Quantitative and qualitative profiles of circulating monocytes may help identifying tuberculosis infection and disease stages

Abstract

Tuberculosis (TB) is one of the most important cause of morbidity and death among infectious diseases, and continuous efforts are needed to improve diagnostic tools and therapy. Previous published studies showed that the absolute cells number of monocytes or lymphocytes in peripheral blood or yet the ratio of monocytes to lymphocytes displayed the ability to predict the risk of active TB. In the present study we evaluated the ratio of monocytes to lymphocytes variation and we also analyzed the ex-vivo expression of CD64 on monocytes as tools to identify biomarkers for discriminating TB stages. Significant differences were found when the average ratio of monocytes to lymphocytes of active TB patients was compared with latent TB infection (LTBI) subjects, cured TB and healthy donors (HD). By the receiver operator characteristics (ROC) curve analysis the cut-off value of 0.285, allowed the discrimination of active TB from HD, with a sensitivity of 91.04% and a specificity of 93.55% (95% of confidence interval: 0.92-0.99). The ROC curve analysis comparing TB patients and LTBI groups, led to a sensitivity and the specificity of the assay of 85.07% and 85.71%, respectively (95% of confidence interval: 0.85 to 0.96). The upregulation of CD64 expression on circulating monocytes in active TB patients could represent an additional biomarker for diagnosis of active TB. In conclusion, we found that the ML ratio or monocyte absolute count or phenotypic measures show predictive value for active TB.

Fig 1. Ratio of monocytes to lymphocytes (ML ratio) of patients with active TB disease, LTBI subjects, cured TB patients and HD.

A. Each dot represented ML ratio of a studied individuals. A horizontal bar indicates the median of each group. Significance of differences between groups was compared using Kruskal-Wallis test. *p<0.00001. B. Monocyte absolute count of active TB patients at diagnosis, LTBI subjects, cured TB patients and HD. Each dot representes the value of an individual subject. Each horizontal bar represents the median of each group. p<0.00001 were considered statistically significant. C. Lymphocyte absolute count of active TB patients at diagnosis, LTBI subjects, cured TB patients and HD. Each dot represents the value of an individual subject. Each horizontal bar represents the median of each group. p<0.00001 were considered statistically significant.

Fig 2. Correlation between the ML ratio and absolute monocyte and lymphocyte counts.

A) The potential association between the ML ratio and absolute monocyte count, were analyzed by Spearman rank correlation test. Data shown are the values of the individual subjects. B) The potential association between the ML ratio and absolute lymphocyte count were analyzed by Spearman rank correlation test. Data shown are the values of the individual subjects. C) Receiver operating characteristic (ROC) curve for the ML ratio. The solid line shows the result for the value of ML ratio comparing active TB vs HD. D) The solid line shows the result for the value of ML ratio comparing active TB patients with LTBI subjects.

Fig 3. Receiver operating characteristic (ROC) curve for the ML ratio index.

The solid line shows the result for the ML ratio value comparing patients with active TB disease to cured TB patients.

Fig 4. Surface molecules expression on circulating monocytes of patients with active TB disease, LTBI subjects, cured TB patients and HD.

A) Cumulative data of the percentage expressed as median of surface expression and IQR. B) Geometric mean fluorescence intensity (geo-mean) of different surface molecules on monocytes and S.E. C) Representative FACS analysis of surface markers expression on circulating monocytes of one representative subject from each cohort group. D) Receiver operating characteristic (ROC) curve for the MFI value of CD64. The solid line shows the result for the value of comparing active TB vs LTBI.

Fig 5. Receiver operating characteristic (ROC) curves for the ML-CD64 index and the Monocyte-CD64 index.

A) The solid line shows the result for the ML-CD64 index value comparing patients with active TB disease to HD. B) The solid line shows the result for the ML-CD64 index value comparing patients with active TB disease to LTBI subjects. C) The solid line shows the result for the monocyte-CD64 index value comparing patients with active TB disease to HD. D) The solid line shows the result for themonocyte-CD64 index value comparing patients with active TB disease to LTBI subjects.

Fig 6. Receiver operating characteristic (ROC) curve for the monocyte-CD64 index.

The solid line shows the result for the monocyte-CD64 value comparing patients with active TB disease to cured TB patients.