1-Nonanethiol-capped silver nanoparticles of about 4.18 nm in diameter were prepared using a liquid−liquid two-phase method. Two-dimensional ordered superlattices of the nanoparticles were formed on carbon films coated on transmission electron microscopy (TEM) copper grids by evaporating a drop of the dispersion in chloroform. The formation process of the silver nanoparticles was investigated by UV−visible absorption spectroscopy and TEM. A blue shift of the maximum absorption peak position of the UV−vis spectra occurred at the beginning of the reaction, followed by a red shift. This result indicated that large thiol-capped silver nanoparticles were formed at the beginning, then the large particles were decomposed into small particles, and in the final stage the small particles enlarged slightly again. The TEM images show directly the same process with the results from the UV−vis spectra. In addition, the UV−visible spectra of the silver nanoparticle colloidal phase obtained finally show that the system is monodisperse and can remain stable for several weeks.

Summary Diet has a substantial impact on cellular metabolism and physiology. Animals must sense different food sources and utilize distinct strategies to adapt to diverse diets. Here we show that C. elegans lifespan is regulated by their adaptive capacity to different diets, which is controlled by alh-6, a conserved proline metabolism gene. alh-6 mutants age prematurely when fed an E. coli OP50 but not HT115 diet. Remarkably, this diet-dependent aging phenotype is determined by exposure to food during development. Mechanistically, alh-6 mutation triggers diet-induced mitochondrial defects and increased generation of ROS, likely due to accumulation of its substrate 1-pyrroline-5-carboxylate. We also identify that neuromedin U receptor signaling is essential for diet-induced mitochondrial changes and premature aging. Moreover, dietary restriction requires alh-6 to induce longevity. Collectively, our data reveal a novel mechanism that animals employ to cope with potential dietary insults and uncover an unprecedented example of lifespan regulation by dietary adaptation.

Mechanisms that coordinate different metabolic pathways, such as glucose and lipid, have been recognized. However, a potential interaction between amino acid and lipid metabolism remains largely elusive. Here we show that during starvation of Caenorhabditis elegans, proline catabolism is coupled with lipid metabolism by SKN-1. Mutation of alh-6, a conserved proline catabolic enzyme, accelerates fat mobilization, enhances the expression of genes involved in fatty acid oxidation and reduces survival in response to fasting. This metabolic coordination is mediated by the activation of the transcription factor SKN-1/Nrf2, possibly due to the accumulation of the alh-6 substrate P5C, and also requires the transcriptional co-regulator MDT-15. Constitutive activation of SKN-1 induces a similar transcriptional response, which protects animals from fat accumulation when fed a high carbohydrate diet. In human cells, an orthologous alh-6 enzyme, ALDH4A1, is also linked to the activity of Nrf2, the human orthologue of SKN-1, and regulates the expression of lipid metabolic genes. Our findings identify a link between proline catabolism and lipid metabolism, and uncover a physiological role for SKN-1 in metabolism.

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Metabolic pathways are regulated to fuel or instruct the immune responses to pathogen threats. However, the regulatory roles for amino acid metabolism in innate immune responses remains poorly understood. Here, we report that mitochondrial proline catabolism modulates innate immunity in Caenorhabditis elegans. Modulation of proline catabolic enzymes affects host susceptibility to bacterial pathogen Pseudomonas aeruginosa. Mechanistically, proline catabolism governs reactive oxygen species (ROS) homeostasis and subsequent activation of SKN-1, a critical transcription factor regulating xenobiotic stress response and pathogen defense. Intriguingly, proline catabolism-mediated activation of SKN-1 requires cell-membrane dual-oxidase Ce-Duox1/BLI-3, highlighting the importance of interaction between mitochondrial and cell-membrane components in host defense. Our findings reveal how animals utilize metabolism of a single amino acid to defend against a pathogen and identify proline catabolism as a component of innate immune signaling.

Average levels of melatonin in the brain and the gastrointestinal (GIT) tissues of newborn mice declined dramatically during the first week postnatally. Food consumption increased considerably in mice bearing subcutaneous serotonin (5-HT) implants (2 mg). Melatonin implants (2 mg) also increased overall consumption but to a lesser degree. Both 5-HT and melatonin implants (2 mg) increased water content of mice fecal pellets, albeit the melatonin effect was less pronounced. Serotonin implants (2,4,6 mg/mouse) increased melatonin levels in brain, jejunum, ileum, and colon, but the effect was not dose-dependent. Intraperitoneally administered melatonin (5, 20 and 200 ug/mouse) elevated melatonin levels in brain and GIT tissues more than 100 times that of the controls, but the effect was not dose-dependent. In contrast, intraperitoneal administration of melatonin (5, 50, and 200 ug) in mice bearing a 5-HT implant (2 mg) resulted in only 3-7 times higher melatonin levels in the GIT as compared to controls, and the brain levels of melatonin were actually lower. A feedback system between 5-HT and melatonin is proposed that regulates appetite and digestive processes by endocrine as well as paracrine effects in both the brain and the GIT.

Pancreatic-derived factor (PANDER) is a cytokine-like peptide highly expressed in pancreatic beta-cells. PANDER was reported to promote apoptosis of pancreatic beta-cells and secrete in response to glucose. Here we explored the effects of glucose on PANDER expression, and the underlying mechanisms in murine pancreatic beta-cell line MIN6 and primary islets. Our results showed that glucose up-regulated PANDER mRNA and protein levels in a time- and dose-dependent manner in MIN6 cells and pancreatic islets. In cells expressing cAMP response element-binding protein (CREB) dominant-negative construct, glucose failed to induce PANDER gene expression and promoter activation. Treatment of the cells with calcium chelator [EGTA, 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl)ester (BAPTA/AM)], the voltage-dependent Ca(2+) channel inhibitor (nifedipine), the protein kinase A (PKA) inhibitor (H89), the protein kinase C (PKC) inhibitor (Go6976), or the MAPK kinase 1/2 inhibitor (PD98059), all significantly inhibited glucose-induced PANDER gene expression and promoter activation. Further studies showed that glucose induced CREB phosphorylation through Ca(2+)-PKA-ERK1/2 and Ca(2+)-PKC pathways. Thus, the Ca(2+)-PKA-ERK1/2-CREB and Ca(2+)-PKC-CREB signaling pathways are involved in glucose-induced PANDER gene expression. Wortmannin (phosphatidylinositol 3-kinase inhibitor), ammonium pyrrolidinedithiocarbamate (nuclear factor-kappaB inhibitor and nonspecific antioxidant), and N-acetylcysteine (antioxidant) were also found to inhibit glucose-induced PANDER promoter activation and gene expression. Because there is no nuclear factor-kappaB binding site in the promoter region of PANDER gene, these results suggest that phosphatidylinositol 3-kinase and reactive oxygen species be involved in glucose-induced PANDER gene expression. In conclusion, glucose induces PANDER gene expression in pancreatic beta-cells through multiple signaling pathways. Because PANDER is expressed by pancreatic beta-cells and in response to glucose in a similar way to those of insulin, PANDER may be involved in glucose homeostasis.

Melatonin receptors were studied in isolated mouse hepatocytes using the 2[¹²⁵I]iodomelatonin binding assay. The binding of 2[¹²⁵I]iodomelatonin to hepatocytes isolated from the mouse using collagenase was stable, saturable, reversible and of high affinity. The equilibrium dissociation constant (K_d) obtained from saturation studies was 10.0 ± 0.4 pmol/l (n = 16), which was comparable to the K_d obtained from kinetics studies (6.9 ± 1.2 pmol/l, n = 3), and the maximum number of binding sites (B_max) was 2.9 ± 0.4 fmol/mg protein (n = 16). The relative order of potency of indoles in competing for 2[¹²⁵I]iodomelatonin binding was 2-iodomelatonin > 2-phenylmelatonin > 6-chloromelatonin > melatonin > 6-hydroxymelatonin > N-acetylserotonin, indicating that the binding was mediated by the ML₁ receptor subtype. The linear Rosenthal plots, the close proximity of the Hill coefficient to unity and the monophasic competition curves suggest that a single class of 2[¹²⁵I]iodomelatonin binding sites is present in the mouse hepatocytes. Guanosine 5′-O-(3-thiotriphosphate) dose-dependently inhibited 2[¹²⁵I]iodomelatonin by lowering the affinity of binding, while no inhibitory effects of adenosine nucleotides were observed, suggesting that the binding sites are G-protein linked. Western immunoblotting was used to identify the melatonin receptor subtype in mouse hepatocytes using anti-Mel_1a and anti-Mel_1b. Hepatocyte membrane extract reacted with anti-Mel_1b but not anti-Mel_1a giving a peptide-blockable band of 36 kD, supporting the hypothesis that the melatonin receptors in mouse hepatocytes are of the Mel_1b subtype. Melatonin injection and a high plasma glucose level affected 2[¹²⁵I]iodomelatonin binding in the whole mouse liver homogenates. Plasma glucose was elevated by mid-light intraperitoneal injection of melatonin (4 and 40 mg/kg body weight) in a dose-dependent manner with maximum elevation achieved 1 h after injection. 2[¹²⁵I]Iodomelatonin binding at this time showed increased K_d with no changes in B_max. When the plasma glucose returned to normal within 2 h, the binding remained lowered with increased K_d but no changes in B_max. Elevation of plasma glucose by 2-deoxyglucose injection (500 mg/kg), on the other hand, decreased the binding by decreasing the B_max without affecting the K_d. Suppression of plasma glucose by insulin injection (3 IU/kg) did not change the binding. Thus, melatonin may act directly on the liver to elevate the plasma glucose level, and changes in plasma glucose level itself may in turn affect hepatic melatonin binding.