OBJECTIVEThis study is performed to investigate the cell topographies and biomechanical properties of two different types of temporomandibular joint (TMJ) discs from goats by using JPK Nano Wizard 3 biological atomic force microscopy (AFM). This process provides a guideline for selecting seed cells for TMJ disc tissue engineering.

METHODSTMJ disc cells from primary goats were cultured by monolayer culture method. AFM was used to contact scan the topographies of the two types of TMJ disc cells under physiological environment. Approximately 20 chondrocyte-like and fibroblast-like cells were selected randomly to plot the force-versus-distance curves of the cytoplasm and nucleus. Young's modulus and adhesion were analyzed by JPK Data Processing.

RESULTSThe triangle-shapednucleus of the chondrocyte-like cell occupied a large portion of the cell. Cytoskeleton was arranged dendritically on the surface. Pseudopodia were extended from cell edges. The spindle-shaped nucleus of the fibroblast-like cell occupied a significantly larger region compared with the cytoplasmic region. Cytoskeleton was arranged regularly. Cell edges were smooth with less pseudopodia extended. No difference was found in the surface roughness between the two types of cells. According to the force-versus-distance curves, the Young's moduli of the two types of cells were not statistically different (P>0.05), but differences were found in the cytoplasmic regions (P=0.047). No statistical difference was found in the adhesions between the two types of cells (P>0.05).

CONCLUSIONThe AFM topography and curves were compared and analyzed. The two types of TMJ disc cells exhibited significantly different topographies, but only slight difference in their mechanical abilities.

Objective To evaluate the values of CD64 expression in diagnosis of infected patients referred to intensive care unit.Method Sixty febrile children referred to the hospital intensive care unit from 2009.11 to 2010.03 were enrolled for a retrospective study.Fever was defined as a body temperature reaching 38℃ or higher with specifically bacterial infection or highly suspected with bacterial infection or viral infection.There were 28 patients with bacterial infection and 32 with viral infection.The non-infectious diseases such as juvenile rheumatoid arthritis and Kawasaki disease were excluded.The controls were 50 healthy children asking for physical examination.On admission,CD64 were measured by using flow cytometry,and blood routine examination,ESR,PCT,blood cultures and sputum cultures were simultaneously detected in all febrile patients.Data were statistically analyzed by using SAS 16.0 software.Data are given as means±SE.Categorical variables were analyzed using X2 test and continuous variables were compared by applying paired 1-tailed t test,Significance level was set at less than 0.05.Results of them,57.1%bacterial infection patients and 71.9%viral infection patients contracted pneumonia.CD64 in bacterial infection patients、viral infection patients and the subjects of control group were(12.6±9.7),(5.4±2.42)and (2.9±0.77),respectively.The CD64 in the bacterial infection patients were significantly higher than those in the virus infection patients(F=11.002,P=0.004).Conclusions CD64 in infected children referred to a hospital intensive care unit can be clearly distinguished between bacterial infections and viral infections, providing an important guidance and a flexible strategy for clinical treatment and determine the timing of withdrawal.

BACKGROUND:Our preliminary studies have shown that basic fibroblast growth factor can induce the differentiation of bone marrow mesenchymal stem cells into disc cells of the temporomandibular joint, and for basic fibroblast growth factor, 10μg/L is superior to 5μg/L in col agen synthesis.
OBJECTIVE:To observe ultrastructural changes of bone marrow mesenchymal stem cells after being induced by different concentrations of basic fibroblast growth factor.
METHODS:We cultured primary sheep bone marrow mesenchymal stem cells and selected passage 3 and 4 cells at good growth state. Bone marrow mesenchymal stem cells were stimulated with 5 and 10μg/L basic fibroblast growth factor and their growth state was observed under inverted phase contrast microscope. Uninduced cells served as controls. The slides with cellcrawling pieces were stained with Safranin O, picrosirius and type I col agen immunohistochemistry at days 7, 14 and 21, respectively. Simultaneously we col ected the cells at day 21 to observe the ultrastructural changes of bone marrow mesenchymal stem cells.
RESULTS AND CONCLUSION:After being induced with different concentrations of basic fibroblast growth factor, bone marrow mesenchymal stem cells were able to differentiate into disc cells of the temporomandibular joint;and after being induced with 10μg/L basic fibroblast growth factor, cells were more like fibroblast-like cells of the temporomandibular joint disc. These findings indicate that bone marrow mesenchymal stem cells have morphological basis for differentiation to the fibroblast-like cells of the temporomandibular joint disc.

Objective To investigate CD38 expression characteristic and the relation of clinic prognosis in children with acute lymphoblastic leukemia,in order to improve individual treatment.Methods Seventy-nine patients with childhood acute lymphoblastic leukemia(B-lineage) were enrolled into this study.Four-color fluorochrome labeled monoclonal antibodies were applyed to analyze the cell immunophenotypes and minimal residual disease screening.When CD38 low-expression was considered to be the effective screening marker and be used to continue monitoring.All patients were divided into CD38 low-expression groups and CD38 high-expression groups,to compared the immunophenotyping characteristic,risk stratification and survive rate of the two groups.All datas were assessed by means of SPSS16.0 and a P value of 0.05or less was considered to indicate statistical significance.Results All of 79 newly diagnosed ALL-B,The group of CD38 low-expression were 50/79 (63.3％) while the other group were 29/79(36.7％).of all patients,11 chilldren showed only a screening indicator-CD38/CD10/CD34/CD19,while 46 belonged to more than one markers (Such as TdT/CD10/CD34/CD19,CD66c/CD10/CD34/CD19 and CD45/CD10/CD34/CD19) and 18 no markers.The stratification of CD38 low-expression and CD38 high-expression groups as follows:21/5 patients with low-risk,14/15 with medium risk and 15/9 with high-risk.In the CD38 low-expression group,Early Pre-B 33,Pre-B 12,Mature-B 5,while in the CD38 high-expression group,Early Pre-B 21,Pre-B 5,Mature-B 3.This study showed that the high-risk stratification in the CD38 high-expression group was obviously higher than the CD38 low-expression group(F=6.24,P=0.044),but the survival time was signicantly shorter than CD38 low-expression group (x2 ＝ 5.22,P =0.022) and the difference was statistically significant.Conclusion CD38 as a MRD monitoring indicator of most acute lymphoblastic leukemia when it low-expression,CD38 high-expression in newly diagnosis childhood acute lymphoblastic leukemia(B-lineage) may be an independent risk factor for predicting poor prognosis.

Objective To analyzed the expression and clinical characteristics of CD 20 marker in children with B-lineage acute lymphoblastic leukemia ( B-ALL) and evaluated its medical significance in assessing the prognosis of disease.Methods From November 2008 to July 2012,125 cases of children with B-lineage acute lymphoblastic leukemia were collected from Shanghai Children ′s Hospital,including 79 males and 46 females, aged between 2 months to 14 years old.Flow cytometry based immunophenotyping and Minimal Residual Disease ( MRD) screening were applied to these children when newly diagnosed ,and MRD monitoring was again carried out after 35 days of induction remission therapy to those bears the MRD markers.These 125 patients were divided into CD20-positive group and CD20-negative group, and the corresponding clinical characteristics ,stage of immunophenotype ,MRD,risk stratification,and overall survival rates were recorded and compared.Data were statistically analyzed by using SPSS 16.0 software including χ2 test,t-test,standard deviation test and survival test.Results A total of 125 children with ALL-B,the group of CD20-positive were 48 while CD20-negative groups were 77,with a median age of 6 years old,and the median follow-up time of 30 months.Multivariate Cox regression Analysis showed that there was no clear correlation between CD20 expression level with age ,sex,white blood cell count at diagnosis ,fusion-gene,the stage of immunophenotype as well as risk stratification.The MRD-positive incidence at 35 days in the CD20 positive group was 35.4%,much higher than that of the CD20-negative group (16.9%),which is statistical significance (χ2 =5.236,P<0.05),while the overall survival rate (OS) for the CD20 positive group is 75.0%,much lower than that of the CD20 negative group (84.4%,χ2 =4.160,P<0.05).Conclusions CD20 positive expression level in children with B-lineage acute lymphoblastic leukemia at diagnosis demonstrates negative correlation with the overall survival rate of the patient ,indicating its usefulness as an additional joint marker for the current regimens to incorporate CD 20-targeted monoclonal therapy.

ObjectiveTo understand the pathological and physiological roles of Presenilin 1 (PS1) in Alzheimer's disease (AD) recurrence, and the interaction between PSI and carhoxyl terminus of Hsc70 interacting protein (CHIP). MethodsThe yeast two-hybrid system was applied to identify a novel PS1 interacting protein as CHIP. After pGBKT7-PS1-C203 bait plasmid and full fragement CHIP of pACT2-CHIP expression vector were constructed, the interaction between PSI and CHIP was tested by β-galactosidase assay, pGBKT7-PS1-C203 was co-transfected with pACT2-CHIP into 293T cells and the interaction between PS1 and CHIP was tested by co-immunoprecipitation and Western blot. ResultsSpecificity of the interaction between PS1 and CHIP was identified by β-galactosidase assay and co- immunoprecipitation. ConclusionsCHIP is able to modulate chaperone functions and the pathway of protein ubiquitination/degradation. CHIP may regulate a proper assembly of the γ-secretase complex through its interaction with PSI, which is helpful to elucidate the mechanism of AD pathology.

In order to investigate the levels of stem cell factor (SCF) and its receptor c-kit protein and mRNA in pediatric aplastic anemia (AA) and their relevance to the pathogenesis, immunocytochemical and in situ hybridization were utilized to detect the expression of SCF and its receptor c-kit gene protein and mRNA, respectively in 59 children with AA and 51 normal controls. The relationship between SCF and c-kit and the pathogenesis of AA was analyzed subsequently. The results showed that the positive rate of SCF protein and mRNA expression in children with AA was significantly lower than that in healthy controls (P < 0.05). However, there was no significant difference in the positive rate of c-kit protein and mRNA expression between children with AA and control group (P > 0.05). It was concluded that the expression of SCF is significantly decreased in children with AA, which may be closely associated with the pathogenesis of the AA. c-kit may be unrelated to the development of pediatric AA. Therefore, AA in children may have abnormalities at SCF/c-kit signal transduction levels.
Anemia, Aplastic/etiology ; Anemia, Aplastic/*metabolism ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics ; Receptors, Colony-Stimulating Factor/*biosynthesis ; Receptors, Colony-Stimulating Factor/genetics ; Stem Cell Factor/*biosynthesis ; Stem Cell Factor/genetics

Objective Using chemically synthesized small interfering RNA (siRNA) transfected HepG2.2.15 cells to construct a cell model in interfering hepatitis B virus (HBV) X gene, studying the inhibi-tion of HBV replication and antigen expression in vitro. Methods After transfection of HepG2.2.15 cell for 24 h, 48 h, 72 h, detecting the cell supernatant of HBsAg and HBeAg by chemiluminescence immunoassay, the cell supernatant HBxAg protein by ELISA , the HBx mRNA relative expression of transfected cell was detected by fluorescence quantitative polymerase chain reaction (PCR), the ability of cell proliferation was detected by CCK-8 assay. Results After HBx-siRNA transfected HepG2.2.15 cells, cell proliferation ability was inhibited. The cell of HBx mRNA and the cell supernatant of HBxAg expression decreased (P < 0.05); at the same time it in-hibited the expression of HBsAg and HBeAg. The suppressed peak and the inhibition rate were 66% and 58%respectively at 72 h. The fluorescence quantitative PCR confirmed that expression of HBV DNA in the super-natant was decreased. Conclusion The HepG2.2.15 cell interference model of HBV X gene has been success-fully constructed, which has an effect of inhibiting proliferation of HepG2.2.15 cells and replication and expres-sion of HBV gene in vitro.

Objective To evaluate the efficacy of different treatment regimens for children with acute promye-locytic leukemia (APL)with positive PML -RARa fusion gene.Methods Thirty -two newly diagnosed APL patients were included in this study,treated either with all -trans -retinoic acid (ATRA)and chemotherapy (CT)(group A) or with ATRA and arsenic trioxide (ATO)(group B).Clinical situation and clinical efficacy were analyzed in patients in different groups.They were also separated into low risk group,intermediate risk group and high risk group according to different risk criteria.Clinical characteristics,complete remission,long -time survival and urine arsenic concentra-tion were analyzed and compared.Results (1 )Fourteen of 1 5 patients (93.3%)in group A achieved hematological complete remission (HCR)with a median time of 38 days (28 -63 days).Sixteen of 1 7 patients (94.1 %)in group B achieved HCR with a median time of 29 days (1 0 -42 days),which was significantly shorter than group A,and there was a significant difference between 2 groups(t =3.53,P =0.002).(2)The 5 -year event -free survival (EFS)of group A and group B was (60.0 ±1 2.6)% and (81 .9 ±9.5)%,respectively;the 5 -year EFS of group B was almost 20% higher than group A;while there was no significant difference between the 2 groups(χ2 =1 .1 5,P =0.28).The 5 -year overall survival (OS)of group A and group B was (72.2 ±1 1 .9)% and (94.1 ±5.7)%,respectively,the 5 -year OS of group B was almost 20% higher than group A;while there was no significant difference between the 2 groups(χ2 =2.88,P =0.1 6).(3)The 5 -year EFS of low plus intermediate group and high risk group patients was (74.0 ±1 0.1 )% and (64.8 ±1 4.3)%,the 5 -year EFS of low plus intermediate group was almost 1 0% higher than high risk group,but there was no significant difference between the 2 groups(χ2 =0.1 4,P =0.71 ).The 5 -year OS of low plus intermediate group and high risk group patients was (84.7 ±8.1 )% and (71 .3 ±1 4.1 )%,the 5 -year OS of low plus intermediate group was almost 1 0% higher than high risk group,while there was no significant difference be-tween the 2 groups(χ2 =0.36,P =0.55).(4)ATO related side effects were mild,including abnormal liver tests and e-lectrocardiogram,but were invertible after supportive therapy.At the end of each chemotherapy course,the urine arsenic concentration remained low and no chronic arsenic toxicity or second malignancies were found during the follow -up period.Conclusions The ATRA plus ATO regimen is a promising and better treatment for childhood APL with positive PML -RARa fusion gene compared with conventional chemotherapy.It was necessary to take risk stratification in APL patients.

A 48-year-old female presented with a 6-year history of papules and plaques all over the body and with 1-year history of blurred vision in the right eye.Physical examination showed porcelain-white atrophic papules with peripheral erythematous halo and telangiectnsia.She also suffered from exotropia,visual deterioration,visual field defects of the right eye,as well as numbness of the left index finger,thumb and right anterior tibia.Skin biopsies of abdominal lesions revealed dermal necrosis with mucoid degeneration,inflammatory infiltration predominated by lymphocytes around several small blood vessels and occlusion of some blood vessels in deep dermis.Colonoscopy of the whole colon demonstrated scattered patches of hyperemia and erosions with the formation of shallow ulcers.Nerve electromyologram revealed damage to the nerves of right quadriceps femoris muscles.Fecal analysis showed that occult blood was strongly positive.A diagnosis of malignant atrophic papulosis was made based on the characteristic clinical presentation,laboratory and histopathological findings.She was treated with dipyridamole and aspirin for three months,which resulted in no clinical improvement or deterioration.