Study On The Clinical And Mechanism Of Self-made "Yangyinqingfei Huoxue Decoction" In The Treatment Of Radiation-induced Pneumonitis

BackgroundRadiotherapy is one of the most commonly used methods in the clinical treatment of malignant tumors.It is characterized by definite curative effect,small trauma,low toxicity and side effects,and is widely used in clinic.However,because radiation does not have the ability to distinguish,it can also kill tumor cells,but also affect normal tissue cells,so there will be varying degrees of side effects.These side effects are clinically called Radiation Induced Injury(RII).For patients who have received chest radiotherapy,this side effect is called Radiation Induced Lung Injury(RILI).Radiation Induced Lung Injury(RILI)[1]is divided into Radiation-induced Pneumonitis(RP)and Radiation Pulmonary Fibrosis(RPF)according to the duration of the disease.RP not only limits the dose of radiation received by patients,leading to a decline in local control rate of tumors,but also leads to intolerable adverse reactions that force the treatment to be interrupted,affecting later treatment and rehabilitation,and more serious will lead to related deaths.At present,western medicine can only take empirical treatment(symptomatic treatment)combined with hormone therapy to improve the symptoms of patients,which can only play a role in curing symptoms,and can not fundamentally solve this problem.The mechanism of RP is generally acknowledged as "cytokine cascade theory".This theory[2]holds that pulmonary parenchymal cells have oxidative damage after irradiation,leading to local aggregation of various inflammatory factors and chemokines,inducing inflammation,further causing aggregation of white blood cells and neutrophils in the body,forming a series of cascade-like reactions,and ultimately leading to the formation of a cascade reactiou,than RP occurs.Autophagy[3],as a highly conserved metabolic pathway in evolution,is an important mechanism for maintaining cell stability and activity under extreme conditions.It is characterized by the formation of a bilayer membrane structure of phagocytosis vesicles,encapsulating damaged proteins or organelles,forming a separate enclosed space,and then ombined with lysosomes to degrade and reuse the encapsulated substances to maintain cell homeostasis.Research data show that[4]oxidative stress injury induced by radiation can inhibit autophagy to a certain extent,thereby aggravating the damage response.Chaachouay H[5]further confirmed that up-regulation of autophagy activity can resist radiation-induced damage to a certain extent.Traditional Chinese medicine classifies RP into the category of "pulmonary mpotence" and "pulmonary arthralgia".It considers that RP belongs to the henomenon of deficiency of the original and excess of the original.There is a great orrelation between the occurrence of RP and insufficient lung fluid,exogenous heat oxicity,phlegm and blood stasis.Based on this theory,Professor Hou Wei, epartment of Oncology,Guang’anmen Hospital,Chinese Academy of Traditional hinese Medicine,has achieved good clinical results in the treatment of radiation neumonia by using the methods of Curative effect.Yangyin Qingfei Huoxue ecoction is composed of Ophiopogon japonicus,Asparagus,Polygonatum,Shengdi,huanbei powder,Almond,Houttuynia cordata,Panax quinquefolium,Panax otoginseng Powder and Licorice.Previous clinical studies[6]show that Yangyin ingfei Huoxue Decoction can effectively improve clinical symptoms such as hortness of breath and fatigue,reduce Watters score,and basic experiments[7]also how that Yangyin Qingfei Huoxue Decoction can significantly improve the ulmonary hemorrhage and edema in rats with radiation pneumonia,and also reduce he expression of local TNF-α,TGF-β and IL-6,thus reducing the lung validation eaction.In order to further explore the specific mechanism of Yangyin Qingfei uoxue Decoction in treating radiation pneumonia,we suggest that Yangyin Qingfei uoxue Decoction may improve radiation pneumonia by intervening alveolar acrophage autophagy,and carry out experimental verification.Research Purposes1.To verify the clinical efficacy of Yangyin Qingfei Huoxue Decoction in the treatment of radiation pneumonia through retrospective clinical research.2.To establish a cell model of radiation pneumonia of rat alveolar macrophages.observe the regulating effect of Yangyin Qingfei Huoxue Decoction on its autophagic activity,reveal the mechanism of Yangyin Qingfei Huoxue Decoction in treating radiation pneumonia at molecular level,explain the scientific mechanism of Yangyin Qingfei Huoxue Decoction in treating radiation pneumonia,and lay a scientific theoretical foundation and factual basis for the clinical application of Yangyin Qingfei Huoxue Decoction.Method1.Clinical researchRadioactive pneumonia cases from January 2010 to December 2017 in Guang’anmen Hospital of Chinese Academy of Traditional Chinese Medical Sciences were divided into treatment group and control group according to their treatment methods.The patients in treatment group were treated with Yangyin Qingfei Huoxue Decoction plus hormone(prednisone acetate tablet),while those in control group were treated with hormone only(prednisone acetate tablet).To verify the clinical efficacy of Yangyin Qingfei Huoxue Decoction through retrospective study.2.Basic ResearchⅠ Rat alveolar macrophage NR-8383 was used to construct the radiation pneumonia cell model by X-ray accelerator and to verify it.Ⅱ The effects of Yangyin Qingfei Huoxue Decoction on the proliferation of rat alveolar macrophages were detected by MTT assay.The effects of Yangyin Qingfei Huoxue Decoction on the oxidative damage of rat alveolar macrophages were determined by DNA oxidative damage index 8-OHdG and lipid oxidative damage index MDA.Ⅲ The autophagic flow in cells was observed by scanning electron microscopy.Ⅳ Western-blot was used to detect the expression of LC3 and P62 protein,and to observe the regulation of Yangyin Qingfei Huoxue Decoction on the autophagic activity of alveolar macrophages in rats.V The effect of Yangyin Qingfei Huoxue Decoction on LKB1/AMPK/mT OR/ULK1 pathway in rat alveolar macrophages was detected by Western-blot.VI Real-time PCR was used to detect the effect of Yangyin Qingfei Huox ue Decoction on LKB1,an upstream regulatory gene of AMPK pathway.Results1.Clinical researchEighty patients were included in the study.There were no significant diffe rences in age,sex,disease type,Watters score,RTOG score,KPS score and T CM symptom score between the two groups before treatment.After one course of treatment,in terms of Watters score,the Watters score of the treatment grou p was 7.95±2.62 and the Watters score of the control group was 9.5±2.69.T here was statistical difference between the two groups(p=0.015<0.05).In term s of RTOG rating,the total effective rate of the treatment group was 75%,the RTOG rating was 1.5±0.82,the total effective rate of the control group was 60%,and the RTOG rating was 1.65±0.83.there was no statistical difference between the two groups(P=0.43>0.05).In terms of KPS score,the average KPS score of the treatment group was 83.5±6.22 and that of the control gro up was 79.5±8.75,with statistical difference between the two groups(p=0.02<0.05).In the aspect of TCM symptoms,the score of cough symptoms in the tr eatment group was 1.1±0.81 and that in the control group was 1.05 ±0.78.Th ere was no statistical difference between the two groups(p=0.77>0.05).The sc ore of shortness of breath in the treatment group was 1.13±0.88 and that in t he control group was 1.65±0.95.The difference was statistically significant(p=0.01<0.05).The score of fatigue symptoms in the treatment group was 0.83±0.84 and that in the control group was 1.55±1.10.The difference was statis tically significant(p=0.001<0.05).2.Basic ResearchⅠ Radioactive pneumonia cell model can be simulated by irradiating alve olar macrophages of rats with 6Gy X-ray accelerator.Ⅱ Compared with the model group,Yangyin Qingfei Huoxue Decoction c an significantly improve the proliferation of alveolar macrophages in rats after irradiation,and the difference is statistically significant.Ⅲ Yangyin Qingfei Huoxue Decoction can significantly reduce the express ion of oxidative damage index 8-OHdG and MDA in rat macrophages after irr adiation,and the difference is statistically significant.Ⅳ By scanning electron microscopy,autophagosomes were found in the c ells of Yangyin Qingfei Huoxue group,which confirmed that Yangyin Qingfei Huoxue Decoction could activate autophagy.Ⅴ.Compared with the model group,the expression of P62 in macrophages of rats treated with Yangyin Qingfei Huoxue Decoction decreased,and the rat io of LC3II/LC3I increased.The difference was statistically significant.Ⅵ Compared with the model group,the expression of LKB1,AMPK and ULK1 increased and the expression of m-TOR decreased in the traditional Ch inese medicine Yangyin Qingfei Huoxue group,with statistical significance.Ⅵ Compared with the model group,the expression of LKB1 gene in the cells of Yangyin Qingfei Huoxue group was significantly increased,and the dif ference was statistically significant.Conclusion1.The therapeutic effect of hormone therapy plus Yangyin Qingfei Huoxue Decoction on radiation pneumonia is similar to that of hormone alone in redu cing RTOG grade and improving clinical symptoms of cough.It is superior to hormone alone in improving clinical symptoms such as fatigue and shortness of breath,lowering Watters score and improving quality of life.There are no obvious adverse reactions and good clinical safety.2.After 6Gy X-ray irradiation of alveolar macrophages in rats,the cell mo del of radiation pneumonia in vitro can be simulated.3.During the course of radiation pneumonia,oxidative damage of pulmona ry parenchymal cells occurred.4.Yangyin Qingfei Huoxue Decoction can activate AMPK-induced autopha gy by directly activating LKB1 and UKL1 and inhibiting the dual pathway of mTOR.5.Yangyin Qingfei Huoxue Decoction may alleviate oxidative stress injury by activating AMPK pathway to induce autophagy,thereby improving radiatio n pneumonia.