AIM: To study the expression of cytokines and their receptors in leukemia cell lines and normal blood cells. METHODS: RT-PCR was used to detect expression of mRNA for cytokines in leukemia cell lines(HL-60,U937,K562,HEL,DAMI,MEG-01,HUT78 and CA) and normal blood cells, including CD34 + cells, megakaryocytes,platelets, peripheral mononucleates cells and granulocytes. RESULTS: ①CD34 + cells simultaneously expressed mRNA for IL-1(?,?),IL-3, IL-6 , G-CSF, GM-CSF and their receptors and SCFR,MPL as well. The granulocytes only expressed IL-6,IL-6R,G-CSFR,GM-CSF. Megakaryocytes and platelets only expressed IL-3R,IL-6,IL-6R,MPL.Interestingly, TGF? 1 ,TNF? and their receptors sustained to express in normal cells.②Most leukemia cell lines were found to simultaneously express at least two or more stimulating cytokines and receptors ,while TGF? 1 , TNF? and their receptors were expressed in all the leukemia cell lines we observed. CONCLUSIONS: ①Multi-autocrine loops exist in leukemia cells;②Imbalance of autocrine loops of positive and negative cytokines may be related to leukemia.

OBJECTIVETo investigate the expression of zinc finger E-box binding homebox 1 (ZEB1) in the prepuce of hypospadias children and its relationship to the incidence of hypospadias.

METHODSPrepuce tissues were collected from 37 children aged 6-15 months undergoing hypospadias repair and 11 age-matched controls receiving circumcision. Based on the position of the urethral meatus, the hypospadias cases were classified as severe (n = 13) and mild-moderate (n = 24). The mRNA and protein expressions of ZEB1 were determined by immunohistochemistry and RT-PCR.

RESULTSThe expression of the ZEB1 protein was remarkably higher in the severe (100% [13/13]) and mild-moderate hypospadias patients (75.0% [18/24]) than in the controls (9.1% [1/11]), with statistically significant differences between any two groups (P < 0.05). RT-PCR showed the integrated density value (IDV) of the ZEB1 mRNA expression to be (0.67 ± 0.21), (0.81 ± 0.24), and (1.55 ± 0.29) in the control, mild-moderate, and severe hypospadias patients, respectively, significantly higher in the severe hypospadias than in the control and mild-moderate hypospadias groups (P < 0.05), but with no significant difference between the latter two (P = 0.64).

CONCLUSIONThe expression of ZEB1 is significantly increased in hypospadias patients, and its upregulation is positively correlated with the severity of hypospadias, which suggests that the overexpression of ZEB1 may contribute to the development of hypospadias.

Biomarkers ; metabolism ; Case-Control Studies ; Circumcision, Male ; Foreskin ; metabolism ; Homeodomain Proteins ; genetics ; metabolism ; Humans ; Hypospadias ; classification ; etiology ; metabolism ; Immunohistochemistry ; Infant ; Male ; Penis ; RNA, Messenger ; metabolism ; Transcription Factors ; genetics ; metabolism ; Up-Regulation ; Urethra ; Zinc Finger E-box-Binding Homeobox 1

Objective To establish a TaqMan real-time fluorescent quantitative PCR to detect Vibrio vulnificus based on hemolysin gene(vvhA)that coding cytolysin.Method By using software Primer Express, the PCR primers and TaqMan probe,which located in the conserved region of vvhA gene sequence,were designed for establishment of a TaqMan real-time fluorescent quantitative PCR to detect 100 bp amplicon from V.vulnificus DNA.A recombinant plasmid pMD19-vvhA100 as a positive control during detection was constructed using gene cloning technique.Minimal amplification cycles(Ct value)and fluorescence intensity enhancement (△Rn value)were used as observing index to optimize the reaction conditions of the TaqMan real-time fluorescent quantitative PCR.The DNAs with different concentrations from V.vulnificus and other eight bacteria and pMD19- vvhA100 were applied as templates to determine the specificity,sensitivity and reappearance of the TaqMan real- time fluorescent quantitative PCR.ICR mice were intraperitoneally,subcutaneously and orally infected with V. vulnificus,respectively.The detection effect of the TaqMan real-time fluorescent quantitative PCR was measured using the specimens of peripheral blood,subcutaneous tissue and intestinal content collected from the infected mice.Results The established TaqMan real-time fluorescent quantitative PCR showed positive results only for V. vulnificus DNA and pMD19-vvhA100.The detection effectiveness of the TaqMan real-time fluorescent quantitative PCR was as high as 0.01 ng of V.vulnificus DNA or 103 copies of pMD19-vvhA100.The SD values of the detection results repeated for three times using pMD19-vvhA100 with different concentrations were lease than 0.79. The detection results of TaqMan real-time fluorescent quantitative PCR were positive for all the specimens of peripheral blood and subcutaneous tissue.Conclusions The TaqMan real-time fluorescent quantitative PCR established in this study for V.vulnificus vvhA gene detection has advantages such as quickness,stability, sensitivity and specificity,indicating this method can be used for clinical laboratory diagnosis of septicemia and wound infection caused by V.vulnificus.

Objective To assess alternations in left ventricular volume and systolic synchrony in patients with frequent premature ventricular complexes(PVCs) from the right ventricular outflow tract(RVOT).Methods Twenty-nine patients with frequent isolated PVCs from RVOT were included and 30 healthy subjects as control.Instantaneous full-volume imaging(IFI) was performed to evaluate left ventricle volumetric parameters,including end-systolic volume (ESV),end-diastolic volume (EDV),stroke volume (SV),ejection fraction (EF),and systolic synchrony parameters,including systolic dyssynchrony index (SDI),dispersion end-systole (DISPES),mean end-systolic time (MES),pre-contraction time volume (PreContr) and post-contraction time volume (PostContr).Contraction front mapping was performed to visualize volumetric contraction sequence.All values of patients with PVCs were recorded during sinus beats (PVC-S) and premature ventricular beats (PVC-V) respectively.Results Significant differences were observed in left ventricular systolic volumetric and synchrony parameters between PVC-V and control subjects (P＜0.01),as well as in MES and PreContr between PVC-S and control subjects (P＜0.01).Conclusions Left ventricular systolic dysynchrony was demonstrated in patients with PVCs from RVOT.IFI was a novel tool to analyze left ventricular global and regional volumetric alternations.

Objective: To investigate the incidence of CMV infection(CMV-I) and CMV related diseases (CMV-D) after allogeneic hematopoietic stem cells transplantation in 70 consecutive allogeneic hematopoietic stem cells transplantation(allo-HSCT) patients and to search for the optimal prophylactic strategy.Methods: Blood samples were monitored using the CMV pp65 antigenemia assay.Of the 70 patients observed,30 patients with chronic myeloid leukemia［CML:CP(27),AP(2),BC(1)］,12 with acute myeloblastic leukemia(AML),10 with acute lymphoblastic leukemia(ALL)and other cases were NHL(3), AA(5), MDS(7), SCLC with pancytopenia (1),CLL(1), and MF (1). Sixty six patients received HLA - identical siblings transplantation and four received tranplants from their HLA- haploidentical donors. Seventy cases included allo-PBPCT (64 cases) , allo-BMT (4 cases) and allo-PB+BMT (2). Before transplantation, all patients and donors received CMV serological examination except 4 pairs of donors/recepients. All 66 patients (3 cases were CMV IgM positive) and 64/66 donors were CMV IgG positive. Results：After transplantation, 64/70 patients developed CMV viremia during monitoring period. Forty three of 70 patients developed CMV-D.Thirty five of them suffered from CMV-associated interstitial pneumonia(CMV-IP). The high peak levels of CMV antigenemia were associated with development of CMV disease　. Close correlation was found between acute graft vs host disease(GVHD) and CMV disease. The patients were followed up for 2 to 24 months. The patients who received preemptive therapy(group A)had significantly better outcome than CMV disease group(group B, P=0.0001). Conclusions: The results suggest that CMV antigenemia has high predictive value for subsequent CMV disease and CMV pp65 antigenemia -guided early therapy has particular advantage for avoiding morbidity and mortality caused by CMV disease.

BACKGROUND: Blood hemoperfusion with resin adsorption can clean larger molecules that exceed the molecular weight cutoff of combined continuous veno-venous hemofiltration (CVVH). Hence blood hemoperfusion with resin adsorption combined CVVH (HP+CVVH) has higher ability of mediator clearance, and can improve clinical outcomes in theory. This study aimed to investigate the effect of blood hemoperfusion with resin adsorption combined continuous veno-venous hemofiltration (HP+CVVH) on plasm cytokines like TNF-α, IL-1β, IL-6, cellular immunity and prognosis in patients with multiple organ dysfunction syndrome (MODS). METHODS: This was a prospective, randomized clinical trial. A total of 30 patients who had been diagnosed with MODS were enrolled in this study. Patients were randomly allocated to routine treatment+HP+CVVH group (treatment group) and routine treatment+only CVVH group (control group). In the treatment group, patients received blood hemoperfusion with resin adsorption for 2 hours, and then received CVVH for 10 hours every day. In the control group, patients received CVVH for 12 hours only every day. The patients in the two groups received blood purification therapy for three days. The plasma of patients in the treatment group was obtained at 0, 2, 12, 24, 26, 36, 48, 50, 60 hours, 5th day, 7th day and 10th day, respectively. The plasma of patients in the control group was obtained at 0, 12, 24, 36, 48, 60 hours, 5th day, 7th day and 10th day, respectively. APACHE II score, T-lymphocytes subpopulations, blood lactate acid concentration, heart rate, breathing rate, and oxygenation index were observed. RESULTS: Plasma cytokines like TNF-α, IL-1β, IL-6 decreased markedly after HP (P<0.01);T-lymphocytes subpopulations CD3+, CD4+, CD8+, CD4+/CD8+ increased after HP+CVVH or only CVVH. The plasma concentrations of TNF-α, IL-1β and IL-6 in the two groups were not markedly different at 12, 36, and 50 hours. But on the 5th day, the plasma concentrations of TNF-α, IL-1β and IL-6 in the treatment group were lower than those in the control group (P<0.05). On the 28th day, 5 patients died in the treatment group, and 6 patients in the control group. CONCLUSIONS: Both HP+CVVH and CVVH can clean plasma cytokines like TNF-α, IL-1β, and IL-6, and improve cellular immunity and clinical symptoms and signs of patients. Compared with only CVVH, the plasma concentrations of TNF-α, IL-1β and IL-6 were lower on the 5th day, and patients have an increased survival rate on the 28 day in the HP+CVVH group.

Objective To analysis the influence factors of standardization in the hierarchical chain management of type 2 diabetes and to enhance the hierarchical chain management of type 2 diabetes.Methods ( 1 ) Six hundred and ninty patients with type 2 diabetes completed 1 years management were divided into well-controlled glycosylated hemoglobin ( HbAlc ) group (＜7.0％ ) and bad-controlled glycosylated hemoglobin (HbAlc) group ( ≥ 7.0％ ).The conditions of diet,physical activity,medication,self-blood sugar monitoring and participation in health seminars were investigated and analyzed.(2) The patients were divided into standardized management group and not standardized management group.Their age,sex,educational background,occupation,monthly income per person,medical security,the course,cognition for glycuresis,two-way transfer,and chronic complications were investigated and statistically analyzed.Results ( 1 ) The proportions of physical activity (70.1％ vs 54.2％,x2=6.163,P=0.018),self-blood sugar monitoring(60.4％ vs 43.8％,x2=6.268,P=0.016) and participation in health seminars (56.0％ vs 41.7％,x2=4.577,P=0.045) in the well-controlled HbAlc group were significantly higher than those in the bad-controlled HbAlc group.(2) Their age [(61.08 ±10.04) years old vs ( 57.75 ± 9.89 ) years old,t=2.539,P=0.012],educational background ( ratio of low educational attainment:8.3 ％ vs 17.2％,x2=6.426,P=0.041 ),medical security (own expense ratios:4.6％ vs 11.5％,x2=3.543,P=0.048 ),awareness of diabetes ( ratio of poor awareness of diabetes:19.4％ vs 41.0％,x2=17.518,P=0.000 ),two-way transfer ( ratio of not transfer treatment:4.6％ vs 14.8％,x2=7.662,P=0.022) and chronic complications ( ratio of chronic complication:41.7 ％ vs 26.2％,x2=6.130,P=0.017) were significantly different between the standardized management group and not standardized management group.(3) Logistic regression analyses indicated that the age ( OR=0.954,P=0.006),monthly income per person ( OR=4.101,P=0.018 ),medical security ( OR=7.617,P=0.003 ),cognition for glycuresis ( OR=0.030,P=0.000),two-way transfer ( OR=9.079,P=0.000) and chronic complications ( OR=0.456,P=0.031 ) were the risk factors of standardized management.Conclusion We should focus on the impact factors affecting the standardized management of patients including age,monthly income per person,medical security,awareness of diabetes,ratio of not transfer treatment,positive strategies for chronic complications,improve the hierarchical chain management of type 2 diabetes,and then make the diabetic patients to early participate in standardization management of diabetes mellitus and delay the appearance of complications.

BACKGROUND: Seed cell exerting its function is required to depend on the extracellular matrix in tissue engineering, so that biocompatible material is important to be selected. OBJECTIVE: To prepare a novel composite scaffolds of chitosan/collagen/nano-hydroxyapatite (HA-CS/Col/nHAP) and to optimize the technology of tissue engineered-stents according to the circumstances of cell adhesion.METHODS: Chitosan was modified by hyaluronate acid. The structure was observed by differential scanning calorimetry and the Fourier transformed infrared spectroscopy. Three composites of HA-CS/Col/nHAP according to different ratio of chitosan and collagen solution (1: 2; 1: 1 and 2: 1) were prepared. The composite scaffolds were co-cultured with osteoblast MC3T3-E1, and the proliferation and cell growth curve were measured by CCK-8 method. RESULTS AND CONCLUSION: Hyaluronic acid and chitosan were crosslinked with amide linkage. Pore size was on the range from 50 μm to 250 μm. Porosity was increased with increased collagen level and elastic modulus, but density was reduced. Increased collagen content was beneficial for cell adhesion and proliferation on stent in the primary phase of cell co-culture. However, from day 10, no significant difference was determined among three samples. At the beginning of cell culture, cells adhered to the airspace insides the composite scaffolds. In the following days, cells grew in a colony manner, and cell-cell junction could be easily observed. These indicate that HA-CS/Col /nHAP composite scaffolds can improve the adhesion and proliferation of osteoblast. The ratio of chitosan to collagen volume at 1: 1 was optimal.

Purpose To investigate the pathological characteristics of loco-regional recurrent nasopharyngeal carcinoma ( rNPC ) . Methods Nasopharyngeal biopsy specimens of 46 rNPCs and 63 primary NPCs were collected. HE staining, immunohistochemistry and EBV small RNAs ( EBERs) in-situ hybridization were performed. Results The over-expression rates of both p63 and CK5/6 in rNPC were significantly higher than those of primary NPCs (P=0. 005, P=0. 026), while no statistical significance of Ki-67 over-ex-pression existed between the two groups ( P=0. 387 ) . More necrotic tissues, inflammatory exudates, giant bizarre carcinoma cells, desmoplastic stroma, giant bizarre tumor cells and higher degree of squamous differentiation were found in rNPCs. The carcinoma cells of 5 rNPCs were negative for both EBERs in-situ hybridization and LMP-1 immunohistochemical staining. Conclusion The loco-re-gional rNPC has two peaks of latency interval:2~5 and 9~11 years. The loco-regional rNPC cells have higher degree of squamous differentiation with higher expression of p63 and CK5/6, as well as more invasive ability. In addition, both EBERs in-situ hybridization and LMP-1 immunostaining are negative in 10. 87% (5/46) of loco-regional rNPC.

Objective To prepare a novel bioactive and degradable scaffold with mineralized collagenpolyose based composite by biomimetic synthesis for bone tissue engineering and explore the compatibility of osteoblast culturing on the scaffold.Methods Using the cross-linking product of collagenⅠ and sodium hyaluronate as the template,the calcium phosphate was deposited on it to produce a mineralized composite.The 3-D porous scaffolds were prepared by liquid phase separation after the mineralized composite combining with polylactic acid (PLA) and NaCl.The materials and scaffolds were investigated by x-ray diffraction (XRD),scanning electronic microscopy (SEM) and universal testing machine.In addition,inverted microscope,fluorescence microscope,SEM,Cell Counting Kit-8 assay and alkaline phosphatase (ALP) assay were introduced to analyze the growth,function and compatibility(morphology,proliferation and differentiation ) of osteoblast-like cell on the scaffolds.Results The degree of hydroxyapatite (HA) crystals in the composite was low and the size was tiny,which were similar to that of nature bone.The SEM micrographs showed that the scaffolds possessed 82% of porosity and the pore size was about 200 μm to 650 μm.Cells on the scaffolds spread well and presented a high proliferation rate and differentiation level.Conclusion The novel scaffolds are simiar to nature cancellous (spongy) bone both on structure and in property and might be used as one of the optimal scaffolds material for bone tissue engineering.