Uric acid, high density lipoprotein cholesterol levels and their ratio are related to microbial enterotypes and serum metabolites in females with a blood stasis constitution.

BACKGROUND: Blood stasis constitution in traditional Chinese medicine (TCM) is believed to render individuals more susceptible to metabolic diseases. However, the biological underpinnings of this constitutional imbalance remain unclear. METHODS: This study explored the association between blood stasis constitution, serum metabolic markers including uric acid (UA), high-density lipoprotein cholesterol (HDLC), their ratio (UHR), serum metabolites, and gut microbiota. Clinical data, fecal and serum samples were acquired from 24 individuals with a blood stasis constitution and 80 individuals with a balanced constitution among healthy individuals from Guangdong. Gut microbiota composition analysis and serum metabolomics analysis were performed. RESULTS: Females with a blood stasis constitution had higher UA levels, lower HDLC levels, and higher UHR in serum, suggesting a higher risk of metabolic abnormalities. Analysis of the gut microbiome revealed two distinct enterotypes dominated by Bacteroides or Prevotella. Intriguingly, blood stasis subjects were disproportionately clustered within the Bacteroides-rich enterotype. Metabolomic analysis identified subtle differences between the groups, including lower phenylalanine and higher trimethylaminoacetone levels in the blood stasis. Several differential metabolites displayed correlations with HDLC, UA, or UHR, unveiling potential new markers of metabolic dysregulation. CONCLUSIONS: These findings elucidate the intricate interplay between host constitution, gut microbiota, and serum metabolites. The concept of blood stasis offers a unique perspective to identify subtle alterations in microbiome composition and metabolic pathways, potentially signaling underlying metabolic vulnerability, even in the presence of ostensibly healthy profiles. Continued investigation of this TCM principle may reveal critical insights into the early biological processes that foreshadow metabolic deterioration.

Effects of flora deficiency on the structure and function of the large intestine.

The significant anatomical changes in large intestine of germ-free (GF) mice provide excellent material for understanding microbe-host crosstalk. We observed significant differences of GF mice in anatomical and physiological involving in enlarged cecum, thinned mucosal layer and enriched water in cecal content. Furthermore, integration analysis of multi-omics data revealed the associations between the structure of large intestinal mesenchymal cells and the thinning of the mucosal layer. Increased Aqp8 expression in GF mice may contribute to enhanced water secretion or altered hydrodynamics in the cecum. In addition, the proportion of epithelial cells, nutrient absorption capacity, immune function and the metabolome of cecum contents of large intestine were also significantly altered. Together, this is the first systematic study of the transcriptome and metabolome of the cecum and colon of GF mice, and these findings contribute to our understanding of the intricate interactions between microbes and the large intestine.

The impact of Traditional Chinese Medicine on mouse gut microbiota abundances and interactions based on Granger causality and pathway analysis.

Objectives: The intestinal microbiota is essential in absorbing nutrients and defending against pathogens and is associated with various diseases, including obesity, type 2 diabetes, and hypertension. As an alternative medicine, Traditional Chinese Medicine (TCM) has long been used in disease treatment and healthcare, partly because it may mediate gut microbiota. However, the specific effects of TCM on the abundance and interactions of microbiota remain unknown. Moreover, using TCM ingredients and data detailing changes in the abundance of gut microorganisms, we developed bioinformatic methods that decipher the impact of TCM on microorganism interactions. Methods: The dynamics of gut microorganisms affected by TCM treatments is explored using a mouse model, which provided the abundance of 70 microorganisms over time. The Granger causality analysis was used to measure microorganism interactions. Novel "serial connection" and "diverging connection" models were used to identify molecular mechanisms underlying the impact of TCM on gut microorganism interactions, based on microorganism proteins, TCM chemical ingredients, and KEGG reaction equations. Results: Codonopsis pilosula (Dangshen), Cassia twig (Gui Zhi), Radices saussureae (Mu Xiang), and Sijunzi Decoction did not cause an increase in the abundance of harmful microorganisms. Most TCMs decreased the abundance of Bifidobacterium pseudolongum, suggesting a Bifidobacterium pseudolongum supplement should be used during TCM treatment. The Granger causality analysis indicated that TCM treatment changes more than half the interactions between the 70 microorganisms, and "serial connection" and "diverging connection" models suggested that changes in interactions may be related to the reaction number connecting species proteins and TCM ingredients. From a species diversity perspective, a TCM decoction is better than a single herb for healthcare. The Sijunzi Decoction only significantly increased the abundance of Bifidobacterium pseudolongum and did not cause a decrease in the abundance of other species but was found to improve the alpha diversity with the lowest replacement rate. Conclusions: Because most of the nine TCMs are medicinal and edible plants, we expect the methods and results presented can be used to optimize and integrate microbiota and TCMs into healthcare processes. Moreover, as a control study, these results can be combined with future disease mouse models to link variations in species abundance with particular diseases.

Association of the characteristics of the blood metabolome and gut microbiome with the outcome of methotrexate therapy in psoriasis.

Metabolic status and gut microecology are implicated in psoriasis. Methotrexate (MTX) is usually the first-line treatment for this disease. However, the relationship between MTX and host metabolic status and the gut microbiota is unclear. This study aimed to characterize the features of blood metabolome and gut microbiome in patients with psoriasis after treatment with MTX. Serum and stool samples were collected from 15 patients with psoriasis. Untargeted liquid chromatography-mass spectrometry and metagenomics sequencing were applied to profile the blood metabolome and gut microbiome, respectively. We found that the response to MTX varied according to metabolomic and metagenomic features at baseline; for example, patients who had high levels of serum nutrient molecular and more enriched gut microbiota had a poor response. After 16 weeks of MTX, we observed a reduction in microbial activity pathways, and patients with a good response showed more microbial activity and less biosynthesis of serum fatty acid. We also found an association between the serum metabolome and the gut microbiome before intervention with MTX. Carbohydrate metabolism, transporter systems, and protein synthesis within microbes were associated with host metabolic clusters of lipids, benzenoids, and organic acids. These findings suggest that the metabolic status of the blood and the gut microbiome is involved in the effectiveness of MTX in psoriasis, and that inhibition of symbiotic intestinal microbiota may be one of the mechanisms of action of MTX. Prospective studies in larger sample sizes are needed to confirm these findings.

Altered metabolome and microbiome features provide clues in understanding irritable bowel syndrome and depression comorbidity.

Irritable bowel syndrome (IBS) is one of the functional gastrointestinal disorders characterized by chronic and/or recurrent symptoms of abdominal pain and irregular defecation. Changed gut microbiota has been proposed to mediate IBS; however, contradictory results exist, and IBS-specific microbiota, metabolites, and their interactions remain poorly understood. To address this issue, we performed metabolomic and metagenomic profiling of stool and serum samples based on discovery (n = 330) and validation (n = 101) cohorts. Fecal metagenomic data showed moderate dysbiosis compared with other diseases, in contrast, serum metabolites showed significant differences with greater power to distinguish IBS patients from healthy controls. Specifically, 726 differentially abundant serum metabolites were identified, including a cluster of fatty acyl-CoAs enriched in IBS. We further identified 522 robust associations between differentially abundant gut bacteria and fecal metabolites, of which three species including Odoribacter splanchnicus, Escherichia coli, and Ruminococcus gnavus were strongly associated with the low abundance of dihydropteroic acid. Moreover, dysregulated tryptophan/serotonin metabolism was found to be correlated with the severity of IBS depression in both fecal and serum metabolomes, characterized by a shift in tryptophan metabolism towards kynurenine production. Collectively, our study revealed serum/fecal metabolome alterations and their relationship with gut microbiome, highlighted the massive alterations of serum metabolites, which empower to recognize IBS patients, suggested potential roles of metabolic dysregulation in IBS pathogenesis, and offered new clues to understand IBS depression comorbidity. Our study provided a valuable resource for future studies, and would facilitate potential clinical applications of IBS featured microbiota and/or metabolites.

Metagenomic Analysis Reveals the Distribution of Antibiotic Resistance Genes in a Large-Scale Population of Healthy Individuals and Patients With Varied Diseases.

The human gut microbiome is a reservoir for antibiotic resistance gene (ARG). Therefore, characterizing resistome distribution and potential disease markers can help manage antibiotics at the clinical level. While much population-level research has highlighted the strong effect of donor geographic origin on ARG prevalence in the human gut, little is known regarding the effects of other properties, such as age, sex, and disease. Here we employed 2,037 fecal metagenomes from 12 countries. By quantifying the known resistance genes for 24 types of antibiotics in each community, we showed that tetracycline, aminoglycoside, beta-lactam, macrolide-lincosamide-streptogramin (MLS), and vancomycin resistance genes were the dominant ARG types in the human gut. We then compared the ARG profiles of 1427 healthy individuals from the 2,037 samples and observed significant differences across countries. This was consistent with expectations that regional antibiotic usage and exposure in medical and food production contexts affect distribution. Although no specific uniform pattern of ARG was observed, a significant increase in resistance potential among multiple disease groups implied that the disease condition may be another source of ARG variance. In particular, the co-occurrence pattern of some enriched bacterial species and ARGs that were obtained in type 2 diabetes (T2D) and liver cirrhosis patients implied that some disease-associated species may be potential hosts of enriched ARGs, which could be potential biomarkers for the prediction and intervention of such diseases. Overall, our study identifies factors associated with the human gut resistome, including substantial effects of region and heterogeneous effects of disease status, and highlights the value of ARG analysis in disease research and clinical applications.

Built form and depression among the Chinese rural elderly: a cross-sectional study.

OBJECTIVES: Few data on the association between housing structure and depression among rural elders in China are available. We examined the impact of built forms on depression. DESIGN: This is a cross-sectional study. SETTING: A representative sample of rural residents aged 60 years or older in China. PARTICIPANTS: A total of 5090 older adults in 2019 in rural Suzhou, China. OUTCOME MEASURES: Associations of built form with odds of probable and possible depression. RESULTS: There was significant difference among elders living in varied sizes of house. Older age (vs 60-64 years: 75-79 years AdjOR, 1.737; 95% CI, 1.309 to 2.305; ≥80 years AdjOR, 2.072; 95% CI, 1.439 to 2.981), male sex (AdjOR, 0.719; 95% CI, 0.593 to 0.871), single (AdjOR, 1.303; 95% CI, 1.032 to 1.646), self-care disability (AdjOR, 4.761; 95% CI, 3.960 to 5.724), three or more chronic diseases (AdjOR, 2.200; 95% CI, 1.657 to 2.920), living alone (AdjOR, 1.443; 95% CI, 1.059 to 1.966), living in cottage (AdjOR, 1.426; 95% CI, 1.033 to 1.967), living space (vs <50 m2: 201-250 m2 AdjOR, 0.566; 95% CI, 0.359 to 0.893; >250 m2 AdjOR, 0.337; 95% CI, 0.223 to 0.511) and space per person (vs <30 m2: 30- m2 AdjOR, 0.502; 95% CI, 0.362 to 0.697; 40- m2 AdjOR, 0.473; 95% CI, 0.347 to 0.646; 50- m2 AdjOR, 0.418; 95% CI, 0.339 to 0.515) were associated with risk of depression among Chinese rural elders. CONCLUSION: The built form was significantly and meaningfully associated with depression among Chinese rural elders. More attention should be paid to preventing mental illness among the rural elderly living in the small housing area and cottages in China.

A comprehensive investigation of metagenome assembly by linked-read sequencing.

BACKGROUND: The human microbiota are complex systems with important roles in our physiological activities and diseases. Sequencing the microbial genomes in the microbiota can help in our interpretation of their activities. The vast majority of the microbes in the microbiota cannot be isolated for individual sequencing. Current metagenomics practices use short-read sequencing to simultaneously sequence a mixture of microbial genomes. However, these results are in ambiguity during genome assembly, leading to unsatisfactory microbial genome completeness and contig continuity. Linked-read sequencing is able to remove some of these ambiguities by attaching the same barcode to the reads from a long DNA fragment (10-100 kb), thus improving metagenome assembly. However, it is not clear how the choices for several parameters in the use of linked-read sequencing affect the assembly quality. RESULTS: We first examined the effects of read depth (C) on metagenome assembly from linked-reads in simulated data and a mock community. The results showed that C positively correlated with the length of assembled sequences but had little effect on their qualities. The latter observation was corroborated by tests using real data from the human gut microbiome, where C demonstrated minor impact on the sequence quality as well as on the proportion of bins annotated as draft genomes. On the other hand, metagenome assembly quality was susceptible to read depth per fragment (CR) and DNA fragment physical depth (CF). For the same C, deeper CR resulted in more draft genomes while deeper CF improved the quality of the draft genomes. We also found that average fragment length (µFL) had marginal effect on assemblies, while fragments per partition (NF/P) impacted the off-target reads involved in local assembly, namely, lower NF/P values would lead to better assemblies by reducing the ambiguities of the off-target reads. In general, the use of linked-reads improved the assembly for contig N50 when compared to Illumina short-reads, but not when compared to PacBio CCS (circular consensus sequencing) long-reads. CONCLUSIONS: We investigated the influence of linked-read sequencing parameters on metagenome assembly comprehensively. While the quality of genome assembly from linked-reads cannot rival that from PacBio CCS long-reads, the case for using linked-read sequencing remains persuasive due to its low cost and high base-quality. Our study revealed that the probable best practice in using linked-reads for metagenome assembly was to merge the linked-reads from multiple libraries, where each had sufficient CR but a smaller amount of input DNA. Video Abstract.

Risk factors for depressive symptoms among older Chinese adults: A meta-analysis.

BACKGROUND: The study aims to explore the risk factors for depressive symptoms among older Chinese adults. METHODS: PubMed, PsycINFO, Cochrane Library, EMbase, Google Scholar, Chinese National Knowledge Infrastructure (CNKI), Chinese BioMedical Literature Database, and Wanfang data were searched for potentially relevant articles published before September 1st, 2019. Stata/IC 15 was used to perform a meta-analysis and subgroup analysis to compute the pooled odds ratio. RESULTS: The retrieve strategy yielded 11 studies that met the inclusion criteria. The total sample size was 31528 across seven districts, including Mainland China, Hong Kong, and Taiwan. Patients with depression were included in the sample size. Fourteen risk factors were extracted for at least having two or more relative studies. The combined odds ratio ranged from 0.70 to 4.75. Female, poor self-perceived financial condition, single, average and poor self-perceived health status, diabetes, adverse life events, poor social support, two or more numbers of cardiovascular diseases, and functional disability are risk factors of depressive symptoms among older Chinese adults. Fair or good social support is a protective factor. LIMITATIONS: These findings may be somewhat limited by (i) quality of studies included, (ii) a finite number of studies met inclusion criteria. CONCLUSIONS: Despite the methodological limitations of the studies and this meta-analysis, average or poor self-perceived health status, functional disability, poor social support, poor self-perceived financial condition, negative life events, and diabetes appear to be significant risk factors for depressive symptoms among the aged population in China. Social support can mitigate depressive symptoms.

A Clostridia-rich microbiota enhances bile acid excretion in diarrhea-predominant irritable bowel syndrome.

An excess of fecal bile acids (BAs) is thought to be one of the mechanisms for diarrhea-predominant irritable bowel syndrome (IBS-D). However, the factors causing excessive BA excretion remain incompletely studied. Given the importance of gut microbiota in BA metabolism, we hypothesized that gut dysbiosis might contribute to excessive BA excretion in IBS-D. By performing BA-related metabolic and metagenomic analyses in 290 IBS-D patients and 89 healthy volunteers, we found that 24.5% of IBS-D patients exhibited excessive excretion of total BAs and alteration of BA-transforming bacteria in feces. Notably, the increase in Clostridia bacteria (e.g., C. scindens) was positively associated with the levels of fecal BAs and serum 7α-hydroxy-4-cholesten-3-one (C4), but negatively correlated with serum fibroblast growth factor 19 (FGF19) concentration. Furthermore, colonization with Clostridia-rich IBS-D fecal microbiota or C. scindens individually enhanced serum C4 and hepatic conjugated BAs but reduced ileal FGF19 expression in mice. Inhibition of Clostridium species with vancomycin yielded opposite results. Clostridia-derived BAs suppressed the intestinal FGF19 expression in vitro and in vivo. In conclusion, this study demonstrates that the Clostridia-rich microbiota contributes to excessive BA excretion in IBS-D patients, which provides a mechanistic hypothesis with testable clinical implications.

Simultaneous amplification of exons 18 to 21 of the EGFR gene using 5' tailed primers and a two-stage protocol.

OBJECTIVE: Reduction of non-specific amplification and achievement of efficient amplification of multiple gene fragments under the same reaction condition is the basic goal of PCR diagnosis; however, this is often difficult. This study was conducted to establish a highly specific and effective amplification of the epidermal growth factor receptor (EGFR) gene's exons, 18-21, simultaneously. METHODS: The 5'-tailed primers were synthesized by adding 10 to 20 bp of a non-specific sequence to the 5'-terminus of sequence-specific primers (tailless primers). The two-stage protocol consisted of 5-10 cycles of a conventional 3-step cycling, which was then followed by 30-35 cycles of two-step cycling. The exons 18-21 of EGFR gene were amplified in 28 non-small cell lung cancer (NSCLC) patients using an optimized PCR that combined 5' tailed primers with a two-stage protocol. RESULTS: The 5' tailed primers exhibited a wider range of suitable annealing temperatures, similar range of primer concentration, similar sensitivity, specificity, and reproducibility, as well as a reduced, non-specific amplification compared with the corresponding tailless primers. The amplification of exons 18-21 of EGFR gene in NSCLC patients revealed that a combination of 5' tailed primers with two-stage protocol (optimized PCR) had a similar PCR success rate (P = 0.873) but had significantly reduced non-specific amplification (P <0.001) compared to conventional PCR. CONCLUSION: 5' tailed primers exhibited a wider range of suitable annealing temperatures and improved specificity compared with conventional PCR primers. An optimized PCR was established with 5' tailed primers and a two-stage protocol to amplify exons 18-21 of the EGFR gene in NSCLC patients.

TET-Mediated Sequestration of miR-26 Drives EZH2 Expression and Gastric Carcinogenesis.

DNA demethylases of the TET family function as tumor suppressors in various human cancers, but their pathogenic contributions and mechanisms of action in gastric carcinogenesis and progression remain unclear. Here, we report that TET is transcriptionally upregulated in gastric cancer, where it correlates with poor prognosis. Mechanistic investigations revealed that TET facilitated gastric carcinogenesis through a noncoding function of the 3'UTR, which interacted with miR-26. This interaction resulted in sequestration of miR-26 from its target EZH2, which released the suppression on EZH2, and thereby led to EZH2 overexpression in gastric cancer. Our findings uncover a novel noncoding function for TET family proteins in facilitating gastric carcinogenesis. Cancer Res; 77(22); 6069-82. ©2017 AACR.

miR-218 suppresses gastric cancer cell cycle progression through the CDK6/Cyclin D1/E2F1 axis in a feedback loop.

Studies in several cancers have suggested that miR-218 has anti-tumor activities, but its function is yet to be elucidated. In this study, we investigated the regulation and function of miR-218 (miR-218-5p) in the cell cycle progression of gastric cancer (GC). We found that miR-218 could suppress proliferation of gastric cancer cells, induce cell cycle arrest at the G1 phase and inhibit tumor growth and metastasis in vivo. We also demonstrated that miR-218 specifically targeted the 3'-UTR regions of CDK6 and cyclin D1 and inhibited the expression of these molecules, which in turn repressed the pRb/E2F1 signaling pathway. Overexpression of CDK6 and Cyclin D1 reversed miR-218-mediated inhibition of pRB/E2F1 signaling and attenuated the miR-218-induced cell cycle arrest. More importantly, miR-218 expression was significantly reduced and inversely correlated with the levels of CDK6 and Cyclin D1 in gastric cancer tissues. Decreased miR-218 expression was also correlated with advanced clinical stage, lymph node metastasis, and poor prognosis in gastric cancer patients. Furthermore, we showed that miR-218 expression was directly activated by E2F1 through the transactivation of miR-218 host genes, SLIT2 and SLIT3, revealing a negative feedback regulation of miR-218 expression. Taken together, our results describe a regulatory loop miR-218-CDK6/CyclinD1-E2F1 whose disruption may contribute to cell cycle progression in gastric cancer and indicate the potential application of miR-218 in cancer therapy.

Mutation in SSUH2 Causes Autosomal-Dominant Dentin Dysplasia Type I.

Dentin dysplasia type I (DDI) is an autosomal-dominant genetic disorder resulting from dentin defects. The molecular basis of DDI remains unclear. DDI exhibits unique characteristics with phenotypes featuring obliteration of pulp chambers and diminutive root, thus providing a useful model for understanding the genetics of tooth formation. Using a large Chinese family with 14 DDI patients, we mapped the gene locus responsible for DDI to 3p26.1-3p24.3 and further identified a missense mutation, c.353C>A (p.P118Q) in the SSUH2 gene on 3p26.1, which co-segregated with DDI. We showed that SSUH2 (p.P118Q) perturbed the structure and significantly reduced levels of mutant (MT) protein and mRNA compared with wild-type SSUH2. Furthermore, MT P141Q knock-in mice (+/- and -/-) had a unique partial obliteration of the pulp cavity and upregulation or downregulation of six major genes involved in odontogenesis: Dspp, Dmp1, Runx2, Pax9, Bmp2, and Dlx2. The phenotype of missing teeth was determined in zebrafish with morpholino gene knockdowns and rescued by injection of normal human mRNA. Taken together, our observations demonstrate that SSUH2 disrupts dental formation and that this novel gene, together with other odontogenesis genes, is involved in tooth development.

Effects of VBMDMP on the reversal of cisplatin resistance in human lung cancer A549/DDP cells.

Tumor drug resistance is a major obstacle to cancer chemotherapy. We previously constructed a fusion protein based on two tumstatin-derived sequences named recombinant VBMDM (rVBMDMP). We preliminarily confirmed its inhibition of HUVEC and colon cancer cell growth. The present study further systematically observed the inhibitory effect of rVBMDMP on lung cancer cell growth and analyzed a possible mechanism to provide a theoretical basis for the development of new antitumor protein drugs. The effect of rVBMDMP on human lung adenocarcinoma (A549) and cisplatin-resistant human lung adenocarcinoma (A549/DDP) cell proliferation was evaluated by MTS assay. Hoechst 33342 staining performed together with fluorescence microscopy and immunoblot analysis were used to examine the effects of rVBMDMP on the apoptosis of A549/DDP cells. A protein phosphorylation chip was used to identify changes in rVBMDMP-induced signaling protein phosphorylation. Changes in the phosphatidylinositol 3 kinase (PI3K)/Akt signal transduction pathway and expression of multidrug resistance protein (MRP-2)-related molecules following rVBMDMP treatment in A549/DDP cells were evaluated by western blot analysis. A lung cancer xenograft model was used to evaluate the reversal effect of rVBMDMP on drug-resistance of A549/DDP cell tumors to cisplatin in vivo. The results demonstrated that rVBMDMP increased the phosphorylation of 79 signaling proteins, including focal adhesion kinase (FAK), caspase-6, Fas, FasL and FAF1 and downregulated 30 signaling proteins, including integrin αV, integrin ß3, PI3K/Akt, NF-κB and MRP-2 compared with the controls. rVBMDMP also increased the sensitivity of A549 and A549/DDP cells to cisplatin and directly induced apoptosis, which may be related to MRP-2 and Bcl-2 downregulation. The effects of growth inhibition and apoptosis induction of rVBMDMP on A549/DDP cells may be related to the inhibition of integrin αVß3 and PI3K/Akt protein phosphorylation. Finally, we observed an increase in cancer cell sensitivity to cisplatin by rVBMDMP using the A549/DDP cell xenograft model in nude mice. Our study suggests that rVBMDMP may be an effective potential chemotherapy sensitizer and may be a viable drug candidate in anticancer therapies.

Evidence of recent natural selection on the Southeast Asian deletion (--(SEA)) causing α-thalassemia in South China.

BACKGROUND: The Southeast Asian deletion (--(SEA)) is the most commonly observed mutation among diverse α-thalassemia alleles in Southeast Asia and South China. It is generally argued that mutation --(SEA), like other variants causing hemoglobin disorders, is associated with protection against malaria that is endemic in these regions. However, little evidence has been provided to support this claim. RESULTS: We first examined the genetic imprint of recent positive selection on the --(SEA) allele and flanking sequences in the human α-globin cluster, covering a genomic region spanning ~410 kb, by genotyping 28 SNPs in a Chinese population consisting of 76 --(SEA) heterozygotes and 138 normal individuals. The pattern of linkage disequilibrium (LD) and the long-range haplotype test revealed a signature of positive selection. The network of inferred haplotypes suggested a single origin of the --(SEA) allele. CONCLUSIONS: Thus, our data support the hypothesis that the --(SEA) allele has been subjected to recent balancing selection, triggered by malaria.