Objective To investigate the effect of specific small interfering RNA (siRNA) silencing the expression of casitas B cell lymphoma-b (Cbl-b) gene on the immune activity of the mouse spleen T lymphocytes.Methods T lymphocytes were isolated by Nylon Wool Fiber Columns.By using anti-CD3 antibodies (1 mg/L) as the first signal of T lymphocyte activation in vitro,and at 12,24 and 48 h after transfecting by liposome,the expression of CD69,CD25 and CD71 was detected and analyzed by using fluorescence conjugated monoclonal antibodies and flowcytometry.At 48 h after transfection,the proliferation of the T lymphocyte was measured with CCK-8 kit.Results Specific Cbl-b siRNA could effectively silence the expression of Cbl-b gene.At 12 h after transfection,CD69 molecular expression levels in transfection group,negative control group and blank group were (46.78 ±4.89)％,(21.36 ±2.06)％,and (25.09 ±2.27)％ respectively (P ＜ 0.01).At 24 h after transfection,CD25 molecular expression levels in transfection group,negative control group and blank group were (19.27 ± 1.84)％,(8.18 ±0.71)％,and (9.25 ± 0.65) ％ respectively (P ＜ 0.01).At 48 h after transfection,CD71 molecular expression levels in transfection group,negative control group and blank group were (43.26 ± 3.89)％,(23.99 ±1.97) ％,and (28.09 ± 2.15)％ respectively (P ＜ 0.01).At 48 h after transfection,A values detected by CCK-8 kit in transfection group,control group,and blank group were 0.763 ± 0.063,0.356 ± 0.039,and 0.383 ± 0.015 respectively (P ＜ 0.01).Conclusion Specific siRNA silencing Cbl-b gene can markedly promote the immune activity of mouse T lymphocytes.

Objective To design, synthesize screen small interfering RNA (siRNA) targeting to Casitas B-cell lineage lymphoma-b (cbl-b).Methods Four pairs of 21 nucleotide siRNAs directed to Cbl-b mRNA were designed and synthesized by utilizing RNA design software to simulate secondary structure of cbl-b mRNA in mice. These siRNAs were respectively transfected into lymphocytes in 96 shadows mask by oligofectamine package, and untreated and unspecific siRNA-transfected lymphocytes served as controls. The expression of cbl-b protein was detected by Western blotting.Results When the work concentration of siRNA was 100 nmol/L, transfection efficiency of lymphocytes was highest, up to (87.48±1.94)% and the mean fluorescence intensity was strongest, up to 33.09±1.77. Compared with bland controls, the expression of cbl-b protein level was markedly down-regulated in siRNA-transfected lymphocytes. The inhibitory rate of the siRNA of the target-4 was highest, up to 85%. The expression of cbl-b protein in unspecific siRNA-transfected lymphocytes had no significant changes.Conclusion siRNA-4, which can highly effectively inhibit protein expression of cbl-b gene, was screened successfully, and its inhibition effect can maintain near 48 h. It is hopeful that the cbl-b siRNA will activate lymphocytes directly by cbl-b gene silencing, and kill tumor by activate immunization.