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ObjectiveTo investigate the effect and mechanism of Dendrobium mixture （DMix）-containing serum on high glucose-induced podocyte injury in mice. MethodThe MPC5 mouse glomerular podocytes were cultured in vitro， and the optimal glucose concentration for modeling， modeling time， and concentration of DMix-containing serum for administration were determined. The cells were classified into normal （5.5 mmol·L-1 glucose+10% blank serum）， model （30 mmol·L-1 glucose+10% blank serum）， DMix-containing serum （30 mmol·L-1 glucose+10% DMix-containing serum）， ferroptosis inhibitor （Fer-1， 30 mmol·L-1 glucose+10% blank serum+1 μmol·L-1 Fer-1） groups. The corresponding kits were used to measure the levels of Fe2+ and lactate dehydrogenase （LDH） in cells. Enzyme-linked immunosorbent assay was employed to determine the content of glutathione （GSH） and lipid peroxide （LPO） in cells. Fluorescence probe was used to measure the reactive oxygen species （ROS） level. Real-time fluorescence quantitative polymerase chain reaction and Western blotting were employed to determine the mRNA and protein levels， respectively， of Wilms' tumor-1 （WT-1）， desmin， long chain acyl-CoA synthase 4 （ACSL4）， and glutathione peroxidase 4 （GPX4） in podocytes. ResultCompared with the blank group， the intervention with 30 mmol·L-1 glucose for 48 h reduced podocyte viability （P<0.01）， and the 10% DMix-containing serum showed the most significant improvement in podocyte viability （P<0.01）. Compared with the normal group， the model group presented elevated levels of Fe2+， LDH， LPO， and ROS， lowered GSH level， up-regulated mRNA and protein levels of desmin and ACSL4， and down-regulated mRNA and protein levels of WT-1 and GPX4 （P<0.01）. Compared with the model group， the DMix-containing serum lowered the Fe2+， LDH， LPO， and ROS levels， elevated the GSH level， down-regulated the mRNA and protein levels of desmin and ACSL4， and up-regulated the mRNA and protein levels of WT-1 and GPX4 in podocytes （P<0.05， P<0.01）. ConclusionDMix-containing serum exerts a protective effect on high glucose-induced podocyte injury by inhibiting ferroptosis.
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ObjectiveTo explore the effect of Babaodan （BBD） on the NOD-like receptor pyrin domain containing 3/cysteine aspartate-specific protease-3 （NLRP3/Caspase-1） pathway proteins in mice with acetaminophen （APAP）-induced acute liver injury. MethodC57BL/6 mice were randomly grouped， and BBD （75， 150， 300 mg·kg-1， ig） was administered twice a day for three days. After 2 hours of the last administration， the mice were treated with APAP （400 mg·kg-1， ip）， and the eyeballs were removed to collect blood after 14 hours. Then they were sacrificed by cervical dislocation for sample collection. Hematoxylin-eosin （HE） staining was used to observe the morphological changes of liver tissue cells， and biochemical methods were used to detect the activities of alanine aminotransferase （ALT）， aspartate aminotransferase （AST）， superoxide dismutase （SOD）， malondialdehyde （MDA） and myeloperoxidase （MPO） in serum of mice in each group. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was performed to determine the mRNA expression of tumor necrosis factor-α （TNF-α）， interleukin-1β （IL-1β） and IL-6， and Western blot was performed to determine the protein expression of cyclooxygenase-2 （COX-2）， inducible nitric oxide synthase （iNOS）， NLRP3， Caspase-1 and IL-18 in the liver of mice. ResultCompared with the conditions in normal group， the hepatic lobule structure of mice in the model group was partially destroyed， and the hepatic sinusoids were dilated. And the expression levels of ALT and AST in serum， the protein levels of NLRP3， Caspase-1， iNOS， IL-18 and COX-2 and the mRNA levels of IL-1β， IL-6 and TNF-α were increased （P<0.05， P<0.01）. Compared with the model group， the administration groups had improvement in liver cell rupture and hepatic sinusoidal compression， and a dose-dependent decrease in the levels of ALT and AST in serum as well as the protein levels of NLRP3， Caspase-1， iNOS， IL-18 and COX-2 and the the mRNA levels of IL-1β， IL-6 and TNF-α in liver tissue （P<0.05， P<0.01）. ConclusionBBD can reduce APAP-induced acute liver injury in mice. The mechanism may be related to anti-oxidative stress， inhibition of NLRP3/Caspase-1 pathway， and decreased expression levels of IL-1β， IL-18， TNF-α and IL-6.
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OBJECTIVE To establish a method for simultaneous determination of 15 bile acids in Tongren niuhuang qingxin pills， and to determine the contents of 15 batches of samples. METHODS Using dehydrocholic acid as internal standard， the determination was performed by ultra-high performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS） method. The determination was performed on Hypersil GOLD C18 column with methanol-0.1% formic acid solution as the mobile phase by gradient elution at the flow rate of 0.2 mL/min. The column temperature was 40 ℃ ， and the sample size was 2 µL. Using heated electrospray ion source， parallel reaction monitoring mode scanning was performed in negative ion mode. SPSS 24.0 software was used for chemical pattern recognition analysis of content determination results. RESULTS The 15 bile acid components had a good linear relationship with peak area （all R2≥0.998 9）； their precision， repeatability and stability were all good （all RSD≤5.49%）； the average recoveries were 93.8%-105.7% （RSD was 0.5%-5.8%）. The average contents of taurocholic acid， 7-oxodeoxycholic acid， 12-dehydrocholic acid， glycocholic acid， 3-oxo-7α， 12α-hydroxy-5β-cholanoic acid， taurochenodeoxycholic acid， 3α-hydroxy- 7-oxo-5β -cholanic acid， hyocholic acid， taurodeoxycholic acid sodium salt hydrate， hyodeoxycholic acid， cholic acid， glycochenodeoxycholic acid， glycodeoxycholic acid， chenodeoxycholic acid and deoxycholic acid were 670.56， 25.97， 10.54， 280.12， 4.04， 29.81， 182.98， 813.55， 120.95， 220.31， 797.37， 18.37， 68.59， 30.13， 59.82 μg/g， respectively. Both cluster analysis and principal component analysis divided 15 batches of Tongren niuhuang qingxin pills into 2 categories， S1-S12 as one category and S13-S15 as the other category. CONCLUSIONS The established method is accurate， sensitive and specific， and can determine many types of bile acids. It also can quickly achieve the quantitative analysis of 15 bile acids in Tongren niuhuang qingxin pills， which is suitable for the quality control of this drug.
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Objective:To analyze the causes of puncture wound infections induced by the high pressure resistant injectable PICC catheter in patients undergoing hematopoietic stem cell transplantation and management measures.Methods:linical data of 75 patients undergoing hematopoietic stem cell transplantation who were treated with the high pressure resistant injectable PICC catheter in our hospital from Nov.2017 to Nov.2019 were retrospectively analyzed.According to whether there were puncture wound infections, patients were divided into the infection group(n=26)and the non-infection group(n=49). Bacterial culture results of the infection group were recorded, and the related factors for puncture wound infections caused by the injectable PICC catheter were analyzed.Effective strategies to prevent high-risk factors, treatment frequency, treatment effect and healing time for patients with different degrees of puncture wound infections were discussed.Results:There were 26 patients in the infection group.The proportions of bacteria types associated with PICC catheter-related infections, in descending order, were as follows: Staphylococcus aureus(46.51%), Klebsiella pneumoniae(30.77%), Corynebacterium(15.38%)and others(7.69%). Significant differences were found in materials used, season of tube placement, timing of dressing changes, duration of catheterization, success rate of first tube placement and condition of dressing films between the non-infection and infection groups( t=5.5, 4.9, 5.0, 13.6, 9.4 and 6.2, all P<0.05). Logistic multi-factor analysis showed that non-U-shaped fixation, delay in dressing changes, long duration of tube placement, low success rate of first tube placement, and loose dressing films were the high-risk factors for PICC catheter-related infections( OR=2.78, 2.42, 3.16, 2.66 and 2.32, all P<0.05). Compared with patients with moderate and mild infections, patients with severe infections had a higher frequency of treatment, a lower total effectiveness rate and a longer healing time( F=10.353, 8.775 and 12.341, all P<0.05). Conclusions:Materials, timing of dressing changes, catheterization time, success rate of first tube placement and condition of dressing films are the high-risk factors for puncture wound infections caused by high pressure resistant injectable PICC catheters in patients undergoing hematopoietic stem cell transplantation.Developing effective intervention strategies can help control the incidence of wound infections.
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Traditional Chinese medicine (TCM) formulas have attracted increasing attention worldwide in the past few years for treating complex disease including rheumatoid arthritis. However, their mechanisms are complex and remain unclear. Guan-Jie-Kang (GJK), a prescription modified from "Wu Tou Decoction," was found to significantly relieve arthritis symptoms in rats with adjuvant-induced arthritis after 30-day treatment, especially in the 24 g/kg/day group. By analyzing 1749 targets related to 358 compounds in the five herbs of GJK, we identified the possible anti-arthritis pathways of GJK, including the calcium signaling and metabolic pathways. Bone damage levels were assessed by micro-computed tomography, and greater bone protective effect was observed with GJK treatment than with methotrexate. Receptor activator of nuclear factor κB ligand (RANKL)-RANK signaling, which is related to calcium signaling, was significantly regulated by GJK. Moreover, a target metabolomics assay of serum was conducted; 17 metabolic biomarkers showed significant correlations with treatment. An integrated pathway analysis revealed that pyruvate metabolism, purine metabolism, and glycolysis metabolism were significantly associated with the effects of GJK in arthritis treatment. Thus, this study establishes a new omics analytical method integrated with bioinformatics analysis for elucidating the multi-pathway mechanisms of TCM.
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Objective@#To explore the value of preventive nursing in the treatment of neonatal respiratory distress syndrome (NRDS) mechanical ventilation in premature infants.@*Methods@#A total of 45 NRDS preterm infants who received treatment during the preventive care program from February 1, 2018 to November 1, 2018 were selected as group A, 45 cases of NRDS preterm infants who received treatment during the preventive care program from January 1, 2017 to January 10, 2018 were included in group B for retrospective study. Baseline data, mechanical ventilation index, complications, and family satisfaction of the children were observed.@*Results@#The number of pulmonary surfactant applications, mechanical ventilation time, hospitalization time and hospitalization cost were (1.5±0.5) times, (92.5±13.8) h, (12.5±1.3) d, (26±4) thousand yuan in group A, and (2.6±0.4) times, (131.4±23.1) h, (16.0±2.8) d, (33±5) thousand yuan in group B, the highest oxygen saturation value was 468.9±42.1 in group A,401.2±22.3 in group B, there were significant difference between the two groups(t=7.334-11.524, P<0.05). The total incidence of complications during treatment was 15.6%(7/45) in group A and 48.9%(22/45), there was significant difference between the two groups (χ2=11.447, P<0.05). The total satisfaction rate of family members was 95.6%(43/45) in group A, which was significantly higher than that in group B (64.4%, 29/45). The difference was statistically significant (χ2=13.611, P<0.05).@*Conclusions@#Prophylactic care can be used in the mechanical ventilation of premature infants with NRDS, which can significantly reduce the incidence of complications, reduce the amount of pulmonary surfactant, shorten the mechanical ventilation time, and save the cost of treatment. It is worth promoting.
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Objective To explore the value of preventive nursing in the treatment of neonatal respiratory distress syndrome (NRDS) mechanical ventilation in premature infants. Methods A total of 45 NRDS preterm infants who received treatment during the preventive care program from February 1, 2018 to November 1, 2018 were selected as group A, 45 cases of NRDS preterm infants who received treatment during the preventive care program from January 1, 2017 to January 10, 2018 were included in group B for retrospective study. Baseline data, mechanical ventilation index, complications, and family satisfaction of the children were observed. Results The number of pulmonary surfactant applications, mechanical ventilation time, hospitalization time and hospitalization cost were (1.5±0.5) times, (92.5±13.8) h, (12.5±1.3) d, (26±4) thousand yuan in group A, and (2.6±0.4) times, (131.4±23.1) h, (16.0±2.8) d, (33± 5) thousand yuan in group B, the highest oxygen saturation value was 468.9±42.1 in group A,401.2±22.3 in group B, there were significant difference between the two groups( t=7.334-11.524, P<0.05). The total incidence of complications during treatment was 15.6% (7/45) in group A and 48.9% (22/45) , there was significant difference between the two groups (χ2=11.447, P<0.05). The total satisfaction rate of family members was 95.6%(43/45) in group A, which was significantly higher than that in group B (64.4%, 29/45). The difference was statistically significant (χ2=13.611, P<0.05). Conclusions Prophylactic care can be used in the mechanical ventilation of premature infants with NRDS, which can significantly reduce the incidence of complications, reduce the amount of pulmonary surfactant, shorten the mechanical ventilation time, and save the cost of treatment. It is worth promoting.
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OBJECTIVE@#To investigate the expressions of secreted frizzled-related protein 4 (SFRP4) in stage Ⅱ DNA mismatch repair-deficient (dMMR) and mismatch repair- proficient (pMMR) colorectal cancers and explore their clinical significance.@*METHODS@#We collected fresh stage Ⅱ colon cancer tissues with different MMR status detected by immunohistochemistry (IHC). The differentially expressed mRNAs between dMMR and pMMR tumors were identified by Affymetrix Human oeLncRNA gene chip, and the expression of SFRP4 in these cancer tissues and in colorectal cancer cell lines were detected using Western blotting and real- time quantitative PCR. The apoptosis rates of HCT116 cells with and without siRNA- mediated transient SFRP4 knockdown were determined using flow cytometry. We further investigated the expression pattern of Ki-67 and its correlation with SFRP4 expression.@*RESULTS@#Compared with pMMR colon cancer tissues or cells, both dMMR colon cancer tissues (=0.014) and cells (=0.0079) showed significantly increased expression of SFRP4, which was in negative correlation with Ki-67 (=0.041). In HCT116 cells, transient SFRP4 knockdown resulted in decreased cell apoptosis, including both early apoptosis (=0.003) and late apoptosis (=0.024).@*CONCLUSIONS@#Up-regulation of SFRP4 in dMMR stage Ⅱ colon cancer promotes apoptosis and inhibits proliferation of the cancer cells, and may improve the prognosis of dMMR colon cancer.
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Humanos , Apoptose , Proliferação de Células , Colo , Metabolismo , Patologia , Neoplasias do Colo , Genética , Metabolismo , Patologia , Neoplasias Colorretais , Genética , Metabolismo , Patologia , Reparo de Erro de Pareamento de DNA , Técnicas de Silenciamento de Genes , Células HCT116 , Antígeno Ki-67 , Metabolismo , Prognóstico , Proteínas Proto-Oncogênicas , Genética , Metabolismo , Regulação para CimaRESUMO
Objective To design a portable respirator that can facilitate both cardiopulmonary resuscitation and auxiliary oxygen inhalation.Methods Positive pressure design as well as pneumatically powered and controlled design were adopted for this respirator.Theoretical calculation of key components was carried out.The virtual prototype was established and the structure checked using the three dimensional design software Solidedge.Finally,the prototype was processed,assembled and tested.Results Experimental tests proved that the designed respirator was capable of cardiopulmonary resuscitation and auxiliary oxygen inhalation when oxygen supply pressure reached 3 -5.5 bar.It could achieve the expected technical indicators,such as the oxygen flow rate that was between 10 and 12 L/min.Conclusion The portable oxygen supply respirator has the advantages of a small size,light weight,simple operation,and automatic power supply.It has broad prospects of application.
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Objective To detect EPPB41L3 methylation frequency difference between esophageal squamous cell carcinoma (ESCC) tissues and the normal tissues and between ESCC patients′plasma and healthy volenteers′plasma, and to analyze the correlation with clinicopathological parameters. Methods We collected esophageal squamous cell carcinoma tissues (n = 42 patients) and adjacent surrounding normal tissues (n = 42 patients), and plasma from 42 patients with ESCC and from 50 healthy individuals. We used methylation specific PCR (MSP) combined with agarose gel electrophoresis to detect the methylation status of the EPB41L3. We used the SPSS 13.0 software for statistical analysis by χ2 test and Fisher′s exact test. Results EPB41L3 frequency of methylation was significantly higher in tumor tissues than in the adjacent tissues (59.5% vs. 4.8%), the difference was statistically significant (χ2 = 28.873, P < 0.001). For plasma, EPB41L3 methylation frequency was 31.0%in cancer patients, while was not detectable in the healthy volunteers. Methylation of EPB41L3 in tissues was more frequently found in patients with tumor size of ≥ 5 cm or T3 than in patients with tumor size of < 5 cm or T1-2. Conclusions The methylation frequency of EPB41L3 is higher in ESCC tissues than in control normal tissues, and higer in plasma from ESCC patients than that from the healthy volunteers. EPB41L3 methylation is more frequently found in patients with more advanced disease.
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Objective To explore the Kano model analysis in the application of midwife outpatient service quality management. Methods Kano questionnaire was consisted of 24 items, including five parts, environment facilities, service evaluation, service technology, the team service, delivery service′quality. Using the Kano model analysis, the impact factors of the midwife outpatient service quality in five different quality attributes were determined. It included property and had the opposite answer to the question, charisma, essential properties and one property. Results By identifing the quality index classification of the pregnant women with satisfactory service quality to midwife outpatient service, attributable to a property were 1,2,3,4,8,9,10,14,20,21;attributable to mandatory attributes were 13, belonging to the charm of the property had 5,6,7,11,12,15,16,17,18,19,22,23,24. Midwives outpatient had most of charisma attribute, 54.2%(13/24) of the total;The second was one attribute, 41.7% (10/24) of the total. The technology level of midwife was essential attribute. Conclusions Kano mode analysis technology will be introduced to the midwife outpatient service quality management to provide decision reference for improving the quality of the hospital management level, improve maternal satisfaction, improve the obstetric operation efficiency.
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Objective To investigate the clinical features,treatment,medication and abnormal metabolism control status in elderly benign prostatic hyperplasia (BPH) with metabolic syndrome (MS).Methods The 397 patients with BPH aged ＞60 years (294 with MS) were retrospectively analyzed.The clinical features,treatment and control status were analyzed.Results Body mass index,waist circumference (BMI),2 hPBG,systolic pressure,diastolic pressure,TG and uric acid,urinary albumin/creatinine ratio,urinary albumin were higher and HDL-C was lower in BPH with MS than without MS (P＜0.05).Prostate volume,PSA level,IPSS score,quality of life score were higher in BPH patients with MS than without MS (P＜0.05).Treatment modality analysis showed that lower urinary tract symptoms was mild,watchful waiting accounted for 22.3％,urinary retention or obstruction related operation treatment for 8.7％,moderate to severe symptoms related oral drugs treatments for 67％,the oral drug treatment was still the main mode of BPH.In the patients with abnormal levels of glucose metabolism,the target arrival rate (TAR) on FBG,2 hPBG and HbAlc were 49.6％,34.9％ and 37.5％,TAR on all the three was 19.1％.In the hypertensive population,TAR on systolic and diastolic pressure were 44.1％ and 82.5％,TAR on both two was 41.7％.TG,TC,LDL-C and HDL-C were 31.3％,35.4％,48.2％ and 64.9％,on all the four was 29.9％.Conclusions The ratio of elderly BPH with MS is high,but the target arrival rate on control of urinary tract symptoms and abnormal metabolism component is not optimistic in spite of a positive drug administration.
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Primary cell culture, techniques of gene transfection, gelatin zymography, and Western blot were used to investigate the effect of hypoxia on the secretion of MMP-2 and MMP-9 in pulmonary artery endothelial cells (PAEC) and smooth muscle cells (PASMC), and the role of HIF-1. Our results showed that (1) after exposure to hypoxia for 24 h, the protein content and activity of MMP-2 in the PAEC medium as well as these of MMP-2 and MMP-9 in PASMC medium (P<0.01) decreased significantly in contrast to those in normoxic group (P<0.05); (2) after transfection of wild type EPO3'-enhancer, a HIF-1 decoy, the content and activity of MMP-2 and MMP-9 in hypoxic mediums became higher than those in normoxic group (P<0.01), while transfection of mutant EPO3'-enhancer didn't affect the hypoxia-induced down-regulation. It is concluded that hypoxia could inhibit the secretion and activity of MMP-2 and MMP-9 in PAEC and PASMC, which could be mitigated by the transfection of EPO3'-enhancer and that HIF-1 pathway might contribute to hypoxia-induced down-regulation of MMP-2 and MMP-9.
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BACKGROUND: Ligustrazine inhibits discharge of cerebral hippocampal neuron, penetrates blood-brain barrier effectively after absorbed in the body and is distributed extensively in cerebral cortex, brain stem, striate body, hippocampus, cerebellum and midbrain.OBJECTIVE: To probe into the influence of ligustrazine and its different concentrations after abdominal injection on cerebral cortical neural cell structure in epileptic rats induced by penicillin.DESIGN: Randomized control experiment.SETTING: Physiological Department of Xianning Medical College.MATERIALS: The experiment was performed in Anatomy Department of Tongji Medical College of Huazhong University of Science and Technology from September 2004 to March 2005. Forty healthy SD rats of clean grade were employed, of either sex, mass weighted varied from 200 to 250 g.They were randomized into 5 groups, named operation control, penicillininduced epilepsy group and ligustrazine groups of 10 mg/kg, 20 mg/kg and 40 mg/kg, 8 rats in each group.METHODS: After anesthetized, the cranium was opened to expose cerebral cortical record region. BL-410 biofunctional experimental system was used to record brain electricity bilaterally and epileptic discharge of cerebral cortex in penicillin-induced epilepsy group and ligustrazine groups of 10 mg/kg,20 mg/kg and 40 mg/kg. In the control, 1 hour after anesthesia and craniotomy, cerebrum was collected. In penicillin-induced epilepsy group, 1 hour after induction, cerebrum was collected. In ligustrazine groups of 10 mg/kg,20 mg/kg and 40 mg/kg, after penicillin-induced epileptic discharge was stable, ligustrazine of 10 mg/kg, 20 mg/kg and 40 mg/kg was injected abdominally successively, and cerebrum was collected when the most remarkable inhibition was achieved. Brain tissue section was prepared separately, with HE staining, the observation was done under optic microscope.MAIN OUTCOME MEASURES: Structure changes in cerebral cortical neural cells in rats of each group.In the control, the morphological structure of cerebral cortical neural cell alternations on cerebral cortical neural cell structure, karyopykosis, plasmarrhexis and vacuolar structure, but there was no Nissel bodies in cytomarrhexis, vacuolar structure and decreased Nissel bodies in cytoplasm with the control, there were decreased vacuoles in neural cell, increased cytoplasm and few Nissel bodies in cytoplasm and cell structural morpholcontrol, karyon was big, round and light stained; clot-like Nissel bodies were visible and cell structural morphology was in tendency to be normal.CONCLUSION: In penicillin-induced epilepsy, morphological structure of cerebral cortical neural cell in rats is abnormal. Tetramethylpyrazine of various dosages may improve at different degrees morphological structure of cerebral cortical neural cell, especially significantly at high dosage, by which, its inhibition on epileptic discharge in rats is achieved.
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<p><b>OBJECTIVE</b>To investigate the effects of anti-HPV16E6-ribozyme (HRz) on phenotype and gene expression of a cervical cancer cell line.</p><p><b>METHODS</b>HRz was designed by computer programs. HRz's activity was identified by cleavage experiments in vitro. HRz and empty eukaryotic plasmids were transfected into CaSKi cells with lipofectin, then renamed CaSKi-R and CaSKi-P, respectively. The expression of ribozyme in transfected cells was observed by RNA dot blot. The amounts of E6 mRNA in three kinds of cells lines were detected by Northern blot. Cell growth curves and soft agar forming ability were studied. The ability of each cell line to form tumors was assessed in nude mice. Apoptosis rates and expression of c-myc, bcl-2, p53 and Fas were detected by flow cytometry (FCM). Antigens of tumor cells, HLA-1, HLA-2, B7-1 and B7-2 were also detected. NK, LAK, and CD(3)AK cells were induced. Their cytotoxicities were detected in CaSKi-R, CaSKi-P, and CaSKi cells.</p><p><b>RESULTS</b>In vitro cleavage reaction demonstrated that HRz could cleave HPV16E6 mRNA in a site-specific manner. HRz could be expressed stably in transfected CaSKi cells. Northern blot analysis showed that E6 mRNA levels were lower in CaSKi-R than in CaSKi. The growth rate of CaSKi-R was slower than those of CaSKi and CaSKi-P. The soft agar-forming rate of CaSKi-R was lower compared with those of CaSKi and CaSKi-P cells. The ability of CaSKi-R to form tumors in nude mice was also poor. The apoptosis rate of CaSKi-R cells was much higher than those of CaSKi and CaSKi-P. HRz could reduce the expression of E6, c-myc and bcl-2 proteins, and increase the expression of p53 as well. HRz could increase the expression of HLA-2, B7-1 and B7-2 antigens. The cytotoxicity of NK, LAK and CD(3)AK cells was much higher in CaSKi-R than in CaSKi-P and CaSKi cells.</p><p><b>CONCLUSION</b>HRz not only reverses the malignant phenotype of CaSKi cells partially, but also induces apoptosis in the cells, and increases sensitivity of CaSKi cells to immune cells.</p>
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Animais , Feminino , Humanos , Camundongos , Apoptose , Divisão Celular , Citotoxicidade Imunológica , Citometria de Fluxo , Expressão Gênica , Terapia Genética , Células Matadoras Naturais , Alergia e Imunologia , Camundongos Nus , Proteínas Oncogênicas Virais , Genética , Fenótipo , RNA Catalítico , Usos Terapêuticos , RNA Mensageiro , Proteínas Repressoras , Células Tumorais Cultivadas , Neoplasias do Colo do Útero , Genética , Patologia , TerapêuticaRESUMO
Purpose:During the malignant transformation, most tuomr cells will become resistant to anoikis (the apoptosis induction upon loss of anchorage), thereby gaining an important growth advantage and providing the basis for tumor spread, growth at distant places, and formation of metastasis.The present study focus on the role of soluble fibronectin, integrin and their signal transduction in the process of tumor cells anoikis apoptosis.Methods:Flow cytometry detected integrin subunits' expression on the human squamous cell carcinoma cell (HSC 3) surface. Establish anoikis induction model. The role of different integrin subunits during anoikis induction were detected by antibody blocking method, p53 and Bcl 2/Bax expression were measured during this anoikis process.Results:There is no or low expression of integrin av on the normal epithelial cells, whilst it expresses relative higher on the surface of HSC 3 cells. The treatment of soluble fibronectin decreased p53 expression, increased the ratio of Bcl 2/Bax and then protected HSC 3 cell from anoikis. Anti integrin ?v blocking antibody resumed the anoikis that was inhibited by soluble fibronectin evidently. Conclusions:We now provide the evidence that the binding of integrin ?v and soluble fibronectin protected HSC 3 cells from anoikis apoptosis, via decreasing the expression of p53, increase the ratio of Bcl 2/Bax.
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Objective: To investigate the effects of HPV16E6-ribozyme on telomerase activity in cervical carcinoma cell line CaSKi and the related mechanisms. Methods: Anti-HPV16E6-ribozyme and blank eucaryotic plasmids were transfected into CaSKi cells via lipofectin, and the resultant cells were named as CaSKi-R and CaSKi-P, respectively. The expression of ribozyme in transfected cells was observed by RNA dot blotting. The expression of E6 mRNA and protein in the 3 kinds of cells were detected by Northern blotting and Western blotting, respectively. Telomerase activity was determined by TRAP-Elisa method; the expression of P53, c-myc, hTERT and hRT mRNA were examined by RT-PCR.Results: RNA dot blotting showed that anti-HPV16E6-ribozyme was stably expressed in transfected CaSKi-R cells. Western blotting showed that the expression of E6 mRNA and protein in CaSKi-R cells was obviously lower than that in CaSKi and CaSKi-P cells. The telomerase activities in CaSKi,CaSKi-P and CaSKi-R cells were (0.89?0.14), (0.90?0.11) and(0.36?0.06),respectively. The inhibitory rate of telomerase activity in CaSKi-R cells was 59.55%, which was significantly lower than those in CaSKi and CaSKi-P cells (P
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Objective: To investigate the characteristics of the cultured cervical cancer cell line transfected with anti HPV16E6 ribozyme, and to investigate the possibility and practicality of ribozyme in treatment of cervical cancer. Methods: The anti HPV16E6 ribozyme and empty eucaryotic expressing plasmids were transfected by lipofectin transfection into CaSKi cell, which named as CaSKi R, CaSKi P respectively. The morphology and the soft agar forming ability were studied. The expression of E6, PCNA and C erbB 2 genes was studied through Flow Cytometry. The tumorgenicity of each cell was detected by injecting cells into the nude mice skin. Three groups of nude mice were injected by CaSKi, CaSKi R and CaSKi P cell separately. Another group of mice was injected by CaSKi cell on right side and CaSKi R cell on left side. Results: There is no distinct difference of the morphology and growth rate between CaSKi and CaSKi P, but the growth rate of CaSKi R decreased. The soft agar forming rate of CaSKi P was similar with that of CaSKi cells, while that of CaSKi R was found decreased. The result of flow cytometric analysis showed that anti HPV16E6 ribozyme could reduce the expression of E6, PCNA and C erbB 2 genes on CaSKi R cells, while this phenomenon was not found on the CaSKi P cells. The tumorgenicity of CaSKi R in nude mice was decreased compared with CaSKi and CaSKi P cells. Conclusion: Anti HPVE6 rivozyme could partly reverse the malignant phenotypes of CaSKi cells. The reason may be the decrease of E6 gene expression, and the succeeding decrease of the PCNA and C erbB 2 genes′ expression.
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Objective: To study the characterization of the cultured cervical cancer cell line transfected with anti- HPV16E6-ribozyme, and to investigate the effect of ribozyme on proliferation and apoptosis of cervical cancer cell. Metliods: Anti-HPV16E6-ribozyme had been designed to cleave the HPV16E6 gene. With the method of lipofectin transfec- tion, the anti-HPVI6E6-ribozyme and empty eucaryotic expressing plasmids were transfected into CaSKi cell, which named as CaSKi-R, CaSKi-P respectively. The amounts of E6 mRNA in the three kinds of cells were detected by northern blot. Cell cycle was detemined by flow cytometry, and cell apoptosis was examined by fluorescent (Hoechst) staining and TUNEL. The expression of some genes, including c-myc, bcl-2, p53, and fas, was also detected by flow cytometry analy- sis. Results: Northern blot showed that E6 mRNA was less in CaSKi-R than in CaSKi. In CaSKi-R cells, cycle was arres- ted in G1 phase, with decreasing in percentage of S phase cells. The apoptosis rate of CaSK1-R cell was much higher than those of CaSKi and CaSK1-P. Anti-HPV16E6-ribozyme could reduce the expression of E6, c-myc, bcl-2 genes on CaSKi- R cells, and increased the expression of p53. While this phenomenon was not found on the CaSK1-P cells. The expression of fas was similar in the three kinds of cells. Conclusion: Anti-HPVE6-rivozyme induces apoptosis of human cervical cancer CaSKi cells. The mechanisms may be the decrease of E6 gene's expression, and the succedent changing of some genes'expression.