S1PR3 Promotes The Progression Of T Cell Acute Lymphocytic Leukemia In Vitro And In Vivo

PART Ⅰ THE CONSTRUCTION OF THE T-ALL CELL LINES WITH THE S1PR3 EXPRESSION KNOCK-DOWNObjective: To construct T-ALL cell lines(Jurkat,Cutll1,and Molt-4cells)with the S1PR3 expression knock-down through infecting with retroviral that expressed siRNA targeting the S1PR3 mRNA.Methods: S1PR3 siRNA retroviral vector was packaged and infected the T-ALL cell lines(Jurkat,Cutll1 and Molt-4 cells);The S1PR3 expression of T-ALL cells was detected by qPCR and Western blot.Results: The S1PR3 siRNA retroviral vector was successfully constructed;The results of q-PCR and Western blot showed that multiple T-ALL cell lines(Jurkat,Cutll1,and Molt-4 cells)stably silenced S1PR3 were successfully established.Conclusion: The expression of the S1PR3 in T-ALL cells was silenced by the retroviral that expressed the siRNA fragment.PART Ⅱ THE EFFECT OF S1PR3 ON CELLAPOPTOSIS AND CYCLE OF T-ALL CELLSObjective: To investigate the role of S1PR3 on the apoptosis and cell cycle of T-ALL cell lines(Jurkat,Cutll1 and Molt-4 cells).Methods: The Annexin V staining was used to detect the apoptosis of T-ALL cells.The Propidium iodide(PI)staining was used to detect the cell cycle of T-ALL cells.The flow cytometry was used to detect the staining cells.Results: The data from Annexin V staining showed that the apoptosis levels of several T-ALL cell lines(Jurkat,Cutll1 and Molt-4 cells)respectively increased 24.7%(p<0.01),10.49%(p<0.05)and 11.61%(p<0.001)after S1PR3 silencing.The results of PI staining showed that after silencing S1PR3,the proportion of G2 phase in Jurkat cells was increased 7.22%(p<0.01),and the proportion of S phase was decreased23.04%(p<0.01),but the proportion of G0 phase increased 11.3%(p<0.05).The results of PI staining showed that the proportion of S phase in Cutll1 cells was decreased 12.41% after silencing S1PR3(p<0.05).Conclusion: The S1PR3 inhibits the apoptosis of cells while facilitating them proliferation in T-ALL cells.PART Ⅲ S1PR3 PROMOTES THE PROGRESSION OF T-ALL IN THE MICE XENOGRAFT MODELObjective: To investigate the role of S1PR3 on the progression of the Jurkat and Molt-4 cells in the NCG mice xenograft model.Methods: The Jurkat cells were tagged with Firefly-Luciferase(F-Luc)by lentiviral infection.The luminous intensity of the cells was detected by luminescence detector.The Jurkat-Luc,Jurkat-Luc si,Molt-4 and Molt-4 si cells were injected into the NCG mice with sub-lethal irradiation.The body weight and survival time of the xenograft mice were observed.The progression of T-ALL in mice was detected by the fluorescence imaging system.Results: The Jurkat cells with Firefly-Luc were tagged successfully.After transplantation,the T-ALL cells progressed in the recipient miceand caused the decreasion of the body gradually.The survival time of Jurkat-Luc si group recipient mice transplanting with Jurkat-Luc si cell was significantly longer than that of observation of the mice with the Jurkat-Luc cell(p<0.05);The survival time of the mice with Molt-4 si cells was significantly longer than that of mice with Molt-4 cells(p<0.01);The results of fluorescence imaging showed the fluorescence intensity of Jurkat-Luc si in mice was significantly weaker than that of the Jurkat-Luc cell in mice(p<0.05).Conclusion: The S1PR3 promotes the progression of Jurkat and Molt-4 cells in mice.