c6 nbd cer  (Croda International Plc)

Journal: The Journal of Biological Chemistry

Article Title: Differentiation-linked changes in the biosynthesis and turnover of sphingomyelins in rat male germ cells: Genes involved and effects of testosterone

doi: 10.1016/j.jbc.2023.103058

Figure Lengend Snippet: Formation of SM, GlcCer, and Cer from NBD-labeled substrates in rat spermatogenic cells in primary culture.A, PtS, RS, and LS were incubated up to 5 h in the presence of NBD-C6-Cer. B and C, total spermatogenic cells (TC) were incubated up to 25 h in the presence of NBD-C6-Cer and NBD-C6-SM, respectively. The fluorescent lipid products were extracted, separated by HPTLC, and their fluorescence was expressed on the basis of cell protein content (mean values ± SD, n = 3). Cer, ceramide; GlcCer, glucosylceramide; LS, late spermatids; PtS, pachytene spermatocytes; RS, round spermatids; SM, sphingomyelin.

Article Snippet: The NBD-C6-Cer and NBD-C6-SM (Molecular Probes) used as substrates were diluted to a concentration of 1 mM in ethanol, from which aliquots were taken to reach a final concentration of 2 μM when added to cell suspensions.

Techniques: Labeling, Incubation, High Performance Thin Layer Chromatography, Fluorescence

Journal: The Journal of Biological Chemistry

Article Title: Differentiation-linked changes in the biosynthesis and turnover of sphingomyelins in rat male germ cells: Genes involved and effects of testosterone

doi: 10.1016/j.jbc.2023.103058

Figure Lengend Snippet: Formation of Cer, SM, and GlcCer from NBD-labeled substrates by rat germ cells in the presence of testosterone.A, formation of NBD-C6-SM from NBD-C6-Cer (left panel) and of NBD-C6-Cer from NBD-C6-SM (right panel) in total spermatogenic cells (TCs) in the absence and presence of testosterone (Tes) or estradiol (E2). The cells were incubated during 5 h in the medium containing either the vehicle ethanol (controls) or a 10−7 M concentration of each hormone. Then, the NBD-labeled substrates were added, and incubations continued for 3 h. B, effect of D609, inhibitor of SM synthase(s), on the amounts of NBD-C6-SM formed from NBD-C6-Cer in the absence and presence of testosterone. TCs were preincubated for 5.5 h without (controls) or with 15 μM D609. At t = 30 min during this period, testosterone or its vehicle were added, and incubations proceeded as before for 5 h, followed by NBD-C6-Cer addition and incubation for 3 h. C, formation of NBD-C6-SM and NBD-C6-GlcCer from NBD-C6-Cer in isolated PtS, RS, and LS, treated as described for TC in part A, i.e., preincubated for 5 h with the vehicle or the hormone (Tes or E2), followed by addition of NBD-C6-Cer and incubation for 3 h. The products in A, B, and C were isolated by TLC their fluorescence was quantified, and the data are presented as percentages with respect to the corresponding controls (mean values ± SD, n = 3). Significant differences with controls (p < 0.01) are indicated by asterisks (∗). Cer, ceramide; GlcCer, glucosylceramide; SM, sphingomyelin.

Article Snippet: The NBD-C6-Cer and NBD-C6-SM (Molecular Probes) used as substrates were diluted to a concentration of 1 mM in ethanol, from which aliquots were taken to reach a final concentration of 2 μM when added to cell suspensions.

Techniques: Labeling, Incubation, Concentration Assay, Isolation, Fluorescence