CXCL9, CXCL11 And Tuberculous Pleural Effusion: Diagnostic Value, Origin, And Biological Function

PurposeThis study aims to evaluate the diagnostic accuracy of pleural fluid(PF)levels of C-X-C motif chemokine ligand 9(CXCL9)and C-X-C motif chemokine ligand 11(CXCL11)for tuberculous pleural effusion(TPE).In addition,we aimed to identify the cell origins of the CXCL9 and CXCL11 in the pleural fluid of TPE patients and their chemotaxis towards CD4~+T cells.We also investigated the underlying mechanism of helper T cell(Th)cells’entry into the thoracic cavity.Methods(1)Clinical study:The PF concentrations of CXCL9 and CXCL11 were compared between 20 TPE patients and 133 non-TPE patients(Hohhot cohort)to assess their diagnostic value.Receiver operating characteristic(ROC)curves and decision curve analysis(DCA)were used to evaluate their diagnostic accuracy.The diagnostic value of CXCL9 and CXCL11was further validated in another independent cohort(Changshu cohort,n=58).We also explored the effect of age on the diagnostic value of PF CXCL9 and CXCL11 in the Hohhot cohort.(2)Basic research:Flow cytometry was used to detect and compare the cell origins of the PF chemokine CXCL9 in TPE patients and non-TPE patients.The source was then further verified by in vitro experiments using Bacillus Calmette-Guérin(BCG)to stimulate CD14~+mononuclear-derived macrophages(M0),and quantitative real-time PCR(q RT-PCR)and Enzyme linked immunosorbent assay(ELISA)to detect CXCL9 and CXCL11,respectively.The gene expression of CXCL9 and CXCL11 in stimulated cells and the concentrations of CXCL9 and CXCL11 in the cell supernatant were determined by q RT-PCR and ELISA,respectively.Next,ELISA was used to determine the concentrations of CXCL9 and CXCL11in the PF and serum to clarify the differential expression of CXCL9 and CXCL11.In vitro chemotaxis assays were performed to further investigate the chemotactic properties of the chemokines CXCL9 and CXCL11 on CD4~+T cells.Results(1)Clinical study:The area under the ROC curves(AUCs)of PF CXCL9 and CXCL11in the Hohhot cohort was 0.70(95%CI:0.55–0.85)and 0.68(95%CI:0.52–0.84),respectively.In the Changshu cohort,the AUCs of PF CXCL9 and CXCL11 were 0.96(95%CI:0.92–1.00)and 0.97(95%CI:0.94–1.00),respectively.The decision curves for PF CXCL9 and CXCL11 were also significantly different in the Hohhot and Changshu cohorts.In the Hohhot cohort,the AUCs of PF CXCL9 and CXCL11 decreased with the advancement of age.(2)Basic research:Flow cytometry revealed that PF CXCL9 of TPE patients was predominantly derived from CD14~+monocytes(4.92%).The m RNA expression of CXCL9and CXCL11(both p<0.001)and their concentrations in the supernatant(p<0.0001,p<0.001)were significantly increased after BCG stimulation of M0 cells.In addition,the concentrations of both chemokines CXCL9 and CXCL11 were significantly higher in TPE than in serum.The results of the in vitro chemotaxis assay showed that TPE and the BCG-stimulated M0 cell culture supernatant exerted a chemotactic effect on peripheral CD4~+T cells through potential chemotactic activity,which was significantly inhibited when anti-CXCL9 or/and anti-CXCL11-monoclonal-neutralizing-antibodies were added.The inhibition of the chemotactic process was even more pronounced when both neutralizing antibodies were added together.ConclusionsOur study indicates that PF levels of CXCL9 and CXCL11 have diagnostic values for TPE.However,the diagnostic performance of PF CXCL9 and CXCL11 may be compromised in elderly patients.Therefore,the age of the patient should be taken into consideration when interpreting the results.In addition,the chemokines CXCL9 and CXCL11 in the PF of TPE patients are mainly derived from CD14~+monocytes.Chemokines CXCL9 and CXCL11 may be able to induce the migration of peripheral blood Th cells to TPE patients’chest cavities.