H-ferritin and proinflammatory cytokines are increased in the bone marrow of patients affected by macrophage activation syndrome

Abstract

Macrophage activation syndrome (MAS) is hyperinflammatory life-threatening syndrome, associated typically with high levels of serum ferritin. This is an iron storage protein including heavy (H) and light (L) subunits, categorized on their molecular weight. The H-/L subunits ratio may be different in tissues, depending on the specific tissue and pathophysiological status. In this study, we analysed the bone marrow (BM) biopsies of adult MAS patients to assess the presence of: (i) H-ferritin and L-ferritin; (ii) CD68⁺ /H-ferritin⁺ and CD68⁺ /L-ferritin⁺ ; and (iii) interleukin (IL)-1β, tumour necrosis factor (TNF) and interferon (IFN)-γ. We also explored possible correlations of these results with clinical data. H-ferritin, IL-1β, TNF and IFN-γ were increased significantly in MAS. Furthermore, an increased number of CD68⁺ /H-ferritin⁺ cells and an infiltrate of cells co-expressing H-ferritin and IL-12, suggesting an infiltrate of M1 macrophages, were observed. H-ferritin levels and CD68⁺ /H-ferritin⁺ cells were correlated with haematological involvement of the disease, serum ferritin and C-reactive protein. L-ferritin and CD68⁺ /L-ferritin⁺ cells did not correlate with these parameters. In conclusion, during MAS, H-ferritin, CD68⁺ /H-ferritin⁺ cells and proinflammatory cytokines were increased significantly in the BM inflammatory infiltrate, pointing out a possible vicious pathogenic loop. To date, H-ferritin and CD68⁺ /H-ferritin⁺ were associated significantly with haematological involvement of the disease, suggesting biomarkers assessing severity of clinical picture.

Keywords: cytokine; ferritin; hyperferritinaemic syndrome; macrophage; macrophage activation syndrome.

Figure 1

Expression of H‐ferritin in bone marrow samples of macrophage activation syndrome (MAS) patients. (a) high H‐ferritin (red) and low tissue expression of L‐ferritin (green) may be observed, respectively; (b) low H‐ferritin expression may be reported in healthy controls (HCs); (c) H‐ferritin was increased significantly when compared with L‐ferritin increased and HCs, analysing immunostaining optical density (***P < 0·0001).

Figure 2

Correlation of H‐ferritin with clinical features. (a,b) Increased values of H‐ferritin were correlated with decreased white blood cells (WBC) and platelet (PLT) counts, respectively; (c) significantly and (d) increased values of H‐ferritin tissue were correlated significantly with increased values of serum ferritin and C‐reactive protein (CRP), respectively.

Figure 3

Expression interleukin (IL)‐1β in bone marrow of macrophage activation syndrome (MAS) patients and its correlation with H‐ferritin. (a) Increased IL‐1β (green) levels and co‐localization (arrows) between H‐ferritin (red), and this cytokine may be observed in the inflammatory bone marrow infiltrate; (b) reduced IL‐1β expression in healthy controls (HCs); (c) in MAS patients, IL‐1β was increased significantly when compared with HCs (***P < 0·001); (d) IL‐1β was correlated significantly with H‐ferritin.

Figure 4

Increased expression of tumour necrosis factor (TNF) and interferon (IFN)‐γ in bone marrow of macrophage activation syndrome (MAS) patients. (a) Increased IFN‐γ (green) levels and H‐ferritin (red) may be described in MAS; (b) reduced IFN‐γ levels in healthy controls (HCs); (c) IFN‐γ was increased significantly when MAS patients were compared with HCs (***P < 0·001); (d) increased TNF (green) levels and H‐ferritin (red) may be observed in MAS; (e) reduced TNF levels in HCs; (f) TNF was increased significantly when MAS patients were compared with HCs (***P < 0·001).

Figure 5

CD68 macrophages expressing H‐ferritin in bone marrow infiltrate of macrophage activation syndrome (MAS) patients. (a) CD68⁺/H‐ferritin⁺ cells (arrows) may be described in the bone marrow (BM) of MAS patients; (b) reduced CD68⁺/H‐ferritin⁺ cells in BM of healthy controls (HCs); (c) CD68⁺/H‐ferritin⁺ cells was increased significantly in MAS when compared with HCs (***P < 0·001); (d) co‐localization of H‐ferritin (red) and interleukin (IL)‐12 (green) may be observed, suggesting M1 macrophages; (e) CD163⁺ cells (green) showed weaker co‐localization with H‐ferritin (red).

Figure 6

Correlation between CD68⁺/H‐ferritin⁺ cells, haematological involvement and inflammatory markers. (a,b) Increased CD68⁺/H‐ferritin⁺ was correlated with decreased white blood cell (WBC) and decreased platelet (PLT) counts, respectively; (c,d) CD68⁺/H‐ferritin⁺ was correlated significantly with serum ferritin and C‐reactive protein (CRP).