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Osteophyte formation, a key indicator of osteoarthritis (OA) severity, remains poorly understood in its relation to gut microbiota and metabolites in knee osteoarthritis (KOA). We conducted 16S rDNA sequencing and untargeted metabolomics on fecal and serum samples from 20 healthy volunteers, 80 KOA patients in Guangdong, and 100 in Inner Mongolia, respectively. Through bioinformatics analysis, we identified 3 genera and 5 serum metabolites associated with KOA osteophyte formation. Blautia abundance negatively correlated with meat, cheese, and bean consumption. The 5 serum metabolites negatively correlated with dairy, beef, cheese, sugar, and salt intake, yet positively with age and oil consumption. Higher Blautia levels in the gut may contribute to KOA osteophyte formation, with serum metabolites LTB4 and PGD2 potentially serving as biomarkers. KOA patients in Inner Mongolia exhibited lower Blautia levels and reduced expression of 5 serum metabolites, possibly due to cheese consumption habits, resulting in less osteophyte formation.
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Environmental disasters like drought reduce agricultural output and plant growth. Redox management significantly affects plant stress responses. An earlier study found that PbPIP1;4 transports H2O2 and promotes H2O2 downstream cascade signaling to restore redox equilibrium. However, this regulatory mechanism requires additional investigation. In this search, the AP2 domain-containing transcription factor was isolated by screening Y1H from the wild pear (Pyrus betulaefolia) cDNA library, named PbERF3. The overexpression of PbERF3 in pear callus and Arabidopsis enhanced plant resistance to drought and re-established redox balance. The transcripts of the NCEDs gene were upregulated under drought stress. The drought stress-related abscisic acid (ABA) signaling pathway modulates PbERF3. PbERF3 silencing lowered drought tolerance. Furthermore, yeast 2-hybrid, luciferase, bimolecular fluorescence complementation, and co-immunoprecipitation assays verified that PbERF3 physically interacted with PbHsfC1a. The PbERF3-PbHsfC1a heterodimer coordinately bound to PbPIP1;4 and PbNCED4 promoter, therefore activating both the H2O2 and the ABA signaling pathway. This work revealed a novel PbERF3-PbHsfC1a-PbNCED4-PbPIP1;4 regulatory module, in which PbERF3 interacts with PbHsfC1a to trigger the expression of target genes. This module establishes an interaction between the H2O2 signaling component PbPIP1;4 and the ABA pathways component PbNCED4, enabling a response to drought.
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Flavonoids are plant pigments that play a major role in plant defense and have significant health benefits to humans. Chalcone synthase (CHS) is an important enzyme in flavonoid biosynthesis and investigation transcription factors (TFs) regulating its expression and downstream targets is critical to understanding its mechanism. Here, a novel TF, PbWRKY18, was isolated from the pear Pyrus betulaefolia. Its expression was evaluated in various tissues by RT-PCR, particularly in response to Alternaria alternata, the pathogen responsible for black spot disease, and exogenous hormone administration. The PbWRKY18 protein was primarily found in the nucleus where it regulated transcriptional activity. Yeast one-hybrid and dual-luciferase reporter assays showed a strong association between PbWRKY18 and the PbCHS3 promoter, which drives PbCHS3 expression. It was also found that PbCHS3 was critical for the development of resistance against black spot disease. In addition, PbWRKY18 was found to significantly increase the expression of PbCHS3 and salicylic acid-related genes, as well as defense enzyme activity and tolerance to black spot disease. PbWRKY18 or PbCHS3 knockdown in pear attenuates resistance to Alternaria alternata. In summary, the study identified a novel WRKY18-CHS3 axis involved in resistance against black spot disease in pear.
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Aciltransferasas , Pyrus , Humanos , Pyrus/genética , Alternaria , Regiones Promotoras GenéticasRESUMEN
Parkinson's disease is the second most prevalent age-related neurodegenerative disease and disrupts the lives of people aged >60 years. Meanwhile, single-target drugs becoming inapplicable as PD pathogenesis diversifies. Mitochondrial dysfunction and neurotoxicity have been shown to be relevant to the pathogenesis of PD. The novel synthetic compound J24335 (11-Hydroxy-1-(8-methoxy-5-(trifluoromethyl)quinolin-2-yl)undecan-1-one oxime), which has been researched similarly to J2326, has the potential to be a multi-targeted drug and alleviate these lesions. Therefore, we investigated the mechanism of action and potential neuroprotective function of J24335 against 6-OHDA-induced neurotoxicity in mice, and in PC12 cell models. The key target of action of J24335 was also screened. MTT assay, LDH assay, flow cytometry, RT-PCR, LC-MS, OCR and ECAR detection, and Western Blot analysis were performed to characterize the neuroprotective effects of J24335 on PC12 cells and its potential mechanism. Behavioral tests and immunohistochemistry were used to evaluate behavioral changes and brain lesions in mice. Moreover, bioinformatics was employed to assess the drug-likeness of J24335 and screen its potential targets. J24335 attenuated the degradation of mitochondrial membrane potential and enhanced glucose metabolism and mitochondrial biosynthesis to ameliorate 6-OHDA-induced mitochondrial dysfunction. Animal behavioral tests demonstrated that J24335 markedly improved motor function and loss of TH-positive neurons and dopaminergic nerve fibers, and contributed to an increase in the levels of dopamine and its metabolites in brain tissue. The activation of both the CREB/PGC-1α/NRF-1/TFAM and PKA/Akt/GSK-3ß pathways was a major contributor to the neuroprotective effects of J24335. Furthermore, bioinformatics predictions revealed that J24335 is a low toxicity and highly BBB permeable compound targeting 8 key genes (SRC, EGFR, ERBB2, SYK, MAPK14, LYN, NTRK1 and PTPN1). Molecular docking suggested a strong and stable binding between J24335 and the 8 core targets. Taken together, our results indicated that J24335, as a multi-targeted neuroprotective agent with promising therapeutic potential for PD, could protect against 6-OHDA-induced neurotoxicity via two potential pathways in mice and PC12 cells.
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Enfermedades Mitocondriales , Enfermedades Neurodegenerativas , Fármacos Neuroprotectores , Humanos , Ratas , Ratones , Animales , Oxidopamina/farmacología , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Células PC12 , Glucógeno Sintasa Quinasa 3 beta , Simulación del Acoplamiento Molecular , Dopamina , Neuronas DopaminérgicasRESUMEN
The aggregation of misfolded proteins, such as α-synuclein in Parkinson's disease (PD), occurs intracellularly or extracellularly in the majority of neurodegenerative diseases. The immunoproteasome has more potent chymotrypsin-like activity than normal proteasome. Thus, degradation of α-synuclein aggregation via immunoproteasome is an attractive approach for PD drug development. Herein, we aimed to determine if novel compound, 11-Hydroxy-1-(8-methoxy-5-(trifluoromethyl)quinolin-2-yl)undecan-1-one oxime (named as J24335), is a promising candidate for disease-modifying therapy to prevent the pathological progression of neurodegenerative diseases, such as PD. The effects of J24335 on inducible PC12/A53T-α-syn cell viability and cytotoxicity were evaluated by MTT assay and LDH assay, respectively. Evaluation of various proteasome activities was done by measuring the luminescence of enzymatic activity after the addition of different amounts of aminoluciferin. Immunoblotting and real-time PCR were employed to detect the expression of various proteins and genes, respectively. We also used a transgenic mouse model for behavioral testing and immunochemical analysis, to assess the neuroprotective effects of J24335. J24335 inhibited wild-type and mutant α-synuclein aggregation without affecting the growth or death of neuronal cells. The inhibition of α-synuclein aggregation by J24335 was caused by activation of immunoproteasome, as mediated by upregulation of LMP7, and increased cellular chymotrypsin-like activity in 20S proteasome. J24335-enhanced immunoproteasome activity was mediated by PKA/Akt/mTOR pathway activation. Moreover, animal studies revealed that J24335 treatment markedly mitigated both the loss of tyrosine hydroxylase-positive (TH-) neurons and impaired motor skill development. This is the first report to use J24335 as an immunoproteasome enhancing agent to antagonize pathological α-synuclein-mediated neurodegeneration.
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Enfermedades Neurodegenerativas , Enfermedad de Parkinson , Ratones , Animales , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Quimotripsina/uso terapéutico , Enfermedad de Parkinson/genética , Ratones Transgénicos , Enfermedades Neurodegenerativas/tratamiento farmacológico , Modelos Animales de EnfermedadRESUMEN
ABSTRACT: Mosaicism can be observed in karyotype analyses of amniotic fluid cells. Differentiating between true mosaicism and pseudomosaicism and determining mosaic proportions can help avoid misjudgments by doctors and effectively reduce mental and physical harm to pregnant women. However, the detection of mosaicism and mosaic proportions via karyotype analysis and fluorescence in situ hybridization (FISH) is extremely complex. We have developed a novel approach, "segmental duplication quantitative fluorescent PCR" (SD-QF-PCR), to detect mosaicism and mosaic proportions.In this study, twenty control samples and fourteen mosaic samples were tested by first-line karyotype analysis; by second-line karyotype analysis, SD-QF-PCR and FISH were used to reassess fetal sex chromosome mosaicism and mosaic proportions.Detection of the 20 control samples by second-line karyotype analysis via FISH and SD-QF-PCR showed normal and consistent results. Among the 14 mosaic samples, the numbers of samples showing true mosaicism and pseudomosaicism detected by the three methods were 6 and 8, respectively.Our study demonstrates that SD-QF-PCR can be used as a complementary method to traditional cytogenetic analysis of amniotic fluid mosaics and has clinical application value.
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Cariotipificación/métodos , Mosaicismo , Reacción en Cadena de la Polimerasa/métodos , Diagnóstico Prenatal/métodos , Aberraciones Cromosómicas Sexuales , Amniocentesis , Líquido Amniótico/citología , Células Cultivadas , Estudios de Factibilidad , Femenino , Humanos , Hibridación Fluorescente in Situ , Embarazo , Cultivo Primario de CélulasRESUMEN
A ceramic-based coating with a hierarchical surface structure was synthesized via solid-state reaction enabled by a double cathode glow discharge technique. This innovative coating comprises two distinct layers, specifically an outer layer with a well-aligned micro-pillar array and a dense inner layer. Both are composed of a face-centered cubic Cu(Co,Ni,Fe) solid solution phase together with a spinel-type Fe(Al,Cr)2O4 oxide. This coating exhibits superhydrophobicity and, yet, a very strong adhesion to water, i.e., the so-called "rose petal effect". This coating also exhibits highly efficient antibacterial ability against both Staphylococcus aureus and Escherichia coli bacteria under both dark and visible light conditions. The excellent antibacterial property originates from the synergistic effects through the release of Cu ions coupled with photothermal activity upon light activation.
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Somatostatin (SST) has a protective role in intestinal injury, inflammatory response, and intestinal mucosal barrier in rats with acute pancreatitis. However, its function in sepsis-induced intestinal barrier dysfunction remains largely unknown. A mouse sepsis model was constructed, and SST was injected into the tail vein. Then, hematoxylin and eosin staining (HE) was used to detect the intestinal barrier dysfunction. Enzyme-linked immunosorbent assay was used to detect the level of tumor necrosis factor α- (TNF-) α, interleukin- (IL-) 6, and interleukin- (IL-) 10 in the ileum. Expressions of tight junction proteins, zonula occludens- (ZO-) 1 and Claudin-1, and NF-κB p65 in the ileum were detected using western blot and immunohistochemistry as needed. Furthermore, JSH-23 as an inhibitor of the NF-κB pathway was injected into sepsis mice with SST or not. Mice with sepsis showed an obvious intestinal barrier dysfunction with decreasing specific somatostatin receptor subtype (SSTRs), and increasing TNF-α, IL-6, and IL-10 in the ileum. SST could relieve the injury, the decrease of SSTRs, and the increase of TNF-α and IL-6 induced by sepsis and also further enhanced the expression of IL-10. Further analysis showed that ZO-1 and Claudin-1 were reduced in the ileum by sepsis but enhanced by SST. NF-κB p65 was promoted in the ileum by sepsis but inhibited by SST. Further experiments confirmed that NF-κB inhibitor JSH-23 could repair the intestinal barrier dysfunction and enhance the protective effect of SST on the intestinal barrier. SST, with a protective effect on intestinal barrier dysfunction through suppression of NF-κB, could be a potential therapeutic drug for sepsis-induced intestinal barrier dysfunction.
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To investigate the associations between prenatal exposure to single metal and multiple metals and the risk of low birth weight (LBW), we conducted a nested case-control study of 246 LBW and 406 NBW mother-infant pairs based on a prospective birth cohort study. 22 serum metals were detected by inductively coupled plasma quadruple mass spectrometry (ICP-MS). Categorical analyses showed serum Co and Ti were associated with LBW (Co: 3rd vs 4th. quartile: ORâ¯=â¯1.83, 95%CI: 1.14-2.92, Ptrendâ¯=â¯0.043; Ti: 2nd vs. 4th quartile: ORâ¯=â¯0.51, 95% CI: 0.32-0.81, P trendâ¯=â¯0.051), especially gestational age >13 weeks (Co: 3rd vs. 4th quartile: OR = 1.94, 95% CI: 1.13 - 3.32, Ptrend = 0.043; Ti: 2nd vs. 4th quartile: OR = 0.50, 95% CI: 0.30 - 0.84, P trend= 0.073). Cubic spline analyses showed serum Co and serum Ti had non-linearity associations with LBW (Co: P for overallâ¯=â¯0.048, P-nonlinearityâ¯=â¯0.014; Ti: P for overallâ¯=â¯0.015, P- nonlinearityâ¯=â¯0.008). In multi-metal compound exposure model, 15 metals selected by elastic net model were significantly associated with the increased risk of LBW and OR (95%CI) was 5.14 (2.81-9.40). Our study suggested that lower level serum Co was positively associated with LBW and lower level serum Ti was negatively associated with LBW, especially gestational age >13 weeks, and both of them had non-linearity dose-relationships with LBW. And multi-metal compound model was significantly associated with LBW compared with single metal model.
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Contaminantes Ambientales/sangre , Recién Nacido de Bajo Peso/metabolismo , Exposición Materna/estadística & datos numéricos , Metales/sangre , Adulto , Peso al Nacer , Estudios de Casos y Controles , China , Estudios de Cohortes , Femenino , Edad Gestacional , Humanos , Lactante , Recién Nacido , Madres , Embarazo , Estudios Prospectivos , Adulto JovenRESUMEN
The study aimed to investigate the associations between maternal lifestyles and antenatal stress and anxiety. 1491 pregnant women were drawn from the Guangxi birth cohort study (GBCS). A base line questionnaire was used to collect demographic information and maternal lifestyles. The Pregnancy Stress Rating Scale (PSRS) and Self-Rating Anxiety Scale (SAS) were used to assess prenatal stress and anxiety, respectively. Regression analyses identified the relationship between maternal lifestyles and prenatal stress and anxiety: (1) Hours of phone use per day was positively correlated to prenatal stress and anxiety and increased with stress and anxiety levels (all P trend < 0.05). In addition, not having baby at home was positively correlated to prenatal stress. (2) Self-reported sleep quality was negative with prenatal stress and anxiety, and decreased with stress and anxiety levels (all P trend < 0.01). Moreover, not frequent cooking was negatively correlated to prenatal stress and having pets was negatively correlated to prenatal anxiety (P < 0.05). However, having pets was not correlated to prenatal stress (P > 0.05). Our results showed that adverse lifestyles increase the risk of antenatal stress and anxiety, a regular routine and a variety of enjoyable activities decreases the risk of prenatal stress and anxiety.
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Ansiedad , Estilo de Vida , Mujeres Embarazadas/psicología , Estrés Psicológico , Adulto , China , Estudios de Cohortes , Estudios Transversales , Femenino , Humanos , Embarazo , AutoinformeRESUMEN
Dot-blot hybridization and high-resolution melting curve methods are used to detect G6PD gene mutations; however, the performance and throughput limitations of these methods hinder their use for screening large populations. For simple screening, we developed a novel approach called "Amplification Refractory Mutation System combined with Melting Curve Analysis (ARMS-MC)," which enables rapid and batch-based detection of the 6 most common G6PD mutations.In this method, we established 4 PCR reaction systems that can be used to detect the 6 most common G6PD mutations (c.95A>G, c.392G>T, c.871G>A, c.1024C>T, c.1376G>T, and c.1388G>A) in the Chinese population.The ARMS-MC method was evaluated with 174 cases of clinical G6PD-deficient samples, and the results were verified by direct sequencing at G6PD gene exons. The results showed that 170 samples had ≥1 of the 6 mutations, which accounted for 97.70% of all mutations. These results were consistent with the results of direct sequencing with 100% accuracy and specificity. Sequencing validation revealed other mutations in the 4 samples in which no mutation was detected by the ARMS-MC method.ARMS-MC provides a rapid, simple, inexpensive, and accurate screening method for detecting the most common G6PD mutations in Chinese people.
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Deficiencia de Glucosafosfato Deshidrogenasa/genética , Glucosafosfato Deshidrogenasa/genética , Reacción en Cadena de la Polimerasa/métodos , Pueblo Asiatico , Femenino , Humanos , Masculino , Polimorfismo de Nucleótido SimpleRESUMEN
Different types of deletional α-thalassemia (α-thal) have been reported by researchers in China. This study describes one family carrying -α21.9 (NG_000006.1: g.14373_36299delinsGGGAAGGGTGGGTGGGAATAACAGCTTTT), -α2.4 (NG_000006.1: g.36860_39251del) and - -THAI (Thailand) (NG_000006.1: g.10664_44164del) alleles in Guangxi Zhuang Autonomous Region, People's Republic of China (PRC), and reports the frequencies of these types in the population of this region. The proband was a 4-year-old girl, who screened positive for thalassemia, although the thalassemia genotype results were normal when screened using the routine kits. Samples of the proband's parents were also collected to perform further analyses. Two real-time gap-polymerase chain reaction (gap-PCR) systems were designed for separate detection of - -THAI and screening for -α21.9 and -α2.4. The genotype of the proband was -α21.9/-α2.4, and the two variants were inherited from her parents. In the frequency study, five - -THAI, four -α21.9 and 11 -α2.4 positive individuals were detected in the 3410 random samples. Thus, allele frequencies of -α21.9, - -THAI and -α2.4 in the population of southern Guangxi were determined as 0.059, 0.073 and 0.161%, respectively. This is the first report of an individual carrying the -α21.9/-α2.4 genotype, and the first report of the detection of -α21.9, -α2.4 and - -THAI in a single family. The total frequency for these alleles was 0.293% in southern Guangxi, suggesting that the thalassemia clinical center in this region should utilize a screening kit that allows detection of these types of deletions for a more comprehensive evaluation of thalassemia risk.
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Frecuencia de los Genes , Eliminación de Secuencia , Talasemia alfa/genética , Preescolar , China , Femenino , Genotipo , Humanos , LinajeRESUMEN
OBJECTIVE: To establish stoma and stoma-free murine models of heterotopic small intestine transplantation in order to choose a more effective and reliable model. METHODS: A total of 140 male 8-10 weeks age C57BL/6(B6) mice weighted 25-30 g were enrolled in the experiment. Syngeneic heterotopic small intestine transplantation was performed between C57BL/6 mice, and recipient mice were divided into either stoma or stoma-free group. Heterotopic small intestine transplantation was performed in 70 mice, with 35 mice in each group. After closing the proximal end of the graft by ligation, the distal end of graft was exteriorized as a stoma then secured to the skin of the abdominal wall in stoma group. In stoma-free group, the distal end of graft was anastomosed end-to-side to the recipient ileum. Successful rate of operation, two-week survival rate, operation time, associated complications, postoperative care time and body weight change were recorded and compared between two groups. RESULTS: The successful rate of stoma group was 65.7%, while it was 80.0% of stoma-free group (χ(2)=1.806, P=0.179). The operation time of donor in stoma group was (48.1±6.6) minutes, while it was (47.2±5.9) minutes in stoma-free group (t=0.598, P=0.552). The operation time of recipient in stoma group was (77.9±9.1) minutes, while it was (76.4±8.3) minutes in stoma-free group (t=0.683, P=0.497). The cold ischemic time of graft in stoma group was (34.7±4.0) minutes, while it was (33.9±4.6) minutes in stoma-free group(t=0.667, P=0.507). The two-week survival rate of stoma group was 45.7%, and it was 77.1% of stoma-free group(χ(2)=7.295, P=0.007). The stoma group had more complications[54.3%(19/35) vs. 22.9%(8/35), χ(2)=7.295, P=0.007], which needed more postoperative care time(191 min vs. 35 min). The weight loss in stoma group in the third day after operation was more significant [(81.52±5.20)% vs. (85.46±4.65)%, t=2.856, P=0.006]. By 2 weeks after operation, the weight of mice in both groups retruned to 95% of the postoperative wight. CONCLUSION: The murine heteropotic small intestine transplantation model with stoma-free appears to be more reasonable and reliable.
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Intestino Delgado/trasplante , Estomas Quirúrgicos , Trasplante Heterotópico/métodos , Animales , Procedimientos Quirúrgicos del Sistema Digestivo , Íleon/cirugía , Masculino , Ratones , Ratones Endogámicos C57BL , Trasplante IsogénicoRESUMEN
A novel 3D printing method for voxel-by-voxel metal printing is presented. Hollow atomic force microscopy (AFM) cantilevers are used to locally supply metal ions in an electrochemical cell, enabling a localized electroplating reaction. By exploiting the deflection feedback of these probes, electrochemical 3D metal printing is, for the first time, demonstrated in a layer-by-layer fashion, enabling the fabrication of arbitrary-shaped geometries.
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Metales/química , Nanotecnología/métodos , Sulfato de Cobre/química , Galvanoplastia , Microscopía de Fuerza Atómica , Microscopía Electrónica de Rastreo , Nanotecnología/instrumentación , Impresión TridimensionalRESUMEN
Severe acute pancreatitis (SAP) is normally related to multiorgan dysfunction and local complications. Studies have found that local pancreatic renin-angiotensin system (RAS) was significantly upregulated in drug-induced SAP. The present study aimed to investigate the effects of angiotensin II receptors inhibitor valsartan on dual role of RAS in SAP in a rat model and to elucidate the underlying mechanisms. 3.8% sodium taurocholate (1 ml/kg) was injected to the pancreatic capsule in order for pancreatitis induction. Rats in the sham group were injected with normal saline in identical locations. We also investigated the regulation of experimentally induced SAP on local RAS expression in the pancreas through determination of the activities of serum amylase, lipase and myeloperoxidase, histological and biochemical analysis, radioimmunoassay, fluorescence quantitative PCR and Western blot analysis. The results indicated that valsartan could effectively suppress the local RAS to protect against experimental acute pancreatitis through inhibition of microcirculation disturbances and inflammation. The results suggest that pancreatic RAS plays a critical role in the regulation of pancreatic functions and demonstrates application potential as AT1 receptor antagonists. Moreover, other RAS inhibitors could be a new therapeutic target in acute pancreatitis.
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Glypican3 (GPC3) is a membrane heparan sulfate proteoglycan involved in cell proliferation, differentiation, adhesion, migration and the development of the majority of mesodermal tissues and organs. GPC3 has been found to be important for the occurrence and development of hepatocellular carcinoma (HCC). Therefore, it may be suitable for use as a novel molecular marker for the diagnosis of primary liver cancer. In the present study, the role of GPC3 in the occurrence and development of HCC was determined. GPC3 recombinant vector was transfected into two HCC cell lines, Huh7 and SKHEP1, to upregulate the expression of GPC3 and examine changes in the biological behavior of the cells. Results indicate that overexpression of GPC3 in Huh7 and SKHEP1 cells effectively inhibited cell proliferation and cell invasion through induction of apoptosis. However, cotreatment of the cells with insulinlike growth factor 2 (IGF2) and fibroblast growth factor 2 (FGF2) was found by Annexin VPI flow cytometric analysis to significantly inhibit the apoptotic cell death induced by GPC3 overexpression. These observations indicate that GPC3 may act as a negative regulator of IGF2 and FGF2 pathways. Taken together, these results demonstrate that overexpression of GPC3 inhibits the occurrence and development of HCC.
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Apoptosis , Glipicanos/metabolismo , Apoptosis/efectos de los fármacos , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Factor 2 de Crecimiento de Fibroblastos/farmacología , Vectores Genéticos/metabolismo , Glipicanos/genética , Humanos , Factor II del Crecimiento Similar a la Insulina/farmacología , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , ARN Mensajero/metabolismo , TransfecciónRESUMEN
Glypican-3 (GPC3), a membrane-associated heparan sulfate proteoglycan, is frequently upregulated in hepatocellular carcinoma (HCC). Yes-associated protein (YAP) is also found over-expressed in HCC and has been identified as a key effector molecule in Hippo pathway, which could control the organ size in animals through the regulation of cell proliferation and apoptosis and plays an important role in the development of malignant tumors. Studies have reported that GPC3 and YAP might collaborate to regulate the development of HCC. To elucidate the role of GPC3 in the development of HCC and its relationship with YAP, siRNA technique was employed to knock down GPC3 in Huh7 HCC cells. Moreover, recombinant human YAP-1 was used to examine the effects of GPC3 on Huh7 cells. The results of flow cytometric analysis and Annexin-V-FLUOS apoptosis assay showed that knockdown of GPC3-induced apoptosis in Huh7 cells, resulting in inhibition of cell proliferation as examined by EdU incorporation assay, migration, and invasion. GPC3 knockdown also suppressed the expression of YAP in mRNA and protein levels, as examined by fluorescence quantitative PCR and Western blot analysis. Moreover, addition of recombinant human YAP-1 effectively rescued the cells from apoptosis triggered by GPC3 knockdown. Taken together, our findings suggest that GPC3 regulates HCC cell proliferation with the involvement of Hippo pathway.
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Proteínas Adaptadoras Transductoras de Señales/metabolismo , Carcinoma Hepatocelular/metabolismo , Glipicanos/genética , Fosfoproteínas/metabolismo , Anexina A5/análisis , Apoptosis/genética , Carcinoma Hepatocelular/genética , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Regulación hacia Abajo , Fluoresceínas , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Invasividad Neoplásica/genética , Interferencia de ARN , ARN Mensajero/biosíntesis , ARN Interferente Pequeño , Proteínas Recombinantes/metabolismo , Factores de Transcripción , Proteínas Señalizadoras YAPRESUMEN
Cellular automata models have historically been a major approach to studying the information-processing properties of self-replication. Here we explore the feasibility of adopting genetic programming so that, when it is given a fairly arbitrary initial cellular automata configuration, it will automatically generate a set of rules that make the given configuration replicate. We found that this approach works surprisingly effectively for structures as large as 50 components or more. The replication mechanisms discovered by genetic programming work quite differently than those of many past manually designed replicators: There is no identifiable instruction sequence or construction arm, the replicating structures generally translate and rotate as they reproduce, and they divide via a fissionlike process that involves highly parallel operations. This makes replication very fast, and one cannot identify which descendant is the parent and which is the child. The ability to automatically generate self-replicating structures in this fashion allowed us to examine the resulting replicators as their properties were systematically varied. Further, it proved possible to produce replicators that simultaneously deposited secondary structures while replicating, as in some past manually designed models. We conclude that genetic programming is a powerful tool for studying self-replication that might also be profitably used in contexts other than cellular spaces.