AIM: To explore the ex vivo expansion characteristics of the endothelial progenitor cells (EPCs). METHODS: CD34+ cells were selected from umbilical cord blood mononuclear cells (MNC) by MiniMACS system, expanded at the same conditions as that for total MNC, coincubation of CD34+ and CD34- from the same donation for EPCs. In addition, we tested the effect of vessel endothelial growth factor (VEGF) and passage on cell differentiation, expansion kinetics and apoptosis. EPCs were determined and quantified by immunocytochemistry and flow cytometry. RESULTS: Coculture of CD34+ and CD34-,total MNC led to a significant increase in the expansion of CD34+ cells compared with CD34 enrichment (P0.05). These differentiated EPCs were stained positive for CD34+, von Willebrand factor (vWF), KDR, CD31 and incorporate acetylated low-density lipoprotein (LDL). CD34+ and AC133+cells accounted for 68.2%?6.3% (n=6) and 57.2%?9.8% (n=6) of attaching (AT) cells at day 7 of culture, respectively. CONCLUSIONS: Coculture of CD34+ and CD34- or culture of MNC enhances ex vivo expansion of EPCs. Early passage decreases apoptosis rate, VEGF has no significant effect on ex vivo expansion of EPCs.

The characteristics for the ex vivo expansion of the endothelial progenitor cells (EPCs) were explored. CD34+ cells were selected from umbilical cord blood mononuclear cells (MNC) by MiniMACS system, expanded under the same conditions as those for total MNC, coincubation of CD34+ and CD34- from the same donor for EPCs. In addition, the effects of vessel endothelial growth factor (VEGF) and passage on cell differentiation, expansion kinetics and apoptosis were examined. EPCs were determined and quantified by immunocytochemistry and flow cytometry. The results showed that both coculture of CD34+ and CD34- and total MNC led to a significant increase in the expansion of CD34+ cells as compared with CD34 enrichment (P < 0.05). There was a tendency toward decreased apoptosis in cultures when early passage was performed immediately after cord like structures appeared. VEGF had no significant effect on apoptosis (P > 0.05). These differentiated EPCs were positive for CD34+, von Willebrand factor (vWF), KDR, CD31 staining and phagocytized acetylated low-density lipoprotein (LDL). CD34+ cells accounted for (68.2 +/- 6.3)% of attaching (AT) cells at day 7 of culture. It was suggested the most efficient method to ex vivo expansion of EPCs was coculture of CD34+ and CD34- or total MNC. Early passage makes cell apoptosis rate decrease. VEGF had no significant effect on ex vivo expansion of EPCs.

Objective To examine whether the increase of carbon monoxide (CO) induced by oral methylene chloride (MC) administration in recipients before heart transplantation would protect heart grafts against isehemia/reperfusion (I/R) injury associated with transplantation and to explore the possible mechanism.Methods Inbred male Balb/c mice were used as donors and recipients to establish cervical heart transplantation model Recipients were treated with either MC (100 mg/kg or 500 mg/kg,per os)(group MC 100 mg,n=10;group MC 500 mg,n=12) or olive oil(0.15 ml,per os.group olive,n=10) 3 h prior to anesthesia.Age-matched norwlal mice served as controls (group N,n=5).The serum COHb and the CO content of myocardial tissue were measured at 0,1,3,6,12,24 h after oral MC administration.Half of recipients were killed at 3 and 24h after transplantation for senum or cardiac graft samples.The serum cTnI levels,the mRNA levels of TNF-α,IL-10,Bcl-2,Bax.the protein levels of NF-κB and the ultrastructures of myocardium were examined.Results As tompared with group olive.the serum COHb and tissue CO were increased significantly and peaked within 3 h in group MC 100 mg and group MC 500 mg.The serum cTnI levels in group MC 100 mg and group MC 500 mg were significantly decreased (P<0. 01 ), especially in group MC 500 mg. The increase of CO in recipients of group MC100 mg and group MC 500 mg significantly inhibited the proinflammatory gene expression of TNF-α mRNA and the pro-apoptotic gene expression of Bax mRNA (P<0. 01), and increased the anti-apoptotic gene expression of Bcl-2 mRNA (P<0. 01), but did not increase the anti-inflammatory gene expression of IL-10 mRNA (P>0. 05) in the heart grafts. As compared with group N, the myocardial NF-κB activation was increased significantly in group olive,group MC 100 mg and group MC 500 mg (P<0. 01 ), but there was no significant difference among the later three groups (P>0. 05). The myocardial ultrastructure was also alleviated significantly in group MC 100 mg and group MC 500 mg as compared with group N. Conclusion The increase of CO induced by MC in recipients suppresses pro-inflammatory and pro-apoptotic gene expression and efficiently ameliorates transplant-induced heart I/R injury. The possible mechanism does not seem to be associated with down-regulation of the NF-κB signaling pathway.

Objective To explore the impact of hypoxia/reoxygenation stimulation on phenotype and immune activity of dendritic cells(DCs) cultured from murine bone marrow. Methods Mouse DCs were generated from bone marrow cells and were divided into control group and hypoxia/reoxygenation group. DC in control group was cultured at normal condition, and in hypoxia/reoxygenation group was cultured at hypoxic condition for 4 h followed by cultured at normal condition for 24 h. Flow cytometry and mixed lym-phocyte reaction(MLR) was used to detect the phenotype and functional properties of DCs. ELISA was used to detect the concentration of TNF-α, IFN-γ and IL-12 in the supernalant, Imrounochemistry was used to de-tect the concentration of NF-κB. Results Hypoxia/reoxygen stimulation increased the CD80, CD86, MHC Ⅱ in the cytomembrane of DCs and TNF-α, IFN-γ, IL-12 concentration in the supernalant. Hypoxia/reoxy-gen stimulation also promoted the shift of NF-κB to karyon. Conclusion Under hypoxia/reoxygen stimula-tion, DCs express high level of surface molecules, and possess strong immune activity.

AIM: To investigate the expression of heat shock protein 60(HSP60) and Toll-like receptor 4 transduction system in mouse cardiac transplantation. METHODS: The mouse cervical heart transplantation model was established. The animals were divided into control group (the donor and recipient were all C57BL/6 mice) and experimental group (the donor was BALB/c mice and recipient was C57BL/6 mice). The heart and blood were collected for study at 3 d and 7 d. The pathological analysis of the hearts was performed. The levels of cytokines in the serum were determined using ELISA. The expression of HSP60, TLR4, MyD88 and NF-κB in the myocardium was determined by immunohistochemistry and Western blotting. RESULTS: The expression levels of HSP60, TLR4, MyD88 and NF-κB were higher in experimental group than those in control group. Severe rejection was observed in experimental group, whereas no distinct rejection in control group was found. The cytokines (TNF-α, IFN-γ, IL-12) increased significantly in experimental group as compared to those in control group. CONCLUSION: HSP60 increases significantly after heart transplantation, which may activate Toll-like receptor 4 transduction system in a MyD88-dependent pathway and promote allograft rejection. Regulation of HSP60 signal transduction may be a novel way for treating allograft rejection.

Objective Pentamethylquercetin (PMQ) has a role in cardiovascular protection. We investigate the effects of 3,3' ,4' ,5,7-pentamethylquercetin, a derivative of PMQ, on intimal hyperplasia of the vein grafts in rats both in vivo and in vitro. Methods The proliferation of vascular smooth muscle cells ( VSMC ) was induced with Ang Ⅱ (0. 1μmol/L, 24 h)while PMQ was administrated at six different dosages (0. 1, 0.3, 1,3, 10 and 30 μmoL/L). Cell viability was identified with MTT; ROS was measured with DCFH-DA; and the expression of NADPH oxidase subunits Nox1, p47phox, and p22phox mRNA were measured with real-time PCR. For the experiment in vivo, 24 SD rats were randomly assigned to control group and PMQ groups, the latter was further divided into three different dosage groups. In the control group, solvent was administrated daily via gavage. In PMQ groups, PMQ ( 12.5 mg/kg, 25 mg/kg, 50 mg/kg) was administrated daily respectively in the same way.All SD rats received operation performed by one person. Reversed external jugular vein was implanted into the external carotid of the same side with interrupted suture. 4 weeks after operation, all vein grafts were harvested. Status of the vein grafts was observed and tissue sections were analyzed with HE staining. The intimal hyperplasia ( intima/media area index and intima/media thickness index) of the vein grafts was assessed. Results Cell viability and ROS of VSMC induced by Ang Ⅱ were suppressed by PMQ. Cell viability and ROS of VSMC were increased substantially when treated with Ang Ⅱ. The therapeutic effects of PMQ could be initially identified at dose of0. 3 μmol/L, with a peak at 3 μmol/L. The effects decreased from 30μmol/L to 10 μmol/L. PMQ at dose of 0.1 μmol/L had no effect on cell viability and ROS of VSMC induced by Ang Ⅱ. PMQ also downregulated the mRNA expression of NADPH oxidase subunits Nox1, p47phox and p22phox induced by Ang Ⅱ. A peak effect was observed at 3μmoL/L and decreased at 30 μmol/L. PMQ at o. 1 μmol/L had no effect on mRNA expression of NADPH oxidase subunits induced by Ang Ⅱ. As compared with control group, PMQ decreased intima/media area index ( 1. 64 ±0.20 in control, 0. 74 ±0.18 at 12.5 mg/kg, 1.09 ±0.17 at 25 mg/kg, 1.21 ± 0. 21 at 50 mg/kg) and intima/media thickness index ( 1.34 ± 0. 24 in control, 0.67 ± 0. 17 at 12.5 mg/kg, 0. 74 ± 0.14 at 25 mg/kg, 0.93 ± 0. 18 at 50mg/kg) at three dosages after implantation. Conclusion PMQ may suppress the proliferation of VSMC and inhibit neointima hyperplasia of vein grafts in rats. The effects may be attributed to the anti-oxidative activity and the downregulation of mRNA expression of NADPH oxidase subunits Noxl, p47phox and p22phox.

OBJECTIVE:To provide references for the application and management of anti-infectives.METHODS:Data on sale volume,DDDs etc.of anti-infection drugs in6hospitals of Guang'an city from2001to2004were statistically analyzed.RESULTS:The DDDs of oral lomefloxacin dominated the first or2nd places for the4years;The DDDs rank orders of isoniazid and rifampin were No.4and No.5respectively in the4years;The DDDs of penicillin for injection dominated the first place on the injectable preparation lists(2001～2003);The latecomer levofloxacin assumed the tendency of surpassing the early starters not only in sale volume but also the DDDs.CONCLUSION:The proportion for anti-infectives in sales volume to the sum total drug consumption of the above mentioned area were close to those of the other medical institutions in southwest area,there are less new drug varieties and the grade of drugs used in the area was low.

Objective To investigate the correlation between survival motor neuron (SMN) gene deletion and Chinese patients with sporadic amyotrophic lateral sclerosis (SALS).Methods A total of 141SALS patients and 134 unrelated controls were recruited from the Chinese population.Polymerase chain reaction (PCR) and restriction fragment length polymorphisro (RFLP) analysis were performed to screen SMN gene deletion.Frequencies of deletion were coropared by Chi-square test.Results Four patients and 3 controls were detected to have horoozygous SMN2 deletion.The frequencies of SMN2 deletion were 2.84%(4/141) and 2.24% (3/134), respectively, which was not significantly different (χ2= 0.0001, P =1.000).No subjects were found to have homozygous SMN1 deletion.Condusion There is no correlation between SMN gene deletion and Chinese patients with SALS.

This article deals with the diagnostic values of IFAT and IEST in human filariasis with both frozen sections of Brugia malayi and Setaria cervi adult worms as antigens. The average positive and the false positive rates of IFAT with two antigens were 84.9-97.1%(45/53-102/105)and 2.9-9.7%(1/35-3/31) respectively, while no cross-reaction was observed in subjects infected with ascaris or hookworm. The average positive and the false positive rates of IEST with two antigens were 94.3-94.9% (99/105-56/59) and 0-2.9% (0/35-l/35) respectively. Both IFAT and IEST with two antigens for the diagnosis of human filariasis were considered to be of higher sensitivity and specificity, frozen sections of adult Setaria cervi being more economical and effective.

A new method of two-direction counter-immunoelectrophoresis (TD-CIEP and TD-ELACIEP) for detection of specific antigen and antibody of enzyme-dissociated schisto-some circulating immune complexes (SCIC) in infected rabbits and advanced schistosomia-sis cases was reported. Results showed that the detection rate was markedly increased in the enzyme-dissociated sera as compared with that of non-dissociated sera. The levels of antigen and antibodies were found to be correlated to the period and intensity of infection. The detected antigen was solely originated from the adult worm. Comparison of sensitivity of both methods showed that TD-ELACIEP was much more sensitive than TD-CIEP either for detection of antigen or antibodies. Of 54 sera of advanced schistosomiasis cases the detection rate of schistosome antigen or antibodies by TD-ELACIEP in enzyme-dissociated and non-dissociated serum samples was 50 and 25.9%, respectively. Only 9 cases( 16.7%) were COP positive with COP rate^5%. Sera from 25 healthy persons revealed no false positive reaction by either method. The present results indicated that TD-ELACIEP of dissociated SCIC may be helpful in immunological diagnosis of advanced schis-tasomiasis.