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Abstract This study evaluated color stability (CS), anti-adherence effect (AAE), and cell viability of microorganisms on acrylic resin (AR) surface, treated associated or not with sodium percarbonate (SP). AR specimens were prepared, and color analysis was performed before and after the treatments and the CS was calculated. For analysis of AAE, the samples were sterilized by radiation in a microwave oven. Then samples were randomly distributed: phosphate-buffered saline (PBS - control), 0.5% sodium hypochlorite (SH), phytosphingosine (PHS), and phytosphingosine + SP (PHS+Na2CO3). The specimens remained in contact with solutions for 30 minutes and were later contaminated by Candida albicans. Aliquots were seeded in Petri dishes with Sabouraud Dextrose agar and incubated at 37°C for 24 hours. After the incubation, the number of colonies was counted. The cell viability of adhered microorganisms on the AR was evaluated and 20 fields were observed under an epifluorescence microscope, and the percentage of adhered viable cells was calculated. Data were compared (One-way ANOVA, Tukey, p<.05). As for CS, PHS+ Na2CO3 (0.4±0.1) resulted in less change than PBS (0.9±0.2), similar to the other groups (SH [1.0±0.3)]; PHS [0.9±0.2)]). There was no difference for all tested solutions regarding the ability to avoid microorganism adherence (p>0.05), but PHS (11.2±4.1) resulted in a smaller area of adhered viable cells, statistically different from SH (18.2±7.6) and PBS (26.4±10.8). It was concluded that PHS resulted in lower adhered viable cells and when associated with Na2CO3, also shows a lower effect on the CS of AR.
Resumo Este estudo avaliou estabilidade de cor (EC), efeito antiaderente (EAA) e viabilidade celular de microrganismos em superfície de resina acrílica (RA), tratada com solução de fitoesfingosina, associada ou não ao percarbonato de sódio (PS). Espécimes RA foram preparados e análise de cor foi realizada antes e após os tratamentos e EC foi calculada. Para análise de EAA, as amostras foram esterilizadas por radiação em forno de micro-ondas. Então foram distribuídas aleatoriamente: solução salina tamponada com fosfato (PBS - controle), hipoclorito de sódio 0,5% (SH), fitoesfingosina (PHS) e fitoesfingosina + SP (PHS+Na2CO3). Os espécimes permaneceram em contato com as soluções por 30 minutos e posteriormente foram contaminados por Candida albicans. Alíquotas foram semeadas em placas de Petri com ágar Sabouraud Dextrose e incubadas a 37°C por 24 horas. Após a incubação, o número de colônias foi contado. A viabilidade celular dos microorganismos aderidos na RA foi avaliada e 20 campos foram observados em microscópio de epifluorescência, e a porcentagem de células viáveis aderidas foi calculada. Os dados foram comparados (One-way ANOVA, Tukey, p<0,05). Quanto a EC, o PHS+ Na2CO3 [0,4 (0,1)] resultou em menor alteração que o PBS [0,9 (0,2)], semelhante aos demais grupos (SH [1,0 (0,3)]; PHS [0,9 (0,2)]). Não houve diferença para todas as soluções testadas quanto à capacidade de evitar a aderência de microorganismos (p>0,05), mas o PHS [11,2 (4,1)] resultou em uma área menor de células viáveis aderidas, estatisticamente diferente do SH [18,2 (7,6)] e PBS [26,4 (10,8)]. Concluiu-se que o PHS resultou em menor número de células viáveis aderidas e, quando associado ao Na2CO3, também apresenta menor efeito sobre o EC da RA.
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Snuhi (Euphorbia neriifolia Linn.) is a conventional herb used broadly in several disease conditions as indicated in classical texts of Ayurveda. As per literature review ascertained, no literature was accessible regarding anticancer activity of Snuhi Kshara. Thus, present work was designed to evaluate the anticancer activity of Snuhi Kshara in HCT-15 (Human Colon Cancer cell line). Anticancer activity was evaluated using MTT assay by % cell viability and IC50. Anticancer activity was compared with standard drug capecitabine. A positive correlation between Concentration and % cell viability was noticed. Lowest cell viability was noted at 5000 µg concentration. Results obtained through the study indicates towards anticancer activity of Snuhi Kshara.
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@#As a new research field,gel has been paid more attention and widely used for studies on tissue engineering,drug delivery and biosensor. Hydrogel is the carrier of cells,while the cell survival and death are keys to the construction of tissues and organs. However,the cell viability and biological behavior are limited by the exchange of hydrogel and nutrients in medium. This review summarizes the types of hydrogel,exchange mode of hydrogel and nutrients in medium and the relevant influencing factors,which will provide a reference for the development and research of tissue bioengineering.
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Objective:To investigate the effects of long non-coding RNA (lncRNA) RP11-1212A22.4 on the cell viability and invasive ability of esophageal cancer cell lines by targeting miRNA-483-5p (miR-483-5p).Methods:The expression of RP11-1212A22.4 in esophageal cancer tissues was analyzed by using GEPIA online database. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of RP11-1212A22.4 in human esophageal cancer cell lines EC9706, KYSE30, TE-13, Eca109 and normal esophageal epithelial cell line HET-1A. The lowest expression level of EC9706 cell line in RP11-1212A22.4 was divided into RP11-1212A22.4 group (transfected with pcDNA-RP11-1212A22.4 plasmid) and the control group (transfected with pcDNA-NC plasmid). The cell viability of EC9706 cell was analyzed by using methyl thiazolyl tetrazolium (MTT) method, and the invasion ability of EC9706 cell was detected by using Transwell assay. The targeting relationship between RP11-1212A22.4 and miR-483-5p was verified by using StarBase database prediction and dual luciferase reporter assay. The relative expression level of miR-483-5p of EC9706 cell in two groups was detected by using qRT-PCR. Western blot was used to detect the expressions of cyclin-dependent kinase 6 (CDK6), matrix metalloproteinase 2 (MMP-2), cyclin-dependent kinase 4 (CDK4), and matrix metalloproteinase 9 (MMP-9) proteins in two groups.Results:In GEPIA online database, compared with adjacent tissues, the relative expression level of RP11-1212A22.4 in esophageal cancer tissues was decreased, and the difference was statistically significant ( P < 0.001). The relative expression levels of RP11-1212A22.4 in esophageal cancer cell lines EC9706, KYSE30, TE-13, Eca109 and normal esophageal mucosal epithelial cell line HET-1A were 0.11±0.08, 0.32±0.09, 0.72±0.09, 0.59±0.13 and 0.97±0.12, and the difference was statistically significant ( F = 40.42, P < 0.001). The relative expression levels of RP11-1212A22.4 in EC9706 cells of RP11-1212A22.4 group and the control group were 11.9±2.4 and 1.0±0.3, respectively, and the difference was statistically significant ( t = 8.89, P < 0.001). Compared with the control group, the cell viability of EC9706 cell in RP11-1212A22.4 group was decreased (all P < 0.05). The number of invasive cells in RP11-1212A22.4 group was lower than that in the control group (48±12 vs. 106±22, t = 4.63, P < 0.001). StarBase database prediction and dual luciferase reporter assay both showed that RP11-1212A22.4 targeted miR-483-5p. The relative expression level of miR-483-5p in RP11-1212A22.4 group was lower than that in the control group (0.24±0.11 vs. 1.02±0.23, t = 5.98, P = 0.001). Compared with the control group, the expressions of CDK6, MMP-2, CDK4 and MMP-9 proteins in the RP11-1212A22.4 group were decreased. Conclusions:RP11-1212A22.4 is lowly expressed in esophageal cancer tissues and cell lines, and it inhibits the cell viability and invasive ability of esophageal cancer cells by targeting miR-483-5p.
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Objective:To investigate the effects of orlistat on the viability of human gall-bladder cancer (GBC) cells.Methods:The experimental study was conducted. The human GBC NOZ cells with high expression of FSAN was screened out through in vitro cultivating human GBC-SD, SGC-996 and NOZ cells. The cell proliferation assay, clone formation assay and protein detection experiment were used to analysis of the effects of orlistat on the viability of human GBC cells. Cell grouping: NOZ cells cultured with medium were set as the control group, cultured with medium + 10 μmol/L orlistat were set as the low-dose orlistat group, cultured with medium + 100 μmol/L orlistat were set as the high-dose orlistat group, respectively. Observation indicators: (1) expression of FASN protein in human GBC cells; (2) effects of orlistat on the proliferation of human GBC NOZ cells; (3) effects of orlistat on apoptosis of human GBC NOZ cells. Measurement data with normal distribution were represented as Mean± SD, the ANOVA test was used for comparison between groups and the least significant difference method was used for pairwise comparison. Results:(1) Expression of FASN protein in human GBC cells. Results of western blot showed that the relative expression of FASN protein in human GBC NOZ, GBC-SD and SGC-996 cells was 0.57±0.06, 0.12±0.04 and 0.10±0.02, respectively, showing a significant difference among them ( F=115.67, P<0.05). There were significant differences between the NOZ cells and the GBC-SD or the SGC-996 cells ( P<0.05), and there was no significant difference between the GBC-SD cells and the SGC-996 cells ( P>0.05). (2) Effects of orlistat on the proliferation of human GBC NOZ cells. ① Results of cell proliferation assay showed that the absorbance value of NOZ cells was 2.34±0.12, 1.57±0.08 and 1.07±0.13 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a significant difference among them ( F=205.88, P<0.05). ② Results of clone formation assay showed that the number of NOZ cells clones was 257±23, 153±11 and 83±11 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a significant difference among them ( F=92.64, P<0.05). ③Results of western blot showed that the relative expression of Cyclin-D1 protein of NOZ cells was 2.31±0.10, 1.52±0.05 and 1.23±0.11 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a significant difference among them ( F=120.73, P<0.05). The relative expression of CDK-4 protein of NOZ cells was 1.58±0.04, 1.21±0.02 and 1.19±0.04 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a signifi-cant difference among them ( F=110.45, P<0.05). (3) Effects of orlistat on apoptosis of human GBC NOZ cells. Results of western blot showed that the relative expression of Bcl-2 protein of NOZ cells was 1.07±0.03, 0.36±0.03 and 0.15±0.02 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a significant difference among them ( F=1 242.93, P<0.05). The relative expression of Bax protein of NOZ cells was 0.51±0.03, 0.38±0.05 and 1.38±0.04 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a signifi-cant difference among them ( F=583.51, P<0.05). Conclusion:Orlistat can inhibit the growth of human GBC NOZ cells and promote their apoptosis.
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Aim To investigate the role of FKBP38 in inhibiting apoptosis in a rotenone-induced Parkinson's disease(PD)cell model. Methods In vivo experiments:MPTP-induced PD in vivo models were constructed,and the expressions of α-synuclein,TH and FKBP38 in brains of PD mice were detected. In vitro experiments:Dopaminergic neuron MN9D cells were stimulated with rotenone to construct an in vitro model of PD; Western blot was used to detect the expression levels of α-synuclein,TH,Tom20 and FKBP38 in PD in vitro model; FKBP38 lentivirus was transferred into MN9D cells to construct stable overexpression and FKBP38 knockdown cell lines; CCK-8 assay was used to detect the cell viability of FKBP38 overexpression and knockdown cells stimulated by rotenone; Western blot was used to detect anti-apoptotic protein Bcl-2 and apoptosis protein in PD cell model expression levels of Bax. Results The expression level of FKBP38 was significantly down-regulated in both in vitro and in vivo models of PD(P<0.01). Knockdown of FKBP38 aggravated the decline of dopaminergic neuron cell viability caused by rotenone(P<0.05),while overexpression of FKBP38 significantly ameliorated the decline of dopaminergic neuron cell viability caused by rotenone(P<0.05). Western blot results showed that overexpression of FKBP38 could significantly up-regulate the expression level of anti-apoptotic protein Bcl-2 and increase the ratio of Bcl-2/Bax in PD dopaminergic neurons(P<0.05). Conclusion In the PD cell model regulation of FKBP38 can improve the apoptosis of dopaminergic neurons.
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This paper aimed to study the chemical constituents from the root bark of Schisandra sphenanthera. Silica, Sephadex LH-20 and RP-HPLC were used to separate and purify the 80% ethanol extract of S. sphenanthera. Eleven compounds were identified by ~1H-NMR, ~(13)C-NMR, ESI-MS, etc., which were 2-[2-hydroxy-5-(3-hydroxypropyl)-3-methoxyphenyl]-propane-1,3-diol(1), threo-7-methoxyguaiacylglycerol(2),4-O-(2-hydroxy-1-hydroxymethylethyl)-dihydroconiferylalcohol(3), morusin(4), sanggenol A(5), sanggenon I(6), sanggenon N(7), leachianone G(8),(+)-catechin(9), epicatechin(10), and 7,4'-dimethoxyisoflavone(11). Among them, compound 1 was a new compound, and compounds 2-9 were isolated from S. sphenanthera for the first time. Compounds 2-11 were subjected to cell viability assay, and the results revealed that compounds 4 and 5 had potential cytotoxicity, and compound 4 also had potential antiviral activity.
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Schisandra , Plant Bark , Antiviral Agents , Biological Assay , Catechin , PhenolsABSTRACT
G‐quadruplexes (G4) are structures formed at the ends of telomeres rich in guanines and stabilized by molecules that bind to specific sites. TMPyP4 and thymoquinone (TQ) are small molecules that bind to G4 and have drawn attention because of their role as telomerase inhibitors. The aim of this study was to evaluate the effects of telomerase inhibitors on cellular proliferation, senescence, and death. Two cell lines, LC‐HK2 (non-small cell lung cancer - NSCLC) and RPE‐1 (hTERT-immortalized), were treated with TMPyP4 (5 μM) and TQ (10 μM). Both inhibitors decreased telomerase activity. TMPyP4 increased the percentage of cells with membrane damage associated with cell death and decreased the frequency of cells in the S‐phase. TMPyP4 reduced cell adhesion ability and modified the pattern of focal adhesion. TQ acted in a concentration-dependent manner, increasing the frequency of senescent cells and inducing cell cycle arrest in G1 phase. Thus, the present results showed that TMPyP4 and TQ, although acting as telomerase inhibitors, had a broader effect on other signaling pathways and processes in cells, differing from each other. However, they act both on malignant and immortalized cells, and further studies are needed before their anti-cancer potential can be considered.
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In the current scenario, microbial infections are major concern and they multiply so rapidly throughout the world; also, they cause serious bio illness and can even results in death among humans. Complexes framed by crystalline metal ions have been used as medications, because of a wide range of organic activities against harmful microorganisms. In the present study, greenish blue potassium tetrachlorocuprate (II) dihydrate K2CuCl4.2H2O crystalline precipitates were prepared by solvent evaporation method at ambient temperature. This complex was found in nature as a rare mineral Mitscherlichite. For finding the crystalline parameters of the sample, a Single-crystal XRD analysis was used and the data confirms. K2CuCl4.2H2O crystallizes into a tetragonal structure. The formation of bonds and the presence of functional groups in the complex were determined from Surface morphology studies, the very clear morphology of the complex has smoothing surface, defect-free, small microstructures and stacklike shapes of a well-defined crystalline pattern. Also, addition, optical analysis supports that its possibility in antimicrobial applications. The sample shows comparatively good results in positive control of antibacterial and antifungal activities, namely Gentamicin, Chloramphenicol and Clotrimazole. IC50 and optical density (OD) values were used to determine the cytotoxicity and cell viability, respectively. In vitro cytotoxicity of MTT assay shows in graphical abstract.
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Objective:To identify the morphological alterations in the Golgi apparatus of skin fibroblasts in spinocerebellar ataxia type 3 (SCA3) patients.Methods:In this study, 3 SCA3 patients and 3 healthy volunteers were obtained in the First Affiliated Hospital of Zhengzhou University from 2016 to 2020. The cytosine, adenine, and guanine repeats of 3 SCA3 patients were 14/76, 20/80 and 21/82, respectively. Tissue mass culture was used to amplify skin fibroblasts derived from SCA3 patients and healthy volunteers. Cell viability and apoptosis were detected using cell counting kit-8 assay and flow cytometry, respectively. Western blotting and immunofluorescence assay were used to detect the protein expression of ataxin-3, Golgi reassembly stacking protein 2 (GORASP2), and Golgi matrix protein 130 (GM130) in the skin fibroblasts. The morphology of the Golgi apparatus in skin fibroblasts was detected using transmission electron microscopy.Results:Tissue culture of skin fibroblasts of both SCA3 patients and healthy volunteers was successfully established. The patient-derived dermal fibroblasts expressing mutant ataxin-3 protein exhibited reduced cell viability ( t=5.06,P=0.007), increased apoptosis ( t=3.77, P=0.020), fragmentation of the Golgi apparatus, increased expression of GM130 ( t=5.23, P=0.006), and decreased expression of GORASP2 ( t=4.35, P=0.012). Transmission electron microscopy revealed that the Golgi apparatus was disorganized in skin fibroblasts. Conclusion:Fragmentation of the Golgi apparatus occurs in the skin fibroblasts of SCA3 patients, and abnormal morphology and structure of the Golgi apparatus may be involved in the pathogenesis of SCA3.
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Objective: To explore the impact of fucoxanthin on oxidized low-density lipoprotein (OxLDL)-induced stress and inflammation in human endothelial cells and its underlying mechanisms. Methods: HUVECs were treated with OxLDL and/or fucoxanthin for a range of time points and concentrations. We evaluated the effects of fucoxanthin on OxLDL-induced HUVECs using the MTT assay, reactive oxygen species accumulation assay, ELISA, RT-PCR, immunofluorescence, and Western blotting. Results: Fucoxanthin enhanced the cell viability in a dose dependent manner after OxLDL exposure. Furthermore, fucoxanthin pretreatment significantly decreased OxLDL-induced reactive oxygen species production and prevented the activation of the nuclear factor kappa-B pathway, which led to substantial suppression of pro-inflammatory gene expressions. OxLDL-induced upregulation of interleukin-6, intercellular adhesion molecule-1, vascular cell adhesion molecule-1, interleukin-1β, monocyte chemotactic protein-1, cyclooxygenase-1, and tumor necrosis factor-α was significantly reduced by fucoxanthin. Conclusions: Fucoxanthin can inhibit OxLDL-induced vascular inflammation and oxidative stress in HUVECs by targeting Nrf2 signaling pathways.
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Aim To explore and optimize the different separation and purification methods of neural stem cells (NSCs) , and compare the cell viability and purity after separation and purification.Methods (X) Separation of NSCs: The brains of ICR mice born within 24 h were isolated ( the cerebellum and olfactory bulb removed) , chopped and digested by mechanical pipetting, trypsin digestion, and PDD enzyme digestion.After sieving and centrifugation, the brain tissues were inoculated at a concentration of 1 X 10 • L , and cultured in serum-free medium to observe and compare the activity, proliferation and number of miscellaneous cells.(2) Purification of NSCs: The brain tissues of mice born within 24 h were isolated, and NSCs were purified by differential centrifugation and sieving, and the purity was identi-fied by immunofluorescence.Results NSCs separated by PDD enzyme digestion had high survival rate and rapid proliferation rate; the Nestin positive rate of NSCs in both Sieving assay and Differential centrifugation assay groups was higher than that of primary NSCs (59.14% ± 0.16% , P <0.05) , and the former (93.54% ± 0.02%) was higher than the latter (86.12% ± 0.04% ) .Conclusions The three separation methods can obtain a large amount of NSCs.Among them, the PDD enzymatic digestion method has little damage to the NSCs, and the neuro-sphere proliferation speed is fast; the NSCs obtained by the sieving assay have higher purity.
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ABSTRACT@#This study evaluated the cytotoxicity of four bioceramic root canal sealers (bioceramic sealers): GuttaFlow Bioseal (GB), MTA Fillapex, CeraSeal Bioceramic root canal sealer (CS), and iRoot SP root canal sealer (iRSP). The viability of human gingival fibroblast (HGF) cells was used to evaluate the cytotoxicity of these bioceramic sealers. HGF cells were cultured and exposed to bioceramic sealer extracts for 24 hours, 48 hours and 72 hours at 37°C in an incubator humidified with 5% CO2. The 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide or MTT assay was conducted to determine cell viability at each incubation period and compared among all bioceramic sealers. The Kruskal-Wallis test revealed statistically significant differences between the positive control group and MTA Fillapex, MTA Fillapex and GB, and between GB and iRSP with p < 0.05. However, no statistical differences were found in cell viability for each material across all the incubation periods. GB was the least cytotoxic bioceramic sealer with cell viability exceeding 90% throughout the 72-hour incubation followed by CS, iRSP, and MTA Fillapex with non-cytotoxicity after 72-hour incubation, mild cytotoxicity after 72-hour incubation, and mild cytotoxicity after 72-hour incubation, respectively. However, iRSP showed moderate cytotoxicity, and MTA Fillapex was severely cytotoxic (< 30% cell viability) after 24-hour incubation.
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Root Cause Analysis , Dental Pulp TestABSTRACT
@#In this study, the conjugate of eicosapentaenoic acid (EPA) and hyaluronic acid (HA) was synthesized and the anti-hepatoma activities in vitro were evaluated.The hyaluronic acid-eicosapentaenoic acid (HA-EPA)nanoparticle was synthesized by linking eicosapentaenoic acid with hyaluronic acid with cystamine.The structure of HA-EPA was characterized by nuclear magnetic resonance (1H NMR) and Fourier transform infrared spectroscopy (FT-IR).Laser particle sizer and Zeta potential analyzer were used to detect the size and potential of HA-EPA.MTT assay was used to detect the anti-proliferative effect of HA-EPA on HepG2, Huh-7 and LX-2 cells in vitro.The effects of HA-EPA nanoparticles on the proliferation and apoptosis of HepG2 cells in vitro were investigated by EdU staining and TUNEL staining. The apoptosis was further confirmed by flow cytometry.The effect of HA-EPA nanoparticles on the migration and invasion of HepG2 cells was demonstrated by transwell and invasion experiments.The results of 1H NMR showed that HA-EPA was successfully synthesized, and the grafting rate of EPA on HA was (40 ± 5) %. The structure of HA-EPA was further confirmed by FT-IR.The particle size was (162.5 ± 10.2) nm, and the potential was -(4.47 ± 0.31) mV.MTT results showed that, with the prolongation of drug treatment time, HA-EPA showed a better inhibitory effect on the activity of HepG2 and Huh-7 cells than EPA under the same EPA content.After treated for 48 hours, the toxicity of HA-EPA to LX-2 cells was less than that of EPA.The results of 24-hour proliferation, apoptosis, migration and invasion of HepG2 showed that, the graft of hyaluronic acid improved the ability of EPA to inhibit proliferation, promote apoptosis, migration and invasion of HepG2 cells (P < 0.001), indicating that grafting of HA can significantly enhance the inhibitory effect of EPA on liver cancer with some role in reducing toxicity.
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OBJECTIVE@#To investigate the effect of PPP2R5C to the activity of Molt-4 cells in childhood acute T lymphocytic leukemia and its mechanism.@*METHODS@#The small interfering RNA (siRNA) technology targeting PPP2R5C gene was used to down-regulate the expression of PPP2R5C in Molt-4 cells. At the same time, a blank control group, a negative control group and a 17-DMAG group were set up. The cells in the negative control group were transfected with siRNA-NC, the cells in 17-DMAG group were treated with the HSP90 inhibitor 17-DMAG at a final concentration of 6.4 μmol/L for 48 h. Real-time fluorescent quantitative PCR (RT-qPCR) and Western blot were used to detect transfection efficiency; CCK-8 method was used to detect the proliferation activity of the cells in each group, EdU was used to detect the proliferation level of the cells in each group, flow cytometry was used to detect the cell cycle distribution ratio of the cells in each group, Annexin V-FITC/PI staining was used to detect the apoptosis of the cell, RT-qPCR and Western blot were used to detect the expression changes of heat shock protein 90 (HSP90) and glucocorticoid receptor (GR) of the cells in each group.@*RESULTS@#After Molt-4 cells were transfected with siRNA-PPP2R5C, the expression of PPP2R5C mRNA and protein in the cells were down-regulated significantly compared with those in the blank control group and the si-NC group (P<0.05); compared with cells in the blank control group and the si-NC group, the proliferation activity of the cells in the siRNA-PPP2R5C group and the 17-DMAG group significantly decreased (P<0.05), and the rate of EdU positive cells was significantly reduced (P<0.05); the proportion of the cells in G1 phase decreased while the proportion of the cells in G2 phase increased (P<0.05), the apoptosis rate of the cells also increased significantly (P<0.05); in addition, the expression of PPP2R5C mRNA and protein of the cells in siRNA-PPP2R5C group was significantly down-regulated compared with those in the blank control group and si-NC group (P<0.05). The expressions of PPP2R5C mRNA and protein in the 17-DMAG group were also significantly down-regulated compared with those in the blank control group and si-NC group (P<0.05).@*CONCLUSION@#Down-regulation of PPP2R5C gene expression can inhibit Molt-4 cell activity in childhood acute T lymphocytic leukemia, block the cells in G2 phase, and promote cell apoptosis, the mechanism may be related to the inhibition of HSP90-GR signaling pathway.
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Child , Humans , Apoptosis , Cell Line, Tumor , Cell Proliferation , Cell Survival , HSP90 Heat-Shock Proteins , Precursor T-Cell Lymphoblastic Leukemia-Lymphoma , RNA, Small Interfering , Receptors, GlucocorticoidABSTRACT
Abstract Objective: To assess the effects of different peracetic acid (PAA) formulations on smear layer (SL) removal, dentine erosion, cytotoxicity, and antibiofilm activity. Methodology: SL removal and dentine erosion were assessed using 90 premolars, distributed into six groups, according to final irrigation: PAA formulations (1% Sigma, 1% Bacterend OX, 1% Arposept, and 0.09-0.15% Anioxyde), 17% ethylenediaminetetraacetic acid (EDTA), and water (control). Cytotoxicity was assessed by methyl-thiazol-tetrazolium (MTT) and neutral red assays. Antibacterial and antibiofilm effectiveness was evaluated against Enterococcus faecalis. For cytotoxicity and antibiofilm activity assessment, the 2.5% NaOCl was also included. Results: EDTA, Sigma, and Bacterend OX removed more SL than Arposept, Anioxyde, and water (p<0.05). EDTA caused more severe dentine erosion than Sigma and Bacterend OX (p<0.05). Sigma and Bacterend OX had higher cytotoxicity than the other solutions (p<0.05). NaOCl, Bacterend OX, Sigma, and Anioxyde significantly reduced E. faecalis colony-forming units (CFU) (p<0.05). The 2.5% NaOCl solution promoted greater biofilm biomass reduction (p<0.05) than the other solutions. All PAA formulations promoted greater biomass reduction than 17% EDTA (p<0.05). Conclusions: Although Sigma and Bacterend OX had higher cytotoxicity, they had a SL removal capability similar to that of EDTA, were as effective as NaOCl against E. faecalis biofilm, and promoted less dentine erosion than EDTA. Arposept and Anioxyde failed to remove the SL, had lower cytotoxicity, and showed less bacterial activity than NaOCl.
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Objective:To determine any effect of repeated thermal stimulation on the viability and functioning of inflamed human umbilical vein endothelial cells (HUVECs).Methods:Well-cultured HUVECs were divided into a normal group, a model group, a thermal stimulation 5 times group (group A), a thermal stimulation 9 times group (group B) and a thermal stimulation 13 times group (group C) and cultured under the same conditions. The normal group was not given any intervention. The model group was stimulated with 1μg/mL lipopolysaccharide for 1 hour. Groups A, B and C were first subjected to 5, 9 and 13 rounds of repeated thermal stimulation, each round lasting 4 minutes at 43℃ and 1 minute at room temperature. They were then incubated for one hour at 37℃ under a 5% CO 2 atmosphere with 1μg/mL lipopolysaccharide. Cell viability and the expression of NF-κB were evaluated using methyl thiazolyl tetrazolium and immunofluorescence assays. The levels of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were determined by enzyme-linked immunosorbent assay. Results:After the intervention, the average cell viability of the model group and of groups A and C was significantly lower than that of the normal group, while that of group B was significantly higher. After the intervention, the average NF-κB expression in the normal group was significantly different from that in the others, with group B′s level significantly different from that of the model group. After the treatment, the average expression of ICAM-1 and VCAM-1 in the model group had increased significantly, while that in groups A, B and C had decreased significantly compared with the normal group. The levels of groups A, B and C were then significantly different from that of the model group. The average ICAM-1 level of group B was significantly different from those of groups A and C.Conclusions:Repeated thermal stimulation can protect inflamed HUVECs and reduce the expression of HUVEC adhesion molecules.
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Objective:To screen 17-AAG-M-induced differentially expressed miRNAs in human non-small cell lung cancer (NSCLC) A549 cells under X-ray and evaluate its effect on radio-sensitivity.Methods:A549 cells were treated with 17-AAG-M and 4 Gy. Total RNA was extracted for microarray screening. The expression of the miRNAs of interest in the tumor was observed by public database. The target miRNAs were analyzed by using GO and KEGG pathways, and verified by qPCR. The effect of target miRNAs on the survival rate and proliferation of A549 cells under X-ray was evaluated by MTT and clone formation assays. The radio-sensitivity of the target miRNAs was analyzed by the single-hit multi-target model formula.Results:20 differentially expressed miRNAs were screened. The down-regulated hsa-miR-30a-3p showed a close correlation with lung cancer in the database. It was involved with 50 biological processes including cell proliferation and affected the MAPK signaling pathway, cancer-related pathways and cell cycle, etc. Compared with the 17-AAG-M group, the relative expression level of hsa-miR-30a-3p under the action of 17-AAG-M and X-ray was down-regulated from 2.42 to 0.16. hsa-miR-30a-3p inhibited the survival rate of A549 cells (survival rate: 78.52%) and further decreased to 69.00% under X-ray. Up-regulation of hsa-miR-30a-3p expression inhibited the proliferation of tumor cells and increased the radio-sensitivity of A549 cells. The radio-sensitization ratio was 1.18. The above performance became more obvious under the action of 17-AAG-M.Conclusions:In A549 cells, hsa-miR-30a-3p is differentially expressed under the action of 17-AAG-M and X-ray. Moreover, up-regulation of the expression level of hsa-miR-30a-3p in A549 cells can reduce the viability and proliferation of tumor cells, and increase the radio-sensitivity of tumor cells. The inhibition effect of X-ray combined with 17-AAG-M upon tumors can be strengthened.
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Objective:To compare the effects of different intraocular infusion solutions on histology and function of retina.Methods:Human corneal endothelial cells (HCEC), human retinal pigment epithelium (HRPE) cells and rat retinal ganglion cells (RGC) were divided into normal control group, balanced saline solution (BSS) group and compound electrolyte intraocular irrigating solution (CEIIS) group, and the cells were cultured in 10% DMEM/F12 medium, BSS and CEIIS for 12, 24 and 48 hours, respectively, according to grouping.The proliferation absorbance value of cultured cells was measured by cell counting kit-8 (CCK8) method.The expression of apoptosis related proteins in cultured cells was detected by cellular immunofluorescence staining.The cell apoptosis rate and cell cycle were measured by flow cytometry.The mitochondrial damage was detected by lactate dehydrogenase (LDH) and succinate dehydrogenase (SDH) quantitative detection kit.Fifteen New Zealand white rabbits were randomly divided into control group ( n=3), BSS group ( n=6) and CEIIS group ( n=6). The left eyes were taken for vitrectomy and different intraocular perfusion fluids were used during vitrectomy according to grouping.The retinal function of operative eyes was measured by flash electroretinogram (ERG) before operation and 24 hours after operation, and the structural changes of each layer of retina were detected by optical coherence tomography (OCT). The early apoptosis of retinal cells was detected by TUNEL staining.The expressions of cytochrome C and bax protein in retina were detected by immunohistochemical staining.The ultrastructural changes of retina were observed under a transmission electron microscope.The use and care of animals complied with the ARVO statement.This study protocol was approved by an Ethics Committee of Peking University People's Hospital (No.2019PHE059). Results:The three kinds of cultured cells in BSS and CEIIS groups were damaged in various degrees.With the extension of culture time, proliferated cells were decreased and the number of apoptotic cells was increased.Compared with the BSS group, cultured cells in the CEIIS group were dense and in orderly arrangement with uniform morphology and size.The apoptosis rates of HRPE cells and RGC in the BSS group were (37.157±6.918)% and (29.993±12.330)%, respectively, which were significantly higher than (4.163±1.310)% and (6.337±1.903)% in the CEIIS group ( P=0.003, 0.045). There was no significant difference in G0/G1+ S phase ratio of HCEC and HRPE cells among the normal control group, BSS group and CEIIS group (HCEC: F=2.226, P=0.189; HRPE: F=2.634, P=0.151), and the proportion of G2/M division arrest phase of RGC in the BSS group was significantly higher than that in the normal control group and CEIIS group ( P=0.047, 0.024). The proliferation absorbance values of HCEC, HRPE cells and RGC in the CEIIS group were significantly higher than those in the BSS group at each culture time point (all at P<0.05). The fluorescence intensity of cytochrome C, bax, caspase-3 and caspase-9 proteins in the BSS group was stronger than that in the normal control group and CEIIS group, and the fluorescence intensity of bcl-2 was weaker than that in the CEIIS group, and the fluorescence intensity of zonula occluden-1 (ZO-1) was weaker than that in the normal control group and CEIIS group.The release level of LDH in the BSS group was significantly higher than that in the CEIIS group at different time points (all at P<0.001). After 48 hours of culture, the release level of SDH in the BSS group was significantly higher than that in the CEIIS group ( P<0.05). No retinal histological abnormalities was found through OCT examination of rabbit eyes after vitrectomy in the two groups, but transmission electron microscopy showed that there were different degrees of loose arrangement of retinal photoreceptor cells, a large number of photoreceptor outer membrane discs falling off and vacuolar degeneration in the two groups, especially in the BSS group.TUNEL staining showed that the apoptotic cells were mainly located in the inner nuclear layer and RGC layer.The number of apoptotic retinal cells was (135.2±22.8)/high-power field of vision in the BSS group, which was significantly higher than (81.3±17.7)/high-power field of vision in the CEIIS group ( t=4.175, P=0.002). Full field flash ERG showed that the amplitudes of scotopic 3.0 ERG a- and b-wave in the CEIIS group after operation were significantly lower than those before operation, but the differences were not statistically significant (all at P>0.05). The amplitudes of scotopic 3.0 ERG a- and b-wave in the BSS group after operation were significantly lower than those before operation ( P=0.026, 0.010). Conclusions:In vivo and in vitro research results show that compared with BSS, there were few apoptotic cells in retinal tissue after vitrectomy perfused by CEIIS.
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Objective To observe the effect of cryptotanshinone on the ferroptosis of human liver cancer HepG2 cells. Methods The viability of the HepG2 cells cultured