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1.
China Occupational Medicine ; (6): 657-661, 2016.
Article in Chinese | WPRIM | ID: wpr-877002

ABSTRACT

OBJECTIVE: To explore the effects of UVB and Nano-CaCO_3 co-treatment in inhibiting cell proliferation and inducing cell apoptosis in human HaCaT keratinocytes,and to explore its possible mechanisms. METHODS: HaCaT cells( logarithmic growth phase) were divided into control group,UVB group,Nano-CaCO_3 group and co-treatment group. UVB group and co-treatment group were irradiated with UVB irradiation with cumulative exposure dose of 2. 97 × 10~(-2)J / cm~2.Control group and Nano-CaCO_3 group were irradiated with UVB irradiation with cumulative exposure dose of zero on equal terms. After that,control group and UVB group were treated with 10. 00% phosphate buffer solution in high-sugar Dulbecco's modified Eagle's medium( DMEM) and incubated,Nano-CaCO_3 group and co-treatment group were treated with high-sugar DMEM with Nano-CaCO_3 at 250 mg / L mass concentration and incubated. Subsequently,HaCaT cells were harvested at incubation time of 0,6,12,18 and 24 hours. Then methyl thiazolyl tetrazolium assay was performed to estimate cellular proliferative activity,flow cytometry was used to detect cell apoptosis,Western blot was used to detect the expression of P53 and Caspases-3 protein. RESULTS: Contrast to control group at the parallel incubation time points of 6-24 hours,the cell viability of HaCaT cells was significant decreased in the other three groups( P < 0. 05) except for UVB group at incubation time of 6 hours( P > 0. 05). The cell viability of co-treatment group was lower than UVB group at all the incubation time( P < 0. 05),and lower than Nano-CaCO_3 group at incubation time of 18 and 24 hours( P < 0. 05).The apoptosis rate of HaCaT cells in UVB group was higher than control group( P < 0. 05),and which in co-treatment group was higher than the other three groups( P < 0. 05). Contrast to control group,the protein expressions of P53 and Caspases-3 in HaCaT cells were upregulated in UVB group and Nano-CaCO_3 group. In co-treatment group,the protein expressions of P53 and Caspases-3 were upregulated contrast to the other three groups. CONCLUSION: Contrast to single damage of UVB or Nano-CaCO_3,co-treatment of UVB with Nano-CaCO_3 increased damage to HaCaT cells,likely by inhibiting proliferative activity,inducing apoptosis,and enhancing protein expressions of P53 and Caspases-3.

2.
China Occupational Medicine ; (6): 400-407, 2016.
Article in Chinese | WPRIM | ID: wpr-876963

ABSTRACT

OBJECTIVE: To explore the role of 14-3-3σ and TPD52L1 in apoptosis,proliferation and cell cycle in HaCaT cells treated with low dose ultraviolet B( UVB). METHODS: i) HaCaT cells in logarithmic growth phase were exposed to UVB irradiation( cumulative exposure dose was 2. 97 ×10~(-2)J /cm~2) and were harvested after culture for 6-48 hours.HaCaT cells with no UVB irradiation were set as the control( pseudo-irradiation). The apoptosis of cells was detected by flow cytometry. The cells were divided into the control group and the UVB group exposed to UVB irradiation. They were harvested after being cultured for 0-72 hours,and monotetrazolium assay was used to detect the proliferation ability of cells.ii) HaCaT cells were randomly divided into control group( given pseudo-irradiation) and UVB group. Cells were cultured for 0-24 hours and then harvested. Flow cytometry was used to detect the proportion of G2 / M phase of cells. iii) After HaCaT cells were exposed to low dose of UVB irradiation and cultured for 3-24 or 3-30 hours,they were harvested. In addition,the control was treated with pseudo-irradiation. Real-time fluorescence quantitative polymerase chain reaction was used to detect the mRNA relative expression levels of 14-3-3σ and TPD52L1. The Western blotting was used to detect the protein relative expression of 14-3-3σ and TPD52L1 in cells. RESULTS: i) With 24 hours after UVB irradiation as the cell cycle observation time,the apoptosis of HaCaT cells reached to the maximum value and the proliferation ability to the minimum value( P < 0. 05). ii) After irradiation,the proportion of G2 / M phase in the control group showed a periodic change that it maintained at the highest levels after 6-12 hours of exposure to pseudo-irradiation,and at the time point of 18 hours it was decreased to the lowest value,the proportion of G2 / M phase in the UVB group was higher than that in control group at time points of 6,12 and 18 hours after irradiation( P < 0. 05),and sustained at the highest value,which resulted in significant G2 / M phase block. iii) After UVB treatment,the relative expression of mRNA and protein of 14-3-3σ first increased and then decreased. The relative expression level of mRNA of 14-3-3σ reached the highest level in 3 hours after irradiation( P < 0. 05),and maintained at the peak level at time points of 3-12 hours( P < 0. 05),and then gradually decreased until it returned to normal level 24 hours after irradiation; the relative expression of 14-3-3σ protein began to rise at time point of 6 hours after irradiation( P < 0. 05),and reached the peak value at time point of 18 hours( P < 0. 05),and then gradually decreased,but it did not recover to normal level after irradiation for 30 hours. The relative expression of mRNA of TPD52L1 first decreased and then increased. It reached the lowest level in 12 hours after irradiation( P <0. 05),and then increased gradually and reached the peak value after 24 hours( P < 0. 05). The relative expression of TPD52L1 protein first increased and then decreased. It got to the peak 24 hours after irradiation( P < 0. 05),and 30 hours after irradiation it returned to normal level( P < 0. 05). CONCLUSION: 14-3-3σ and TPD52L1 might jointly play an important role in cell apoptosis,proliferation and G2 / M phase block in HaCaT cells induced by low dose UVB irradiation.

3.
Article in English | WPRIM | ID: wpr-320395

ABSTRACT

<p><b>OBJECTIVE</b>To examine UVB-induced responses in normal human keratinocytes (HaCaT) and epidermoid carcinoma cells (A431) at the cellular and molecular level, and investigated the protective effect of salidroside.</p><p><b>METHODS</b>Cells irradiated by UVB at various dosage and their viability was assessed by MTT assays, cell cycle was analysed by flow cytometry. The expression of NF-κB, BCL-2, and CDK6 after 50 J/m(2) UVB irradiation were detected by RT-PCR and western blotting.</p><p><b>RESULTS</b>Our results confirmed greater tolerance of A341 cells to UVB-induced damage such as cell viability and cell cycle arrest, which was accompanied by differential expression changes in NF-κB, BCL-2, and CDK6. UVB exposure resulted in HaCaT cells undergoing G(1)-S phase arrest. When treated with salidroside, HaCaT survival was significantly enhanced following exposure to UVB, suggesting great therapeutic potential for this compound.</p><p><b>CONCLUSION</b>Taken together, our study suggests that A431 respond differently to UVB than normal HaCaT cells, and supports a role for NF-κB, CDK6, and BCL-2 in UVB-induced cell G(1)-S phase arrest. Furthermore, salidroside can effectively protect HaCaT from UVB irradiation.</p>


Subject(s)
Antioxidants , Pharmacology , Apoptosis , Radiation Effects , Carcinoma, Squamous Cell , Cell Cycle Checkpoints , Cell Line, Tumor , Cell Survival , Radiation Effects , Gene Expression Regulation, Neoplastic , Glucosides , Pharmacology , Humans , Keratinocytes , Radiation Effects , Phenols , Pharmacology , Ultraviolet Rays
4.
Article in Chinese | WPRIM | ID: wpr-267673

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the biological effects of ultraviolet B (UVB) irradiation on human bronchial epithelial cells (16HBE cells) and explore the possible mechanism.</p><p><b>METHODS</b>The survival rates of 16HBE cells were detected by MTT assay at 12 h after UVB irradiation at different doses (0, 10, 30, 50, 70, and 100 J/m(2)) or at 50 J/m(2) for different durations (2, 4, 8, 12, and 24 h). The DNA ladder was detected by agarose gel electrophoresis, the cell cycle changes were analyzed by flow cytometry, and the expression of nuclear factor-κB (NF-κB)/p65 protein was assayed by Western blotting following the exposures.</p><p><b>RESULTS</b>UVB irradiation of the cells resulted in lowered cell survival rates, DNA fragmentation, S phase arrest and up-regulation of NF-κB/p65 protein expression.</p><p><b>CONCLUSIONS</b>UVB irradiation can induce growth inhibition and apoptosis of 16HBE cells, in which process NF-κB protein may play a key role.</p>


Subject(s)
Apoptosis , Radiation Effects , Bronchi , Cell Biology , Cell Line , Cell Survival , Radiation Effects , Endothelial Cells , Cell Biology , Radiation Effects , Humans , NF-kappa B , Metabolism , Ultraviolet Rays
5.
Article in Chinese | WPRIM | ID: wpr-340810

ABSTRACT

<p><b>OBJECTIVE</b>To study the effect of radiation injury on nitric oxide (NO) concentration in mouse peripheral blood and liver.</p><p><b>METHODS</b>NIH mice were subjected to gamma-ray exposure at 9.0 Gy and transferred immediately in room temperature condition. NO concentrations in the liver and peripheral blood were examined before and at different time points after the exposure.</p><p><b>RESULTS</b>Compared to that before exposure, NO concentration in the peripheral blood and liver significantly increased after gamma-ray exposure. NO concentration in the peripheral blood began to increase 3 h after the exposure, but that in the liver increased till 6 h after the exposure.</p><p><b>CONCLUSION</b>Radiation can cause the increase of NO concentration in the peripheral blood and liver, but different tissues may exhibit different response intensities to radiation.</p>


Subject(s)
Animals , Gamma Rays , Liver , Metabolism , Radiation Effects , Male , Mice , Nitric Oxide , Blood , Metabolism , Radiation Injuries, Experimental , Blood , Metabolism , Time Factors
6.
Article in Chinese | WPRIM | ID: wpr-270191

ABSTRACT

<p><b>OBJECTIVE</b>To explore the involvement of p38 and ERK signal transduction pathways in UVB-induced cell apoptosis.</p><p><b>METHODS</b>HaCat cells were exposed to UVB irradiation for 1, 3, 5, 10, and 15 min, respectively, after which the cell survival was assessed using MTT assay, and the cell apoptosis observed under fluorescent microscope with Hoechst staining. Western blotting was used to examine the possible signal transduction pathway involved in the cell apoptosis following the exposures.</p><p><b>RESULTS</b>For the same incubation time following the exposure, the cell survival rate decreased gradually with the increase of UVB irradiation dose. At a fixed UVB irradiation dose, prolonged cell incubation following the exposure resulted in decreased cell survival rate, which, however, began to increase after the minimum rate was reached. At different UVB doses, cell exposure for 5 min caused the highest cell apoptosis rate, which peaked at 12 h during the post-irradiation incubation. The expressions of p38 and p53 were significantly decreased while p44/42 expression remained unchanged in the exposed cells.</p><p><b>CONCLUSION</b>UVB irradiation can induce growth inhibition and apoptosis of HaCat cells in a dose- and time-dependent manner, and p38 pathway other than ERK pathway is probably involved in UVB-induced cell apoptosis.</p>


Subject(s)
Apoptosis , Radiation Effects , Cells, Cultured , Extracellular Signal-Regulated MAP Kinases , Metabolism , Humans , Keratinocytes , Cell Biology , Metabolism , Radiation Effects , Signal Transduction , Radiation Effects , Ultraviolet Rays , p38 Mitogen-Activated Protein Kinases , Metabolism
7.
Article in Chinese | WPRIM | ID: wpr-337372

ABSTRACT

<p><b>OBJECTIVE</b>To study the biodistribution of L-[S-methyl-(11)C]-methioine ((11)C-MET) and explore its clinical application in positron emission tomography (PET) for brain tumor detection.</p><p><b>METHODS</b>Twenty-four Wistar rats and divided into 6 equal groups and injected with (11)C-MET through the tail vein and killed by decollation at 5, 10, 20, 30 and 40 min after injection, respectively. The liver, brain, blood, heart, lung, kidney, and spleen were harvested to measure the radioactivity and calculate the biodistribution of (11)C-MET. PET imaging with (11)C-MET was performed in 6 normal volunteers and 30 patients with pathologically confirmed brain gliomas.</p><p><b>RESULTS AND CONCLUSION</b>(11)C-MET showed high blood uptake and a long retention in the tumor mass, therefore can be a valuable tracer for PET imaging of brain tumor and the hypophysis.</p>


Subject(s)
Adult , Aged , Animals , Brain , Diagnostic Imaging , Metabolism , Pathology , Brain Neoplasms , Diagnosis , Diagnostic Imaging , Metabolism , Carbon Radioisotopes , Female , Glioma , Diagnosis , Diagnostic Imaging , Metabolism , Humans , Injections, Intravenous , Male , Middle Aged , Positron-Emission Tomography , Methods , Radiopharmaceuticals , Pharmacokinetics , Rats , Rats, Wistar , Sensitivity and Specificity , Tissue Distribution , Vitamin U , Pharmacokinetics
8.
Article in English | WPRIM | ID: wpr-337357

ABSTRACT

The aromatic-ring-hydroxylating dioxygenase is a key enzyme that initiates the biodegradation of polycyclic aromatic hydrocarbons in bacteria. In the present study, a novel dioxygenase sequence was cloned from Terrabacter sp. FLO using a genome walking method. The dioxygenase was cloned into pET17 and actively expressed in E.coli BL21 (DE3) in co-expression with electron transfer chain proteins. The recombinant dioxygenase was found to transform phenanthrene, fluorene, pyrene and fluoranthene into the cis-dihydrodiol metabolites.


Subject(s)
Actinomycetales , Genetics , Bacterial Proteins , Genetics , Metabolism , Biodegradation, Environmental , Cloning, Molecular , Dioxygenases , Genetics , Metabolism , Electrophoresis, Polyacrylamide Gel , Escherichia coli , Genetics , Metabolism , Fluorenes , Metabolism , Hydroxylation , Molecular Sequence Data , Phenanthrenes , Metabolism , Polycyclic Aromatic Hydrocarbons , Metabolism , Pyrenes , Metabolism , Recombinant Proteins , Metabolism
9.
Article in Chinese | WPRIM | ID: wpr-268104

ABSTRACT

<p><b>OBJECTIVE</b>To obtain specific small interfering RNAs (siRNA) for hepatitis B virus (HBV) and evaluate their interfering effect.</p><p><b>METHODS</b>Three siRNAs were transfected into HepG2.2.15 cells, and the amount of HBV mRNA in the cell culture medium was quantified with real-time fluorescence quantitative RT-PCR. HBsAg in the culture media was assayed with Western blotting at different time points after transfection.</p><p><b>RESULTS</b>The cells transfected with specific siRNAs showed decreased levels of HBV mRNA and HBsAg (P<0.05), but those with nonspecific siRNA transfection as the negative control did not show such changes (P>0.05).</p><p><b>CONCLUSION</b>Specific siRNA can significantly inhibit protein expression and mRNA synthesis of HBV in HepG2.2.15 cells in vitro.</p>


Subject(s)
Cell Line, Tumor , Hepatitis B Surface Antigens , Hepatitis B virus , Physiology , Humans , RNA, Messenger , RNA, Small Interfering , Pharmacology , RNA, Viral , Reverse Transcriptase Polymerase Chain Reaction , Transfection , Virus Replication
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