Experimental Research Of Wtp53and Rb94Gene Transfected Into The Retinoblastoma Of Nude Mice By Ultrasound-targeted Microbubble Destruction

Background and Objective Retinoblastoma (RB) is the mostcommon malignant intraocular tumor that occurs in preschool children.When the doctor found more children has entered into the serious harm tovision and life. The treatment of our country is ophthalmectomy,complementary with radiation therapy, chemotherapy, cryotherapy andother, et al. The gene therapy, especially the research on efficiency andsecurity vector of targeted gene, becomes a main research focus for it canfundamentally solve the problem. In recent years，ultrasound microbubblebecame a new research objects of gene transporter.The Rb gene isrecognized the most clost relations with tumor suppressor genes to the RBform, while at present P53gene is recognized a tumor-suppressor geneswith the highest correlation to the human malignant tumor. In our study, theRB as the research object,and observe the inhibition of wtp53and Rb94gene into transfected the retinoblastoma of cell and nude mices by a kind ofneoteric transporters which named ultrasound microbubble. Methods1.HXO-Rb44were cultured in RPMI-1640Mediumcontaining10%FBS.When they grow to logarithmic period, collect andadjust the cell density for (5.5~6.5) x106/mL.40nude mice weretransplanted with human RB cell(HXO-Rb44) into eyes by subretinalinjection,observed the tumor growth each three days.2.After the animalmodle done successfully,mice were randomly divided into five groups:①Blank control group;②blank plasmid group；③wtp53plasmid group；④Rb94plasmid group；⑤wtp53+Rb94plasmid group，each groupseyeballs were exposed to ultrasound with intensity of0.5W/cm2immlediately，and the time were all60s that working time control at20％．Removed the tumor tissue in the seven day，the expression of wtp53and Rb94gene was detection by RT-PCR and Western-blot.3.Theexpression of VEGF were observed by immunohistochemical staining andWestern-blot. The tumor tissue of MVD was detected byimmunohistochemical. TUNEL method was used to analyze the apoptosisof retinoblastoma cells.Results1.40nude mice was injected the HXO-Rb44cell by subretinalinjection,and32nude mice was successful.2.The single gene groups andjoint group were detected gene expression by RT-PCR. WB can detect thesingle gene groups and joint group of wtp53and Rb94protein expression.3.Detecting every group of tumor tissue,we found single gene groups andjoint gene groups have an effect on VEGF, MVD and the apoptosis, but the joint group has more effective than the single gene groups, and thedifference between the three groups was statistically significant(P<0.05).Conclusion As a new-style transfection medium, ultrasoundmicrobubble can achieve the purpose of transfection with a single gene andunited gene. We have observed that transfected double gene into RB tumortissues by ultrasound microbubbles, wtp53cooperate with Rb94genes havemore obvious function in apoptosis of retinoblastoma. Our research was toprovide more effective choice for the treatment of RB, and also expand theidea of the mechanism of RB development.