Expression of small nucleolar RNA SNORD15A in acute leukemia and its clinical significance / 白血病·淋巴瘤

Objective:To investigate the expression level of small nucleolar RNA SNORD15A in bone marrow of patients with acute leukemia (AL) and its relationship with clinical characteristics and prognosis of patients.Methods:Bone marrow blood samples of 53 newly treated AL patients and 29 healthy subjects without clinical diagnosis of hematologic diseases or other malignant diseases (control group) at the Affiliated Hospital of Guangdong Medical University from March 2018 to December 2021 were collected. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the relative expression of SNORD15A in bone marrow blood mononuclear cells of the two groups. The median relative expression of SNORD15A (0.148) was used as the boundary, and AL patients were divided into low expression group (<0.148) and high expression group (≥0.148). The relationship between the expression level of SNORD15A and the clinical characteristics, clinical indicators and overall survival (OS) of AL patients was analyzed. Kaplan-Meier method was used for survival analysis and log-rank test was performed; Cox proportional hazards model was used for univariate and multivariate analyses of OS of patients.Results:The relative expression of SNORD15A was 0.148 (0.012-1.376) in newly treated AL patients and 0.921 (0.513-2.288) in the control group, and the difference was statistically significant ( Z = -6.85, P < 0.01). The differences in SNORD15A relative expression between patients with different prognostic stratification, efficacy and with or without fever and bleeding were statistically significant (all P < 0.05). The differences in platelet count, plateletcrit and albumin levels between SNORD15A low expression group and high expression group were statistically significant (all P < 0.05), and the differences in molecular biology and cytogenetic characteristics were not statistically significant (all P > 0.05). The patients in SNORD15A high expression group had better OS than the low expression group ( P < 0.05). The results of univariate Cox regression analysis showed that SNORD15A was an influencing factor for patients' OS ( HR = 0.063, 95% CI 0.005-0.766, P < 0.05); the results of multivariate Cox regression analysis showed that fatigue ( HR = 4.754, 95% CI 1.014-22.290), fever ( HR = 0.147, 95% CI 0.029-0.746) and hemoglobin ( HR = 0.970, 95% CI 0.944 -0.998) were independent influencing factors for OS (all P < 0.05). Conclusions:SNORD15A is lowly expressed in AL and may be an indicator for disease monitoring and prognostic assessment in AL patients.

The impact of α-particles irradiation on src kinase activity and autophagy system mediated by ROS / 中华放射医学与防护杂志

Objective To investigate the modulation role of autophagy in radiation-induced cell death by detecting the response of src kinase activity and autophagy in HEK293 cells irradiated with different dose of α-particle.Methods HEK293 cells were irradiated by contral group (0 cGy) a low dose group (10 cGy) and high dose group (300 cGy) α-particles.Molecular probe 2',7'-dichlorofluorescin (DCFHDA) was used to detect the cell ROS.The src kinase activity and endogenous protein level of LC3Ⅰ/Ⅱ were monitored by Western Blot.Cell death rate of irradiated cells pretreated with autophagy inducer of rapamycin was tested by flow cytometry.Results Compared with control group,the ratio of LC3Ⅰ/Ⅱ decreased (t =4.07,P ＜ 0.05) and the percentage of cells with GFP-LC3 punctuate dots increased (t =12.29,P ＜0.05) under 10 cGy irradiation,indicating the induction of autophagy.On the contrary,the ratio of LC3Ⅰ/Ⅱ increased (t =2.93,P ＜ 0.05) and the GFP-LC3 morphology had no obvious change under 300 cGy irradiation.The cellular ROS level reached to the maximum value at 4 h postirradiation.Both 10 cGy and 300 cGy irradiation could elevate the ROS level (t =17.93,22.88,P ＜0.05),whereas the amplitude of elevation of 300 cGy irradiation was higher than that of 10 cGy irradiation (t =15.76,22.66,14.22,P ＜ 0.05).Compared with control group,the 419th site of tyrosine residue in src kinase manifested hyper-phosphorylation (t =5.66,P ＜0.05) under 10 cGy irradiation whereas it had hypo-phosphorylation under 300 cGy irradiation (t =4.67,P ＜ 0.05).Treatment of cells with DMSO could partly restore the impact on src kinase activity under high or low dose irradiation.Pre-treating the cells with autophagy inducer rapamycin could reduce cell death under 300 cGy irradiations (t =12.14,P ＜ 0.05).Conclusions High or low dose of α-particles irradiation could inhibit or activate src kinase and autophagy system,respectively.ROS mediated the response of src kinase activity and autophagy system induced by irradiation.Modulation of autophagy could desensitize cell responses to irradiation.

Affinity maturation of anti-TNF-alpha scFv with somatic hypermutation in non-B cells

Activation-induced cytidine deaminase (AID) is required for the generation of antibody diversity through initiating both somatic hypermutation (SHM) and class switch recombination. A few research groups have successfully used the feature of AID for generating mutant libraries in directed evolution of target proteins in B cells in vitro. B cells, cultured in suspension, are not convenient for transfection and cloning. In this study, we established an AID-based mutant accumulation and sorting system in adherent human cells. Mouse AID gene was first transfected into the human non-small cell lung carcinoma H1299 cells, and a stable cell clone (H1299-AID) was selected. Afterwards, anti-hTNF-α scFv (ATscFv) was transfected into H1299-AID cells and ATscFv was displayed on the surface of H1299-AID cells. By 4-round amplification/flow cytometric sorting for cells with the highest affinities to hTNF-alpha, two ATscFv mutant gene clones were isolated. Compared with the wild type ATscFv, the two mutants were much more efficient in neutralizing cytotoxicity of hTNF-alpha. The results indicate that directed evolution by somatic hypermutation can be carried out in adherent non-B cells, which makes directed evolution in mammalian cells easier and more efficient.

Study on the antithrombotic effect of fibrinolytic proteins from Scolopendra subspinipes mutilans / 中国药理学通报

Aim The aim is to purify fibrinolytic enzyme from Scolopendra subspinipes mutilans L.Koch and to study the thrombolytic and anticoagulant effect.Methods Scolopendra subspinipes mutilans L.Koch fibrinolytic enzyme(SSFE) was purified by ammonium sulphate precipitation,DEAE-cellulose and SephadexG-75 column chromatography from Scolopendra subspinipes mutilans L.Kochby.And fibrinolytic activity was determined by fibrin plate.The anticoagulant effect was measured on mice with haemolytic test and hemorrhagic test.The thrombolytic effect was measured with rats In vitro and in vivo,and the activated partial thromboplastin time(APTT),plasma prothrombin time(PT),thrombin time(TT) were measured.Results SSFE was single component with fibrinolytic activity and without any hemolyzation and hemorrhagic activity.All doses of SSFE(2,5,10 mg?kg-1) could obviously prolong activated partial thromboplastin time(APTT) and thrombin time(TT) ;Middle dose of SSFE(5 mg?kg-1) could prolong plasma prothrombin time(PT) while high dose of SSFE(10 mg?kg-1) didn't prolong obviously.Conclusion SSFE has obvious thrombolytic effect and anticoagulant effect.