| Malignant pleural effusion(MPE)is a kind of pleural effusion characterized by the presence of malignant cells.The incidence rate of cancer patients is as high as 15%.It is a common complication leading to the loss of living ability in patients with malignant tumors.The common symptoms of most MPE patients include dyspnea,cough,chest pain and so on.Western medicine treatment mainly focuses on symptomatic treatment,including thoracic puncture,pleural fixation,thoracic perfusion and other methods.However,MPE is still difficult to cure,and it is unable to effectively control the accumulation of MPE again and improve the quality of life of patients.MPE belongs to the category of "Xuanyin" and "Piyin"in traditional Chinese medicine.On the basis of the original cancer disease,the Qi loss of Sanjiao and the dysfunction of lung,spleen,liver and kidney lead to the abnormal metabolism of water and fluid and the formation of phlegm and retained fluid.Finally,it is formed by competing with the pathological products of cancer and resting in the chest.The pathological nature belongs to the deficiency of origin and the excess of signs.On the basis of positive deficiency,such as deficiency of Qi and Yin,deficiency of lung and spleen,there are phlegm,blood stasis,Qi stagnation,cancer toxin and so on.In terms of treatment,most of them take Yiqi Xiaoshui method as the treatment method,combined with promoting blood circulation and removing blood stasis,regulating Qi and dispersing knot,anti-cancer and detoxification,etc.Yiqi Xiaoshui method is widely used in the clinical treatment of MPE.The early clinical trial proved that the effective prescription of Yiqi Xiaoshui method Xiaoshui formula combined with cisplatin chest perfusion could effectively improve the clinical symptoms and improve the quality of life of patients.On this basis,the experimental study found that Xiaoshui formula with the function of Yiqi Xiaoshui method might inhibit the formation and development of MPE by affecting the polarization of tumor-associated macrophages(TAMs).Recent studies have proved that tumor derived exosomes played an important role in mediating immunosuppression and regulating tumor microenvironment.Exosome derived microRNA-21(miRNA-21,miR-21)can promote the progress of MPE by changing the immune microenvironment and other mechanisms.Preliminary experiments also suggested that Xiaoshui formula,which had the effect of Yiqi Xiaoshui,could regulate miR-21,an exosome derived from tumor cells.Based on the previous research results and the latest research progress,we further refine the high-level evidence of the clinical efficacy of Yiqi Xiaoshui method in the treatment of MPE.Taking TAMs as the starting point,to explore whether the inhibitory effect of Xiaoshui formula on MPE was related to the regulation of miR-21 in tumor cell-derived exocrine secretion,and then affect the polarization of TAMs.Objective:Through data mining,the experience of Professor Hua Baojin in the treatment of MPE will be analyzed.And carry out systematic review and meta-analysis of the clinical efficacy and safety of Yiqi Xiaoshui method in the treatment of MPE in order to provide high-level evidence,and support for the prevention and treatment of MPE by Yiqi Xiaoshui method.On this basis,with the help of molecular biology technology,the internal mechanism of Yiqi Xiaoshui method in the prevention and treatment of MPE will be explored to find the mechanism of regulating tumor cell-derived secretion by Xiaoshui formula,affecting the polarization of TAMs in vivo by miR-21,and then inhibiting the occurrence and development of MPE.And provide scientific basis for the prevention and treatment of MPE by Xiaoshui formula.Method:1 Through the methods of systematic review and meta-analysis,this paper evaluated and summarized the efficacy and safety of Yiqi Xiaoshui method in the treatment of MPE,in order to provide high-level evidence supporting for the clinical application of Yiqi Xiaoshui method in the treatment of MPE;2 Based on the method of data mining,this paper analyzed and summarized the experience of Professor Hua Baojin in the treatment of MPE;3 The MPE model of mice bearing Lewis lung cancer cells was established.Xiaoshui formula,an effective prescription of Yiqi Xiaoshui method,was intervened alone(high,medium and low doses)or combined with cisplatin.The volume of MPE,the mass of tumor in situ and the weight of mice were observed to verify the inhibitory effect of Xiaoshui formula,an effective prescription of Yiqi Xiaoshui method,on MPE in mice;4 The proportion of M1 and M2 TAMs in spleen and the expression of TAMs marker protein in pleural effusion or pleural implant tumor of MPE mice were detected by flow cytometry,Western blot and immunohistochemistry,respectively.The effects of Xiaoshui formula alone(high,medium and low doses)or combined with cisplatin on the phenotype of TAMs were observed;5 To construct the Lewis cell line stably transfected by miR-21 gene overexpression and miR-21 gene interference,so as to provide technical support for subsequent experiments;6 The MPE model of Lewis lung cancer bearing mice,the MPE model of Lewis lung cancer bearing overexpression of miR-21 gene and the MPE model of Lewis lung cancer bearing interference of miR-21 gene were constructed.Through the intervention of Xiaoshui formula,an effective prescription of Yiqi Xiaoshui method,the formation of MPE and the quality of tumor in situ in mice were observed.The proportion of M1 and M2 TAMs in spleen of MPE mice,the expression of TAMs marker protein in pleural effusion and pleural implant tumor,and the expression of TAMs cytokines in pleural effusion were detected by flow cytometry,Western blott,immunohistochemistry and ELISA to explore whether Xiaoshui formula can affect the polarization of TAMs and inhibit the formation and development of MPE by regulating tumor cell miR-21.At the same time,extracted the exosomes from mouse pleural effusion,detected the expression of miR-21 in exosomes by fluorescence quantitative PCR,and explored the effect of Xiaoshui formula on the expression of miR-21 in tumor cell exosomes;7 The serum containing Xiaoshui formula was prepared,and the overexpression of miR-21 gene and the interference of miR-21 gene with Lewis and Lewis lung cancer cells were intervened.Extracted exosomes from cell culture supernatant of each group,detected the expression of miR-21 in exosomes of each group by fluorescence quantitative PCR,and observed the effect of Xiaoshui formula on the expression of miR-21 in exosomes from tumor cells;8 The exosomes extracted in research method 7 were co-cultured with macrophages,and the ratio of M1 and M2 TAMs cells was detected by flow cytometry fluorescence sorting technology.It was observed whether Xiaoshui formula affected the polarization of TAMs by regulating miR-21 in exosomes derived from tumor cells.Results:1 The meta-analysis included 21 randomized controlled studies on the treatment of MPE with Yiqi Xiaoshui method,involving 1698 subjects.The results showed that Yiqi Xiaoshui method combined with systemic chemotherapy or thoracic perfusion could significantly improve the effective rate and KPS score of MPE,reduce the incidence of adverse reactions and improve the quality of life of subjects compared with systemic chemotherapy or thoracic perfusion alone,and the differences between the above groups were statistically significant(P<0.05).However,GRADE evidence quality evaluation showed that the quality grade of effective rate was medium,and the quality grade of KPS score and adverse reaction rate were low;2 A total of 410 outpatient medical records of MPE patients were included in the study.Based on the statistical analysis of data mining,it was found that the syndrome types of traditional Chinese medicine were mainly lung and spleen deficiency,drinking and stopping chest threat syndrome,and the treatment was based on the method of Yiqi Xiaoshui.Commonly used prescriptions included Buzhong Yiqi Decoction,Linggui Zhugan decoction,Tingli Dazao xiefei decoction,Xuanfu Daizhe Decoction,etc.A total of 194 traditional Chinese medicines were involved,of which deficiency tonifying drugs were the most commonly used,and Qi tonifying drugs were the most frequently used.There were also cough relieving,phlegm resolving and asthma relieving drugs,diuretic and hygroscopic drugs and food reducing drugs.In the aspect of nature and taste,warm and sweet drugs were used most frequently,and they mainly returned to the spleen and lung meridians.The top 20 drugs used frequently include Tinglizi,Jiaomu,Shenghuangqi,Fuling,Baizhu,Zexie,Chenpi,Shengjiang,Dazao,Chaoguya,Chaomaiya,Fangji,Hongjingtian,Chaoshenqu,Chaoshanzha,Shengyiyiren,Shengshigao,Zhuling,Gancao and Shegan.The drug combination of Tinglizi-Jiaomu-Huangqi had the highest frequency.The core formula included Shenghuangqi,Baizhu,Fuling,Chenpi,Fangji,Zexie,Jiaomu,Tinglizi,Chaoshenqu,Chaoshanzha,Chaoguya,Chaomaiya,Shengjiang,Dazao,Hongjingtian,and five new formula combinations for the treatment of MPE were obtained based on the formula composition law of entropy method;3 After Xiaoshui formula intervention,the volume of pleural effusion and the mass of thoracic implant tumor in MPE model mice decreased.With the increase of Xiaoshui formula concentration and dose,the effect of inhibiting MPE and thoracic implant tumor was more obvious.Xiaoshui formula combined with cisplatin could inhibit the growth of MPE and thoracic implant tumor better than cisplatin alone.Xiaoshui formula had little effect on the weight of mice while controlling the development of MPE,while cisplatin and cisplatin combined Xiaoshui formula had a greater effect on the weight of mice;4 Xiaoshui formula alone(high,medium and low doses)could inhibit the progress of MPE.The high dose of Xiaoshui formula had the best effect on increasing the proportion of M1 TAMs in mouse spleen,reducing the proportion of M2 TAMs,increasing the expression of M1 TAMs marker protein in mouse pleural effusion,reducing the expression of M2 TAMs marker protein,increasing the expression of M1 TAMs marker protein in mouse thoracic implant tumor and reducing the expression of M2 TAMs marker protein,which was better than cisplatin.The intervention effect of Xiaoshui formula combined with cisplatin group was better than that of cisplatin alone group;5 The Lewis cell line stably transfected by miR-21 gene overexpression and miR-21 gene interference were successfully constructed by lentivirus stable transfection technology,and verified from the gene level;6 Overexpression of miR-21 gene in tumor cells could significantly increase the volume of mouse MPE and the quality of thoracic implant tumor,increase the proportion of M2 TAMs in mouse spleen,reduce the proportion of M1 TAMs,increase the expression of M2 TAMs marker protein in mouse MPE,reduce the expression of M1 TAMs marker protein,increase the expression of M2 TAMs marker protein in mouse thoracic implant tumor,reduce the expression of M1 TAMs marker protein,and increase the expression of M2 TAMs cytokine.The expression of cytokines in M1 TAMs was down regulated.The expression of miR-21 gene in tumor cells was disturbed or the results after Xiaoshui formula intervention were opposite to the above situation.It was found that the expression of miR-21 in MPE exosomes of mice in miR-21 overexpression group increased,and the expression of miR-21 decreased after Xiaoshui formula intervention;7 The overexpression of miR-21 after the intervention was successfully extracted.Lewis and miR-21 interfered with Lewis and Lewis cell exosomes.It was found that the serum containing Xiaoshui formula could inhibit the expression of miR-21 in tumor cell-derived exosomes in vitro;8 Co-culture of miR-21 overexpression Lewis cell exosomes with macrophages could increase the proportion of M2 TAMs and reduce the proportion of M1 TMAs.After the intervention of serum containing Xiaoshui formula,the proportion of M2 TAMs decreased and M1 TMAs increased after the co-culture of Lewis cell exosomes and macrophages with miR-21 overexpression.Conclusions:1 The method of Yiqi Xiaoshui of traditional Chinese medicine was widely used in the clinical treatment of MPE.It could effectively improve the effective rate of MPE treatment and the KPS score of patients,reduce the incidence of adverse reactions and improve the quality of life of patients.It had definite clinical curative effect.It was an effective treatment method of traditional Chinese medicine for the prevention and treatment of MPE;2 Professor Hua Baojin mainly used the method of Yiqi Xiaoshui to treat MPE;3 Xiaoshui formula,an effective prescription of the method of Yiqi Xiaoshui,could effectively inhibit the occurrence and development of MPE.Its mechanism might be related to inhibiting the expression of miR-21 in the secretion from tumor cells,thereby inhibiting the polarization of tumor related macrophages to M2 and increasing the proportion of M1 TAMs. |
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