| Obj ectiveStudy on CFSD(compound Fritillaria Scutellaria Decoction)can play a role in anti-leukemia and relieve inflammation by down-regulating the relative expression of WiplMethodsPart ?:The M1 cells were adjusted to low,medium and high doses to establish a mouse acute myeloid leukemia model by tail vein inoculation.The general condition of the mice was observed.On the day of modeling,on the 10th,20th,and 30th day after modeling,blood routine,peripheral blood smear white blood cell classification,and flow analysis of CD33 positive cells in peripheral blood were measured.Mice were sacrificed on day 30 or at the time of sudden death for pathological tissue sections to further identify the model.Drug intervention in leukemia mice,wherein the model group was intragastrically administrated with the other three groups of distilled water per day;the traditional Chinese medicine group was administered with CFSD once a day for 21 times;Intraperitoneal injection of doxorubicin hydrochloride lmg/kg,once every other day,a total of 7 times;Chinese medicine+chemotherapy group CFSD combined with doxorubicin hydrochloride intraperitoneal injection,once every other day,a total of 14 days.Peripheral blood sampling and peripheral blood smear leukocyte classification were performed in mice.After the animals were sacrificed by eyelids,the proportion of CD33 positive cells in blood and bone marrow,liver and spleen pathological tissue sections,PCR detection of blood and bone marrow were detected by flow cytometry.The expression level of Wipl was detected by Western Blot and the expression level of Wipl protein was detected by Western Blot.Part ?:A mouse infection model was established by intraperitoneal injection of Staphylococcus aureus at a concentration of 1.5×108CFU/ml and 4×108 CFU/ml at a dose of 0.2ml/mouse.Mice were observed for activity,symptoms,body weight,general condition,and death.MSS scores were obtained.Blood samples from tail veins were used to detect blood routine and blood smear white blood cell morphology and classification.The model mice were randomly divided into observation group and traditional Chinese medicine group.The observation group was given normal saline 0.012ml/g daily,and the Chinese medicine group started CFSD three days in advance,each time the dosage was 0.012ml/g,once a day,a total of 10 days.Observe the general condition of the mice,complete blood cell analysis,and rt-pcr to detect the expression level of Wip1.Part ?,the peripheral blood of 46 patients was collected and detected by ?-actin as the internal reference.The results were in accordance with the mean±standard deviation of the normal distribution,and the median(interquartile range)was not consistent with the normal distribution.The relative expression levels of Wipl mRNA in neutrophils in normal group,initial treatment/remission group and relapsed/refractory group were 0.002071(0.001627,0.011372),0.012964(0.004613,0.026645),0.010601(0.002145,0.0560517),respectively;The relative expression levels of Wipl mRNA in mononuclear cells in the group,the initial treatment/remission group and the relapsed/refractory group were 0.002461(0.001289,0.003140),0.00876(0.003105,0.171932),0.003808(0.002464,0.0108125),respectively.There was a statistically significant difference between the initial treatment/remission group and the normal group,the relapsed/refractory group and the normal group(P<0.05).The expression levels of wild-type p53 mRNA and ?-actin in the normal group,the initial treatment/remission group,and the relapsed/refractory group were 0.003754(0.001698,0.00876),0.482498(0.075047,0.1047654),and 0.0056179(0.001549,0.021865),respectively.The relative expression levels of p53 mRNA in mononuclear cells in the normal group,the initial treatment/remission group,and the relapsed/refractory group were 0.002071(0.001627,0.011372),0.040667(0.001882,0.095611),and 0.001845(0.000790,0.004543),respectively.The expression of p53 mRNA in neutral granules was statistically significant(P<0.05).The Wip1 gene in peripheral blood neutrophils and mononuclear cells of leukemia patients is highly expressed,suggesting that high expression of Wipl may affect the pathogenesis of leukemia.ResultsPart I:The established leukemia model was randomly selected into a model group,a Chinese medicine group,a chemotherapy group,and a combined group,and a group of healthy mice was set as a blank group.1 Survival status:After the mice were modeled,the appetite was poor,the spirits were wilting,the sedative movements,the skin was rough,and the skin wrinkles increased.2 Body weight(g)at 30 days of experiment:model group,Chinese medicine group,chemotherapy group,combined group,blank group,body weight were:14.50±1.17,15.66±2.35,15.11±3.65,17.08±1.84,19.79±0.38.The weight loss of the model group was the most significant,and the weight loss of the Chinese medicine+chemotherapy group was slower(P<0.05).Peripheral blood white blood cell count(×109/L):The white blood cell counts of the model group,the Chinese medicine group,the chemotherapy group,the combined group and the blank group were 9.33±2.20,6.55±2.26,4.09±2.70,6.67±2.04,2.68±0.28,respectively.The peripheral blood leukocytes of the model mice were significantly higher than the other groups(P<0.05).Liver and spleen weight(g):The spleen weights of the model group,Chinese medicine group,chemotherapy group,combined group and blank group were:1.167574±0.153007,0.992438±0.041468,1.059987±0.403035,1.096196±0.192931,1.094403±0.069661,P>0.05 The spleen weights of the model group,the Chinese medicine group,the chemotherapy group,the combined group and the blank group were:0.25568±0.030474,0.182473±0.007355,0.21829±0.0.013861,0.21337±0.008098,0.156837±0.009639,and the spleen weight of the Chinese medicine group was the lowest(P<0.05).Liver and spleen morphology and pathological sections:Necrotic foci and varying degrees of splenomegaly were observed in the spleen of each group of leukemia mice,and the liver showed white spots due to leukemia cell infiltration.After HE staining,the infiltrating foci formed by polynuclear cells and tumor cells were observed in the spleen of the microscope.The spleen structure of the combined group was relatively complete,followed by the Chinese medicine group.The other two groups were basically spleen structure.Destruction;leukemia cell infiltration can be seen in the liver of each leukemia mouse.Relative expression of Wipl mRNA:Wipl mRNA in bone marrow solution of model group,Chinese medicine group,chemotherapy group,combination group and blank group were:0.511956±0.179827,0.064957±0.078797,0.086938±0.122472,0.00418±0.005286,0.007271±0.009748,model group mice The expression of Wipl mRNA was higher in the bone marrow,and the expression level of Wipl mRNA in the bone marrow of the other groups was significantly lower.The expression level of Wipl mRNA in the bone marrow of the combined group was the lowest(P<0.05).Part ?The weight of the control group was normal before and after the experiment(P>0.05).On the 7th day after injection,the peripheral blood leukocyte counts of the control group and the Chinese medicine group were(5.15±1.09)×109/L and(8.70±2.66)×109/L,respectively,and the Chinese medicine group was higher than the control group(P<0.05).The percentage of peripheral blood neutrophils in the control group and the Chinese medicine group on the 7th day after injection was 25.29±2.04 and 18.54±8.80,respectively,and the Chinese medicine group was lower than the control group(P<0.05).Using ?-actin as an internal reference,RT-PCR showed that the expression of Wipl mRNA in the Chinese medicine group was significantly lower than that in the control group.The relative expression levels of Wipl mRNA in the control group and the Chinese medicine group were 1.64±1.16 and 0.098±0.13,respectively.The relative expression of Wipl mRNA in the group was lower than that in the control group(P<0.05).Part ?The peripheral blood of 46 patients was collected and detected by ?-actin as the internal reference.The results were in accordance with the mean±standard deviation of the normal distribution,and the median(interquartile range)was not consistent with the normal distribution.The relative expression levels of Wipl mRNA in neutrophils in normal group,initial treatment/remission group and relapsed/refractory group were 0.002071(0.001627,0.011372),0.012964(0.004613,0.026645),0.010601(0.002145,0.0560517),respectively;The relative expression levels of Wipl mRNA in mononuclear cells in the group,the initial treatment/remission group and the relapsed/refractory group were 0.002461(0.001289,0.003140),0.00876(0.003105,0.171932),0.003808(0.002464,0.0108125),respectively.There was a statistically significant difference between the initial treatment/remission group and the normal group,the relapsed/refractory group and the normal group(P<0.05).The expression levels of wild-type p53 mRNA and(3-actin in the normal group,the initial treatment/remission group,and the relapsed/refractory group were 0.003754(0.001698,0.00876),0.482498(0.075047,0.1047654),and 0.0056179(0.001549,0.021865),respectively.The relative expression levels of p53 mRNA in mononuclear cells in the normal group,the initial treatment/remission group,and the relapsed/refractory group were 0.002071(0.001627,0.011372),0.040667(0.001882,0.095611),and 0.001845(0.000790,0.004543),respectively.The expression of p53 mRNA in neutral granules was statistically significant(P<0.05).The Wipl gene in peripheral blood neutrophils and mononuclear cells of leukemia patients is highly expressed,suggesting that high expression of Wipl may affect the pathogenesis of leukemia.ConclusionCFSD can play an anti-leukemia effect by down-regulating the relative expression of Wipl,and at the same time can relieve the inflammatory response. |
PDF Full Text Request