IgVH mutation status in patients with chronic lymphocytic leukemia / 中华血液学杂志

<p><b>OBJECTIVE</b>To evaluate the frequency and mutation status of IgVH gene expression in patients with chronic lymphocytic leukemia (CLL) in China.</p><p><b>METHODS</b>IgVH mutation was detected by multiplex PCR and directly sequencing in 29 CLL patients. IgH somatic hypermutation and mutation site were analysed by IMGT/V-QUEST.</p><p><b>RESULTS</b>Of 29 CLL patients, 21 had IgVH mutation (72%). The most frequently expressed VH gene family was found to be VH3 (55%) followed by VH4 (38%), VH2 (3.5%) and VH7 (3.5%), with no expression of VH1, VH5 and VH6.</p><p><b>CONCLUSIONS</b>The expression frequency of IgVH gene families in Chinese CLL patients is significantly different from that in Western CLL patients, suggesting the involvement of ethnic and/or environmental factors in CLL development, which might partly explain the different incidence of CLL between China and Western countries.</p>

Expression of CLLU1 in patients with chronic lymphocytic leukemia and its prognostic significance / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate CLLU1 expression and its relationship with clinical stage, expression of CD38 and ZAP-70, and chromosome abnormalities in patients with chronic lymphoid leukemia (CLL).</p><p><b>METHODS</b>Fifty CLL patients were studied. Semiquantitative RT-PCR was performed to detect CLLU1 expression levels; three-color flow cytometry to detect the ZAP-70 and CD38 expression, interphase fluorescence in situ hybridization (FISH) to detect chromosomal aberrations.</p><p><b>RESULTS</b>The expression of CLLU1 mRNA was detected in 26 of the 50 cases (52%), of them 7 cases (26.92%) in Binet A and 19 (73. 08%) in Binet B + C. The expression levels of CLLU1 were significantly higher in Binet stage B + C than in Binet stage A (P < 0. 01). Among 24 CD38+ CLL cases, 17 (70. 83%) expressed high CLLU1 mRNA, whereas in 26 CD38- CLL patients only 9 (34.62%) were CLLU1 positive. The expression of CLLU1 in CD38+ CLL was significantly higher than that in CD38- CLL (P < 0.05). However, no significant difference of CLLU1 expression was found between ZAP-70+ and ZAP-70- patients (P > 0. 05), and between chromosomal aberrations (P > 0. 05).</p><p><b>CONCLUSIONS</b>CLLU1 expression was significantly associated with clinical stage and CD38 expression, and might be an important prognostic factor in CLL patients.</p>

Detection of IgVH mutation status in patients with chronic lymphocytic leukemia by multiplex PCR / 中国实验血液学杂志

IgVH mutation status is one of the most important independent prognostic factor of chronic lymphocytic leukemia (CLL). In order to evaluate IgVH mutation status in patients with CLL, IgVH mutation was detected by multiplex PCR in 9 CLL patients and purified PCR amplification products were directly sequenced, IgH somatic hypermutation and mutation site were analysed by IMGT/V-QUEST. The results showed that 5 patients had mutated IgVH, and IgVHs were IGHV3-11*03, IGHV3-9*01, IGHV3-23*01, IGHV4-59*01, IGHV4-34*02, respectively; whereas 4 others had unmutated IgVH, these IgVHs were IGHV3-53*01, IGHV3-23*03, IGHV3-33*05 and IGHV3-7*01. It is concluded that multiplex PCR is a rapid and easy method to detect IgVH mutation status, and it solves the limitations and pitfalls of routine PCR, and it is worth being extensively used both in clinic and scientific researches.

Multiple myeloma cell line U266 apoptosis induced by velcade / 中国实验血液学杂志

To investigate the effect of velcade on multiple myeloma cell line U266 apoptosis and its mechanism, cell viability was estimated by trypan blue dye exclusion. Annexin-V, mitochondrial transmembrane potential (delta psi m) and reactive oxygen species (ROS) labeled by DCFHDA were examined by flow cytometry, the expression of bcl-2 mRNA was detected by semi-quantitative RT-PCR. The results showed that the velcade inhibited the growth of U266 cells and reduced cell viability accompanied by appearance of morphologic characteristics of apoptosis. Velcade at 50 nmol/L increased Annexin V positivity and fluorescence intensity of DCF because of ROS generation while it decreased the delta psi m of U266 cells. Expression of anti-apoptotic gene bcl-2 mRNA also decreased. It is concluded that velcade inhibited the growth and reduce cell viability of U266 cells. Velcade can induce U266 cells apoptosis by intrinsic cell apoptotic pathway.

Application of a four antibody (cMPO/cCD79aalpha/cCD3/CD45) combination to the diagnosis of acute leukemia expressing cross-lineage antigens / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the diagnostic value of intracellular antibody combination in acute leukemia (AL) expressing cross-lineage cell-surface antigens.</p><p><b>METHODS</b>Flow cytometric immunophenotyping using intracellular antibody combination (cMPO/cCD79alpha/cCD3/CD45) was performed additionally in 60 patients who expressed cross-lineage antigens from 269 previously untreated adult AL.</p><p><b>RESULTS</b>Fifty-four of 269 previously untreated adult AL patients who expressed only one kind of intracellular antigen were diagnosed as cross-lineage AL, the percentage of cross-lineage AL in T cell acute lymphoblastic leukemia (T-ALL), B-ALL and acute myeloid leukemia (AML) was 28.6%, 43.6% and 13.4%, respectively. The positive rate of CD7, CD19, CD5 and CD20 in cross-lineage AML was 65.4%, 15.4%, 11.5%, and 7.7%, respectively. The positive rate of CD13, CD33 and CD15 in cross-lineage ALL was 89.3%, 21.4% and 3.6%, respectively. Six (2.3%) patients expressed two-lineage intracellular antigens were diagnosed as biphenotypic AL: 2 of T/B type and 4 B/M (B/myeloid) type.</p><p><b>CONCLUSION</b>Intracellular antibodies possess lineage specificity and four-color combination flow cytometric immunophenotyping can provide fast and multi-parameter data. To ensure accuracy of the results, CD45/SSC gating and normal cells as internal reference should be used in the immunophenotyping of abnormal cells.</p>