Target-specific oxidation processes in LDL generate molecular epitopes that are more atherogenic than the native forms and are able to elicit an immunological reaction leading to the formation of anti-oxLDL autoantibodies (oxLDL-Ab) that may participate in the overall process of atherogenesis. Thus, the detection of oxLDLAb, in addition to mirroring the occurrence of in vivo LDL oxidation, will give valuable information on the occurrence of this immune response. Plasma oxLDLAb (IgG and IgM) were measured in 72 control subjects (CS) and in 80 patients with chronic renal failure (CRF), undergoing repetitive hemodialysis (N = 56) or peritoneal dialysis (N = 24), with an ELISA method using native LDL, CuSO4-oxidized LDL (oxLDL) or malondialdehyde-derivatized LDL (MDA-LDL) as antigens. To monitor cross reactivity of the antibodies detected with other oxidatively-modified proteins, human serum albumin (HSA) and MDA-derivatized HSA (MDA-HSA) were also employed as antigens. The antibody titer was calculated as the ratio of antibodies against modified versus native proteins. CRF patients had an antibody ratio significantly higher than CS as concerning anti-oxLDL IgG (1.39 +/- 0.36 vs. 1.05 +/- 0.3, P < 0.05) and IgM (2.15 +/- 0.75 vs. 1.43 +/- 0.43, P < 0.01), and anti-MDA-LDL IgG (3.05 +/- 0.74 vs. 2.04 +/- 0.42, P < 0.01) and IgM (5.55 +/- 1.79 vs. 2.9 +/- 0.85, P < 0.01). The anti-MDA-HSA antibody titer was also higher in CRF patients than in CS (2.49 +/- 0.5 vs. 1.46 +/- 0.39, P < 0.01 for IgG and 2.80 +/- 1.03 vs. 1.26 +/- 0.43, P < 0.01 for IgM).(ABSTRACT TRUNCATED AT 250 WORDS)
beta H-crystallin was exposed to radiolytically generated hydroxyl radicals at defined radical concentrations, and its capacity to act as an amine-acceptor substrate and as an amine-donor substrate for transglutaminase were investigated. [14C]Methylamine was used as a probe for labelling amine-acceptor sites; a novel biotinylated hexapeptide was used to label amine-donor sites. The results demonstrate that both primary amine incorporation and hexapeptide incorporation by transglutaminase are considerably increased after oxidative attack on the crystallin. The identity of the labelled subunits was established, and it is shown that, in both cases, this increased incorporation is not due to the production of new substrates, but that the existing incorporation sites become more susceptible. Moreover, using the newly developed probe, we could identify, for the first time, the major crystallin subunits active as amine-donor substrates (both before and after treatment) to be beta B1-, beta A3- and beta A4-crystallin. These data support the proposal that oxidative stress and transglutaminase activity may be jointly involved in the changes found in lens crystallins with age and in the development of cataract.
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