Experimental Study On The Relationship Between Cooling Effect Of Saikosaponin A And AC,cAMP,PKA And NF-?B

At the present stage,antipyretic therapy in Western Medicine is the main clinical fever treatment.While,the contraindications of various complications,obvious side effects and the unsatisfactory treatment effect are the main obstacles in clinical treatment all the time.For nearly a thousand years,Bupleurum is the main drug used in fever treatment by traditional Chinese medicine science.It has remarkable efficiency and little side effect in the treating the patients who developed fever with contraindications or poor treatment of western medicine.It has greatly improved the quality of the patient's life,laying a foundation for the subsequent rehabilitation.The application traditional Chinese medicine instead of traditional western medicine is a new path for treating the fevered patients in the development of medical treatment in the future,and it is also a further research goal of the health workers.Bupleurum as a kind of traditional Chinese medicine is mainly used to treat fever clinically.It has many advantages such as stable curative effect,less repetition,better symptoms,less side effects,and so on.Unfortunately,the mechanism of treating fever is still unclear.EP3 signal transduction pathway plays an important role in body temperature regulation.In this study,SD rats were used as targets,and AC,cAMP,PKA,and NF-?B were used as observation indexes.The contents of each protein or ion were analyzed by histochemistry,ELISA,western blot,Real-time PCR,and EP3 signal transduction pathways.Changes and expressions were discussed to investigate the effect of Bupleurum on the signal transduction pathway of EP3 down-regulation,and to elucidate the mechanism of the treatment of fever by Bupleurum.This work provides theoretical and experimental basis for the correct and extensive application of clinical treatment of fever by Bupleurum,laying a foundation for the promotion of Bupleurum in the international medical field.MethodsIn this experiment,the healthy SD rats with the mass varying from 180 g to 220 g were selected as the experimental subjects.After the rats were anesthetized,a stereotaxic apparatus was placed into the rat's brain,then the lateral ventricle tube placed and fixed on the rat's brain bone.To increase the stability between the lateral ventricle tube and the brain bone,a small screw was fixed on the brain bone.After the surgery,the rates were injected by anti-inflammatory drugs to avoid the wound inflection.The rats were allowed to return to their normal state by normal feeding for one week.The temperature of the rats were measured in succession,to decrease the stress reaction to a minimum by acquainting the rats with the operator's skill.When the surgery was conducted,the temperature of the rats was measured for 3 times and the average value was used as the basal body temperature.Then,the drug was injected directly into the lateral ventricle of the rat by the tube.During the surgery,the temperature of the rat's rectum was measured and recorded every 30 min.At the end of the experiment,the rats were sacrificed by the decapitation method and the perfusion method.After being killed by decapitation,the rats' hypothalamus was quickly placed in liquid nitrogen.The total protein,cytoplasmic protein and nuclear protein were extracted according to the instructions of the protein extraction kit,and the mRNA was extracted according to the kit.Then the ELISA,Western and RT-PCR experiments were performed.After being sacrificed by perfusion method,hypothalamus tissue was taken from the rats for embedding,sectioning,and finally the immunohistochemical experiment was conducted.ResultsAfter injecting saikosaponin A into the fever rats,the body temperature of rats showed a significant downward trend(P<0.05).The content of AC and cAMP in hypothalamus of IL-? group was significantly higher than that of artificial cerebrospinal fluid group(P<0.05).When the temperature of hypothermia rats induced by saponin A decreased,the content of AC and cAMP in the hypothalamus was lower than that of the IL-1? group(P<0.05)obviously.The expression of fine PKA in the IL-1? group was obviously higher than that of the artificial cerebrospinal fluid group and the bupleurum Saponin A group(P<0.05).The expression of NF-?B in IL-1? group was obviously higher than that in artificial cerebrospinal fluid group and saikosaponin A group(P<0.05).The expression of total protein,cytosolic protein and nuclear protein in NF-?B in IL-1? group was significantly increased.The amount of NF-?B mRNA was obviously higher than that of artificial cerebrospinal fluid group and saikosaponin A group(P<0.05).ConclusionsThe cooling effect of saikosaponin A may be related to the decreased expression of AC,cAMP,PKA and NF-?B in EP3 signal transduction pathway.Saikosaponin A may play a role in cooling by inhibiting the EP3 signal transduction pathway.