ABSTRACT
BACKGROUND: The central position of dendritic cells (DCs) has aroused increasing attention due to strong antigen presentation capability in anti-tumor. However, how to obtain enough functional DCs and reports regarding immunomodulator with low toxicity are few. OBJECTIVE: To investigate the effect of mycobacterium phlei F.U.36 (Utilins) on the proliferation and maturity of DCs derived from human umbilical cord blood in vitro.METHODS: The mononuclear cells were isolated from human umbilical cord blood using Ficoll-Hypaque method and cultured with Utilins, cytokine (human recombinant granulocyte/macrophage colonystimulating factor + recombinant human tumor necrosisfactor-α + recombinant human interleukin-4), or combination cytokine + Utilins, respectively. In addition, cells cultured with RPMI-1640 served as controls. Morphological features and growth of DCs were observed by an inverted microscope. Thephenotype changes of DCs, such CD1a and HLA-DR, were detected by flow cytometry at 9 days after culture. Moreover, DCs were stained by Wright-Giemsa and observed under oil lens. RESULTS AND CONCLUSION: Typical DCs with high expressions of CD1a and HLA-DR were obviously detected in all experimental groups except the control group. The positive rates of CD1a and HLA-DR in the Utilins group were higher than those in the control group, but lower than the cytokine group (P < 0.05). The HLA-DR positive rate of the combination group was higher than that of the cytokine group (P < 0.05). The results revealed that, Utilins can not only promote the proliferation of DCs derived from human umbilical cord blood in vitro but also cooperate with cytokines to induce the maturity of DCs.
ABSTRACT
BACKGROUND: The existence of minimal residual leukemia cells is the main cause for the recurrence of acute leukemia in children, and immunological biological therapy has attracted more and more attentions in the various methods from eliminating minimal residual disease. Previous studies have found that interferon α-2b can effectively inhibit the increase of tumor cells in vivo in children with neuroblastoma and malignant lymphoma, whether it can inhibit the increase of leukemia cells?OBJECTIVE: To investigate the effects of interferon α-2b in vitro on leukemia cells.DESIGN: A comparative observation taking human promyelocytic leukemia HL-60 cell line as the material.SETTING: Cell Culture Room; Immunological Laboratory; Cell Room, Institute of Pediatrics, Affiliated Hospital,Medical College of Qingdao University.MATERIALS: HL-60 cell line was provided by Shandong Institute of Basic Medical Sciences. Interferon α-2b was purchased from Megagene Company Fluorescein isothiocyanate (FTTC) rabbit-anti-rat Ig solution (CatEK001) and CD13 anti-human monoclonal antibody solution (Cat. DK013Y) were purchased from Union Stem Cell & Gene Engineering Co.,Ltd.METHODS: The experiments were carried out in the Institute of Pediatrics, Affiliated Hospital, Medical College of Qingdao University from March to September 2005. HL-60 cells culture system was established in vitro, and the oncentration was adjusted to 1×109 L-1. The cells were divided into control group and experimental group. In the experimental group, each well was added by interferon-α-2b with the terminal concentration of 5×105, 1×106, 2×106,5×106 and 1×107 U/L, respectively. In the control group, each well was added by saline of the same volume. The cells were cultured continuously for 48 hours. The morphological changes of HL-60 cells were observed using Wright's staining under light microscope; Cell apoptosis was observed using acridine orange/ethidium bromide double staining; Antigen expression and maturation and differentiation on cell membrane were observed by determining CD13 protein expression; Proliferation and activity of HL-60 were detected with methyl-thiazol-tetrazolium (MTT) assay.MAIN OUTCOME MEASURES: The occurrence of apoptosis was judged according to the uniformity and staining of HL-60 nuclear chromatin; HL-60 cell proliferation was judged according to the absorbance (A) value; The maturation of HL-60 cells was judged according to the number of positive CD13 cells.RESULTS: ① HL-60 cell apoptosis: The cells were cultured for 48 hours. When the concentration of interferon α-2b was 5×105 U/L, there were mainly early apoptotic HL-60 cells; When the concentration was 1×107 U/L, there were mainly late apoptotic cells, and the apoptotic rate was significantly higher than those in the control group (P ＜ 0.01 ).② HL-60 cell proliferation: The A values in the experimental groups treated with interferon α-2b of 2×106 U/L and 1 ×107 U/L were significantly lower than that in the control group (P ＜ 0.01). ③ Maturation of HL-60 cells: The percentages of positive CD13 cells in the experimental groups treated with interferon α-2b of 1 ×106 and 1 ×107 U/L were significantly lower than that in the control group (P ＜ 0.01).CONCLUSION: It is concluded that interferon α-2b can enhance the apoptosis, inhibit the proliferation and promote maturation and differentiation of HL-60 cells.
ABSTRACT
BACKGROUND: Cardiac fibrosis, which results from the loss of balance between synthesis and degradation of cardiac matrix component, is the structural foundation of the stiffness of damaged myocardial tissues. Astragalus membranaceus, a traditional Chinese herb, has multiple functions such as exerting a tonic effect on the heart to induce diuresis. However,the effect of astragaloside Ⅳ on cardiac collagen is poorly known in practice.OBJECTIVE: To observe the effects of astragaloside Ⅳ on myocardial collagen and cardiac function in ischemic rats and to investigate the dosage and time effects of astragaloside Ⅳ.DESIGN: A completely randomized grouping design and randomized controlled trial.SETTING: Department of Pediatrics, the Affiliated Hospital of Medical College of Qingdao University.MATERIALS: The study was conducted in the Encephalopathy Research Institute, Medical College of Qingdao University, from July 2003 to February 2004. Totally 132 Wistar rats of cleaning grade were randomized into three groups: control group (n=11), ischemic group (n=10) and astragaloside Ⅳ group (n=121).METHODS: Rats in control group had thoracotomy, but did not have their left anterior descending coronary artery ligated; rats in ischemic group had thoracotomy and had their left anterior descending coronary artery ligated to establish acute myocardial infarction model; rats in astragaloside Ⅳ group were given astragaloside Ⅳ after surgical ligature of left anterior descending coronary artery. The changes in hemodynamic parameters, cardiac function and myocardial collagen were determined. The dosage and time effects of astragaloside Ⅳ on myocardial collagen and cardiac function were observed.MAIN OUTCOME MEASURES: ① The dosage and time effect of astragaloside Ⅳ on the content of myocardial collagen in the left ventricle of rats with myocardial infarction; ② The dosage and time effects of astragaloside Ⅳ on hemodynamics and cardiac function of rats with myocardial infarction.RESULTS: One hundred rats entered the results analysis. There were 10 in control group and ischemic group, respectively, and 80 in astragaloside Ⅳ group. The five dosage groups of astragaloside Ⅳ [2.5, 5.0, 10.0, 15.0 and 20.0 mg/(kg·d)] and the five postoperative time points (3, 7, 14, 21 and 28 days) had eight rats for each. Astragaloside Ⅳ at a dose of 15.0 mg/kg per day was found to have the most marked effect on ischemic myocardium, so this dose was chosen for observing time effect. ① After administration of astragaloside Ⅳ, the content of collagen in myocardial tissues of the infarcted area of left ventricle, the serum concentration of carboxyterminal procollagen type Ⅰ propeptide and aminoterminal procollagen type Ⅲ propeptide decreased gradually with the increased dose of astragaloside Ⅳ and with the prolonged action time of astragaloside Ⅳ [15 mg/(kg·d)] (P ＜ 0.05-0.01). The serum concentration of carboxyterminal procollagen type 1 propeptide and aminoterminal procollagen type Ⅲ propeptide returned to the level of control at a dose of 10 mg·kg-1per day and at 21 days after astragaloside Ⅳ administration, respectively. The content of collagen in myocardial tissues of the infarcted area of left ventricle was higher than that of non-infarcted area (P＜ 0.01); there were no significant changes in the content of cardiac collagen of right ventricle and non-infarcted area of left ventricle before and after astragaloside Ⅳ administration. ② The cardiac function of ischemic rats significantly improved after astragaloside Ⅳ administration (P ＜ 0.05-0.01); cardiac output, heart rate, stroke volume,mean aortic pressure, systolic aortic pressure, and the stroke work of left ventricle gradually returned to the level of control with the increased dose of astragaloside Ⅳ and with the prolonged action time of astragaloside Ⅳ.CONCLUSION: Astragaloside Ⅳ can inhibit the proliferation of cardiac collagen and improve cardiac function in rats with myocardial infarction.The content of myocardial collagen gradually decreases and cardiac function gradually improves with the increased dose of astragaloside Ⅳ and the prolonged action time of astragaloside Ⅳ.