Transcatheter arterial chemoembolization enhances expression of Nm23-H1 and TIMP-2 in the tumor tissue of patients with hepatocellular carcinoma.

BACKGROUND/AIMS: To investigate the effects of transcatheter arterial chemoembolization (TACE) on expression of nm23-H1 and TIMP-2 in the tumor tissue and prognosis of patients with hepatocellular carcinoma (HCC). METHODOLOGY: Seventy-two patients with resectable HCC were randomized into two equal groups with 36 patients in each: TACE before surgical resection of HCC (Group A) and direct surgical resection of HCC (Group B). All samples were subjected to pathological examination and immunohistochemical staining using nm23-H1 and TIMP-2 antibodies. Expression level and distribution of nm23-H1 and TIMP-2 in tumor and adjacent tissue were assessed. Extrahepatic metastasis and survival time of patients in both groups were evaluated through 36 months follow-up. RESULTS: Immunohistochemical analysis showed that the tumor tissues from patients in Group A had a higher positive expression of nm23-H1 than Group B (chi2=15.52, p<0.01). Group A also showed a higher positive expression of TIMP-2 than Group B (chi2=9.00, p<0.05). Patients in Group A had a longer mean survival time (36 vs. 28 months in Groups A and B, respectively) and higher survival rate (chi2=5.734, p=0.017). CONCLUSIONS: Preoperative TACE enhances the expression of metastasis suppressors nm23-H1 and TIMP-2, and may potentially inhibit metastasis of HCC and increase the survival time of patients with the resectable HCC.

[An infrared imaging system for detecting electrophoretic mobility shift of DNA-protein complexes].

OBJECTIVE: To establish a new non-radioactive method for electrophoretic mobility shift assay (EMSA) to investigate the binding between glucocorticoid induced leucine zipper (GILZ) and peroxisome proliferator-activated receptor-gamma 2 (PPARgamma2) promoter oligonucleotides. METHODS: GILZ protein prepared by prokaryotic expression was linked to PPARgamma2 promoter oligonucleotides end-labeled with IRDye 800 infrared dye. The DNA-protein complex was separated with non-denatured polyacrylamide gel and scanned with the Odyssey. Infrared Imaging System. RESULTS: One lane of DNA-protein complex was clearly presented, and the signal intensity increased along with the increment of the protein load. CONCLUSION: This infrared imaging system can be used for EMSA for detecting the DNA-protein complex with high sensitivity efficiency and allows easy operation.

[Construction of a recombinant adenoviral vector carrying integrin-alpha v and the influence of integrin-alpha on the adhesion characteristics of SMMC-7721 cells].

OBJECTIVE: To establish human integrin-alpha v adenovirus expressed in human liver cancer SMMC-7721 cells and analyze the characteristics of integrin-alpha adhesion to liver cancer cells. METHODS: Human integrin-alpha v cDNA was cloned into the plasmid pAdTrack-CMV. Then, pAdTrack-integrin-alpha v was cotransformed with adenoviral backbone vector into E. coli strain BJ5183. The recombinant adenoviral plasmid was identified by restriction analysis with Pac I and transfected into HEK-293A cells to package and amplify recombinant adenovirus particles. Western blot showed that integrin-alpha v gene was exactly transcript and expressed in SMMC-7721 cells. Twenty four hours after transfection, the effect of integrin-alpha on adhesion of SMMC-7721 cells was detected by adhesion experiment. RESULTS: Recombinant adenovirus vector of integrin-alpha v gene was successfully constructed and high titers of recombinant adenovirus was obtained. Adhesion cell count was significantly higher in integrin-alpha v transfected cells than in untransfected and Ad-null-transfected cells (P<0.01). CONCLUSION: Over expression of integrin-alpha v promotes adhesion of SMMC-7721 cells, is an important molecular mechanism of tumor metastasis.

[Construction of GFP-AWP1 fusion gene vector and its expression in 293 cells].

OBJECTIVE: To construct green fluorescent protein (GFP)-AWP1 (a novel human protein associated with protein kinase C-related kinase 1) fusion gene vector for observing the expression and localization of AWP1 in 293 cells. METHODS: The coding region in AWP1 cDNA was amplified by RT-PCR from human endothelial cell line ECV304 and recombined into pEGFP-C2 plasmid expressing GFP. After identification with restriction endonucleases and sequence analysis, the recombinant plasmid was transfected into 293 cells with the cationic liposome DOTAP as the transfection reagent. The expression and localization of AWP1 were observed under a fluorescence microscope. RESULTS: Restriction endonuclease assay and sequence analysis verified the successful construction of the recombinant vector pEGFP-C2/AWP1, and GFP-AWP1 fusion protein was highly efficiently expressed in 293 cells. Under fluorescent microscope, green fluorescence was seen homogeneously distributed in the entire cell body of the cells transfected by the empty vector pEGFP-C2, but diffusely in the cytoplasm of the cells transfected by the recombinant vector pEGFP-C2/AWP1. CONCLUSION: GFP-AWP1 fusion gene vector is successfully constructed and the fusion protein expressed in the cytoplasm of 293 cells.

[Effect of astrocyte-conditioned medium on tert-butyl hydroperoxide-induced apoptosis of PC12 cells].

OBJECTIVE: To investigate the effect of astrocyte-conditioned medium (ACM) on PC12 cell apoptosis induced by reactive oxygen species tert-butyl hydroperoxide (tbOOH). METHODS: PC12 cells attacked by tbOOH were cultured with Sprague Dawley rat cerebral cortex ACM, and the cell apoptosis was observed by fluorescent microscopy and transmission electron microscopy. The cell apoptosis rate was evaluated by flow cytometry and malondialdehyde (MDA) concentration measured using thiobarbituric acid (TBA)-reacting substances (TRABS). RESULT: TbOOH induced PC12 cell apoptosis, and ACM significantly decreased the apoptosis rate and MDA concentration in tbOOH-treated cells. CONCLUSION: ACM may increase the anti-oxidation capacity of PC12 cells and inhibit cell apoptosis induced by tbOOH.