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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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Objective:To study the effect of cyclin Y (CCNY) on the lesion degree of patients with liver cancer and its correlation with preoperative and postoperative liver injury.Methods:The clinical data of 60 patients with liver cancer (liver cancer group) and 69 patients with liver cirrhosis (liver cirrhosis group) in Hangzhou Hospital of Zhejiang Medical and Health Group from January 2015 to October 2017 were retrospectively analyzed. In liver cirrhosis group, Child-Pugh liver function grade A was in 33 cases (liver cirrhosis grade A group), grade B was in 21 cases (liver cirrhosis grade B group), grade C was in 15 cases (liver cirrhosis grade C group). The serum total bilirubin (TBIL), alanine aminotransferase (ALT), albumin, cholinesterase, γ-glutamyl transpeptidase (GGT) and total bile acid were detected by automatic biochemical analyzer. The serum CCNY was detected by WB method, and compared with 40 healthy subjects (healthy control group).Results:Compared with those in healthy control group, the albumin and cholinesterase in liver cirrhosis grade A, B and C groups were significantly decreased, the ALT, TBIL, GGT, total bile acid and CCNY were significantly increased, , and the changes were more obvious with the severity of liver disease, and there were statistical differences ( P<0.05). Pearson correlation analysis result showed that the CCNY was positive correlation with TBIL, ALT, total bile acid and GGT in patients with liver cancer ( r = 0.544, 0.612, 0.553 and 0.539; P<0.05), and CCNY was negative correlation with albumin and cholinesterase ( r = - 0.478 and - 0.620, P<0.05). In patients with liver cancer, before operation and 1, 2 and 7 d after operation, the CCNY was 3.01 ± 1.10, 7.24 ± 2.57, 6.29 ± 1.78 and 4.01 ± 1.52, ALT was (98.74 ± 16.79), (430.55 ± 197.62), (255.73 ± 26.77) and (121.89 ± 20.42) U/L, respectively; the CCNY and ALT 1 and 2 d after operation were significantly higher than those before operation, those 2 d after operation were significantly lower than those 1 d after operation, those 7 d after operation were significantly lower than those 2 d after operation, and there were statistical differences ( P<0.05); there were no statistical difference between 7 d after operation and before operation ( P>0.05). The expression of CCNY before operation and 1 d, 2 d, 7 d after operation was positive correlation with ALT in patients with liver cancer ( r = 0.669, 0.821, 0.663 and 0.642; P<0.01). Conclusions:The more severe the degree of liver lesions in patients with liver cancer, the higher the serum CCNY, and the higher the expression of CCNY, the more severe the degree of liver injury in patients with liver cancer due to surgery, which is positively correlated with liver injury indexes.
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@#Protein kinases (PKs) are regulators of protein phosphorylation in many infectious diseases, including malaria. However, the cellular functions of majority of PKs in Plasmodium falciparum remain unknown. The mechanisms involved in P. falciparum cell cycle progress are not fully understood. The activation of cyclin-dependent kinases (CDKs), which constitute a PK family that includes crucial regulators of cell cycle progression in eukaryotes, is strictly being coordinated by the interaction with specific cyclins at well-defined points within the cell cycle. These cyclin/CDK complexes are very well characterised in humans, but little is known in P. falciparum. This review expand our understanding of the characteristic of CDKs and cyclins in P. falciparum, and paves the way for further investigations on the precise molecular role of these crucial regulatory proteins in mosquito and human. This represents a valuable step towards the elucidation of cell cycle control mechanisms in malaria parasites.
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AIM:To investigate the role of Bcl-2-associated athanogene 2(BAG2)in the proliferation of human lung adenocarcinoma A549 cells and its clinical implications.METHODS:The abundance of BAG2 protein in A549 and lung bronchial epithelium(HBE)cells were measured by Western blot.After down-regulation of BAG2 by transfection of siRNA in A549 cells, the expression of cell proliferation and cell cycle related proteins were detected by CFSE assay, WST-1 assay and Western blot,respectively.Moreover,the expression of BAG2 in cDNA array which contained 10 pairs of lung cancer and adjacent tissue was verified.Meanwhile, BAG2 expression in GEO database, which included the human lung cancer and adjacent tissue microarray data was analyzed.The prognosis power of BAG2 was evaluated by the Kaplan-Meier survival curve analysis.RESULTS:BAG2 had remarkably higher expression level in A 549 cells than that in HBE cells.Knockdown of BAG2 resulted in significantly inhibition of proliferation in A 549 cells,accompany with the significant-ly down-regulation of cyclin B1 and cyclin E1.BAG2 was over-expressed in the lung cancer tissues,as compared with the adjacent normal tissues.Kaplan-Meier plotter and cDNA microarray results showed that patients with higher BAG 2 expres-sion were significantly associated with poorer survival.CONCLUSION:The BAG2 gene tends to regulate A549 cells pro-liferation via cyclin B1 and cyclin E1.BAG2 has significantly prognostic power on the survival of lung cancer patients.
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Objective To evaluate the effect of berberine on the expression of cell cycle-related miRNA and its target gene in human melanoma A375 cells.Methods In vitro cultured A375 cells were classified into several groups to be treated with berberine at different concentrations of 0 (blank control group),20,40,60,80,and 100 μmol/L,respectively,for 48 hours.qRT-PCR was performed to determine the mRNA expression of miRNA-582-5p and its target gene cyclin-dependent kinase 1 (CDK1),miRNA-188-5p and its target gene CDK2,Cyclin D1 and Cyclin A.Western blot analysis was conducted to measure the protein expression of cell cycle-related proteins CDK1,CDK2,Cyclin D1 and Cyclin A.Results qRT-PCR showed that compared with the blank control group,the 20 and 40 μmol/L berberine groups had a similar expression of miRNA-582-5p (both P ＞ 0.05),but the 60 and 80 μmol/L berberine groups had a significantly up-regulated expression of miRNA-582-5p (both P ＜ 0.05).Compared with the blank control group,all the 5 berberine groups had a significantly increased expression of miRNA-188-5p (F =22.600,P =0.002).However,the mRNA expression of CDK2,CDK1,Cyclin A and Cyclin D1 gradually decreased along with the increase of berberine concentrations (F =51.976,248.510,626.671 and 312.740,respectively,all P ＜ 0.001).Western blot analysis revealed that berberine decreased the protein expression of CDK1,CDK2,Cyclin D1 and Cyclin A (F =138.124,110.966,278.772 and 140.167,all P ＜ 0.001).Conclusion Berberine can decrease the expression of cell cycle-related proteins CDK1,CDK2,Cyclin D1 and Cyclin A,likely by decreasing their mRNA expression and increasing the expression of miRNA-582-5p and miRNA-188-5p,and then block the cell cycle progression of A375 cells and inhibit the growth of tumor.
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Objective To investigate the optimal time and molecular mechanism of gene therapy for promoting distraction osteogenesis by observing the effect of gene transfecting at different time on expression of cyclins in mandibular distraction area.Methods Forty eight New-Zealand rabbits were employed.After accomplishing bilateral mandibular distraction osteogenesis model,the rabbits were randomly divided into the group A,B,C and D.The group A,B and C were transfected by recombinant plasmids pIRES-hBMP2-hVEGF165 2 μg(0.1 μg/μL)at the distraction area instantly after operation,on postoperative 4,14 d and given local electroporation stimulation.The four groups entered the consolidation stage after 10 d continuous traction at a rate of 1 mm/d on postoperative 4 d.Three rabbits in each group were respectively sacrificed on 7,14,28,56 d of the consolidation stage.The expression of cyclin A,D1,and E of fresh bone tissue in distraction area were detected by immunohistochemical staining.Results Cyclin A,D1,and E were strongly expressed in the traction gap on 7,14 d of consolidation stage found,which was strongest on 7 d,moreover the group A,B and C were stronger than the group D.The expression in each group was weakened on 28,56 d.The expression on 7 d in the group B was stronger than that in the4 group A,C and D(P0.05),but all were stronger than the group D(P0.05).Conclusion The high expression of cyclin A,D1 and E can promote the cellular division,proliferation and differentiation in distraction area,thus accelerates the new bone formation in the distraction area,prompting that the distraction period is the best time for distraction osteogenesis under gene therapeutic intervention.
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Objective To investigate the potential mechanism that melatonin at higher concentrations inhibits the proliferation of human MG-63 osteosarcoma cells,so as to provide a certain experimental basis for the better application of melatonin in the treatment of diseases in Department of Orthopedics. Methods MG-63 cells cultured in vitro were treated with melatonin at a concentration of 4 mmol/L . Western blotting and real-time PCR method were used to analyze the effect of melatonin on the expression of cyclins and CDKs at protein and mRNA levels ,respectively. Results Western blotting and real-time PCR analyses showed that melatonin's inhibitory effect was possibly through the downregulation of cyclin D1 and CDK4 that related to the G1 phase,and downregulation of cyclin B1 and CDK1 that related to the G2/M phase. However,there was no obvious dif-ference of cyclin E,CDK2,and cyclin A,which were related to G1/S transition and S phase. Conclusion Melatonin may significantly inhibit hu-man osteosarcoma cell proliferation by inducing cell cycle arrest in a time-dependent manner,which is related to the downregulation of cyclin D1, CDK4,cyclin B1 and CDK1.
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Background As the main cellular constituent of corneal epithelium,corneal epithelial cells play critical roles in regulating and controlling the migration, proliferation and differentiation of cells during the repair of damage.MicroRNA (miRNA) is endogenously expressed small non-coding RNAs, which participates in a variety of biological processes.Previous studies demonstrated that miR-204 is highly expressed in normal corneal epithelium,but its function is still unclear.Objective This study was to investigate the function and mechanism of miR-204 in corneal epithelial cell proliferation.Methods Corneal epithelial tissue was collected during the corneal refractive surgery from the patients with refractive error under the informed consent, and human corneal epithelial cells(HCECs) were cultured and passaged.The relative expressing levels of miR-204 mRNA in the normal corneal epithelium and HCECs were detected by real time quantitative PCR.Cultured HCECs were evenly divided into three groups.The liposome with miR-204 mimic was transfected into the cells of the miR-204 mimic group, and blank liposome was transfected in the cells of the positive control group,and regularly cultured cells served as the normal control group.Cell proliferation capability was evaluated by colony-forming assay, and the percentage of the cells in different cell cycles was analyzed by flow cytometry.Western blot assay was employed to detect the expression levels of p-RB, E2F1 ,p27,CyclinA, CDC2, p-CDC2 and CDK2 proteins in the cells.Results The relative expression levels of miR-204 mRNA were 1.077 ±0.268 in the normal corneal epithelium and 0.041 ±0.018 in the HCECs, showing a significant difference between them (t =7.700,P＜0.001).The cloning cell number was evidently decreased in the miR-204 mimic group in comparison with the positive control group and normal control group.The percentage of cells in the G1 phase was 47.75％ in the miR-204 mimic group, which was significantly higher than 37.23％ in the positive control group and 40.72％ in the normal control group.The expression levels of E2F1 and p27 proteins in the cells were elevated (t=14.87,25.11;both at P＜0.01) and those of CDK2 and p-CDC2 proteins were decreased (t=5.39,10.65;both at P＜0.01) in the miR-204 mimic group in comparison with the positive control group.Conclusions The overexpression of miR-204 in the normal corneas probably is associated with non-proliferation status of corneal epithelial cells.Transfection of miR-204 into corneal epithelial cells can inhibit the proliferation of corneal epithelial cells probably by up-regulating the expression of E2F1 and p27 and suppressing the expression of CDK2/CyclinA and p-CDC2/CyclinA,which lead to cell arrest in G1 phase.
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BACKGROUND:Cdh1 has been shown to express in rat hippocampus and cortex in a large number. Moreover, in vitro test demonstrated that Cdh1 expression was higher in neurons than in neural stem cel s, which possibly associated with the differentiation of neural stem cel s into neurons. However, the effects of anaphase promoting complex Cdh1 on ischemic neuronal damage remain unclear. OBJECTIVE:To investigate the expression of Cdh1 and its downstream substrate in primary cultured neurons with oxygen-glucose deprivation. METHODS:Primary neurons from cortex of postnatal 24-hour rat pups were cultured in vitro, and identified by immunofluorescence staining. The oxygen-and glucose-deprived models were established by three gas incubator fil ed with nitrogen in sugar-free Earle’s solution. After 1 hour of hypoxia, reoxygenation was conducted. Real-time fluorescent quantitative PCR was used to detect the mRNA expression of Cdh1 and its downstream substrates Skp2, Cyclin B1 before hypoxia, 6 hours, 1, 3, 7 days after oxygen glucose deprivation. RESULTS AND CONCLUSION:After oxygen glucose deprivation, the expression of Cdh1 and Cyclin B1 in primary neurons was increased (P<0.05), while Skp2 expression was decreased (P<0.05). Above data indicated that Cdh1 expression in neurons increased after oxygen-glucose deprivation. It may degrade Skp2 and participate in hypoxic neuronal apoptosis by ubiquitination.
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Objective To construct the luciferase reporter gene vector of cell division cycle 2 (Cdc2) gene promoter and determine its transcriptional activity. Methods Primers were designed based on human Cdc2 promoter sequence from UCSC software. Then Cdc2 promoter from human genome DNA was replicated. After pGL3-Basic vector and Cdc2 promoter were digested with restriction enzymes SacⅠand XhoⅠseparately, Cdc2 promoter was inserted into pGL3-Basic vector. The recombinant plasmid named pGL3-Cdc2-promoter was transiently co-transfected into U2OS cells with control vector pRL-SV40, and then the activity of dual luciferase was detected. Results pGL3-Cdc2-promoter was constructed successfully. The restriction analysis and sequencing proved the entirely correct sequencing results. The luciferase activity was higher in pGL3-Cdc2-promoter/pRL-SV40 group than that of pGL3-Basic/pRL-SV40 group (1.591 5±0.199 8 vs 0.049 9±0.010 4). Conclusion pGL3-Cdc2-promoter can be transcribed and activated in U2OS cells. This study provided an important basis for screening and evaluation of anticancer drugs.
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BACKGROUND:Cyclin A2 is a key regulator of cellcycle. Location and expression of cyclin A2 in neonatal mouse myocardium is not clear. OBJECTIVE:To observe the location and expression of cyclin A2 in neonatal mouse cardiomyocytes and its relationship with the exit of cardiomyocytes from cellcycle. METHODS:Neonatal mice were kil ed to take myocardial tissues at 0, 3, 7, 14 and 28 days after birth. Western blot were used to detect the expression of cyclin A2, proliferating cellnucleus antigen and Phospho-histone H3. Immunohistochemitry detection was used to detect the location of cyclin A2 and expression of proliferation cellnucleus antigen at different time after birth. RESULTS AND CONCLUSION:Western blot showed the decrease of cyclin A2 after birth til disappeared at day 4 (P=0.001). Cyclin A2 located mainly in the nucleus after birth and exported to the cytoplasm at day 14, and basical y disappeared at day 28. Proliferating cellnucleus antigen showed gradual y decreased tendency after birth. Mitosis specific marker, Phospho-histone H3, exhibited a gradual decrease after birth, which was consistent with cyclin A2 in expression intensity.
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BACKGROUND:Proliferation, migration and phenotypic changes of vascular smooth muscle cells is the core of the occurrence of atherosclerosis, and a series of related genes via methylation are involved in the process. OBJECTIVE:To investigate the effects of oxidized low density lipoprotein (ox-LDL) on DNA methylation in the promoter region of the p21 gene and its potential mechanism in the pathogenesis of atherosclerosis. METHODCultured human vascular smooth muscle cells were treated with different concentrations of ox-LDL (0, 10, 20, 40 mg/L) for 24 hours. The degree of DNA methylation was assayed by methylation-specific polymerase chain reaction, the expression of p21 mRNA was measured by reverse transcriptional polymerase chain reaction and the proliferative activity of vascular smooth muscle cells was determined by the MTT assay. RESULTS AND CONCLUSION:The ox-LDL treatment resulted in a promotion in the methylation in the promoter region of the p21 gene and a decrease in mRNA expression with a concentration-dependent manner;it also induced a dose-dependent promoting effect on vascular smooth muscle cellproliferation. The atherogenic mechanism of ox-LDL might promote vascular smooth muscle cellproliferation by the hypermethylation of the p21 gene that may lead to the occurrence and development of atherosclerosis.
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Objective: To investigate the effect of a small interfering RNA (si-RNA) targeting signal transducer and activator of transcription-3 (STAT3) on the biological function of human endometrial cancer cell line HEC-1B. Methods: The HEC-1B cells were transfected with chemically synthesized si-RNA targeting STAT3 (si-STAT3). Then the expression of STAT3 mRNA was detected by real-time fluorescence quantitative-PCR, the proliferation ability was determined by cell counting kit-8 (CCK-8) assay, the cell cycle distribution and apoptosis were measured by flow cytometry, the migration and invasion were examined by Transwell chamber, and the expression levels of STAT3, phospho-STAT3 (p-STAT3), and cyclin D2 (CCND2) and metalloproteinase-2 (MMP-2) were detected by Western blotting. Results: As compared with the blank control group (only liposome was added) or transfection with si-control group, the expression level of STAT3 mRNA in HEC-1B cells after transfection with si-STAT3 was down-regulated (P < 0.01), the proliferation ability was inhibited (P < 0.01), the percentage of the cells in G1-phase was increased and which in S1-phase was decreased (P < 0.01). The abilities of migration and invasion of HEC-1B cells after transfection with si-STAT3 were inhibited (P < 0.01), and the expression levels of STAT3, p-STAT3, CCND2 and MMP-2 proteins were down-regulated (P < 0.01). Conclusion: Si-RNA targeting STAT3 gene can inhibit the expressions of STAT3 mRNA and protein in HEC-1B cells, and also inhibit the cell proliferation, the abilities of migration and invasion, and apoptosis. Copyright © 2013 by TUMOR.
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Cyclin E is expressed starting from the middle G1 phase of the cell cycle,and is accumulated in the G1/S boundry.Cyclin E binds to and activates the cyclin-dependent kinase CDK2.Cyclin E-CDK2 complex initiates a cascade of events that leads to DNA replication by phosphorylating its substrates,such as Rb,CDC6,NPAT and P107,etc.Additionally,cyclin E plays an important role in the regulation of genomic stability,spindle-organizing structure and centrosome cycle.Cyclin E expression is trans-activated by members of the transcription factor E2F family and degrades via the ubiquitin-proteasome pathway.At the same time,it is also negatively regulated by the CIP/KIP proteins.Cyclin E highly expressed in the initiation and progression of different human cancers,such as breast cancers,lung cancers,leukemia,lymphomas and others.
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ObjectiveTo evaluate the relationship of human papillomavirus(HPV) intection with expressions of cell cycle regulatory proteins cyclin D1,E and their dependent kinase inhibitor p27 in bowenoid papulosis(BP).MethodsTissue specimens were obtained from the lesions of 44 patients with BP,and circumcised foreskin tissue from 10 males served as the control.Gene chip was used to determine the genotypes of HPV,and immunohistochemistry to quantify the expressions of cyclin D1,E,p27,in these specimens.Results Of the 44 BP specimens,all were positive for HPV DNA,38(86.36％) for high risk types of HPV,and 6 for low risk types of HPV.Of the high risk HPV-positive specimens,30(68.18％) harbored HPV16,16 harbored single HPV 16,14 harbored other types of HPV besides HPV 16,8 harbored other high risk types of HPV.HPV 6 predominated in low risk HPV-positive specimens.The expression of cyclin D1 was significantly higher in patients with high-risk HPV(u =53.00,P＜ 0.05),with both high and low risk HPV(u =5.00,P＜ 0.01) and with low risk HPV (u =22.50,P＜ 0.05) than in normal human controls,and higher in patients with both high and low risk HPV (u =44.00,P＜ 0.01) and with low risk HPV (u =22.50,P＜ 0.05) than those with high risk HPV.In the case of cyclin E expression,patients with high risk HPV (u =0.00,P ＜ 0.01 ),with both high and low risk HPV (u =4.00,P ＜ 0.01 ),and with low risk HPV(u =1.50,P ＜ 0.01 ) were higher than normal human controls,and patients with high risk HPV were higher than those with low risk HPV(u =11.00,P ＜ 0.01).No significant difference was observed in the expression of p27 between patients with high and low risk types of HPV.A significant increase was observed in the expression of p27 in patients aged ＞ 50 years compared with patients aged 20-30 years(u =47.00,P＜ 0.05) and aged 31-50 years (u =55.50,P＜ 0.05),as well as in the expression of cyclin E in patients aged ＞ 50 years compared with those aged 20-30 years(u =45.50,P ＜ 0.05),and in female patients compared with male patients (u =137.50,P＜ 0.05).ConlusionThere is a significant difference in the expression of cyclin D1,E and p27 among patients with BP infected with different types of HPV.
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Objective: To investigate the effect of 5-Aza-dC (5-aza-2'-deoxycytidine) on cell cycle of multidrug resistant cells and its mechanism. Methods: Inhibitory effects of 5-Aza-dC on proliferation of breast cancer MCF-7/ADR cells and oral epidermoid carcinoma KBV/200 cells were detected by MTT method. The changes of cell cycle distribution were examined by flow cytometry when the cells were treated with 5-Aza-dC at different concentrations for 72 h or treated with the same concentration of 5-Aza-dC (10 μmol/L) for different time. The expression levels of cyclin A, cyclin E and p21waf1/cip1 mRNAs and proteins were detected by real-time fluorogenic quantitative-PCR and Western blotting, respectively. Results: The proliferation of MCF-7/ADR and KBV/200 cells was suppressed significantly after 5-AzadC treatment. The cell cycle was arrested at G2/M phase in dose- and time-dependent manners. The expression levels of cyclin A, cyclin E and p21waf1/cip1 mRNAs and proteins were elevated. Conclusion: The cell cycle of MCF-7/ADR and KBV/200 cells can be arrested at G2/M phase which consequently results in the inhibition of proliferation of multidrug resistant cells induced by 5-Aza-dC. The molecular mechanisms may be related with up-regulation of expressions of cyclin A, cyclin E and p21waf1/cip1 mRNAs and proteins. Copyright © 2012 by TUMOR.
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PURPOSE: Cyclins, and their associated cyclin dependent kinases, regulate progression of the cell cycle through the G1 phase and into the S-phase during the DNA replication process. Cyclin E regulation is an important event in cell proliferation. Despite its importance, abnormalities of these genes and their protein products have yet to be found in lits asoociation with lung cancer. MATERIALS AND METHODS: The relationships between expression of cyclin A, cyclin B1, cyclin D1, cyclin D3, and cyclin E and clinicopathologic factors were investigated in 103 cases with non-small cell carcinomas, using immunohistochemical analysis. RESULTS: The positive immunoreactivity was observed in 51 cases (50%) for cyclin A, 33 cases (32%) for cyclin B1, 83 cases (81%) for cyclin D1, 19 cases (18%) for cyclin D3, and 11 cases (11%) for cyclin E. Expression of cyclin E was significant for lymph node metastasis (p=0.004, Chi-square test). There was no relationship between cyclin A, B1, D1, and E and histological typing, tumor size, lymph node metastasis, or pathological tumor, node and metastasis staging. CONCLUSION: These findings suggest that the expression of cyclin E played a role, to some degree, in the lymph node metastasis.
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Adenocarcinoma , Carcinoma, Squamous Cell , Cell Cycle , Cell Proliferation , Cyclin A , Cyclin B1 , Cyclin D1 , Cyclin D3 , Cyclin E , Cyclin-Dependent Kinases , Cyclins , DNA Replication , G1 Phase , Lung , Lung Neoplasms , Lymph Nodes , Neoplasm MetastasisABSTRACT
Objective To observe the effect of intermedin (IMD) preconditioning on cyclin D1, cyclin E and CDKs expression, and explore its role in.promoting kidney tissue regeneration after renal ischemiareperfusion injury. Methods One hundred and forty-four healthy male Wistar rats were randomly divided into four groups: sham operation (S) group, ischemia-reperfusion injury (IR) group, empty plasmid (EP) group and IMD group. In the IR group, after the right kidney was excised, the aorta abdominalis and left renal artery were bluntly dissected in EP and IMD group, empty plasmid and IMD plasmid were transfected into the left kidney using ultrasound-micro-bubble (SonoVue) mediated system, respectively. One week later, renal IRI model was made by clasping the left renal artery for 45 min. After 1, 2, 3, 4, 7 and 14 day of reperfusion, the kidney in each group was collected to detect the expression of cyclin D1, cyclin E, CDK4 and CDK2 by western blot analysis or enzyme-linked immunosorbent assay (ELISA). Results Compared with S group, the expression of cyclin D1, cyclin E, CDK4 and CDK2 was significantly up-regulated in day 1, 2,3, 4, 7 and 14 in IR group. And the above index increased gradually after reperfusion, and reached the peak at day 7 (F=54.92, 69.60, 61.28, 77.38, P＜0.05). While in IMD group, these indexes reached the peak at day 1, then progressively declined, and could not be detected at day 14 (compared with the IR group, F=54.92, 69.60, 61.28, 77.38, P＜0.05). Conclusion IMD preconditioning can up-regulate the expression of cyclin D1, cyclin E, CDK2 and CDK4 in the early phase of renal ischemia-reperfusion injury that may accelerate repair of renal tissue, at least by part, by enhancinge cell proliferation.
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Objective To explore whether the change of S phase kinase-associated protein 2 (Skp2) expression could regulate mesangial cell proliferation. Methods Skp2 siRNA and control siRNA, pIRES-GFP-Skp2 plasmid and pIRES-GFP plasmid were designed and synthesized. Cell transfection was performed by Lipofectamine 2000. Skp2 mRNA and protein levels were detected by semiquantitative PCR and Western blotting respectively. Primary culture rat mesangial cells were divided into 6 groups: 0%FCS, 20%FCS, 10%FCS+pIRES-GFP plasmid, 10%FCS+pIRES-GFP-Skp2 plasmid, 20%FCS+control siRNA, 20%FCS+Skp2 siRNA. Cell number was detected by MTT. S phase entry was measured by BrdU labeling. Cell cycle profile was determined by flow cytometric analysis. Results Skp2 mRNA level was significantly down-regulated by Skp2 siRNA compared to control siRNA. Skp2 protein level increased after pIRES-GFP-Skp2plasmid transfection compared to pIRES-GFP plasmid. MTT, BrdU labeling and cell cycle profile demonstrated that cell number (A: 0.419±0.088 vs 0.305±0.036, P＜0.01) and S-phase cells (BrdU labeling positive cell: 0.21±0.04 vs 0.15±0.03, P＜0.01;S-phase cell number:20.18±0.64vs 14.33±0.37, P＜0.01) obviously increased after Skp2 plasmid transfection, while decreased after Skp2 siRNA transfection compared to control groups (A: 0.328±0.069 vs 0.482±0.133, P＜0.01;BrdU labeling positive cell: 0.17±0.01 vs 0.24±0.00, P＜0.01; S-phase cell number: 16.52±0.75vs 23.81 ±1.25, P＜0.01). Conclusion Over-expression of Skp2 stimulates mesangial cell proliferation while down-regulated expression of Skp2 inhibits mesangial cell proliferation.
ABSTRACT
OBJECTIVE: The objective was to assess the effects of oral administration of sodium butyrate on colon carcinogenesis. METHODS: Carcinogenesis in adult male Wistar rats was induced with 1.2-dimethylhydrazine injections at a dose of 40mg/kg of body weight. A solution of sodium butyrate (3.4 percent) was given ad libitum for 4 weeks (butyrate group, n=16) instead of water (control group, n=9). Rats were killed 17 weeks after 1.2-dimethylhydrazine administration. Aberrant crypt foci and expression of the messenger ribonucleic acid (mRNA) of cyclins D1 and E were quantified in the colon. Alterations in the fatty acid profile of the colon, liver, intra-abdominal fat and feces were also analyzed. RESULTS: A significant decrease in aberrant crypt foci was found in the group taking butyrate. No differences were found between the groups in the mRNA expression of cyclins D1 and E. Nevertheless, butyrate intake decreased the content of stearic and oleic acids in the intra-abdominal fat and docosahexaenoic acid in the liver. Moreover, these rats presented higher percentages of linoleic acid in the intra-abdominal fat than control rats. CONCLUSION: The data indicate that butyrate use in rats reduced preneoplastic lesions and changes in the intra-abdominal fat and fatty acid profile of the liver, commonly found in colon carcinogenesis.
OBJETIVO: Avaliar o efeito da administração oral de butirato de sódio na carcinogênese do cólon. MÉTODOS: A carcinogênese em ratos Wistar foi induzida com injeções de 1,2-dimetilhidrazina na dose de 40mg/kg de peso corporal. A solução de butirato de sódio (3,4 por cento) foi oferecida ad libitum por 4 semanas (grupo butirato, n=16), em substituição à água (grupo controle, n=9). Os ratos foram sacrificados na 17ª semana após tratamento com a 1,2-dimetilhidrazina. Focos de criptas aberrantes e a expressão dos genes para o ácido ribonucléico mensageiro (RNAm) das ciclinas D1 e E foram quantificadas no cólon. Alterações no perfil de ácidos graxos do cólon, no fígado, na gordura intra-abdominal e nas fezes foram analisadas. RESULTADOS: Uma significante diminuição nos focos de criptas aberrantes foi encontrada no grupo que recebeu butirato. Nenhuma diferença foi encontrada na expressão do RNAm das ciclinas D1 e E entre os grupos. Contudo, a ingestão de butirato diminuiu a quantidade de ácido esteárico e ácido oléico na gordura intra-abdominal e do ácido docosahexanóico no fígado. Além disso, o grupo butirato apresentou maior percentual de ácido linoléico na gordura intra-abdominal, comparado com os ratos do grupo controle. CONCLUSÃO: Os dados indicam que o uso de butirato reduz lesões pré-neoplásicas em ratos e diminui as alterações no perfil de ácidos graxos da gordura intra-abdominal e do fígado, comumente encontradas na carcinogênese colônica.