of 10 M pregnenolone formation of EC was almost completely Away constantly and does not accumulate esterified cholesterol in the lysosomes of macrophages. After removal of pregnenolone by SB-207499 Cilomilast inhibitor washing the cells with buffer cholesterol metabolism of lysosomal CE was repeated in the presence or absence of beauveriolides. As shown in FIG. 3, I and III inhibits the synthesis of EC beauveriolides dosedependent manner with IC50 values of 0.37 and 0.21 m, respectively, from the very Similar to those of synthesis Ls Ure EC are. Second under the same conditions, Fig Effect beauveriolides I and III and beauvericin on neutral lipid synthesis 14C Ls Acid by macrophages. Monolayers of macrophages obtained from 5105 cells per well in a microtiter plate in 0.
25 ml medium were incubated with a liposome A phospholipid cholesterol consists of phosphatidylcholine, phosphatidylserine, and cholesterol in Pelitinib a molar dicetylphosphate Ratio of Ls Acid 10:10:2 : 15 and I in the absence or presence of indicated amounts beauveriolide or III or beauvericin. After incubation for 14 h oleate, cholesterol and triglycerides were determined on TLC with radio scanner, as described in Materials and Methods and plotted separately percent embroidered. FIG third Effect beauveriolides I and III lysosomal cholesterol metabolism. Macrophages in 48-well microtiter plates were incubated with 10 l plastic cultured cholesterol liposomes for 12 hours in the presence of 10 M pregnenolone erg Incubated complements.
After incubation, the medium was removed and the cells in each well were washed twice with buffer B and once bufferBcontainingBSA, they were then incubated in 0.25 ml of medium A containing beauveriolide I, III, 283 546 CL or cytochalasin D. After incubation for 5 h was separated by TLC and the radioactivity CE t was measured by a radio scanner, as described in Materials and Methods. The results were expressed as percent stitched on. Namatame et al. PNAS 20th January 2004, vol. No. 3 involved 739 101 E RY BIOCHIMIST cytochalasin D, an inhibitor of actin polymerization in endocytosis of liposomes did not affect lysosomal cholesterol synthesis even EC 20 M, wherein the compound inhibits the synthesis of EC Ls Acid with an IC50 value of 2.4 M. Likewise inhibited CL 283,546, a synthetic ACAT inhibitor, the synthesis of cholesterol and synthesis of lysosomal CE ls ure with the EC almost anything similar IC50 values.
These data show that the process of cholesterol metabolism beauveriolides block postlysosomal entered Ing inhibition of the synthesis of CE in macrophages. The inhibition of macrophage ACAT Beauveriolides mouse. Indicated experience with pregnenolone that inhibition beauveriolides place between the start of cholesterol from lysosomes and the point of cholesterol esterification in the ER is. Therefore, their effect on ACAT activity Judged t because Cyclodepsipetides known as beauvericin and other Enniatins with gr Eren cyclic skeletons to the activity t Inhibit ACAT. To do so, microsomes prepared from mouse peritoneal macrophages, mouse liver and human Caco 2 cells were used as the enzyme source. Beauveriolides I and III was found ACAT activity T inhibit in microsomes of mouse macrophage