIRP expression is evidently reduced in cortical neurons; the expression of activated caspase-3 is significantly enhanced with reduced level of TRX expression. The rate of neuronal apoptosis reaches 53.5±1.7% (P < 0.05, compared to that in 32 °C group). CONCLUSIONS: The induction of CIRP protein in rat cortical neurons under hypothermia inhibits H2O2-induced neuronal apoptosis and thereby exerts neuroprotective effect, which forms one of the cerebral protective pathways under hypothermia.

Asunto(s)

Apoptosis/fisiología , Corteza Cerebral/metabolismo , Peróxido de Hidrógeno/toxicidad , Neuronas/metabolismo , Proteínas de Unión al ARN/biosíntesis , Animales , Apoptosis/efectos de los fármacos , Células Cultivadas , Corteza Cerebral/efectos de los fármacos , Femenino , Peróxido de Hidrógeno/antagonistas & inhibidores , Hipotermia Inducida/métodos , Neuronas/efectos de los fármacos , Embarazo , Ratas , Ratas Sprague-Dawley

RESUMEN

BACKGROUND: Cerebral vasospasm (CVS) is a common serious complication after the spontaneous subarachnoid hemorrhage (SAH). Despite recent advances in medical and surgical treatments, the 30-day mortality rate of SAH remains high, and there is lack of especially effective clinical treatment to alleviate and improve CVS. The present study has investigated the therapeutic effect of insulin and vitamin C on CVS after SAH. RESULTS: Five days after SAH, there is obvious basilar artery spasm in SAH group, whose average vascular cross-sectional area (233,099 ± 16,750 µm²) is significantly smaller than that in control group (462,128 ± 74,756 µm²), which is also significantly different from those in SAH + insulin group (221,114 ± 43,457 µm²) and SAH + vitamin C group (237,820 ± 21,703 µm²). SAH + insulin + vitamin C group shows no evident vasospasm and maintains a vascular cross-sectional area of 425,530 ± 45,503 µm², which is significantly different from that in SAH group. Insulin receptor α (InRα) expression is significantly downregulated in the vascular endothelial cells of SAH, SAH + insulin, and SAH + vitamin C groups (P < 0.01) but remains unchanged in vascular endothelial cells of SAH + insulin + vitamin C group (P > 0.05). Five days after SAH, serum and cerebrospinal fluid NO levels in SAH, SAH + insulin, and SAH + vitamin C groups decrease significantly (P < 0.01) compared to that in control group, whereas the reduction is not evident in SAH + insulin + vitamin C group (P > 0.05). CONCLUSION: Combinatorial treatment with insulin and vitamin C has effectively relieved the CVS after SAH in rabbit, possibly through increasing the InRα expression and NO level, whereas treatment with insulin or vitamin C alone fails to do so.

Asunto(s)

Ácido Ascórbico/uso terapéutico , Hipoglucemiantes/uso terapéutico , Insulina/uso terapéutico , Hemorragia Subaracnoidea/complicaciones , Vasoespasmo Intracraneal/tratamiento farmacológico , Vitaminas/uso terapéutico , Animales , Modelos Animales de Enfermedad , Quimioterapia Combinada , Insulina/sangre , Conejos , Vasoespasmo Intracraneal/etiología

RESUMEN

A new ultraviolet (UV) labeling reagent, p-acetamidobenzenesulfonyl fluoride (PAABS-F), was designed and synthesized to label and determine the amino acids by capillary electrophoresis (CE) with diode-array detector (DAD). PAABS-F is very stable and easy to synthesize. It reacted with primary or secondary amino acids very quickly under facile conditions to give corresponding derivatives in high yield with excellent sensitivity and stability. No by-products were observed in amino acid derivatives when stored at room temperature under natural daylight for at least 7 days. Both amino acids standard solution and real samples reacted with this new UV labeling reagent smoothly to form high UV-absorption derivatives. The labeled 20 standard amino acids were efficiently separated by CE and the mass detection limits (S/N=3) were ranged from 59.3 fmol for L-tryptophan to 1.70 pmol for L-histidine.