Уважаемый Владимир Яковлевич,
Ваше письмо "не в Science" нуждается в доработке. Очень рекомендую к чтению (в полном тексте) новую работу про холестерин и озон. А также работу о масс спектрометрическом анализе продуктов озонолиза холестерина.
Formation of Biologically Active Oxysterols During Ozonolysis of Cholesterol Present in Lung Surfactant
Melissa K. Pulfer and Robert C. Murphy
Department of Pharmacology
University of Colorado Health Sciences Center
Denver, CO 80206
Correspondence to: Dr. Robert C. Murphy
National Jewish Medical and Research Center
1400 Jackson Street
Denver, CO 80206
Tel: (303) 398-1849 Fax: (303) 398-1694
INTRODUCTION
Human exposure to 0.2 ppm levels of ozone in ambient air has been shown to cause numerous pulmonary effects such as increased airway inflammation and decreased pulmonary function (1,2). Studies of ozone in animals using up to 3 ppm ozone have been shown to cause
increased airway hyperresponsivenesss and epithelial cell death. It has been hypothesized that the very high chemical reactivity of ozone limits the distribution of this gas in the pulmonary system, preventing direct exposure to the cellular components of the lung. In part ozone may react with the various components of the epithelial cell lining fluid in the lung, also known as pulmonary surfactant, which includes proteins, lipids and single electron antioxidant agents such as ascorbic acid (3-5). Due to the very high reactivity of ozone with lipids containing double bonds, considerable emphasis has been placed on the reaction of ozone with lipid compounds in the lungs and the possibility that the adverse effects of ozone are mediated by lipid ozonized products (6). Evidence in support of this theory has been accumulating with the identification of biologically active phospholipids (7) such as 1-hexadecanoyl-2-(9-oxo-nonanoyl)-glycerophosphocholine, found following ozone exposure to lung surfactant (8). This oxidized phospholipid which eluted as a somewhat polar product on normal phase HPLC was found to initiate apoptotic death in monocytes and macrophages. However, a relatively non-polar component was also found to elute from this normal phase HPLC separation that was also cytotoxic, and preliminary data suggested that several oxidized neutral lipid products were present in this fraction.
Cholesterol is the most abundant neutral lipid present in pulmonary surfactant and this molecule has a double bond that would be susceptible to attack by ozone (9,10). While there has been some controversy about the exact chemical structure of the major ozonolysis product when cholesterol is ozonized in solution at high ozone concentrations (>0.1%) (11-14), electrospray 5 tandem mass spectrometry was recently used to characterize this chemically reactive cholesterol ozonolysis product as 5-hydroperoxy-B-homo-6-oxa-cholestane-3$,7a-diol (Scheme 1) (15). Reduction of 5-hydroperoxy-B-homo-6-oxa-cholestane-3$,7a-diol has been shown to primarily
yield 3$-hydroxy-5-oxo-5,6-secocholestan-6-al (5,6-secosterol) (11-13). In fact, this reduced product has been detected in lung homogenate and broncheoalveolar lavage fluid of rats exposed to ozone, and it has thus been suggested as a biomarker for ozone exposure (16). The recent observation of 5,6-secosterol in arterial plaques has provided evidence that the formation of ozone may also occur as the result of normal biochemical events taking place in vivo during an inflammatory response (17).
It is important to consider that the interaction between ozone and cholesterol has primarily been studied in organic solvents with high levels of ozone, where 5-hydroperoxy-B-homo-6-oxacholestane-3$,7a-diol is the major product (11,14,15). However, environmentally relevant concentrations of ozone acting on lipid cellular membranes or in lipid rich pulmonary surfactant could involve different chemistry, because of the ordered nature of the lipid bilayer, yielding alternate products. Isolated bronchoalveolar lavage fluid was exposed in vitro to precise levels of ozone in a carefully controlled ozone chamber to study the formation of cholesterol derived
ozonolysis products. This revealed the formation of 5$,6$-epoxycholesterol ($-epoxide) (Scheme1) as a more abundant product than 5-hydroperoxy-B-homo-6-oxa-cholestan-3$,7a-diol in this system. The ability of this compound and its cellular metabolites to cause cytotoxicity and to inhibit cholesterol synthesis in cultured human bronchial epithelial cells was subsequently studied.Both 5$,6$-epoxycholesterol and cholestan-6-oxo-3$,5"-diol were shown to be cytotoxic to cultured 16-HBE cells. A possible mechanism for cytotoxicity is the ability of these oxysterols to inhibit isoprenoid based cholesterol biosynthesis in these cells.
P.S. Ссылочка номер 6 также очень примечательна. Вы знакомы с работами доктора William Pryor?
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