Systematic Analysis of Impact of Sampling Regions and Storage Methods on Fecal Gut Microbiome and Metabolome Profiles.

The contribution of human gastrointestinal (GI) microbiota and metabolites to host health has recently become much clearer. However, many confounding factors can influence the accuracy of gut microbiome and metabolome studies, resulting in inconsistencies in published results. In this study, we systematically investigated the effects of fecal sampling regions and storage and retrieval conditions on gut microbiome and metabolite profiles from three healthy children. Our analysis indicated that compared to homogenized and snap-frozen samples (standard control [SC]), different sampling regions did not affect microbial community alpha diversity, while a total of 22 of 176 identified metabolites varied significantly across different sampling regions. In contrast, storage conditions significantly influenced the microbiome and metabolome. Short-term room temperature storage had a minimal effect on the microbiome and metabolome profiles. Sample storage in RNALater showed a significant level of variation in both microbiome and metabolome profiles, independent of the storage or retrieval conditions. The effect of RNALater on the metabolome was stronger than the effect on the microbiome, and individual variability between study participants outweighed the effect of RNALater on the microbiome. We conclude that homogenizing stool samples was critical for metabolomic analysis but not necessary for microbiome analysis. Short-term room temperature storage had a minimal effect on the microbiome and metabolome profiles and is recommended for short-term fecal sample storage. In addition, our study indicates that the use of RNALater as a storage medium of stool samples for microbial and metabolomic analyses is not recommended.IMPORTANCE The gastrointestinal microbiome and metabolome can provide a new angle to understand the development of health and disease. Stool samples are most frequently used for large-scale cohort studies. Standardized procedures for stool sample handling and storage can be a determining factor for performing microbiome or metabolome studies. In this study, we focused on the effects of stool sampling regions and stool sample storage conditions on variations in the gut microbiome composition and metabolome profile

Infection and Infertility

Infection is a multifactorial process, which can be induced by a virus, bacterium, or parasite. It may cause many diseases, including obesity, cancer, and infertility. In this chapter, we focus our attention on the association of infection and fertility alteration. Numerous studies have suggested that genetic polymorphisms influencing infection are associated with infertility. So we also review the genetic influence on infection and risk of infertility

Interactions between Exposure to Environmental Polycyclic Aromatic Hydrocarbons and DNA Repair Gene Polymorphisms on Bulky DNA Adducts in Human Sperm

BACKGROUND: Nucleotide excision repair (NER) and base excision repair (BER) are the primary mechanisms for repair of bulky adducts caused by chemical agents, such as PAHs. It is expected that polymorphisms in NER or BER genes may modulate individual susceptibility to PAHs exposure. Here, we evaluate the effects of PAHs exposure and polymorphisms in NER and BER pathway, alone or combined, on polycyclic aromatic hydrocarbon-DNA (PAH-DNA) adducts in human sperm. METHODOLOGY/PRINCIPAL FINDINGS: Sperm PAH-DNA adducts were measured by immunofluorescent assay using flow cytometry in a sample of 465 infertile adults. Polymorphisms of XPA, XPD, ERCC1, XPF, and XRCC1 were determined by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) techniques. The PAHs exposure was detected as urinary 1-hydroxypyrene (1-OHP) levels. In multivariate models adjusted for potential confounders, we observed that XRCC1 5'pUTR -T/C, Arg194Trp, Arg399Gln polymorphisms were associated with increased sperm adduct levels. Furthermore, the stratified analysis indicated that adverse effects of XRCC1 Arg194Trp, Arg399Gln polymorphisms on PAH-DNA adducts were detected only in the high PAHs exposure group. CONCLUSIONS/SIGNIFICANCE: These findings provided the first evidence that polymorphisms of XRCC1 may modify sperm PAH-DNA adduct levels and may be useful biomarkers to identify individuals susceptible to DNA damage resulting from PAHs exposure

SLIT2/ROBO1-miR-218-1-RET/PLAG1: a new disease pathway involved in Hirschsprung\u27s disease.

Hirschsprung\u27s disease (HSCR) is a rare congenital disease caused by impaired proliferation and migration of neural crest cells. We investigated changes in expression of microRNAs (miRNAs) and the genes they regulate in tissues of patients with HSCR. Quantitative real-time PCR and immunoblot analyses were used to measure levels of miRNA, mRNAs, and proteins in colon tissues from 69 patients with HSCR and 49 individuals without HSCR (controls). Direct interactions between miRNAs and specific mRNAs were indentified in vitro, while the function role of miR-218-1 was investigated by using miR-218 transgenic mice. An increased level of miR-218-1 correlated with increased levels of SLIT2 and decreased levels of RET and PLAG1 mRNA and protein. The reductions in RET and PLAG1 by miR-218-1 reduced proliferation and migration of SH-SY5Y cells. Overexpression of the secreted form of SLIT2 inhibited cell migration via binding to its receptor ROBO1. Bowel tissues from miR-218-1 transgenic mice had nerve fibre hyperplasia and reduced numbers of gangliocytes, compared with wild-type mice. Altered miR-218-1 regulation of SLIT2, RET and PLAG1 might be involved in the pathogenesis of HSCR

Mechanism of crocin I on ANIT-induced intrahepatic cholestasis by combined metabolomics and transcriptomics

Background: Intrahepatic cholestasis (IC) is a disorder of bile production, secretion, and excretion with various causes. Crocin I (CR) is effective in the treatment of IC, but its underlying mechanisms need to be further explored. We aimed to reveal the therapeutic mechanism of crocin I for IC by combining an integrated strategy of metabolomics and transcriptomics.Methods: The hepatoprotective effect of CR against cholestasis liver injury induced by α-naphthylisothiocyanate (ANIT) was evaluated in rats. The serum biochemical indices, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bile acid (TBA), total bilirubin (TBIL), direct bilirubin (DBIL), tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), and interleukin 1β (IL-1β), as well as the liver oxidative stress indexes and the pathological characteristics of the liver were analyzed. In addition, we also performed a serum metabolomics study using UPLC-Q Exactive HF-X technology to investigate the effect of CR on the serum of rats with ANIT-induced IC and screened potential biomarkers. The enrichment analysis of differential expressed genes (DEGs) was performed by transcriptomics. Finally, the regulatory targets of CR on potential biomarkers were obtained by combined analysis, and the relevant key targets were verified by western blotting.Results: CR improved serum and liver homogenate indexes and alleviated liver histological injury. Compared with ANIT group, the CR group had 76 differential metabolites, and 10 metabolic pathways were enriched. There were 473 DEGs significantly changed after CR treatment, most of which were enriched in the retinol metabolism, calcium signaling pathway, PPAR signaling pathway, circadian rhythm, chemokine signaling pathway, arachidonic acid metabolism, bile secretion, primary bile acid biosynthesis, and other pathways. By constructing the “compound-reaction-enzyme-gene” interaction network, three potential key-target regulation biomarkers were obtained, including 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR), ATP-binding cassette transporter G5 (ABCG5), and sulfotransferase2A1(SULT2A1), which were further verified by western blotting. Compared with the ANIT group, the CR group significantly increased the expression of ABCG5 and SULT2A1, and the expression of HMGCR significantly decreased.Conclusion: Combined metabolomic and transcriptomic analyses show that CR has a therapeutic effect on IC through regulation of the biosynthesis of bile acids and bilirubin in the bile secretion pathway and regulation of the expression of HMGCR, ABCG5, and SULT2A1