Proliferating cell nuclear antigen of Ulva prolifera is involved in the response to temperature stress

Ulva prolifera is the most common specie causative to green tide,and its growth is sensitive to temperature stress.However,the mechanisms of U.prolifera response to temperature stress remain elusive.In this study,high temperature(36℃)stimulus promoted the death of unformed cell wall protoplasts and delayed the division of formed cell wall protoplasts,while low-temperature(4℃)stimulus did not,suggesting that the mechanisms of the response of U.prolifera to high and low temperature stresses are different.Transcriptome results show that proliferation-related genes were differentially expressed under high and low-temperature stresses,especially the proliferating cell nuclear antigen(PCNA)and cyclins(CYCs).Subsequently,the interaction between PCNA and Cyclin A was confirmed by Co-immunoprecipitation,yeast two-hybrid,and so on.Furthermore,high-and low temperature stresses induced the expression of PCNA and Cyclin A in varying of degrees,and activated extracellular signal-regulated kinase(ERK)signal pathway.These results suggest,PCNA,Cyclin A,and ERK signal pathway played important roles in the resistance of U.prolifera to temperature stress.Interestingly,high-temperature stress induced an increase of miR-2916 in abundance,and exhibiting reverse expression of PCNA;and PCNA was target gene of miR-2916,suggesting that miR-2916 protected U.prolifera from high-temperature stress via post-transcriptionally regulation of PCNA.This study laid a foundation for understanding the function of PCNA and Cyclin A,moreover,it has a guiding significance to explore the mechanisms of the response to temperature stress from proliferation-related genes regulatory networks in U.prolifera.

Proliferating cell nuclear antigen clamp associated factor,a potential proto-oncogene with increased expression in malignant gastrointestinal tumors

Gastrointestinal(GI)cancers,including malignancies in the gastrointestinal tract and accessory organs of digestion,represent the leading cause of death worldwide due to the poor prognosis of most GI cancers.An investigation into the potential molecular targets of prediction,diagnosis,prognosis,and therapy in GI cancers is urgently required.Proliferating cell nuclear antigen(PCNA)clamp associated factor(PCLAF),which plays an essential role in cell proliferation,apoptosis,and cell cycle regulation by binding to PCNA,is a potential molecular target of GI cancers as it contributes to a series of malignant properties,including tumorigenesis,epithelial-mesenchymal transition,migration,and invasion.Furthermore,PCLAF is an underlying plasma prediction target in colorectal cancer and liver cancer.In addition to GI cancers,PCLAF is also involved in other types of cancers and autoimmune diseases.Several pivotal pathways,including the Rb/E2F pathway,NF-κB pathway,and p53-p21 cascade,are implicated in PCLAF-mediated diseases.PCLAF also contributes to some diseases through dysregulation of the p53 pathway,WNT signal pathway,MEK/ERK pathway,and PI3K/AKT/mTOR signal cascade.This review mainly describes in detail the role of PCLAF in physiological status and GI cancers.The signaling pathways involved in PCLAF are also summarized.Suppression of the interaction of PCLAF/PCNA or the expression of PCLAF might be potential biological therapeutic strategies for GI cancers.

Mast cell density and the context of clinicopathological parameters and expression of p185,estrogen receptor,and proliferating cell nuclear antigen in gastric carcinoma

AIM:To investigate the relationship between the mast cell density(MCD)and the context of clinicopathological parameters and expression of p185,estrogen receptor(ER), and proliferating cell nuclear antigen(PCNA)in gastric carcinoma. METHODS:Mast cell,p185,ER,and PCNA were detected using immunohistochemical S-P labeling method.Mast cell was counted in tissue of gastric carcinoma and regional lymph nodes respectively,and involved lymph nodes(ILN)were examined as usual. RESULTS:MCD was significantly related to both age and depth of penetration(x^2=4.688,P<0.05 for age and x^2=9.350, P<0.01 for depth of penetration)between MCD>21/0.03 mm^2 and MCD≤21/0.03 mm^2 in 100 patients;MCD in 1-6 ILN group patients was significantly higher than that in 7-15 ILN or>15 ILN group patients(u=6.881,8.055,P<0.01); There were significant differences intergroup in positive expression rate of p185,ER and PCNA between MCD>21/ 0.03 mm^2 and MCD≤21/0.03 mm^2 in 100 patients. CONCLUSION:Mast cell may have effect on inhibiting invasive growth of tumor,especially in the aged patients; The number of mast cells,in certain degree,may predicate the number of involved lymph nodes,which is valuable for assessment of prognosis;MCD was related to the expression of p185,ER,and PCNA in gastric carcinoma.It suggests that mast cell accumulation may inhibit the proliferation and the dissemination of the gastric carcinoma. INTRODUCTION Recently,many studies have reported on the association of mast cell with various tumorst.In several malignancies,mast cell has been found to correlate with growth,penetration and prognosis of tumor.Therefore,our study was undertaken to investigate the relationship between the mast cell density (MCD)and the context of clinicopathological parameters and expression of p 185,estrogen receptor(ER),and proliferating cell nuclear antigen(PCNA)in gastric carcinoma.

Clinical significance of expression of proliferating cell nuclear antigen and E-cadherin in gastric carcinoma

AIM to investigate the expression of proliferating cell nuclear antigen(p CNA)and E-cadherin in gastric carcinoma and to analyze their clinical significance.METHODS A total of 146 patients were selected for this study,including 38 patients with intestinal metaplasia,42with dysplasia,and 66 with primary gastric cancer.In addition,40 patients with normal gastric tissues were selected as controls.the expression of p CNA and E-cadherin was detected by immunohistochemistry.Differences in p CNA and the E-cadherin labeling indexes among normal gastric mucosa,intestinal metaplasia,dysplasia,and gastric carcinoma were compared.Subjects with normal gastric tissues were assigned to a normal group,while gastric cancer patients were assigned to a gastric cancer group.the difference in p CNA and E-cadherin expression between these two groups was compared.the relationship between expression of p CNA and E-cadherin and clinicopathological features was also explored in gastric cancer patients.furthermore,prognosis-related factors,as well as the expression of p CNA and E-cadherin,were analyzed in patients with gastric cancer to determine the 3-year survival of these patients.RESULTS the difference in p CNA and the E-cadherin labeling indexes among normal gastric mucosa,intestinal metaplasia,dysplasia,and gastric carcinoma was statistically significant(p<0.05).During the transition of normal gastric mucosa to gastric cancer,the p CNA labeling index gradually increased,while the E-cadherin labeling index gradually decreased(p<0.05).the p CNA labeling index was significantly higher and the E-cadherin labeling index was significantly lower in gastric cancer than in dysplasia(p<0.05).the expression of p CNA was significantly higher in the gastric cancer group than in the normal group,but E-cadherin was weaker(p<0.05).there was a negative correlation between the expression of p CNA and E-cadherin in gastric carcinoma(r=-0.741,p=0.000).p CNA expression differed significantly between gastric cancer patients with and without lymph node metastasis and between patients at different t stages.E-cadherin expression also differed significantly between gastric cancer patients with and without lymph node metastasis(p<0.05).High t stage and positive p CNA expression were risk factors for the prognosis of patients with gastric cancer(RR>1),while the positive expression of E-cadherin was a protective factor(RR<1).the sensitivity,specificity,and accuracy of p CNA positivity in predicting the 3-year survival of patients with gastric cancer were 93.33%,38.89%,and0.64,respectively;while these values for E-cadherin negativity were 80.0%,41.67%,and 0.59,respectively.When p CNA positivity and E-cadherin negativity were combined,the sensitivity,specificity,and accuracy were66.67%,66.67%,and 0.67,respectively.CONCLUSION Combined detection of p CNA and E-cadherin can improve the accuracy of assessing the prognosis of patients with gastric cancer.

Evaluation of germ-cell kinetics in infertile patients with proliferating cell nuclear antigen proliferating index

Aim: To explore the usefulness of proliferating cell nuclear antigen proliferating index (PCNA PI) in the pathologicaldiagnosis and treatment of male infertility. Methods: Testicular biopsy specimen obtained from 48 cases of male in-fertility and 2 normal controls were fixed and embedded. The sections were stained with anti-PCNA monoclonal anti-bodies or haematoxylin/eosin. Proliferating index (PI), expressed as the percentage of germ-cell nuclei positivelystained with PCNA antibody, was assessed from more than 20 seminiferous tubules or 600 germ-cells. Results: Theinfertile patients were divided into 4 groups: Group 1, normal spermatogenesis (14 cases); Group 2, hypospermato-genesis (16 cases); Group 3, germinal arrest (10 cases); Group 4, Sertoli cell only syndrome (8 cases). The PCNAPI of normal control testis was 86.5 % (mean value). Group 3 had a significantly lower PCNA PI (29.8 %) than nor-mal testis; Group 1 and 2 had similar Pis (82.3% and 82.3%, respectively) as the control testis. PI of the negativecontrol (Group 4) was 0 as no germ-cells were found. Conclusion: PCNA PI is useful for assessing germ-cell ki-netics, especially for pathological diagnosis of germinal arrest which is difficult to differentiate by routine HE stainingtechnique. In germinal arrest, there is a significantly lowered PCNA PI, which is an indication of DNA synthesis dete-rioration, suggesting the use of therapies be different from those for hypospermatogenesis. (Asian J Androl 2001 Mar;3: 63-66)

The Relationship of Expression of bcl-2, p53, and Proliferating Cell Nuclear Antigen (PCNA) to Cell Proliferation and Apoptosis in Renal Cell Carcinoma

To investigate the relationship of bcl-2, p53, proliferating cell nuclear antigen (PCNA) to cell proliferation, apoptosis and pathological parameters, the patterns of cell growth and turnover in renal cell carcinoma (RCC), formalin-fixed and paraffin-embedded tissue blocks from 34 patients with RCC were examined. Cell proliferation activity was detected by PCNA immunostaining and the proliferation index (PI) was expressed as a percentage of the PCNA-positive cells in the tumor cells. Apoptosis was detected by terminal deoxy- nucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), and the apoptotic index (AI) was expressed as a percentage of the TUNEL-positive cells in the tumor cells. Expressions of bcl-2 and p53 were assessed immunohistochemically. Our results showed that the PI ranged from 6.0 % to 24.0 % (median 12.3 %) and the AI from 2.0 % to 8.0 % (median 5.4 %) in RCC. The expression of the bcl-2 protein was demonstrated in 15 cases (44.1 %); the expression of the p53 protein, however, was seen in only 3 case. bcl-2 positivity was not associated with PI or AI or any pathological parameters. There were close associations between PI and tumor grade and stage, and a significant relationship between AI and the tumor grade of RCC. Our study suggests that bcl-2 positivity was not associated with PI or AI or any pathological parameters. There are close associations between PI and AI and tumor grade and stage of RCC. Active cell proliferation may be accompanied by frequent apoptosis in RCC.

Influence of norcantharidin on proliferation,proliferation-related gene proteins prolifera-ting cell nuclear antigen and Ki-67 of human gallbladder carcinoma GBC-SD cells

BACKGROUND: Gallbladder carcinoma is a highly lethal and aggressive disease with early metastasis, strong invasion and poor prognosis. Most patients with this disease are at the advanced and un-resectable stage and should be consi- dered for palliative treatment such as chemotherapy and ra- diotherapy. Unfortunately, reports of chemotherapy and radiotherapy for gallbladder carcinoma are disappointing. We investigated the influence of norcantharidin (NCTD) on proliferation, proliferation-related gene proteins PCNA and Ki-67 of human gallbladder carcinoma GBC-SD cells in vitro. METHODS: GBC-SD cell lines of human gallbladder carci- noma were cultured by the cell culture technique. The ex- periment was divided into NCTD group and control group. The tetrazolium-based colorimetric assay was used to evaluate cell growth. The streptavidin-biotin complex method was used to determine the expressions of prolifera- tion-related gene proteins PCNA and Ki-67 of human gall- bladder carcinoma GBC-SD cells. RESULTS: NCTD inhibited the growth and proliferation of GBC-SD cells from 10 mg/L or after 6 hours in a dose- and time-dependent manner, with the IC50 value of 56.18 μg/ ml at 48 hours. After treatment with NCTD, the expression of PCNA (0.932 ±0.031 vs. 0.318 ±0.023, P<0.001) and Ki-67 (0.964 ±0.092 vs. 0.297 ±0.018, P<0.001) proteins were decreased significantly. CONCLUSION: NCTD inhibits the proliferation of human gallbladder carcinoma GBC-SD cells in vitro and the expres- sion of their proliferation-related gene proteins PCNA and Ki-67.

The Relationship between Apoptosis and the Expression of Proliferating Cell Nuclear Antigen and the Clinical Stages in Gastric Carcinoma

The relationship between the apoptosis and the expression of proliferating cell nuclear antigen (PCNA) and the clinical stages in gastric cancers was studied. By using terminal deoxynucleotidyl transferase mediated nick end labelling (TUNEL) technique and PCNA immunohistochemical staining, the apoptosis and the expression of PCNA in tissue of gastric carcinoma were assayed in situ, the index of apoptosis (AI), index of PCNA (PI) and the rate of AI/PI were calculated. AI and PI in gastric cancer tissues were (6.5±3.7) % and (49.8±15.9) % respectively, and the rate of AI/PI was 0.13±0.05, which were obviously different from those of normal gastric mucosa in paragastric cancer ( P <0.01). With the advanced TNM stages of gastric carcinoma, the AI was decreased, PI was increased and the rate of AI/PI decreased in gastric carcinoma. There was significant difference in them between the gastric cancer tissues and normal gastric mucosa in pericarcinoma in TNM stage Ⅱ to Ⅳ ( P <0.05). It was suggested that the decreased apoptotic cells and the increased proliferating cells were obviously related to the tumor genesis and tumor progression in gastric carcinoma. The AI, PI and the rate of AI/PI would become the prognostic factors in advanced gastric carcinoma.

Cloning and characterization of proliferating cell nuclear antigen gene of Alexandrium catenella (Dinoflagellate) with respect to cell growth

Harmful algal blooms （HABs） have been affecting negatively the shellfish and aquaculture industries around the world. Though a lot of efforts have been made to disclose the changes of environmental factors involved and their effects on the HABs events, the molecular mechanism of this process remains unclear. To address this problem, proliferating cell nuclear antigen gene （pcna） was isolated and characterized from Alexandrium catenella. It showed high homology to those of other dinoflagellates （89% and 91% homology to Pfiesteria piscicid and Pyrocystis lunula, respectively）, and also 42%–43% homology to those of plant and animals. The expression level of pcna revealed by quantitative real time PCR was the lowest at the late lagging cell growth phase, increased to the highest at the late exponential phase, and then decreased at the stationary phase. Though the cell growth rate was also changing, no positive correlation between pcna expression level and cell growth rate was displayed throughout the whole cell growth stages （r 2 =0.024 6）. However, the pcna expression level had the similar trend with the change of cell growth rate throughout the whole growing process, e.g., from increasing at the earlier cell growth stage to decreasing at the following stages, though slightly lagging to the latter.

EXPRESSION OF P53 PROTEIN AND PROLIFERATING CELL NUCLEAR ANTIGEN IN HUMAN GESTATION TROPHOBLASTIC DISEASE

To study the relationship between p53 protein, proliferating cell nuclear antigen (PCNA) expression and benign or malignant gestational trophoblastic disease (MGTD). Methods: The histotomic sections of 48 patients with gestational trophoblastic disease and 24 patients of normal chorionic villi were stained using immunohistochemistry. The monoclonal antibodies were used to determine p53 protein and PCNA. Results: The frequency of p53 and PCNA positive expression were significantly different among the chorionic villi of normal pregnancy, hydratidiform mole (HM) and MGTD. But neither p53 nor PCNA has any relation with the clinical staging or metastasis of MGTD. Conclusion: Both P53 and PCNA are valuable in diagnosis of human gestational trophoblastic disease.

Neuron specific enolase,p53蛋白和proliferating-cell nuclear antigen在肺癌组织中的表达及意义

目的：研究神经元特异性烯醇化酶（neuronspecificenolase，NSE）的表达，与p53蛋白和增殖细胞核抗原（prolif－erating－cellnuclearantigen，PCNA）表达的关系及意义．方法：用特异性鼠抗人单克隆抗体，按LSAB免疫组织化学方法检测石蜡包埋的肺癌标本中NSE，p53蛋白和PCNA的表达．结果：在小细胞性肺癌（SmallCellLungCancer，SCLC）中，NSE阳性表达者，PCNA标记指数（LabellingIndex，LI）高于NSE阴性者，而p53蛋白的表达与NSE的表达无关．在非小细胞性肺癌（non－smallcelllungcancr，non－SCLC）中，p53蛋白的表达和PCNALI均与NSE的表达无关．结论：在SCLC的发生及发展过程中，神经内分泌功能可能起了部分作用．而p53抑癌基因在SCLC的发生中并不起着重要的作用．

Abnormal expressions of proliferating cell nuclear antigen and P27 protein in brain glioma

Both proliferating cell nuclear antigen and P27 protein are important factors to regulate cell cycle. While, the combination of them can provide exactly objective markers to evaluate prognosis of patients with brain glioma needs to be further studied based on pathological level. OBJECTIVE： To observe the expressions of proliferating cell nuclear antigen and P27 protein in both injured and normal brain glioma tissues and analyze the effect of them on onset and development of brain glioma. DESIGN： Case contrast observation. SETTING： Department of Neurosurgery, the Second Affiliated Hospital of Xi＇an Jiaotong University. PARTICIPANTS： A total of 63 patients with brain glioma were selected from Department of Neurosurgery, the Second Affiliated Hospital of Xi＇an Jiaotong University from July 1996 to June 2000. There were 38 males and 25 females and their ages ranged from 23 to 71 years. Based on pathological classification and grading standards of brain glioma, patients were divided into grade I - II （n=30） and grade III- IV （n = 33）. All cases received one operation but no radiotherapy and chemiotherapy before operation. Sample tissues were collected from tumor parenchyma. Non-neoplastic brain tissues were collected from another 12 non-tumor subjects who received craniocerebral trauma infra-decompression and regarded as the control group. There were l0 males and 2 females and their ages ranged from 16 to 54 years. The experiment had got confirmed consent from local ethic committee and the collection was provided confirmed consent from patients and their relatives. All samples were restained with HE staining so as to diagnose as the brain glioma. While, all patients with brain glioma received radiotherapy after operation and their survival periods were followed up. METHODS： Primary lesion wax of brain glioma was cut into serial sections and stained with S-P immunohistochemical staining. Brown substance which was observed in tumor nucleus was regarded as the positive expressions of both proliferating cell nuclear antigen and P27 protein. Automatic imaging analytic system was used to quantitatively analyze staining results of tumor. MAIN OUTCOME MEASURES： To compare the expressions of proliferating cell nuclear antigen and P27 protein in brain glioma tissues and non-tumor brain tissues and investigate the effect of various sexes, ages, survival periods and severities on the expressions of them in brain tissues. RESULTS： There was no significant difference of sexes and ages in the expressions of proliferating cell nuclear antigen and P27 protein （P 〉 0.05）; however, the expressions of proliferating cell nuclear antigen and P27 protein were milder in non-tumor brain tissues than those in the brain glioma tissues （P 〈 0.05）. Expression of proliferating cell nuclear antigen in brain tissue of grade III- IV severity was stronger than that of grade I - II severity, and the expression in ≥ 5-year survival periods were also stronger than that in 〈 5-year survival periods （P 〈 0.05）. In addition, expression of P27 protein in brain tissue of grade III- IV severity was stronger than that of grade I - II severity, and the expression in ≥ 5-year survival periods were also stronger than that in 〈 5-year survival periods （P 〈 0.05）. CONCLUSION： Abnormal expressions of proliferating cell nuclear antigen and P27 protein in human brain glioma are closely related to onset, development and prognosis of tumor.

Effects of UVB on the expression of proliferating cell nuclear antigen (PCNA) in dinoflagellate Prorocentrum donghaiense Lu

PCNA has been considered as a useful marker for the estimation of growth rates of marine phytoplankton at the species level. Since dinoflagellates are noted for having many prokaryotic features in that they are the only eukaryotes to have permanently condensed chromosomes as well as lacking histones and a nucleosome, the sensitivity to UVB radiation and the validity of PCNA as a maker of growth rate in dinoflagellate need to be evaluated. Prorocentrum donghaiense Lu was investigated to valuate the UVB sensitivity in relation to cellular and molecular aspects of PCNA as a growth indicator. The effects of UVB radiation on PCNA were studied using the methods of western blots technology and whole-cell immunoflurescence with one mono-antibody. UVB changed the cell morphology, halted the growth and increased the cell size, even caused cell death to a certain extent after treatment with UVB radiation in P. donghaiense. Compared with the control, treating the algal cultures in exponential phases with UVB radiation for 24 h caused chromatin release and increases in protein levels of PCNA.

The Effect of the LysoPC-induced Endothelial Cell Conditioned Medium on Proliferating Cell Nuclear Antigen Expression of the Calf Thoracic Aorta Smooth Muscle Cells

In order to study the effect of and mechanism of lysophosphatidylcholine (LysoPC) on proliferation of the calf thoracic aorta smooth muscle cells (ASMCs), the ASMCs were used to observe the effects of LysoPC induced endothelial cell conditioned medium on the DNA content and proliferating cell nuclear antigen (PCNA) expression in the calf thoracic ASMCs by flow cytometry and Western Blot technique. It was found that LysoPC induced endothelial cell conditioned medium could significantly promote PCNA expression of the calf ASMCs, induce the converting of ASMCs from G 0 /G 1 phase to S phase of DNA synthesis, and increase the tyrosine phosphorylation protein expression. Tyrosine protein kinase inhibitor (TPKi) RG50864 could obviously inhibit proliferation of LysoPC induced ASMCs in a dose dependence manner. The results indicated that the effect of LysoPC promoting the proliferation of ASMCs is partly evoked by endothelial cell derived growth factors such as PDGF and so on.

Evaluation of proliferation kinetics in large intestinal neoplasms:Comparative study on the expression of proliferating cell nuclear antigen and AgNOR counting

Expression of proliferating cell nuclear antigen (PCNA) in human colorectal lesions was studied immunohistologically and compared with the results of silver stained nucleolar organized regions (AgNOR)counting for evaluating the changes of proliferative kinetics in adenoma-carcinoma sequence of the large intestine.Ninety-six formalin-fixed paraffin-embedded specimens were used, which consisted of 21 normal colon mucosa,23 adenocarcinomas and 52 adenomas (34 cases with mild atypia, 12 morderate and 6 severe). Three types of PCNA distribution pattern were identified. 73. 9% normal mucosa was type A pattern. Type C pattern was characteristic of adenocarcinoma, and adenoma was of type B or C pattern. Labeling index (LI) of PCNA was an important parameter to evaluate the proliferation activity. In normal mucosa, LI of PCNA was 0. 34±0. 10, while in malignant lesions, it was 0. 71±0. 09. Furthermore, the LI had a tendency to elevate with the degree of atypia in adenoma from 0. 52±0. 10 in mild atypia to 0. 68±0. 07 in severe atypia. The pattern of AgNOR counting changes, which were 1. 97±0. 36 and 4. 16±1. 24 in the normal mucosa and malignant mucosa respectively, was found to be similar to Ll changes of the PCNA in different lesions of the same section. A good correlation was found between the mean number of AgNOR per nucleus and the LI of PCNA in 23 adenocarcinomas (r=0. 59, P <0. 05). These results suggested that proliferation kinetics could be comprehensively evaluated by combining immunohistological staining for PCNA and a simple silver staining for NOR, which would be advantageous for the diagnosis of colorectal lesions and for evaluating the degree of histopathological atypia of adenomas.

The effect of Chinese herbal medicine“heche assisted pregnancy recipe”on endometrial estrogen and progesterone receptor,proliferating cell nuclear antigen and vascular endothelial growth factor in the patients with infertility

Objectives:To investigate the effect of Chinese herbal medicine"heche assisted preg-nancy recipe (HCAPR)" on estrogen receptor(ER), progesterone receptor (PR), pro-lifierating cell nuclear antigen(PCNA) and vascular endothelial growth factor (VEGF)in endometrium of infertile women.Methods: The S-P immunohistochemical assay was used to observe expression ofER, PR , PCNA and VEGF in late proliferative phase before and after the HCAPR treat-ment.Results: After the treatment, the expression of ER,PR,PCNA and VEGF in nucleiof glandular epithelium and stromal cells was significantly stronger (all P＜0. 001) re-spectively than that before treatment , especially the expression of PCNA and VEGF.Conclusions: These results suggest that traditional Chinese medicine HCAPR oftonifying kidney and regulating menstruation increased the synthesis of ER,PR, PCNAand VEGF, which may promote normal growth and development of the endometrium ,improve the micro-environment of the endometrium, and enhance uterine receptivity.The evidence may provide theoretical basis for therapy infertility with Chinese herbalmedicine.

Effect of aloe-emodin on expression of proliferating cell nuclear antigen of vascular smooth muscle cells in culture after arterial injury

Objective To observe the effect of aloe-emodin on the expression of proliferating cell nuclear antigen (PCNA) in smooth muscle cells (SMCs) after arterial injury and study the molecular mechanism of inhibition of aloe-emodin on SMC proliferation.Methods Deendothelialization was performed at the abdominal aorta in Japanese white rabbits using a 3F Fogarty arterial embolectomy catheter. 48 hours later, the medium of abdominal aorta was isolated and primary SMCs culture was performed. Cells were synchronized to G0 by serum starvation, then aloe-emodin at a concentration of 20?μg/ml was added to the culture medium containing 10%［v/v］ fetal calf serum. Vehicle was also added to the medium as a control. After 18 hours, the expression of PCNA at the level of mRNA and protein were examined using techniques of RT/PCR, Western blotting and inmmunocytochemistry respectively. Results Compared with the control group, the expression of PCNA mRNA and protein was prominently decreased after addition of aloe-emodin. Conclusion The inhibition of aloe-emodin on SMCs proliferation may be caused by inhibiting the expression of the PCNA gene.

Bladder cancer therapy using combined proliferating cell nuclear antigen antisense oligonucleotides and recombinant adenovirus p53

Objective To evaluate the antitumor efficacy of proliferating cell nuclear antigen antisense oligonucleotide (PCNA-ASO) in combination with recombinant adenovirus p53 (Ad-p53) against bladder cancer EJ and BIU-87 cells in vitro and in vivo. Methods Cells were transfected with Ad-p53 (100 MOI), and PCNA-ASO (1.6 μmol/L) was then introduced into the cells using a cationic lipid (lipofectamine, 20 μl/ml). In vitro and in vivo antitumor effects of combining PCNA-ASO with Ad-p53 were measured using the MTT assay, flow cytometry, clone formation, and a nude mice model. Results The combination of PCNA-ASO and Ad-p53 inhibited cell viability in both the EJ (89.3%) and BIU-87 (78.6%) cell lines. The ability of the cells to form foci was also reduced by 74.8% in EJ cells and by 67.5% in BIU-87 cells (P<0.01). A significant decrease of cells in the S phase (11.4% in EJ cells, 14.6% in BIU-87 cells) and a significant increase of cells in G1 phase (62.2% in EJ, 56.8% in BIU-87) were noted. The mean tumor volume after 7 days of treatment with PCNA-ASO or Ad-p53 in combination decreased to 47.6% or 36.4% of the initial tumor size in the two cell lines respectively. Conclusion These results indicate that combined PCNA-ASO and Ad-p53 in the treatment of bladder cancer with mutant p53 has important therapeutic potential, significantly suppressing the growth of human bladder cancer both in vitro and in vivo.

The Influence of the Total Flavonoids of Hedysarum Polybotry on the Proliferation,Cell Cycle,and Expressions of p21^(Ras) and Proliferating Cell Nuclear Antigen Gene in Erythroleukemia Cell Line K562

Objective： To investigate the effect of total flavonoids of Hedysarum polybotry on the proliferation, cell cycle, and expressions of p21Ras and proliferating cell nuclear antigen （PCNA） gene in erythroleukemia cell line K562. Methods： The effect of total flavonoids of Hedysarum po/ybotry on K562 cell line survival was determined by the 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium （MTT） reduction assay. The time- and dose- dependent manner was also observed. The cell cycle and apoptosis were analyzed with flow cytometry （FCM）. The immunocytochemistry method was applied to quantitatively analyze the effects of flavonoids of Hedysarum polybotry on changes p21Ras and PCNA gene expressions. Results： Flavonoids of Hedysarum polybotry （20-100 g/mL） significantly inhibited the proliferation of K562 cells in a time- and dose-dependent manner. After K562 cells were cultured for 48 h, total flavonoids of Hedysarum polybotry had no significant effect on the apoptosis of K562 cells but showed significantly inhibition （P〈0.01）, indicating that total flavonoids of Hedysarum polybotry could induce K562 cells arrested at Go/G1 and G2/M phases. Compared with the control group, p21Ras and PCNA gene expressions were decreased significantly in K562 cells treated with total flavonoids of Hedysarum polybotry （40 and 80 μg/mL, respectively） for 48 h. Conclusion： The inhibitory effect on proliferation of K562 cells was observed in the groups treated with flavonoids of Hedysarum polybotry, which might be related to cells arresting.

RELATIONSHIP BETWEEN PROLIFERATING CELL NUCLEARANTIGEN EXPRESSION AND ITS MALIGNANCY POTENTIAL IN COLORECTAL CARCINOMA

Objective: To study the relationship between proliferating cell nuclear antigen expression and its malignancy potential in colorectal carcinoma. Methods: Paraffin sections of 86 patients with advanced colorectal carcinoma were assessed by immunohistochemical study, using a mouse monoclonal antibody (pc-10, DAKO Co. USA) to check proliferating cell nuclear antigen (PCNA). To compare PCNA with conventional clinicopathologic factor, including p53 overexpression, tissue carcinoembnyonic antigen immunoreactivity pattern and flow cytometric DNA ploidy for assessing tumor malignancy potential. In addition, recurrence and survival of patients with advanced colorectal carcinoma after curative resection were analyzed in accordance with degree of PCNA expression. Results: PCNA-labeling index (PCNA-LI) increased significantly as the tumor stage advanced (p=0.0001). Strong correlations were observed between PCNA-LI and various pathologic parameters, including histologic differentiation (P=0.0027), lymphatic invasion (P=0.0001), vascular invasion (P=0.0001), lymph node metastasis (P=0.0001), and liver metastasis (P=0.0036). Mean PCNA-LI was also significantly higher in tumor with DNA aneuploidy (P=0.0006) and negative (P=0.01). Linear relationships were demonstrated between PCNA-LI and clinical outcomes; Recurrence rate was significantly greater in the group with higher than the mean PCNA-LI, who underwent curative resection (P<0.01), and three-year survival rates for curative cases with higher than the mean PCNA-LI were significantly poorer than those with lower than mean PCNA-LI (P<0.005). Conclusion: There were correlations between PCNA-LI and various pathologic parameters, PCNA-LI increased significantly as the tumor stage advanced in colorectal carcinoma, the rates of recurrence and death got higher as PCNA-LI increased after curative resection for colorectal carcinoma.