Reconstruction of the genome-scale metabolic network model of Sinorhizobium fredii CCBAU45436 for free-living and symbiotic states.

Sinorhizobium fredii CCBAU45436 is an excellent rhizobium that plays an important role in agricultural production. However, there still needs more comprehensive understanding of the metabolic system of S. fredii CCBAU45436, which hinders its application in agriculture. Therefore, based on the first-generation metabolic model iCC541 we developed a new genome-scale metabolic model iAQY970, which contains 970 genes, 1,052 reactions, 942 metabolites and is scored 89% in the MEMOTE test. Cell growth phenotype predicted by iAQY970 is 81.7% consistent with the experimental data. The results of mapping the proteome data under free-living and symbiosis conditions to the model showed that the biomass production rate in the logarithmic phase was faster than that in the stable phase, and the nitrogen fixation efficiency of rhizobia parasitized in cultivated soybean was higher than that in wild-type soybean, which was consistent with the actual situation. In the symbiotic condition, there are 184 genes that would affect growth, of which 94 are essential; In the free-living condition, there are 143 genes that influence growth, of which 78 are essential. Among them, 86 of the 94 essential genes in the symbiotic condition were consistent with the prediction of iCC541, and 44 essential genes were confirmed by literature information; meanwhile, 30 genes were identified by DEG and 33 genes were identified by Geptop. In addition, we extracted four key nitrogen fixation modules from the model and predicted that sulfite reductase (EC 1.8.7.1) and nitrogenase (EC 1.18.6.1) as the target enzymes to enhance nitrogen fixation by MOMA, which provided a potential focus for strain optimization. Through the comprehensive metabolic model, we can better understand the metabolic capabilities of S. fredii CCBAU45436 and make full use of it in the future.

A carboxymethyl chitosan/oxidized hyaluronic acid composite hydrogel dressing loading with stem cell exosome for chronic inflammation wounds healing.

Stem cell exosomes (Exo) play an important role in the transformation of macrophages, but the rapid clearance of Exo in vivo limits their therapeutic effects for chronic inflammation wounds healing. Here, stem cell Exo was isolated and introduced to a composite hydrogel including carboxymethyl chitosan (CMCS) and oxidized hyaluronic acid (OHA) through chemical cross-linking, which formed an Exo-loaded (CMCS/OHA/Exo) hydrogel. The CMCS/OHA/Exo hydrogel exhibited a function of Exo sustained release and an Exo protection within 6 days. This CMCS/OHA/Exo hydrogel was much better than CMCS/OHA hydrogel or Exo solution in macrophage cell phagocytosis, proliferation and migration in vitro, especially, played an obviously positive role in the transformation of macrophages compared with the reference groups. For the treatment of the chronic inflammation wounds in vivo, the CMCS/OHA/Exo hydrogel had the best results at wound heal rate and inhibiting the secretion of inflammatory factors, and it was far superior to reference groups in wound re-epithelization and collagen production. CMCS/OHA/Exo hydrogels can promote Exo release based on hydrogel degradation to regulate macrophages transformation and accelerate chronic wound healing. The study offers a method for preparing Exo-loaded hydrogels that effectively promote the transformation of macrophages and accelerate chronic inflammatory wound healing.

Construction and functional evaluation of oral long-acting insulin hydrogel microparticles based on physical and chemical double crosslinking.

The objective of this study was to enhance the convenience and effectiveness of diabetes treatment by developing hydrogel microparticles as an oral insulin delivery system, aiming to reduce the necessity for frequent treatments. The hydrogel microparticles were prepared with polysaccharides through a combination of physical and chemical crosslinking method, they achieved good results in insulin loading efficiency (70 %), insulin release efficiency (98 %) and sustained release time (>20 h). The effective transmembrane transport was validated using an intestinal epithelial cell model, which demonstrated a continuous hypoglycemic effect lasting from 6 to 26 h in a type 2 diabetes mouse model. Additionally, the relative bioavailability of insulin reached 30.14 ± 2.62 %, representing a significant breakthrough in the field of oral insulin delivery carriers. Furthermore, oral insulin hydrogel exhibited a substantial improvement in insulin resistance, organ damage, and diabetes-related complications stemming from hyperglycemia. These compelling findings underscore the potential of hydrogel microparticles as a cost-effective and valuable strategy for oral drug delivery in diabetes treatment.

GMMAD: a comprehensive database of human gut microbial metabolite associations with diseases.

BACKGROUND: The natural products, metabolites, of gut microbes are crucial effect factors on diseases. Comprehensive identification and annotation of relationships among disease, metabolites, and microbes can provide efficient and targeted solutions towards understanding the mechanism of complex disease and development of new markers and drugs. RESULTS: We developed Gut Microbial Metabolite Association with Disease (GMMAD), a manually curated database of associations among human diseases, gut microbes, and metabolites of gut microbes. Here, this initial release (i) contains 3,836 disease-microbe associations and 879,263 microbe-metabolite associations, which were extracted from literatures and available resources and then experienced our manual curation; (ii) defines an association strength score and a confidence score. With these two scores, GMMAD predicted 220,690 disease-metabolite associations, where the metabolites all belong to the gut microbes. We think that the positive effective (with both scores higher than suggested thresholds) associations will help identify disease marker and understand the pathogenic mechanism from the sense of gut microbes. The negative effective associations would be taken as biomarkers and have the potential as drug candidates. Literature proofs supported our proposal with experimental consistence; (iii) provides a user-friendly web interface that allows users to browse, search, and download information on associations among diseases, metabolites, and microbes. The resource is freely available at http://guolab.whu.edu.cn/GMMAD . CONCLUSIONS: As the online-available unique resource for gut microbial metabolite-disease associations, GMMAD is helpful for researchers to explore mechanisms of disease- metabolite-microbe and screen the drug and marker candidates for different diseases.

hUC-MSC transplantation therapy effects on lupus-prone MRL/lpr mice at early disease stages.

BACKGROUND: The efficacy of human umbilical cord mesenchymal stem cell (hUC-MSC) transplantation in treating systemic lupus erythematosus (SLE) has been confirmed by small-scale clinical trials. However, these trials focused on severe or refractory SLE, while few studies focused on mild SLE. Therefore, this study focused on the therapeutic effects of hUC-MSC transplantation in early-stage or mild MRL/lpr lupus model mice. METHODS: Commercially available hUC-MSCs were transplanted into 8-week-old MRL/lpr mice by tail vein injection. Flow cytometry was used to analyze B cells and their subsets in the peripheral blood. Further, plasma inflammatory factors, autoantibodies, and plasma biochemical indices were detected using protein chip technology and ELISA kits. In addition, pathological staining and immunofluorescence were performed to detect kidney injury in mice. RESULTS: hUC-MSC transplantation did not affect the mice's body weight, and both middle and high dose hUC-MSC transplantation (MD and HD group) actually reduced spleen weight. hUC-MSC transplantation significantly decreased the proportion of plasmablasts (PB), IgG1- PB, IgG1+ PB, IgG1+ memory B (MB) cells, IgG1+ DN MB, and IgG1+ SP MB cells. The hUC-MSC transplantation had significantly reduced plasma levels of inflammatory factors, such as TNF-α, IFN-γ, IL-6, and IL-13. Pathological staining showed that the infiltration of glomerular inflammatory cells was significantly reduced and that the level of glomerular fibrosis was significantly alleviated in hUC-MSC-transplanted mice. Immunofluorescence assays showed that the deposition of IgG and IgM antibodies in the kidneys of hUC-MSC-transplanted mice was significantly lower than in the control. CONCLUSION: hUC-MSC transplantation could inhibit the proliferation and differentiation of peripheral blood B cells in the early-stage of MRL/lpr mice, thereby alleviating the plasma inflammatory environment in mice, leading to kidney injury remission. The study provides a new and feasible strategy for SLE treatment.

Mslar: Microbial synthetic lethal and rescue database.

Synthetic lethality (SL) occurs when mutations in two genes together lead to cell or organism death, while a single mutation in either gene does not have a significant impact. This concept can also be extended to three or more genes for SL. Computational and experimental methods have been developed to predict and verify SL gene pairs, especially for yeast and Escherichia coli. However, there is currently a lack of a specialized platform to collect microbial SL gene pairs. Therefore, we designed a synthetic interaction database for microbial genetics that collects 13,313 SL and 2,994 Synthetic Rescue (SR) gene pairs that are reported in the literature, as well as 86,981 putative SL pairs got through homologous transfer method in 281 bacterial genomes. Our database website provides multiple functions such as search, browse, visualization, and Blast. Based on the SL interaction data in the S. cerevisiae, we review the issue of duplications' essentiality and observed that the duplicated genes and singletons have a similar ratio of being essential when we consider both individual and SL. The Microbial Synthetic Lethal and Rescue Database (Mslar) is expected to be a useful reference resource for researchers interested in the SL and SR genes of microorganisms. Mslar is open freely to everyone and available on the web at http://guolab.whu.edu.cn/Mslar/.

Essential Oil Nanoemulsion Hydrogel with Anti-Biofilm Activity for the Treatment of Infected Wounds.

The formation of a bacterial biofilm on an infected wound can impede drug penetration and greatly thwart the healing process. Thus, it is essential to develop a wound dressing that can inhibit the growth of and remove biofilms, facilitating the healing of infected wounds. In this study, optimized eucalyptus essential oil nanoemulsions (EEO NEs) were prepared from eucalyptus essential oil, Tween 80, anhydrous ethanol, and water. Afterward, they were combined with a hydrogel matrix physically cross-linked with Carbomer 940 (CBM) and carboxymethyl chitosan (CMC) to prepare eucalyptus essential oil nanoemulsion hydrogels (CBM/CMC/EEO NE). The physical-chemical properties, in vitro bacterial inhibition, and biocompatibility of EEO NE and CBM/CMC/EEO NE were extensively investigated and the infected wound models were proposed to validate the in vivo therapeutic efficacy of CBM/CMC/EEO NE. The results showed that the average particle size of EEO NE was 15.34 ± 3.77 nm with PDI ˂ 0.2, the minimum inhibitory concentration (MIC) of EEO NE was 15 mg/mL, and the minimum bactericidal concentration (MBC) against S. aureus was 25 mg/mL. The inhibition and clearance of EEO NE against S. aureus biofilm at 2×MIC concentrations were 77.530 ± 7.292% and 60.700 ± 3.341%, respectively, demonstrating high anti-biofilm activity in vitro. CBM/CMC/EEO NE exhibited good rheology, water retention, porosity, water vapor permeability, and biocompatibility, meeting the requirements for trauma dressings. In vivo experiments revealed that CBM/CMC/EEO NE effectively promoted wound healing, reduced the bacterial load of wounds, and accelerated the recovery of epidermal and dermal tissue cells. Moreover, CBM/CMC/EEO NE significantly down-regulated the expression of two inflammatory factors, IL-6 and TNF-α, and up-regulated three growth-promoting factors, TGF-ß1, VEGF, and EGF. Thus, the CBM/CMC/EEO NE hydrogel effectively treated wounds infected with S. aureus, enhancing the healing process. It is expected to be a new clinical alternative for healing infected wounds in the future.

A Schiff base hydrogel dressing loading extracts from Periplaneta Americana for diabetic wound healing.

As a common complication of diabetic patients, the chronic wound of diabetes has a high incidence, expensive treatment, and recurrence probability, which causes long-term negative impacts on patients' daily life. In this study, the hydrogel was formed by Schiff base reaction between oxidized hyaluronic acid (OHA) and carboxymethyl chitosan (CMCS), and the composite hydrogel dressing was prepared by adding the active polypeptides extract of Periplaneta Americana (PAE). By mass spectrometer determined, PAE mainly includes vitellogenins that can trigger an immune response. The composite hydrogel has good swelling properties, proper fluidity, and a regular 3D network structure. The hydrogel has good cytocompatibility and can promote cell proliferation by L929 fibroblast assay. Finally, it was used to evaluate the effect of diabetic wound repair. The results showed that it could effectively promote wound healing, promote tissue and vascular regeneration, inhibit inflammatory factors, and promote the expression of growth factors. The OHA/CMCS/PAE hydrogels would be promising candidates for chronic wound healing applications.

Prediction of protein-coding small ORFs in multi-species using integrated sequence-derived features and the random forest model.

Proteins encoded by small open reading frames (sORFs) can serve as functional elements playing important roles in vivo. Such sORFs also constitute the potential pool for facilitating the de novo gene birth, driving evolutionary innovation and species diversity. Therefore, their theoretical and experimental identification has become a critical issue. Herein, we proposed a protein-coding sORFs prediction method merely based on integrative sequence-derived features. Our prediction performance is better or comparable compared with other nine prevalent methods, which shows that our method can provide a relatively reliable research tool for the prediction of protein-coding sORFs. Our method allows users to estimate the potential expression of a queried sORF, which has been demonstrated by the correlation analysis between our possibility estimation and codon adaption index (CAI). Based on the features that we used, we demonstrated that the sequence features of the protein-coding sORFs in the two domains have significant differences implying that it might be a relatively hard task in terms of cross-domain prediction, hence domain-specific models were developed, which allowed users to predict protein-coding sORFs both in eukaryotes and prokaryotes. Finally, a web-server was developed and provided to boost and facilitate the study of the related field, which is freely available at http://guolab.whu.edu.cn/codingCapacity/index.html.

A composite hydrogel scaffold based on collagen and carboxymethyl chitosan for cartilage regeneration through one-step chemical crosslinking.

The number of cases of cartilage damage worldwide is increasing annually and this problem severely limits an individual's physical activities, subsequently contributing to additional medical problems. Hydrogels can repair cartilage defects and promote cartilage regeneration. In this study, a composite hydrogel scaffold was prepared with collagen (COL), carboxymethyl chitosan (CMC), and the Arg-Gly-Asp (RGD) peptide through one-step chemical crosslinking, in which the three compositions ratio was especially investigated. The hydrogel scaffold performed well in cell adhesion and biocompatibility experiments, mainly due to the favorable porosity (the aperture was concentrated at 100 µm and the porosity was >70 %) and RGD concentration (2 mM RGD was the optimal concentration, which could effectively improve the attachment of BMSCs to the stent). Moreover, bone marrow mesenchymal stem cells (BMSCs) filled in the hydrogel scaffold, together with transforming growth factor TGF-ß3, which was applied to evaluate the feasibility on the repair of the injured cartilage of the rat. In vitro and in vivo study, according to the results of cell proliferation and cytotoxicity, the hydrogel material had no toxic effect on cells, and the COL2/CMC1 hydrogel scaffold had the most obvious role in promoting cell proliferation. The results of pathological section showed that the cell scaffold complex group provided good mechanical properties for the wound and supplemented the stem cells derived from chondrocytes and showed good cartilage defect repair effect; In the scaffold group, the surface fibrosis of the injured area was mainly filled with fibrocartilage and other collagen fibers The hydrogel/BMSCs complex based on COL and CMC can be beneficial for the regeneration of cartilage.

The development and improvement of immunodeficient mice and humanized immune system mouse models.

Animal models play an indispensable role in the study of human diseases. However, animal models of different diseases do not fully mimic the complex internal environment of humans. Immunodeficient mice are deficient in certain genes and do not express these or show reduced expression in some of their cells, facilitating the establishment of humanized mice and simulation of the human environment in vivo. Here, we summarize the developments in immunodeficient mice, from the initial nude mice lacking T lymphocytes to NOD/SCID rgnull mice lacking T, B, and NK cell populations. We describe existing humanized immune system mouse models based on immunodeficient mice in which human cells or tissues have been transplanted to establish a human immune system, including humanized-peripheral blood mononuclear cells (Hu-PBMCs), humanized hematopoietic stem cells (Hu-HSCs), and humanized bone marrow, liver, thymus (Hu-BLT) mouse models. The different methods for their development involve varying levels of complexity and humanization. Humanized mice are widely used in the study of various diseases to provide a transitional stage for clinical research. However, several challenges persist, including improving the efficiency of reconstructing the human B cell immune response, extending lifespan, improving the survival rate of mice to extend the observation period, and improving the development of standardized commercialized models and as well as their use. Overall, there are many opportunities and challenges in the development of humanized immune system mouse models which can provide novel strategies for understanding the mechanisms and treatments of human disease.

Life factors acting on systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a highly heterogeneous autoimmune disease that primarily affects women. Currently, in the search for the mechanisms of SLE pathogenesis, the association of lifestyle factors such as diet, cigarette smoking, ultraviolet radiation exposure, alcohol and caffeine-rich beverage consumption with SLE susceptibility has been systematically investigated. The cellular and molecular mechanisms mediating lifestyle effects on SLE occurrence, including interactions between genetic risk loci and environment, epigenetic changes, immune dysfunction, hyper-inflammatory response, and cytotoxicity, have been proposed. In the present review of the reports published in reputable peer-reviewed journals and government websites, we consider the current knowledge about the relationships between lifestyle factors and SLE incidence and outline directions of future research in this area. Formulation of practical measures with regard to the lifestyle in the future will benefit SLE patients and may provide potential therapy strategies.

Discovery of Drug Candidates for Specific Human Disease Based on Natural Products of Gut Microbes.

The beneficial metabolites of the microbiome could be used as a tool for screening drugs that have the potential for the therapy of various human diseases. Narrowing down the range of beneficial metabolite candidates in specific diseases was primarily a key step for further validation in model organisms. Herein, we proposed a reasonable hypothesis that the metabolites existing commonly in multiple beneficial (or negatively associated) bacteria might have a high probability of being effective drug candidates for specific diseases. According to this hypothesis, we screened metabolites associated with seven human diseases. For type I diabetes, 45 out of 88 screened metabolites had been reported as potential drugs in the literature. Meanwhile, 18 of these metabolites were specific to type I diabetes. Additionally, metabolite correlation could reflect disease relationships in some sense. Our results have demonstrated the potential of bioinformatics mining gut microbes' metabolites as drug candidates based on reported numerous microbe-disease associations and the Virtual Metabolic Human database. More subtle methods would be developed to ensure more accurate predictions.

EcoliGD: An Online Tool for Designing Escherichia coli Genome.

Synthetic biology is an important interdisciplinary field that has emerged in this century, focusing on the rewriting and reprogramming of DNA through the cycles of "design-edit", and so, the cell's own operating system, its genome, is naturally coming into focus. Here, we propose EcoliGD, an online genome design tool with a visual interactive interface and the function of browsing information, as well as the ability to perform insertion, exchange, deletion, and codon replacement operations on the E. coli genome and display the results in real-time. Users can utilize EcoliGD to check various functional characteristic about E. coli genes, to help them build their genomes. Furthermore, we also collected experimentally verified large genomic segments that have been successfully deleted from the genome for users to choose from and simplify the genome. EcoliGD can help recode the entire E. coli genome, providing a novel way to explore the diversity and function of this microorganism. The EcoliGD web tool is available at http://guolab.whu.edu.cn/EcoliGD/.

T-G-A Deficiency Pattern in Protein-Coding Genes and Its Potential Reason.

If a stop codon appears within one gene, then its translation will be terminated earlier than expected. False folding of premature protein will be adverse to the host; hence, all functional genes would tend to avoid the intragenic stop codons. Therefore, we hypothesize that there will be less frequency of nucleotides corresponding to stop codons at each codon position of genes. Here, we validate this inference by investigating the nucleotide frequency at a large scale and results from 19,911 prokaryote genomes revealed that nucleotides coinciding with stop codons indeed have the lowest frequency in most genomes. Interestingly, genes with three types of stop codons all tend to follow a T-G-A deficiency pattern, suggesting that the property of avoiding intragenic termination pressure is the same and the major stop codon TGA plays a dominant role in this effect. Finally, a positive correlation between the TGA deficiency extent and the base length was observed in start-experimentally verified genes of Escherichia coli (E. coli). This strengthens the proof of our hypothesis. The T-G-A deficiency pattern observed would help to understand the evolution of codon usage tactics in extant organisms.

Anti-CRISPRdb v2.2: an online repository of anti-CRISPR proteins including information on inhibitory mechanisms, activities and neighbors of curated anti-CRISPR proteins.

We previously released the Anti-CRISPRdb database hosting anti-CRISPR proteins (Acrs) and associated information. Since then, the number of known Acr families, types, structures and inhibitory activities has accumulated over time, and Acr neighbors can be used as a candidate pool for screening Acrs in further studies. Therefore, we here updated the database to include the new available information. Our newly updated database shows several improvements: (i) it comprises more entries and families because it includes both Acrs reported in the most recent literatures and Acrs obtained via performing homologous alignment; (ii) the prediction of Acr neighbors is integrated into Anti-CRISPRdb v2.2, and users can identify novel Acrs from these candidates; and (iii) this version includes experimental information on the inhibitory strength and stage for Acr-Cas/Acr-CRISPR pairs, motivating the development of tools for predicting specific inhibitory abilities. Additionally, a parameter, the rank of codon usage bias (CUBRank), was proposed and provided in the new version, which showed a positive relationship with predicted result from AcRanker; hence, it can be used as an indicator for proteins to be Acrs. CUBRank can be used to estimate the possibility of genes occurring within genome island-a hotspot hosting potential genes encoding Acrs. Based on CUBRank and Anti-CRISPRdb, we also gave the first glimpse for the emergence of Acr genes (acrs). DATABASE URL: http://guolab.whu.edu.cn/anti-CRISPRdb.

Geptop 2.0: Accurately Select Essential Genes from the List of Protein-Coding Genes in Prokaryotic Genomes.

Computational tool composites alternative way to identify essential genes and it is low-cost and time-efficient. Based on experimental essentiality sets deposited in the databases DEG and OGEE as reference, we developed an automatically computational tool named Geptop to select essential genes from the set of protein-coding genes in a prokaryotic genome, which utilizes the strategy of reciprocally best hit for homology search and evolutionary distance for weight assigning. The latest version of Geptop is 2.0 ( http://guolab.whu.edu.cn/geptop ), which can predict gene essentiality with the mean AUC 0f 0.84 in prokaryotes and is more stable. The chapter is to briefly introduce the tool and tell how to use it.

Consistent Clustering Pattern of Prokaryotic Genes Based on Base Frequency at the Second Codon Position and its Association with Functional Category Preference.

In 2002, our research group observed a gene clustering pattern based on the base frequency of A versus T at the second codon position in the genome of Vibrio cholera and found that the functional category distribution of genes in the two clusters was different. With the availability of a large number of sequenced genomes, we performed a systematic investigation of A2-T2 distribution and found that 2694 out of 2764 prokaryotic genomes have an optimal clustering number of two, indicating a consistent pattern. Analysis of the functional categories of the coding genes in each cluster in 1483 prokaryotic genomes indicated, that 99.33% of the genomes exhibited a significant difference (p < 0.01) in function distribution between the two clusters. Specifically, functional category P was overrepresented in the small cluster of 98.65% of genomes, whereas categories J, K, and L were overrepresented in the larger cluster of over 98.52% of genomes. Lineage analysis uncovered that these preferences appear consistently across all phyla. Overall, our work revealed an almost universal clustering pattern based on the relative frequency of A2 versus T2 and its role in functional category preference. These findings will promote the understanding of the rationality of theoretical prediction of functional classes of genes from their nucleotide sequences and how protein function is determined by DNA sequence.

High-Fat Diet-Induced Renal Proximal Tubular Inflammatory Injury: Emerging Risk Factor of Chronic Kidney Disease.

Nowadays, with the improvements in living standards and changes in living habits, high-fat diet (HFD) has become much more common in the populations worldwide. Recent studies have shown that HFD could induce lipid accumulation, and structural and functional abnormalities, accompanied by the release of large amounts of pro-inflammatory cytokines, in proximal tubular epithelial cells (PTECs). These findings indicate that, as an emerging risk factor, PTEC injury-induced by HFD may be closely related to inflammation; however, the potential mechanisms underlying this phenomenon is still not well-known, but may involve the several inflammatory pathways, including oxidative stress-related signaling pathways, mitochondrial dysfunction, the myeloid differentiation factor 2/Toll like receptor 4 (MD2/TLR4) signaling pathway, the ERK1/2-kidney injury molecule 1 (KIM-1)-related pathway, and nuclear factor-κB (NF-κB) activation, etc., and the detailed molecular mechanisms underlying these pathways still need further investigated in the future. Based on lipid abnormalities-induced inflammation is closely related to the development and progression of chronic kidney disease (CKD), to summarize the potential mechanisms underlying HFD-induced renal proximal tubular inflammatory injury, may provide novel approaches for CKD treatment.