Validation Of Bevilacqua Breast Cancer Nomogram To Predict Lymphedema And The Expression Of Lipid Metabolism Gene In Lymphedema

Background:Breast cancer is the highest incidence and mortality of female worldwide,and the incidence of breast cancer is still on the rise in most countries,including China.Surgery is very important in the treatment of breast cancer.Breast surgery includes surgery to breast surgery and axillary.Upper arm lymphedema is a common complication after axillary lymph node dissection with regional enlargement of one or both arms in breast cancer patient.Once upper arm lymphedema occurs,there is no effective way to cure it.Therefore,establish a quantitative nomogram to predict the risk using the risk factors of upper arm lymphedema will help us realize the early prevention of lymphedema.The research team of Bevilacqua have established nomogram to predict the risk,but it yet not was validated in Chinese breast cancer patients in present.The study of lymphedema is mostly focused on the clinical risk factors,while the molecular mechanism is rarely studied.Some scholars believe the upper arm lymphedema was due to abnormal deposition of fat,chronic inflammation and fibrosis,which led to the obstruction of lymphatic reflux.The molecular markers that regulate adipocyte metabolism were significantly increased in the mouse model of lymphedema.As the regulatory genes of lipid metabolism,FADS1,FASN,SCD1 and COX-2 are widely involved in energy conversion and lipid synthesis,but whether they play an important role in lymphedema were unknown.Part 1: Validation of Bevilacqua breast cancer nomogram to predict lymphedemaObjective:To validate the Bevilacqua nomogram to predict the risk of lymphedema in Guangdong breast cancer patients.Methods:A total of 409 patients who underwent breast surgery and axillary lymph node dissection(Level I and II)were included.The median follow-up was 68 months(range: 60-83 months).Upper arm lymphedema was confirmed if a difference of volume between two arms were greater than 200 m L.The baseline data were compared using t test,chi square test,Mann-Whitney U test and Kruskal-Wallis H test when appropriate.Cox regression analysis was used to identify the risk factors for upper arm lymphedema in Guangdong breast cancer population.Kaplan-Meier survival curve analysis of the relationship between baseline data and prognosis of patients included.Kaplan-Meier analysis the 5-year cumulative incidence of upper arm lymphedema.ROC curve was used to evaluate the discriminative ability of the model.The calibration plots was used to calculated and plotted average prediction probability.Hosmere-Lemeshow test was used to evaluate the fitting degree between the predictive value and the actual value,P > 0.05 indicates the model works well.STATA(SE13.0,Stata Corp,LP,College,Station,TX)was applied for all statistical analyses.P values <0.05 were considered significant.Results:1.With a median follow-up of 68 months(range: 60-83 months),The patients included were relatively young,with a mean age of 47.2 years old.2.Multivariate COX proportional hazard regression model analysis showed that tumor size(p<0.0001,HR:6.681,95%CI:2.388-18.691),the expression of Ki-67(p<0.0001,HR:0.028,95%CI:0.013-0.063)and TNM stage(p=0.001,HR:3.791,95%CI:1.706-8.421)were the independent prognostic factors of relapse free survival(RFS)of breast cancer patients.The tumor size(p=0.001,HR:0.035,95%CI:0.005-0.259),lymph node staging(p<0.0001,HR:5.917,95%CI:2.694-12.993)and the expression of Ki-67(p<0.0001,HR:30.319,95%CI:,13.250-69.374)were the independent prognostic factors of overall survival(OS).3.The 5-year cumulative incidence of upper arm lymphedema was 22.3% in 409 samples included.The annual incidence of upper arm lymphedema began to increase significantly after the 1 year follow-up and increased almost linearly.4.Higher body mass index(HR=1.06,95%CI:1.00-1.13),neoadjuvant chemotherapy(HR= 3.76,95% CI:2.29-6.20),larger extend of axillary surgery(I/ II / III / vs I II: HR=2.39,95% CI:1.30-4.37)and radiotherapy(HR=4.90,95%CI:1.90-12.5)were independently associated with upper arm lymphedema in our populations.5.In ROC analysis,the AUC value was 0.706(95% CI: 0.648-0.752),Model 2establised by Bevilacqua had significant discriminative efficacy in our population.6.The cumulative risks of upper arm lymphedema in the high-risk subgroup were significantly higher than those in the other subgroups(P < 0.01).7.The calibration plots suggested that the nomogram is well calibrated,with the predicted risks of LE not significantly differed from the actual risks of LE(Hosmere-Lemeshow test,P=0.0634),upper arm lymphedema risk prediction and the actual risk did not differ significantly(Hosmer-Lemeshow test,P=0.0634),indicating that the nomogram 2 to predict the risk of lymphedema established by Bevilacqua perform well.Conclusion:1.This study confirmed that the Bevilacqua nomogram 2 to predict the risk of lymphedema has a good discriminative capacity in Guangdong breast cancer population.2.The Bevilacqua nomogram 2 to predict the risk of lymphedema was accurate and effective in Guangdong breast cancer population.3.Further studies should be performed to evaluate the impact of other factors(lifestyle,behavior,etc.)on the predictive model.Part 2: The expression of lipid metabolism regulation gene in lymphedemaObjective:To investigate the changes in lipid metabolism and the expression of lipid metabolism gene FADS1,FASN,SCD1,COX-2 and its clinical significance in the tissue specimens of upper limb lymphedema after axillary lymph node dissection.Methods:60 patients as experimental group with upper arm lymphedema within 3 years after surgery from 2010—2016 in our Hospital were reviewed,60 patients without upper arm lymphedema within 3 years were paired according to the clinical characteristics as control group.The morphological changes of adipocyte in tissue samples of edema group and non edema group were observed by oil red O staining,and the staining of lipid metabolic genes in two groups was observed by immunohistochemistry.The expression of lipid metabolism genes in the edema group and non edema group were detected by PCR(q RT-PCR)method to understand the correlation between gene and upper limb lymphedema.Results:Through the oil red O staining,it was found that in non-lymphedema group,the red dye area was less and the red dye was relatively shallow.While in lymphedema group,the red dye range was wider and the color was deeper.Also,in the adjacent tumor tissue,a large amount of fatty cell was gathered.Compared with the non-lymphedema group,the staining intensity of gene FASN,SCD1 and COX-2 in the lymphedema group was higher(P<0.05),while the expression of FADS1 in the two groups was not significant(P=0.453).By further examination of the RT-q PCR test,it showed that FASN,SCD1 and COX2 genes were higher expressed in the tissue of lymphedema group.Conclusion:The upper limb lymphedema after axillary lymph node dissection may be associated with abnormal fat metabolism in the patient.The high expression of lipid metabolism gene FASN,SCD1 and COX-2 in the carcinoma tissue and adjacent tumor tissue in the edema group may be closely related to the occurrence of upper limb lymphedema,which provides a way of studying the follow mechanism.