1. Volume 57, Issue 2, Pages 697-708 (October 2000)
Transforming growth factor-β is involved in the pathogenesis of dialysis-related amyloidosis 
Kenzo Matsuo, T. Alp Ikizler, Richard L. Hoover, Masahiko Nakamoto, Chikao Yasunaga, Lara B. Pupim, Raymond M. Hakim 
Kidney International 
Volume 57, Issue 2, Pages (October 2000)
DOI: /j x
Copyright © 2000 International Society of Nephrology Terms and Conditions

2. Figure 1
Immunohistochemical study for transforming growth factor betas (TGF-βs) and their receptors in the synovial tissues of dialysis related amyloidosis (DRA). (A) Anti-AGE immunostaining. (B) Anti-β2-microglobulin (anti-β2m) immunostaining. (C) Anti-CD68 immunostaining. (D) Anti–TGF-β1 immunostaining. (E) Anti–TGF-β2 immunostaining. (F) Anti–TGF-β3 immunostaining. (G) Anti–TGF-β receptor type I immunostaining. (H) Anti–TGF-β receptor type II immunostaining. (I) Anti–TGF-β receptor type III immunostaining. (J) Antinon-immunized IgG immunostaining. Most of AGE was detected in the amyloid deposition. CD68-positive macrophages mainly infiltrated around the amyloid deposition and in the synovial lining layer. TGF-β1, -2, and -3 were mainly detected in the infiltrated CD68-positive macrophages and synovial lining cells. They were also localized in the vessel walls, although faintly detected in the synovial intimal cells. Their receptors of types I, II, and III were also almost localized in the same area of TGF-βs. Original magnifications of (A–J) were ×100.
Kidney International  , DOI: ( /j x)
Copyright © 2000 International Society of Nephrology Terms and Conditions

3. Figure 1
Immunohistochemical study for transforming growth factor betas (TGF-βs) and their receptors in the synovial tissues of dialysis related amyloidosis (DRA). (A) Anti-AGE immunostaining. (B) Anti-β2-microglobulin (anti-β2m) immunostaining. (C) Anti-CD68 immunostaining. (D) Anti–TGF-β1 immunostaining. (E) Anti–TGF-β2 immunostaining. (F) Anti–TGF-β3 immunostaining. (G) Anti–TGF-β receptor type I immunostaining. (H) Anti–TGF-β receptor type II immunostaining. (I) Anti–TGF-β receptor type III immunostaining. (J) Antinon-immunized IgG immunostaining. Most of AGE was detected in the amyloid deposition. CD68-positive macrophages mainly infiltrated around the amyloid deposition and in the synovial lining layer. TGF-β1, -2, and -3 were mainly detected in the infiltrated CD68-positive macrophages and synovial lining cells. They were also localized in the vessel walls, although faintly detected in the synovial intimal cells. Their receptors of types I, II, and III were also almost localized in the same area of TGF-βs. Original magnifications of (A–J) were ×100.
Kidney International  , DOI: ( /j x)
Copyright © 2000 International Society of Nephrology Terms and Conditions

4. Figure 2
Effect of advanced glycation end-product modified β2m (AGE-β2m) on TGF-β1 secretion by macrophages. Levels of TGF-β1 in supernatants from in vitro-derived macrophages cultured under various conditions of normal β2m (12.5, 25, 50 μg/mL), AGE-β2m (12.5, 25, and 50 μg/mL), anti–TGF-β1 antibody (10 μg/mL), AGE-BSA (50 μg/mL), and LPS (0.1 μg/mL) for 24 hours. TGF-β1 levels (pg/mL) were determined by ELISA. Data are shown as means ± SD from four separate representative experiments using monocytes from different donors (N = 4). Abbreviation Ab is anti–TGF-β1 antibody. *P < 0.01 vs. normal β2m. **P < 0.01 between 12.5 and 50 μg/mL of AGE-β2m.
Kidney International  , DOI: ( /j x)
Copyright © 2000 International Society of Nephrology Terms and Conditions

5. Figure 3
Effect of TGF-β1 on TNF-α secretion by macrophages. Levels of TNF-α in supernatants from in vitro-derived macrophages cultured under various conditions of normal β2m (12.5, 25, and 50 μg/mL), AGE-β2m (12.5, 25, and 50 μg/mL), AGE-β2m and TGF-β1 (0.1, 1, and 10 ng/mL), AGE-β2m and anti–TGF-β1 antibody (10 μg/mL), AGE-BSA (50 μg/mL), and LPS (0.1 μg/mL) for 24 hours. TNF-α levels (pg/mL) were determined by ELISA. Data are shown as means ± SD from four separate representative experiments using monocytes from different donors (N = 4). Ab is anti–TGF-β1 antibody. *P < vs. normal β2m. **P < between those of 12.5 and 50 μg/mL of AGE-β2m.
Kidney International  , DOI: ( /j x)
Copyright © 2000 International Society of Nephrology Terms and Conditions

6. Figure 4
Effect of TGF-β1 on IL-1Ra secretion by macrophages. Levels of IL-1Ra in supernatants from in vitro-derived macrophages cultured under various conditions of normal β2m (12.5, 25, and 50 μg/mL), AGE-β2m (12.5, 25, and 50 μg/mL), AGE-β2m and TGF-β1 (0.1, 1, and 10 ng/mL), AGE-β2m and anti–TGF-β1 antibody (10 μg/mL), AGE-BSA (50 μg/mL), and LPS (0.1 μg/mL) for 24 hours. IL-1Ra levels (pg/mL) were determined by ELISA. Data are shown as means ± SD from four separate representative experiments using monocytes from different donors (N = 4). Ab is anti–TGF-β1 antibody. *P < 0.05 vs μg/mL normal β2m. **P < 0.01 vs. 50 μg/mL normal β2m. ***P < between those of 12.5 and 50 μg/mL of AGE-β2m.
Kidney International  , DOI: ( /j x)
Copyright © 2000 International Society of Nephrology Terms and Conditions

7. Figure 5
Effect of TGF-β1 on IL-1β secretion by macrophages. Levels of IL-1β in supernatants from in vitro-derived macrophages cultured under various conditions of normal β2m (12.5, 25, and 50 μg/mL), AGE-β2m (12.5, 25, and 50 μg/mL), AGE-β2m and TGF-β1 (0.1, 1, and 10ng/mL), AGE-β2m and anti–TGF-β1 antibody (10 μg/mL), AGE-BSA (50 μg/mL), and LPS (0.1 μg/mL) for 24 hours. IL-1β levels (pg/mL) were determined by ELISA. Data are shown as means ± SD from four separate representative experiments using monocytes from different donors (N = 4). Ab is anti–TGF-β1 antibody.
Kidney International  , DOI: ( /j x)
Copyright © 2000 International Society of Nephrology Terms and Conditions

8. Figure 6
Involvement of TGF-β1 in AGE-β2m induced monocyte chemotaxis. Monocyte migration was tested in response to increased concentrations of (A) TGF-β1 (0.1, 1, and 10 ng/mL), normal β2m (12.5, 25, and 50 μg/mL) alone and those various combinations (B) AGE-β2m (12.5, 25, and 50 μg/mL), TGF-β1 (0.1, 1, and 10 ng/mL) alone and those various combinations, AGE-BSA (50 μg/mL; (C) various combinations of AGE-β2m (12.5, 25, and 50 μg/mL) and anti–TGF-β1 antibody (Ab; 0.1, 0.5, 1, and 10 μg/mL), and nonimmune IgG. FMLP (10-6 mol/L) and medium itself were used as a positive, negative control, respectively. Chemotactic activity is defined as the mean number of monocytes that had migrated to the lower surface of the filters. The number of cells per eight high-power fields (hpf; ×400) was counted through a microscope. Cell migration assay was duplicated using healthy human PBMCs. TGF-β1 had the same level of chemotactic activity during this range. Data are shown as means ± SD from four representative experiments (N = 4).
Kidney International  , DOI: ( /j x)
Copyright © 2000 International Society of Nephrology Terms and Conditions