ABSTRACT
OBJECTIVE@#To investigate the inhibitory effect of NF-kappaB p65siRNA on human laryngeal carcinoma xenograft model in nude mice.@*METHOD@#Human laryngeal carcinoma cell line Hep-2 was seeded in the subcutaneous layer of 15 nude mice to build laryngeal carcinoma xenograft model. Then they were randomly divided into three groups. NF-kappaB p65siRNA was given in siRNA group and FAM-Control siRNA was given in negative control group while phosphoric-buffered saline (PBS) was used in normal control group for 3 weeks. Tumor size and body weight of the mice were measured. TUNEL method and immunohistochemical S-P method were used for detecting the expression of NF-kappaB p65 and Bcl-xL protein.@*RESULT@#The volume of tumors in siRNA group was reduced and the average weight of tumors in siRNA group was lower than the other two groups (P < 0.05). In siRNA group, the expression of NF-kappaB p65 and Bcl-xL protein was down-regulated and the apoptotic rate was increased obviously as compared with the negative control group and the normal control group.@*CONCLUSION@#NF-kappaB p65siRNA can significantly inhibit the expression of NF-kappaB p65 and the growth of human laryngeal carcinoma xenograft in nude mice. Its mechanism may be related to inducing the apoptosis in tumor cells by down-regulating the expression of Bcl-xL protein.
Subject(s)
Animals , Female , Humans , Mice , Apoptosis , Carcinoma, Squamous Cell , Genetics , Pathology , Cell Line, Tumor , Gene Expression Regulation, Neoplastic , Laryngeal Neoplasms , Genetics , Pathology , Mice, Inbred BALB C , Mice, Nude , RNA, Small Interfering , Transcription Factor RelA , Genetics , Xenograft Model Antitumor Assays , bcl-X Protein , MetabolismABSTRACT
OBJECTIVE@#To detect the expression of c-myc in the tissue of laryngeal squamous cell carcinoma. RNA interference(RNAi) was employed to inhibit the expression of c-myc in Hep-2 cells and to evaluate the effects of c-myc as a target for gene therapy in laryngeal carcinoma.@*METHOD@#Immunohistochemistry was used to determine the protein levels of c-myc and Rb in 80 cases of laryngeal squamous cell carcinoma and 30 cases of polyp of vocal cord. Hep-2 cells were transfected with c-myc siRNA, c-myc protein and mRNA levels were detected using Western Blotting and RT-PCR. Cell viability was detected by MTT after the Hep-2 cells were transfected with c-myc siRNA for different times or transfected with different concentrations c-myc siRNA. The sensitivity of Hep-2 cells to 5-Fu transfected with or without c-myc siRNA was evaluated also by MTT. Hep-2 cells were transfected with c-myc siRNA in combination with 5-Fu for 48 h and then analyzed cell apoptosis by flow cytometry.@*RESULT@#Immunohistochemical analysis showed that c-myc was highly expressed in the tissues of laryngeal squamous cell carcinoma while the expression of Rb was lower. The protein and mRNA levels of c-myc decreased after transfected with c-myc siRNA. The results of MTT showed that the c-myc siRNA inhibited Hep-2 cells growth in a concentration-dependent manner. When transfected with c-myc siRNA(50 nmol/L), the cells were inhibited in a time-dependent manner. Compared with the untransfected cells, the viability of transfected Hep-2 cells was significantly suppressed at the same concentration of 5-Fu (P < 0.05). C-myc siRNA combination with 5-Fu could obviously increase cell apoptosis, even in the low concentration of 5-Fu (P < 0.05).@*CONCLUSION@#The protein level of C-myc has highly expressed in tumor tissues. C-myc siRNA can effectively inhibit the expression of c-myc and has anti-proliferation effects, increasing the sensitivity of Hep-2 cells to 5-Fu. Therefore,c-myc might be a good target for cancer treatment.
Subject(s)
Adult , Aged , Female , Humans , Male , Middle Aged , Apoptosis , Carcinoma, Squamous Cell , Metabolism , Pathology , Cell Line, Tumor , Cell Proliferation , Laryngeal Neoplasms , Metabolism , Pathology , Proto-Oncogene Proteins c-myc , Genetics , Metabolism , RNA Interference , RNA, Messenger , Genetics , RNA, Small Interfering , GeneticsABSTRACT
OBJECTIVE@#To study siRNA inhibited the expression of mRNA and protein of NF-kappaBp65 in the Hep -2 cell line.@*METHOD@#Hep-2 were transfected with p65SiRNA. Western blotting was used to examine the protein levels of NF-kappaBp65. RT-PCR method was adopted to determine the mRNA expression of NF-kappaBp65. MTT method was adopted to investigate the proliferation of the Hep-2 cells after the transfection of p65siRNA.@*RESULT@#The western blotting result showed that the level of NF-kappaBp65 protein was gradually declined after transfection of p65siRNA. The RT-PCR result showed that transfection with p65siRNA caused special degradation of the p65mRNA in Hep-2 cells at 24, 48 and 72 h. After transfection with p65siRNA.@*CONCLUSION@#p65siRNA has significant inhibition effects on the proliferation of the Hep-2 cells and expression of purpose gene mRNA and protein. The inhibition effects are time depended.