Gene polymorphism and frequencies of the NPC1L1 gene (rs2072183, rs217434 and rs217428) in Japanese patients with dyslipidemia.

WHAT IS KNOWN AND OBJECTIVE: Niemann-Pick C1-Like 1 (NPC1L1) plays a pivotal role in intestinal cholesterol absorption. Ezetimibe is known as an inhibitor for NPC1L1 and decreases concentration of low-density lipoprotein cholesterol (LDL-C) in blood. Responses of the decrease of serum LDL-C levels to ezetimibe have been reported to be different among NPC1L1 variants. However, there are still limited data concerning the genetic variation in the NPC1L1 gene, specifically, in Japanese patients with dyslipidemia. The purpose of this study is to elucidate genotype and allele frequencies of the NPC1L1 gene in Japanese patients with dyslipidemia. METHODS: Written informed consent was obtained from all participants. All patients were administered ezetimibe at the dose of 10 mg for once a day either alone or coadministered with statins. Patient's data were retrospectively obtained from their medical records. Genomic DNA was extracted from whole blood samples and analysed three NPC1L1 SNPs (rs2072183, rs217428 and rs217434) by the direct sequencing method. RESULTS AND DISCUSSION: We found that there is a significant difference of genotype frequencies between healthy Japanese and dyslipidemic subjects in rs2072183. No significant differences were observed in rs217428 and rs217434; however, comparison of our data with literature reports suggests that there are significant differences in the frequencies of rs217428 and rs217434 between Canadian and Japanese dyslipidemic patients. WHAT IS NEW AND CONCLUSION: Our study is the first report concerning the genotype and allele frequencies of the gene coding for NPC1L1 in Japanese patients with dyslipidemia. The most notable result was to demonstrate that there exists a significant difference in rs2072183 variant between healthy Japanese and dyslipidemic subjects and also found that there exists genetic variation of rs2072183 between Japanese and Canadian patients with dyslipidemia. Our results are expected to facilitate research in the proper use of ezetimibe-based mono- or combination therapies. Further studies will be required to evaluate the effects of rs2072183 on the efficacy of LDL cholesterol reduction by ezetimibe.

Amino acid sequence and oligosaccharide attachment sites of the glycosylated domain of dog erythrocyte glycophorin.

Glycophorin was prepared from dog erythrocyte membranes by extraction with lithium diiodosalicylate and partition in aqueous phenol. Tryptic and chymotryptic treatments of the glycophorin produced two major glycopeptides labeled T1 and CH1, respectively. The glycopeptides were isolated by gel chromatography followed by ion-exchange chromatography, and subjected to amino acid sequence analysis. Both glycopeptides represented the amino-terminal domain of the major dog glycophorin; T1 of 52 residues and CH1 of 43 residues. The amino-terminal sequence of dog glycophorin does not have significant homology with those of human, horse or porcine glycophorins. This result is in good agreement with our previous proposal that there is no homology in the sequence of the amino-terminal glycosylated domain of glycophorin.

Primary structure of horse erythrocyte glycophorin HA. Its amino acid sequence has a unique homology with those of human and porcine erythrocyte glycophorins.

The complete amino acid sequence of the major sialoglycoproteins of horse erythrocyte membranes, glycophorin HA, was determined by manual sequencing methods, using tryptic, chymotryptic, and cyanogen bromide fragments. Glycophorin HA is a polypeptide chain of 120 amino acid residues and contains 10 oligosaccharide units attached to the amino-terminal side of the molecule. Its amino terminus is pyroglutamic acid. All of the oligosaccharides are linked O-glycosidically to threonine or serine residues. The amino acid sequence is consistent with the transmembrane orientation of glycophorins. There is no significant homology between the glycosylated domains of horse, human, and porcine glycophorins, but there is a considerable homology between the hydrophobic domains of the three glycophorins, which interact with the lipid bilayer of the erythrocyte membrane.

Isolation and characterization of two glycophorins from horse erythrocyte membranes.

Crude glycophorin fraction was prepared from horse erythrocyte membranes by extraction with lithium diiodosalicylate and partition in aqueous phenol. Two glycophorins, designated glycophorins HA and HB, were isolated by two different techniques: preparative gel electrophoresis in the presence of sodium dodecyl sulfate and ion-exchange chromatography in the presence of the nonionic detergent Ammonyx LO. Each glycophorin formed at least two bands on gel electrophoresis, which corresponded to a dimeric form and a monomeric form. Glycophorin HA, the major component, had a blocked amino-terminus and consisted of 70% protein and 30% carbohydrate. Glycophorin HB, the minor component, had threonine as the amino-terminus and consisted of 80% protein and 20% carbohydrate. Since glycophorin HB showed a chemical composition distinct from that of glycophorin HA, glycophorin HB was not a partially degraded form of glycophorin HA.