ABSTRACT
To investigate the inhibitory effect of isobutyrylshikonin on the growth of human colon carcinoma cells and its effect on the PI3K/Akt/m-TOR pathway. MTT assay was used to detect the inhibitory effect of different concentrations (0, 6.25, 12.5, 25, 50, 100 mg·L⁻¹) of isobutyrylshikonin on the proliferation of human colon carcinoma cell HT29 at 24, 48 h. CCK-8 method was used to detect the inhibitory effect of isobutyrylshikonin on HT29, HCT116, DLD-1 and Caco-2 at 48 h. AnnexinV/propidium iodide staining was applied in detecting the apoptoticrate of HT29 cells treated with different concentrations of isobutyrylshikonin at 24 h and 48 h. Cycletest plus DNA was employed to analyze HT29 apoptosis and cell cycle after 48 h treatment with isobutyrylshikonin at different concentrations. Western blot and RT-PCR assay were used to examine the protein and mRNA expressions of PI3K, p-PI3K, Akt, p-Akt and m-TOR. The results showed that isobutyrylshikonin inhibited the proliferation of different human colon carcinoma cells, and the inhibition rate was in a dose-dependent manner. Isobutyrylshikonin induced apoptosis mainly in the early stage and blocked cells in the G₀/G₁ or G₂/M phase. Isobutyrylshikonin reduced the protein expressions of PI3K, p-PI3K, Akt, p-Akt, m-TOR and the mRNA expressions of PI3K, Akt, m-TOR in a dose-dependent manner. Isobutyrylshikonin can significantly inhibit the proliferation, induce the early apoptosis and change the cycle distribution in colon carcinoma cells.This biological effect may be correlated with the inhibition of PI3K/AKT/m-TOR pathway.
ABSTRACT
The objective of this study was to detect the level of plasma stromal cell-derived factor-1 (SDF-1) and the expression of CXCR4 (SDF-1 receptor in bone marrow cells) in children with Acute Leukemia (AL) and to investigate the relationship between the expression of CXCR4 and extramedullary infiltration. 48 children with acute leukemia and 20 with non-hematologic malignancies were selected into the AL group and the control group respectively. The peripheral plasma and bone marrow cells were collected. The level of SDF-1 in peripheral plasma was detected by ELISA and the expression of CXCR4 in bone marrow cells was determined by flow cytometry. The results showed that the levels of SDF-1 in peripheral plasma and the expression of CXCR4 in bone marrow cells of AL group was significantly higher than that of control group, among which the level of SDF-1 of the acute lymphoblastic leukemia (ALL) group was also higher than that of the acute myeloid leukemia (AML) group, the expression level of CXCR4 in the bone marrow cells of the extramedullary infiltration (EI) group was higher than that of the non-extramedullary Infiltration (NI) group, and all the differences between the both groups were significant. It is concluded that SDF-1 and CXCR4 express a high level in children with AL, which closely relates with the type of leukemia and the migration and infiltration of leukemia cells in bone marrow.
Subject(s)
Adolescent , Child , Child, Preschool , Female , Humans , Infant , Male , Acute Disease , Bone Marrow , Metabolism , Bone Marrow Cells , Metabolism , Case-Control Studies , Chemokine CXCL12 , Blood , Metabolism , Flow Cytometry , Leukemia, Myeloid, Acute , Metabolism , Pathology , Precursor Cell Lymphoblastic Leukemia-Lymphoma , Metabolism , Pathology , Receptors, CXCR4 , MetabolismABSTRACT
The purpose was to study the responses of AML cell treated with cytochrome C and to explore the influence of cytochrome C on apoptosis of AML cell induced by daunorudicine (DNR). The differentiation of AML cell was detected by Wright-Giemsa staining and NBT test, the apoptosis of AML cell was assayed by flow cytometry and fluorescence microscopy. The results showed as follows: (1) different concentrations of cytochrome C could induce different effects on AML cells. Concentration of cytochrome C for differentiation was 10 microl/ml, for apoptosis was 20 microl/ml, and for necrosis was 40 microl/ml. (2) the apoptosis of AML cells decreased with the administration of cytochrome C in 10.0 microg/ml before treating AML cells with DNR (P < 0.01), but no change was shown with the administration of cytochrome C in 20.0 microg/ml (P > 0.05). (3) in reverse sequence, administrating of cytochrome C in 10 microl/ml and 20 microl/ml after treating AML cells with DNR, two different concentrations of cytochrome C could increase the apoptosis of AML cells (P < 0.01). It is suggested that cytochrome C may probably affect the apoptosis of AML cells induced by DNR.
Subject(s)
Humans , Antibiotics, Antineoplastic , Pharmacology , Apoptosis , Cytochromes c , Pharmacology , Daunorubicin , Pharmacology , Dose-Response Relationship, Drug , Drug Synergism , Flow Cytometry , Leukemia, Myeloid, Acute , Pathology , Microscopy, Fluorescence , Tumor Cells, CulturedABSTRACT
To study the effect of interleukin-15 (IL-15) on the proliferation, differentiation and apoptosis of MDS CD34(+) cells, CD34(+) cells of high enrichment were separated by MACS system, and cultured in liquid media with different concentration of IL-15 in treated group and without IL-15 in the control group. Apoptosis of hematopoietic precursors was assayed by propidium iodine staining and cell by FCM, and the other MDS CD34(+) cells were stained by cytochemical staining after culture. The results showed that after culture with IL-15 the proliferation and differentiation of MDS CD34(+) cells were obviously promoted. It was found the every lineage of mature cells developed, the expressions of cell surface antigens CD71, CD33 and CD19 all increased in the MDS CD34(+) cell treated with IL-15. It is suggested that IL-15 stimulates the proliferation and differentiation of MDS CD34(+) cells, and partly shows anti-apoptosis effects which may be applicable to the therapy MDS.
Subject(s)
Humans , Antigens, CD , Allergy and Immunology , Antigens, CD19 , Allergy and Immunology , Antigens, CD34 , Allergy and Immunology , Antigens, Differentiation, Myelomonocytic , Allergy and Immunology , Apoptosis , Bone Marrow Cells , Allergy and Immunology , Pathology , Cell Cycle , Cell Differentiation , Cell Proliferation , Cells, Cultured , Flow Cytometry , Interleukin-15 , Pharmacology , Microscopy, Fluorescence , Myelodysplastic Syndromes , Blood , Allergy and Immunology , Pathology , Receptors, Transferrin , Allergy and Immunology , Sialic Acid Binding Ig-like Lectin 3ABSTRACT
<p><b>OBJECTIVE</b>To study the inhibition of the expression of bcl-xL gene induced by RNA interference in CNE-2Z cell line in addition to the inhibition of its proliferation and apoptotic induction.</p><p><b>METHODS</b>Small interfering RNAs targeting bcl-xL gene were synthesized by using web design software provided by Amnion and the silencer short interfering RNA (siRNA) construction kit; fluorescein-labeled siRNAs were done by FAM-silencer siRNA labeling kit; siRNAs were transfected into CNE-2Z cells by using lipofectamine 2000 reagent; siRNA transfection efficiencies were analyzed by fluorescent microscopy; down-regulation of bcl-xL was detected by RT-PCR; thiazolyl blue (MTT) assay was used to assess the cell growth; apoptosis of CNE-2Z cells was analyzed by flow cytometry.</p><p><b>RESULTS</b>Green fluorescence in the cells was seen clearly in FAM-labeled siRNA transfected group under the fluorescent microscope while none in the untransfected group. Different down-regulations of bcl-xL mRNA expression were found in the transfected groups. The expression of bcl-xL mRNA decreased by 10% - 70% in the siRNAs transfected CNE-2Z by RT-PCR scan analysis. The inhibitory rate of cell proliferation depended on time and concentrations to some extent. Different cell apoptosis could be induced by different concentrations of siRNA4.</p><p><b>CONCLUSIONS</b>The synthesized siRNAs in vitro were able to down-regulate the expression of bcl-xL There were different capabilities of the specific siRNAs down-regulation. The transient transfected bcl-xL siRNA4 could effectively inhibit the growth of the cancer cells and induce theirs apoptosis. It was suggested that the siRNA technique provide not only an extremely powerful tool for the functional analysis of genome but also a new method for anti-nasopharyngeal carcinoma gene therapy.</p>
Subject(s)
Humans , Apoptosis , Cell Line, Tumor , Cell Proliferation , Gene Expression Regulation, Neoplastic , Nasopharyngeal Neoplasms , Genetics , Pathology , RNA Interference , RNA, Messenger , Genetics , Transfection , bcl-X Protein , GeneticsABSTRACT
Objective To analyze the chemokine receptor CXCR4 expression in acute leukemic cells of children and its relationship with extramedullary infiltration.Methods The immunotypes of cases of acute leukemia in children and the expression of CXCR4 in marrow leukemic cells were studied by flow cytometry respectively. The relationship between CXCR4 expression and extramedullary infiltration of leukemic cells were analyzed by statistical method.Results The expression rates of CXCR4 in ALL children were higher than those in NALL children(P