ABSTRACT
Objective To observe the effects of triptolide(TPL) on the anti-oncogene-APC gene of acute lymphoblastic leukemia cell line Jurkat in vitro. Methods The effects of TPL on proliferation Jurkat cells were assayed by using cell culture, MTT. The effects of TPL on APC gene of Jurkat cells were analyzed by nested methylation specific PCR and RT-PCR. The effects of TPL on the proteinum expression of APC gene were detected by Western blotting analysis. Results Following the treatment of TPL, the cell proliferation rate was degraded as the treatment concentration increased and the culture time extended. The effects were dose and time-dependent. The 48 hour IC50 was 19.7 ng/ml. TPL can reverse hypermethylation of APC gene,and induce the expression of the mRNA and the proteinum. Conclusion Low dose TPL could depress the proliferation rate of Jurkat. The possible mechanism might be its reversing the hypermethylation of APC gene and activiting the expression of APC gene.
ABSTRACT
Objective: To verify that COX-2 promoter can drive its downstream genes specifically in COX-2-positive o-varian cancer cells; Moreover, comparing with CMV promoter, analyze the transcript efficiency of COX-2 promoter. Methods: Contacting the recombinant plasmids named COX-2-BAX and CMV-Luc. After transient transfection liposome-mediated with the plasmids COX-2-Luc and CMV-Luc, respectively, the expression of Luciferase reporter gene was measured in COX-2-positive ovarian cancer cell line-SKOV3 and COX-2-negative colon cancer cell line-SW480. SKOV-3 and SW480 were transfected with COX-2-BAX and CMV-BAX in the same way, respectively. The apoptosis rates were measured through flow cytometry. Results: The recombinant plasmids named COX-2-BAX and CMV-Luc were constructed successfully. The expression efficiency of reporter gene was 1554 ? 86. 5 in SKOV3 and 53. 7 ? 10.9 in SW480 after 24 hours transfected with phPES2, 9851. 7 ? 129. 5 in SKOV3 and 8831. 0 ? 167. 3 in SW480 after 24 hours transfected with CMV-Luc in the same way. The apoptosis rate was 10.4% in SKOV3 and 3.7% in SW480 after transfected with COX-2-BAX, 21.7%in SKOV3 and 25. 6% in SW480 after 36 hours transfected with pcDNA3-BAX in the same way. Conclusions: COX-2 promoter can drive its downstream genes specifically in COX-2-positive ovarian cancer cell lines, but its expression efficiency wasmarkedly lower than CMV promoters. With proper modification, COX-2 promoter is expected to be useful in gene therapy of ovarian cancers.