| ObjectiveSesquiterpene lactones of Inula helenium L.is an active ingredient of traditional Chinese medicine Inula helenium L..It has antibacterial,deworming,liver-protecting and other pharmacological effects,and is widely used as traditional Chinese medicine.However,there is currently no systematic toxicological evaluation of the alkaloids.This study was conducted to observe the damage of forestomach mucosa to the gastric mucosa,to observe the mechanism of injury,and to compare the changes of gastric mucosal injury before and after stopping the treatment of Inula helenium L.total lactone,which is a reasonable application of lnula helenium L.as a traditional Chinese medicine resource.Provide test basis.Method150 SD rats,half male and half female,were randomly divided into 5 groups,and were intragastrically administered with normal saline,solvent CMC-Na and 20 mg/kg,120 mg/kg,and 600 mg/kg.The test animals died 10 days in each group after the gavage,and the remaining 5 animals stopped the administration and continued to feed for 28 days、Then the stomach tissues were sacrificed for gross observation and HE staining for histopathological diagnosis.TUNEL was used.Methods The apoptosis of gastric mucosa was detected.Immunohistochemical technique was used to detect the expression of TNF-α,TNFR1,TRADD,CK14 and caspase 8 in the forestomach mucosa.Statistical analysis Using SPSS 19.0 statistical software,Fisher’s exact probability method was used for gross anatomy findings and histological findings;TUNEL was analyzed by ANOVA,and immunohistochemical staining results were statistically analyzed using Fisher’s exact probability method and semi-quantitative method.P<0.05 was considered to be a significant difference.Result1.Sesquiterpene lactones of Inula/helenium L.can induce forestomach lesions in rats.The anatomical observation showed that the gastric mucosa of the control group,the vehicle group and the 20 mg/kg group had normal appearance before administration.In the 120 mg/kg and 600 mg/kg groups,the forestomach mucosa was pale and thick,nodules or masses.All gastric mucosa returned to normal after the recovery period.Microscopically,the control group,the vehicle group,the 20 mg/kg group had normal forestomach mucosa,and the 120 mg/kg group had keratinization in the forestomach,squamous epithelial simple hyperplasia and papillary hyperplasia,which occurred in the submucosa.Different degrees of congestion,edema,and inflammatory cell infiltration were observed in the mucosal layer.In addition to the above lesions,a squamous epithelial Pseudoepitheliomatous hyperplasiawas observed in the 600 mg/kg group.At the end of the recovery period,the gastric mucosa of each group was basically normal.It indicated that the phytolactone showed toxic effects on SD rats,and the target organs were in the forestomach,which could induce forestomach mucosal inflammation and proliferative lesions.2.The relationship between the pre-gastric mucosal lesions and the dose of the alkaloids:After the end of the administration period,70%of the rats in the 120 mg/kg group had an angle of the forestomach mucosa.60%of the rats had proliferative lesions in the forestomach mucosa;60%of the rats had hyperemia in the forestomach mucosa and 40%had edema.The incidence of these lesions was higher than that of the control group,the vehicle group and the 20 mg/kg group(P<0.05).In the forestomach mucosal injury of rats in the 600 mg/kg group,proliferative lesions occurred in 100%of the forestomach mucosa of rats,50%of which had squamous epithelial Pseudoepitheliomatous hyperplasiasignificantly higher than 120 mg/kg group;submucosal congestion and edema The incidence of inflammatory cell infiltration and mucosal hyperemia and inflammatory cell infiltration was also significantly higher than that in the 120 mg/kg group.The forestomach mucosa of the rats in the recovery period were normal.The unobserved toxic dose(NOAEL)of the eucalyptus lactone in this test was 20 mg/kg.This dose did not cause forestomach mucosal lesions in rats,and the lowest observed dose(LOAEL)was 120 mg/kg.The severity of the lesion increases with increasing dose.The lesion can be recovered after stopping the drug.3.The change of apoptotic index in the proliferative lesions induced by Tuyuxiang total lactone:the average apoptotic index of normal gastric mucosa in rats was 23.61%;the simple hyperplasia was 20.99%;the papillary hyperplasia was 18.63%,scale The epithelial Pseudoepitheliomatous hyperplasiawas 11.77%,which was significantly lower than that of the normal forestomach mucosa(23.61%)(P<0.01).It indicated that the apoptotic index of forestomach proliferative lesions induced by the alkaloids showed a decrease with the severity of hyperplasia..4.Immunohistochemical method was used to detect the expression levels of TNF-a,TNFR1,TRADD,CK14 and caspase 8 in normal gastric mucosa and proliferative lesions.The high expression rate of TNF-a was related to the severity of the lesion.From the normal gastric mucosa(0%),squamous epithelial simple hyperplasia(18%),squamous epithelial papillary hyperplasia(45%)to squamous epithelial Pseudoepitheliomatous hyperplasia(73%)gradually increased,the difference was statistically significant(P<0.05).The high expression rate of TNR1 is in normal gastric mucosa(0%),squamous epithelial simple hyperplasia(9%),squamous epithelial papillary hyperplasia(36%),squamous epithelial Pseudoepitheliormatous hyperplasia(73%),with proliferation And gradually increased,the difference was statistically significant(P<0.05).The high expression rate of TRADD is in normal gastric mucosa(0%),squamous epithelial simple hyperplasia(18%),squamous epithelial papillary hyperplasia(36%),squamous epithelial Pseudoepitheliomatous hyperplasia(64%),with degree of hyperplasia The increase was statistically significant(P<0.05).The high expression rate of CK14 was in normal gastric mucosa(0%),squamous epithelial hyperplasia(55%),squamous epithelial papillary hyperplasia(82%),squamous epithelial Pseudoepitheliomatous hyperplasia(91%),with degree of hyperplasia The increase was statistically significant(P<0.05).The positi’ve expression rate of caspase 8 was in normal gastric mucosa(100%),squamous epithelial simple hyperplasia(55%),squamous epithelial papillary hyperplasia(36%),squamous epithelial Pseudoepitheliomatous hyperplasia(9%),with The degree of hyperplasia decreased and the trend was consistent with the TUNEL results.The difference was statistically significant(P<0.05).By observing the difference in the expression of TRADD and CK14 between normal tissues and squamous epithelial papilloma,it was found that TRADD and CK14 were co?expressed in squamous epithelial Pseudoepitheliomatous hyperplasiainduced by Sesquiterpene lactones of Inula helenium L.ConclusionSesquiterpene lactones of Inula helenium L.has a toxic effect on SD rats,and the target organ of toxic effect is in the forestomach,which can induce inflammation and proliferative lesions of the forestomach.The unobserved toxic dose(NOAEL)of the general toxic effect of the alkaloids in this test was 20 mg/kg,which exceeded the clinical maximum dose by 6 times.The lowest observed toxic dose(LOAEL)was 120 mg/kg.Its toxic damage to the forestomach is reversible,and the lesion can be recovered after stopping the drug.Induction of squamous epithelial proliferative lesions in the forestomach may be related to the TNFa-TNFR1-TRADD-FADD-caspase 8 pathway-mediated apoptotic pathway,which may be due to a decrease in the dissociation of CK14 and TRADD.Thus,competitive inhibition of TNF-a-TNFRl and TRADD forms a complex,prevents the apoptosis signal from transmitting downward,reduces the activation of the downstream protein caspase 8,mediates apoptosis,and leads to the occurrence of squamous epithelial hyperplasia leisions. |
PDF Full Text Request