Icariin Promotes Osteogenic Differentiation Of Bone Marrow Stromal Cells And Osteoblasts Via PI3K/AKT-eNOS Pathway

Osteoporosis is a common disease that threatens the health of middle-aged and oldpeople greatly. With the continuing shift in population toward an older society, the incidenceof osteoporosis has been increasing steadily. Herba Epimedii from many species of genusEpimedium (family Berberidaceae) has been widely used as a tonic, aphrodisiac andantirheumatic in China. It also is one of the most frequently used herbs in treatingosteoporosis in traditional Chinese medicines. Icariin (ICA) is a flavonol glycoside isolatedfrom Epimedium sagittatum, which has been identified to own anti-osteoporosis activity.However, its mechanism is still unknown.To investigate the pathway mechanism of icariin, the effect of Icariin on the osteogenicdifferentiation of rat bone marrow stromal cells (rBMSCs) and osteoblasts（OB） wereinvestigated. The subcultured cells grown to about80%confluences were randomly dividedfour groups: ICA group (1×10-5mol·L-1), LY294002group (5×10-5mol·L-1), ICA+LY294002group and control group. The mRNA expression was detected by quantitativereal-time RT-PCR after24hours. The protein expression levels of p-AKT, iNOS and eNOSwere examined by Western blots. The activity of TNOS and inos as well as the NO contentswere measured by commercial kits.Our results were what follows next:1. The ICA group expression level of ALP and p-AKT were significantly improved(P<0.01), while LY294002strongly inhibited the increase of ALP expression level, anddecreased the p-AKT protein expression level. The results indicate that LY294002inhibitedosteogenic differentiation of bone marrow stromal cells by Icariin. The downstream target ofPI3K is AKT, Icariin promotes osteogenic differentiation of bone marrow stromal cells viaPI3K/AKT pathway.2. Icariin increased the expression levels of both eNOS and iNOS. LY294002inhibitedeNOS increasing, but had no obvious effect on iNOS. This results revealed the downstreamtarget of AKT is eNOS, Icariin promotes osteogenic differentiation of bone marrow stromalcells via PI3K/AKT-eNOS pathway.3. The change tendency of TNOS and NO content was similar with eNOS and ALP, theresults further indicate that Icariin promotes osteogenic differentiation of bone marrow stromal cells via PI3K/AKT-eNOS pathway.4. The change tendency of Osteoblast content was similar with bone marrow stromalwhich indicate that Icariin promotes osteogenic differentiation of osteoblasts via PI3K/AKT-eNOS pathway.The present data showed that Icariin promotes osteogenic differentiation of bone marrowstromal cells and Osteoblast differentiation via PI3K/AKT-eNOS pathway.