Cholesterol synthesis inhibitor U18666A and the role of sterol metabolism and trafficking in numerous pathophysiological processes.

The multiple actions of U18666A have enabled major discoveries in lipid research and contributed to understanding the pathophysiology of multiple diseases. This review describes these advances and the utility of U18666A as a tool in lipid research. Harry Rudney's recognition that U18666A inhibited oxidosqualene cyclase led him to discover a pathway for formation of polar sterols that he proved to be important regulators of 3-hydroxy-3-methyl-glutaryl coenzyme A reductase.

Inhibition of fatty acid synthase by Orlistat accelerates gastric tumor cell apoptosis in culture and increases survival rates in gastric tumor bearing mice in vivo.

Orlistat, an anti-obesity drug, is a potent inhibitor of fatty acid synthase (FAS) and tumor cell viability. It can also induce apoptotic cancer cell death. We examined the effects of Orlistat on cultured NUGC-3 gastric cancer cells.

Status of caveolin-1 in various membrane domains of the bovine lens.

Recent studies of the distribution and relative concentration of caveolin-1 in fractions of bovine lens epithelial and fiber cells have led to the novel concept that caveolin-1 may largely exist as a peripheral membrane protein in some cells. Caveolin-1 is typically viewed as a scaffolding protein for caveolae in plasma membrane. In this study, membrane from cultured bovine lens epithelial cells and bovine lens fiber cells were divided into urea soluble and insoluble fractions.

Comparison of effects of U18666A and enantiomeric U18666A on sterol synthesis and induction of apoptosis.

Treatment of animals or cells with the amphipathic tertiary amine U18666A {3beta-[2-(diethylamino) ethoxy]androst-5-en-17-one} provides models for several human diseases (e.g., cataracts, Niemann-Pick disease, and epilepsy).

Multiple forms of 22 kDa caveolin-1 alpha present in bovine lens cells could reflect variable palmitoylation.

Two-dimensional immunoblots of immunoprecipitated caveolin-1 from cultured bovine lens epithelial cells revealed four to five-22 kDa forms of caveolin-1 alpha with isoelectric points of between pH values 5.5 and 6.6.

Concentration and distribution of ubiquinone (coenzyme Q), the endogenous lipid antioxidant, in the rat lens: effect of treatment with simvastatin.

Purpose: Ubiquinone (Ub) is the only known endogenously synthesized lipid soluble antioxidant. It is synthesized from intermediates in the cholesterol metabolic pathway. Our goal was to identify the Ubs and determine the concentration and distribution of Ubs in the rat lens and the effect of treatment with simvastatin, a cholesterol synthesis inhibitor, on lens levels.

Diethylstilbestrol increases intracellular calcium in lens epithelial cells.

The effects of diethylstilbestrol (DES) on steady-state intracellular calcium concentration ([Ca(2+)](i)) and resting Ca(2+) influx were examined in primary cultures of bovine lens epithelial cells using conventional fluorometric techniques (Fura-2). At low concentrations (10 microM), DES usually induced relatively rapid increases in [Ca(2+)](i) that occurred over an interval of 10-50 s and that persisted for several minutes in the continued presence of the drug. In about 10% of the cells, cyclic oscillations in [Ca(2+)](i) were seen after adding 10 microM DES.

Direct perturbation of lens membrane structure may contribute to cataracts caused by U18666A, an oxidosqualene cyclase inhibitor.

Induction of cataracts in experimental animals is a common toxic feature of oxidosqualene cyclase (OSC) inhibitors. U18666A has been shown to produce irreversible lens damage within a few weeks of treatment. Drug actions, besides reducing the availability of cholesterol, could contribute to cataract formation.

Distribution of caveolin-1 in bovine lens and redistribution in cultured bovine lens epithelial cells upon confluence.

The distribution of caveolin-1 in the lens and lens epithelial cells was determined to assess possible roles in cholesterol trafficking, cell to cell communication and signal transduction. Bovine lenses and cultured bovine lens epithelial cells (BLEC) were divided into subcellular fractions and the distribution of proteins recognized by three different caveolin-1 antibodies determined. The immunolocalization of caveolin-1 in the lens epithelium and in subconfluent and confluent cultured BLEC was probed by fluorescence microscopy and laser scanning confocal microscopy.

Discordant expression of the sterol pathway in lens underlies simvastatin-induced cataracts in Chbb: Thom rats.

Simvastatin rapidly induced cataracts in young Chbb:Thom (CT) but not Sprague Dawley (SD) or Hilltop Wistar (HW) rats. Oral treatment for 14 but not 7 days committed CT rat lenses to cataract formation. The cholesterol to phospholipid molar ratio in lenses of treated CT rats was unchanged.

Rapid, non-genomic actions of progesterone and estradiol on steady-state calcium and resting calcium influx in lens epithelial cells.

The effects of steroids on the steady-state intracellular [Ca(2+)] ([Ca(2+)](i)) and resting Ca(2+) influx in Fura-2-loaded bovine lens epithelial cells were examined to identify potential rapid, non-genomic actions. When administered in the presence of 1-2 mM extracellular Ca(2+) ([Ca(2+)](o)), 100 micro M progesterone produced large (up to 12-fold) and transient (5 min) increases in [Ca(2+)](i). These effects were abolished in EGTA-containing solutions, and were associated with large increases in the rate at which extracellularly administered Mn(2+) quenched the intracellular Fura signal.

Immunomagnetic capture of lens membrane fractions containing steroid binding protein.

This study describes the use of magnetic Dynabeads to purify microsomes from a crude microsomal fraction. A 28 kDa membrane-associated protein is proposed to mediate the binding of progesterone and other steroid hormones to ocular lens membranes and the rapid-nongenomic actions of these steroids. The subcellular location of this membrane steroid binding protein (MSBP) was probed by capture of organelles containing MSBP by magnetic beads displaying an antibody to a cytoplasmic domain of the protein.