| Objective1.To construct animal models of lymphoma/leukemia biological characteristics with high expression of miR-17-92 L1210 cells.2.To observe the chemotherapy sensitivity of L1210 leukemia mouse models and hig h expression of miR-17-92 L1210 leukemia mouse models in different treatment and cla rify the correlation between miR-17-92 and drug resistance.Observe the curative effect o f CZBG in lymphoma or leukemia.3.To observe inhibition effect of CZBG-medicated serum on drug resistance induced by high expression of miR-17-92 L1210 cells and to investigate the effect of CZBG on apoptosis gene expression of FASL, BAX and FAS.4.Analysis the understanding of ancient and modern physicians on etiology and pathogenesis, syndrome differentiation, treatment and prescription of lymphoma,in order to provide guidance to standardize TCM diagnosis and treatment of lymphoma.Method1.B-leukemia miR-17-92 high expression L1210 cells was conventionally infused into each DBA/2 mouse through the vena caudalis at the cellular number of 100×104 (A group), at the cellular number of 300×104 (B group),and subcutis at the cellular number of 300×104 in the initial part of right leg(C group).The process of leukemia in these mice was investigated.The amount of peripherial leucocytes in mouse blood was counted and the morphological changes and proportion of peripherial leucocytes were investigated.Mouse bone marrow cells were collected before the first 22 days or before dying,and the liver,spleen,kidney and lung were kept,weighed and made into pathological sections.2.B-leukemia miR-17-92 high expression L1210 cells was conventionally infused into DBA/2 mice through the vena caudalis at the cellular number of 100×104.Make them into 5 groups with a randomized method:miR-17-92 observation group,miR-17-92 chemotherapy group,miR-17-92 traditional Chinese medicine and chemotherapy group,L1210 chemotherapy group, normal control group.The drug:CTX dose was at the concentration of 8mg/ml,two days delivery times, continuous medication three times;CZBG dose was at the concentration of 0.4g/ml,gavage once daily, continuous medication for fourteen days.The process of leukemia in these mice was investigated.The amount of leucocytes in mouse blood and leukemia cell percent in blood and bone marrow was counted and the morphological changes were investigated.The difference of the amount of P-gp and spleen weight in five groups was analysised at last.3.First step was to produce CZBG dose containing serum(4g d-1·kg-1)with eight wistar male rats.Second,divided doxorubicin group and CZBG adriamycin group with cultured L1210 cells, miR-17-92 high expression L1210 cells, L1210/CDDP cells. Doxorubicin group was divided into control group, low-dose,medium-dose, high-dose group; CZBG adriamycin group was divided into control group, low-dose,medium-dose, high-dose group. The culture was blank control group,the remaining three groups were added to the dose CZBG containing serum and low-dose, medium-dose,high dose of doxorubicin. After training 24,48,72h,then used AnnexinV-FITC/PI double staining, detected the rate of apoptosis and detected the rate of apoptosis by flow cytometry (FCM).4.1n ancient Chinese medicine,lymphoma was often named of’Stone gangrene","Yin gangrene","evil-core","sputum core","lost glory","scrofula".Using with 5th Edition "China Medical Code" software systems and China National Knowledge Chinese journals database, we retrievalled original ancient and modern literature with keyword of disease names above.Name of disease,pathogenesis, syndrome differentiation, therapy herbs was sortted and analyzed in order to summarize ancient and modern Chinese medicine theory and treatment of lymphoma.Result1.The rate of tumorigenesis of vena caudalis injection group and subcutaneous group was 100%.The average survival time of the three groups was(24.30±3.64), (14.20±3.42), (26.40±5.56)d. The amount of leucocytes after seven days infusion was(9.96±2.03)×109/L, (10.81±1.70)×109/L,(12.76±2.06)×109/L,and the ratio of leukemia cell was (21.33±2.78)%, (32.17±6.50)%, (10.00±2.00)%.After fourteen days infusion it was(8.07±1.31)×109/L, (5.02±1.22)×109/L, (13.28±2.08)×109/L, and the ratio was(32.83±3.50)%,(49.67±7.00)%,(12.66 ±2.00)%.In pathological sections of the liver and spleen of dying mice,diffused leukemia cell infiltration could be observed,accompanied by destruction to the normal tissue and structure.A small quantity of leukemia cells was found in lungs and kidneys.The destruction to their normal tissue and structure was not obvious and hemorrhage was easily seen in lungs.2.The mice of five groups were killed before tail vein blood and bone marrow fluid specimens on medication for 15 days or dying.The amount of leucocytes in mice blood was counted and the morphological changes were investigated.The spleen of the mice were kept,weighed and made into pathological sections.The amount of leucocytes in each group was (7.09±2.93)×109/L,(15.30±11.40)×109/L,(15.31=11.42)×109/L,(11.47±4.25)×109/L,(11.40±2.58) ×109/L.The ratio of leukemia cell in blood was (37.17±13.08)%, (5.67±3.93)%,(1.33±0.82)%, (0.83±0.75)%,0%.The HGB in each group was(130.33±20.32),(159.00±13.10),(145.67±17.07), (156.17±13.93),(171.67±13.50).The amount of platelet count in each group was(664.17±284.11 )×109/L, (1884.17±335.35)×109/L,(1673.17±428.42)×109/L,(1279.67±650.68)×109/L, (973.67 ±42.70)×109/L.The ratio of leukemia cell in bone marrow was (26.92±10.98)%, (6.83±1.97)%, (3.17±1.03)%, (1.67±1.21)%,0.00%. The spleen weight in each group is was(0.69±0.18),(0.19± 0.17), (0.44±0.47),(0.23±0.19),(0.60±0.18).The value of P-gp in each group was(32.68±10.88), (26.67±10.87),(31.99±11.25), (20.31±11.12), (17.39±2.64).3.(1)Result the early apoptosis rate and total apoptosis rate induced by doxorubicin showed high expression of miR-17-92 L1210 cells low-dose group was lower than L1210 cells low-dose group and L1210/CDDP cells low-dose group(P<0.05). (2)The 24h late and total apoptosis rate induced by all dose ADM and CZBG medicated serum of miR-17-92 high expression L1210 cells group was higher compared to the ADM group alone(P<0.01).The total apoptosis rate induced by all dose ADM and CZBG medicated serum of L1210 cells group was significantly higher compared to the ADM group alone(P<0.05).The 24h late and total apoptosis rate induced by high dose ADM and CZBG medicated serum of L1210/CDDP cells group was higher compared to the ADM group alone(P<0.01). (3)Administration with ADM for 24h and 72h, FASL,BAX,FAS expression level of miR-17-92 high expression L1210 group was significantly lower than the L1210 group. Administration with CZBG and ADM for 24h,FASL,BAX,FAS expression level of miR-17-92 high expression L1210 group was significantly higher compared with ADM treatment alone.4.(1)Pathogenesis in lymphoma was considered of empty,cold, phlegm, poison (immunoto-xin),stasis,stagnation as much.(2)According to different syndromes,there were more unilateral prescriptions.(3)Shamisen medicine in CZBG had long been documented in ancient books of lymphoma.Conclusion1.Inoculation of high expression of miR-17-92 L1210 cells with the number of 100×104 into DBA/2 mice is more in line with the biological characteristics of clinical leukemia and meet the test cycle.2.High expression of miR-17-92 can reduce sensitivity to chemotherapy in mouse lymphoma cells.CZBG can improve remission rates and reversal of drug resistance partially.3.CZBG-medicated serum has a certain effect on reversing of drug resistance and promoting apoptosis in high expression of miR-17-92 L1210 cells.4.Lymphoma has long been documented in ancient Chinese medicine, and Chinese medicine treatment of lymphoma have a certain effect. |
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