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Background: Diet-induced hepatic steatosis is highly associated with nonalcoholic fatty liver disease, which is related to the development of metabolic syndrome. While advanced stage nonalcoholic hepatic steatosis and steatohepatitis (NASH) result ultimately in fibrosis and cirrhosis, the molecular basis for lipid droplet formation is poorly understood. Common pathways underlie the pathology of mammalian obesity and the zebrafish diet-induced obesity model (DIO-zebrafish) used in this study. Methods: Our analysis involved a combination of transcriptome (DNA microarray) and proteome (two-dimensional electrophoresis) methods using liver tissue from DIO-zebrafish to find candidate genes involved in hepatic steatosis. We conducted intraperitoneal injection (i.p.) of morpholino antisense oligonucleotides (MOs) for each gene into DIO-zebrafish. We also conducted in vitro overexpression in human cells. Additionally, we examined gene expression during feeding experiments involving anti-obesity compounds, creatine and anserine. Results: We found that fatty acid binding protein 3 (fabp3) and E2F transcription factors were upregulated in hepatic steatosis. E2f8 MO i.p. suppressed fabp3 expression in liver, and ameliorated hepatic steatosis. In human cells (HepG2), E2F8 overexpression promoted FABP3 expression. Additionally, co-administration of creatine and anserine suppressed obesity associated phenotypes including hepatic steatosis as indicated by e2f8 and fabp3 down regulation. Conclusion: We discovered that the e2f8--fabp3 axis is important in the promotion of hepatic steatosis in DIO-zebrafish. The combination of transcriptome and proteome analyses using the disease model zebrafish allow identification of novel pathways involved in human diseases. [ABSTRACT FROM AUTHOR]

Dissolved proteins in seawater samples collected from a coastal area of Tokyo Bay, Sagami Bay and a location off the Kuroshio Current were investigated by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and high resolution two-dimensional electrophoresis (2-DE). Four to nine protein bands were detected in SDS-PAGE in the apparent molecular weight (MW) range from 12 kilo Dalton (kDa) to 49 kDa. The 2-DE technique distinguished 10 to 46 protein spots exhibiting isoelectric point (p I)/MW ranging 4.3–9.2/12–63 kDa. The elecrophoretic patterns were similar between the coastal and pelagic samples, as well as previously reported patterns from various pelagic areas. The close similarity of electrophoretic mobility on both SDS-PAGE and 2-DE gels indicates the compositional homogeneity of dissolved proteins in seawater throughout a broad range of marine environments. Proteinaceous dissolved organic matter (DOM) that was unresolved and smeared staining characteristics on both SDS-PAGE and 2-DE gels was first observed in Tokyo Bay waters in the present study and its possible sources are discussed. Although the two protein species, 48 kDa and 39 kDa proteins, have been identified as homologues of Porin P and low molecular weight-alkaline phosphatase of Pseudomonas aeruginosa PAO1, respectively, four strains of P. aeruginosa and two species of Pseudomonas spp. have been newly identified as the source organisms of these proteins using the N-terminal amino acid sequence data determined in previous studies. [ABSTRACT FROM AUTHOR]

Objectives: To evaluate improved protein extraction and two-dimensional electrophoresis (2DE) separation methods with Japanese reference human hair (JRH); to determine whether fibre curvature is related to protein composition in curly and straight Japanese women's human hair (JHH) samples; and to identify proteins from JRH 2DE maps and expression differences between curly and straight JHH., Methods: Hair keratin and keratin-associated proteins (KAPs) were extracted intact with dithiothreitol or tris(2-carboxyethyl) phosphine from JRH or from curved or straight JHH. Extracted proteins were isoelectric-focused on first-dimensional pH gradient gel strips, then separated by molecular weight on laboratory-made, second-dimension, large format gels. The software compared protein abundance between duplicate 2DE gels of curved and straight JHH. Thirty-eight proteins from a JRH 2DE gel were enzyme-cleaved for MALDI-TOF-MS analysis to determine peptide composition, and where possible, de novo sequencing gave peptide sequence data. An in-house human hair protein database incorporating ninety-eight annotated protein sequences assisted MS analysis., Results: 2DE gels of tris(2-carboxyethyl) phosphine-extracted JRH improved keratin and KAP resolution and number compared to those of dithiothreitol-extracted JRH and published commercially made second-dimensional gels. Silver-stained 2DE gels of the straight or curved JHH sets were remarkably similar. Over-staining to reveal basic proteins caused poor resolution of the major acidic protein classes. Software comparisons of fifty-nine resolved proteins revealed two were significantly different in abundance between curved and straight hairs but in insufficient amounts for MS analysis. MS identified twelve proteins from a JRH CBBG-stained 2DE gel: six type II keratins, three type I keratins and three high sulphur proteins. A further eight were potential conformational isoforms and isoelectric variants of the identified proteins bringing the total to twenty identified or partially identified proteins., Conclusion: Root-end human hair extraction with tris(2-carboxyethyl) phosphine improves protein resolution and visualizes more proteins on large format 2DE gels. The two minor protein differences between duplicate straight or curved JHH 2DE gels were unlikely to change fibre structure from straight to curved hair. MS results confirmed that multiple isoforms exist of various hair proteins. Low sequence coverage prevented distinction between members in rows of homologous protein spots of similar molecular weight., (© 2020 Society of Cosmetic Scientists and the Société Française de Cosmétologie.)