Acyl modifications in bovine, porcine, and equine ghrelins.

Ghrelin is a peptide hormone with various important physiological functions. The unique feature of ghrelin is its serine 3 acyl-modification, which is essential for ghrelin activity. The major form of ghrelin is modified with n-octanoic acid (C8:0) by ghrelin O-acyltransferase. Various acyl modifications have been reported in different species. However, the underlying mechanism by which ghrelin is modified with various fatty acids remains to be elucidated. Herein, we report the purification of bovine, porcine, and equine ghrelins. The major active form of bovine ghrelin was a 27-amino acid peptide with an n-octanoyl (C8:0) modification at Ser3. The major active form of porcine and equine ghrelin was a 28-amino acid peptide. However, porcine ghrelin was modified with n-octanol (C8:0), whereas equine ghrelin was modified with n-butanol (C4:0) at Ser3. This study indicates the existence of structural divergence in ghrelin and suggests that it is necessary to measure the minor and major forms of ghrelin to fully understand its physiology.

New insights into the role of microheterogeneity of ZP3 during structural maturation of the avian equivalent of mammalian zona pellucida.

The egg coat including mammalian zona pellucida (ZP) and the avian equivalent, i.e., inner-perivitelline layer (IPVL), is a specialized extracellular matrix being composed of the ZP glycoproteins and surrounds both pre-ovulatory oocytes and ovulated egg cells in vertebrates. The egg coat is well known for its potential importance in both the reproduction and early development, although the underlying molecular mechanisms remain to be fully elucidated. Interestingly, ZP3, one of the ZP-glycoprotein family members forming scaffolds of the egg-coat matrices with other ZP glycoproteins, exhibits extreme but distinctive microheterogeneity to form a large number of isoelectric-point isoforms at least in the chicken IPVL. In the present study, we performed three-dimensional confocal imaging and two-dimensional polyacrylamide-gel electrophoresis (2D-PAGE) of chicken IPVLs that were isolated from the ovarian follicles at different growth stages before ovulation. The results suggest that the relative proportions of the ZP3 isoforms are differentially altered during the structural maturation of the egg-coat matrices. Furthermore, tandem mass spectrometry (MS/MS) analyses and ZP1 binding assays against separated ZP3 isoforms demonstrated that each ZP3 isoform contains characteristic modifications, and there are large differences among ZP3 isoforms in the ZP1 binding affinities. These results suggest that the microheterogeneity of chicken ZP3 might be regulated to be associated with the formation of egg-coat matrices during the structural maturation of chicken IPVL. Our findings may provide new insights into molecular mechanisms of egg-coat assembly processes.

Characterization of putative tachykinin peptides in Caenorhabditis elegans.

Tachykinin-like peptides, such as substance P, neurokinin A, and neurokinin B, are among the earliest discovered and best-studied neuropeptide families, and research on them has contributed greatly to our understanding of the endocrine control of many physiological processes. However, there are still many orphan tachykinin receptor homologs for which cognate ligands have not yet been identified, especially in small invertebrates, such as the nematode Caenorhabditis elegans (C. elegans). We here show that the C. elegans nlp-58 gene encodes putative ligands for the orphan G protein-coupled receptor (GPCR) TKR-1, which is a worm ortholog of tachykinin receptors. We first determine, through an unbiased biochemical screen, that a peptide derived from the NLP-58 preprotein stimulates TKR-1. Three mature peptides that are predicted to be generated from NLP-58 show potent agonist activity against TKR-1. We designate these peptides as C. elegans tachykinin (CeTK)-1, -2, and -3. The CeTK peptides contain the C-terminal sequence GLR-amide, which is shared by tachykinin-like peptides in other invertebrate species. nlp-58 exhibits a strongly restricted expression pattern in several neurons, implying that CeTKs behave as neuropeptides. The discovery of CeTKs provides important information to aid our understanding of tachykinin-like peptides and their functional interaction with GPCRs.

The pathological role of visceral fat accumulation in steatosis, inflammation, and progression of nonalcoholic fatty liver disease.

BACKGROUND: Our previous studies have indicated a close association between visceral fat accumulation and hepatic steatosis in nonalcoholic fatty liver disease (NAFLD). This study investigated whether visceral fat accumulation was related to the pathogenesis and disease progression of nonalcoholic steatohepatitis (NASH)/NAFLD. METHODS: First, a total of 550 subjects who underwent a health checkup and measurement of visceral fat accumulation, done with a bioelectrical impedance analyzer (X-SCAN; Owa Medical, Fukuoka, Japan), were included. The relationship between visceral fat accumulation and biochemical parameters was examined. Second, a total of 74 patients with NASH/NAFLD who underwent liver biopsy were reviewed. Visceral fat accumulation was determined by abdominal computed tomography. The association between visceral fat accumulation and the histopathological grade/stage determined by the NAFLD activity score and Brunt's classification was evaluated. RESULTS: There was a significant relationship between visceral fat accumulation and glucose, triglyceride, and alanine aminotransferase (ALT; r = 0.423, P < 0.01). In stepwise regression analysis, visceral fat area (VFA), serum triglyceride level, and serum low-density lipoprotein (LDL)-cholesterol level were selected as predictor variables for serum ALT level, in a continuous manner (serum ALT level = -1.359 + 0.143 × VFA + 0.046 × triglyceride + 0.059 × LDL, R(2) = 0.217, P < 0.001). In patients with NASH, there was no correlation between histological grade and the visceral fat volume. Visceral fat accumulation in patients with stage 3/4 advanced NASH was greater than that in patients with stage 1/2 early NASH (P < 0.05). CONCLUSIONS: These results suggest that visceral fat accumulation plays a role in steatosis and fibrosis in the pathogenesis and prognosis of NAFLD.

A triplet carbene surviving a week in solution at room temperature.

A stable triplet carbene, having a lifetime at 25 degrees C of 14.5 days in a dilute benzene solution, was realized by simply changing the substituent at the 10 position of the previously most persistent carbene, di[9-(10-phenyl)anthryl]carbene, from a phenyl to a 2,6-dimethyl-4-tert-butylphenyl group.