Clinical Significance Of ?-SMA On Microvessel And Lymphatic Vessel Density In Lung Adenocarcinoma

Objective : We aimed to determine whether cancer associated fibroblasts(CAFs)are associated with micro vessel density(MVD)and lymphatic vessel density(LVD)in lung adenocarcinoma(ADC)and correlate with prognosis.Methods : A total of 93 formalin-fixed and paraffin-embedded tumor samples from patients who underwent curative surgical resection from stage I-III lung adenocarcinoma at the Department of Thoracic Surgery in the Qingdao Municipal Hospital from 1 January,2010 to 31 June,2011 were selected.The 93 blocks available were cut into 5-um-thick sections and were stained with ?-SMA monoclonal antibody for histopathological evaluation of CAFs.Lymphatic vessels were assessed by D2-40 staining and micro vessels were stained with CD34.?-SMA positive stained tissues were graded as CAF-rich and CAF-poor groups according to CAF staining density.The ?-SMA positivity was calculated as percentage of ?-SMA positive cell numbers / total cell count.A value ?40%was considered high CAFs group,and < 40% was defined as CAF-poor group.vessel hot spot regions were identified under 40× magnifications and then 400× magnification was applied for counting micro vessels in 3 separate hot spots and the average was calculated and expressed as mean.the lumen in ribbon-shaped or lacuna shaped structures,formed by endothelial cells and positively stained by D2-40 are identified as lymphatic vessels.Hot spot regions were selected under 40× and 100× magnification.The density of lymph-vessels was evaluated under 400× magnification and the average numbers in 5selected hot spots were calculated and expressed as mean.The follow-up information was collected until death or disease progression.A routine protocol including detailed inquiry of medical history and radiological examination was proceeded over the course of the follow-up period.Statistical package SPSS version 20.0 was used for data analyses.The Pearson's Chi-square test and Logistic regression were used for exploring the correlation between clinical data and immunohistochemical results.Kaplan Meier method wasemployed to calculate the survival rate.Inter-group difference was preceeded by the Log-Rank test.The Cox proportional hazards model was used for multiple factors analysis to predict survival and a P-value < 0.05 was considered statistically significant.Results : ?-SMA positive staining was mainly detected in tumor stroma,indicating involvement of CAFs in tumor pathogenesis.Out of the 93 lung adenocarcinoma cases studied,72 were defined as high CAF group and 21 were low CAF expressing.CAF density,MVD and LVD are significantly correlating with age,tumor size,degree of differentiated,clinical stages and lymph node metastasis(P<0.05).Whereas were not influenced by parameters such as gender,tumor location and smoking history(P>0.05).MVD was significantly higher in CAF-rich group than in the CAF-poor group(P=0.02),LVD area was significantly higher in CAF-rich group than in the CAF-poor group(P<0.001),suggesting a close relationship between CAFs and lymphangiogenesis.3 years survival of CAFs-poor group was 64.5%,which is significantly higher than in CAF-rich group(41.9%).Kaplan Meier analyses showed that age,clinical stage,CAF density,MVD,LVD,tumor size,degree of differentiation and lymph node metastasis all had impact on the overall survival(P<0.05),while gender,smoking history and tumor location had little influence(P>0.05).Cox multi-factor analysis identified clinical stage as an independent prognostic factor in lung adenocarcinoma(P<0.05).Conclusion:CAF density,identified by ?-SMA staining,is associated with occurrence,development and metastasis of lung adenocarcinoma and influences disease outcome of the patient.