Effects Of Id1Knockdown By Small Interfering RNA On Cell Proliferation, Migration And Apoptosis Of Mouse Osteosarcoma Cell

ObjectiveTo detect the expression of Id1in K7M2-WT, MC3T3-E1, C3H10cellsand then investigate the effect of siRNA specific for Id1gene on theproliferation, migration and apoptosis of mouse osteosarcoma cell lineK7M2-WT, as well as the possible mechanisms, for providing a new targetfor genetic treatment of osteosarcoma.MethodsK7M2-WT、MC3T3-E1、C3H10cells thaw recovery, and incubated incomplete medium supplemented with10%fetal bovine serum at37℃in5%CO2．Logarithmic growth of cells，using Trizol lysate extracted totalRNA，the total RNA reverse transcription of cDNA，The expression of IdlmRNA in the three cell lines was detected by using Id1primers forRT-PCR．After extraction of total protein，the expression protein level of Id1gene in three kinds of cell lines was detected by Western Blot．The mouseosteosarcoma cell line K7M2-WT was infected with AdsimId1 (experimental group) and AdsiRNA (AdsiRNA group) respectively,untreated cells were set up as Control group. The mRNA and proteinexpression of Id1were detected by RT-PCR and Western Blot. Cellproliferation, migration, and apoptosis were detected by methylthiazolyltetrazolium (MTT) assay, wound closure assay(the scratch test) andtranswell, as well as DAPI staining was performed to detect apoptosis,respectively.Results1. Id1exhibited a high expression in C3H10and K7M2-WT cells.2. The mRNA and protein expression of Id1were down-regulated byadenovirus mediated siRNA specific for mouse Id1gene after3daysinfection (P<0.05).3. The MTT OD values of the Control group, AdsiRNA group andAdsimId1group were (0.52±0.032),(0.49±0.01),(0.22±0.0063),respectively. Compared with control group and AdsiRNA group, theinhibition rate of cells in the experimental group significantly decreasedby58%and55%(P<0.05).4. Wound healing assay showed that the migration capacity of K7M2-WTcells decreased with AdsimId1infection.5. The number of migratory cells in AdsimId1group (44.7±3.7) wassignificantly lower than that of the control group (100.3±3.8) orAdsiRNA group (103.8±4.4)(P<0.05). 6. The inhibition of Id1genes resulted in increased apoptosis.ConclusionsThe expression of Id1gene is highest in mesenchymal stem cells(C3H10)； The expression of Id1in K7M2-WT cells are higher thanMC3T3-E1cells. Silencing of Id1can effectively inhibit the proliferationand migration ability of mouse osteosarcoma cells and induced K7M2-WTcells apoptosis. Id1gene may play an important role to regulate thedevelopment of osteosarcoma. Our findings suggest that Id1gene may beprovide a new target for the genetic treatment of osteosarcoma.