Roland C. Aloia

Characterization of highly purified, inactivated HIV-1 particles isolated by anion exchange chromatography

This report characterizes inactivated, gp120 depleted, HIV-1 particles purified by an anion exchange chromatography production process. This antigen formulated with incomplete Freunds adjuvant constitutes Remune, which is being evaluated in a phase III clinical endpoint trial to determine the effect of this immune-based therapy on clinical progression of HIV-1 seropositive patients. Multiple production lots of the inactivated HIV-1 antigen strain HZ321, isolated by anion exchange chromatography, exhibit purity of > 95% by gel filtration. These findings are corroborated by thin section electron microscopy showing a homogenous field of intact particles. Analyses of the purified virus particles for protein, lipid, carbohydrate and RNA show structural retention of the envelope proteins, lipid bilayer and core components after large scale processing. The qualitative identification of at least 85% of total HIV-1 protein is determined by ELISA, Western blot, HPLC and amino acid sequencing analyses. Quantitative values are assigned to 50% of these proteins. The data confirm the presence of virally encoded proteins p6, p7, pI15, p17, p24, p32, pI39Gag, gp41, pp55Gag, p66/51, Vpr, Vif and Nef. Excellent consistency between production lots and equivalency to HIV-1 preparations purified by sucrose density gradient sedimentation has been established for protein and lipid composition, and overall purity. These findings further establish that non-viral encoded proteins and lipids are integral structural components of the intact virion and are not contaminants unique to a particular isolation method. The data confirm the presence of multicomponent antigens in the viral particles for stimulating a broad HIV-1 specific immune response. Finally, the work demonstrates that the two inactivation procedures (beta-propiolactone and gamma irradiation), which achieve efficient viral inactivation meeting US FDA guidelines, do not damage the protein antigens of the viral particles.

Effect of a high fat diet on phospholipid class distribution and fatty acid composition in rat liver

1. Long term consumption (20 weeks) of a high fat diet (65% of the energy content as fat) rich in either saturated [30% (w/w) coconut oil] or unsaturated [30% (w/w) sunflower oil] fatty acids resulted in strikingly similar alterations in the phospholipid class distribution and fatty acid composition in the liver of male Wistar rats. 2. The effect of these two diets was compared to a control group maintained on a 2% fat diet (w/w) for the same time interval. 3. In spite of the difference in the PUFA/SFA (polyunsaturated fatty acid/saturated fatty acid) ratio between the two high fat diets (0.1, saturated fatty acid diet; 5.4, unsaturated fatty acid diet), both diets resulted in a similar PUFA/SFA ratio in liver phospholipids, a similar reduction in palmitic acid (16:0), oleic acid (18:1, n-9) and arachidonic acid (20:4, n-6) and an elevation in stearic acid (18:0), linoleic acid (18:2, n-6) and docosahexaenoic acid (22:6, n-3). 4. Further, changes in the phospholipid classes were also similarly affected by both high fat diets.

Effect of a high fat diet on plasma lipids, lipoprotein lipase, lecithin:cholesterol acyltransferase, and insulin function in adult rabbits

Abstract The influence of a high fat diet (19% wt./wt.) vs. a standard, chow diet (2% fat) on plasma lipids, lipoprotein lipase (LPL), lecithin:cholesterol acyltransferase (LCAT), glucose, and glucose tolerance, was investigated. Both diets had a similar fatty acid pattern and a polyunsaturated:saturated (P:S) fatty acid ratio of 2.7. The high fat diet elevated plasma triglycerides, phospholipids, and cholesterol concentrations and changed the percent distribution of cholesterol and phospholipids among the lipoprotein fractions. Additionally, LCAT and plasma glucose increased, while lipoprotein lipase and its products, free fatty acids and glycerol, were not altered. Glucose tolerance was significantly inhibited in animals on the high fat diet, which also exhibited a diminished insulin secretion. As such, the high fat diet seemed to evoke a diabetogenic situation. The return to the standard chow diet appeared to normalize, to a great extent, the alterations evoked by the high fat diet.

Effect of skim milk and whey diets on plasma lipid levels of rabbits in a cross-over study

The influence of a skim milk and a whey-based diet on plasma lipids and lipoproteins in rabbits was investigated in a crossover design, in which the diets were switched after 12 weeks. The whey diet increased plasma lipid levels regardless of when it was consumed in the feeding protocol. The milk diet, on the other hand, modestly elevated the lipid levels if provided in the first half of the feeding protocol, but reduced lipid levels if given in the second half of the feeding regime. Both diets also induced similar changes in lipoprotein patterns when given in the first half of the feeding protocol, but opposite effects during the second half. Atherosclerotic changes were also observed in the aorta of animals receiving the whey diet during the second half of the experimental protocol.