Metaproteomics Characterizes the Human Gingival Crevicular Fluid Microbiome Function in Periodontitis.

Periodontitis is the leading cause of tooth loss in adults worldwide. The human proteome and metaproteome characterization of periodontitis is not clearly understood. Gingival crevicular fluid samples were collected from eight periodontitis and eight healthy subjects. Both the human and microbial proteins were characterized by liquid chromatography coupled with high-resolution mass spectrometry. A total of 570 human proteins were found differentially expressed, which were primarily associated with inflammatory response, cell death, cellular junction, and fatty acid metabolism. For the metaproteome, 51 genera were identified, and 10 genera were found highly expressed in periodontitis, while 11 genera were downregulated. The analysis showed that microbial proteins related to butyrate metabolism were upregulated in periodontitis cases. In particular, correlation analysis showed that the expression of host proteins related to inflammatory response, cell death, cellular junction, and lipid metabolism correlates with the alteration of metaproteins, which reflect the changes of molecular function during the occurrence of periodontitis. This study showed that the gingival crevicular fluid human proteome and metaproteome could reflect the characteristics of periodontitis. This might benefit the understanding of the periodontitis mechanism.

A qualitative and quantitative analysis of the human gingival crevicular fluid proteome and metaproteome.

Gingival crevicular fluid (GCF) is an integral part of oral fluid that plays a special role in maintaining the structure of junctional epithelium and defending against bacterial infection. In this study, we comprehensively analysed the composition of the human GCF proteome and metaproteome simultaneously to obtain multidimensional information about GCF. A total of 3680 human proteins (2540 with at least two unique peptides) were identified in the normal GCF sample, and their functions were mainly associated with immune function and inflammation. Among these proteins, 1874 proteins could be quantified by the iBAQ algorithm, and their abundances spanned a dynamic range of six orders of magnitude. For the GCF metaproteome, a total of 3082 proteins and 69 genera were found. In addition, 16 genera were not identified by GCF metagenomic analysis. Compared to the saliva metaproteome, 32 genera were found to be in common. The protein quantitative analysis showed that the abundance of GCF metaproteome contributed to approximately 4.17% of the total GCF proteome. The top three most abundant genera were Fusobacterium, Corynebacterium, and Leptotrichia. The above data will be useful for future research on GCF-related diseases.

A comparative proteomic analysis to define the influencing factors on gingival crevicular fluid using LC-MS/MS.

Gingival crevicular fluid (GCF) is a promising biofluid for disease identification and biomarker searching in periodontology. This study aimed to investigate the possible influencing factors, including tooth site, sex and age, on the normal GCF proteome. Forty periodontal healthy adults were randomly divided into a training group and a testing group. In the training group, GCF samples from 12 adults were analyzed using the iTRAQ 2D LC-MS/MS method. The influencing factors, tooth site (including periodontitis-susceptible and -insusceptible tooth sites), sex and age, and related differential proteins were defined and functionally annotated. The important differential proteins from 28 adults in the testing group were then validated by PRM analysis. An average of approximately 5 differential proteins were found between tooth sites of periodontitis-susceptible and -insusceptible sites. Eighty-five differentially expressed proteins were obtained between sexes in the young group, while only 7 sex-associated proteins were found in the old group. A total of 203 and 235 age-associated proteins were found in the male and female groups, respectively. The differential protein functional annotation showed that sex-related proteins were mainly related to immune function and metabolism, and age-related proteins were primarily associated with inflammation, lipid metabolism and immune function. In the testing group, a total of 4 sex-related proteins and 12 age-related proteins were validated by PRM analysis. SIGNIFICANCE: The influences of tooth site, sex and age in GCF proteomics in periodontal health were firstly analyzed using LC-MS/MS. Tooth site showed a small influence on the GCF proteome. The sex effect was significant in young adults, but its influence in old adults is small. Age is an important impact factor for the GCF proteome. These findings enrich the knowledge about the normal GCF proteome and might benefit future disease analyses.

Effects of fertilizer practice on fungal and actinobacterial cellulolytic community with different humified particle-size fractions in double-cropping field.

Cellulose plays an important role in maintaining or improving soil carbon (C) cycling and soil fertility of paddy field. There had close relationship between functional cellulose genes (cbhI and GH48) with characterize of soil organic matter chemical components (fulvic acid and humic acid) and soil physical fractions. However, there is still limited information about how functional cellulose degradation response to long-term fertilizer management and their relative importance for C sequestration under the double-cropping rice paddy field in southern of China. Therefore, the objective of this study were investigated the effects of 34-years long-term fertilizer regime on community abundance of cbhI and GH48 genes in five soil particle-size fractions (> 2000 µm, 2000-200 µm, 200-50 µm, 50-2 µm and 2-0.1 µm) by using polarization magic angle spinning 13C nuclear magnetic resonance spectroscopy. The field experiment was included four different fertilizer treatments: chemical fertilizer alone (MF), rice straw and chemical fertilizer (RF), 30% organic manure and 70% chemical fertilizer (OM), and without fertilizer input as a control (CK). The results showed that distribution of soil humus and cellulolytic microbial community abundance was significant increased under long-term application of crop residue and organic manure condition. And the FA, HA and HM C contents in > 2000 µm and 2000-50 µm fractions with MF, RF and OM treatments were significant higher than that of CK treatment. Meanwhile, the alkyl C and Oalkyl C groups of FA and HA in > 2000 µm fraction with MF, RF, OM and CK treatments were higher than that of the other fractions. There had higher AL% and lower ARO% of FA and HA in different particle-size fractions with MF, RF, OM and CK treatments. The results indicated that abundance of cbhI and GH48 genes in different particle-size fractions with RF and OM treatments were significant increased, compared with CK treatment. There had significant positive correlation between soil humus C components (FA and HA) with abundance of cbhI and GH48 genes, and the o-alkyl C and AL% of FA were positively correlated with abundance of cbhI and GH48 genes. As a result, the community abundance of cbhI and GH48 genes were significant increased under combined application of crop residue and organic manure with chemical fertilizer condition.

Comparative proteomic analysis of the influence of gender and acid stimulation on normal human saliva using LC/MS/MS.

PURPOSE: Human saliva is an important source for disease biomarker discovery. This study is to investigate the influence of gender and acid stimulation on the normal human salivary proteome. EXPERIMENTAL DESIGN: Unstimulated and acid-stimulated saliva samples from 5 males and 5 females were labeled with 4-plex iTRAQ and analyzed by 2-DLC MS/MS. By bioinformatics analysis the gender and acid stimulation related proteins were defined. According to protein annotation the important proteins were validated by multiple reaction monitor analysis. RESULTS: A total of 1770 proteins were identified, and 82 proteins in unstimulated saliva were found to be gender-specific, mainly associated with immune function, metabolism and inflammation. However, no gender-specific proteins were found in acid-stimulated saliva. In addition, 182 and 307 differential proteins were found to be acid stimulation-specific in male samples and female samples, respectively, mainly participated in the process of cellular movement, immune function and inflammatory response. Besides, it was found that acid stimulation caused more significant alteration and played a more important role in the human salivary proteome than gender. Gender-specific (IGHG2 and TIMP1) and acid stimulation (PERL, ENOA, ACTB, B4E022 and CALL3) related proteins were validated by MRM analysis. CONCLUSIONS AND CLINICAL RELEVANCE: The results indicate that gender differences exist in the unstimulated salivary proteome, and the influence of acid stimulation on the salivary proteome was more significant than that of gender. The above results may be helpful for salivary proteome research in the future.