[Improvement of Preparation Method of Rats Platelet-Rich Plasma and Quality Detection].

OBJECTIVE: To optimize the single centrifugation preparation protocol of rat platelet-rich plasma (PRP). METHODS: The arterial blood of rats obtained by carotid artery intubation was collected by heparin sodium anticoagulant tubes, and then the blood divided into sterile EP tubes, adjusting the red blood cell concentration with normal saline, while rat PRP was prepared by centrifugation under different conditions (the centrifugal force was 200×g-240×g, and the centrifugal time was 8-12 min). Subsequently, the blood cell count and quality evaluation of anticoagulat whole blood and PRP were performed by hematology analyzer and flow cytometry, respectively, and the differences between different groups were compared. RESULTS: The red blood cell concentration to (5.5-6.5)×1012/L after anticoagulation of rat whole blood was good for PRP extraction. When the blood samples was centrifuged at 220×g for 10 min, the platelet recovery rate was the highestï¼»(53.52±0.63)%ï¼½. The level of apoptosis and activation of plateles in PRP were not significantly different compared to whole blood(P>0.05), and the release level of growth factor was significantly increased(P<0.05). CONCLUSION: It is a key to improve the PRP extraction efficiency by reducing the amount of mixed red blood cell, and this study successfully modified the preparation method of rat PRP, with platelets high recovery rate and stable quality.

[Factors Affecting the Preservation of Human Peripheral Blood Mononuclear Cells at 4 ℃].

OBJECTIVE: To analyze the preservation effect and related influencing factors of human peripheral blood mononuclear cells under serum-free condition at 4 ℃. METHODS: Human peripheral blood mononuclear cells were isolated by density gradient centrifugation, and stored at 4 ℃ under different cell concentrations, supplemented with human serum albumin, and glucose. The cell viability, total cell number, viable cell number and cell phenotype were detected during preservation of 72 h. RESULTS: With the prolongation of storage time, the number of human peripheral blood mononuclear cells gradually decreased(r=0.982). Compared with the cell concentration of (5-6)×106 cells/ml, the cell number decreased more slowly when the cell storage concentration was (1-2)×106 cells/ml; Adding human serum albumin and glucose can effectively improve the survival rate of human peripheral blood mononuclear cells, among which 2% human serum albumin has a better preservation effect; Compared with the blank control group, the analysis results of cell subsets showed that the downward trends of NK cells and T cells were significantly slowed after adding albumin and glucose. CONCLUSION: The cell density of (1-2)×106/ml and 2% human serum albumin are more suitable for the preservation of PBMC, and 5% glucose can improve the preservation effect of human peripheral blood mononuclear cells at 4 ℃.

[Correlation between expression of matrix metalloproteinase-2 and angiogenesis in esophageal carcinoma].

OBJECTIVE: To study expression of MMP-2 in relation to microvessel density (MVD) in esophageal carcinoma. METHODS: Forty-eight specimens of esophageal carcinoma (Ec) and 17 specimens of grade II + III epithelial dysplasia (Dy) close to the tumor and 12 specimens of normal tissue (Nt) along the incisional margin were examined by S-P immunohistochemical staining with anti-MMP-2 monoclonal antibody. An anti-CD34 monoclonal antibody was used to show MVD. RESULTS: MMP-2 expression in Ec was remarkably higher than that in Dy, which was higher than that in Nt. MMP-2 expression in Ec and Dy was significantly correlated with MVD in the tumor and nearby tissue. MMP-2 expression and MVD in Ec significantly associated with lymph node metastasis. CONCLUSION: Expression of MMP-2 plays an important role in angiogenesis and lymph node metastasis of esophageal cancer.

[The effects of pcDNA3.1-IL-15 transfection on the surface molecules and immune function of murine bone marrow-derived dendritic cells].

AIM: To explore the effects of pcDNA3.1-IL-15 transfected on co-stimulatory molecule expression and immune function on murine bone marrow-derived dendritic cells (DCs). METHODS: Recombinant plasmid pcDNA3.1-IL-15 was constructed and used to transfect DCs. The expression of CD40, CD80 and CD86 on the transfected DCs was analyzed by flow cytometry. Murine splenocytes were stimulated with the transfected DCs. CD4+ and CD8+ T cell subsets in the splenocytes were analyzed by flow cytometry. The proliferation of splenocytes was detected by MTT colorimetry. The IFN-gamma in the culture supernatant of the splenocytes was detected by ELISA. RESULTS: pcDNA3.1-IL-15-transfected DCs expressed higher level of CD40, CD80 and CD86, and induced proliferation of CD4+ T cells and CD8+ T cells in the splenocytes. But the ratio of CD4+/CD8+ T cells was lower than that in the spleen cells stimulated by untransfected DCs or DCs transfected with pcDNA3.1. CONCLUSION: pcDNA3.1-IL-15 can improve the expression of co-stimulatory molecules on DCs and enhance their immune function.

[Effects of ginsenoside Rh2(GS-Rh2) on cell cycle of Eca-109 esophageal carcinoma cell line].

OBJECTIVE: To investigate the effects of ginsenoside Rh2 (GS-Rh2) on growth inhibition and cell cycle of Eca-109 esophageal carcinoma cell line in culture. METHOD: The effects of GS-Rh2 on cell growth inhibition was detected by MTT assay. Cell cycle was analyzed by flow cytometry (FCM). Cell morphology was observed by a light microscope after HE staining. The protein expression of cell cycle components (cyclinE, CDK2, p21WAF1) were examined by immunocytochemistry and Western blot. The mRNA expression were examined by semiquantitative RT-PCR. RESULT: GS-Rh2 inhibited the proliferation of Eca-109 cells in dose and time-dependent manners. The inhibition rate was about 50% after 1-day treatment with 20 microg x mL(-1) GS-Rh2 x 20 microg x mL(-1) GS-Rh2 induced the mature differentiation and morphological reversion. With increasing dose of GS-Rh2 treatment, the cell number of G0/G1 phase was increased, whereas it decreased at S and G2/M phase. There was significant difference between 10, 20 microg x mL(-1) GS-Rh2 groups and the corresponding group without GS-Rh2 treatement. After treating cells by 20 microg x mL(-1) GS-Rh2 for 1, 2, 3 days individually, the protein and mRNA expression of both cyclinE and CDK2 reduced, while the expression of p21WAF1 enhanced gradually. CONCLUSION: GS-Rh2 could arrest Eca-109 cells at G0/G1 phase and induce cell differentiation tending to normal. Furthermore, GS-Rh2 had an effect on expression of cell cycle components (cyclinE, CDK2 and p21WAF1) to inhibit Eca-109 cell proliferation.

STUDDZS OF SCREENING MICROORGANISMS DEGRADING LINEAR SODIUM ALKYLBENZENESULFONATE (LAS) AND ITS DEGRADINGCHARACTERISTICS / 微生物学通报

A bacterium which can effectively degrade LAS (Linear Sodium Alkylbenzenesulfonate) was isolated from washing powder manufacturing effluent and was preliminarily identified as Corynebacterium jeikeium GZ6. The bacterium can degrade LAS up to till 700 mg/L, and the optimum pH, temperature and concentration of LAS are 7.0, 30℃ and 400 mg/L, respectively. The biodegradation rate can reach 98.7% after 24 hours'cultivation in the suitable conditions. Experiments also showed that some heavy metal ions such as Hg2+ , Co2+ , Cd2+ can differently inhibit the degradation of LAS.