Objective:To observe the effect of Chinese medicine of nourishing yin and clear heat for prevention of chronic graft-versus-host disease(cGVHD) in mice.Methods:Mice in the Chinese medicine groups were administrated with high-dose,middle-dose and low-dose yin-nourishing and heat-clearing Chinese medicine respectively from the day of model induction.Positive control group and negative control group were administrated with the same volume of normal saline.At the 2nd,5th,8th,12th week of model induction,the number of Y chromosome in bone marrow cells was detected for the analysis of to chimerism.The body weight,hair and incidence of cGVHD were observed every 3 days.One hundred days after model induction,the pathological changes of mice skin,liver and intestine were observed.Results:Stable chimerism formed in all Chinese medicine groups and positive control group.Incidence of cGVHD in high-dose and middle-dose Chinese medicine groups was lower than that in low-dose Chinese medicine group and positive control group.As for the symptomatic grading and pathological grading,statistical difference was found between high-dose Chinese medicine group and positive control group while not found between low-dose Chinese medicine group and positive control group.Conclusion:Appropriate dose of Chinese medicine of nourishing yin and clear heat can prevent cGVHD in mice with haploidentical bone marrow transplantation.

OBJECTIVE:To study the effects of small interfering RNA (siRNA)-mediated silencing of CD158b expression on the efficiency of the natural killer (NK) cells in killing allogeneic dendritic cells.METHODS:After the knockdown of CD158b by CD158b -SiRNA, the CD158b mRNA expression in natural killer cells was examined by qRT-PCR and the CD158b protein expression by flow cytometer. The cytotoxic activity of RNAi-NK cells and normal NK cells against the allogeneic dendritic cells was detected by LDH release assay.RESULTS:The CD158b mRNA expression and its protein expression were decreased significantly in the NK cells by CD158b siRNA (P/0.05). The cytotoxic activities of alloreactive NK cells generated by RNAi CD158b expression against allogeneic dendritic cells were increased significantly.CONCLUSION:Silencing CD158b gene can inhibit the NK cell CD158B mRNA and protein expression. Alloreactive NK cells generated by RNAi CD158b expression have the potential for use in interventions of GVHD.

Objective To study whether special Ly49C-SiRNA can inhibit the Ly49C gene expression in mice natural killer cells.Methods After the knockdown of Ly49C by Ly49C-SiRNA, the Ly49C mRNA expression was examined by SYBR Real-time PCR and Ly49C protein expression by flow cytometry in mice natural killer cells. Results The Ly49C mRNA and its protein expression significantly decreased in mice natural killer cells by Ly49C-SiRNA(P<0.05).Conclusion Silencing Ly49C gene inhibits mice natural killer cell Ly49C mRNA and protein expression.

OBJECTIVE:To investigate the in vitro proliferation of CD4(+)CD25(+) T cells from the peripheral blood mononuclear cells (PBMCs) of the chronic myelocytic leukemia patients and the inhibitory effect on CD4(+)CD25(-) T cells.METHODS:Magnetic-activated cell sorting (MACS) was used to separate CD4(+)CD25(+) T and CD4(+)CD25(-) T cells from the PBMCs of patients with chronic myelocytic leukemia, and the purity and activity of CD4(+)CD25(+) T cells were analyzed with flow cytometry. After stimulation with anti-CD3 mAb, anti-CD28 mAb and recombinant human interleukin-2 (rhIL-2), the CD4(+)CD25(-) and CD4(+)CD25(+) T cells were cocultured to observe the inhibitory effect of CD4(+)CD25(+)T on CD4(+)CD25(-)T cells using MTT assay.RESULTS:After cell sorting, the purity of CD4(+)CD25(+)T cells from healthy control and chronic myelocytic leukemia patients were (84.93-/+2.55)% and (86.32-/+2.40)%, respectively, showing no significant difference between them (P>0.05). The activity of CD4(+)CD25(+) and CD4(+)CD25(-) T cells from healthy control and the leukemic patients was also comparable [(98.12-/+0.68)% vs (97.33-/+0.78)%, P>0.05). In the coculture, CD4(+)CD25(+) T cells obviously inhibited CD4(+)CD25(-) T cell proliferation in vitro, and the maximum inhibition occurred when CD4(+)CD25(+)T cells were cocultured with CD4(+)CD25(-)T cells at the ratio of 1:1.CONCLUSION:The MACS system can effectively isolate CD4(+)CD25(+) and CD4(+)CD25(-) T cells. CD4(+)CD25(+) T cells obviously inhibit the proliferation of CD4(+)CD25(-)T cells in vitro, and the effect displays an effector-target ratio relationship.

目的:探讨扩增后的小鼠CD4+CD25+T细胞(Treg)输注对异基因骨髓移植(allo-BMT)后移植物抗宿主病(GVHD)的影响.方法:利用免疫磁珠法分选小鼠Treg细胞;以羊抗鼠CD3ε单抗、羊抗鼠CD28单抗、鼠重组IL-2及辐射过的BALB/c小鼠脾细胞为共刺激因子,扩增Treg细胞;建立C57BL/6→BALB/c小鼠allo-BMT模型,移植受鼠随机分为3组,移植后6～8 h后分别予尾静脉输注扩增的Treg细胞、CD4+cD25-T细胞和RPMI1640培养液.以移植后GVHD表现,肝、脾、小肠组织病理形态、生存期为观察指标并进行组间比较.结果:经免疫磁珠法分选可获得高纯度(91.80±2.15)%及具较强活力(97.58±1.23)%的Treg细胞;移植后(4周左右)的扩增后Treg细胞移植组小鼠肝、脾、小肠GVHD病理损害较同期cD4+CD25-T细胞移植组与空白对照组小鼠有一定程度的减轻.3组小鼠移植后平均存活时间分别为(61.45±27.88)d、(18.58±12.39)d和(26.37±15.65)d,扩增后Treg细胞移植组小鼠平均存活时间较CD4+CD25-T细胞、空白对照组小鼠延长(JP<0.05).结论:allo-BMT后输注扩增的Treg细胞可以减轻移植后GVHD程度,延长小鼠生存时间.