[Influence of crocin on proliferation in vitro and function of dendritic cells derived from bone marrow of children with acute leukemia].

This study was purposed to investigate the effect of crocin on the proliferation in vitro and immune function of dendritic cells (DC) derived from the bone marrow of children with acute leukemia. The mononuclear cells were isolated from bone marrow of leukemia children by Ficoll-Hypaque. The experiment was divided into six groups: blank control group (A), crocin 1.25 mg/ml group (B), cytokines (rhGM-CSF 75 ng/ml+rhIL-4 75 ng/ml+rhTNF-α 50 ng/ml) group (C), cytokines+crocin 0.3125, 1.25 or 5.0 mg/ml groups (D, E, F). The numbers of DC were counted and the phenotypes of DC were determined by flow cytometry on the ninth day of culture. The DC of different groups were mixed with T cells just separated from peripheral blood of another children with acute lymphoblastic leukemia, and cultured with rhIL-2 200 U/ml for 5 d. The function of DC was detected by mixed lymphocyte reaction (MLR). The results indicated that the test groups and control group all obtained a certain amount of typical DC, but the DC numbers in test groups were all higher than those in control group (P < 0.01). Cultured for 9 days, the rates of CD1a(+), CD83(+), and HLA-DR(+) in group C, D, E, F were higher than group A (P < 0.01). There was no statistically significant difference between A and B groups (P > 0.05). MLR showed that with the increasing of DC, the stimulation index of T cells in group A and B was not rising (P > 0.05); the stimulated index of T cells in group C and E was significantly rising, there was statistically significant difference between them (P < 0.01). When the number of stimulated cells was the same, the stimulation index of T cell in group E was the highest (P < 0.01). It is concluded that the capability of DC proliferation promoted by crocin alone is lower than that of its combination with rhGM-CSF, rhIL-4 and rhTNF-α, but the crocin can synergically promote the maturity of DC cooperating with rhGM-CSF, rhIL-4 and rhTNF-α. The DC induced by crocin can particularly enhance the proliferation of T cells.

[Imbalance of Th17/Treg cells ratio in peripheral blood of patients with immune thrombocytopenia].

The aim of this study was to investigate the expression of Th17 cells and regulatory T (Treg) cells in peripheral blood of patients with immune thrombocytopenia (ITP) and to clarify the role of the Th17/Treg cell ratio imbalance in pathogenesis of ITP. Patients were divided into the pre-treatment group (active group) (n = 38) and post-treatment group (remission group) according to the platelet count and curative effect. Post-treatment group was further divided into remission group (n = 24), partial remission group (n = 10), and non-remission group (n = 4). 30 healthy subjects were enrolled in control group. Flow cytometry was used to detect the percentages of peripheral blood Th17 cells and Treg cells in CD4(+) T cells from ITP patients and controls respectively. The results showed that the percentages of Th17 cells in active group and non-remission group were significantly higher than those in control group (p < 0.05). The percentages of Th17 cells in remission group, partial-remission group were also higher than those in control group, but there were no statistically significant differences between these groups. The percentage of Th17 cells in remission group was lower than that in active group, but there was also no statistically difference between two groups. The percentages of Treg cells in active group, partial-remission group and non-remission group significantly decreased, compared with in control group (p < 0.01). The percentage of Treg cells in remission group was lower than that in control group, but there was no statistically significant difference. The ratio of peripheral blood Th17/Treg cells in active group, partial-remission group and non-remission group was higher, as compared with in control group. The ratio of peripheral blood Th17/Treg cells in remission group was higher than that in control group, but there was no statistically difference between two groups. It is concluded the percentage of Th17 cells and the ratio of Th17/Treg cells are higher in active group. The percentage of Treg cells is low in active group, partial remission and non-remission groups. The imbalance of Th17/Treg ratio may play a critical role in ITP pathogenesis.

Muramyl Dipeptide modulates differentiation, maturity of dendritic cells and anti-tumor effect of DC-mediated T cell in acute leukemia children.

BACKGROUND: Targeted therapy is a potentially useful approach for antileukemic therapy, in particular eliminating minimal residual disease(MRD) to prevent tumor relapse. This study was aimed to find out an effective, nontoxic dendritic cell (DC) maturating agent for the immunotherapy of acute leukemia. RESULTS: MDP-matured DCs(M-DCs) expressed higher level of phenotypic markers and secreted higher cytokine level, while lower than TNF-α-matured DCs (T-DCs) and co-administration of MDP and TNF-α-matured DCs (MT-DCs). MT-DCs promoted significantly allogeneic T-cells reaction. As a result, allogeneic T-cell proliferated significantly and secreted higher amount of IFN-γ. HL60-derived antigens were presented more effectively by MT-DCs to cytotoxic T lymphocytes (CTLs) to induce more beneficial anti-tumor effects in a dose-dependent manner. METHODS: Purified mononuclear cells (MNCs) from bone marrow of acute leukemia children were differentiated by granulocyte-macrophage colony stimulating factor (GM-CSF) and recombinant human interleukin-4 (rhIL-4) and further matured by either Muramyl Dipeptide (MDP), tumor necrosis factor-alpha (TNF-α) or co-administration of MDP and TNF-α. CONCLUSIONS: These results demonstrate MDP can be used as a candidate clinical agent for antigen specific cancer immunotherapy.

[Effect of Bacillus Calmette-Guerin on the expansion of dendritic cells from peripheral blood of pediatric patients with leukemia in vitro].

This study was purposed to investigate the effect of bacillus calmette-guerin (BCG) on the expansion of human dendritic cells (DC) from peripheral blood of pediatric patients with leukemia in vitro. The experiment was divi-ded into two groups: the control and the test group, and the latter group was divided into 3 subgroups: BCG (only BCG), GTI (GM-CSF, TNF-α, IL-4) and GTIB (GM-CSF, TNF-α, IL-4 plus BCG). On day 9 of culture the DCs were counted in each groups, the phenotypes of DC were determined by flow cytometry and these DC were stained with Wright-Giemsa for observation and photography under microscopy. The results showed that the test groups all obtained a certain amount of typical DC; the number of DC in the BCG subgroup is lower than that in the GTI and GTIB subgroups (t=4.20; 6.36, p<0.01); there was no significant difference between the GTI and the GTIB subgroups (t=2.25; p>0.05). The rate of CD1a+ in the BCG subgroup was obviously higher than that in the control group (t=3.04, p<0.05), but was lower than that in the GTI and the GTIB subgroups (t=2.79, 6.41, p<0.05), there was no significant difference between the GTI and the GTIB subgroups (t=0.65, p>0.05). The rate of HLA-DR+, CD83+ in the BCG group was higher than that in the control group (t=4.77, 4.15; p<0.05), but lower than that in the GTI and the GTIB subgroups (t=6.65, 3.19; p<0.05). The rate of HLA-DR+, CD83+ in the GTI subgroup was lower than that in the GTIB subgroup (t=5.64, 2.98; p<0.05). It is concluded that BCG not only promotes the proliferation of DC derived from human peripheral blood of leukemia patients in vitro, but also cooperates with rhGM-CSF, rhTNF-α and rhIL-4 in promoting the maturation of DCs.

[Proliferative inhibition and apoptotic induction effects of crocin on human leukemia HL-60 cells and their mechanisms].

This study was to investigate the proliferative inhibition and apoptosis of human leukemia HL-60 cells induced by crocin and their possible mechanisms. The cell viability was tested by cell counting. The morphology of HL-60 cells was observed by fluorescence microscopy. The MTT assay was used to evaluate the inhibitory effect of crocin on the growth of HL-60 cells. Flow cytometry was used to measure the cell cycle. RT-PCR was used to detect bcl-2 and bax expression. The results indicated that the growth of HL-60 cells was inhibited remarkably in the dose and time dependent way. When the crocin concentration was higher than 5 mg/ml, the percentage of apoptotic HL-60 cells was not increased, on the contrary this percentage decreased, the cells manifested necrosis. Flow cytometry profiles revealed that cells were blocked in G0/G1 phase, the cell proliferation was inhibited obviously at 5 mg/ml. RT-PCR detection revealed that the expression of bcl-2 was down-regulated strikingly and bax was up-regulated. It is concluded that the crocin can inhibit the proliferation of HL-60 cells effectively, and block cells in G0/G1 phase. The mechanisms by which crocin induced apoptosis in HL-60 cells may be related to the inhibition of bcl-2 and activation of bax.

[Influence of Interferon alpha-2b on proliferation inhibition and apoptosis induction in HL-60 cells].

To investigate the effects of interferon alpha-2b on proliferation and apoptosis in HL-60 cells, HL-60 cells were cultured in different concentrations of IFN alpha-2b. The morphologic changes were observed by Wright's and acridine orange (AO) and ethidium bromide (EB) staining respectively. Inhibition of proliferation was detected by MTT. Expression of CD13(+) was checked by indirect fluoroimmunoassay. The results showed that apoptosis rate of HL-60 cells assayed by the above-mentioned two methods was (51 +/- 2)% and (78 +/- 3)% respectively and OD(570) values of proliferation inhibited were 1.8 +/- 0.1 and 1.0 +/- 0.1 respectively when the concentrations of the IFN(alpha-2b) were 500 and 10,000 U/ml in culture for 48 hours. Morphology and count of CD13(+) cells were changed. CD13(+) cell expression rate was (62 +/- 2)% and (30 +/- 3)% respectively when the concentrations of the IFN(alpha-2b) were 500 and 10,000 U/ml in culture for 48 hours. It is concluded that IFN(alpha-2b) can enhance the apoptosis of HL-60 cells, inhibit their proliferation, promote their maturation and differentiation.

[Effect of nimodipine on mechanisms of HL-60 cell apoptosis induced by cytarabine].

The aim was to study the mechanisms of HL-60 cell apoptosis induced by nimodipine (NMDP) and cytarabine (Ara-C). The DNA fragment was detected by agarose gel electrophoresis. The expressions of bcl-2 and bax gene proteins related with apoptosis were investigated by immunohistochemistry. The results showed that HL-60 cell apoptosis rate had been increasing in the experimental groups compared with the control group since culturing 8 hours. The expression of Bcl-2 protein was lower and the expression of Bax protein was higher in the experimental groups than that in the control group, while ratio of bcl-2/bax was lower in the experimental groups than that in the control group. It is concluded that NMDP and Ara-C induce apoptosis of HL-60 cells, and the mechanism of apoptosis induced by them may down-regulate the expression of bcl-2 gene and up-regulate the expression of bax gene. The mechanism of HL-60 cell apoptosis induced by Ara-C and NMDP is probably associated with the down-regulation of Bcl-2 protein expression.

[Infection status and clinical significance of Epstein-Barr virus in pediatric leukemia---a report of 35 cases].

BACKGROUND & OBJECTIVE: Epstein-Barr virus (EBV) is associated with genesis of many human tumors. This study was to detect EBV infection in pediatric leukemia, and to explore its clinical significance. METHODS: EBV DNA in peripheral blood mononuclear cells in 35 pediatric leukemia patients, including 26 cases of acute lymphoblastic leukemia (ALL) (24 received initial treatment and 2 received retreatment), 8 cases of acute non-lymphocytic leukemia (ANLL) and 1 case of chronic lymphocytic leukemia (CLL), and in 14 healthy children was detected by fluorescent quantitative polymerase chain reaction (FQ-PCR). Its clinical significance was analyzed according to the clinical manifestations, prednisone sensitivity test, and complete remission (CR) rate after induction chemotherapy. RESULTS: EBV DNA was detected in 8 (22.86%) of the 35 pediatric leukemia patients. The positive rate of EBV DNA was 26.92% (7/26) in ALL with quantity of (5.144+/-6.91)x10(5) copies/ml, and 12.5% (1/8) in ANLL patients with quantity of 4.031x10(3) copies/ml. No EBV DNA was detected in CLL patients and healthy controls. The occurrence rates of peripheral leukocytosis and hepatosplenomegaly were significantly higher in the patients with EBV infection than in the patients without EBV infection (P <0.001). In ALL, the rate of no response to prednisone was significantly higher in the patients with EBV infection than in the patients without EBV infection (100% vs. 26.32%, P =0.001); CR rate after induction chemotherapy was significantly lower in the patients with EBV infection than in the patients without EBV infection (28.57% vs. 84.21%, P=0.003). In ANLL, the differences of CR rate and relapse rate were not significant between the patients with and without EBV infection (P=0.5). CONCLUSIONS: Pediatric leukemia patients with EBV infection have higher incidence of peripheral leukocytosis and hepatosplenomegaly. ALL patients with EBV infection have poor prednisone response and low CR rate.

[The progress of research on ginlcgobiobal flavone].

OBJECTIVE: The research on flavonoid in the recent years is extensive, such as Soybeans flavone, Baicalensis flavone, Epimedium flavone. Experiments show the effects of flavonoid is closely related to human health. There are a lot of reports about Ginkcobiobal flavone. In order to make further progress research on ginlcgobiobal flavone, we sum up the articles and reports on ginlcgobiobal flavone in the recent years. METHOD: To searche the articles about ginlcgobiobal flavone studies in the past five years. RESULT: Ginlcgobiobal flavone is not only a vasodilator, but also has the effects of anti-inflammation, analgesia, lowering blood lipids, preventing senile and inhibiting tumor, treating leukaemia, regulating gene and biotransformation. CONCLUSION: Ginlcgobiobal flavone has the potential value for drug research and development.