Purpose To study changes of cell cycle of vascular endothelial cell in non-proliferative diabetic retinopathy. Methods Alloxan induced Wistar-rats were employed and immunohistochemistry,Western blotting methods were used. Results The vascular endothelial cells of retinas of 8～20 weeks diabetic rats were observe to be cyclinD1,cyclinD3,cyclinB1,p21 and p27 positive stained with light and electronmicroscopies.CyclinE immuno-stained vascular endothelial cells was observed occasionally.Meanwhile,the evidences of morphologic changes of the vascular en dothelial cells were proved:less plasma,thinner cell,more bubble organelles than those of controls.But,the ultra-structures of pericytes and other type of retinal cells did not change and they also immunostain negative.Komas blue and Western blotting methods also proved that the vascular endothelial cells of retina of 20th week diabetic rats expressed cyclinD1,cyclinB1,p21 and p27 protein. Conclusion Glucose induced retinal vascular endothelial cells of 8～20th weeks diabetic rats enter cell cycle and were arrested at G1/S restriction point.This study also suggested that retinal vascular endothelial cells may possess the ability to resist glucose damage and mechanism of selfstability during very early stage of diabetes. (Chin J Ocul Fundus Dis,2000,16:173-176)
Objective To investigate cell cycle as a new tool to evaluate the biocompatibility of biomaterials.Methods The cell cycle and the expression of related genes were analyzed by the methods of immunocytochemistry, protein blotting, RT PCR and flow cytometry. Results The physical properteis, chemical properties and topological properities of biomaterials could not only influence cell cycle of the cells attached onto biomaterials but also affect the expression of related genes of target cells. Conclusion As an important extension of routine proliferation epxeriments, the study of cell cycle control will be great help for us to to study the cell group as an organic society. It revealed the balance between cell proliferation, cell differentiation and apotosis. It is suggested that the study of cell cycle control will play a key role in the research of tissue engineering.
Objective To investigate the effect of exogenous Rb gene on the cell cycle of vitreous retinoblastoma (RB) transplantation tumor in nude mouse. Methods Based on establishing vitreous RB transplantation tumor in nude mouse,constructing retrovirus vector of Rb gene PBabe-Rb and transfecing it into the RB transplantation model by liposome Dosper,the change of cell cycle of the RB transplantation tumor by flow cytometry(FCM)was analysed． Results FCM showed that the cells of G1phase of the treated eyes were obviously more than the control eyes with the value of DNA index(DI)and S phase fraction(SPF) decreased by the Rb gene expression. Conclusion The exogenous PBabe-Rb gene can partially suppress the progress of the cell cycle of RB transplantation tumor in vivo. (Chin J Ocul Fundus Dis，2000，16：1-70)
ObjectiveTo construct DPC4 gene recombinant expression vector and to study the inhibitory effect of DPC4 on the growth of human pancreatic adenocarcinoma cell line (PC3) cells.MethodsDPC4 cDNA was amplified from K562 cell line using RTPCR, and was cloned into the pcDNA3.1 vector to construct a recombinant expression vector plasmid pcDNA3.1DPC4. The recombinant expression plasmid was transferred into PC3 cells by liposome method. After G418 selection, cell cycle and apoptosis were assessed by flow cytometry, then the cell growth rate was estimated by cell count. The cells being not transferred plasmid and transferred pcDNA3.1 plasmid were used as controls.ResultsThe DPC4 gene recombinant expression vector was constructed. Wildtype DPC4 gene attributed to the increase of G1 phase cells and the decrease of S phase cells in PC3 cells,and could inhibit the growth of PC3 cells, the cell growth rates was reduced to 34.3%-41.1% of that of the controls, but cell apoptosis was not observed on all groups. ConclusionWildtype DPC4 gene could inhibit the proliferation of human pancreatic adenocarcinoma cells and could become one of the target genes of pancreas adenocarcinoma gene therapy
Objective To observe the replicative senescence of rat articular chondrocyte cultured in vitro so as to provide reference for the succeeding experiment of using medicine interfere and reverse the cataplasia of tissue engineering cartilage or probing cataplasia mechanism．Methods Different generations(P1, P2, P3 and P4) of the chondrocytes were detected with the methods of histochemistry for β-galactosidase (β-gal)， electronmicroscope for ultromicrostructure， immunocytochemistry for proliferating cell nuclear antigen (PCNA)，alcian blue stain for content and structure of sulfatglycosaminoglycan (GAG) of extracellular matrix (ECM)，reverse transcriptionpolymerase chain reaction (RTPCR) for content of collagen Ⅱ，flow cytometry for cell life cycle and proliferative index(PI) to observe senescence of chondrocytes．Results In the 4th passage，the chondrocytes emerging quantitively positive express of β-gal，cyto-architecture cataplasia such as caryoplasm ratio increasing and karyopycnosis emerging under electronmicroscope ，cell life cycle being detented on G1 phase(83．8％)，while in P1, P2, P3 the content of G1 phase was 79.1%, 79.2%, 80.8% respectively. In the 4th passage, PI decreased(16．2％)，while in P1, P2, P3, it was 20.9%, 20.8%, 19.2%. The positive percentage of PCNA，the content of GAG(long chain molecule) and the positive expression of collagen Ⅱ diminished,all detections above were significantly different (Plt;0.01) when compared the 4th passage with the preceding passages．Conclusion Chondrocytes show the onset of senescence in the 4th passage．
Objective To investigate the effect of the 8-bromum-cyclic adenosine monophosphate (8-Br-cAMP) on the telomerase activity and changes of cell cycle in retinoblastoma （RB） cells. Methods The cultured RB cells were divided into the experimental group (8-Br-cAMP) and control group. After cultured for 24, 48 and 72 hours in vitro, the telomerase activity of RB cells was detected by polymerase chain reaction enzyme-linked immunosorbent assay (PCR-ELISA) and the changes of cell cycle were detected by flow-cytometry. Results The difference of telomerase activity was significant between the experimental groups and control group (Plt;0.01). There was a negative correlation between the A value of absorbance and the time in the experimental groups (r=-0.778 9, F=33.936, Plt;0.01). The changes of the cell cycle were that the percentages increased in G1 phase and decreased in S phases. Conclusion 8-Br-cAMP may weaken telomerase activity, affect the cell cycle, and inhibit the proliferation of RB cells. (Chin J Ocul Fundus Dis，2004，20：358-360)
Objective To evaluate the role of the cell cycle related genes cyclinD1 and Bcl-2 protein expression in the pathogenesis and infilt ration of the uveal melanoma． Methods Using immunohis tochemistry to detect the cyclinD1 and bcl-2 protein expression in 96 cases of uveal melanoma． Results The expression content of bcl-2 was high in uveal melanoma, and there wasn't any relationship between bcl-2 cell positivity and tumor cell type and extrascleral extension． In contrast, cyclinD1 expression was higher in epithelial cell uveal melanoma than mix cell and spindle cell varieties． There was a positive correlation between cyclinD1 cell positivity and extrascleral extension． Conclusion The expression of bcl-2 protein is important for the survival of the uveal melanoma． CyclinD1 may serve as a sensitive index of its malignancy． (Chin J Ocul Fundus Dis, 2001,17:44-46)
ObjectiveTo study the effects of recombinant human growth hormone (rhGH) on proliferation of human rectal cancer cell in vitro. MethodsThe experiment was divided into control group,rhGH group,Oxaliplatin (LOHP) group and rhGH+LOHP group. The double proliferation time of cells,cell inhibition rate,cell cycle, proliferation index (PI) and DNA inhibition rate of human rectal cancer line,HR8348,were studied by cell culture, MTT assay and flow cytometry on different concentration of rhGH. ResultsIn vitro the markedly accelerated effects of rhGH on multiplication of HR8348 cell line were not found: there was no statistical significance as compared rhGH group with control group or compared rhGH+LOHP group and LOHP group （Pgt;0.05）. The double proliferation time of cells was markedly lengthened, cell inhibition rate and the cells arrested in G0-G1 phase were obviously increased, meanwhile, the cells in S phase （P＜0.05） and G2-M phase and PI were markedly decreased and DNA inhibition rate was obviously risen as compared rhGH+LOHP group with control group or rhGH+LOHP group and rhGH group （P＜0.01）.ConclusionIn vitro rhGH does not accelerate the multiplication of human rectal cancer cells.
Objective To study the effects of L-arginine （L-Arg） on cell proliferation, inducible nitric oxide synthase (iNOS) expression and cell cycle in human colon carcinoma cell line LS174 through nitric oxide (NO) pathway. Methods LS174 cells were cultured in medium with L-Arg at different concentrations for different times. MTT method was employed to evaluate the level of the cell proliferation. The production of NO in culture supernatants of LS174 cell was detected with enzyme reduction of nitrate. The distribution of the cell cycle was detected with the flow cytometry (FCM). The expression level of iNOS in the cells was determined by Western blot and SP immunocytochemical staining method. Results The growth of LS174 was promoted by the L-Arg at low concentration (0.125 mmol/L) and inhibited at high concentrations (0.5， 2， 8 and 32 mmol/L). The level of NO was increased with the increasing concentration of L-Arg in culture medium. To compare with the control group, the ratio of cells at S phase was increased after 48 hours’ treatments with high concentrations (0.5, 2, 8 and 32 mmol/L) of L-Arg (P＜0.05, P＜0.01); while there was no obvious difference after treatments with low concentration (0.125 mmol/L) of L-Arg (Pgt;0.05). With the increase of the concentration of L-Arg, the expression of iNOS was increased as compared with control group. The higher the concentration of L-Arg was, the better the effect. Conclusion L-Arg can induce the expression of iNOS resulting in increase the production of nitric oxide (NO). Low concentration of L-Arg can promote the growth of LS174 cells, while high concentration ones can inhibit growth and proliferation. The high concentration of L-Arg could induce S phase arrestion in the cell cycle.
【Abstract】ObjectiveTo investigate the effect of CDAⅡ on the cell cycle progression of breast cancer cells.MethodsThe effects of CDAⅡ on growth curve, cell cycle progression and morphology of breast cancer cell lines MCF7 and MDAMB231 were observed when CDAⅡ and MCF7 or CDAⅡ and MDAMB231 were blended to cultivate in vitro, in comparison with the classical cell differentiation inducer ATRA. ResultsCDAⅡ decreased the growth speed and inhibit proliferation ability in breast cancer cell lines MCF7 and MDAMB231.It caused G0/G1 phase block of cell cycle and reduced the rate of S phase of breast cancer cells. ConclusionCDAⅡ has remarkable effect of anticellproliferation and can induce cell cycle block of G0/G1 on breast cancer cells. This results provide experimental bases for the treatment of breast cancer with CDAⅡ.