The Value Of MUC16/CA125 And Targeted Radionuclide Probes In The Diagnosis And Risk Prediction Of Interstitial Lung Diseas

Part Ⅰ:The diagnosis and clinical prognosis of CA125 in interstitial lung diseaseBackground:Interstitial lung disease(ILD)is characterized by different degrees of inflammation and fibrosis in the pulmonary.The interaction of multiple immune cells and stromal cells is involved in the pathological and physiological mechanisms of the disease.Carbohydrate antigen 125(CA125)is formed by the extracellular portion of mucin 16(MUC16),which is cleaved by proteases and released from the cell membrane into the serum or extracellular matrix.Studies have shown that high levels of serum CA125 expression are associated with increased mortality in patients with IPF.The aim of this study was to analyze the diagnostic role,correlation with clinical features and the impact on disease prognosis of serum CA125 levels among different types of interstitial lung disease,including idiopathic pulmonary fibrosis and other types of interstitial lung disease(connective tissue disease-related interstitial lung disease(CTD-ILD)and sarcoidosis).Methods:Ninety-three patients with IPF,108 patients with sarcoidosis,144 patients with connective tissue disease(CTD)and 112 healthy controls were enrolled in this study.Serum CA125 levels were detected,clinical information and follow-up information was collected.The disease progression prediction model(Nomogram)was constructed for patients with IPF.The results of tumor markers in patients with CTD were collected,and the imaging HRCT manifestations were evaluated to determine whether ILD occurred in the patients,and the HRCT score of pulmonary fibrosis was performed to evaluate the ability of serum CA125 to differentiate CTD-ILD and CTD without ILD.The correlation of CA125 with clinical characteristics,pulmonary function,imaging Scadding stage and prognosis of sarcoidosis patients were analyzed.Results:Serum CA125 was significantly higher in patients with IPF than in healthy controls(median 49.82 vs 16.97 U/mL,P<0.0001),and the area under the ROC curve(AUC)was 90.03%(SE 0.02;95%CI 0.86-0.94),and the optimal cut-off value was 37.15 U/mL(sensitivity 67.7%,specificity 99.1%).CA125 levels were significantly higher in the group of IPF patients with progressive events than in the group without events(71.48±12.63 vs 64.47±14.21,P=0.014).The Event-free survival(EFS)of IPF patients was analyzed by univariate and multivariate Cox proportional hazards models.The results showed that the forced vital capacity(FVC)as a percentage of predicted(FVC%predicted)(P=0.026,hazard ratio(HR)0.98,95%confidence interval(CI)0.95-0.99)and CA125 level(p<0.001,HR 1.028,95%CI 0.014-1.043)were independent predictors of EFS.There was a significant positive correlation between the expression level of serum CA125 and the HRCT score of IPF(r=0.2544,P=0.0138).In this study,a disease progression prediction model(SCAPH score)for IPF patients was constructed,and the C-index index of the calculation model reached 0.71,indicating that it has good partition degree and accuracy,and can effectively predict whether IPF patients have progressive events.In other ILD patients,serum CA125 levels were significantly higher in CTD-ILD patients(P=0.001)than in CTD patients without ILD(AUC=0.67,optimal cut-off value of 26.50 U/mL).Spearman correlation test showed that CA125 level was significantly correlated with HRCT pulmonary fibrosis score in CTD-ILD patients(R=0.468,P<0.0001).In the sarcoidosis group,the expression level of serum CA125 was significantly higher than that in the healthy control group(P<0.001),the area under the ROC curve was 0.9833,and the optimal cut-off value was 32.33 U/mL.The expression level of CA125 was significantly correlated with the Scadding stage of sarcoidosis(P<0.05).Conclusion:This study found that CA125 is a potential diagnostic marker of IPF,and CA125 expression in IPF patients is positively correlated with the degree of pulmonary fibrosis.High serum CA125 levels can be used as an independent predictor of progression prognosis in patients with IPF.In addition,in other interstitial lung diseases,serum CA125 level is a potential diagnostic marker to help determine whether CTD patients occurred with ILD,and serum CA125 level is related to the severity of pulmonary fibrosis imaging in CTD-ILD patients.Serum CA125 levels were also elevated in patients with sarcoidosis and correlated with lung involvement.Part Ⅱ The mechanism of MUC16 in the pathogenesis of IPFBackground:According to the results of previous studies,the serum level of CA125 is significantly increased in patients with IPF,and CA125 is the part of MUC16 extracellular domain that enters the serum or extracellular matrix after enzyme digestion.However,it is still unclear whether MUC16 is overexpressed in the lung tissue of patients with IPF and whether it is involved in the pathogenesis of IPF pulmonary fibrosisMethods:Firstly,18 C57BL/6 male mice aged 6-8 weeks were randomly divided into control group and pulmonary fibrosis group.Pulmonary fibrosis was induced by intratracheal injection of bleomycin in the experimental group.The lung tissues of the mice were collected 2 and 3 weeks after the model was established.Immunohistochemical H&E staining,Masson staining and MUC16 expression in lung tissue were performed.The expressions of MUC16 and phospho-Smad3(pSmad3)in A549 cells treated with transforming growth factor-β1(Transforming growth factor β1,TGFβ1)were observed by immunofluorescence.In addition,A549 cell line was constructed by small interfering RNA(siRNA)silencing MUC16,and the expression of downstream molecules of EMT induced by TGFβ1 was detected by RT-PCR.The lung tissues of IPF patients(n=6)and healthy controls(n=6)were collected,and the degree of pulmonary fibrosis was observed by immunohistochemical H&E staining and Masson staining,and the expressions of MUC16,Galectin-3,α-SMA and Collagen 1 in human lung tissues were determined.Results:In pulmonary fibrosis model mice,compared with the control group,MUC16 was overexpressed on the surface of bronchoalveolar epithelial cells by H&E staining,Masson staining and MUC16 immunohistochemical method.The expression of MUC16 and pSmad3 on the surface of A549 cells was increased after stimulation with TGF-β1 observed by immunofluorescence.MUC16/pSmad3 molecules were colocalized in A549 cells stimulated by TGFβ1,and the expression of MUC16/pSmad3 was higher than that in the control group.After Transfection of siRNA-MUC16,the expression of Col-1 and zinc finger family transcriptional repressor 1(SNAI1)in A549 cell line stimulated by TGFβ1 was significantly decreased compared with siRNA(-)group,suggesting that MUC16 played a role in promoting EMT mechanism induced by TGFβ1/Smad3 pathway in pulmonary fibrosis alveolar epithelial cells.The results of immunohistochemistry showed that the expression level of MUC16 was significantly increased in the honeycomb area and the mucus area of IPF lung tissue.Conclusion:The expression of MUC16 was increased in the injured bronchoalveolar epithelial cells of patients with IPF.The results of animal experiments showed that the expression of MCU16 in bleomycin-induced pulmonary fibrosis mice was higher than that in the control group,and it was mainly distributed on the surface of bronchial epithelial cells in the pulmonary fibrosis area.MUC16 can mediate the EMT mechanism of TGFβ1/Smad3 pathway in alveolar epithelial cells and promote the occurrence of pulmonary fibrosis.Part Ⅲ Pilot Study of 99mTc-FAPI SPECT/CT Imaging Probe in the Diagnosis and the Evaluation of Idiopathic Pulmonary FibrosisBackground:Previous studies have shown that MUC16 can mediate TGFβ1induced fibroblast activation,leading to fibrosis progression.Fibroblast activation associated protein(FAP),as a specific protein secreted by activated pulmonary fibroblasts,has suggestive value for the active status of pulmonary fibrosis.In this study,99mTc-FAPI radionuclide was used to target FAP in patients with IPF,and to evaluate the imaging characteristics of 99mTc-FAPI SPECT/CT targeted radionuclide probe in patients with IPF and its correlation with clinical characteristics.Methods:The 99mTc-FAPI targeted radionuclide probe was synthesized by the Department of Nuclear Medicine of Peking Union Medical College Hospital,and 11 IPF patients recruited in the respiratory outpatient department of our hospital from November 2021 to January 2022 were included in the study.After enrollment,they were injected with 99mTc-FAPI in the Department of Nuclear Medicine of our hospital,and the imaging chest HRCT examination and pulmonary function test were completed at the baseline level.The standardized uptake value(SUV)was calculated.The SUVmax and lesion to background ratio(LBR)of the affected lung parenchyma with positive FAP in 5 lung planes were recorded.For each level of SUVmax and LBR,total SUVmax and total LBR were used to score the degree of pulmonary fibrosis on 5 slices of chest HRCT,and the weighted total HRCT score of pulmonary fibrosis was included.The correlation of radionuclide imaging score with pulmonary function and imaging parameters of pulmonary fibrosis was analyzed.Results:All patients with IPF showed increased uptake of 99mTc-FAPI,which was mainly distributed in the peripheral subpleural area and the basal part of both lungs,and the corresponding areas also showed pulmonary fibrosis on chest HRCT.HRCT score was significantly associated with total SUVmax or LBR(P<0.05).With the increase of HRCT staging score,the total SUVmax and LBR increased significantly(P<0.05),except that the pulmonary fibrosis stage was stage 4.Total SUVmax and LBR had no significant difference with lung function parameters CPI,DLCO%predicted,FEV1%predicted and FEV1/FVC(P>0.05).Conclusion:Targeted molecular nuclide imaging is an effective imaging technique for pulmonary fibrosis.The imaging effects of IPF patients with different degrees of pulmonary fibrosis are different.99mTc-FAPI SPECT/CT can achieve the accurate diagnosis of IPF patients.The uptake of radionuclides in the areas of pulmonary fibrosis shown by HRCT is higher,which has a certain correlation with the degree of pulmonary fibrosis.This study shows that 99mTc-FAPI SPECT/CT detection technology can provide new ideas for clinical treatment and prognosis evaluation of IPF patients.