Construction of lignan glycosides biosynthetic network in Escherichia coli using mutltienzyme modules.

BACKGROUND: Due to the complexity of the metabolic pathway network of active ingredients, precise targeted synthesis of any active ingredient on a synthetic network is a huge challenge. Based on a complete analysis of the active ingredient pathway in a species, this goal can be achieved by elucidating the functional differences of each enzyme in the pathway and achieving this goal through different combinations. Lignans are a class of phytoestrogens that are present abundantly in plants and play a role in various physiological activities of plants due to their structural diversity. In addition, lignans offer various medicinal benefits to humans. Despite their value, the low concentration of lignans in plants limits their extraction and utilization. Recently, synthetic biology approaches have been explored for lignan production, but achieving the synthesis of most lignans, especially the more valuable lignan glycosides, across the entire synthetic network remains incomplete. RESULTS: By evaluating various gene construction methods and sequences, we determined that the pCDF-Duet-Prx02-PsVAO gene construction was the most effective for the production of (+)-pinoresinol, yielding up to 698.9 mg/L after shake-flask fermentation. Based on the stable production of (+)-pinoresinol, we synthesized downstream metabolites in vivo. By comparing different fermentation methods, including "one-cell, one-pot" and "multicellular one-pot", we determined that the "multicellular one-pot" method was more effective for producing (+)-lariciresinol, (-)-secoisolariciresinol, (-)-matairesinol, and their glycoside products. The "multicellular one-pot" fermentation yielded 434.08 mg/L of (+)-lariciresinol, 96.81 mg/L of (-)-secoisolariciresinol, and 45.14 mg/L of (-)-matairesinol. Subsequently, ultilizing the strict substrate recognition pecificities of UDP-glycosyltransferase (UGT) incorporating the native uridine diphosphate glucose (UDPG) Module for in vivo synthesis of glycoside products resulted in the following yields: (+)-pinoresinol glucoside: 1.71 mg/L, (+)-lariciresinol-4-O-D-glucopyranoside: 1.3 mg/L, (+)-lariciresinol-4'-O-D-glucopyranoside: 836 µg/L, (-)-secoisolariciresinol monoglucoside: 103.77 µg/L, (-)-matairesinol-4-O-D-glucopyranoside: 86.79 µg/L, and (-)-matairesinol-4'-O-D-glucopyranoside: 74.5 µg/L. CONCLUSIONS: By using various construction and fermentation methods, we successfully synthesized 10 products of the lignan pathway in Isatis indigotica Fort in Escherichia coli, with eugenol as substrate. Additionally, we obtained a diverse range of lignan products by combining different modules, setting a foundation for future high-yield lignan production.

Prevalence and predictors of non-alcoholic liver disease on MRI among patients with Crohn's disease.

BACKGROUND: It has been documented that Crohn's disease (CD) patients were prone to develop non-alcoholic liver fatty liver disease (NAFLD) with less metabolic factors. Our purpose is to investigate the prevalence, clinical characteristics and possible indicators for NAFLD in a cohort of Chinese patients with CD. METHODS: Established CD patients who underwent magnetic resonance enterography (MRE) at the gastroenterology unit of our hospital were consecutively enrolled between June 2018 and May 2020. The diagnosis of NAFLD was made by magnetic resonance proton density fat fraction (MR-PDFF) maps. Medical records during hospitalization were collected and examined by univariate and multivariate analyses. Then a predictive model was constructed based on logistic regression analysis to evaluate the risk of developing NAFLD. RESULTS: A total of 340 CD subjects were enrolled in this study, 83 (24.4%) suffered from NAFLD. Compared with those without NAFLD, patients with NAFLD showed longer disease duration, higher body mass index (BMI), more frequent use of corticosteroid and pronouncedly elevated liver function tests. The comparison showed no difference in terms of prolonged anti tumor necrosis factor-α (TNF-α) use (> 54w). Multivariate analysis demonstrated that BMI, serum transaminase, pre-albumin and disease duration could independently predict hepatic steatosis. CONCLUSION: NAFLD is frequent in chronic CD patients, while long term use of anti TNF-α seems to have no impact on the development of NAFLD in this population. The model incorporating duration, serum transaminase and body mass index presented as a clinical nomogram could well predict the risk of NAFLD in patient with CD.

Efficacy of early intervention on the bowel damage and intestinal surgery of Crohn's disease, based on the Lémann index.

BACKGROUND: Clinicians aim to prevent progression of Crohn's disease (CD); however, many patients require surgical resection because of cumulative bowel damage. The aim of this study was to evaluate the impact of early intervention on bowel damage in patients with CD using the Lémann Index and to identify bowel resection predictors. METHODS: We analyzed consecutive patients with CD retrospectively. The Lémann Index was determined at the point of inclusion and at follow-up termination. The Paris definition was used to subdivide patients into early and late CD groups. RESULTS: We included 154 patients, comprising 70 with early CD and 84 with late CD. After follow-up for 17.0 months, more patients experienced a decrease in the Lémann Index (61.4% vs. 42.9%), and fewer patients showed an increase in the Lémann Index (20% vs. 35.7%) in the early compared with the late CD group. Infliximab and other therapies reversed bowel damage to a greater extent in early CD patients than in late CD patients. Twenty-two patients underwent intestinal surgery, involving 5 patients in the early CD group and 17 patients in the late CD group. Three independent predictors of bowel resection were identified: baseline Lémann index ≥ 8.99, disease behavior B1, and history of intestinal surgery. CONCLUSIONS: Early intervention within 18 months after CD diagnosis could reverse bowel damage and decrease short-term intestinal resection. Patients with CD with a history of intestinal surgery, and/or a Lémann index > 8.99 should be treated aggressively and monitored carefully to prevent progressive bowel damage.

Assessment of patient-centered outcomes (PROs) in inflammatory bowel disease (IBD): a multicenter survey preceding a cross-disciplinary (functional) consensus.

BACKGROUND: With a shift in the healthcare paradigm towards a more patient-centered approach, data on inflammatory bowel disease (IBD) needs to be further explored. This study aimed to determine patient perspectives on the effect of IBD and features of patients with lower satisfaction level and compare patient and physician perception of IBD-related Quality of Care (QoC). METHODS: A previously developed pre-standardised set of questions regarding patient-centered outcome (PRO) measures for IBD, comprising 36 items, was administered in five centers, and a concomitant questionnaire for specialised physicians was adapted and administered. RESULTS: Overall, 1005 patients with IBD met the inclusion criteria. Sixty-five questionnaires were administered to specialised physicians. Both patients and physicians perceived the IBD-related QoC as being satisfactory. Furthermore, this study revealed areas of shortcomings where it comes to patient perceptions. Female sex and the presence of negatively impacting disease characteristics (presence of significant pain or discomfort, lack of energy, feeling fatigued most of the time, experiencing anxiety or depression in the last 2 weeks) were associated with lower satisfaction levels. CONCLUSIONS: Our findings can be used in establishing strategies aimed at improving patient QoC and defining strategic priorities. These data can aid in improving the communication of the pressing needs of IBD patients, to both the public payers and health authorities.

[Advances in three important signaling pathways related to aging in Drosophila melanogaster and screening of anti-aging traditional Chinese medicine].

Aging is the most basic life feature of organisms. It is a phenomenon of dysfunction of cells,tissues and organs under the influence of external environment and internal factors during the growth of organisms. During the process,many cellular signaling pathways and biologically active substances,such as insulin/insulin-like growth factor( IGF)-1 signaling( IIS) pathway,apoptotic signaling pathway,mTOR signaling pathway,AMPK pathway,sirtuin pathway,deacetylases have been found to be closely related to the molecular mechanism of aging. Modern studies have indicated that anti-aging natural compounds can cause great side effects,while delaying aging and even inducing another disease,which is against with the purpose of delaying aging and achieving healthy aging. Therefore,the researches of anti-aging traditional Chinese medicines with fewer side effects are extremely important. Based on the different mechanisms and theories of aging,many traditional Chinese medicines have been discovered to be related to anti-aging. As one of the most important model organisms,Drosophila melanogaster has been widely used in studies of aging process in recent years. In this paper,we reviewed three important signaling pathways related to aging,such as insulin/insulin-like growth factor( IGF)-1 signaling( IIS) pathway,m TOR pathway,AMPK pathway,and screened out anti-aging traditional Chinese medicines based on D. melanogaster in recent years.

Effect of pinoresinol-lariciresinol reductases on biosynthesis of lignans with substrate selectivity in Schisandra chinensis.

Schisandra lignans are the main bioactive compounds found in Schisandra chinensis fruits, such as schisandrol lignans and schisandrin lignans, which play important roles in organ protection or other clinical roles. Pinoresinol-lariciresinol reductase (PLR) plays a pivotal role in plant lignan biosynthesis, however, limited research has been conducted on S. chinensis PLR to date. This study identified five genes as ScPLR, successfully cloned their coding sequences, and elucidated their catalytic capabilities. ScPLR3-5 could recognize both pinoresinol and lariciresinol as substrates, and convert them into lariciresinol and secoisolariciresinol, respectively, while ScPLR2 exclusively catalyzed the conversion of (+)-pinoresinol into (+)-lariciresinol. Transcript-metabolite correlation analysis indicated that ScPLR2 exhibited unique properties that differed from the other members. Molecular docking and site-directed mutagenesis revealed that Phe271 and Leu40 in the substrate binding motif were crucial for the catalytic activity of ScPLR2. This study serves as a foundation for understanding the essential enzymes involved in schisandra lignan biosynthesis.

Up-regulation and pre-activation of TRAF3 and TRAF5 in inflammatory bowel disease.

OBJECTIVE: TRAF3 and TRAF5 share a common ancestral gene, and interact as essential components of signaling pathways in immunity. TRAF3 and TRAF5 are overexpressed in the colon of rat/mouse models with colitis. However, the expressions of TRAF3 and TRAF5 in patients with inflammatory bowel disease have not been elucidated. The aim of the present study is to explore the potential roles of TRAF3 and TRAF5 in patients with inflammatory bowel disease. METHODS: Plasma levels of TRAF3 and TRAF5 proteins were detected by Enzyme-linked Immunosorbent Assay (ELISA). Colonic expression of TRAF3 and TRAF5 proteins was detected by western blot analysis. Quantitative Real-time PCR (qRT-PCR) was applied for gene expression. Inflamed intestinal mucosa and non-inflamed intestinal mucosa in patients with inflammatory bowel disease and normal mucosa was analyzed from healthy controls. RESULTS: The plasma levels of TRAF3 and TRAF5 were significantly higher both in patients with Crohn's disease and ulcerative colitis than in healthy controls. Only soluble TRAF5 showed a weak correlation with endoscopic disease activity index (Baron score) in patients with ulcerative colitis (spearman's r=0.358, P=0.022). Gene expressions of TRAF3 and TRAF5 in peripheral blood mononuclear cells were significantly higher both in patients with Crohn's disease and ulcerative colitis than in healthy controls (all P<0.0001). Gene and protein expressions of TRAF3 and TRAF5 were significantly higher in inflamed colonic mucosa of patients with Crohn's disease and ulcerative colitis than in non-inflamed colonic mucosa and normal mucosa of healthy controls (all P<0.0001). Furthermore, gene and protein expressions of TRAF3 and TRAF5 were also significantly higher in non-inflamed colonic mucosa of patients with Crohn's disease and ulcerative colitis than in normal mucosa of healthy controls. CONCLUSIONS: TRAF3 and TRAF5 are overexpressed in inflammatory bowel disease. Although the endoscopic appearance can be normal, TRAF3 and TRAF5 pre-activation can be detected in non-inflamed colonic segments.

Different activation of TRAF4 and TRAF6 in inflammatory bowel disease.

In recent years, interests combining the exploration of tumor necrosis factor receptor-associated factor 4 (TRAF4) and TRAF6 in immune cells and transgenic mice are emerging. Although it has been found that TRAF4 and TRAF6 share the same TRAF binding sites, comprehensive study of TRAF4 and TRAF6 in inflammatory bowel disease (IBD) is still lacking. This paper shows similar and different expression patterns of TRAF4 and TRAF6 in patients with IBD. The results indicate that TRAF4 and TRAF6 are overexpressed in IBD. TRAF4 and TRAF6 play different roles in the pathogenesis of IBD. Moreover, TRAF4 may be an indicator of endoscopic disease activity of UC and TRAF6 preactivation can be detected in noninflamed colonic segments.

Low vaccination and infection rate of Omicron in patients with inflammatory bowel disease: a comparative study of three unique cohorts.

Background: The SARS-CoV-2 Omicron variant caused a large-scale outbreak of COVID-19 in Shanghai, China. Patients with inflammatory bowel disease (IBD) are at high risk of infection due to immunosuppressive interventions. We aimed to investigate the vaccination information of patients with IBD and update a vaccination guide based on a comparison of vaccination in asymptomatic carriers and healthy individuals. Methods: This retrospective study was conducted during an Omicron variant wave. We assessed the vaccination status in patients with IBD, asymptomatic carriers and healthy individuals. Factors with unvaccinated status and adverse events following vaccination were also determined in patients with IBD. Results: The vaccination rate was 51.2% in patients with IBD, 73.2% in asymptomatic carriers, and 96.1% in healthy individuals. Female sex (p = 0.012), Crohn's disease (p = 0.026), and disease behavior of B3 (p = 0.029) were factors that indicated a lower vaccination rate. A significantly higher proportion of healthy individuals had received one booster dose (76.8%) than asymptomatic carriers (43.4%) and patients with IBD (26.2%). Patients with IBD received vaccination without an increased risk of adverse events (p = 0.768). Conclusion: The vaccination rate of patients with IBD remains much lower than that of asymptomatic carriers and healthy individuals. The COVID-19 vaccine has been found to be safe among all three groups and patients with IBD are not more susceptible to adverse events.

Phenotyping of Salvia miltiorrhiza Roots Reveals Associations between Root Traits and Bioactive Components.

Plant phenomics aims to perform high-throughput, rapid, and accurate measurement of plant traits, facilitating the identification of desirable traits and optimal genotypes for crop breeding. Salvia miltiorrhiza (Danshen) roots possess remarkable therapeutic effect on cardiovascular diseases, with huge market demands. Although great advances have been made in metabolic studies of the bioactive metabolites, investigation for S. miltiorrhiza roots on other physiological aspects is poor. Here, we developed a framework that utilizes image feature extraction software for in-depth phenotyping of S. miltiorrhiza roots. By employing multiple software programs, S. miltiorrhiza roots were described from 3 aspects: agronomic traits, anatomy traits, and root system architecture. Through K-means clustering based on the diameter ranges of each root branch, all roots were categorized into 3 groups, with primary root-associated key traits. As a proof of concept, we examined the phenotypic components in a series of randomly collected S. miltiorrhiza roots, demonstrating that the total surface of root was the best parameter for the biomass prediction with high linear regression correlation (R2 = 0.8312), which was sufficient for subsequently estimating the production of bioactive metabolites without content determination. This study provides an important approach for further grading of medicinal materials and breeding practices.

Functional divergence of CYP76AKs shapes the chemodiversity of abietane-type diterpenoids in genus Salvia.

The genus Salvia L. (Lamiaceae) comprises myriad distinct medicinal herbs, with terpenoids as one of their major active chemical groups. Abietane-type diterpenoids (ATDs), such as tanshinones and carnosic acids, are specific to Salvia and exhibit taxonomic chemical diversity among lineages. To elucidate how ATD chemical diversity evolved, we carried out large-scale metabolic and phylogenetic analyses of 71 Salvia species, combined with enzyme function, ancestral sequence and chemical trait reconstruction, and comparative genomics experiments. This integrated approach showed that the lineage-wide ATD diversities in Salvia were induced by differences in the oxidation of the terpenoid skeleton at C-20, which was caused by the functional divergence of the cytochrome P450 subfamily CYP76AK. These findings present a unique pattern of chemical diversity in plants that was shaped by the loss of enzyme activity and associated catalytic pathways.

Construction of a neural network diagnostic model and investigation of immune infiltration characteristics for Crohn's disease.

Objective: Crohn's disease (CD), a chronic recurrent illness, is a type of inflammatory bowel disease whose incidence and prevalence rates are gradually increasing. However, there is no universally accepted criterion for CD diagnosis. The aim of this study was to create a diagnostic prediction model for CD and identify immune cell infiltration features in CD. Methods: In this study, gene expression microarray datasets were obtained from the Gene Expression Omnibus (GEO) database. Then, we identified differentially expressed genes (DEGs) between 178 CD and 38 control cases. Enrichment analysis of DEGs was also performed to explore the biological role of DEGs. Moreover, the "randomForest" package was applied to select core genes that were used to create a neural network model. Finally, in the training cohort, we used CIBERSORT to evaluate the immune landscape between the CD and normal groups. Results: The results of enrichment analysis revealed that these DEGs may be involved in biological processes associated with immunity and inflammatory responses. Moreover, the top 3 hub genes in the protein-protein interaction network were IL-1ß, CCL2, and CXCR2. The diagnostic model allowed significant discrimination with an area under the ROC curve of 0.984 [95% confidence interval: 0.971-0.993]. A validation cohort (GSE36807) was utilized to ensure the reliability and applicability of the model. In addition, the immune infiltration analysis indicated nine different immune cell types were significantly different between the CD and healthy control groups. Conclusion: In summary, this study offers a novel insight into the diagnosis of CD and provides potential biomarkers for the precise treatment of CD.

Aqueous Extract from Astragalus membranaceus Can Improve the Function Degradation and Delay Aging on Drosophila melanogaster Through Antioxidant Mechanism.

Astragali Radix is the dry root of the leguminous plants Astragalus membranaceus (Fisch.) Bge. Var. mongholicus (Bge.) Hsiao and A. membranaceus (Fisch.) Bge. Astragali Radix is mostly used clinically as a decoction. A number of pharmacological studies show that Astragalus extract can increase telomerase activity and has antioxidation, anti-inflammatory, immune regulation, anticancer, lowering blood lipid, lowering blood sugar, and other effects. However, the anti-aging mechanism of aqueous extract from Astragali Radix (ARE) is still unclear. In this study, we evaluated the anti-aging effect of ARE on Drosophila melanogaster and investigated the underlying mechanism. The results of life span assay showed that 1.25 mg/mL of ARE can significantly prolong the life span of D. melanogaster in a natural aging model and protect against H2O2 and paraquat. Meanwhile, ARE can improve the climbing ability and food intake of flies. Metabolomics and the glutamate content assay suggested that ARE prevented an age-dependent increase in glutamate levels in D. melanogaster. Furthermore, ARE showed a dose-dependent effect on the scavenging ability of α, α-diphenyl-ß-picrylhydrazyl in vitro. Superoxide dismutase and catalase activities in the aging group also increased after the intervention of ARE. The data and the findings described here support the notion that ARE may play a preventive role in aging by improving the climbing ability, eliminating harmful free radicals accumulated in D. melanogaster and triggering antioxidant responses.

Relating the transcriptome and microbiome by paired terminal ileal Crohn disease.

Management of terminal ileal Crohn disease (CD) is difficult due to fibrotic prognosis and failure to achieve mucosal healing. A limited number of synchronous analyses have been conducted on the transcriptome and microbiome in unpaired terminal ileum tissues. Therefore, our study focused on the transcriptome and mucosal microbiome in terminal ileal tissues of patients with CD with the aim of determining the role of cross-talk between the microbiome and transcriptome in the pathogenesis of terminal ileal CD. Mucosa-attached microbial communities were significantly associated with segmental inflammation status. Interaction-related transcription factors (TFs) are the panel nodes for cross-talk between the gene patterns and microbiome for terminal ileal CD. The transcriptome and microbiome in terminal ileal CD can be differently related to the local inflammatory status, and specific differentially expressed genes may be targeted for mucosal healing. TFs connect gene patterns with the microbiome by reflecting environmental stimuli and signals from microbiota.

MRI-Based Radiomic Signature Identifying Secondary Loss of Response to Infliximab in Crohn's Disease.

Up to 50% of patients with Crohn's disease (CD) experience secondary loss of response (SLR) to infliximab. Patients with SLR may show clinical signs of iron deficiency as a result of inflammation despite being iron-replete. The magnetic resonance imaging (MRI)-based radiomic index, R2*, can detect changes in iron metabolism. Therefore, the R2* parameter has considerable potential for detection of SLR to infliximab. The aims of this study were to explore the correlation between R2* and inflammation and to develop a non-invasive nomogram based on R2* to identify SLR to infliximab in patients with CD. Three hundred and twenty-two infliximab-treated patients with CD who underwent magnetic resonance enterography within 2 weeks before or after 54 weeks of infliximab therapy were divided into training and validation datasets at a ratio of 8:2. Point-biserial analysis was conducted to confirm the relationship between R2* and inflammation. A multivariate logistic regression model was created using R2*, CRP and hemoglobin (OR, 1.10, 1.04 and 0.98; P < 0.05). Receiver-operating characteristic curves and the Hosmer-Lemeshow test were used to assess the performance of the model. A correlation between R2* and inflammation was identified. Different trends in R2* and iron status indices were observed between patients with responsive and non-responsive CD, which is worthy of further study. The model was converted to a visualized nomogram that had a good ability to discriminate the outcomes of infliximab therapy with an area under the curve of 0.723 (95% CI, 0.661-0.785) in the training dataset and 0.715 (95% CI, 0.587-0.843) in the validation dataset. We confirmed a correlation between R2* and inflammation in patients with CD. Based on the MRI-based radiomic signature, a novel nomogram was established and validated to facilitate individualized identification of SLR to infliximab in patients with CD.

Microcystis blooms aggravate the diurnal alternation of nitrification and nitrate reduction in the water column in Lake Taihu.

To explore the effects of Microcystis blooms on nitrogen (N) cycling in the water column, the community structures of the Microcystis-attached and free-living bacteria in Lake Taihu were assessed and a mesocosm experiment was further conducted on the shore of Lake Taihu. The bacterial communities of Microcystis-attached and free-living bacteria were dominated by heterotrophic bacteria, such as Pseudomonas and Massilia, while the relative abundances of the genera related to traditional autotrophic nitrification were surprisingly low. However, the dramatic increase in nitrate (NO3-) levels at the daytime suggested that in the mesocosms nitrification did occur, during which the heterotrophic nitrifiers played a predominant role as revealed by the acetylene inhibition experiment. The ammonium (NH4+) concentrations were always maintained at a low level, indicating that most of the substrates for daytime nitrification originated from organic N. The total N being removed during the experiment was much less than the sum of daily NO3- reduction, while the decrease in NO3- concentration was much higher than the increase in NH4+ concentration during the night, indicating that assimilation was the main explanation for nocturnal NO3- reduction. Thus, the cycling of organic N (remineralization) - heterotrophic nitrification - NO3- assimilation (reduction) promoted by Microcystis blooms aggravates the diurnal variation of NO3- in the water column.

Astragaloside IV Extends Lifespan of Caenorhabditis elegans by Improving Age-Related Functional Declines and Triggering Antioxidant Responses.

Astragaloside IV (AS-IV) is a representative component of astragaloside saponins in dried roots of Astragali Radix. Astragaloside possesses a broad spectrum of pharmacological activities, including antibacterial, anti-fibrosis, antioxidant, anti-inflammatory, and neuroprotective effects. However, the role of AS-IV in antiaging remains unclear. In this article, we studied the function of AS-IV in antiaging by using the Caenorhabditis elegans (C. elegans) model. We showed that AS-IV can prolong the lifespan of C. elegans in a natural aging model, a paraquat injury model, and a heat stress model and improve the movement capacity of nematodes. 1H-NMR data indicate an improvement of glutamate content and a decrease in glucose in the AS-IV treatment group compared with the control. Further investigation revealed that AS-IV can induce the mRNA expression of superoxide dismutase (SOD) and catalase (CAT) genes and increase the activities of SOD and CAT in the nematode. Interestingly, AS-IV could not extend the lifespan of sod-1, sod-2, sod-3, sod-4, sod-5, ctl-1, ctl-2, ctl-3, and daf-16 mutants. These data indicate that AS-IV prevents aging via mainly improving age-related functional declines, the antioxidant capacity of nematodes and partially modulating the insulin/insulin growth factor 1 signaling pathway activity. Our results provide new insights into how AS-IV prevents and treats aging.

Potential influential factors on incidence and prevalence of inflammatory bowel disease in mainland China.

The incidence of inflammatory bowel disease (IBD) has increased annually in newly industrialized countries. In mainland China, there is also a trend of increasing incidence and prevalence of IBD. Epidemiological studies have shown that the incidence of IBD in mainland China is higher in the south and lower in the north. However, the existing research results are affected by the factor of floating population. Some socioeconomic factors also seem to play a role in the rising trend of IBD incidence and prevalence. These underlying factors provide opportunities and challenges for our IBD epidemiological studies over the next decade.