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K Ohta, T Kikuchi, N Yoshida, A Sato, S Gaun, N Yoshimura; Protective Role of Heme Oxygenase-1 Against Endotoxin-Induced Uveitis in Rats . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1555.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:Heme oxygenase-1 (HO-1) is a heat shock protein induced by oxidative stress. HO-1 metabolizes heme to the antioxidant bilirubin and carbon monoxide, and represents a powerful endogenous defensive mechanism against free radical scavenger in many diseases. In the present study, we examined the effect of hemin, an inducer of HO-1, on ocular inflammation induced by lipopolysaccharide (LPS). Methods: Endotoxin-induced uveitis (EIU) was induced by footpad injection of 200 micro g LPS in male Lewis rats. Hemin (50 mg/kg) was injected intraperitoneally 1 hour prior to LPS administration. After collecting aqueous humor (AqH) at 24 hours, the number of infiltrating cells and protein concentration in it were counted. Immunohistochemical staining with polyclonal antibody against HO-1 or HO-2 was performed to evaluate the effect of hemin. Western blot analysis was performed on homogenized iris-ciliary body (ICB) samples using same antibodies. HO-1, HO-2, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-6 and iNOS mRNA expression on ICB was determined by using RT-PCR. The levels of TNF-alpha and IL-6 in AqH were also evaluated by ELISA. The concentration of nitrate/nitrite in AqH was also measured by using Griess reagents. Results:The cell number in AqH was 27039 cells/ml of rats injected LPS, whereas 306.8 cells/micro l of those treated with hemin (p<0.001). The concentration of protein was significantly lower in AqH from rats treated with hemin than those from rats without it (3.10.6 mg/ml vs. 8.60.8 mg/ml) (p<0.001). Positive staining with anti-HO-1 in ICB observed by both immunohistochemistry and western blot analysis at 24 hours after LPS administration was enhanced with hemin treatment. ICB at 6 hours after LPS injection exhibited increased expression of TNF-alpha, IL-6 and iNOS mRNA, which was decreased with hemin treatment. On the other hand, HO-1 mRNA was up-regulated with hemine. Hemin also significantly diminished the levels of TNF-alpha, IL-6 and nitrate/nitrite in AqH. Conclusion:These results suggest that hemin is effective in inducing HO-1 and in reducing ocular inflammation afflicted with EIU by down regulating at least pro-inflammatory cytokine expression. We suggest that induction of endogenous HO-1 may be an effective treatment of uveitis or other ocular inflammation.
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