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. 2011:17:2665-71.
Epub 2011 Oct 15.

Combined treatment with antioxidants and immunosuppressants on cytokine release by human peripheral blood mononuclear cells - chemically injured keratocyte reaction

Kayoung Yi  1 , Tae Young Chung, Joon Young Hyon, Jae Woong Koh, Won Ryang Wee, Young Joo Shin
Affiliations

Combined treatment with antioxidants and immunosuppressants on cytokine release by human peripheral blood mononuclear cells - chemically injured keratocyte reaction

Kayoung Yi et al. Mol Vis. 2011.

Abstract

Purpose: To investigate the effect of antioxidants and immunosuppresants on mixed peripheral blood mononuclear cells (PBMC) - chemically injured keratocytes reaction (MLKR).

Methods: The PBMC stimulation assay was performed using chemically injured keratocytes treated with 0.05 N NaOH for 90 s (MLKR). MLKR were treated with various drugs including rapamycin, dexamethasone, mycophenoleic acid (MPA), alpha lipoic acid (ALA), and N-acetyl cysteine (NAC). Matrix metalloprotease-9 (MMP-9), transforming growth factor-beta 1 (TGF-β1), interleukin-6 (IL-6), and macrophage migration inhibitory factor (MIF) secretion profiles of activated PBMCs stimulated by NaOH-treated keratocytes were determined by ELISA.

Results: Anti-oxidants as well as immunosuppressants suppressed PBMC proliferation. MMP-9 levels were lower in antioxidants group. IL-6 levels decreased in dexamethasone group and anti-oxidants group. Combination of immunosuppressants and antioxidants suppressed more PBMC proliferation except for rapamycin + ALA group, suppressed MMP-9 production except for MPA + ALA group, decreased IL-6 levels and increased MIF levels except for rapamycin + ALA group. TGF-β1 levels were elevated in rapamycin group and rapamycin + ALA group.

Conclusions: Cytokine production was different depending on combination of drugs.Our results suggest that the different drugs should be selected for treatment according to the phases of corneal chemical burn.

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Figures

Figure 1
Figure 1
Identification of keratocytes. Inverted phase-contrast images (A) and immune florescent vimentin staining (B) of human keratocytes. Keratocytes were observed growing parallel to the etched lines. Immunofluorescence with anti-vimentin antibody, a general marker of keratocyte, showed the morphology. The nuclei were stained with Hoechst 33342 (blue). Magnification, 200×.
Figure 2
Figure 2
PBMC stimulation test. NaOH-treated human keratocytes served as the stimulators. Anti-oxidants as well as immunosuppressants suppressed the PBMC proliferation (Top left). The combination of immunosuppressants and NAC was more effective in suppression of PBMC proliferation while the combination of immunosuppressants and ALA increased PBMC proliferation except for dexamethasone.
Figure 3
Figure 3
MMP-9 levels measured by ELISA. MMP-9 levels were lower in ALA group and NAC group (Top left). The combination of immunosuppressants and antioxidants was more effective in suppression of MMP-9 production except for the combination of MPA and ALA.
Figure 4
Figure 4
IL-6 levels measured by ELISA. IL-6 levels decreased in treatment with dexamethasone and anti-oxidants (Top left). The combination of immunosuppressants and antioxidants are more effective to suppress the production of IL-6 .
Figure 5
Figure 5
MIF levels measured by ELISA. MIF levels were not different in treatment with each drug (Top left). MIF levels increased in MLKR treated with the combination of immunosuppressants and antioxidants except for the combination of rapamycin and ALA.
Figure 6
Figure 6
TGF-β1 levels measured by ELISA. TGF-β1 levels were elevated in rapamycin group, rapamycin + ALA group.
See this image and copyright information in PMC

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