The Relationship between the Expression of p27^(Kip1),p53 and the Infiltration,Metastasis and Prognosis in Gastric Carcinoma

Objective: To study the relationship between the expression of p27Kip1 and p53, and the infiltration, metastasis and prognosis in gastric carcinoma. Methods: The expression of p27Kip1 and p53 at protein level was determined by immunohistochemical assay (two-step method) in 100 cases of gastric carcinoma. Results: Of the 100 cases, the positive rate of p27Kip1 and p53 expressions were 44% and 49%, respectively. In the group of gastric carcinomas with deep infiltration (infiltration group), lymph nodes metastasis group (metastasis group) and death-within-5-years group (death group), the expression of p27Kip1 was statistically lower (_P<0.05). In the metastasis group and death group, the expression of p53 was significantly higher (P<0.05). The results of the monovariate analysis revealed that the 5-year survival rate of the high p27Kip1 expression group was 70.59%, which is higher than those of the low p27Kip1 expression group (54.55%) and the negative experession group (26%). The 5-year survival rate of the high p53 expression group was 19.23%, which was lower than those of the p53 low expression group (43.75%) and the negative group (53.19%). Cox multivariate analysis showed that p27Kip1, like p53, was an independent prognostic index. But p27Klpl protein expression was a stronger independent survival predictor (RR=3.06) than p53 expression (RR=2.33). Conclusion: The low expression of p27Kip1 and the high expression of p53 reflected the more frequent invasion and metastasis, which resulted in the reduced survival of patients. As an independent markers of the gastric carcinoma, the expression of p27Kip1 is more useful than that of p53 in the prognosis prediction of gastric carcinoma.

Impact of p27mt gene on transplantation model of human colorectal cancer in nude mice

AIM:To investigate the inhibitory and anti-metastatic effect of mutant p27 gene(p27mt) on the growth of colorectal cancer xenografts in nude mice and its underlying mechanism.METHODS:Inhibitory effect of p27mt gene on the growth of colorectal cancer xenografts was determined by measurement of tumor size before and after direct intratumoral injection of Ad-p27mt in a preestablished transplantation model of human colorectal cancer in nude mice.Cell cycle and apoptosis were detected by flow cytometry performed on single-cell suspension from an isolated tumor.Expression of MMP-9 in tumor tissue was detected by immunohistochemistry.RESULTS:The average sizes of transplantation tumors were 1.94 ± 0.67 cm3,2.75 ± 0.83 cm3 and 3.01 ± 0.76 cm3 in the Ad-p27mt,Ad-LacZ and control groups,respectively(P < 0.05).The average proliferation rates were 37.34% ± 1.45%,53.16% ± 3.27% and 54.48% ± 2.43%,in the Ad-p27mt,Ad-LacZ and control groups,respectively(P < 0.05).The average apoptosis rates were 19.79% ± 3.32%,6.38% ± 4.91% and 7.25% ± 5.20% in the Ad-p27mt,Ad-LacZ and control groups,respectively(P < 0.01).The average MMP-9 expression rates were 20%,75% and 66.7% in the Ad-p27mt,Ad-LacZ and control groups,respectively(P < 0.01).CONCLUSION:p27mt inhibits the growth of transplanted tumor by blocking the proliferation of cancer xenografts and by promoting apoptosis of transplantated tumor cells,as well as decrease transpl-anted tumor metastasis.

Adenovirus expressing p27^(kip1) suppresses growth of established esophageal carcinoma xenografts

AIM： TO investigate the growth suppression of adenovirus expressing p27^kip1 on established esophageal tumors in nude mice. METHODS： Esophageal carcinoma xenografts in nude mice were established by tumor tissue mass transplantation. The successfully constructed recombinant adenoviral vectors carrying p2^kip1 gene （Ad- p27^%kip1） were directly injected into the esophageal tumors in nude mice. Compared to control group, the growth curve of tumor was drawn and the growth inhibition rate of tumor was calculated. The histology of tumors was examined by hematoxylin and eosin （H＆E） staining. The expression of p27^kip1 and survivin was detected in tumors by immunohistochemical technique. RESULTS： The growth of tumors in gene therapy group with Ad-p27^kip1 was obviously suppressed compared to control group （0.42±0.08 g vs 1.17±0.30 g, t=6.39, P〈0.01）, the inhibition rate of tumor growth reached 64.1%. Pathological detection showed that the tumors in nude mice were poorly differentiated esophageal squarnous carcinoma. In addition, the expression of p27^kip1 was increased, while the expression of survivin was decreased in tumors after being transfected with Ad-p27^kip1. CONCLUSION： p27^kip1 gene therapy mediated by adenovirus vector has a significant inhibitory effect on esophageal carcinoma in vivo. Up-regulated p27^kip1 expression and down-regulated survivin expression may be its important mechanisms.

Numerical simulation of avascular tumor growth based on p27 gene regulation

A multi-scale continuous-discrete model based on the effects of the p27 gene control is built to simulate the avascular tumor growth. At the tissue level, the continuous Eulerian model is adopted to determine the distribution of the concentration of oxygen, the extracellular matrix （ECM）, and the matrix-degradative enzyme （MDE）. At the cellular level, the discrete Lagrangien model is adopted to determine the movement, the proliferation, and the death of single tumor cells （TCs）. At the genetic level, whether a cell is committed to mitosis is determined by solving a set of equations modeling the effects of the p27 gene control. The avascular morphological evolution of the solid tumor growth is simulated, including the radius the oxygen distribution over time, and the expression. of the solid tumor, the number of the TCs, inhibiting effect＇ of the up-regulating p27 gene

Influence of human mutant p27 gene on the biological behaviors of colon cancer cells

Objective： To observe the influence of human mutant p27 gene （p27mt） on the growth and apoptosis of colon cancer cells so as to investigate the function mechanism of p27mt in gene therapy for colon cancer. Methods： Colon cancer cell line SW480 was infected with recombinant replication defective adenovirus Ad-p27mt, and expression of p27mt protein was detected by Western blot; the inhibition effect of p27mt on SW480 cells was detected with cytometry. Cell cycle was decided with flow cytometer, and DNA fragment analytic process identified the occurrence of apoptosis. Results： After transfected SW480 cells with Ad-p27mt, high expression of p27 protein was identified with immunoblotting assay. PI staining and flow cytometer assay showed 77.96% colon cancer cells was blocked in phase G0/G1, while in Ad-LacZ group and blank control group, 27.57% and 25.29% cells were blocked in the same phase, respectively. Growth curve showed Ad-p27mt has an obvious inhibition effect on the growth of SW480 cells, DNA fragment assay demonstrated that p27mt was able to induce the apoptosis of colon cancer cells. Conclusion： p27mt has an obvious blocking effect on colon cancer cell cycle, and most cells were blocked in phase G0/G1. This blockage is related with the growth inhibition and apoptosis induction effect of p27mt.

Effect of p27 Gene Combined with Pientzehuang (片仔癀) on Tumor Growth in Osteosarcoma-Bearing Nude Mice

Objective： To observe the effect of p27 gene recombinant adenovirus combined with Chinese medicine Pientzehuang （片仔癀） on the growth of xenografted human osteosarcoma in nude mice. Methods： Tissue transplantation was used to construct the orthotopic model of human osteosarcoma Saos-2 cell in nude mice. Thirty tumor-bearing nude mice were randomly divided into 5 groups with 6 mice in each group： blank control group （model of osteosarcoma）, empty vector group （recombinant adeno-associated virus-multiple cloning site）, Pientzehuang group, p27 gene group and combined treatment group （p27 gene combined with Pientzehuang）. The effect of combined treatment on human osteosarcoma was analyzed through the tumor formation, tumor volume and inhibition rate of tumor growth. The expression of p27 was measured by immunohistochemical staining and Western blot. Results： The orthotopic model of osteosarcoma in nude mice was successfully constructed. The general appearance of tumor-bearing nude mice in Pientzehuang and p27 gene groups was markedly improved compared with the blank control group; and in the combined treatment group it was significantly improved compared with the Pientzehuang and p27 gene groups. The tumor growth in the Pientzehuang and p27 gene groups was significantly inhibited compared with the blank control group （P〈0.05）; while in the combined treatment group it was markedly inhibited compared with the Pientzehuang and p27 gene groups （P〈0.05）. The rates of tumor growth inhibition were 34.1%, 56.5% and 63.8% in the Pientzehuang, p27 gene and combined treatment groups, respectively. Meanwhile, the protein expression of p27 gene in the p27 gene group was significantly increased compared with the blank control group （P〈0.05）; and it was significantly increased in the combined treatment group compared with the p27 gene and Pientzehuang groups （P〈0.05）. Conclusion： p27 gene introduced by adenovirus combined with Pientzehuang can inhibit the growth of human osteosarcoma cell Saos-2 in nude mice.

Effect of adenovirus-mediated p27 gene expression on the proliferation and apoptosis of HL-60 and Raji cell lines

Background p27 is an essential mediator of cell cycle control,which plays a key negative role in the proliferation and tumorigenesis of certain cell types. Here, we designed this study to explore the possible effects of p27 on the proliferation and apoptosis of HL-60 and Raji cell lines.Methods HL-60 and Raji cells were transfected with p27 via an adenovirus-mediated approach. The efficiency of Adp27 infection and the expression of p27 mRNA and protein were evaluated by X-gal staining, RT-PCR, and flow cytometry. The proliferation and apoptosis of HL-60 and Raji cells were estimated by means of trypan blue staining, MTT assay, Annexin V/PI, and DNA ladder electrophoresis. Results The infection efficiencies in HL-60 and Raji cells were 40.3% and 32.0%, respectively. RT-PCR and flow cytometry showed that there was significant expression of p27 mRNA and protein in HL-60 and Raji cells infected with Adp27; on the other hand, uninfected HL-60 cells showed faint traces of p27 mRNA and protein and Raji cells showed nearly no signs of p27 mRNA and protein. As demonstrated by a cell growth curve and by an MTT assay, strong time-dependent proliferation inhibition was apparent in HL-60 and Raji cells infected by Adp27. After 72 hours of infection, the Annexin V+/PI- apoptotic cell rates in HL-60 and Raji cell lines were 46.9% and 35.7%, respectively, significantly higher than in the control groups (4.7% and 5.6%, respectively). Typical DNA ladder bands were detectable in HL-60 and Raji cells after 48 hours of Adp27 infection. Conclusions Adenoviral vector-mediated p27 gene transfection of HL-60 and Raji cells leads to the inhibition of cellular proliferation and the promotion of cell apoptosis. This technique may provide an approach to gene therapy for leukemia or lymphoma.