AIM: To analyze T cell clonality in patients with T cell acute lymphoblastic leukemia (T-ALL). METHODS: The complementarity determining region 3 (CDR3) size of 24 T cell antigen receptor variable β (TCR Vβ) region gene was analyzed in peripheral blood mononuclear cell (PBMC) samples from 6 T-ALL cases and 10 normal individuals by using reverse transcriptase-polymerase chain reaction (RT-PCR). The PCR products were further studied by genescan and sequencing analysis. RESULTS: Some TCR Vβ subfamily T cells display mono- or oligoclonal expansions in 3 cases of T-ALL, predominantly in Vβ2, Vβ3, Vβ6, Vβ9, Vβ21 or Vβ24, respectively. Polyclonal expansions of T cells were found in the other three cases, which could also be found in normal samples. CONCLUSION: A part of T cell acute lymphoblastic leukemia cells may arise from a clonal expansion of TCR Vβ subfamily T cell. This method may be useful for the detection of minimal residual disease in clinical study of the disease.

Objective To explore the mechanism and way for CT10 regulator of kinase like (CRKL) involving in drug resistance in leukemia cells. Methods The four major proteins included Ras protein, signal transducer and activator of transcripton 5 (STAT5) protein, phosphoinositide 3-kinase (PI3K) protein and paxillin protein in leukemia which involved in signal transduction pathway of CRKL. The expressions of those proteins were detected by Western-blot and immunofluorescent staining and confocal laser scanning microscopy. Results Compared with K562/S cells, the expressions of Ras(41.52±15.47 vs. 23.74±8.67) and PI3K (35.60±12.48 vs. 10.09±0.005) protein were up-regulated in K562/ADM cells (t=3.01,6.13;both P<0.05), while there were no significant changes in the expressions of paxillin (20.10±11.89 vs. 23.11±12.40) and STAT5 protein (25.72±14.46 vs. 17.58±9.21) between K562/S cells and K562/ADM cells(t=0. 18,1.43;both P>0. 0S). Conclusions Ras and PI3K protein may play a role in the multidrug resistance of K562 cell line, while paxillin and STAT5 protein may be not involved in the formation of resistant in K562 cells.

Objective To construct a lentiviral vector carrying CRKL gene RNA interfering( RNAi ).Methods The CRKL RNAi was selected and subcloned into the lentiviral vector,pGCL-GFP(including U6 promotor and green fluorescent protein),generating the lentiviral vector LV-shCRKL.The corrected CRKL was confirmed by endoenzyme digestion ,sequencing.Recombinant lentiviruses were produced by 293T cells following the eo-transfection of LV-shCRKL,with the packaging plasmids pHelper1.0 and pHelper2.0.The virus titer was detected by GFP expressions in 293T cells.Results Plasmid LV-shCRKL carried the correct sequence.The recombinant lentiviruse LV-shCRKL could be produced by co-transfection of LV-shCRKL to 293T cells.Conclusion The recombinant lentiviruse vector LV-shCRKL is constructed successful.

Objective To investigate the role of CRKL activity in leukemia cells with muhidrug resistance and find new factor related to multidrug resistance. Methods By flow cytometry, CRKL activity was compared in K562, HL-60 cells and its resistance cells. The change of CRKL activity was observed in sensitive cells treated with and withdrawal daunorubicin. Results With the comparison of K562, HL-60 sensitive cells, in K562, HL-60 resistant cell lines, the level of CRKL phosphorylation in K562, HL-60 resistance cells treated with daunombicin 72 hours increased markedly. The level of CRKL phosphorylation was time-dependent with chemotherapy drugs, not change of CRKL activity was found in Jurkat ceils.Conclusion The level of CRKL activity is new factor related to muhidrug resistance in leukemia cells.

AIM: To analyze T cell clonality in patients with T cell acute lymphoblastic leukemia (T-ALL). METHODS: The complementarity determining region 3 (CDR3) size of 24 T cell antigen receptor variable ? (TCR V?) region gene was analyzed in peripheral blood mononuclear cell (PBMC) samples from 6 T-ALL cases and 10 normal individuals by using reverse transcriptase-polymerase chain reaction (RT-PCR). The PCR products were further studied by genescan and sequencing analysis. RESULTS: Some TCR V? subfamily T cells display mono- or oligoclonal expansions in 3 cases of T-ALL, predominantly in V?2, V?3, V?6, V?9, V?21 or V?24, respectively. Polyclonal expansions of T cells were found in the other three cases, which could also be found in normal samples. CONCLUSION: A part of T cell acute lymphoblastic leukemia cells may arise from a clonal expansion of TCR V? subfamily T cell. This method may be useful for the detection of minimal residual disease in clinical study of the disease.

Objective To investigate the renoprotective effects of autologous transplantation of adipose-derived mesenchymal stem cells (ADMSCs) in diabetic rats. Methods Sprague-Dawley (SD) rats were injected intraperitoneally with 40 mg/kg streptozotocin (STZ) for 5 consecutive days to induce type 1 diabetes. Four weeks following STZ injection, eighteen SD rats were randomized into two groups: the diabetic group (n = 9) and the ADMSCs group (n = 9). Normal nondiaetic rats were set as the normal control (n = 9). Autologous ADMSCs were cultured and identified in vitro , which were intravenously injection to the ADMSCs group rats via the tail vein. At 8 weeks after transplantation, levels of blood glucose, insulin, serum urea nitrogen, serumcreatinine and urine protein were measured. Meanwhile the body weight and kidney weight were examined. Results Mesenchymal cell surface markers were expressed in the cultured ADMSCs. The ADMSCs could differentiate into the adipogenic and osteoblastic lineages. Both the diabetic group and the ADMSCs group rats had higher levels of blood glucose , urea nitrogen , serum creatinine , urine protein and higher ratio of the kidney weight/body weight than those in the normal control group (P < 0.05, respectively). Blood glucose, urea nitrogen and the ratio of kidney weight/body weight in the ADMSCs group rats were significantly decreased compared with the diabetic group (P < 0.05, respectively). The decreased insulin level was attenuated after transplantation of ADMSCs (P < 0.05). Besides, levels of serum creatinine and urine protein in the ADMSCs group were lower than those in the diabetic group with no significant difference. Conclusion Autologous transplantation of ADMSCs can improve metabolic disorder and relieves diabetic renal damage.

AIM: To investigate the expression level of transcriptional factor Elf-1 in mononuclear cells, CD4~+ and CD8~+ T cells from umbilical cord blood (UCB). METHODS: Real-time PCR with SYBR green I technique was used for detecting the Elf-1 expression in mononuclear cells, sorted CD4~+ and CD8~+ T cells from 12 cases of umbilical cord blood. The relative mRNA expression level of Elf-1 was analyzed by a formula of 2~(-△Ct)×100%. The expression level of β_2-microglobulin gene (β_2M) was used as an endogenous reference. The peripheral blood from 10 cases of healthy adults was served as control. RESULTS: Elf-1 mRNA expressed in all blood samples collected from both UCB and healthy adults. The expression level showed apparent diversity in different individuals. The relative mRNA expression of Elf-1 in both mononuclear cells (18.55%±2.48%) and CD8~+ T cells (3.52%±0.45%) from UCB were significantly higher than those from healthy adults (9.16%±1.92%, 2.02%±0.27%, respectively, P<0.01, P<0.05). CONCLUSION: The results of Elf-1 expression level from umbilical cord blood indicate that the over-expression of Elf-1 gene in mononuclear cells and CD8~+ T cells might be one of the features of T cell immune state in umbilical cord blood.

OBJECTIVEThis study aimed at investigating effects of exogenous interleukine-10 (IL-10) on IL-6 and intercellular adhesion molecular (ICAM-1) expression in inflamed gingival tissue.

METHODSThe expression of IL-6 and ICAM-1 was examined using immunohistochemical techniques. Inflammatory gingival tissue treated with IL-10 was taken as the experimental group and the same patient's inflammatory gingival tissue without treatment of IL-10 was included into the control group.

RESULTSIL-6 expression was found mainly in monocytes, macrophages, lymphocytes, endotheliocytes and fibroblasts. The expression of ICAM-1 was found mainly in epithelial cells, monocot-macrophages, lymphocytes, endotheliocytes and fibroblasts. The immunohistochemical optical density (IOD) of the expression of IL-6 and ICAM-1 was detected by using Image-Proplus software, and the results showed that the expression in the experimental group differed significantly from that in the control group.

CONCLUSIONThe exogenous IL-10 may down-regulate IL-6 and ICAM-1 expression in inflammatory gingival tissue.

Adult ; Down-Regulation ; Female ; Gingiva ; metabolism ; Gingivitis ; metabolism ; Humans ; Intercellular Adhesion Molecule-1 ; biosynthesis ; genetics ; Interleukin-10 ; pharmacology ; Interleukin-6 ; biosynthesis ; genetics ; Male ; Middle Aged

Objective To investigate the activation of nuclear factor-B (NF-B)and hypoxia-inducible factor-1 (HIF-1)in patients with obstructive sleep apnea-hypopnea syndrome (OSAHS). Methods Subjects were classified into mild-to-moderate OSHAS group (n = 16), severe OSAHS group (n = 14) and the matched control group (n=30). Gene and protein expressions of NF-B and HIF-1 were measured by RT-PCR, Western blot in peripheral blood mononuclear cells (PBMC). Results NF-κB p65 mRNA and NF-κB p65 protein,HIF-1α mRNA and HIF-1α protein in PBMC were significantly higher in severe OSAHS group than those in mild–to-moderate group and control group (P<0.01). But there were no significant difference in NF-κB p65 mRNA and NF-κB p65 protein between mild-to-moderate group and control group (P=0.068, P=0.254 respectively). Only gene (P<0.05) not the protein (P=0.777) of HIF-1αwas higher in mild-to-moderate as compared with control group. Both NF-κB p65 mRNA and HIF-1αmRNA were positively correlated with AHI (r=0.493, P=0.006, r=0.508, P=0.004), while negatively correlated with nighttime lowest blood oxygen saturation (LSaO2)(r=-0.488, P=0.006, r=-0.46, P=0.011). There was a positive correlation between NF-κB p65 protein level and AHI (r=0.669, P<0.001). HIF-1αprotein level was positively correlated with AHI, ODI (r=0.628, P=0.001;r=0.480, P=0.018). There were positive correlations between NF-κBp65mRNA and HIF-1αmRNA (r=0.543, P=0.002), NF-κBp65 and HIF-1αprotein respectively (r=0.716, P<0.001). Conclusions As the gene and protein of NF-κB and HIF-1 were up-regulated in patients with OSAHS, and also positively correlated with the severity of sickness. We conclude that both NF-κB and HIF-1 were involed in the pathogenesis of OSAHS.

Objective:To explore the relationship between end-dialysis over-weight (edOW) in initial stage of hemodialysis and long-term prognosis in maintenance hemodialysis patients.Methods:The data of initial uremia patients receiving hemodialysis in the First Affiliated Hospital, College of Medicine, Zhejiang University from January 1, 2008 to April 30, 2017 were retrospectively analyzed. The end point of follow-up was death or until April 30, 2018. The general data including age, gender, body mass index, primary disease, complications and laboratory indicators of the patients and the related parameters of dialysis from four to twelve months were collected. Kaplan-Meier method was used to analyze survival rate. Cox multivariate regression was used to analyze the relationship between edOW and all-cause mortality and cardiovascular disease (CVD) mortality.Results:A total of 469 patients (300 males, 64.0%) were enrolled, with age of (56.9±17.1)years old. During the follow-up period of (4.1±2.4) years (1.0-10.3 years), 102 patients died. The main cause of death was cardiovascular and cerebrovascular events, accounting for 44.1% (45/102). The value of edOW was (0.28±0.02) kg. The patients were divided into edOW<0.28 kg group ( n=292) and edOW≥0.28 kg group ( n=177) according to the mean value of edOW. Kaplan-Meier survival analysis showed that the long-term survival rate in edOW<0.28 kg group was higher than that in edOW≥0.28 kg group (Log-rank χ2=4.134, P=0.043), and the CVD mortality in edOW≥0.28 kg group was significantly higher than that in edOW<0.28 kg group (Log-rank χ2=11.136, P=0.001). Cox multivariate regression analysis showed that high edOW was an independent influencing factor for all-cause death and CVD death in hemodialysis patients ( HR=1.541, 95% CI 1.057-2.249, P=0.025; HR=1.930, 95% CI 1.198-3.107, P=0.007). Conclusion:High edOW in early phase is an independent influencing factor of all-cause and CVD death in hemodialysis patients.