Hepatic cell line of murine origin was grown in a humidified atmosphere of 5% CO2 at 37 °C in Dulbecco’s modified Eagle’s minimum essential medium (DMEM) (Thermo Fisher Scientific, Waltham, MA, USA): F12-Ham’s medium (GE Healthcare Life Science, South Logan, UT, USA) enriched with fetal bovine serum and insulin/transferrin/selenium. When cells reached a confluence of 90–100%, medium was removed, cells were washed twice with phosphate buffered saline followed by addition of medium free of fetal bovine serum, insulin, transferrin and selenium. Cells were then incubated for 6 h with 200 µM squalene (Sigma-Merck, Darmstadt, Germany) or with 200 nM lanosterol, dihydrolanosterol, zymostenol or desmosterol (Avanti Polar lipids, Alabaster, AL, USA). Each condition was tested in six replicates. Media were removed, cells were washed twice with phosphate buffered saline then collected and total RNA was extracted using Tri-reagent solution (Ambion, Austin, TX, USA). DNA contaminants were removed by TURBO DNAse treatment using DNA removal kit (Ambion, Austin, TX, USA). squalene and sterol effects were investigated at mRNA level by RT-qPCR assays.
Lanosterol
Manufactured by Avanti Polar Lipids
Sourced in Germany, United States, China
Lanosterol is a tetracyclic triterpenoid alcohol that is a precursor in the biosynthesis of cholesterol and other steroids. It serves as a core structural component in various biological processes and is an important intermediate in the production of many pharmaceutical and chemical compounds.
Automatically generated - may contain errors
Lab products found in correlation
Desmosterol, by Avanti Polar Lipids (3 mentions)
Dihydrolanosterol, by Avanti Polar Lipids (2 mentions)
Zymostenol, by Avanti Polar Lipids (2 mentions)
Zymosterol, by Avanti Polar Lipids (2 mentions)
Cholesterol, by Merck Group (2 mentions)
T mas, by Avanti Polar Lipids (2 mentions)
Ro 48 8071, by Cayman Chemical (2 mentions)
Ham s f12 medium, by GE Healthcare (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Tri reagent solution, by Thermo Fisher Scientific (1 mentions)
Squalene, by Merck Group (1 mentions)
Turbo dnase treatment, by Thermo Fisher Scientific (1 mentions)
Dna removal kit, by Thermo Fisher Scientific (1 mentions)
2h6 lanosterol, by Avanti Polar Lipids (1 mentions)
Lathosterol, by Avanti Polar Lipids (1 mentions)
Live dead cell assay kit, by Thermo Fisher Scientific (1 mentions)
Streptomycin, by Thermo Fisher Scientific (1 mentions)
L glutamine, by Thermo Fisher Scientific (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
4 2 hydroxyethyl 1 piperazineethanesulfonic acid hepes, by Thermo Fisher Scientific (1 mentions)
7 protocols using lanosterol
1
Murine Hepatic Cell Line Squalene and Sterol Response
2
Sterol Biosynthesis Pathway Enzymes
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DMHCA, lanosterol, [26,26,26,27,27,27-2H6]lanosterol, zymosterol, [2,2,3,4,4,-2H5]zymosterol, lathosterol, and desmosterol were from Avanti Polar Lipids, Inc. (Alabaster, AL, United States). Cholesterol and [3H]cholesterol were from Steraloids, Inc. (Newport, RI, United States) and PerkinElmer (Waltham, MA, United States), respectively. [26,26,26,27,27,27-2H6]desmosterol and [1,2,5,6α-2H4]lathosterol were from C/D/N Isotopes Inc. (Pointe-Claire, QC, Canada). Rodent chow (5P76 Prolab Isopro RMH 3000) was from T. R. Last Co. (Saxonburg, PA, United States). All other chemicals were from Sigma-Aldrich (St. Louis, MO, United States) unless otherwise indicated. Recombinant CYP27A1, CYP46A1, adrenodoxin reductase, adrenodoxin, and NADPH-CYP oxidoreductase were expressed and purified as described (Sagara et al., 1992 (link), 1993 (link); Hanna et al., 1998 (link); Mast et al., 2006 (link), 2012 (link)).
3
Lipid-Based Nanoformulation for Ocular Drug Delivery
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Lanosterol, hesperetin, distearoylphosphatidylethanolamine (DSPE)-Polyethylene Glycol 2000, distearoylphosphatidylcholine (DSPC), cholesterol (CHOL) were supplied by Avanti Polar Lipids (Beijing, China). 4–2-Hydroxyethyl-1-piperazineethanesulfonic acid (HEPES) was obtained from Thermo Fisher Scientific, China.
Human ocular epithelial cell line was procured from China. Dulbecco’s minimum essential medium (DMEM), 2.5% fetal bovine serum, L-glutamine, penicillin and streptomycin were obtained from GibcoTM (Beijing, China). Live/dead cell assay kit was procured from Thermo Fisher Scientific, China. The chemicals utilized in this experiment were obtained from Sigma (Shanghai, China). N-trimethyl chitosan was obtained from Chitolife Co., Korea. The substances utilized in the study are of premier analytical standard. The substances were utilized as acquired without added refinement. Milli-Q water was used for all experiments.
Human ocular epithelial cell line was procured from China. Dulbecco’s minimum essential medium (DMEM), 2.5% fetal bovine serum, L-glutamine, penicillin and streptomycin were obtained from GibcoTM (Beijing, China). Live/dead cell assay kit was procured from Thermo Fisher Scientific, China. The chemicals utilized in this experiment were obtained from Sigma (Shanghai, China). N-trimethyl chitosan was obtained from Chitolife Co., Korea. The substances utilized in the study are of premier analytical standard. The substances were utilized as acquired without added refinement. Milli-Q water was used for all experiments.
4
Preparation of Sterol-Cyclodextrin Complexes
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Cholesterol, ergosterol, dihydroCholesterol, coprostanol, and cholesteryl sulfate were purchased from Sigma (St. Louis, MO), epiCholesterol from Steraloids, cholestenone and lanosterol from Avanti Polar Lipids, cholesteryl N-(2-dimethylaminoethyl)carbamate and cholesteryl N-(trimethyl-ammonioethyl) carbamate chloride from Santa Cruz Biotechnology. Sterol stocks were prepared at 20 mM in chloroform, except for cholesteryl sulfate which was prepared in methanol, stored at −20 °C and pre-warmed to 25 °C prior to use. Methyl-β-cyclodextrin (MβCD)-sterol complexes were formed in phosphate buffered saline (10 mM phosphate/90 mM citrate/150 mM NaCl, pH 7.4; PBS-citrate) at a molar ratio of 1:2. Sterol suspensions were sonicated for 30 s and incubated at 37 °C overnight, at which point any insoluble sterol was removed via 0.2 μm filtration.
5
Deuterium Labeled Lipid Synthesis
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Deuterium labeled water (D2O, 99.8% pure) was purchased from Sigma. Cholesterol, lanosterol, dihydrolanosterol, ff-MAS, t-MAS, zymosterol, desmosterol, 7-dehydroCholesterol, d5-zymosterol, d6-lanosterol, and d6-sitosterol were obtained from Avanti Polar Lipids (Alabaster, AL). 25-hydroxyCholesterol (25-OHC) was obtained from Steraloids, Inc. (Newport, RI). Methyl-cyclodextrin (MCD) was obtained from Cyclodextrin Technologies (High Springs, FL). d5-zymosterol, d6-lanosterol were bound to MCD in a 1:10 and 1:20, respectively, stoichiometric ratio for solubilization as previously described (Brown et al., 2002 (link)).
6
Authenticated Small Molecule Chemical Compound Procurement
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The identity and purity of key small molecules were authenticated by LC-MS. The following compounds were purchased from Cayman Chemicals: PF-1163A, APB, SB366791, Ro 48-8071. Fenpyrazamine was purchased through LGC Standards. The following sterols were purchased through Avanti Polar Lipids: T-MAS, zymosterone, MAS-414, lanosterol, and zymostenol. Cholestanone, cholestanol, 17-OHP, and (2-hydroxypropyl)-β-cyclodextrin were purchased through Sigma-Aldrich Company. 3-Hydroxy-N-(3-methyloxyphenyl)-2-naphthamide and N-(3-Methoxyphenyl) cinnamide were purchased through Synthonix US. (2E)-N,3-diphenylprop-2-enamide, (2E)-N-(3-methylphenyl)-3-phenyl-2-propenamide, N-(2,4-dimethyoxyphenyl)-3-pheynylacrylamide, N-mesityl-3-phenylacrylamide, and beta-methyl-N-phenylcinnamamide were purchased through Sigma-Aldrich Company. Z26548267, Z27782563, and Z27782075 were purchased through Enamine Store. Paraformaldehyde was purchased through Ted Pella Inc (cat 18505).
7
Biosynthesis of Cholesterol Precursors
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