[Dynamic Microbial Shifts and Functional Analysis of Saliva Microbial Communities with Caries Children].

Objective: To observe the dynamic changes in the salivary microbiota of children with dental caries and those who were caries-free and to analyze the functional differences in the oral microecology of the two groups during the course of sugar metabolism and the synthesis and transport of multiple amino acids. Methods: Ten children with dental caries and 10 caries-free children were enrolled. We employed Illumina metagenomics technology to analyze the composition and function of salivary microbiome in children with and without caries. Six months later, PacBio single-molecule long-read sequencing technology was used to analyze the changes over time in the oral microbial communities of the two groups. We studied the patterns of change in the oral microbial communities under diseased or healthy conditions and attempted to offer a comprehensive interpretation of children's oral microbiota in terms of its composition and functions. Results: The composition of the oral microbiota of children with or without dental caries changed significantly over time. At the phylum level, changing trends in the salivary microbial communities of children with dental caries were in line with those in caries-free children. In these microbial communities, increased proportions of Firmicutes and decreased proportions of Actinobacteria and Bacteroidetes were found in the two groups. At the genus level, however, the two groups showed significantly different changes of the salivary microbial communities. Upward trends in the abundance of Lactobacillus, Methylobacterium, and Megasphaera were found in the caries group, while the abundance of these genera in the caries-free group showed downward trends. At the species level, L. fermentum, L. gasseri, L. oris, S. downei, and some other species belonging to the genus Lactobacillus showed upward trends in the saliva of children with caries, while they consistently stayed at a relative low level of abundance in caries-free children. The abundance of S. gordonii and S. mutans decreased to a certain extent in children with dental caries, but the abundance of S. gordonii and S. mutans in caries-free children were always at a low level. Species such as S. mutans and C. gracilis were positively correlated to the sum of decayed, missing and filled teeth (dmft), while N. flavescens was negatively correlated to dmft. gltA, icd, and mqo, the key genes related to tricarboxylic acid (TCA) cycle, gudB, a glutamate synthesis-related gene, and argAB/C/J, arginine synthesis-related genes, were significantly increased in caries-free children. In addition, the abundance of the NADH dehydrogenase-related gene nuoB/C/D/E/H/I/J/K/L/M in the electron transport chain increased significantly in caries-free children. Conclusion: Dynamic changes were found in the oral microbiota of children with or without caries. The trends of microbial shifts over time were associated with the oral health status. Oxidative phosphorylation and the synthesis and transport of amino acids such as glutamate and arginine in the oral microecology were more active in caries-free children.

Oleanolic acid derivative DKS26 exerts antidiabetic and hepatoprotective effects in diabetic mice and promotes glucagon-like peptide-1 secretion and expression in intestinal cells.

BACKGROUND AND PURPOSE: Glucagon-like peptide-1 (GLP-1) is an important target for diabetes therapy based on its key role in maintaining glucose and lipid homeostasis. This study was designed to investigate antidiabetic and hepatoprotective effects of a novel oleanolic acid derivative DKS26 in diabetic mice and elucidate its underlying GLP-1 related antidiabetic mechanisms in vitro and in vivo. EXPERIMENTAL APPROACH: The therapeutic effects of DKS26 were investigated in streptozotocin (STZ)-induced and db/db diabetic mouse models. Levels of plasma glucose, glycosylated serum protein (GSP), lipid profiles, insulin, alanine aminotransferase (ALT) and aspartate aminotransferase (AST), oral glucose tolerance (OGT), pancreatic islets and hepatic histopathological morphology, liver lipid levels and expression of pro-inflammatory cytokines were assessed. Intestinal NCI-H716 cells and diabetic models were used to further validate its potential GLP-1-related antidiabetic mechanisms. KEY RESULTS: DKS26 treatment (100 mg·kg-1 ·day-1 ) decreased plasma levels of glucose, GSP, ALT and AST; ameliorated OGT and plasma lipid profiles; augmented plasma insulin levels; alleviated islets and hepatic pathological morphology; and reduced liver lipid accumulation, inflammation and necrosis in vivo. Furthermore, DKS26 enhanced GLP-1 release and expression, accompanied by elevated levels of cAMP and phosphorylated PKA in vitro and in vivo. CONCLUSION AND IMPLICATIONS: DKS26 exerted hypoglycaemic, hypolipidaemic and islets protective effects, which were associated with an enhanced release and expression of GLP-1 mediated by the activation of the cAMP/PKA signalling pathway, and alleviated hepatic damage by reducing liver lipid levels and inflammation. These findings firmly identified DKS26 as a new viable therapeutic option for diabetes control.

Effects and molecular mechanisms of the antidiabetic fraction of Acorus calamus L. on GLP-1 expression and secretion in vivo and in vitro.

ETHNOPHARMACOLOGICAL RELEVANCE: The radix of Acorus calamus L. (AC) is widely used in diabetes therapies in traditional folk medicine from America and Indonesia, and we have previously reported that the ethyl acetate fraction of AC (ACE) acts as an antidiabetic through insulin sensitizing, insulin releasing and alpha-glucosidase inhibitory activities. The present study is designed to investigate the effects and molecular mechanisms of ACE on glucagon-like peptide-1 (GLP-1) expression and secretion related to its hypoglycemic effects. MATERIALS AND METHODS: The hypoglycemic effect of ACE (100mg/kg, i.g.) was confirmed by testing blood glucose levels or via oral glucose tolerance test (OGTT) in streptozotocin (STZ) induced hyperglycemic mice, db/db diabetic mice and diet-induced obese (DIO) mice. Plasma insulin, GLP-1 levels and intestinal GLP-1 related gene expression were determined in STZ-induced and db/db diabetic mice. The in vitro effects of ACE (12.5µg/ml) on the expression and secretion of GLP-1 were detected in NCI-H716 intestinal L-cells, and the correlation between ACE and molecules in the Wnt signaling pathway was further explored. RESULTS: ACE (100mg/kg) significantly lowered fasting blood glucose in STZ-induced and db/db diabetic mice and improved the OGTT in DIO mice. Insulin releasing and islet protective effects, along with the increased secretion of GLP-1, were observed. The expression of proglucagon gene (gcg) and post-translational processing gene prohormone convertase 3 (pc3) and the GLP-1 content in the culture medium of L-cells notably increased after the ACE treatment (12.5µg/ml). At the same time, ß-catenin nuclear translocation occurred, and its downstream protein cyclin D1 was activated, showing the involvement of Wnt signaling. CONCLUSIONS: ACE might activate Wnt signaling to increase the gene expression of gcg and pc3 and exert incretin effects, including insulinotropic and islet protection, to lower blood glucose levels via elevated GLP-1 secretion either directly or indirectly.

[Quantitative real-time PCR for target periodontal bacteria in subgingival plaque before and after local delivery of periocline, scaling and root planning].

PURPOSE: To compare the copy number of Porphyromonas gingivalis (Pg) and Prevotella intermedia (Pi) in subgingival plaque before and after local delivery of periocline (2% minocycline hydrochloride ointment, MO), scaling and root planning (SRP) by quantitative real-time PCR (qRT-PCR) and evaluate the efficacy. METHODS: Sixty-two adults with moderate to severe chronic periodontitis were selected in the study. Microbial samples were taken from pocket before and after MO and SRP(7d). The samples were evaluated by qRT-PCR for Pg and Pi. Microbiological effectiveness of treatments was assessed using Kruskal-Wallis and Wilcoxon rank-sum test. All tests were two-sided with a significance level of 0.05. All analyses were conducted with SAS 9.1.3 software package. RESULTS: The copy number of Pg and Pi in subgingival plaque was 10(3)-10(6) and 10(2)-10(6). Bacterial loads of Pg were reduced in SPR+ MO, SRP and MO site. The counts of Pi decreased in SRP+ MO sites compared with those in the MO or SRP alone sites significantly (P<0.05). CONCLUSIONS: Quantitative real-time PCR (qRT-PCR) is used as a powerful tool with high sensitivity and specificity to quantitatively assess target periodontal bacteria. The results show that subgingival administration of MO and SRP was effective for reducing pathogenic bacteria and improving clinical outcome. Supported by 2011 National Clinical Specialist Construction Project; Natural Science Foundation of Zhejiang Province(LY13H140002); Education Department Funds of Zhejiang Province(20061258) and Medical General Research Project of Zhejiang Province(2012KYB121).