The Study Of Function Of Silencing Scinderin Gene In Osteosarcoma Saos2Cells

Objective1) Construct SCIN (Scinderin, SCIN) gene siRNA lentiviral vector, andtransfected osteosarcoma Saos2cells.2) Observe the impact of function of Saos2cells through silencing SCIN geneexpression in osteosarcoma Saos2cells.Methods1) SCIN-siRNA lentivirus vectors were constructed and then used to transfectthe Saos2osteosarcoma cells;2) Real time-PCR and western blotting were employed to assess the genesilencing efficacy of these recombinants;3) Saos2cells infected with siRNA-SCIN lentiviral vector (KD), blank control(NC), and transfected the lentiviral vector containing a nonsense sequence asa negative control (CON) proliferation assay using CCK-8;4) The migration and invasion potential of the three group cells were analyzedby the wound healing assay and matrigel invasion activities.Result1) After identification confirmation lentiviral vector was constructedsuccessfully, we transfected sarcoma Saos2cells with plasmid which carryingGFP green fluorescent protein gene. With fluorescence microscopy the transfected cells were visible the green fluorescent.The KD group and CONgroup,up to more than90%and85%, respectively with green fluorescentprove that the cells transfected with success.2) Real-time PCR and Western-blot experiments method to verify the silence theSCIN genes efficiency. The results showed the SCIN geng of thesiRNA-SCIN (KD) cells at the mRNA and protein levels compared with CONand NC group was significantly decreased, the difference was statisticallysignificant (P <0.05). The difference between the CON and NC two groupswas not statistically significant (P>0.05).3) By CCK-8growth curve observed silencing the SCIN genes，the Saos2cellproliferative capacity improved significantly compared with the controlgroup, with the control group, a significant difference (P <0.05) since thesecond day.4) The wound healing assay showed greatly increased the distance and thenumber of healing growth of the siRNA-SCIN (KD) cells group compariedwith the other CON and NC groups.the difference was statistically significantdifference (P <0.05and P <0.01).5) Transwell invasion test showed invasion rates of siRNA-SCIN (KD) groupwas significantly increased than the CON and NC groups (P<0.05). Thedifference between the CON and NC two groups was not statisticallysignificant (P>0.05).Conclusion1) SCIN-siRNA lentivirus vectors was constructed successfully and then used totransfect the Saos2osteosarcoma cells. The expression of SCIN gene ofSaos2cell transfection was significantly inhibited.2) The proliferative capacity and invasion rates were significantly increased bySilencing SCIN gene of osteosarcoma Saos2cells. It is suggested that SCINgene expression may decreased or lack of in the high incidence of distant metastasis in patients with osteosarcoma.Therefore, overexpressed theexpression of SCIN gene could become a new treatment method with whichfor osteosarcoma metastasis and relapse prevention is worth further study. ObjectiveTo compare clinical differences between children and adolescent osteosarcomapatients and identify the prognostic factors of the disease．Methods1) The clinical data of166osteosarcoma patients younger than19years wereretrospectively reviewed．The data obtained covered the period from October2003to March2012,and its prognosis were followed up for3to103months,an average of53.0months, divided into age children osteosarcoma (≤14yearsold) and youth (14to19years old) osteosarcoma groups, differences inclinical characteristics between the two groups were compared.TheKaplan—Meier method was used to measure the overall survival rate．2) χ2methods and Fisher exact methods were used to compare clinicaldifferences between children and adolescent patients．The Kaplan—Meiermethod was used to measure the overall survival rate．A log—rank univariateanalysis was used to determine the prognostic factors related to the survivalrate．The Cox model multivariate analysis was used to identify independentprognostic factors.3) The chi-square test the relationships between neoadjuvant chemotherapy withsurgery, neoadjuvant chemotherapy with lung metastasis, neoadjuvantchemotherapy with local recurrence, and surgical method with local recurrence.While observing the impact of prognosis about active treatment oflung metastasis.Results1) The median survival time of166patients in the present study was31.0months(95%CI：22.0to40.0)．The1、2and3-year cumulative survival rateswere90±2%、59±4%and45±4%，respectively．2) No significant difference between children and adolescent patients in terms ofsex，Enneking stage，KPS score，pathological fracture，tumor necrosis rate，histologic type，tumor location，frequency of adjuvant chemotherapy andtumor size was found．However, more child patients received amputationsurgery．3) Log-rank univariate analysis showed that the significant factors were the KPSscore，Enneking surgical staging, neoadjuvant chemotherapy, frequency ofadjuvant chemotherapy, local recurrence and occurrence of metastasis．4) Cox regression analysis revealed that the KPS score，frequency of adjuvantchemotherapy, whether associated with distant metastasis were independentprognostic factors.5) The chi-square test showed correlation between neoadjuvant chemotherapywith limb salvage rate, but neoadjuvant chemotherapy has not associated withlung metastasis and local recurrence. No correlation between the surgery withlocal recurrence; however actively dealing with lung metastases significantlyaffect survival (P=0.000).ConclusionThe clinical characteristics and survival rates of child and ado1escent withosteosarcoma in China were similar，The KPS score，frequency of adjuvantchemotherapy,and occurence of metastasis were the prognostic factors ofosteosarcoma．...