The role of plasma membrane type sialidase gene Neu3 on the pathogenesis of type 2 diabetes mellitusin Japanese.

神经节苷类是存在于细胞膜表面的一类含唾液酸的鞘糖脂。一些证据表明，它们在调节细胞生长、细胞分化和跨膜信号传导等广泛的生物过程中发挥着重要的功能作用。质膜相关唾液酸酶（Neu3）是神经节苷脂水解的关键酶，在细胞表面功能调控中起着重要作用。我们利用人NEU3 cDNA研究了过表达对转基因小鼠的影响。我们证明，过表达人NEU3的小鼠会出现糖尿病表型，并伴有高胰岛素血症、胰岛增生和β细胞质量增加。与野生型相比，胰岛素刺激下胰岛素受体（IR）和胰岛素受体底物I的磷酸化水平显著降低，磷脂酰肌醇3-激酶和糖原合成酶活性降低。在低龄小鼠中，IR磷酸化在出现贫血之前就已经减弱。瞬时转染NEU3至3T3-L1脂肪细胞和L6肌细胞可导致IR信号显著降低。在对胰岛素的反应中，NEU3被发现经历酪氨酸磷酸化并随后与Grb2蛋白结合，从而被激活并引起胰岛素信号的负调控。事实上，转基因组织中可能的唾液酸酶产物GM1和GM2的积累在体外引起IR磷酸化的抑制，并且阻断与Grb2的关联导致L6细胞中受损的胰岛素信号通路的逆转。这些数据表明NEU3确实参与胰岛素信号的控制，可能是通过神经节苷脂的调节和与Grb2的相互作用，并且小鼠可以作为人类胰岛素抵抗型糖尿病的有价值的模型。我们还研究了NEU3中单核苷酸多态性（snp）对2型糖尿病发展的可能贡献。我们调查了298名日本2型糖尿病患者和两组对照日本人群的NEU3基因snp：一组由148名老年人组成，他们符合严格的非糖尿病标准，包括年龄在60岁以上且无糖尿病证据（HbA1c<5.6%），另一组由308名糖耐量正常（NGT）的受试者组成。我们鉴定了8个编码区有氨基酸取代的snp和5个编码区没有氨基酸取代的snp。其中一个snp的等位基因频率在2型糖尿病患者中显著高于老年人正常组和NGT组，而其他snp的等位基因频率在这三组中基本相同。此外，在NGT受试者中，SNP与口服葡萄糖耐量试验中胰岛素敏感性指数显著降低相关。这些结果强烈表明，在日本，NEU32基因中的SNP通过减少胰岛素信号传导与2型糖尿病的发展相关。少

Gangliosides are a family of sialic acid-containing glycosphingolipids present in the cell surface membranes. Several lines of evidence suggest their important functional roles in regulating a wide range of biological processes including cell growth, cell differentiation, and transmembrane signaling. Plasma membrane-associated sialidase (Neu3) is a key enzyme for ganglioside hydrolysis, thereby playing crucial roles in regulation of cell surface functions.We have investigated effects of overexpression in transgenic mice by using the human NEU3 cDNA. We demonstrated that mice overexpressing the human NEU3 develop diabetic phenotype associated with hyperinsulinemia, islet hyperplasia, and increased beta-cell mass. As compared with the wild type, insulin-stimulated phosphorylation of the insulin receptor (IR) and insulin receptor substrate I was significantly reduced, and activities of phosphatidylinositol 3-kinase and glycogen synthase were low in transgenic muscle. IR phosphorylation wa … More s already attenuated in the younger mice before manifestation of cemia. Transient transfection of NEU3 into 3T3-L1 adipocytes and L6 myocytes caused a significant decrease in IR signaling. In response to insulin, NEU3 was found to undergo tyrosine phosphorylation and subsequent association with the Grb2 protein, thus being activated and causing negative regulation of insulin signaling. In fact, accumulation of GM1 and GM2, the possible sialidase products in transgenic tissues, caused inhibition of IR phosphorylation in vitro, and blocking of association with Grb2 resulted in reversion of impaired insulin signaling in L6 cells. The data indicate that NEU3 indeed participates in the control of insulin signaling, probably via modulation of gangliosides and interaction with Grb2, and that the mice can serve as a valuable model for human insulin-resistant diabetes.We also investigate the possible contribution of single nucleotide polymorphisms (SNPs) in NEU3 to the development of type 2 diabetes. We surveyed SNPs in the NEU3 gene in 298 Japanese subjects with type 2 diabetes mellitus and two control Japanese populations : one consisting of 148 elderly subjects who met stringent criteria for being non-diabetic including age above 60 years and no evidence of diabetes (HbA1c<5.6%), and another 308 subjects with normal glucose tolerance (NGT). We identified eight SNPs with and five SNPs without amino acid substitutions in their coding regions. The allele frequency of one of SNPs was significantly higher in type 2 diabetic patients than in both elderly normal and NGT subjects, whereas the allele frequency of other SNPs was essentially identical in these three groups. Furthermore, in the NGT subjects, the SNP was associated with a significantly lower insulin sensitivity index on oral glucose tolerance test. These results strongly suggest that, in Japanese, the SNP in the NEU32 gene is associated with the development of type 2 diabetes, via reduced insulin signaling. Less