| Objective Exploring the relationship between the liver and spleen and the therapeutic effects of related drugs by establishing a rat model of aplastic anemia(AA).To provide a foundation and basis for the understanding of the mutual influence of the five organs in Mongolian medicine and the treatment of deficiency.Methods Randomly divide SPF grade SD male rats into blank group,model group,low dose of Ulan-10,high dose of Ulan-10,bear bile group,ursodeoxycholic acid group,and positive control group.Except for the blank group,all other groups were injected subcutaneously with2% acetylphenyl hydrazine(APH)physiological saline solution(20mg/kg)on the first day of the body mass experiment,and subcutaneously with 2% acetylphenyl hydrazine physiological saline solution(10mg/kg)on the fourth day.After 2 hours and on days 5,6,and 7,a rat model of aplastic anemia was established by intraperitoneal injection of cyclophosphamide(CTX)physiological saline solution(20mg/kg).After successful modeling,the blank group and model group were given CMC-Na solution,and each treatment was given corresponding drugs for 30 consecutive days by gavage.Observe the general situation of fur,food intake,and activity status of rats in each group.After the last administration,fasting for 12 hours was used for sampling.Detection of red blood cells,hemoglobin,and platelets;Spleen index;ALT 、 AST 、 ALP;IL-2,IL-3,IL-6,IL-11,IL-17,EPO,TNF-a,c AMP/c GMP levels;Pathological changes in liver,spleen,thymus,and femoral bone marrow;Western blot method was used to detect the content of TGR5 protein in the liver and spleen tissues of rats in each group.Results 1.After modeling,the rats showed symptoms such as pale mouth,nose,ears,and tail,dry hair,decreased water and food intake,and weight loss.After treatment,the above symptoms were relieved and no deaths occurred.2.Compared with the blank group,the model group showed a significant decrease in red blood cell,hemoglobin,and platelet counts(P<0.01);Except for the low dose Ulan-10 group and the ursodeoxycholic acid group,there was no significant difference in red blood cells(P>0.05),all other groups showed significant differences.Compared with the blank group,the spleen index of the model group significantly increased,with a significant difference(P<0.01);Compared with the model group,there was a significant difference in the decrease in Ulan-10 high-dose group(P<0.01),positive control group(P<0.01),and bear bile group(P<0.05);There was no significant change in the low dose group of Ulan-10 and the ursodeoxycholic acid group.4.There was no difference in ALT,AST,and ALP between the model group and the blank group.5.Compared with the blank group,the serum levels of IL-2,IL-3,IL-6,IL-11,IL-17,and TNF-a in the model group significantly increased(P<0.01),while EPO levels decreased(P<0.01).There was no difference in c AMP/c GMP levels.The IL-2 level significantly decreased compared to the model group after treatment(P<0.01),and there was no difference in the positive control group.The IL-3 level in the treatment group decreased(P<0.01),and there was no difference in the low dose of Ulan-10 compared to the ursodeoxycholic acid group.The levels of IL-6 in each treatment group decreased significantly(P<0.01).The IL-11 level in the treatment group decreased(P<0.01),but there was no difference between the low dose group of Ulan-10 and the bear bile group.The levels of IL-17 in each treatment group decreased significantly,with significant differences(P<0.01).The levels of TNF-a in the treatment group decreased significantly(P<0.01).The levels of EPO in each treatment group increased significantly(P<0.01).c AMP did not change(P>0.05),c GMP showed a downward trend(P=0.05),and there was no difference in c AMP/c GMP levels(P>0.05).6.In the model group,hepatocytes were steatosis,and some hepatocytes had small fat vacuoles.Focal infiltration of inflammatory cells can be observed near the portal area of the Ulan-10 high-dose group.At the low dose portal area of Ulan-10,very few hepatocytes were found with steatosis,and the nuclear fragmentation of hepatocytes disappeared.In the bear bile group,a small amount of steatosis of hepatocytes was occasionally seen in the portal area,and there were small fat vacuoles in the cytoplasm of hepatocytes.In ursodeoxycholic acid group,the portal area occasionally saw proliferation of hepatocyte nuclear epithelial cells,and the gap between hepatocytes was enlarged,without obvious steatosis.In the positive control group,a small amount of inflammatory cells infiltrated near the portal vein,bile duct epithelial cells proliferated,and the gap between hepatocytes increased,without obvious steatosis.In the spleen tissue model group,the arrangement of the boundary between red and white pulp is disordered,the peripheral area around the arteries of white pulp becomes thinner,the number of lymphocytes decreases,the splenic nodules significantly shrink,the germinal center is not obvious,the red pulp area is swollen,the splenic sinus is dilated and congested,and the red pulp contains hemosiderin pigment deposition.The boundaries between red and white pulp in each treatment group are clear.The high dose group of Ulan-10 showed deposition of hemosiderin pigments in both the red and white pulp;In the low dose group of Ulan-10,the peripheral area of the white pulp became thinner,and there were deposits of hemosiderin pigments in both the red and white pulp;There are deposits of hemosiderin pigments in both the red and white pulp of the bear bile group;In the ursodeoxycholic acid group,the peripheral area of the white pulp became thinner,and hemosiderin pigments were deposited in the red pulp.The positive control group showed an increase in red pulp lymphocytes near the cortical area,splenic sinus dilation and congestion,and deposition of hemosiderin pigments in the red pulp area.The thymic tissue model group had unclear boundaries between the thymic cortex and medulla,loose arrangement of lymphocytes,more necrosis and fragmented nuclei in the thymic medulla area,and interstitial edema.Except for the unclear thymic cortex medullary boundary in the low dose group of Ulan-10,the thymic cortex medullary boundary in the other groups is clear.Vascular congestion can be seen in the high-dose medullary area of Ulan-10,and no obvious necrotic cells are observed.Vascular congestion can be seen in the low-dose medullary area of Ulan-10,and partial lymphocyte necrosis can be seen in the cortical area.Vascular congestion and mild interstitial edema can be seen in the medullary area of the bear bile group.In the deoxycholic acid group,vascular congestion and mild interstitial edema were observed in the medullary area.In the positive control group,the medullary boundary of the thymic cortex was clear,and a small amount of lymphocyte necrosis was seen in the cortical area.The number and volume of adipocytes in the bone marrow cavity of the model group increased,with a large number of mature red blood cells entering the blood sinuses,and the number of hematopoietic cells decreased compared to the blank group.The number of adipocytes in the bone marrow cavity of the Ulan-10 high-dose group decreased compared to the model group,while the number of hematopoietic cells increased.There was no significant change in the number of adipocytes in the bone marrow tissue of the Ulan-10 low-dose group,but the volume of adipocytes decreased and the number of hematopoietic cells increased.The volume of adipocytes in the bear bile group decreased and the number of hematopoietic cells increased.The volume of adipocytes in the ursodeoxycholic acid group decreased and the number of hematopoietic cells increased.The volume of adipocytes in the positive control group decreased and the number of hematopoietic cells increased.7.Western blot was used to detect the expression of TGR5 protein in the liver and spleen tissues of rats.The results showed that there was a significant difference in TGR5 content in the liver tissue of the model group compared to the blank group(P<0.01);Compared with the model group,there was no significant difference between the Ulan-10 low-dose group and the Ulan-10 high-dose group(P>0.05);There was a significant difference between the bear bile group and the ursodeoxycholic acid group(P<0.05),and the positive control group(P<0.01).There was a significant difference in TGR5 content in the spleen tissue of the model group compared to the blank group(P<0.01);Compared with the model group,there was no significant difference between the Ulan-10 low-dose group and the Ulan-10 high-dose group(P>0.05);There was a significant difference between the bear bile group and the ursodeoxycholic acid group(P<0.05),and the positive control group(P<0.01).Conclusion 1.The interaction between liver and spleen lesions in a rat model of aplastic anemia.2.Mongolian medicine Wulan-10,bear bile powder and Ursodeoxycholic acid can improve the immune function and bone marrow hematopoietic function of rats with aplastic anemia,and have a certain therapeutic effect on this disease. |
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