1, Silencing Sam68 Gene On The Proliferation Of Acute T Lymphoblastic Leukemia Cells Jurkat 2, Human Endogenous Calcium / Calmodulin-dependent Protein Kinase Ⅱ Inhibitory Protein Expression In Leukemia

Background:T acute lymphoblastic leukemia is a malignant blood disease, caused by an infinite undifferentiated or poorly differentiated lymphocytic proliferation in hematopoietic tissues, occurs in children and adolescents. The new chemotherapy drugs, multi-drug combination of intensive chemotherapy and hematopoietic stem cell transplantation have been carried out so that the treatments of T acute lymphoblastic leukemia greatly improved. However, refractory and recurrent cases still lack effective treatments and adverse events occurring during treatment is a serious problem.Sam68, the substrate of Src in mitosis, belongs to the family of RNA binding proteins.As an adaptor protein the role of Sam68in protein-protein interaction with RNA binding activity is to connect signal transduction of tyrosine kinases with the regulation of RNA metabolism. Recently, a series of published reports stated that Sam68was up-regulated in a variety of human cancers, down-regulation of Sam68remarkably repressed cellular motility and invasion. It indicates that Sam68seem to be implicated in carcinogenesis, progress and prognosis. In our previous study, depletion of Sam68in chicken DT40cells suppressed growth by elongation of the cell cycle. But the effect of Sam68gene on the proliferation of human acute T lymphoblastic leukemia has never been reported. In light of these findings, we investigated the expression of Sam68in human leukemia cells and investigate the effect of down-regulation of Sam68gene on the proliferation of Jurkat cells.Objective:To investigate the effect of Sam68gene on the proliferation of human acute T lymphoblastic leukemia cell line Jurkat, which may provide a theoretical basis for development of novel therapeutic strategies targeting acute T lymphoblastic leukemia.Methods:The sequence of shRNA targeting the site531-552of Sam68mRNA was designed, chemically synthesized and then a single-vector lentiviral, Tet-inducible shRNA-Sam68system (pLKO-Tet-On) was constructed. Next Jurkat cells were infected with lentivirus to create stable cell clones with regulatable Sam68gene expression. The inhibition efficiency of Sam68gene was assayed by Real-time PCR and Western blot; the cells activity of Jurkat cells was detected with MTT assay; the change of colony forming potential of Jurkat cells was analyzed by colony forming unit test; the cell cycle distribution was tested by flow cytometry.Results:The results indicated that the expression of Sam68in experimental cells was statistically decreased compared with that of the control cells; the cells activity and colony forming capacity of the Jurkat cells with Sam68gene silence were significantly inhibited; the percentage of S phase cells was significantly increased, while the percentage of G2phase cells was significantly decreased.Conclusions:It is concluded that Sam68gene silence with shRNA interference could effectively inhibit the proliferation of human acute T lymphoblastic leukemia cell line Jurkat. Background:Calcium/calmodulin-dependent protein kinase Ⅱ (CaMKⅡ) is a multifunctional calcium signaling downstream of serine/threonine protein kinase. It is a widely distributed protein kinase that regulates numerous physiological functions. Inhibition of CaMKⅡ activity, mostly by synthetic reagents, has been proved to suppress cell growth in many cases. hCaMKⅡN is an inhibitor protein (CaMKⅡN) from the human dendritic cell cDNA library by large-scale random sequencing. In recent years, studies have shown that it has some differences in expression of certain normal and abnormal tissue cells. hCaMKⅡN can inhibit tumor cell proliferation and promote apoptosis of tumor cells. Provide a new idea for clinical Oncology therapy.Objective:This study was aimed to investigate the expression of human endogenous CaMKⅡ inhibitory proteins, human endogenous CaMKⅡ inhibitory protein a (hCaMKIINa) and human endogenous CaMKⅡ inhibitory protein β (hCaMKⅡNβ) in acute myelocytic leukemia patients (AML), acute lymphoblastic leukemia (ALL) patients and chronic myeloid leukemia (CML) patients and explore its clinical significance.Methods:The relative expression levels of hCaMKⅡNa and hCaMKⅡNβ in bone marrow mononuclear cells of healthy controls、ALL、AML and CML patients were detected by using Real time PCR.Results:The results showed that the expression levels of hCaMKⅡNa and hCaMKⅡNβ、in ALL、AML and CML patients were significantly lower than healthy controls.Conclusions:It is concluded that hCaMKⅡNa and hCaMKⅡNβ may be involved in the pathogenesis and progression of ALL、AML and CML.