| Research Background:Surgical therapy,chemotherapy,radiotherapy,targeted therapy and immunotherapy are the five main methods of malignant tumor treatment in modern medicine.Radiotherapy(RT),referred to as Radiotherapy,is widely used as first-line therapy in the treatment of lung cancer,esophageal cancer,breast cancer and mediastinal tumor,etc.Although the progress of radiotherapy technology and the emergence of multi-mode therapy have prolonged the survival time of many cancer patients,the Lung toxicity such as radiation-induced Lung Injury(RILI)accompanied by low-dose long-term irradiation is the main complication of thoracic tumor radiotherapy.It is still the main factor to limit the radiation dose in the target area of chest tumor.According to statistics,about 50-70%of patients with chest tumors receive radiation therapy,and about 5%-15%of these patients will develop RILI.RILI can be divided into acute and chronic according to the time of occurrence.Acute Radiation lung injury,also known as Radiation Pneumonitis(RP),occurs mostly in 4-12 weeks after exposure.If the disease progresses without effective treatment,it develops into chronic disease,which becomes radiation-induced pulmonary fibrosis(RIPF).Radiation lung injury is mainly diagnosed on the basis of symptoms,medical history and imaging changes.Its clinical symptoms are cough,fever,chest pain,dyspnea and so on.Imaging examination suggested inflammatory and fibrotic changes in the irradiated site.At present,the main measures for the treatment of acute radiation lung injury are glucocorticoids,antibiotics and other drug therapy,but the effect is not good and the adverse reactions are large.Therefore,it is of great significance to carry out drug therapy research related to radiation lung injury to improve the dose of radiation in tumor target area,and improve the quality of life and prognosis of patients.It was found that ionizing radiation-induced activation of p38 MAPK/NF-κB pathway,various cytokines,such as Interleukin-6(IL-6),Tumor necrosis factor-α,(TNF-α)and Transforming growth factor-β(TGF-β)expression may be the main mechanism of lung injury caused by radiation.Traditional Chinese medicine believes that radiation belongs to the category of "fire poison" and "fiery poison evil",which can burn and consume lung jin and damage lung collateral.Therefore,pulmonary dryness injury is the core pathogenesis of RILI,and can also trap sputum and blood stasis and other pathological products.According to this mechanism,the supervisor combined with years of clinical practice,mainly nourishing Yin,moistening lung and generating fluid,simultaneously clearing heat and removing phlegm,promoting blood circulation and removing blood stasis,on the basis of the prescription of Shashen Maidong Decoction,through addition and reduction,the decoction of Shashen Jiegeng Decoction has achieved good clinical efficacy.However,the specific mechanism of its action is still lacking.Now through the clinical curative effect observation and animal experiment research,from the improvement of the clinical symptoms,histopathology,signaling pathways,cytokines and other aspects of exploration,for the straight ladybell provide theoretical basis for treatment of radioactive lung injury of Shashen Jiegeng Decoction for traditional Chinese medicine in the application of radioactive lung injury,clinical treatment and follow-up have certain guiding significance to the further research direction.Clinical Research:Objective:To observe the clinical efficacy of Shashen Jiegeng Decoction in the treatment of acute radiation lung injury and its effect on serum cytokines.Content and methods:Patients with acute radiation lung injury who were treated in the oncology outpatient department and ward of Xiyuan Hospital,China Academy of Chinese Medical Sciences from November 2018 to February 2022 were selected.A prospective clinical single-arm study was used.Patients who met the inclusion criteria were given 1 dose of Shashen Jiegeng Decoction daily for 28 days.At the beginning of the visit,14 days after treatment and 28 days after treatment,The levels of serum cytokines(IL-6,TNF-α),radioactive lung injury grade,functional status score(KPS score)and clinical symptom score(cough,fever,dyspnea,chest pain)were assessed and recorded according to clinical symptoms,CT imaging and laboratory examination.The clinical treatment effect was evaluated by UICC(Union for International Cancer Control)efficacy grading standard for radioactive lung injury,and the data were statistically analyzed by SPSS26.0 statistical software.Results:30 patients were planned to be enrolled,25 patients were actually enrolled with acute radiation lung injury.After the observation period,5 patients were cured,16 were improved,and 4 were invalid by UICC grading.The total clinical response rate was 84.7%,higher than the general response rate of hormone or antibiotic therapy in previous studies.After treatment,serum inflammatory factors(IL-6,TNF-α),lung injury grade(RTOG grade)and life ability score(KPS score)were improved,and the differences were statistically significant(P<0.05).Symptom scores of cough,fever,dyspnea and chest pain were significantly improved after treatment,with statistical significance(P<0.05).Conclusion:Shashen Jiegeng Decoction can improve the clinical efficacy of acute radiation lung injury,reduce the levels of serum inflammatory factors IL-6,TNF-α and grade of acute radiation lung injury,and improve the clinical symptoms and quality of life of patients with cough,dyspnea,fever,chest pain,with clear efficacy.Animal Experiments:Objective:To investigate whether the mechanism of Shashen Jiegeng Decoction in the treatment of radioactive lung injury is related to the inhibition of P38 MAPK/NF-κB signaling pathway and the down-regulation of related inflammatory factors(IL-6,TNF-α,TGF-β).Content and Methods:Fifty-four male C57BL/6 mice were randomly divided into 6 groups:Control group(Control group),Model group(Model group),Anisomycin group(An group),SB 203580 group(SB group),Shashen Jiegeng Decoction group(SSJG group,equivalent dose),Shashen Jiegeng Decoction+Anisomycin group(SSJG+An group),with 9 rats in each group.Except for the Control group,the other five groups received a single whole-chest irradiation of 15Gy under the linear accelerator to construct an animal model of lung injury in C57BL/6 mice.Mice in the An group were intraperitoneally injected with P38 MAPK pathway-specific activator Anisomycin solution(5mg/kg/day),mice in the SB group were intraperitoneally injected with P38 MAPK path-specific inhibitor SB203580 solution(5mg/kg/day).SSJG group was given 2.365 g/ml Shashen Jiegeng Decoction(equivalent dose)by intragastric administration,SSJG+An group was given Anisomycin solution(5mg/kg/day)by intraperitoneal injection combined with 2.365 g/mL Shashen Jiegeng Decoction(equivalent dose)by intragastric administration.Control group and Model group were given 0.9%normal saline intragastric administration,once a day for 28 days.During the experiment,the general state changes of mice in each group were observed and recorded weekly,including body weight,mental state,activity and depilation.On day 28,the samples were collected after general anesthesia with 5%chloral hydrate(0.1ml/10g).Hematoxylin-eosin(HE)staining and Masson staining were performed on the double lung lobes of mice,and the semi-quantitative analysis of alveolitis and pulmonary fibrosis was conducted under light microscope.The expression levels of inflammatory factors IL-6,TNF-αand TGF-β in peripheral blood were determined by Enzyme-Linked immunosorbent assay(ELISA).Western blot was used to detect the expression of related pathway proteins P38,P-P38,P65 and P-P65 in lung tissues.Results:Body weight:During the feeding period,the body weight of C57BL/6 mice in all groups except the Control group decreased to varying degrees in the first week after radiation exposure,especially the An group and Model group.After 2-4 weeks,the body weight of C57BL/6 mice in the irradiated group gradually increased,especially the SB group and SSJG group.Pathological HE staining:the structure of bronchial mucosa and alveoli in the Control group was clear and complete,and the epithelial cells were neatly arranged,and no inflammatory cell infiltration was observed in the lung interstitium.In Model group and An group,the pulmonary bronchial lumen was dilated,tortured,local destruction,alveolar structure was unclear,neutrophils increased,and mucosal epithelial degeneration and exfoliation were observed under microscope.Other intervention groups had different degrees of relief.SSJG had the least inflammation and no obvious fibrous tissue hyperplasia.Both SB group and SSJG+An group showed mild to moderate inflammation,but the fibrosis degree of SB group was lighter than that of E group.Compared with the Control group,the other 5 groups had higher Szapiel scores in lung tissues,and the differences were statistically significant(P<0.05).Compared with the Model group,the other 4 groups had no significant difference in Szapiel alveolitis scores(P>0.05).The difference was statistically significant(P<0.05).Masson staining:In the Control group,the alveolar and bronchial collagen fibers were relatively less thickened,and the density of blue collagen fibers was lower under the microscope.Compared with the Control group,the lung tissues of mice in the other five groups all showed fibrosis changes to varying degrees.Dense blue optical density areas were observed in the Model group and the An group,with a wide distribution range.The fibrosis degree of SB group,SSJG+An group and SSJG group was mild.Ashcroft score of lung fibrosis in mice showed that compared with the Control group,Ashcroft score of lung tissues in the other four groups except SSJG group was higher,and the differences were statistically significant(P<0.05),suggesting that there was a tendency of lung fibrosis in mice after radiation lung injury modeling.Compared with the SSJG group,The difference was statistically significant(P<0.05),and there was no significant difference in Ashcroft score between the Model group and the other four modeling groups(P>0.05).ELISA results:The levels of inflammatory cytokines IL-6,TNF-α and TGF-β in peripheral serum of mice in Control group were all at low levels.Compared with the Control group,the contents of il-6,TNF-α and TGF-β in peripheral blood of Model group and An group were significantly increased,except the serum IL-6 content of SSJG group,there were statistical differences(P<0.05).Compared with Model group,the levels of IL-6,TNF-α and TGF-β in peripheral blood of SSJG group were decreased,and the differences were statistically significant(P<0.01).The TNF-αcontent in SSJG+An group was lower than that in Model group,and the difference was statistically significant(P<0.01).Compared with An group,the expressions of IL-6,TNF-α and TGF-β in serum of SSJG group were significantly lower than those of An group,the differences were statistically significant(P<0.0001).The expressions of TNF-α and TGF-β in SSJG+An group were lower than those in An group,the difference was statistically significant(P<0.001).Western Blot results:The protein expression levels of p38,P-P38,P65 and P-P65 pathways were the lowest in the Control group.Compared with the Control group,the protein expression levels of P-P38,P65 and P-P65 pathways were higher in the Model group.An group and SB group,and the protein content of the An group was the highest.The differences were statistically significant(P<0.05),indicating that p38 and P65 related proteins were phosphorylated and p38 MAPK/NF-κB signaling pathway was activated in lung tissues of irradiated lung injury model mice.Compared with Model group,the contents of P38,P-P38,P65 and P-P65 in lung tissues of mice in An group were increased,while the proteins of P38,P-P38,P65 and P-P65 in SB group were decreased.Suggesting that P38 inhibitors can reduce the expression of related proteins in MAPK/NF-κB pathway.The expression levels of P-P38,P-P65 and P65 in SSJG group were significantly decreased,and the differences were statistically significant(P<0.05),suggesting that Shashen Jiegeng Decoction could significantly inhibit the protein expressions of P-P38,P-P65 and P65.Although there was no statistical difference between SSJG+An group and Model group,the expression level of SSJG+An group was significantly lower than that of An group.Compared with An group,the relative protein expression levels of P-P3 8 and P65 in lung tissues of mice in SSJG+An group were significantly decreased(P<0.05),and the protein levels of P38 and P-P65 were also decreased,suggesting that Shashen Jiegeng Decoction could reverse the excitatory effect of anisomycin,a pathway activator.Conclusion:(1)Shashen Jiegeng Decoction can inhibit the activation of P38 MAPK/NF-κB pathway,down-regulate the levels of inflammatory factors IL-6,TNF-α and TGF-β by inhibiting the protein expression of P38,P-P38,P65 and P-P65 signaling pathways,and improve the tendency of lung inflammation and pulmonary fibrosis in mice with lung injury caused by radiation.(2)P38 MAPK/NF-κB signaling pathway is an important regulatory pathway in the course of RILI disease,and is also an effective target for the treatment of Shashen Jiegeng Decoction,providing theoretical basis and data support for guiding the clinical prevention and treatment of radioactive lung injury in Traditional Chinese Medicine and drug development. |
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