Bone marrow-derived monocyte infusion improves hepatic fibrosis by decreasing osteopontin, TGF-β1, IL-13 and oxidative stress

Figure 1 Representative FACS histograms of bone marrow mononuclear cells and CD11b⁺ monocytes isolated by immunomagnetic separation. BMMCs: Bone marrow mononuclear cells.

Figure 2 Schematic flowchart of experimental design. A: Male C57BL/6 mice underwent chronic administration of CCl₄ and EtOH solutions for 6 mo; B: Bone marrow mononuclear cells were harvested from C57BL/6 donor mice for CD11b⁺ monocyte isolation using immunomagnetic separation; C: Chronically liver-damaged mice underwent cell therapy; D: After 2 mo, effects of the treatment were evaluated using morphometric, biochemical, immunohistochemistry and sandwich ELISA analysis.

Figure 3 Photomicrographs of histological liver sections stained with picro-Sirius red. A-D: The figure shows hepatic collagens in (A) control mice, mice after CCl₄ administration and treatment with (B) saline, (C) BMMCs and (D) BMMC-derived monocytes (picro-Sirius red; magnification × 100); E: Morphometric evaluation of picro-Sirius Red-stained sections; F: Hydroxyproline in liver fragments of mice undergoing cell transplantation; G: Kupffer cell count in hematoxylin-eosin-stained histological liver sections from mice who underwent CD11b⁺CD14⁺ monocyte therapy and BMMC-treated mice. ^aP < 0.05; ^bP < 0.01; ^cP < 0.001. BMMCs: Bone marrow mononuclear cells.

Figure 4 Immunohistochemistry for detection of α-SMA⁺ hepatic stellate cells in histological sections. A-D: Control (A), saline-treated (B), BMMCs-treated (C) and CD11b⁺CD14⁺ monocyte-treated (D) groups of mice (magnification × 200); E: Measurement of α-SMA⁺ hepatic stellate cells at 2 mo after treatment with CD11b⁺CD14⁺ monocytes and BMMCs. ^aP < 0.05; ^bP < 0.01. BMMCs: Bone marrow mononuclear cells.

Figure 5 Immunohistochemistry for detection of osteopontin in histological sections. A-D: Control (A), saline-treated (B), BMMCs-treated (C) and CD11b⁺CD14⁺ monocyte-treated (D) groups of mice (magnification × 200); E: Levels of hepatic OPN at 2 mo after treatment with CD11b⁺CD14⁺ monocytes. ^aP < 0.05. OPN: Osteopontin. BMMCs: Bone marrow mononuclear cells.

Figure 6 Effects of monocyte therapy on the cytokine profile of tumor necrosis factor-alpha (A), interleukin-1β (B), interleukin-6 (C) and interleukin-13 (D), as measured by enzyme-linked immunosorbent assay. Data are represented graphically as the mean ± SEM of 5 mice/group. ^aP < 0.05; ^bP < 0.01. BMMCs: Bone marrow mononuclear cells; IL: Interleukin; TNF-α: Tumor necrosis factor-alpha.

Figure 7 Effects of monocyte-based therapy on chronically liver-damaged mice. A: Hepatic levels of TGF-β1; B: Splenic levels of TGF-β1; C and D: Hepatic levels of IL-17 (C) and IL-23 (D). ^aP < 0.05; ^bP < 0.01. TGF-β: Transforming growth factor-beta; IL: Interleukin; BMMCs: Bone marrow mononuclear cells.

Figure 8 Effects of monocyte-based therapy on hepatic levels of matrix metalloproteinases-9 (A), tissue inhibitors of metalloproteinase-1 (B), interleukin-10 (C) and glutathione (D), in chronically liver-damaged mice. ^aP < 0.05; ^bP < 0.01. MMP-9: Matrix metalloproteinases-9; TIMP: Tissue inhibitors of metalloproteinase; IL: Interleukin; GSH: Glutathione; BMMCs: Bone marrow mononuclear cells.