Metaproteomic and Metabolomic Approaches for Characterizing the Gut Microbiome.

The gut microbiome has been shown to play a significant role in human healthy and diseased states. The dynamic signaling that occurs between the host and microbiome is critical for the maintenance of host homeostasis. Analyzing the human microbiome with metaproteomics, metabolomics, and integrative multi-omics analyses can provide significant information on markers for healthy and diseased states, allowing for the eventual creation of microbiome-targeted treatments for diseases associated with dysbiosis. Metaproteomics enables functional activity information to be gained from the microbiome samples, while metabolomics provides insight into the overall metabolic states affecting/representing the host-microbiome interactions. Combining these functional -omic platforms together with microbiome composition profiling allows for a holistic overview on the functional and metabolic state of the microbiome and its influence on human health. Here the benefits of metaproteomics, metabolomics, and the integrative multi-omic approaches to investigating the gut microbiome in the context of human health and diseases are reviewed.

Mice Fed a High-Cholesterol Diet Supplemented with Quercetin-3-Glucoside Show Attenuated Hyperlipidemia and Hyperinsulinemia Associated with Differential Regulation of PCSK9 and LDLR in their Liver and Pancreas.

SCOPE: Hepatic LDL receptor (LDLR) and proprotein convertase subtilisin/kexin type 9 (PCSK9) regulate the clearance of plasma LDL-cholesterol (LDL-C): LDLR promotes it, and PCSK9 opposes it. These proteins also express in pancreatic ß cells. Using cultured hepatocytes, we previously showed that the plant flavonoid quercetin-3-glucoside (Q3G) inhibits PCSK9 secretion, stimulated LDLR expression, and enhanced LDL-C uptake. Here, we examine whether Q3G supplementation could reverse the hyperlipidemia and hyperinsulinemia of mice fed a high-cholesterol diet, and how it affects hepatic and pancreatic LDLR and PCSK9 expression. METHODS AND RESULTS: For 12 weeks, mice are fed a low- (0%) or high- (1%) cholesterol diet (LCD or HCD), supplemented or not with Q3G at 0.05 or 0.1% (w/w). Tissue LDLR and PCSK9 is analyzed by immunoblotting, plasma PCSK9 and insulin by ELISA, and plasma cholesterol and glucose by colorimetry. In LCD-fed mice, Q3G has no effect. In HCD-fed mice, it attenuates the increase in plasma cholesterol and insulin, accentuates the decrease in plasma PCSK9, and increases hepatic and pancreatic LDLR and PCSK9. In cultured pancreatic ß cells, however, it stimulates PCSK9 secretion. CONCLUSION: In mice, dietary Q3G could counter HCD-induced hyperlipidemia and hyperinsulinemia, in part by oppositely modulating hepatic and pancreatic PCSK9 secretion.

NARC-1/PCSK9 and its natural mutants: zymogen cleavage and effects on the low density lipoprotein (LDL) receptor and LDL cholesterol.

The discovery of autosomal dominant hypercholesterolemic patients with mutations in the PCSK9 gene, encoding the proprotein convertase NARC-1, resulting in the missense mutations suggested a role in low density lipoprotein (LDL) metabolism. We show that the endoplasmic reticulum-localized proNARC-1 to NARC-1 zymogen conversion is Ca2+-independent and that within the zymogen autocatalytic processing site SSVFAQ [downward arrow]SIP Val at P4 and Pro at P3' are critical. The S127R and D374Y mutations result in approximately 50-60% and > or =98% decrease in zymogen processing, respectively. In contrast, the double [D374Y + N157K], F216L, and R218S natural mutants resulted in normal zymogen processing. The cell surface LDL receptor (LDLR) levels are reduced by 35% in lymphoblasts of S127R patients. The LDLR levels are also reduced in stable HepG2 cells overexpressing NARC-1 or its natural mutant S127R, and this reduction is abrogated in the presence of 5 mm ammonium chloride, suggesting that overexpression of NARC-1 increases the turnover rate of the LDLR. Adenoviral expression of wild type human NARC-1 in mice resulted in a maximal approximately 9-fold increase in circulating LDL cholesterol, while in LDLR-/- mice a delayed approximately 2-fold increase in LDL cholesterol was observed. In conclusion, NARC-1 seems to affect both the level of LDLR and that of circulating apoB-containing lipoproteins in an LDLR-dependent and -independent fashion.