| Objective:On the basis of modern serum pharmacology method combined with ELISA,flow cytometry,cell culture in vitro and so on,through the observation of the content in serum and changes of the IL-2 and SIL-2 receptor from CD34+DC in the bone marrow which comes from the patients of AML-CR by taking the fufang qudu soup,the thesis whit its relations with DC as its breakthrough point does research into the mechanism of action of improving MRD-L patients'immune logic function by taking the Chinese traditional medicine fufang fuzheng qudu soup.Method:Give high medium low three groups of different doses of fuzheng qudu soup to the New Zealand rabbits by gavage in the morning and evening. On the third day in a comlotely sterile environment,take the experimental rabbit heart blood to prepare rabbit serum of different drug concentration. Select the posterior superior iliac spine as puncture point and take bone marrow biopsy on patients who is standard in such experiments. Then take the bone marrow and cellect by heparin sodium biochemical tube. In the laboratory bench use lymphocytes separation medium and CD34+ magnetic beads to separate and collect theCD34+hematopoietic stem cells in bone marrow. Then use four kinds of hematopoietic stem cell amplification factor Thrombopoietin,Stem Cell Factor,Fms-like tyrosine kinase,Interleukin-4 to amplify the CD34+cells separated and collected before in vitro for 6-9days and make a logarithmic growth after CD34+cells into the DC induction. Under the optical microscope adjust the number of CD34+cells to 1 x 106/ml by direct count method. Cultivate the four kinds of cytokines GM-CSF,IL-4,IFN-A,TNF-A matched with differnet concentration of rabbit serum medicated and cytokines. Divide them into pure rabbit serum cytokines+group,pure cytokines, fuzheng fufang qudu soup of high medium and low dose+cells factor group. On the day 0,day 6,day 9 take a dish upper clear liquid and detect the content of IL-2 and SIL-2R in the supernatant fluid by ELISA assay. After the ninth day using flow cytometry instrument to detect the DC of the expression of surface antigen.Result:1.Through the inverted microscope we observe CD34+cells appear swelled as time goes by and get together into colomy.form the relatively typical DC form. Compared with the pure serum group and cytokines group, the drug containing serum group is more typical and the number of DC is larger.2.CD34+induced DC can express the immunophenotype of HLA-DR?CD86?CD80? CD1a and CD83. The expression of CD80 between the groups is not obvious (P>0.05). The expression in the medicated serum+factor group of CD80,CD83,CD86,CD1a and HLA-DR is significantly higher than that of pure rabbit serum+factor group and pure factor group (p<0.05). The difference between pure factor group and pure rabbit serum+factor group is not obvious(p>0.05). In the medicated serum+factor group, fuzheng fufang qudu soup high does+factor group of CD83 expression quantity is dominant (p<0.05). The expression of CD86 in fuzheng qudu soup low doses+factor group is higher(p<0.05). The difference of CA1a and HLA-DR in fuzheng fufang qudu soup low medium high does+factor is not obvious.3.Monitor the content of IL-2 and SIL-2R on day 0, day 6 and day 9.?As time goes on it shows the progress of the content of IL-2 significantly(p<0.05). In the same time,compared with the pure rabbit serum+factor group and pure factor group medicated serum+factor group has higher content of IL-2(p<0.05). The comparation between the pure rabbit serum+factor group and pure factor group have no statistical significance(p>0.05). At the same time the content of IL-2 in the high doses medicated serum+factor group is higher than the middle and low group(p<0.05).? The content of SIL-2R in medicated serum+factor group has decreased as time goes by (p<0.05). The difference of the content of SIL-2R of the pure rabbit serum+factor group and pure factor group between day 0 and day 6 is not obvious(p>0.05). But the content of SIL-2R in day 9 is lower than that in day 0 and day 6(p<0.05). And the difference of the content of SIL-2R in day 0 is not obvious(p>0.05). The content of SIL-2R in medicated serum+factor group in day 6 and day 9 is obvious lower than the the pure rabbit serum+factor group and pure factor group(p<0.05).The difference between the pure rabbit serum+factor group and pure factor group is not obvious(p>0.05). And the content of SIL-2R in the fuzheng qudu high doses+factor group is obvious lower than the middle and low doses group(p<0.05).Conclusion1. Combining fuzheng qudu soup with the cytokines can induce the MRD-L patients with CD34+source DC mature and can make the cell surface of HLA-DR,CD86,CDla and CD83 antigen had a higher expression rate.2. Fuzheng qudu soup can promote the increase of IL-2in the serum and reduce the concentration of serum SIL-2R. |
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