The pathways of cell cycle regulation in retinoblastoma / As vias de regulação do ciclo celular em retinoblastoma

Retinoblastoma is a childhood ocular tumor often caused by the biallelic inactivation of the RB1 gene affecting children up to 5 years of age. A retinoblastoma protein (pRB), encoded by the tumor suppressor gene RB1, is responsible for the regular progression of the G1 phase to the phase S of the cell cycle. This protein forms a complex with the transcriptional factor E2F causing the cell cycle to remain in the G0/G1 stage. With a phosphorylation of cyclin-dependent kinases (CDK), the phosphorylation of the RB protein is activated and the complex formed with E2F is disrupted, with the advancement of the cell cycle to an S phase and cell proliferation. All the control of cell proliferation is regulated not only by the complex formed by RB and E2F proteins, but also by other proteins that participate in and/or interfere in this cell division control mechanism, such as mdm2, mdm4 and p21 proteins.

O retinoblastoma é um tumor ocular infantil ocasionado, frequentemente, pela inativação bialélica do gene RB1 acometendo crianças até os 5 anos de idade. A proteína retinoblastoma (pRB), codificada pelo gene supressor tumoral RB1, é responsável por regular a progressão da fase G1 para a fase S do ciclo celular. Essa proteína forma um complexo com o fator transcricional E2F fazendo com que o ciclo celular permaneça no estágio G0/G1. Com a fosforilação de quinases dependentes de ciclinas, a fosforilação da proteína RB é ativada e o complexo formado com o E2F é desfeito, havendo o avanço do ciclo celular para a fase S e a proliferação celular. Todo esse controle da proliferação celular é regulado não só pelo complexo formado pela proteína RB e E2F, mas também por outras proteínas que participam e/ou interferem neste mecanismo de controle da divisão celular, como, por exemplo, as proteínas mdm2, mdm4, p21

Effect of ERH gene on migration and invasion of human bladder cancer cells T24 and 5637 / 中国医师杂志

Objective To study the effect of enhancer of rudimentary homolog (ERH) gene on migration and invasion in human bladder cancer T24 and 5637 cells.Methods After knocking out the ERH gene of human bladder cancer T24 and 5637 cells,Wound healing assay,Transwell cell migration assay and Transwell cell invasion assay were used to verify the migration and invasion function.Cell migration related protein was detected by Western blot.Nude mouse tail vein transfer assay was used to study the metastasis ability of bladder cancer cells in vivo.Results (1) The Wound healing assay showed that there were significant differences in the migration cell counts of human bladder cancer 5637 and T24 (P ＜ 0.05).(2) There were significant differences in migration and invasion cell counts of Transwell assay (P ＜0.05).(3) Western blot showed that the expression of E-Cadherin in human bladder cancer 5637 cells and T24 cells was significantly increased (P ＜ 0.05) after knocking out ERH gene,while the expression of Fibronectin,Twist,Vimentin and Snail2 protein were significantly decreased (P ＜ 0.05).(4) Nude mouse tail vein transfer assay showed that lung metastases were significantly reduced in the ERH knockout group compared with the normal ERH group.Conclusions Both in vitro and in vivo experiments suggest that ERH knockout affects the migration and invasion of human bladder cancer T24 and 5637 cells.

Effect of microRNA-1180 transfection on growth of renal cell carcinoma / 重庆医学

Objective To investigate the effect of microRNA-1180 transfection on the growth of renal cell carcinoma lines 786-O and ACHN.Methods The renal carcinoma cells were divided into the two groups:control group (transfecting dsControl) and experimental group (transfecting miR-1180).The expression change of p21 mRNA was detected by qRT-PCR.Western blot was conducted to analyze the expression changes of p21,CDK4,CDK6 and CyclinD1 proteins.Flow cytometry (FCM) was used to detect the cell cycle change.The MTS assay was conducted to detect the cell viability and the colony forming assay was performed to examine the cell proliferation ability.Results The qRT-PCR results showed that compared with the negative control dsControl group,after miR-1180 transfection,the expression level of·p21 mRNA in 786-O and ACHN cells was up-regulated to 2.54-fold and 2.49-fold respectively(P＜0.01).The expression trend of p21 protein was consistent with qRT-PCR results.The expression of CDK4,CDK6 and CyclinD1 proteins were significantly down-regulated.The FCM results showed that the proportion of cells in G0/ G1 phase was significantly increased after transfection of miR-1180,but the proportion of cells in S phase and G2/M phase was decreased significantly,indicating that the cell cycle was arrested in G0/G1 phase.The MTS assay results showed that compared with the dsControl group,the viability of the two kinds of renal carcinoma cells was significantly decreased.The colony formation assay showed that the number of colonies formed in the miR-1180 group was smaller,indicating the proliferation ability of miR-1180 transfected cells was decreased.Conclusion miR-1180 can significantly activate the p21 protein expression and inhibit the growth of renal carcinoma cell lines 786-O and ACHN.

Overexpression of Cell Cycle Proteins of Peripheral Lymphocytes in Patients with Alzheimer's Disease

OBJECTIVE: Biological markers for Alzheimer's disease (AD) will help clinicians make objective diagnoses early during the course of dementia. Previous studies have suggested that cell cycle dysregulation begins earlier than the onset of clinical manifestations in AD. METHODS: We examined the lymphocyte expression of cell cycle proteins in AD patients, dementia controls (DC), and normal controls (NC). One-hundred seventeen subjects (36 AD, 31 DC, and 50 NC) were recruited. The cell cycle proteins CDK2, CDK4, CDK6, cyclin B, and cyclin D were measured in peripheral lymphocytes. Cell cycle protein expression in the three groups was compared after adjusting for age and sex. RESULTS: The levels of cell cycle proteins CDK2, CDK4, CDK6, cyclin B, and cyclin D were significantly higher in AD patients than in the NC subjects. The DC group manifested intermediate levels of cell cycle proteins compared with the AD patients and the NC subjects. The present study indicates that cell cycle proteins are upregulated in the peripheral lymphocytes of AD patients. CONCLUSION: Cell cycle dysregulation in peripheral lymphocytes may present a promising starting point for identifying peripheral biomarkers of AD.

Correlation between TRIB3 gene rs2295490 polymorphism and coronary heart disease with dyslipidemia / 中华检验医学杂志

Objective To explore the correlation between the polymorphism of TRIB3 gene rs2295490 and coronary heart disease (CHD) with dyslipidemia.Methods In the case control study,we enrolled 220 CHD patients as observation group (60.15 years mean age,150 males and 70 females) and 151 age and gender matched healthy individuals as control group (59.22 years mean age,102 males and 49 fe males).Genotypes of TRIB3 gene rs2295490 polymorphisms in the two groups were detected by PCR and sequence-based testing.At the same time,blood glucose (GLU),total cholesterol (TC),triglyceride (TG),high density lipoprotein cholesterol (HDL-C),low density lipoprotein cholesterol (LDL-C) were measured.The correlations between the polymorphism of TRIB3 gene rs2295490 and CHD,biochemical indicators of two groups were analyzed,respectively.The comparison of genotype and allele frequency between two groups used x2 test,while the comparison of biochemical indexes between two groups used t test.Results In the control group,genotypic frequencies of AA,AG and GG of TRIB3 gene rs2295490 were 57％,35％ and 8％,and the frequencies of A and G alleles were 75％ and 25％,respectively.In the CHD group,genotypic frequencies of AA,AG and GG of TRIB3 gene rs2295490 were 50％,41％ and 9％,and the frequencies of A and G alleles were 71％ and 39％,respectively.There was no significant difference of genotype and allele frequencies of polymorphism of TRIB3 gene rs2295490 between CHD and healthy controls(x2 =1.74,P =0.42;x2 =1.26,P =0.243).In the CHD group,the HDL-C levels of the AA genotype were higher than those of AG + GG genotype (t =-9.78,P =0.00);The TG,LDL-C levels of the AA genotype were lower than those of AG + GG genotype (t =2.59,P =0.01;t =6.15,P =0.00).All the differences were statistically significant.Conclusions TRIB3 gene rs2295490 polymorphism has no correlation with CHD,but the G allele may be correlated with dyslipidemia in CHD patients.

Expresión simultánea de los marcadores p16INK4a y Ki-67 en extendidos de citología cérvico-vaginal anormales de archivo / Simultaneous expression of markers in smears of abnormal cervical cytology previously stored

Los biomarcadores Ki-67 y p16INK4a se utilizan en citología cérvico-vaginal para facilitarla detección de células anormales, debido al incremento en su expresión durante la transformación neoplásica de células epiteliales escamosas de cuello uterino. El hallazgo de estos marcadores es mutuamente excluyente en una célula normal, por tanto, la expresión dual p16INK4a/Ki-67 se considera un marcador sensible y específico de lesiones premalignas o malignas. Objetivo: Determinar la expresión simultánea de los marcadores p16INK4a y Ki-67 en extendidos de citología cérvico-vaginal convencionalanormales almacenadas en archivo.

The p16INK4a and Ki-67 biomarkers are used in cytology to help the detection of abnormal cells, due to their overexpression during the neoplastic transformation of cervix squamous epithelial cells. The expression of both markers in normal cells, is mutually exclusive, therefore, p16INK4a/Ki-67 dual-stained has shown high sensitivity and specificity in pre-malignant and malignant lesions. Objective: The aim of this study was to determine the simultaneous expression of p16INK4a and Ki-67 markers in conventional cervical abnormal cytology previously stored.

The synergistic effects of docetaxel and adenovirus-mediated NDRG2 gene on prostate cancer cell line DU145 / 中国医师杂志

Objective To investigate the antitumor activities of adenovirus-mediated NDRG2 gene (Ad-NDRG2) and docetaxel on human prostate cancer DU145 cells.Methods The protein expressions of cyclin D1,cycliu E,and NDRG2 in the cells were determined by Western blot.MTT and flow cytometry were used to observe the effects of docetaxel (10-6 mol/L,10-7 mol/L,and 10-s mol/L) and Ad-NDRG2 on prostate cancer cell line DU145 in single or synergistic administration ways for 24 and 48 hours in vitro.Male BALB/C-nu mice with DU145 prostate cancer cell lines were treated by docetaxel and Ad-NDRG2 singly or synergistically in vivo.Results After infected by adenovirus,the protein expression of NDRG2 increased,but cyclin D1 and cyclin E decreased in DU145 cells.Ad-NDRG2 inhibited the cell growth (inhibition ratio =41.8％,t =4.18,P ＜0.01),promoted apoptosis (apoptosis ratio =32.4％,x2 =11.66,P ＜0.05),changed the ratio of G2/M phase from 50.2％ to 23.6％,and reversed partially the G2/M arrest,of DU145 cells induced by 10-7 mol/L docetaxel.In vivo experiment showed that docetaxel,Ad-NDRG2,and combination of docetaxel and Ad-NDRG2 inhibited tumor growth with a inhibition rate of 30.7％,28.2％,and 55.8％,respectively.The coefficient of drug interaction (CDI) of docetaxel and Ad-NDRG2 was 0.89.Conclusions Ad-NDRG2 can enhance the growth suppression and apoptosis induced by docetaxel in synergistic way in vitro and in vivo.It demonstrated the great potential of Ad-NDRG2 in the treatment of androgen-independent prostate carcinoma.

Expression of growth arrest and DNA damage-inducible 45α and p38 mitogen-activated protein kinase in placentas of women complicated with preeclampsia / 中华围产医学杂志

Objective To evaluate the expression of Gadd45α and p38 MAPK in placentas and the correlations of Gadd45α protein and serum soluble vascular endothelial growth factor receptor-1 (sFlt-1) and soluble endoglin (sEng) in preeclampsia(PE). Methods Fifty-four pregnant women who delivered from September 2009 to March 2010 in the First Affiliated Hospital of Chongqing Medical University were chosen as the subjects. They were classified into mild preeclampsia group (n=20),severe preeclampsia group (n=16) and the control group (normal pregnant women underwent elective cesarean sections at term without labor and perinatal complications, n=18). Western blot and immunohistochemistry were employed to determine the expression and localization of Gadd45α and p-p38 MAPK protein respectively. Gadd45α mRNA level was determined by quantitative real-time PCR. The levels of seum sFlt-1 and sEng were measured by enzyme-linked immunosorbent assay (ELISA). One-way ANOVA and LSD-t test were applied for statistical analysis. Results (1)Immunohistochemistry identified that the positive stained cells were mostly located in trophoblast cells in normotensive placentas, whereas in preeclamptic placentas Gadd45α protein and p-p38 MAPK protein were detected in trophoblast and endothelial cells, as well as a few stromal cells at increased levels.(2)The mRNA levels of Gadd45α was significantly elevated in mild and severe preeclampsia groups compared to the control group (2.10±0.11 and 3.33±0.13 vs 1.01±0.18, P＜0.05), and Gadd45α mRNA level in severe group was significantly higher than in mild group (P＜0.05).(3)The data of Western blot revealed that the Gadd45α protein levels in each group were 0.22±0.11, 0.65±0.15 and 1.34±0.17, respectively, with significant differences between each group(P＜0.05). The p-p38 MAPK protein levels in each group were 0.32±0.08, 0.72±0.12 and 1.45±0.21, respectively, with significant differences between each group (P＜0.05). p38 MAPK protein levels in the total groups showed no difference(P＞0.05).(4)Compared with the control group, sFlt-1 and sEng concentrations in maternal circulation were significantly increasing in mild and severe preeclampsia groups, and concentrations in severe group were significantly higher than those in mild group (P＜0.05).(5) There were positive correlations between Gadd45α protein levels and the concentrations of serum sFlt-1 and sEng in each group( r=0.88 and 0.87, respectively all P＜0.05). Conclusions Upregulation of Gadd45α in preeclampsia placentas may play an important role in the pathogenesis of preeclampsia. It may induce the increased maternal serum levels of sFlt-l and sEng by activating p38 MAPK signaling pathway, leading to deficient cytotrophoblastic invasion and abnormal placental vascular reconstruction during pregnancy.

Expression and significance of CDK4,p18,p19 in canceration of esophageal epithelium / 肿瘤研究与临床

Objective To investigate the role of cell cycle regulatory protein CDK4,p18,p19 in the genesis and development of esophageal squamous cell carcinoma (SCC).Methods Tissue microarray and immunohistochemical method (Envision) were used to detect the protein expression of CDK4,p18,p19 in 120 cases of esophageal tissues.The results were statistically analyzed.Results The positive rate of CDK4 protein expression in normal esophageal epithelium was low [28.3 % (34/120)],it increased in esophageal intraepithelial neoplasia [32.5 % (39/120)],and it was high in esophageal SCC [84.2 % (101/120)],which increased with the degree of SCC differentiation decreasing gradually.There was significant differences between the SCC and normal esophageal epithelium or esophageal intraepithelial neoplasia (x2= 76.004,P ＜0.05; x 2= 65.897,P ＜ 0.05).The expression of CDK4 in group with lymphatic metastasis [93.88 % (46/49)]was higher than without it [71.43 % (55/71)] (x2= 5.860,P ＜ 0.05).The positive rates of p18,p19 protein expression in normal esophageal epithelium were high [34.2 % (41/120),29.2 % (35/120)],it decreased in esophageal intraepithelial neoplasia [19.2 % (23/120),15.0 % (1 8/120)] (x 2= 134.481,P ＜ 0.05; x 2 = 141.376,P ＜ 0.05),but it were high in esophageal SCC [63.3 % (76/120) and 61.7 % (74/120)] which decreased with the degree of SCC differentiation gradually increased.There were significant differences between the normal esophageal epithelium and esophageal intraepithelial neoplasia,esophegeal intraepithelial neoplasia and SCC,normal esophageal epithelium and SCC (p 18:x 2 = 6.903,48.296,20.429,P ＜ 0.05; p1 9:x2 = 6.998,55.276,25.565,P＜ 0.05).CDK4 protein expression was correlated with both p18 and p19 (r =0.696,0.630,P ＜0.05),and there was significant positive correlation between the protein expression of p18 and p19 (r =0.833,P ＜0.05).Conclusion Cell cycle regulatory gene CDK4,p18,p19 get involved in the genesis and development of esophageal squamous cell carcinoma.Their protein expressions are closely related to canceration of esophageal epithelium.

Anticancer Effects of Astaxanthin and alpha-tocopherol in Esophageal Cancer Cell Lines

BACKGROUND/AIMS: Astaxanthin (AX) has been attributed with potential for protecting the organism against different types of cancer due to its anti-oxidant activity. Also several in vivo and in vitro studies suggest certain naturally occurring vitamin E (i.e. alpha-tocopherol) as promising anticancer agents. We assessed the effect of AX and alpha-tocopherol (AT) respectively and their combination on human esophageal cancer cell lines to investigate the mechanism of anticancer effect and their therapeutic potential. MATERIALS AND METHODS: Two human esophageal cancer cell lines (TE-1, TE-4) were exposed to AX (6 to 10 microg/mL) and AT (20 to 100 microM) for 24 hours. Quantification of proliferation was performed by MTT assay. Cell cycle machinery proteins such as p-AKT, p-p38, cyclin D1, p27 and caspase-3 were investigated by Western blot. RESULTS: Significant inhibition of cell proliferation of AX and AT was observed in TE-4 cell line by a dose-dependent manner. Furthermore, AX and AT as single agents increased the protein expression of p27 and cleaved caspase-3 in TE-4 cell line. The combination of the two agents decreased the expression of cyclin D1, however they did not demonstrate pro-apoptotic effect. CONCLUSIONS: AX and AT as single agents are effective at inhibition of cell proliferation and induce apoptosis by the modulation of cell cycle machinery proteins in esophageal cancer cell lines. However, our data could not suggest that their combination has any cooperative apoptotic effect.

Expressão das proteínas p53, p16 e ki67 no adenocarcinoma da cabeça do pâncreas e sua relação com a sobrevida e diferenciação celular / Expression of p53, p16 and Ki67 proteins in ductal adenocarcinoma of the pancreatic head and their relation with survival and cell differentiation

Objective: To determine the expression of p53, p16 and Ki-67 and its relevance in survival and cell differentiation. Methods: Fifteen duodenopancreatectomized patients were included. Immunohistochemical expression of p53, p16 and Ki-67 was determined in paraffin embedded tumor blocks. The relation of these expressions with different variables was studied. Results: Ninetythree per cent of tumors showed expression of p53 and p16. Ki- 67 was expressed in 86.66% of tumors (labeling index plus or minus LI 11.91 ± 9.47). The presence of combined alterations was not related to significant differences in tumor type, stage or survival; similar results were obtained analyzing isolated expressions. When groups of p16 and Ki-67 expressions where created, the median survival was not significant. However, there was a slightly better survival in patientswith focal expression of p16 (median survival 20.75 versus 14.34), when compared to patients with diffuse expression. Conclusion: The overexpression of p53, p16 and Ki-67 was not related to survival or tumor grade, when comparing isolated or combined expressions.

Objetivo: Determinar a expressão de p53, p16 e Ki-67 e sua relevância na sobrevida e diferenciação celular. Métodos: Foram incluídos 15 pacientes submetidos a duodenopancreatectomia. A expressão imunohistoquímica de p53, p16 e Ki-67 foi determinada em blocos tumorais embebidos em parafina. Foi estudada a relação dessas expressões com as variáveis. Resultados: Noventa e três por cento dos tumores apresentaram expressão de p53 e p16. Ki-67 estava expresso em 86,66% dos tumores (índice proliferativo mais ou menos IP 11,91 ± 9,47). A presença de alterações combinadas não estava relacionada a diferenças significativas no tipo tumoral, no estágio ou na sobrevida; resultados semelhantes foram obtidos com a análise de expressões isoladas. Quando foram criados os grupos de expressões de p16 e Ki-67, a sobrevida mediana não era significativa. Entretanto, havia uma sobrevida discretamente melhor nos pacientes com expressão focal do p16 (sobrevida mediana 20,75 versus 14,34) em comparação com pacientes com expressão difusa. Conclusão: A superexpressão das proteínas p53, p16 e Ki-67 não estava relacionada à sobrevida ou ao grau tumoral quando se compararam as expressões isoladas ou combinadas.

Expression changes of the growth arrest and DNA damage 45 alpha gene in placenta in patients with preeclampsia and its clinical significance / 中华妇产科杂志

Objective To evaluate the expression and location of the growth arrest and DNA damage 45 alpha(Gadd45α)gene in human placenta and explore the relationship between Gadd45α and preeclampsia(PE).Methods Thirty-six women with preeclampsia who delivered from September 2009 to March 2010 in the First Affiliated Hospital of Chongqing Medical University were chosen as the study objects.They were classified into mild group( n = 20), severe group( n = 16) and elective term cesarean sections group without previous onset of labor and perinatal complications ( control group, n = 18 ).Placentas were collected after they delivered and immunohistochemical streptravidin-biotin peroxidase(SP) method and histoscore were employed to detect the expression and localization of Gadd45α protein.Gadd45α mRNA level was determined by reverse transcription polymerase chain reaction (RT-PCR) technique and western blot analysis was used to quantify Gadd45α protein expression level.Results (1)Gadd45α protein was detectable in placenta tissues of the mild and severe groups, mostly located in cytoplasm and nuclei of trophoblast, plus nuclei of vascular endothelial cells and a few stromal cells; whereas placenta tissues of control group showed weak staining, and only detectable in trophoblast, undetectable in vascular endothelial cells.(2)The Gadd45α mRNA levels in placenta tissues of the severe and mild groups were 0.75 ±0.07 and 0.44±0.13, respectively, significantly higher than that in control group (0.18 ±0.04, P ＜0.05);compared with mild group, Gadd45α mRNA level was significantly higher in severe group( P ＜ 0.05 ).(3)The Gadd45α protein levels in placenta tissues of the severe and mild groups was 1.34 ±0.17 and 0.65 ±0.15, respectively, compared with mild group, Gadd45α protein level was higher in severe group (P ＜0.05); they were both significantly higher than that in control group ( 0.22 ± 0.11, P ＜ 0.05 ).Conclusions Gadd45α mRNA and protein levels in placenta tissues of patients with preeclampsia are higher than that of normotensive women, and up-regulated with aggravation of preeclampsia; the Gadd45α protein is located in trophoblast cells, which are closely related to pathogenesis of preeclampsia.These results indicate that Gadd45α plays an important role in the pathogenesis and progression of preeclampsia.

Expressão de marcadores de proliferação celular e apoptose em carcinoma basocelular / Markers expression of cell proliferation and apoptosis in basal cell carcinoma

Fundamentos: O carcinoma basocelular é o câncer mais comum em humanos. Estudos que utilizam recursos da biologia molecular e genética, associados à histomorfologia, permitem a identificação de fatores de risco no desenvolvimento de lesões mais recorrentes e agressivas. Objetivo: Correlacionar a expressão dos marcadores de apoptose (p53 e Bcl-2) e proliferação celular (Ki-67 e PCNA) com os indicadores histológicos de gravidade do tumor. Métodos: Estudaram-se cinco amostras das formas nodular, morfeiforme e superficial, respectivamente, e um grupo-controle com três pacientes livres de lesão. Empregou-se o teste de Mann-Whitney na comparação da expressão desses marcadores com a forma de apresentação do carcinoma basocelular. Resultados: Verificou-se que a marcação do Bcl-2 foi expressiva nos CBCs ditos agressivos (variantes morfeiforme e nodular). Dos tumores estudados, 66,7 por cento (n = 10) indicaram fortemente o p53. Nossos resultados mostram maior expressão do Ki-67 no carcinoma basocelular nodular e superficial, sem expressão nos controles. O PCNA mostrou forte marcação em todos os tipos de tumores e nos controles. Conclusão: Os achados nos permitem concluir que o Bcl-2 e o p53 apresentam tendência para diagnosticar gravidade do carcinoma basocelular e o Ki-67, por seu comportamento variável, não pode ser considerado como marcador de gravidade, assim como o PCNA, que não foi um bom marcador de proliferação celular.

Background: - Basal cell carcinoma is the most common form of human cancer. Studies employing molecular and genetic biology techniques, associated with histomorphology, lead to the identification of risk factors in the development of more recurring and aggressive lesions. Objetive - To correlate markers expression of apoptosis (p53 and bcl-2) and cell proliferation (Ki-67 and PCNA) with histological indicators of tumor severity. Methods - Five samples of the nodular, morpheaform and superficial types of carcinoma were studied, espectively.One control group with three lesion-free patients was also included in the study. The Mann-Whitney test was used to compare these markers expression with the manifestation form of basal cell carcinoma. Results - Bcl-2 expression was significant in basal cell carcinomas said to be aggressive (morpheaform and nodular types). Of the studied tumors, 66.7 percent (n =10) strongly expressed p53.Our results show a greater expression of Ki-67 in nodular and superficial basal cell carcinoma, with no expression in the controls. PCNA showed a strong expression in all types of tumors and in the controls. Conclusion - The findings allow us to conclude that Bcl-2 and p53 show a tendency to indicate the severity of basal cell carcinoma. In contrast, Ki-67, due to its variable behavior, cannot be considered a marker of severity. Also, PCNA was not a good marker of cell proliferation.

Effects of adhesion mediated by bone marrow stroma cells from leukemia patient on cell cycle of Jurkat cells / 中国医师杂志

Objective To investigate the effects of adhesion mediated by bone nlalTow stroma celh from leukemia patient on chemotherapeutics sensitivity and cell cycle of Jurkat cells in the co-cultured model.Methods Bone mw stroma cells were isohted and cultured from leukemia patients routinely.To construct the co-cultured model.Jurkat cells were co-cultured with BMSCs the irradiated layer by 60Co,and the model was observed with scanning electron microscope.The IC50 values of Jurkat cells expesured to DNR were quantified by MTT.The cell cycles of Jurkat cells after 24h-adhesion in the co-cultured model were analyzed by Facs.The expression of cyclin A,cyclin E and p27 in Jurkat cells adhered to BMSCs for 4h.24h and 48h were detected by Western blot.Results Jurkat ceUs in the co-cultured model showed a decreased sensitivity to DNR.ICSO values for normal BMSCs,leukemic BMSCs and non-adhered control were of 1.78Ixmol/L,2.30pznol/L and 0.45p,mol/L,respectively.The percentages of Go-Gl phase for leukemic BMSCs group and non-adhered control group were of 48.74％±8.77％and 27.83％壬1.86％.Respectively.The percentages of Gz-M phase for leukemic BMSCs group and non-adhered control group were of2.01％±1.17％and 20.33％±1.84％。Respectively.Compared with eomrol group,the 24h-ad- hesion mediated by BMSCs from leukemia patients up-regulated the percentage of Go-G1 phase of Jurkat cells(P<O.01),but down-regula-ted the percentage of G2-M phase(P<O.01).The expression of cyclin A and cyclin E in Jurkat ceUs was up-regulated when adhered to BMSCs for four hours and the higher expression emerged after adhering for 24h and 48h.Oppositely,the expression of p27 were down-regulated.Especially after 48h-adhesion.Conclusions Drug resistance of leukemia cells csn be mediated by adhering to bone marrow stroma cells,which may be related to the changes of cell cycle.

The correlation in transcriptional expression with promoter hypermethylation of AKAP12 gene in bladder transitional cell carcinoma / 中华检验医学杂志

Objective To investigate transcriptional expression and promoter CpG methylation status of A-kinase anchoring protein 12 (AKAP12) gene and analyze their correlation with clinical pathological stage in bladder transitional cell carcinoma. Methods AKAP12 mRNA expression level and promoter CpG metbylation status was measured by fluorescent quantitative RT-PCR (FQ-RT-PCR) and methylation specific PCR (MSP) in 30 bladder transitional cell carcinoma and adjacent normal tissues. The products of PCR were cloned and bisulfite sequenced. Results Decreased AKAP12 mRNA expression was demonstrated in 22 carcinomas (73. 3% ) and was significantly associated with turnout grade (P =0. 02).The frequency of promoter methylation of AKAP12 gene was 53. 3 % (16/30) and correlated with the tumor stage(r =0.52,Pn =0.03)and grade(r =0.61,Pn =0.01). Conclusion Aberrant promoter methylation of AKAP12 can result in the loss of gene expression and may association with bladder transitional cell carcinoma.

Expression of human spindle mitosis arrest deficiency gene in spontaneous abortion embryo tissues / 中华妇产科杂志

Objective To investigate the expression of human spindle mitosis arrest deficiency gene (hsMAD2)in spontaneous abortion embryos and the relationship between low expression of hsMAD2 and numerical chromosomal aberration. Methods Spontaneous abortion embryo tissues were collected,including 23 cases of once spontaneous abortion tissue and 10 cases of twice or more spontaneous abortion tissue and induced abortion embryos(35 cases)from the Department of Gynaecology and Obstetrics of the Affilisted Hospitals of Chongqing University of Medical Science during the period of March 2006 to March 2007.FQ-PCR and western blot were used to evaluate the endogenous expression level of hsMAD2 mRNA and hsMAD2 protein;primary culturing of cells from the induced abortion embryos was conducted and 5 embryonic cells were selected by chromosomes karyotype analysis.Recombinant shRNA plasmids targeting hsMAD2 gene were constructed to inhibit the expression of endogenous hsMAIY2 genes in embryonic cells which have normal karyotypes;the groups were defined as the first experimental group(transfeeted with pshRNA-hsMAD2-1),the second experimental group(transfected with pshRNA-hsMAD2-2),the third experimental group(transfected with pshRNA-hsMAD2-3),the first control group(transfected with nothing),the second control group(transfected with pTZU6+1)and the independent group(transfected with pshRNA-N1).Interference efficiency was demonstrated by FQ-PCR and western blot:cell prolireration was meagured by methyl thiazolyl tetrazolium(MTT)assay;cell-cycle was assessed by flow cytometry (FCM):the chromosome numbers were calculated to analyze the variation of chromosomes.Results(1) The mRNA levels of hsMAD2 in the once spontaneous abortion tissue,twice or more spontaneous abortion tissue and indueed abortion tissue were 0.00879±0.00035.0.00901±0.00033 and 0.00941±0.00026 respectively,and there Wag no significant ditierence(P＞0.05)compared with each other;however,the protein levels of hsMAD2 in three groups were 0.2791±0.0311.0.0431±0.0020 and 0.5790±0.0331 respectively,and there were significant difierences(P＜0.05)compared with each other.(2)Recombinant shRNA plagmids could significantly and specifically inhibit hsMAD2 gene expression in embryonic cells.Compared with the first control group(4%)and the second control group(3%),the recombinant shRNA could inhibit embryonic cell proliferation to 54% at 48 h after transfection(P＜0.05):compared with the first control group(8.2%)and the second control group(8.0%),the ratios of G2/M phase cells in the experimental group(17.9%)was significantly increased(P＜0.05);compared with the first control group (4.8%),the ratios of abnormal chromosomes in the experimental group was increased to 30.0%(P＜0.05).Conclusions Down-expression of hsMAD2 gene may be one of the mechanisms inducing numerical chromosome aberration,abnormal embryo development and the occurrence of spontaneous abortion.

The inhibitory effect of artesunate on human esophageal carcinoma is associated with modulation of CDC25A and TGFβ / 肿瘤

Objective: To observe the effects of artesunate (Art) on the proliferation of human esophageal carcinoma Eca109 cell line and xenograft tumors in nude mice, and explore the relationship between Art-induced cell cycle arrest and the expressions of CDC25A and transforming growth factor beta (TGFβ). Methods: The inhibitory effect of Art on proliferation of human esophageal carcinoma Eca109 cells and normal human peripheral blood mononuclear cells (hPBMC) at different concentrations were determined by MTT assay. The changes of cell cycle distribution of tumor cells were determined by flow cytometry (FCM). The inhibitory effects of Art on the growth of transplanted human esophageal carcinoma in nude mice was observed. The expression of CDC25A mRNA was examined by reverse transcriptase-polymerase chain reaction (RT-PCR). The protein expressions of CDC25A and TGFβ were measured by Western blotting. Results: Art significantly inhibited the proliferation of Eca109 cells with the IC50, of (68.80 ± 0.76) μmol/L. However it had no apparent effect on the proliferation of hPBMC. Art at low concentrations arrested the Eca109 cells at G0/G1 phase and significantly decreased the number of cells in S phase. Art at 100 μmol/L arrested Eca109 cells at G2/M phase. The volume and weight of xenograft tumors significantly were decreased in Art-treated group compared with model group. The maximal inhibitory ratio was 76.4% for tumor volume and 63.2% for tumor weight, respertively. Art dramatically inhibited mRNA and protein expression of CDC25A, while upregulated the expression of TGFβprotein. Conclusion: Art inhibits the proliferation of tumor cells with no apparent side effects. Art exerts its anti-tumor effect via modulating cell cycle distribution by upregulation of TGFβ and downregulation of CDC25A expression.

Expressions of heat shock protein 70,P27 and Bcl-2 protein in mucoepidermoid carcinoma of salivary glands and clinical significances / 吉林大学学报(医学版)

Objective To study the expressions of Hsp70,P27,Bcl-2 in mucoepidermoid carcinoma of salivary glands and clinical significances.Methods Forty cases of mucoepidermoid carcinoma of salivary glands were chosen and the expressions of Hsp70,P27,Bcl-2 were detected by the method of immunohistochemistry,10 cases of polymorphic adenoma were chosen as control group.Results Both Hsp70 and P27 were stained with aliomycin in nucleus and cytoplasm,the positive rates of them were 75% and 27.5%,respectively,there were significant differences compared with control group(P0.05).The expressions of Hsp70 and P27 had significant differences in different clinical stages,different histological grades and lymph node metastasis(P0.05).Conclusion The expressions of Hsp70 and P27 can be used to estimate prognosis of patients with mucoepidermoid carcinoma.Hsp70 and Bcl-2 can inhibit apoptosis by different ways.

Expression of c-kit and Cell Cycle Regulators in Non-small Cell Lung Carcinoma

BACKGROUND: The abnormal expression of c-kit is implicated in the pathogenesis of a variety of solid tumors. The Rb pathway and p53 act as cell cycle regulators. The purpose of this study was to assess the expression of c-kit, Rb, p53, p16 and cyclin D1 and their relationship to clinical and pathological parameters in patients with non-small cell lung carcinomas (NSCLC(s)). METHODS: Tissue microarrays consisting of 2 mm cores from the corresponding blocks were constructed from 54 NSCLC(s). Immunohistochemical staining for c-kit, Rb, p53, p16 and cyclin D1 was performed. C-kit immunostaining was considered positive if > or =10% of tumor cells were immunoreactive along the membrane and/or in cytoplasm. For Rb, p53, p16 and cyclin D1, tumor cells showing a nuclear staining pattern were interpreted as positive. RESULTS: We found that c-kit was expressed in 13 (24%) cases, Rb was lost in 39 (72%) cases, p53 was expressed in 28 (52%) cases, p16 was lost in 42 (78%) cases and cyclin D1 was expressed in 33 (61%) cases. The c-kit expression was significantly higher in adenocarcinoma (39%) than in squamous cell carcinoma (8%). We did not find any correlation between c-kit, Rb, p53, p16 and cyclin D1 expression and clinicopathological parameters such as: age, tumor size, lymph node involvement, disease stage and distant metastasis. There was a direct correlation between p53 expression and Rb loss. CONCLUSIONS: These results suggest that c-kit may be a useful therapeutic target for patients with c-kit positive tumors, and that the disruption of Rb and p53 pathways may play an important role in the development and progression of NSCLC(s).

Expression of Cell Cycle-Regulatory Proteins in Rectal Cancer: Significance of the Tumor Response to Preoperative Radiochemotherapy and for the Prognosis

BACKGROUND: Preoperative radiochemotherapy (PRCT) improves the outcomes for patients suffering with locally advanced rectal carcinoma, compared with surgery alone. However, there are no reliable factors predicting the survival and therapeutic benefits. METHODS: The cell-cycle regulatory proteins were investigated in the pretreatment biopsies from 68 patients who were suffering with rectal cancer by performing immunohistochemical studies of p53, p21, cyclin D1, Rb and p16 protein. The tumor response was graded on a three-scale grading system: no response (NR), partial remission (PR) and complete remission (CR). RESULTS: The tumors were positive for p53, p21 and cyclin D1 in 46 (67.6%), 32 (47.1%) and 14 (20.6%) cases, respectively. Abnormalities in Rb immunostaining were observed in 9 (13.2%) cases, while an abnormal p16 expression was noted in 59 (86.8%) tumors. Forty-two patients (61.8%) responded to PRCT: 18 (26.5%) cases achieved a CR and 24 (35.3%) cases achieved a PR. None of the above molecular markers were significantly associated with tumor response. However, the altered expression of p16 showed a significant correlation with overall survival (p=0.001). The high expression of p21 demonstrated a trend for longer survival (p=0.061). CONCLUSIONS: Of the cell-cycle regulatory proteins, p16 may be a valuable marker for to predict rectal cancer patients' survival; however, the role of each cell-cycle regulatory protein for the therapeutic benefits of PRCT needs to be further studied.