Effect of SHIP-1 on Invasion, Migration and PI3K-AKT Signaling Pathway of Leukemic Cells / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect of SH2-containing inositol phosphatase-1 (SHIP-1) on the proliferation, invasion and migration of human leukemia cells as well as phosphatidylinositol-3 kinase (PI3K) / protein kinase B (AKT) signaling pathway.</p><p><b>METHODS</b>The overexpression vector pCDNA3.1-SHIP1 was transfected into THP-1 cells by Lipofectamine 2000. The experiment was divided into 3 groups: control group (untreated cells) and empty vector group (transfected with empty vector pCDNA3.1-NC) and overexpression group (transfected with overexpression vector pCDNA3.1-SHIP1). The cell proliferation was tected by CCK-8 assay, Transwell assay was used to evaluate the cell invasion and migration capabilities. The expressions of SHIP-1, AKT, phosphorylated AKT (pAKT), matrix metalloproteinase-9 (MMP-9) protein were analyzed by Western blot.</p><p><b>RESULTS</b>The expression of SHIP-1 in overexpression group was significantly higher than that in the control group(P<0.05). Compared with the control group, the absorbance of the cells in the empty vector group was not statistically different (P>0.05), and the absorbance in overexpression group decreased significantly(P<0.05). The cell numbers of invasion and migration were not significantly different between empty and control groups(P>0.05), but those in overexpression group were significantly lower than those in the control group(P<0.05). Compared with the control group, the expression of AKT, pAKT and MMP-9 in the empty vector group was not statistically different (P>0.05); the AKT protein in overexpression group was not significantly different (P>0.05), but the pAKT and MMP-9 significantly decreased(P<0.05).</p><p><b>CONCLUSION</b>SHIP-1 plays a role in inhibiting the proliferation, invasion and migration of leukemia cells, the mechanism probably relates with supressing the expression of MMP-9 by regulating PI3K/AKT signaling pathway.</p>

Anti-inflammatory effect of interleukin-35 in mice with colitis and its mechanism / 浙江大学学报·医学版

OBJECTIVE@#To investigate the anti-inflammatory effect and mechanisms of interleukin-35 (IL-35) in inflammatory bowel disease.@*METHODS@#BALB/c mice were divided into three groups with 10 mice in each group:control group, model group (oral administration of 4% glucan sodium sulfate for 7 d) and IL-35-treated group (oral administration of 4% glucan sodium sulfate for 7 d, intraperitoneal injection of 2 μg IL-35 at d2-5). Disease activity index (DAI) was scored every day. After 7 d, the mice were sacrificed, and the serum and intestinal tissue samples were collected. The gross morphology of the colon was observed; HE staining was used to observe the pathological changes of colon tissue; flow cytometry was employed to detect the change of macrophage polarization ratio in colon tissue; the mRNA expression levels of cytokines IL-6, TNF-α, IFN-γ, IL-10 and SHIP1 in colon tissue were determined by real-time quantitative RT-PCR; the expression and distribution of SHIP1 in colon tissue was measured by immunohistochemistry; Western blotting was adopted to detect the expression level of SHIP1 protein in colonic intestinal tissues of each group.@*RESULTS@#The DAI scores of the mice in the model group were higher than those in the control group, while the DAI scores in the IL-35-treated group were lower than those in the model group (all 0.05). Compared with the model group, microscopic inflammatory infiltration score was decreased and microscopic crypt destruction and score was significantly lower in IL-35-treated group (all <0.05). The relative expression of proinflammatory cytokines IL-6, TNF-α and IFN-γ in the colon tissue of IL-35-treated group was decreased compared with the model group, while the relative expression of IL-10 mRNA was higher than that of the model group (all <0.05). Compared with the control group, the proportion of M1 macrophages in the model group increased (<0.05), while the proportion of M1 macrophages in the IL-35-treated group was lower than that in the model group (<0.05). The relative expression of SHIP1 mRNA and protein in the colon tissue of IL-35-treated group was higher than that in the model group (all <0.05).@*CONCLUSIONS@#IL-35 can inhibit the polarization of M1 macrophages and regulate inflammatory cytokines to promote anti-inflammatory effect on mice with colitis.

Mutation analysis of SHIP gene in acute leukemia / 中国实验血液学杂志

The SH2 domain containing inositol 5'-phosphatase (SHIP) was initially described as a 145 kD protein phosphorylated on tyrosines upon growth factor and cytokine stimulation. SHIP is predominately expressed in hematopoietic cells, and is a crucial negative regulator in the development of hematopoietic cells. To evaluate the role of the SHIP gene in human leukemogenesis, expression and mutation of SHIP gene in bone marrow and/or peripheral blood from 32 patients with acute myeloid leukemia (AML), 9 patients with acute lymphoblastic leukemia (ALL), as well as human hematopoietic cell lines were analyzed by reverse transcription-polymerase chain reaction (RT-PCR), single strand conformational polymorphism (SSCP) and sequencing. The RT-PCR showed that all samples expressed SHIP gene. Mutations of SHIP gene were detected in 7 out of 32 AML patients (22%) and one out of 9 ALL patients (12%). Interestingly, two missense mutations that had been observed in one AML patient at diagnosis disappeared after complete remission (CR). In addition, Akt phosphorylation was prolonged and increased following IL-3 stimulation in this patient sample. In conclusion, data of this study demonstrate the mutation of the SHIP gene in acute leukemia for the first time and suggest a possible role of the mutation of this gene in the development of acute leukemia. SHIP serves as a tumor suppressor by negatively regulating the PI3K/Akt signaling pathway in hematopoietic cells.