using oigo dT from isoted RN smpesDN ws then used s temptes for poymersechin rection PCR. The PCR coitionsthe sequences of primers re isted in Tbe for the genes encoding Paeonol the foowing proteins:kine phosphtse, corebiing factor , bone sioprotein, peroxisome proifertor ctivted receptor gmm PPR ,gycerdehydephosphte dehydrogense GPDH. The PCR products were resoved bygrose ge eectrophoresisvisuized with ethidium bromide. The retive expression of ech gene ws quntifed by densitometry using the Ge Imge Ver.System Tinon, Shnghi, Chinnormized ginst GPDH. Immunobotting studies Ces were ysed using the RIP ysis buffer contining mM Tris HC p, mM NC,sodium deoxychote,NP mM phenymethysufony fuoride,mM EDT.

ERK, phosphoERK, p, phosphopGDPH ntibodies were used Boster Bioogic Technoogy, Wuhn, Chi Imges were nyzed with Quntity One softwre BioRd, Hercues, Cb intensity ws quntifiednormized ginst GDPH. Sttistic nysis The dt were expressed s menstrd devition s.d. for ee or more iepeent experiments. Sttistic significnce ws estimted by onewy NOV with Bonferroni’s post test mutipe prisons,Student NewmnKeus test prisons between two groups ws used where pproprite. PNS Promote Osteogenic Differentition Ce Physio Tacrolimus Biochem ;: B C DPnx notoginseng sponins potentited the osteogenesis of bone mrrow strom ces. Bone mrrow strom ces were treted with Pnx notoginseng sponins tor μgm fordys,ce vibiity ws ssessed by the MTT test. Dt is expressed s menSDthe experiments were done in qudrupicteB Bone mrrow strom ces were treted with Pnx notoginseng sponins tor μg m for dys. Mineriztion of bone mrrow strom ces ws quntified by izrin red S ssys. Dt in C represents menSD of experiments done in pentpicte. Versus OIM group; b . Versus μ gm group;

Versus μgm group.D Pnx notoginseng sponins enhnced kine phosphtse ctivities of bone mrrow strom ces. kine phosphtse ctivities were mesured by convention methodthe dt represent the menSD in qudrupicte . versus bone mrrow strom ces uergoing simpe osteogenic iuction; b. versus the μgm group; c. versus the μgm group; d. versus the μgm group. Resuts Pnx notoginseng sponins promote the proifertionmineriztion of bone mrrow strom ces We isoted primry bone mrrow strom ces from the femortibi of rtscuture them uer osteogenic coitions. We first investigted whether Pnx notoginseng sponins coud promote the proifertion of bone mrrow strom ces in vitro . Our MTT ssys showed tht, t one week fter the strt of incubtion ,μgm Pnx notoginseng sponins mrkedy incresed the proifertion of bone mrrow strom ces pred with controCe Physio Biochem EPnx masitinib fak inhibitor  notoginseng sponins potentited osteogenesis of bone mrrow strom ces vi the ERKp signing pthwys. izrin red S ssys showed tht PD, but not SP, inhibited Pnx notoginseng sponinspotentited mineriztion of bone mrrow strom ces. B The content of ccium deposit ws quntifiedsttisticy nyzed .

versus bone mrrow strom ces uergoing simpe osteogenic iuction; b . versus μgm group . versus μgm group.C PD, but not SP, inhibited Pnx notoginseng sponins enhnced kine phosphtse ctivity of bone mrrow strom ces. versus OIM ; b . versus μgm PNS group; c no significnt difference versus μgm PNS group; d . versus μgm PNS group.D The expression of genes invoved in osteogenesis of bone mrrow strom ces ws quntified by RTPCR with or without Pnx notoginseng sponins in the bsence or presence of PD orfordys. E masitinib VEGFR-PDGFR inhibitor Densitometric nysis of the genes described in D. . versus bone mrrow strom ces uergoing simpe osteogenic iuction; b . versus the μg m group; c . versus μgm PNS groupesWe further studied whether Pnx notoginseng sponins coud stimute the mineriztion of bone mrrow strom ces by exmining these ces using izrin red S ssys. We fou tht, ee weeks fter the strt of incubtion, bone mrrow strom ces growing in osteogenic medium showed pprent mineriztion .