Modification of low density lipoprotein (LDL) by free radical oidat renders this molecular complex cytotoxic. Oxidized lipoproteins exist in vivo In atherosclerotic lesions and in the plasma of diabetic i , suggesting that Upoprotein-induced tissue damage may occur in cen diseases. We undertook purifiation and Identicati of the major cytotoxin In oxidized LDL. The lipid extract from oxidized LDL was subjected to multiple HPLC separations, and the fractions were assayed for cytotoxicity. Mass spectrometry and nuclear magnetic resonance Identified the purified toxin as 7p-hydroperoxycholest-5-en-3I3-ol Vascular cells are susceptible to the toxic effects of low density lipoprotein (LDL) or very low density lipoprotein (VLDL) after modification of the lipoproteins by free radical oxidation (1-4). Oxidation of LDL leads to cytotoxin formation whether oxidation is mediated by lipoxygenases (5), metal ions (6), or ultraviolet irradiation (7) in cell-free systems or by the action of free radicals from cultured endothelial cells (8), vascular smooth muscle cells (8), neutrophils (9), or stimulated human monocytes (10). The cytotoxic moiety of oxidized LDL (oxLDL) is extractable with organic solvents (4), and numerous candidate substances could be proposed to explain its toxic action (11).Oxidized lipoproteins occur in vivo in vascular lesions (12)(13)(14) and in plasma of certain diabetic subjects (15). OxLDL has been proposed to play a causal role in atherosclerosis (11,(16)(17)(18) is an important step toward testing evolving theories of atherogenesis that include lipoprotein oxidation (11,(16)(17)(18) and vascular injury (20,21) as putative early events.by dialysis against 2-6 /uM cupric sulfate for various times (23). OxLDL preparations were dialyzed against 0.15 M NaCl/0.5 mM EDTA, pH 8.5, to remove cupric ions. Relative oxidation, measured as thiobarbituric acid reactivity (6,24), was equivalent to 4-6 nmol of malondialdehyde (MDA) per mg of LDL cholesterol.Human foreskin fibroblasts were plated in a 1:1 (vol/vol) mixture of Dulbecco's minimal essential medium and Ham's F-12 medium (DME/F-12) supplemented with 5% (vol/vol) fetal bovine serum (6) Aliquots of5 mg ofnative LDL or oxLDL were lyophilized overnight, and the lipid was extracted with 5 ml of acetone. The mixture was sonicated for 10-20 sec, mixed for 1 min, and allowed to stand for 20 min. After centrifugation at 1000x g for 10-20 min, the residue was reextracted twice, as above. Preliminary experiments revealed that the recovery of the cytotoxic activity by this extraction procedure was equivalent or superior to other means, including chloroform/ methanol extraction ofaqueous lipoprotein solutions. Pooled extracts were dried under nitrogen and redissolved in isopropanol/acetonitrile, 1:1 (vol/vol). After centrifugation for 5 min at 1000 x g, the supernatant was analyzed by reversephase HPLC (Waters 1LBondapak C1s preparative column).The solvent gradient for elution consisted of water/ acetonitrile, 1:1 (vol/vol), which was increased over 5 ...

There is ample evidence that oxidized lipoproteins exist in vivo, not only in atherosclerotic lesions, but also associated with some experimental models of diabetes. Whether the lipoprotein oxidation is an epiphenomenon of other atherogenic or diabetogenic agents or processes or whether it is causally related to lesion formation in atherosclerosis or other forms of tissue damage in people with diabetes is unresolved. Intense interest in testing these ideas derives from in vitro observations of the ways in which oxidized lipoproteins interact with cells that are unlike the interactions with native lipoproteins. Many of these altered interactions suggest known features of atherosclerotic lesions, and recent data show that antioxidant treatment reduces the progression of vascular lesions. There are reasons to believe that hyperglycemia may worsen lipid and lipoprotein oxidation. If this observation is the case in vivo, and if it is ultimately proved that lipoprotein oxidation facilitates lesion development, these events may help explain the accelerated atherosclerosis suffered by diabetic patients. The multiple pathways for which there is evidence that hyperglycemia may contribute to oxidative events--for example, by enhancing free radical production in stimulated inflammatory cells or by forming glycation products that can propagate free radical events--suggest avenues for further research and may ultimately indicate points for intervention in the various manifestations of the disease.

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The combination of FSGS with leukemia poses a management challenge to the clinician in terms of further treatment with potentially nephrotoxic drugs, complications of nephrotic syndrome (including infections), and timing of renal transplantation. Future studies should address whether FSGS represents a glomerular response to anthracycline-induced injury in susceptible black persons.

Puromycin-induced nephrotic syndrome is an animal model of progressive renal disease. Both angiotensin converting enzyme inhibitors and lipid-lowering agents have been used to preserve renal structure and function in this model, although neither completely prevents progression. We tested the hypothesis that the combination of the two agents would be more protective than either alone. Rats were divided into five groups; all were uninephrectomized. Four groups were given puromycin at a dose of 10 mg/100 g body weight (BW) with additional doses of 4 mg/100 g BW given intraperitoneally at 4, 5, and 6 weeks thereafter. One group was given enalapril (EN) 50 mg/l dissolved in the drinking water; the second received lovastatin (L) 15 mg/kg given daily by gavage; the third received both agents; the fourth was left untreated, and the final group received no puromycin and served as the control group. Eight weeks after the initial dose of puromycin, glomerular filtration rate (GFR), as inulin clearance, and protein excretion were determined and blood was collected for cholesterol and triglycerides. Blood pressure was not different between any of the groups. At the end of the study period, serum cholesterol [mean +/- SD, 252 +/- 185 mg/dl (L), 135 +/- 101 mg/dl (L + EN)] and triglycerides (239 +/- 200, 148 +/- 158 mg/dl) were significantly lower (P < 0.001) in the lovastatin-treated groups than in the untreated puromycin group (535 +/- 255 mg/dl and 579 +/- 561 mg/dl, cholesterol and triglyceride, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

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Adriamycin induces proteinuria and glomerular changes in rats similar to those found in human focal segmental glomerulosclerosis (FSGS). Progression of this lesion may be slowed by use of angiotensin converting enzyme inhibition. To evaluate this we injected two groups of Sprague-Dawley rats with Adriamycin (2 intravenous doses of 2 mg/kg given at an interval of 3 weeks). One group of rats received enalapril (50 mg/l) in their drinking water. Control rats were injected with saline. After 28 weeks, the mean whole kidney glomerular filtration rate was significantly less and proteinuria and sclerotic index were significantly greater in rats receiving adriamycin compared with controls (P < 0.05). Administration of enalapril did not decrease proteinuria (545 +/- 398 mg/day vs 494 +/- 325 mg/day, P >0.05) or improve the glomerular filtration rate (0.31 +/- 0.18 ml/min per g kidney weight vs 0.41 +/- 0.21 ml/min per g, P = 0.27). However, treatment with enalapril significantly reduced the mean glomerular sclerotic index compared with untreated rats (1.62 +/- 0.88 vs 0.82 +/- 0.49, P = 0.05). Enalapril may be beneficial in preserving glomerular structure in this experimental model of FSGS.

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