Experimental And Clinical Study On Lung Cancer Angiogenesis And Signal Transduction Pathways With Six Spirits Pill

Objective:to observe1.the effect of Six Spirits Pill medicated serum roling in the cells for theproliferation,apoptosis,cells form and growth condition of the lung cancer cell A549,and for the protein expression changes of VEGF,K-RAS,ERK1,ERK2,p-ERK andVEGFmRNA,the correlation between K-RAS, ERK1,ERK2,p-ERK protein expressionand of VEGF expression.2.the clinical effects and the changes of serum CEA,CYFRA21-1and VEGF for the lung cancer patients without any treatment IIIB-IV stage or IIIA stagepatients who can't be operated on by Six Spirits pill combination chemotherapy treatment；The paper discusses whether Six Spirits Pill antiangiogenic is achieved by activating theRAS→Raf→MEK→MAPK signal transduction pathways and Whether serum VEGF canbe used as one of clinical observation curative effect.Methods:1. Experimental studyCulture the lung cancer A549cells in vitro utilizing the method of serumpharmacolog,incubate different concentration of serum which included Six Spirits Pilltogether with A549cells and observe them in group.Detect the effect of the serum with pillto proliferation by the use of the MTT method,observe cells'form with optical microscopeand growth state with inverted phase contrast microscope,detect cell apoptosis with theFlow cytometric,determine the protein expression changes of the VEGF,K-RAS,ERK1,ERK2,p-ERK in immunocytochemical method,test VEGFmRNA by means of retrovirus--polymerase chain reaction method. 2.Clinical studyObserve the clinical effects for the non-small cell lung cancer patient and the effectfor the expression changes of serum CEA,CYFRA21-1,VEGF from the six Spirits pillcombination chemotherapy treatment and chemotherapy treatment group.All case were thein-patients who were from shandong university of traditional Chinese medicine hospital,from May2010to December2011tumor. According to the criteria, there are60cases inall,some were divided into observation group of30cases and the other control group of30cases randomly.The control group uses the TP chemotherapy,the observation group wereadded Six Spirits Pill during chemotherapy.Before the treatment and after the two cycletreatment, Collect5ml peripheral blood of all patients,observe the VEGF level by the useof enzyme-linked immunosorbent assay (ELISA) test method,detect CYFRA21-1,CEAexpression level by radiation immune analysis method,and observe the clinical curativeeffect before and after treatment.Results:1.Experimental study(1)Utilizing medicated serum of Six Spirits Pill to A549cells has obvious cytotoxiceffects,and with the increase of medicated serum concentrations the OD value graduallyreduced,there are obviously differences.(2)The medicated serum group all can destroy thenormal form of A549cells,hinder the growth of proliferation,and have positivecorrelation with serum concentration. Six Spirits Pill and DDP have the synergisticeffect.(3)Early apoptosis rate of medicated serum groups was higher than the controlgroup；As the drug concentration rises,early apoptosis rate obviously increased；The jointgroup's was higher than medicated serum group's and DDP single drug group's.(4)Theexpression amount of VEGF,ERK1,ERK2,p-ERK,K-RAS and the relative amountVEGFmRNA of medicated serum group were significantly lower than the control group,there are significantly differences；Drug concentration was negatively.(5)The statisticalresults show that the Six Spirits Pill medicated serum for A549cells reducing VEGFexpression and it's roling in ERK1,ERK-2,p-ERK,K-ras expression has significantcorrelation (r=0.887,0.835,0.952,0.866).2.Clinical study(1) The chemotherapy accomplishment circumstances of the two groups:the2cycle chemotherapy completion rate of the observation group is93.3%(28/30),higher than86.7%of the control group(26/30)；(2)the recent clinical curative effect:through thechi-square test,there are no significant difference between the effective rate and stabilityrate of two groups.(3)the comparison of serum VEGF,CEA,CYFRA21-1level of twogroups before and after treatment:after treating, the expression levels of VEGF,CEA,CYFRA21-1both descend obviously,compared with those before treatment. Comparedwith the control group,the serum VEGF,CEA expression levels of the observation groupare lower,their ranges decrease more,and they have statistically significant differences.Atthe same time,the CYFRA21-1expression level of observation group is more low too,butthe comparison of two groups is no statistical difference.(4)The correlation analysisbetween serum CEA,CYFRA21-1level and VEGF of two groups:The result is obviouslypositive correlation(r=0.399,0.275,P<0.01).ConclusionSix Spirits Pill plays the role of anti-angiogenesis through inhibition ofRas→Raf→MEK→MAPK signal transduction pathway to reduce lung cancer A549cellexpression of VEGF protein,to inhibit lung cancer A549cell growth and promoteapoptosis,to reduce the serum VEGF levels of lung cancer patients,at the same time,theeffect of Six Spirits Pill combination chemotherapy is better,the two have synergisticeffect.Clinical joint detection of lung cancer patients's serum VEGF,CEA,CYFRA21-1changes plays the role in the observing efficacy and monitoring the patient condition.