PEI-PEG as a siRNA Genetic Vector Demonstrating Interference in the Expression of CD44v6 Protein in Gastric Cancer Cells

OBJECTIVE To investigate the effect of polyethylene imine glycol （PEI-PEG）/siRNA nanocomposites in the in vitro transfection of human gastric cancer SGC7901 cell lines and the down-regulation of gene expression of the adherence factor CD44v6. METHODS PEI-PEG/siRNA nanoparticles, in different N/P ratios, were synthesized and transfected into gastric cancer cells. Lipo2000/siRNA was used in the control group. The transfection efficiencies were observed under fluorescence microscope. The cytotoxicity of the nanoparticles was measured using the MTT assay （mononuclear cell direct cytotoxicity assay）, and the down-regulation effect of siRNA on CD44v6 gene was evaluated by Western blot. Based on the different N/P ratios, PEI-PEG/siRNA composites were synthesized and transfected into gastric cancer cells. Lipo2000/siRNA was used in the controls. The transfection efficiency was observed under fluorescence microscope. The cytotoxicity of the nanoparticles was measured using the MTT assay and the down-regulation effect of siRNA on CD44v6 gene was evaluated by Western blot. RESULTS After transfection, the transfection efficiency of the PEI-PEG/siRNA nanocomposites increased incrementally in N/P ratio value. The transfection efficiency improved with an increase in N/P ratio. When the N/P value was 15, fluorescence became more intense in the PEI-PEG/siRNA group than in the Lipo2000/siRNA group. At the same time, cell viability was （80.4 ± 5.6）% in the MTT reduction assay, which was similar to that in the Lipo2000/siRNA group. The results of Western blot analysis showed that the expression level of CD44v6 protein decreased to （59.7 ± 3.0）% after siRNA-CD44v6 was inhibited. CONCLUSION PEI-PEG could effectively form the nanocomposite in combination with siRNA, be transfected into the SGC7901 gastric cancer cell lines and inhibit CD44v6 protein expression. Moreover, as a genetic carrier, PEI-PEG copolymer has greater advantages, including high transfection e. ciency, less cytotoxicity and an easily alterable vector structure.

EXPRESSION OF INTRON 9 IN CD44 GENE IN CERVICAL CANCER AND CIN AND ITS CLINICAL SIGNIFICANCE

Objective： To detect the retention of intron 9 in CD44 mRNA in cervical cancer tissue, CIN, cervicitis and their exfoliated cells, and to study their clinical significance in diagnosis and treatment of early-stage, non-invasive cervical cancer, Methods： RT-PCR methods were used to detect the retention of intron 9 in CD44 mRNA in 30 cases of cervical cancer tissue, 11 cases of CIN tissue, 30 cases of cervicitis tissue and their exfoliated cells. Results： The retention rate of intron 9 in CD44 gene transcripts were 76.7% in cervical cancer tissue, 89,8% in corresponding exfoliated cells, 70.8% in CIN tissue, and 60.0% in CIN exfoliated cells, but undetected in neither cervicitis tissue nor exfoliated cells. The relative quantity of intron 9 in CD44 gene transcripts was 1.10 ± 0.12 in cervical cancer tissue, 1.21 ± 0.11 in CIN tissue, 1.11 ± 0.19 in cervical cancer exfoliated cells, 1.17 ± 0.12 in CIN exfoliated cells respectively, but undetected in neither cervicitis tissue nor exfoliated cells. The retention rate and relative content of intron 9 in CD44 gene transcripts in cervical cancer and CIN tissue and their exfoliated cells were statistically higher than that in cervicitis and their exfoliated cells （P〈0.005）. There was statistic difference comparing the retention rate of intron 9 in CD44 gene mRNA transcripts （70.8%） with positive rate in cytology （40.0%） in cervical cancer （x^2=5.78, P〈0.05）, but no statistic difference between the retention rate of intron 9 in CD44 gene transcripts （60.0%） with positive rate in cytology （36.4%） in CIN （x^2=0.585, P〉0.05）. Conclusion： Detecting the retention of intron 9 in CD44 mRNA in cervical exfoliated cells was more sensitivity than traditional cytology exam for diagnosing cervical cancer, and the techniques was worth clinical application.

Expression and Clinical Significance of CD44vin Hum an Gastric Cancers

Expression of CD44v mRNA in 20 patients with gastric tumor was investigated to identify the correlation of the CD44 gene splice and abnormal expression with the clinicopathological changes. The results showed that CD44v mRNA was expressed in 16 of 20 gastric cancers. In 5 out of the 10 mucosa tissues adjacent to gastric cancer, However, no expression of CD44v mRNA was found in all 10 normal mucosa specimens; Positive expression was found in 15 of 16 patients with lymph node metastasis, and 1 out of 4 patients without lymph node metastasis, as well as 5 cases with distant metastasis (P<0.05). Positive expression of CD44v was found in 2 out of 5 well differentiated type gastric cancers, and 14 out of 15 poorly differentiated gastric cancers. Differences were found in the expression between the two types of gastric mucosa (P<0.05). All the mucosa samples including normal gastric mucosa, gastric cancer and its adjacent tissues presented the standard CD44v mRNA. The conclusion was that the abnormal expression of CD44v is a universal significance in gastric cancer, which might serve as an early marker of gastric cancer and metastasis.

Bioinformatics research of CD44 and epithelial cell adhesion molecule related genes and pathways in colorectal cancer

Objective To investigate genes and involved biological processes closely associated with stem cell markers of colorectal cancer-epithelial cell adhesion molecule(EpCAM)+and CD44+.Methods By the bioinformatics method,with microarray data of colorectal cancer from gene expression omnibus(GEO)database and R2 platform,the genes significantly related with CD44 and Ep-