AIM:To investigate the differentiation from human mesenchymal stem cells(hMSC) into neuron-like cells.METHODS:hMSC were separated from rib marrow with Ficoll-Paque reagent and expanded in culture medium. hMSC were induced to differentiate with DMEM/monothioglycerol or DMEM/β-mercaptoethanol, respectively. Neuron-specific enolase(NSE), neurofilament(NF), and glial fibrillary acidic protein(GFAP) were detected by immunohistochemistry. RESULTS:hMSC were expanded as undifferentiated cells in culture for more than 5 passages. When treated with monothioglycerol or β-mercaptoethanol for 5 hours, hMSC exhibited neuronal phenotype. The expression of NSE and NF in the neuron-like cells was positive, but the glial astrocyte marker GFAP didn't express. CONCLUSION:hMSC can be induced to differentiate into neurous.

ObjectiveTo evaluate analytical performance of NT-proBNP on Electro-Chemiluminescence Immunoassay system.MethodsThe precision,accuracy,limit of blank ( LoB ),limit of detection (LoD),functional sensitivity (FS),analytical measure range (AMR),maximal dilution rate,clinical reportable range(CRR) and the analytical anti-interference ability of NT-proBNP were evaluated according to EP documents issued by CLSI and related references.The analytical performance data were compared to quality standards declared by the manusfacturers.According to CLSI C28-A2,80 healthy volunteers,aged from 18 to 74, were chosen and divided into 4 groups on average for biological reference intervals verification.Results The within-run CV and total CV were 1.1％ -2.2％and 1.5％ -2.9％ respectively.The deviations from controls distributed by National Center for Clinical Laboratory and affiliated calibrators were 2.7％ -5.9％ and 2.7％ -7.5％,respectively.The results of LoB,LoD and FS were 2.5,7.8 and 8.8 pg/ml,respectively.AMR was 8 -35 126 pg/ml,and the most suitable dilution rate was 1∶ 2,so the CRR was 9 -70 252 pg/ml.428 μmol/L bilirubin,2 g/L haematoglobin and 2 200 FIU chyle didn't interfere with the NT-proBNP assay.Moreover,almost all the data from different age groups were in the range of biological reference intervals declared by the manusfacturers, except one test data (167 pg/ml).Conclusions The analyticalperformance of NT-proBNP analyzed on Roche Cobas E601electrochemiluminescence immunoassay systemisconsistentwiththestandarlswhichmanufacturershas proclaimed.The establishment of LoD,FS,maximal dilution and CRR for NT-proBNP assay could provide the quality assurance for clinical use and the biological reference intervals declared by manusfacturers could meet the clinical needs.

Objective To determine whether pulmonary arterial hypertension(PAH) and pulmonary venous hypertension (PVH) can be differentiated noninvasively by echocardiography. Methods Fifty-six patients with pulmonary arterial systolic pressure(PASP) ≥40 mmHg by echocardiography were involved,and cardiac catheterization performed within 7 days of each other. Based on left ventricular end-diastolic pressure or pulmonary capillary wedge pressure(PCWP) ,30 patients were classified as PAH group and 26 patients as PVH group. The early(E) and late(A) diastolic mitral inflow velocities,E/A ratios,deceleration time(DT),early dastlic mitrial annular velocity(E') and E/E' ratios were measured by conventional and Doppler tissue imaging echocardiography in the two groups. Results Compared with PVH group,the PAH group had significantly higher A,DT,PASP and E',and significantly lower E,E/A ratio and E/E' ratio (P < 0. 01 or P <0. 001). E/E' and E/A ratio was optimal indexes for differentiation of PAH and PVH,the area under receiver operating characteristic curve was 97% and 91 %, respectively. Optimal cutoff for diagnosing PVH was E/E'>9.2 (sensitivity 95%, specificity 97%) and E/A> 1. 7 (sensitivity 75%,specificity 92%). Conclusions PAH and PVH imaging can be reliably differentiated by echocardiography.

AIM: To investigate the differentiation from human mesenchymal stem cells (hMSC) to adipocytes.METHODS: hMSC were separated from rib marrow and expanded in culture medium. To detect the surface antigens, the labeled cells were analysed on a FACScan flow cytometer. hMSC were induced with dexamethasone, insulin, 1-methy1-3-isobutylxanthine and indomethacin which acted as adipocyte differentiation inducer. The cells were stained with Oil Red O. The number of adipocytes were counted on a phase-contrast microscope.RESULTS: hMSC were expanded as undifferentiated cells in culture for more than 5 passages. The isolated cultured MSC comprised a single phenotypic population and displayed a fibroblast-like morphology. These expanded, attached MSC were uniformly positive for CD29, CD44, CD90, CD105, CD166 and didn't express CD14, CD34, CD45, CD11a. After induced with induction medium, lipid vacuoles were first detectable within the cells at 48 hours. Two weeks later, more than 85% MSC differentiated into adipocytes which displayed a perinuclear accumlation of lipid vacuoles, as detected by Oil Red O. CONCLUSION: hMSC can be induced to differentiate into adipocytes. [

AIM: To investigate the differentiation from adult rat and human bone marrow mesenchymal stem cells (BMMSCs) into neuron with musk polypeptide (Mu-P).METHODS: Adult rat and human BMMSCs were induced with Mu-P.Neuron-specific enolase (NSE),neurofilament (NF),Nestin,glial fibrillary acidic protein (GFAP) were detected by immunohistochemistry.RESULTS: Simple methods with Mu-P induced adult rat and human BMMSCs exhibiting a neuronal phenotype,expressing Nestin at 3 hours to 5 hours,and expressing NE and NF at 5 hours to 7 days.But the neuron-like cells didn't express the glial astrocyte marker GFAP.CONCLUSION: Adult rat and human BMMSCs can be induced to differentiate into neurons with Mu-P.

AIM: To investigate the differentiation from human mesenchymal stem cells(hMSC) into neuron-like cells by the increase in intracellular cyclic AMP. METHODS: hMSC were separated from human marrow with Ficoll-Paque reagent and expanded in culture medium. hMSC were induced to differentiate into neurons with Forskolin and 3-isobutyl-1-methyl-xanthine (IBMX). Neuron-specific enolase(NSE), neurofilament(NF), glial fibrillary acaidic protein(GFAP) were detected by immunohistochemistry. RESULTS: hMSC were expanded as undifferentiated cells in culture for more than 10 passages. Forskolin/IBMX can induce hMSC to exhibit a neuronal phenotype, expressing NSE and NF-M in 5 hours. But the neuron-like cells didn't express the glial astrocyte marker GFAP. CONCLUSION: hMSC can be induced to differentiate into neurons by increase in the intracellular cAMP.

AIM: To investigate the differentiation potential of human bone marrow mesenchymal stem cells (hBMMSCs) into hematopoietic cells in vivo. METHODS: hBMMSCs prepared from the bone marrow-aspirate sample obtained from healthy human donors were culture-expanded in vitro with 5-8 passages. hBMMSCs(P5-8, 4.8?10 5 cells/mouse) were injected into the severe combined immuodeficiency (SCID) mice treated by cyclophosphamide(CPA) and various tissues were analyzed at 35 days post-transplant for the presence of differentiated human cells. RESULTS: hBMMSCs(P5-8) viability, which was determined by typan blue staining at the end of the harvest and before infusion, was greater than 95% in every infusate at both time points. Cells characterized by flow cytometry using human MSC-specific monoclonal antibodies were uniformly positive for CD29, CD44, CD90, CD105, CD106, CD166 and negative for CD11a, CD14, CD34, CD38, CD45, CD80, CD86 which are common on cells of the hematopoietic lineages. Analysis of PB demonstrated that 5 of 6 hBMMSCs transplanted SCID mice had low level of circulating human CD45 +/ H-2D d- cells(range from 0.17% to 0.36%)and CD34 +/ H-2D d- cells(range from 0.10% to 0.50%). Analysis of BM for the presence of hematopoietic chimerism demonstrated human CD45 +/ H-2D d- cells and CD34 +/ H-2D d- cells in the marrow of 4 out of 6 hBMMSCs transplanted SCID mice (0.10%-0.19% and 0.03%-0.52%, respectively). Human hematopoietic cells with these same phenotype were also detected in the spleen 4 of the hBMMSCs transplanted SCID mice (range from 0.19% to 1.65% ,from 0.20% to 0.26%, respectively). No human hematopoietic cell was seen either in the PB, BM or spleen of all control animals. CONCLUSION: hBMMSCs have the ability to differentiate into blood cells of multiple lineages, including CD34 + hematopoietic stem cells/progenitor cells (HSC/HPC).

AIM: To induce lymphoid stem cells and/or T-cell precursors to diffe rentiate into functional mature T lymphocyte, and to increase the surface marker of T lymphocytes such as CD + 3, while embryonic stem(ES) cells differentiate d into hematopoietic stem/progenitor cells(HSPCs) in vitro . When they were i njec ted into lethally irradiated mice, these differentiated cells had the advantage in immune reconstitution. METHODS: Embryonic stem cells formed e mbryoid bodies(EBs) in the medium containing methycellulose, hematopoietic growt h factors(HGFs) was added to the culture system on the 6th day, thymopeptide was added at the same time. Flow cytometry were performed to detect the surface mar ker CD 34 and CD 3 of the differentiated cells. Finally the differentiated cells were injected into lethally irradiated mice, 60 days later, the incidence rate of graft versus host disease(GVHD) was taken as the mark of cell mediated immunity, PCR was performed to detect the sex determining region of the Y-chromo some(Sry) in bone marrow cells and spleen cells of the survival host female mice . RESULTS: The percentage of CD + 3 T lymphocytes was 10.52% a nd the incidence rate of GVHD was 0% on the 13th day, but they respectively rose up to 22.93% an d 100% if thymopeptide was added in the procedure of inducing ES cells to differ entiate into HSPC in vitro . CONCLUSION: The quantity of CD + 3 T lymphocytes increased in medium containing thymopeptide when ES cells differe ntiated into CD 34 + HSPC.

AIM: To investigate multi-potential of rat bo ne marrow mesenchymal stem cells (rBMMSC) and mutation inclination, the rBMMSC w ere long passaged in vitro. METHODS: Cellular cycles of diff erent passages were assayed by FA CSan flow cytometry and karyotypes of passage 6, passage 25 and passage 45 were compared by G-binding analysis. RESULTS: The early passages and long-term passages all showed st rong proliferation; passage 6, passage 25 and passage 45 all showed normal karyo type. CONCLUSION: Long-term culture and passage of rBMMSC still remain s strong proliferation. With this capability, the mutation inclination is not enhanced.

AIM: To observe the relationship between the immune deviation of Th1 and Th2 cell clones and class Ⅱ major histocompatibility complex (MHC) antigen expression in different stages of acute rejection in transplanted hearts. METHODS: Heart transplantation were performed in rats.Isografts and non-transplanted animals were used as control group. Donor class II MHC antigen expression were detected with monoclonal antibodies and immunostaining technique and the amount of type Ⅰ and Ⅱ cytokines mRNA expression were detected by semiquantitative RT-PCR in cardiac allografts. RESULTS: Myocardial IL-2 mRNA and donor class Ⅱ MHC antigen expression were significantly in creased, accompanied with development of acute rejection( P