| Objective: The NASH model of SD rats was established by high-fat diet induction to observe the pathological changes of the spleen of NASH rats.To explore the correlation between Spleen-Derived IL-10 and NAFLD,and to further explore the possible ways of spleen and Spleen-Derived IL-10 participating in the protection of NAFLD.Part ONE Animal experimentMethods:(1)20 male SD rats aged 4-5 weeks and weighing 80-120 g were fed adaptively for one week.(2)20 healthy SD rats were randomized into two groups: 8 in the control group were fed regular feed and 12 in the experimental group were fed with high-fat feed.(3)Two rats were randomly killed from the experimental group at the 8th and 12 th week,and liver tissue was extracted for HE staining and microscopic observation was given to see if NASH pathological changes was achieved,12 weekend modeling was observed under microscope.After fasting without water for 12 h,all rats in the experimental group and control group were sacrificed.(4)The blood of inferior vena cava was collected and the biochemical indexes(ALT,AST,GGT,TG,TC,LDL)were detected by ELISA method.(5)Liver tissue and spleen tissue were extracted and paraffin-embedded to make pathological sections,HE staining,microscopic observation of liver and spleen tissue pathological changes.After immunohistochemistry,the expression of IL-10 in the spleen was measured by tissue sectioning digital scanner and imaging system.Results:(1)Histopathological changes of liver: the control group observed under the liver microscope at the end of 12 weeks: the structure of hepatic lobule was intact;the hepatocytes were arranged neatly,there were no lipid droplet vacuoles in the cytoplasm,and there was no inflammatory cell infiltration in the portal area.The observation of the liver microscope at the end of the 12 th week of the experimental group showed that the structure of hepatic lobule was destroyed,the arrangement of hepatic cord was disordered,hepatocyte steatosis was obvious,hepatocyte steatosis appeared in almost all lobules,the cytoplasm was full of lipid droplet vacuoles,and lymphocyte and neutrophil infiltration could be seen in the portal area.(2)Histopathological changes of spleen: The control group and the experimental group were observed under the microscope of the spleen at12 weeks.Compared with the control group,the edge area of the spleen pulp of rats in the experimental group was slightly wider,and hyperemia,lymph nodes increased,and the red pulp was moderately bruised.(3)Comparison of serological indexes: Compared with the control group,the serum levels of ALT,AST,GGT,TG,TC and LDL in the experimental group were higher,and the levels of ALT,GGT,TG and LDL were statistically significant(P < 0.05).(4)Comparison of IL-10 expression in spleen:Compared with the control group,the expression of IL-10 in the spleen of rats in the experimental group decreased significantly,and the difference was statistically significant(P<0.05).Conclusion:High-fat diet-induced changes in spleen histopathology in rats with nonalcoholic steatohepatitis SD,and decreased expression of IL-10 in the spleen of NASH rats.Part Two clinical studyMethods:(1)From November 2021 to August 2022,abdominal ultrasound examinations were selected in the physical examination center,outpatient clinic and inpatient of the Affiliated Hospital of Yan’an University as the research subjects.According to the inclusion criteria,136 cases of research subjects were finally determined,91 cases in the NAFLD group and 45 cases in the healthy control group.According to whether the patients had abnormal blood lipids,they were divided into normal lipid group(81 cases)and dyslipidemia group(55 cases).(2)General data on the study subjects were collected,including age,sex,body mass index(BMI),blood pressure(SBP/DBP),heart rate.(3)Serum biochemical indexes were collected,all on an empty stomach for more than 8 hours,and venous blood samples were drawn in the morning,Including alanine aminotransferase(ALT),aspartate aminotransferase(AST),alkaline phosphatase(ALP),γ-glutamyl transpeptidase(GGT),triglycerides(TG),total cholesterol(TC),high-density lipoprotein(HDL),low-density lipoprotein(LDL),fasting blood glucose(FBG).The ELISA method measures IL-10 and collects the thickness of the spleen of the study subjects.(4)The changes of general data,liver enzymes,blood lipids,fasting blood glucose,serum IL-10 and spleen thickness / BMI were analyzed and statistically analyzed.Results:(1)General demographic characterization: Among the 136 participants,the youngest was 25 years old and the oldest was 75 years,with a mean age of 44.79 ± 12.12 years;85cases were male,accounting for 62.5%,51 cases were female,accounting for 37.5%.The average BMI value of the subjects was 25.38kg/m2.(2)Comparison of general information: Comparing the general data between the two groups,it can be seen that the mean values of age,BMI,SBP,DBP and heart rate in the NAFLD group are higher than those in the healthy control group.There were significant differences in age,BMI and SDP between the two groups(P<0.05),but there were no significant differences in sex,DBP and heart rate(P>0.05).(3)Comparison of liver function enzymology: Compared with the healthy control group,the NAFLD group had higher levels of ALT,AST,ALP and GGT,and the difference was statistically significant(P<0.05).(4)Comparison of lipid: Compared with the healthy control group,the NAFLD group had higher TG,TC,LDL levels and lower HDL levels,and there was a significant difference in TG,TC and HDL levels between the two groups(P<0.05),although the LDL levels in the NAFLD group were higher than those in the healthy control group(P>0.05).(5)Comparison of fasting glucose: Compared with the healthy control group,NAFLD group had a higher level of FBG,and the difference was statistically significant(P<0.05).(6)Comparison of serum IL-10: Compared with the healthy control group,the NAFLD group had a lower level of serum IL-10,and the difference was statistically significant(P<0.05).(7)Comparison of spleen-thickness/BMI between the two groups: The spleenthickness/BMI of the NAFLD group was smaller than that of the healthy control group,and the difference was statistically significant(P <0.05).(8)Correlation analysis between different indexes and serum IL-10: There was a negative correlation between serum IL-10 and systolic blood pressure(r = 0.236,P <0.018)and FBG(r = 0.173,P < 0.048).(9)The results of multivariate logistic regression analysis of NAFLD disease showed that the risk factor of NAFLD was BMI(OR=14.635 2.296,93.301),and the protective factor of NAFLD was serum IL-10 level(OR=0.961(0.937),0.9985).(10)Comparison of serum IL-10,spleen thickness / BMI between normal blood lipid group and dyslipidemia group: The results showed that the IL-10 of dyslipidemia group was lower than that of normal blood lipid group(P>0.05).Spleen-thickness / BMI in dyslipidemia group was lower than that in normal blood lipid group(P<0.05).Conclusion:The decrease of serum IL-10 level is related to the occurrence and development of non-alcoholic fatty liver disease,and Serum IL-10 is a protective factor of NAFLD.Spleen may affect the occurrence and development of NAFLD by regulating lipid metabolism,and the underlying mechanism may involve Spleen-Derived IL-10. |
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