Takagi Yutaka, Nakagawa Hidemi, Matsuo Noboru, Nomura Tomoko, Takizawa Minoru, Imokawa Genji
Department of Dermatology, Jichi Medical University, Tochigi, Japan.
J Invest Dermatol. 2004 Mar;122(3):722-9. doi: 10.1111/j.0022-202X.2004.22307.x.
We have investigated the physiological significance of the glucosylation of ceramides and the subsequent deglucosylation of glucosylceramide in the synthetic pathway of acylceramide. In this metabolic pathway using [14C]-serine in organ culture, newborn murine (BALB/c) epidermis synthesizes several types of ceramides, including acylceramide, as analyzed by thin-layer chromatography. When conduritol-B-epoxide, a specific inhibitor of beta-glucocerebrosidase, was added to the culture medium, the synthesis of acylceramide was significantly suppressed in concert with a significant increase in acylglucosylceramide. Furthermore, addition of d-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol, an inhibitor of glucosyltransferase, also specifically abolished the synthesis of acylceramide whereas non-acylated ceramides were relatively less affected. We further determined whether the physiological substrate of glucosyltransferase is omega-hydroxyceramide (C30) or non-omega-hydroxylated ceramides. Of those, only non-omega-hydroxylated ceramides proved to be good substrates for glucosyltransferase in vitro. Our parallel in vitro study also demonstrated that murine epidermis contains enzymatic activity by which omega-hydroxyglucosylceramide or omega-hydroxyceramide can be converted to acylglucosylceramide or acylceramide. Collectively, these findings indicate that the majority of acylceramides found in the stratum corneum may be synthesized through a distinct sequence of enzymatic reactions consisting of the glucosylation of ceramides by glucosyltransferase, omega-hydroxylation of glucosylceramide, the acylation of omega-hydroxyglucosylceramide (possibly by an omega-acyltransferase), and the deglucosylation of acylglucosylceramide by beta-glucocerebrosidase.
我们研究了神经酰胺糖基化以及随后糖基神经酰胺去糖基化在酰基神经酰胺合成途径中的生理意义。在器官培养中使用[14C]-丝氨酸的这一代谢途径中，通过薄层色谱分析，新生小鼠（BALB/c）表皮合成了几种类型的神经酰胺，包括酰基神经酰胺。当向培养基中添加β-葡萄糖脑苷脂酶的特异性抑制剂conduritol-B-环氧化物时，酰基神经酰胺的合成受到显著抑制，同时酰基葡萄糖神经酰胺显著增加。此外，添加葡糖基转移酶抑制剂d-苏式-1-苯基-2-癸酰氨基-3-吗啉基-1-丙醇也特异性地消除了酰基神经酰胺的合成，而非酰化神经酰胺受到的影响相对较小。我们进一步确定葡糖基转移酶的生理底物是ω-羟基神经酰胺（C30）还是非ω-羟基化神经酰胺。其中，只有非ω-羟基化神经酰胺在体外被证明是葡糖基转移酶的良好底物。我们的平行体外研究还表明，小鼠表皮含有可将ω-羟基葡萄糖神经酰胺或ω-羟基神经酰胺转化为酰基葡萄糖神经酰胺或酰基神经酰胺的酶活性。总体而言，这些发现表明，角质层中发现的大多数酰基神经酰胺可能是通过一系列独特的酶促反应合成的，这些反应包括葡糖基转移酶对神经酰胺的糖基化、糖基神经酰胺的ω-羟基化、ω-羟基葡萄糖神经酰胺的酰化（可能由ω-酰基转移酶催化）以及β-葡萄糖脑苷脂酶对酰基葡萄糖神经酰胺的去糖基化。
