554P Study on the differentially expressed miRNA of drug- contained serum of Bushen decoction on breast cancer bearing nude mice osteoblast by miRNA array

Abstract

Background This study aimed to investigate the effect on the target or related genes of miRNA after serum containing Bushen decoction intervention on breast cancer-bearing nude mice osteoblast cells, at the differential expression of miRNA profile levels. Methods The primary breast cancer-bearing nude mice osteoblasts were identified by alkaline phosphatase staining and collagen type I immunofluorescent staining and mineralized nodules staining. The osteoblasts were cultured in serum containing Bushen decoction. After a given time of intervention, the cultured cells were collected, and the total RNA were isolated and purified and used for follow-up miRNA microarray experiments. Real-time quantitative PCR was used to further validate the expression level of miRNA in samples of Bushen decoction intervention on breast cancer-bearing nude mice osteoblasts. Results The twice-enzyme digestion method was practicable, efficient and ideal technique to obtain a large number of purified primary breast cancer-bearing nude mice osteoblasts in a short time and culture cells with typical osteoblast characteristics. Alkaline phosphatase staining and collagen type I immunofluorescent staining and mineralized nodules staining were positive. Compared with the Bushen decoction containing serum cultured 1d, 229 different expression of miRNA (52 up-regulated and 177 down-regulated more than twice) were found 6 days after Bushen decoction containing serum cultured. The results of real-time PCR were in accordance with the miRNA array results that the expression levels of mmu-miR-205 was over 4-fold up- regulated and mmu-miR-335 over 4-fold down-regulated significantly in the cultured 6d compared with the cultured 1d. Conclusions A set of specific miRNA were obtained after Bushen decoction containing serum intervention on breast cancer-bearing nude mice osteoblast cells for 6d. Target gene prediction software predicts the up-regulated miRNA mainly by regulating bone development, bone morphogenesis, bone resorption and endochondral bone morphogenesis through the target genes, such as Runx2, Wnt, Notch, and Axin2, et al. Legal entity responsible for the study N/A Funding N/A Disclosure All authors have declared no conflicts of interest.