Diallyl trisulfide plays an antifibrotic role by inhibiting the expression of Bcl-2 in hepatic stellate cells.

Hepatic fibrosis is an important early stage in the evolution of liver cirrhosis, and specific medicine and therapeutic measures are unavailable to date. Hepatic stellate cells (HSCs) are the main cells involved in the formation of hepatic fibrosis, and induction of the apoptosis of HSCs is an important strategy for the treatment of hepatic fibrosis. Diallyl trisulfide (DATS) is a natural product and is the main active ingredient in garlic. However, the exact molecular mechanisms underlying HSC apoptosis induced by DATS are not well understood. This study aimed to analyze the efficiency and mechanism of DATS in hepatic fibrosis. Different concentrations (25, 50, 100, and 200 µM) of DATS were used to treat HSCs. Changes in cell morphology and formation of apoptotic bodies were observed under an inverted microscope and an electric microscope. Bcl-2 signaling involving Bax, Caspase-3, Caspase-6, Caspase-8, Caspase-9, p53, Apaf-1, and Cyto-c in fibrosis were examined, which is a critical step in the evaluation of antihepatic fibrosis agents. We also evaluated the effect of DATS on the cellular morphology of HSCs and apoptosis-related factors under different Bcl-2 expression states. Our results suggest that DATS regulates hepatic fibrosis by blocking the Bcl-2 signaling pathway and upregulating the Bax/Bcl-2 ratio.

Transcriptome analysis reveals dynamic changes in coxsackievirus A16 infected HEK 293T cells.

BACKGROUND: Coxsackievirus A16 (CVA16) and enterovirus 71 (EV71) are two of the major causes of hand, foot and mouth disease (HFMD) world-wide. Although many studies have focused on infection and pathogenic mechanisms, the transcriptome profile of the host cell upon CVA16 infection is still largely unknown. RESULTS: In this study, we compared the mRNA and miRNA expression profiles of human embryonic kidney 293T cells infected and non-infected with CVA16. We highlighted that the transcription of SCARB2, a cellular receptor for both CVA16 and EV71, was up-regulated by nearly 10-fold in infected cells compared to non-infected cells. The up-regulation of SCARB2 transcription induced by CVA16 may increase the possibility of subsequent infection of CVA16/EV71, resulting in the co-infection with two viruses in a single cell. This explanation would partly account for the co-circulation and genetic recombination of a great number of EV71 and CVA16 viruses. Based on correlation analysis of miRNAs and genes, we speculated that the high expression of SCARB2 is modulated by down-regulation of miRNA has-miR-3605-5p. At the same time, we found that differentially expressed miRNA target genes were mainly reflected in the extracellular membrane (ECM)-receptor interaction and circadian rhythm pathways, which may be related to clinical symptoms of patients infected with CVA16, such as aphthous ulcers, cough, myocarditis, somnolence and potentially meningoencephalitis. The miRNAs hsa-miR-149-3p and hsa-miR-5001-5p may result in up-regulation of genes in these morbigenous pathways related to CVA16 and further cause clinical symptoms. CONCLUSIONS: The present study elucidated the changes in 293T cells upon CVA16 infection at transcriptome level, containing highly up-regulated SCARB2 and genes in ECM-receptor interaction and circadian rhythm pathways, and key miRNAs in gene expression regulation. These results provided novel insight into the pathogenesis of HFMD induced by CVA16 infection.

Alterations of Gut Microbiota in Pyogenic Liver Abscess Patients with and without Type 2 Diabetes Mellitus.

Purpose: The clinical manifestations of pyogenic liver abscess (PLA) vary between patients with and without diabetes mellitus (DM). However, the relationship between PLA and the gut microbiome remains unknown. This study analyzed the composition of gut microbiota in PLA patients with and without DM and healthy controls (HCs) with the goal of identifying potential reasons for the observed variations in clinical manifestations. Patients and Methods: Using 16S ribosomal RNA(16S rRNA) gene sequencing, we analyzed the compositions of gut microbiota in 32 PLA patients with DM, 32 PLA patients without DM, and 29 matched HCs. Results: In PLA patients with DM, the D-dimer level, fibrinogen degradation products, and thrombin time were significantly higher compared to the PLA patients without DM (P < 0.05). The abundance and diversity of intestinal flora were reduced in both groups of PLA patients compared with the HCs (P < 0.05). Specifically, the PLA patients with DM showed significant decreases in the relative abundances of Bacteroides, Blautia, Prevotella9, and Faecalibacterium, whereas Enterococcus and Escherichia-Shigella were relatively more abundant (P < 0.05). Compared to PLA patients without DM, those with DM had lower relative abundances of Lactobacillus and Klebsiella (P < 0.05) and showed different bacterial flora, including Anaerosporobacter and Megamonas. Conclusion: PLA patients with DM exhibited more severe clinical manifestations of PLA compared to patients without DM. It is important to monitor blood coagulation in PLA patients with DM to prevent the development of thrombotic diseases. Additionally, PLA patients with DM exhibit distinct differences in the composition and diversity of their intestinal flora compared to both PLA patients without DM and HCs.

Role of intestinal flora in the development of nonalcoholic fatty liver disease in children.

In China, 45% of adolescents with obesity develop fatty liver disease, a condition that increases the long-term risk of developing cirrhosis and liver cancer. Although the factors triggering nonalcoholic fatty liver disease (NAFLD) vary in children, the composition of intestinal microflora has been found to play an increasingly important role. However, evidence is limited on the prevalence of nonalcoholic fatty liver (NAFL) and nonalcoholic steatohepatitis (NASH) in Chinese children. Therefore, this study aimed to evaluate the fecal microbiome of Chinese children with NAFLD and further analyze the potential of flora in regulating NAFLD-related symptoms and metabolic functions. Specifically, the study applied a 16S rRNA and metagenomic sequencing to the fecal samples of pediatric patients with NAFLD, NASH, and NAFL, as well as healthy controls, to explore the correlation among NAFLD-related indexes, metabolic pathways, and gut flora. The findings showed that some fecal microbiota had a negative correlation with body mass index, and various NAFLD-related bacteria, including Lachnoclostridium, Escherichia-Shigella, and Faecalibacterium prausnitzii, were detected. Consequently, the study concluded that the variation in gut microbiota might be more important in improving NAFLD/NASH compared with single species, providing a microbiota diagnostic profile of NAFLD/NASH.IMPORTANCEThis study aims to characterize the gut microbiota in Chinese children with nonalcoholic fatty liver disease (NAFLD) through 16S rRNA and metagenomic sequencing. The results highlight the association between fecal microbiota and NAFLD in Chinese children, demonstrating distinct characteristics compared to adults and children from other countries. Based on the sequencing data from our cohort's fecal samples, we propose a microbiota model with a high area under the curve for distinguishing between NAFLD and healthy individuals. Furthermore, our follow-up study reveals that changes in the relative abundance of microbial biomarkers in this model are consistent with variations in patients' body mass index. These findings suggest the potential utility of the microbiota model and microbial biomarkers for diagnosing and treating NAFLD in children.

Gut microbiome and mycobiome in inflammatory bowel disease patients with Clostridioides difficile infection.

Background: Clostridium difficile infection (CDI) is common in patients with inflammatory bowel disease (IBD) and has been reported as a risk factor for poor outcome. However, gut microbiome and mycobiome of IBD patients with CDI have been barely investigated. This study aimed to assess the gut microbiome and mycobiome in IBD patients with CDI. Methods: We collected fecal samples from patients with active IBD and concomitant CDI (IBD-CDI group, n=25), patients with active IBD and no CDI (IBD-only group, n=51), and healthy subjects (HC, n=40). Patients' characteristics including demographic data, disease severity, and medication history were collected. Metagenomic sequencing, taxonomic and functional analysis were carried out in the samples. Results: We found that the bacterial alpha diversity of the IBD-CDI group was decreased. The bacterial and fungal beta diversity variations between IBD patients and HC were significant, regardless of CDI status. But the IBD-CDI group did not significantly cluster separately from the IBD-only group. Several bacterial taxa, including Enterococcus faecium, Ruminococcus gnavus, and Clostridium innocuum were overrepresented in the IBD-CDI group. Furthermore, IBD patients with CDI were distinguished by several fungal taxa, including overrepresentation of Saccharomyces cerevisiae. We also identified functional differences in IBD patients with CDI include enrichment of peptidoglycan biosynthesis. The network analysis indicated specific interactions between microbial markers in IBD-CDI patients. Conclusion: IBD patients with CDI had pronounced microbial dysbiosis. Gut micro-ecological changes in IBD patients with CDI might provide insight into the pathological process and potential strategies for diagnosis and treatment in this subset of patients.

Human functional genetic studies are biased against the medically most relevant primate-specific genes.

BACKGROUND: Many functional, structural and evolutionary features of human genes have been observed to correlate with expression breadth and/or gene age. Here, we systematically explore these correlations. RESULTS: Gene age and expression breadth are strongly correlated, but contribute independently to the variation of functional, structural and evolutionary features, even when we take account of variation in mRNA expression level. Human genes without orthologs in distant species ('young' genes) tend to be tissue-specific in their expression. As computational inference of gene function often relies on the existence of homologs in other species, and experimental characterization is facilitated by broad and high expression, young, tissue-specific human genes are often the least characterized. At the same time, young genes are most likely to be medically relevant. CONCLUSIONS: Our results indicate that functional characterization of human genes is biased against young, tissue-specific genes that are mostly medically relevant. The biases should not be taken lightly because they may pose serious obstacles to our understanding of the molecular basis of human diseases. Future studies should thus be designed to specifically explore the properties of primate-specific genes.

A gene catalogue for post-diapause development of an anhydrobiotic arthropod Artemia franciscana.

BACKGROUND: Diapause is a reversible state of developmental suspension and found among diverse taxa, from plants to animals, including marsupials and some other mammals. Although previous work has accumulated ample data, the molecular mechanism underlying diapause and reactivation from it remain elusive. RESULTS: Using Artemia franciscana, a model organism to study the development of post-diapause embryos in Arthropod, we sequenced random clones up to a total of 28,039 ESTs from four cDNA libraries made from dehydrated cysts and three time points after rehydration/reactivation, which were assembled into 8,018 unigene clusters. We identified 324 differentially-expressed genes (DEGs, P < 0.05) based on pairwise comparisons of the four cDNA libraries. We identified a group of genes that are involved in an anti-water-deficit system, including proteases, protease inhibitors, heat shock proteins, and several novel members of the late embryogenesis abundant (LEA) protein family. In addition, we classified most of the up-regulated genes after cyst reactivation into metabolism, biosynthesis, transcription, and translation, and this result is consistent with the rapid development of the embryo. Some of the specific expressions of DEGs were confirmed experimentally based on quantitative real-time PCR. CONCLUSION: We found that the first 5-hour period after rehydration is most important for embryonic reactivation of Artemia. As the total number of expressed genes increases significantly, the majority of DEGs were also identified in this period, including a group of water-deficient-induced genes. A group of genes with similar functions have been described in plant seeds; for instance, one of the novel LEA members shares ~70% amino-acid identity with an Arabidopsis EM (embryonic abundant) protein, the closest animal relative to plant LEA families identified thus far. Our findings also suggested that not only nutrition, but also mRNAs are produced and stored during cyst formation to support rapid development after reactivation.

Proteomic profiling of rice embryos from a hybrid rice cultivar and its parental lines.

Elite rice hybrids, when compared to their parental lines, exhibit increased yield and other favorable agronomical traits, such as pathogen- and water-stress resistances, which are described as heterosis, and the molecular mechanism of heterosis remains to be elucidated. Since genomic sequences of the paternal (9311) and maternal lines (P64S) of a major rice hybrid variety LYP9 (Liang-You-Pei-Jiu) were acquired recently, we performed a proteomic study on mature embryos of this hybrid triad based on 2-DE and MALDI-TOF MS analyses, and identified 54 differentially expressed proteins involved in major biological processes including nutrient reservoir, response to stress, and metabolism. We observed that most of the storage proteins exhibit overdominance and stress-induced proteins display additivity. We compared proteomic results with transcriptomic data generated from the same embryo samples and found 28 candidate heterosis-associated genes shared by the two datasets. We further traced back to their genomic structures including protein-coding and regulatory regions and found that most of these genes have multiple copies in rice genomes as paralogous genes. Based on alignment of coding and regulation regions, we found that most of the differentially expressed genes at both protein and RNA levels are recent gene duplicates (paralogous genes) with relative little difference in protein-coding regions between orthologous genes (between genes of the two parental genomes) as compared to regulatory regions that harbor numerous indels and base substitutions.

Transcriptomic profiling of mature embryo from an elite super-hybrid rice LYP9 and its parental lines.

BACKGROUND: The mature embryo of rice (Oryza sativa, L.) is a synchronized and integrated tissue mass laying the foundation at molecular level for its growth, development, and differentiation toward a developing and ultimately a mature plant. We carried out an EST (expressed-sequence-tags)-based transcriptomic study, aiming at gaining molecular insights into embryonic development of a rice hybrid triad-an elite hybrid rice LYP9 and its parental lines (93-11 and PA64s)-and possible relatedness to heterosis. RESULTS: We generated 27,566 high-quality ESTs from cDNA libraries made from mature rice embryos. We classified these ESTs into 7,557 unigenes (2,511 contigs and 5,046 singletons) and 7,250 (95.9%) of them were annotated. We noticed that the high-abundance genes in mature rice embryos belong to two major functional categories, stress-tolerance and preparation-for-development, and we also identified 191 differentially-expressed genes (General Chi-squared test, P-value <= 0.05) between LYP9 and its parental lines, representing typical expression patterns including over-dominance, high- and low-parent dominance, additivity, and under-dominance. In LYP9, the majority of embryo-associated genes were found not only abundantly and specifically enriched but also significantly up-regulated. CONCLUSION: Our results suggested that massively strengthening tissue-(or stage-) characteristic functions may contribute to heterosis rather than a few simple mechanistic explanations at the individual gene level. In addition, the large collection of rice embryonic ESTs provides significant amount of data for future comparative analyses on plant development, especially for the important crops of the grass family.

FTO-dependent demethylation of N6-methyladenosine regulates mRNA splicing and is required for adipogenesis.

The role of Fat Mass and Obesity-associated protein (FTO) and its substrate N6-methyladenosine (m6A) in mRNA processing and adipogenesis remains largely unknown. We show that FTO expression and m6A levels are inversely correlated during adipogenesis. FTO depletion blocks differentiation and only catalytically active FTO restores adipogenesis. Transcriptome analyses in combination with m6A-seq revealed that gene expression and mRNA splicing of grouped genes are regulated by FTO. M6A is enriched in exonic regions flanking 5'- and 3'-splice sites, spatially overlapping with mRNA splicing regulatory serine/arginine-rich (SR) protein exonic splicing enhancer binding regions. Enhanced levels of m6A in response to FTO depletion promotes the RNA binding ability of SRSF2 protein, leading to increased inclusion of target exons. FTO controls exonic splicing of adipogenic regulatory factor RUNX1T1 by regulating m6A levels around splice sites and thereby modulates differentiation. These findings provide compelling evidence that FTO-dependent m6A demethylation functions as a novel regulatory mechanism of RNA processing and plays a critical role in the regulation of adipogenesis.

Diverse LEA (late embryogenesis abundant) and LEA-like genes and their responses to hypersaline stress in post-diapause embryonic development of Artemia franciscana.

From post-diapause cysts of Artemia franciscana, we defined fourteen LEA (late embryogenesis abundant) and LEA-like genes, including four novel members (Afrlea1-5, Afrlea3-5, Afrlea3-like1 and Afrlea3-like2), which were classified into four groups: G1, G3, G3-like (LEA group3-like), and SMP-like (seed-maturation-protein-like), based on their conserved and diversified sequence motifs and amino acid compositions among bacteria, plants, and animals. We also validated six representative genes based on quantitative RT-PCR, including three LEA and two LEA3-like genes that are down-regulated when dehydrated cysts hatch to desiccation-intolerant larvae as well as one SMP-like gene that is slightly up-regulated. We further tested their responses to hypersaline stress for four representatives-one from each group-and found that the expression of Afrlea1-5 and Afrlea3-2 were inducible but not Afrlea3-like1 and Afrsmp-like. This result suggested that the LEA and LEA-like genes may play different roles in resistance to hypersaline stress.

Sequence Variation and Expression Analysis of Seed Dormancy- and Germination-Associated ABA- and GA-Related Genes in Rice Cultivars.

Abscisic acid (ABA) and Gibberellic acid (GA) play key roles in regulating seed dormancy and germination. First, when examining germination of different rice cultivars, we found that their germination timing and dormancy status are rather distinct, coupled with different GA/ABA ratio. Second, we studied genomic sequences of ABA and GA dormancy- and germination-associated genes in rice and discovered single nucleotide polymorphisms and insertions/deletions (Indels) in both coding and regulatory sequences. We aligned all these variations to the genome assemblies of 9311 and PA64s and demonstrated their relevance to seed dormancy both quantitatively and qualitatively based on gene expression data. Third, we surveyed and compared differentially expressed genes in dry seeds between 9311 and PA64s to show that these differentially expressed genes may play roles in seed dormancy and germination.

HRGD: a database for mining potential heterosis-related genes in plants.

Heterosis-Related Gene Database (HRGD) is designed to manage the output of comparative transcriptomic studies related to heterosis (or hybrid vigor) among major agricultural crops, providing publicly available query and analysis platform for practical data mining. The database contains information concerning over 5,000 differentially expressed genes (DEGs) among the hybrid-parent tissue panels from rice genomes generated in our institute and other genomes of major cereal crops collected from published literatures. We have annotated relevant genes from manually extracted information includes not only gene sequences, genomic structures, and functional annotations but also empirical expression data generated based on various large-scale genomic methods. Tools for visualization and functional analysis of DEGs from various hybrid-parent tissue panels are also integrated via a user-friendly web interface. HRGD is a useful tool for helping plant biologists and crop breeders to explore biological knowledge that is being generated on a daily basis and in a large scale. The HRGD data resource is publicly available at http://hrgd.genomics.org.cn/ .

A proteomic study on postdiapaused embryonic development of brine shrimp (Artemia franciscana).

Encysted gastrula of brine shrimp (Artemia, Crustacea, and Anostraca) provides an excellent model for studying molecular processes of diapause. We report a proteomic study on early molecular responses of Artemia's postdiapaused cysts and found that dehydrated cysts actually store more proteins, in both kind and amount, than developing cysts. We identified 75 differentially expressed proteins over a course of cyst development, and also exploited PTMs of dehydrate cysts. We further surveyed gene expression of postdiapaused cysts in early developmental phases in a 0.5 h interval up to the seventh hour, and discovered that the activation of cellular activities is ignited as early as 0.5 h after rehydration. We traced nine differentially expressed proteins (COXI, COXIII, heat shock proteins (HSP26, HSP60, and HSP70), CDC48, late embryogenesis abundant (LEA), GS1-like protein, and cathepsin L-associated protein (CLAP)) for quantitative transcriptional changes, monitored by real-time PCR, and found these proteins exhibiting distinct expression patterns that suggest complex gene regulations for cyst reactivation after diapause breakage. Future experiments should be designed to focus on early activation concerning signal transduction, energy generation, and PTMs.