| Deficiency-stasis-poison is the basic pathogenesis of lung cancer,which is the main causes and the key pathological mechanism of development transfer or progression of lung cancer.So,Stasis poison,as a kind of new pathogenic factors,is more and more brought to the attention of the academia In this paper,we summarized the clinical and laboratory characteristics of lung cancer patients with qi-deficiency and blood-stasis syndrome systematically,hoping to find new clues to control lung cancer and give evidence of clinical applications.Meanwhile,We discussed the effect of DiterpenoidTanshinones on lung cancers and its mechanism of PERK-EIF2α signaling pathway.The first part is theoretical researchIn the paper,we systematically sort out and demonstrate the history,etiology,pathogenesis,biological basis of the theory of poison and blood stasis,and focused on the pathogenesis of stasis-poison and the interventional effects of drugs which can activate blood circulation and remove stasis to lung cancer.We concluded that deficiency of body energy is the root,qi and blood running obstacles is the basics generation,the exogenous of pathogenic qi and emotional internal injuries play important inducements of poison and blood stasis.The second part is clinical researchObjective To investigate the clinical and laboratory characteristics of lung cancer patients with qi-deficiency and blood-stasis syndrome,we have used the retrospective clinical study to analyze the correlation of coagulation indexes with pathological type,tumor stage,smoking status,Hoping to provide reference for clinical of lung cancer.Methods Selected 60 cases of the first affiliated hospital of Zhejiang Chinese Medical University with lung cancer who were syndrome differentiation for qi deficiency and blood stasis from January 2014 to October 2015.Through the descriptive analysis,we collected the individual patients data about general clinical information,such as medical history,agglutinative index,and so on.At the same time,we analyed the difference of blood coagulation index with the same period internal hospital check-up for the 60 cases of normal controls.Analysis blood coagulation in lung cancer patients with qi deficiency blood stasis,type of index and the clinical pathological stage,smoking history and so on,to summarize the clinical and laboratory characteristics of lung cancer patients with qi-deficiency and blood-stasis syndrome.Results1.Among 60 cases,male 34 cases,6 cases of women,The male to female ratio was 1.307:1,The median age was 61 years,with adenocarcinoma and squamous carcinoma as the main type.Ⅲ、Ⅳ stage are mainly clinical stages.2.Compared with control group,PT was significantly prolonged(P<0.05),FIB、D-dmier concentrations were increased significantly(P<0.05),the other indexes showed no obvious change(P>0.05)3.Compared with lung adenocarcinoma patients,the mean D-dimer and FIB were significantly higher in patients with squamous carcinoma group,with statistical significance(P<0.05)4.Compared with Ⅰ,Ⅱ stages,all of coagulation indicators were significantly higher in patients with Ⅲ、Ⅳ stages except to the PT.5.PT,FIB,D-dimer level in patients with cancer transferring were significantly higher than the no transfers.(P<0.05)6.Compared with non-smokers,smokers have higher PT,FIB,D-dimer level.The difference,however,was not statistically significant(P>0.05)The third part is experimental studyObjective In this part,we discussed the effects of DiterpenoidTanshinone(diter Tan)to improve microcirculation on orthotopically implanted tumors in nude mice,furtherly determine the role of PERK-EIF2α signaling pathway in Diterpenoid Tanshinones-induced lung cancer PC9 cell apoptosis to expound the mechanism of apoptosis for its clinical application.Methods1.In vivo experiments(1)Orthotopically implanted tumors in nude mice was modeled with PC9 cells(2)After the neck can touch subcutaneous tumors about 0.5×0.5cm,nude mices were randomly divided into 4 groups with 6 in each group:The groups are divided into model group,low、high and middle-dose group of DiterpenoidTanshinone.Chinese medical group was fed with different doses of DiterpenoidTanshinone,with 15、30、60mg/kg,once a day for 4 weeks,while model group was given normal saline by gavage,once a day for 4 weeks.(3)After administration,we anesthetized mice by isoflurane inhaled,using laser doppler MoorFLPI scanner to observe the blood flow changes in peripheral blood and tumors.After the scan,all of mices were killed and observed the sizes of tumor volume to observe the effect of DiterpenoidTanshinone on the growth of PC9 human lung caner subcutaneous transplanting tumor in nude mice.2.In vitro studies(1)Cells were randomly divided into 4 groups,The blank group,low-dose of DiterpenoidTanshinone(2.5ug/ml),middle-dose of DiterpenoidTanshinone(5ug/ml)and high-dose of DiterpenoidTanshinone(7.5ug/ml)(2)Effect of DiterpenoidTanshinones on the proliferation of PC9 cells were observed with CCK8 assay and Hoechst 33258;The apoptosis was detected by Annexin V-FITC/PI double staining method with flow cytometry in different concentration of 0,2.5,5,7.5ug/ml.(3)The expressions of PERK-EIF2α-related proteins and its downstream protein expression levels,such as PERK,EIF2α,CHOP,ATF4,Bax、Bcl-2,et al,were analyzed by Western blot.(4)The RNA interference segments targeting to PERK gene were successfully transfected into PC9 cells,and the expression of PERK genes was detected by Western blot,and selected out the best interference fragment.Since then,we also detected the apoptosis and the expressions of PERK-EIF2α-related proteins by Annexin V-FITC/PI double staining and Western blot separatelyResults1.In vivo experiments(1)Compared with the model group,the tumor body and peripheral blood flow increase are not obvious with low and middle-dose of DiterpenoidTanshinone,but high-doses with statistical significance(P<0.05).(2)Compared with model group,the Carcinomas were reduced obvious in all groups of DiterpenoidTanshinone,which was particularly obvious in the high dosage group(P<0.05)2.In vitro studies(1)Cell proliferations were inhibited and the apoptosis rate were increased significantly,with the increase of the Diterpenoid Tanshinones concentration(P<0.05)(2)As the time went on,the phosphorylation levels of PERK、EIF2α were increased significantly,especially in 8h(P<0.05),ATF decreased,and CHOP increased furtherly the interference of PERK gene can increases cell proliferation,inhibit apoptosis(P<0.05),and the phosphorylation level of PERK、EIF2α decreased obviously(P<0.05),CHOP decreased either,but ATF increasing.Conclusions:1.Lung cancer based on the treatment of poison and blood stasis has more in-depth theoretical basis and the exact clinical curative effect.Qi-deficiency and blood-stagnation caused the microenvironment of poison-blood stasis is the main pathogenesis of lung cancer.2.Qi deficiency and blood stasis syndrome pathogenesis is widespread in lung cancer,particularly in advanced non-small cell lung carcinoma.Coagulopathy has the high correlation with tumor stagings,pathologic type and cancer metastasis,which can be used an important factor that decide the prognosis of lung cancer.The level of FIB,D-dimer responsed the blood hypercoagulability in patients of qi-deficiency and bloodstasis with lung cancer,which can be expected to be a very practical clinical diagnosis of microcosmic index.3.Experiments studies in vivo and in vitro have shown that DiterpenoidTanshinone can reduce the size of our tumors,improve minicirculation,induce apoptosis,which proves the stasis poison pathogenesis is objective existence.The reason is connected with the activation of PERK/EIF2α signaling pathway. |
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