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Jeong-Hyeon Sohn, Henry J. Kaplan, Hye-Jung Suk, Puran S. Bora, Nalini S. Bora; Complement Regulatory Activity of Normal Human Intraocular Fluid Is Mediated by MCP, DAF, and CD59. Invest. Ophthalmol. Vis. Sci. 2000;41(13):4195-4202.
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purpose. To identify the molecules in normal human intraocular fluid (aqueous
humor and vitreous) that inhibit the functional activity of the
methods. Aqueous humor and vitreous were obtained from patients with
noninflammatory ocular disease at the time of surgery. Samples were
incubated with normal human serum (NHS), and the mixture assayed for
inhibition of the classical and alternative complement pathways using
standard CH50 and AH50 hemolytic assays,
respectively. Both aqueous humor and vitreous were fractionated by
microconcentrators and size exclusion column chromatography. The
inhibitory molecules were identified by immunoblotting as well as by
studying the effect of depletion of membrane cofactor protein (MCP),
decay-accelerating factor (DAF), and CD59 on inhibitory activity.
results. Both aqueous humor and vitreous inhibited the activity of the classical
pathway (CH50). Microcentrifugation revealed the major
inhibitory activity resided in the fraction with an M r ≥ 3 kDa. Chromatography on an
S-100-HR column demonstrated that the most potent inhibition was
associated with the high-molecular-weight fractions (≥19.5 kDa). In
contrast to unfractionated aqueous and vitreous, fractions with an M r ≥ 3 kDa also had an inhibitory
effect on the alternative pathway activity (AH50). The
complement regulatory activity in normal human intraocular fluid was
partially blocked by monoclonal antibodies against MCP, DAF, and CD59.
Immunoblot analysis confirmed the presence of these three molecules in
normal intraocular fluid.
conclusions. Our results demonstrate that normal human intraocular fluid (aqueous
humor and vitreous) contains complement inhibitory factors.
Furthermore, the high-molecular-weight factors appear to be the soluble
forms of MCP, DAF, and CD59.
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