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    Effect of cyclodextrin-encapsulated β-carotene on progesterone production by bovine luteal cells
    (Elsevier Science Bv, 2000) Arikan, S; Rodway, RG
    Experiments were conducted to examine the effect of cyclodextrin-encapsulated beta -carotene on basal or cholesterol (cyclodextrin-encapsulated), LH and dibutyryl cyclic AMP (dbcAMP)stimulated progesterone production by bovine corpus luteum cells isolated from mid-luteal heifer ovaries by collagenase digestion. Cells were cultured with serum-free DMEM/Ham's F12 medium in serum pre-treated plastic culture dishes for periods of up to 11 days. Medium was replaced after 24h and thereafter every 48h. beta -carotene was added to cultures in a carrier molecule, dimethyl-beta -cyclodextrin, to facilitate dissolution. All treatments were started on day 3 of culture. Treatment of cells with 1 or 2 mu mol/l beta -carotene resulted in sharp inhibition of progesterone production. On the contrary, treatment of cells with 0.1 mu mol/l beta -carotene resulted in significant stimulation (P < 0.05) of both basal and cholesterol-stimulated progesterone secretion. The effect of -carotene on LH or dbcAMP-stimulated progesterone production was also examined. Treatment of cells with LH or dbcAMP always resulted in stimulation of progesterone secretion (P < 0.001). However, cells treated with LH plus -carotene or dbcAMP plus beta -carotene both produced significantly (P < 0.01) less progesterone relative to those cells treated with LH or dbcAMP alone on days 7, 9 and 11 of culture. These results indicate that -carotene can enhance luteal steroidogenesis when present at low concentrations but is inhibitory at higher concentrations and that encapsulation of beta -carotene in cyclodextrin is an effective method of supplying it to cells in culture. (C) 2000 Elsevier Science B.V. All rights reserved.
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    Effects of high density lipoprotein containing high or low β-carotene concentrations on progesterone production and β-carotene uptake and depletion by bovine luteal cells
    (Elsevier Science Bv, 2000) Arikan, S; Rodway, RG
    Luteal cells were isolated from mid-luteal heifer ovaries by collagenase digestion. Cells were cultured with DMEM/Ham's F12 medium in serum pre-treated plastic culture dishes for periods of up to 11 days. As beta-carotene is almost completely insoluble in all polar solvents, it was added to cultures in either dimethyl sulphoxide (DMSO), tetrahydrofuran (THF) or as high-density lipoprotein (HDL) containing high or low beta-carotene concentrations. Medium was replaced after 24 h, thereafter medium was changed every 48 h, Treatment of cells with DMSO alone or with beta-carotene (5 mu mol/l) in DMSO both resulted in significant (P < 0.01) stimulation of progesterone production. beta-Carotene (5 mu mol/l) in THF did not alter progesterone production but 50 mu mol/l beta-carotene in TNF resulted in significant inhibition (P < 0.02) of progesterone production on days 3 and 7, Cultures were also supplemented with bovine HDL preparations containing equal concentrations of cholesterol (25 mu g/ml) but high or low beta-carotene (12.4 or 0.44 mu g/mg of cholesterol). Both HDL preparations significantly stimulated progesterone production (P < 0.001) but the high beta-carotene HDL was significantly (P < 0.02) more effective than the low beta-carotene HDL. However, when given together with bovine luteinizing hormone (bLH) or dibutyryl cAMP (dbcAMP), the high beta-carotene HDL stimulated progesterone production less than did the low NDL (P < 0.01). Uptake and depletion of beta-carotene by luteal cells were also examined in culture, beta-carotene supplementation increased luteal cell beta-carotene from an initial level of 373 ng per 10(6) cells to 2,030 ng per 10(6) cells by day 6. In contrast, the levels in control cells decreased to 14% of starting values during the same period. Cells treated with HDL containing high p-carotene on day 1 or days 1 and 3 were then incubated with or without bLH or dbcAMP for a further 2 days to investigate the effect of bLH and dbcAMP on depletion of beta-carotene by luteal cells, beta-Carotene depletion in the luteal cells was significantly higher (P < 0.05) in LH- and dbcAMP-treated cells than in the control cells in both groups. These results indicate that the use of solvents such as DMSO or THF may have undesirable effects due to alteration of cell membrane permeability. Supplementation with bLH or dbcAMP may increase the metabolism of beta-carotene in luteal cells, bLH or dbcAMP together with high beta-carotene WDL may, when combined with the effect of increased beta-carotene metabolism, give less stimulation than with low beta-carotene HDL. (C) 2000 Elsevier Science B.V. All rights reserved.