Objective To investigate the possible influence of immunosuppressive therapy,including sirolimus (SRL) and calcineurin inhibitors (CNI, tacrolimus), on level of Treg in the liver allo-graft recipients, Methods Forty-seven liver transplant recipients with stable liver function were assessed for at least 2 years, and divided into two groups: one composed of 15 patients receiving SRL,and another, of those receiving CNI (32 patients with tacrolimus). Thirty-eight age-matched healthy subjects (HS) were used as normal controls. We examined the expression of CD4, CD25, and Foxp3 in peripheral blood monouclear cells. Treg of every group was analyzed using Cell Quest software.Results SRL significantly increased the prevalence of CD4+ CD25high T cells, compared with HS and CNI group. The prevalence of CD4+ CD25high T cells of CNIs group was significantly lower than that of HS. The percentage of CD4+ CD25high T cells in the total CD4+ T cells was 1.88 % (1.56 %-2. 60 %), 1. 15 % (0. 57 %-l. 48 %) and 0. 84 % (0. 46 %-1. 45 %) in SRL, NS and CNI group,respectively (P<0. 01 or P<0. 05). Foxp3 was expressed in more than 95 % of CD4 + CD25high T cells and less than 20 % of CD4+ CD25low T cells, whereas not expressed in CD4+ CD25- T cells.Conclusion Different immunosuppressive therapy (SRL or CNI) might have different roles in tolerance induction in liver transplant recipients. Namely, SRL promoted the induction of allo-antigen tolerance, but CNI hampered the progression.

Objective:To clone soluble human anti-digoxin antibodies from large phage antibody library and construct a vector that expresses diabody.Methods:Soluble ScFvs were prepared through infecting E coli.HB2151 with the selected phage antibodies and induced with IPTG.The diabody vector was modified by enzyme digestion and checked by SDS-electrophoretogram.Results:It was showed by ELISA that soluble ScFv had specific binding ability to digoxin.The vector to express a soluble diabody was obtained by genetic modification,which was shown by Western blot.Conclusion:Soluble human anti-digoxin antibodies were successfully obtained from phage antibodies.The vector is efficient in creating diabody.

Objective To investigate whether Tumor Necrosis Factor-alpha (TNFα) is capable of activating Rho kinase pathway which leads to smooth muscle cell proliferation and the intervention function of Rosuvastatin, and clarify the mechanism and intervention manner of anti-atherosclerosis by Rosuvastatin. Methods Wistar neonate rat smooth muscle cells were cultured, and the activity of cell proliferation was determined by methyl thiazolyl tetrazolium (MTT). The expression of Rho kinase genes after the stimulation of TNFα was evaluated by RT-PCR. Western blot method was used to measure the protein expression of proliferating cell nuclear antigen (PCNA) after TNFα stimulation and Rosuvastatin intervention in smooth muscle cell. Results The TNFα stimulation significantly enhanced the expression of Rho kinase and increased the expression of PCNA protein in smooth muscle cells (P < 0.05). These effects were positively correlated with prolonged treatment whereas additional Rosuvastatin administration inhibited the above-mentioned effects (P < 0.05). Conclusions The activation of TNFα mediated Rho kinase signaling pathway can significantly promote smooth muscle cell proliferation, and Rosuvastatin can not only inhibit this pathway but also the induced proliferation.

Objective To investigate the application value of DFY-Ⅱ ultrasound imaging analysis software in evaluation of fetal lung maturity in different pregnancy.Methods Totally 315 cases of healthy single pregnancies (24-41 weeks) were collected.The sonographic views of fetal right side sagittal plane were obtained.The echo intensity of fetal lung and liver were analyzed and the ratio was canculated with DFY-Ⅱ ultrasound imaging analysis software.Results The ratio of echo intensity from fetal lung and liver had positive correlation with gestational weeks (r=0.94,P＜0.05),the linear regression equation was Y=0.60+0.07X (r2 =0.883).Conclusion The ratio of echo intensity by fetal lung and liver analyzed with DFY-Ⅱ ultrasound imaging analysis software can be an effective method to evaluate fetal lung maturity.

Objective To determine the expression of indole-2,3 dioxygenase (IDO) in human acute leukemia,and to investigate its correlations with clinicopathological parameters and prognosis in acute leukemia.Methods The expression of IDO in protein and RNA levels was detected by immunohistochemistry and real-time quantitative RT-PCR,respectively,and the correlations of IDO with clinicopathologic features and prognosis of acute myeloid leukemia (AML)-M5 were analyzed.Results The positive rate of IDO protein was 63.3 ％ (38/60) in human acute leukemia,while it in AML (34/49,69.4 ％),especially in AML-M5 patients (29/35,82.9 ％),was significantly higher than that of acute lymphoblastic leukemia (4/11,36.4 ％).The expression of IDO protein in healthy human peripheral blood mononuclear cells was negative.The RNA expression level of IDO in AML-M5 or non AML-M5 patients were significantly higher than that of healthy people (P ＜ 0.001),and AML-M5 patients had significantly higher IDO RNA level than that in non AML-M5 patients (P ＜ 0.05).The IDO gene expression was not correlated with sex,age and drug sensitivity,while it was closely related with these factors in the patients without complication of pulmonary infection.IDO could not act as an independent prognostic marker.Conclusion The expression of IDO in AML-M5 patients is significantly higher than that in non AML-M5 patients and healthy people.The positive expression of IDO is associated with poor prognosis of AML-M5 patients,but it is not an independent poor prognostic indicator.

In proteomic research, to improve protein solubility of membrane proteins and nuclear proteins, buffers containing high concentration of detergent, such as 4% SDS, were widely used. However, high concentration of detergent might severely interfere with the downstream proteomic analysis, including protein quantitation and trypsin digestion. To improve the proteomic compatibility of buffers with high concentration of detergent, we used short gel method to pretreat buffers containing detergent. Protein samples were first separated by a short (2-2.5 mm) SDS-PAGE electrophoresis, and proteins were quantitated by comparing with bovine serum albumin standards via optical density analysis. The gel was then cut and peptides were recovered using in-gel digestion. The quantitative linearity range of this method was 1 to 8 μg. The quantitation was accurate and reproducible. After short gel analysis, recovered peptides generated high mass spectrometry signals. In conclusion, short gel method eliminated the interference of high concentration detergent in the proteomics analysis, and it was suitable for protein samples' pretreatment, and was worth to apply in proteomic research.
Detergents ; chemistry ; Electrophoresis, Polyacrylamide Gel ; methods ; Mass Spectrometry ; Membrane Proteins ; chemistry ; Nuclear Proteins ; chemistry ; Proteins ; chemistry ; Proteomics ; methods ; Trypsin

PurposeTo explore the value of prenatal ultrasound in diagnosis of fetuses with limb body wall complex (LBWC).Material and MethodsThe ultrasound images and follow-up results were studied retrospectively in 20 cases of LBWC, and the ultrasonic features of LBWC were analyzed.ResultsAll 20 cases showed rather severe body wall defect and spinal abnormality, including 16 cases of abdominal wall defect, 4 cases of abdominal thoracoabdominal defect, 8 cases of idiopathic scoliosis, 2 cases of kyphosis, 3 cases of twist into a corner, 1 cases of distortion as S shape, and 6 cases of vertebral body arranged in disorder. Moreover, 16 cases presented abnormal umbilical cord,among which 11 had too short umbilical cord, 1 without umbilical cord, and 7 as single umbilical artery (3 cases with short cord). Five cases showed limb abnormalities, among which 2 cases were left lower limb absence, 1 cases as left upper limb absent, 1 cases as left foot varus and right foot slightly varus, and 1 cases as left foot drop.ConclusionPrenatal ultrasonography can accurately diagnose LBWC in time, so as to provide evidence for early clinical intervention.

Objective To explore the correlation of the myeloid antigen expression and clinical characteristics of acute lymphoblastic leukemia (ALL) in children.Methods The clinical data of 77 newly diagnosed ALL patients in Department of Hematology,the People's Hospital of Zhengzhou University from Jan.2010 to Dec.2013 were analyzed.The patients included 53 boys and 24 girls with a median age of 7.73 (2.00-15.00) years old.Based on flow cytometry (FCM) analysis of bone marrow,these patients were divided into 2 groups:one group included 26 patients with positive myeloid antigen expression (MyAg + ALL) and the other group included 51 patients with negative myeloid antigen expressions (MyAg-ALL).The correlation among myeloid antigen expression,clinical features,prednisone experiment,myelogram on the 15th day was analyzed through induction chemotherapy and minimal residual disease (MRD) on the 33rd day,and the rate of disease-free survival (DFS) was compared between the 2 groups.Results There were 26 cases with myeloid antigen expression among 77 patients (33.77％),CD13 + accounting for 19.48％ (15/77 cases),CD33 + 10.39％ (8/77 cases),and CD117 + 5.19％ (4/77 cases).Among these patients,there were 2 patients expressing both CD13 + and CD33 +,and 1 patient expressing both CD33 + and CD117 +.There was no difference between the MyAg + ALL group and MyAg-ALL group in gender (x2 =0.217,P =0.641),age (≥ 10 years old,x2 =0.011,P =0.918),white blood count(≥50 × 109/L,x2 =1.198,P =0.274),lactate dehydrogenase (LDH) (≥500 U/L,x2 =0.317,P =0.573),genetic abnormality (x2 =0.377,P =0.539),immunophenotype (B-ALL/T-ALL,x2 =0.397,P =0.529),and risk stratification (low-risk group,middle-risk group and high-risk group,x2 =0.260,P =0.878).Univariate Logistic regression showed that the reaction rate of prednisone experiment (P =0.023,OR =3.422) and positive rate of MRD (P =0.001,OR =0.133) of MyAg + ALL group were obviously higher than those in MyAg-ALL group.Multivariate Logistic regression showed that positive rate of MRD in CD13 + ALL group was obviously higher than that of CD13-ALL group (P =0.034,OR =120.765).The DFS rate of CD13 + ALL group and CD13-ALL group were (50.4 ± 13.8)％ and (77.4 ±6.7)％ respectively,and there was a significant difference between the 2 groups (x2 =3.928,P =0.047).Conclusions There is no significant correlation between myeloid antigen expression and clinical characteristics of children patients with ALL.For the patients with myeloid antigens,the early reaction of induction chemotherapy is bad,and for patients with CD13,the prognosis is not good.

Objective To evaluate intraoporative ultrasonography (IOUS) of liver tumors. Methods In this study, 45 patients with liver tumors were examined by the IOUS(Logiq 500 scanner with a dedicated 5 ～ 7.5 MHz linear-array intraoperative transducer), and the results of IOUS were compared with those of preoperative ultrasonography (PHILIPS HDI 5000 scanner with a 3.5 ～ 5 MHz convex-array transducer) and computed tomography (CT). Results For tumors with a diameter ≥ 1 cm, the detection rate of IOUS was 100%, which was higher than that of preoperative ultrasonography and CT (100% vs 89.80% and 97.96%), though there was no statistical significance(xc2 = 3.372 ,P >0.05 and Fisher exact probability was 1.00) ; For tumors with a diameter < 1 cm, IOUS had a significantly higher detection rate than preoperative uhrasonography and CT(90.70% vs 62.79% and 74.42%, respectively,the former x2= 9.382, the latter x2 = 3.957, both P < 0.05) ;The total detection rate of IOUS was significantly higher than that of preoperative ultrasonography and CT (95.65% vs 77.17% and 86. 96%, respectively, the former x2 = 13.378, the latter x2 = 4.381, both P < 0.05). All the detected lesions underwent surgical resection or intratumoral injection of ethanol with the guidance of IOUS. Conclusions Compared with preoperative imaging examination, IOUS not only can enhance the detection rates of preoperatively overlooked lesions, but also can provide more valuable information to guide the surgical procedures.