Increase in the percentage of SLAMF7high CD16− monocytes in PB of MPN patients with MF who harbored JAK2V617F and the correlations between SLAMF7high CD16− monocytes and JAK2 allele burden in PB and fibrocytes. (A) Comparison of SLAMF7highCD16− monocytes in PB among HCs and MPN patients with or without MF. The median percentage of SLAMF7highCD16− monocytes in PB of HCs (n = 21), MPN patients without MF (n = 21), and MPN patients with MF (n = 37) were 0.94%, 3.23%, and 19.60%, respectively. The SLAMF7highCD16− monocyte percentage of MPN patients with MF was significantly increased compared with HCs and MPN patients without MF. The results are reported as medians. Statistical significance was calculated using 1-way ANOVA with post hoc Steel-Dwass test. (B) Subgroup analyses by genetic mutation (JAK2V617F, CALR, and triple-negative mutations). In this analysis, we excluded the MPL group because it contained only 1 patient and could not be subjected to statistical analysis. SLAMF7highCD16− monocytes were significantly increased in the MPN patients who harbored a JAK2V617F mutation (n = 28) compared with the MPN patients who harbored a CALR mutation (n = 9) or the MPN patients with triple-negative mutations (n = 20) (median: JAK2V617F 28.75% vs CALR 3.26% vs triple-negative mutations 1.73%; P < .01). The results are reported as medians. Statistical significance was calculated using 1-way ANOVA with post hoc Steel-Dwass test, (C) Subgroup analysis of SLAMF7highCD16− monocytes in PB by the presence or absence of JAK2V617F mutation. MPN patients with MF who harbored JAK2V617F (n = 19) had a significantly higher SLAMF7highCD16− monocyte percentage than those without MF (n = 9) (median: 43.70% vs 7.74%; P < .01). In a similar manner, MPN patients with MF who did not harbor JAK2V617F (n = 18) had a significantly higher SLAMF7highCD16− monocyte percentage than those without MF (n = 12) (median: 1.24% vs 4.32%; P < .05). The contrast between patients with and without MF was clearer in MPN patients who harbored JAK2V617F than in those who did not harbor JAK2V617F. Results are reported as medians. Statistical significance was calculated using the Mann-Whitney U test. (D) ROC curve showing the cutoff value of the percentage of SLAMF7highCD16− monocytes in PB for separating MPN patients with MF who harbored a JAK2V617F mutation from those without MF. The AUC was 0.901, and by setting the cutoff of the percentage of SLAMF7highCD16− monocytes to >25%, the sensitivity and specificity for classifying MF onset were 79.0% and 77.8%, respectively. (E) Correlation between SLAMF7highCD16− monocytes in PB and JAK2 allele burden of PB (n = 28). There was a positive correlation between the SLAMF7highCD16− monocyte percentage in PB and the JAK2V617F allele burden of PB (r = 0.531; R2 = 0.282; P = .0037). Pearson’s correlation coefficient was calculated, and the statistical significance was calculated using linear regression analysis. (F) Correlation between SLAMF7highCD16− monocytes in PB and the JAK2 allele burden of cultured fibrocytes (n = 28). There was a positive correlation between SLAMF7highCD16− monocyte percentage in PB and JAK2V617F allele burden of fibrocytes (r = 0.631; R2 = 0.398; P = .0003). Pearson’s correlation coefficient was calculated, and the statistical significance was calculated using linear regression analysis. (G) The JAK2V617F allele burden of isolated SLAMF7lowCD16− monocytes and isolated SLAMF7highCD16− monocytes from 7 MPN patients who harbored JAK2V617F. The JAK2V617F allele burden of isolated SLAMF7highCD16− monocytes was significantly higher than the JAK2V617F allele burden of isolated SLAMF7lowCD16− monocytes (paired Student t test P < .01). (H) Culture assay of isolated SLAMF7lowCD16− monocytes and isolated SLAMF7highCD16− monocytes collected from 2 MPN patients who harbored a JAK2V617F mutation. The frequency of fibrocytes derived from SLAMF7highCD16− monocytes was significantly increased compared with that from SLAMF7lowCD16− monocytes. Results are reported as the mean ± standard deviation of data obtained from 3 individual experiments. Statistical significance was calculated by unpaired Student t test. *P < .05; **P < .01.