Clinical characteristics and the risk factors for the exacerbation of symptoms in patients with inflammatory bowel disease during the COVID-19 pandemic.

Background: In 2023, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant caused a large-scale outbreak of coronavirus disease 2019 (COVID-19) in China. It is not clear the risk factors that lead to the exacerbation of symptoms in patients with inflammatory bowel disease (IBD) after COVID-19 infection. Our study aims to find out the risk factors for the exacerbation of IBD-related symptoms in IBD patients with COVID-19 infection and to provide guidance for the clinical management of IBD. Methods: This is a retrospective, observational study. The online questionnaire was distributed to conduct a survey to collect demographic, clinical, and IBD related characteristics in IBD patients. Univariate and multivariate regression analyses were conducted to assess the independent effects. Results: In total, 534 cases of IBD patients were analyzed in our study. Among them, 466 (87.3%) cases diagnosed with COVID-19, 160 (34.3%) cases experienced exacerbation of IBD symptoms, and 84 (18.0%) patients opted for medication discontinuation. Male sex (OR 2.04, 95% CI 1.34-3.49, p = 0.001), and the decrease in body mass index (BMI) (OR 0.93, 95% CI 0.87-1.00, p = 0.035) were positively correlated with the exacerbation of IBD symptoms. Furthermore, the medication discontinuation (OR 2.60, 95% CI 1.58-4.30, p < 0.001) was strongly positively correlated with the exacerbation of IBD symptoms. No significant association was seen between age, comorbidities, smoking, disease activity, vaccination, therapy for COVID-19 and the worsening of IBD symptoms. Conclusion: This study confirms that the infection rate of COVID-19 in China IBD patients was comparable to the general population. Male sex, the decrease in BMI and medication discontinuation are significant risk factors for the exacerbation of IBD-related symptoms in IBD patients with COVID-19 infection.

Chromosome-level genome sequencing and multi-omics of the Hungarian White Goose (Anser anser domesticus) reveals novel miRNA-mRNA regulation mechanism of waterfowl feather follicle development.

The Hungarian White Goose (Anser anser domesticus) is an excellent European goose breed, with high feather and meat production. Despite its importance in the poultry industry, no available genome assembly information has been published. This study aimed to present Chromosome-level and functional genome sequencing of the Hungarian White Goose. The results showed that the genome assembly has a total length of 1115.82 Mb, 39 pairs of chromosomes, 92.98% of the BUSCO index, and contig N50 and scaffold N50 were up to 2.32 Mb and 60.69 Mb, respectively. Annotation of the genome assembly revealed 19550 genes, 286 miRNAs, etc. We identified 235 expanded and 1,167 contracted gene families in this breed compared with the other 16 species. We performed a positive selection analysis between this breed and four species of Anatidae to uncover the genetic information underlying feather follicle development. Further, we detected the function of miR-199-x, miR-143-y, and miR-23-z on goose embryonic skin fibroblast. In summary, we have successfully generated a highly complete genome sequence of the Hungarian white goose, which will provide a great resource to improve our understanding of gene functions and enhance the studies on feather follicle development at the genomic level.

DHX9 maintains epithelial homeostasis by restraining R-loop-mediated genomic instability in intestinal stem cells.

Epithelial barrier dysfunction and crypt destruction are hallmarks of inflammatory bowel disease (IBD). Intestinal stem cells (ISCs) residing in the crypts play a crucial role in the continuous self-renewal and rapid recovery of intestinal epithelial cells (IECs). However, how ISCs are dysregulated in IBD remains poorly understood. Here, we observe reduced DHX9 protein levels in IBD patients, and mice with conditional DHX9 depletion in the intestinal epithelium (Dhx9ΔIEC) exhibit an increased susceptibility to experimental colitis. Notably, Dhx9ΔIEC mice display a significant reduction in the numbers of ISCs and Paneth cells. Further investigation using ISC-specific or Paneth cell-specific Dhx9-deficient mice demonstrates the involvement of ISC-expressed DHX9 in maintaining epithelial homeostasis. Mechanistically, DHX9 deficiency leads to abnormal R-loop accumulation, resulting in genomic instability and the cGAS-STING-mediated inflammatory response, which together impair ISC function and contribute to the pathogenesis of IBD. Collectively, our findings highlight R-loop-mediated genomic instability in ISCs as a risk factor in IBD.

Protein profile analysis of Jilin white goose testicles at different stages of the laying cycle by DIA strategy.

BACKGROUND: Jilin white goose is an excellent local breed in China, with a high annual egg production and laying eggs mainly from February to July each year. The testis, as the only organ that can produce sperm, can affect the sexual maturity and fecundity of male animals. Its growth and development are affected and regulated by a variety of factors. Proteomics is generally applied to identify and quantify proteins in cells and tissues in order to understand the physiological or pathological changes that occur in tissues or cells under specific conditions. Currently, the female poultry reproductive system has been extensively studied, while few related studies focusing on the regulatory mechanism of the reproductive system of male poultry have been conducted. RESULTS: A total of 1753 differentially expressed proteins (DEPs) were generated in which there were 594, 391 and 768 different proteins showing differential expression in three stages, Initial of Laying Cycle (ILC), Peak of Laying Cycle (PLC) and End of Laying Cycle (ELC). Furthermore, bioinformatics was used to analyze the DEPs. Gene ontology (GO) enrichment, Clusters of Orthologous Groups (COG), Kyoto Encyclopedia of Genes and Genomes (KEGG) and protein-protein interaction (PPI) network analysis were adopted. All DEPs were found to be implicated in multiple biological processes and pathways associated with testicular development, such as renin secretion, Lysosomes, SNARE interactions in vesicle trafficking, the p53 signaling pathway and pathways related to metabolism. Additionally, the reliability of transcriptome results was verified by real-time quantitative PCR by selecting the transcript abundance of 6 selected DEPs at the three stages of the laying cycle. CONCLUSIONS: The funding in this study will provide critical insight into the complex molecular mechanisms and breeding practices underlying the developmental characteristics of testicles in Jilin white goose.

Dermal FOXO3 activity in response to Wnt/ß-catenin signaling is required for feather follicle development of goose embryos (Anser cygnoides).

Feather is an important economic trait of poultry, and growth and development state of feathers plays an important role in the economic value of poultry. Dermal fibroblasts are required for structural integrity of the skin and for feather follicle development. How FOXO3 affects feather follicle development as skin tissues change during goose embryo (Anser cygnoides) development and growth is not well understood. Here, we demonstrate that in vitro culture of single feathers and skin tissue results in changes in feather morphological structure by adding drugs to the culture medium that affect FOXO3 expression. We used feather follicles to show that during growth, the root location of feathers, the dermis layer, affects cell proliferation and apoptosis and regulates the expression of major genes in the Wingless-types/beta-catenin (Wnt/ß-catenin) signaling pathway through the activity of FOXO3 in dermal fibroblasts. Feathers and dorsal skin tissues develop the correct structure, but feather length and width and feather follicle diameter change significantly (p < 0.05) without significant changes in feather follicle density (p > 0.05). Transfected dermal fibroblasts also showed that FOXO3 affected the formation and development of feather follicles in the embryonic stage by regulating the Wnt/ß-catenin signaling pathway. Therefore, this study reveals the critical role of dermal fibroblast-FOXO3-induced Wnt/ß-catenin signaling in promoting the formation and development of embryonic feather follicles.

miR-140-y targets TCF4 to regulate the Wnt signaling pathway and promote embryonic feather follicle development in Hungarian white goose.

Goose down feather has become one of the most important economical products in the goose breeding industry and it provides several essential physiological roles in birds. Therefore, understanding and regulating the development of skin and feather follicles during embryogenesis is critical for avian biology and the poultry industry. MicroRNAs are known to play an important role in controlling gene expression during skin and feather follicle development. In this study, bioinformatics analysis was conducted to select miR-140-y as a potential miRNA involved in skin and feather follicle development and to predict TCF4 as its target gene. This gene was expressed at significant levels during embryonic feather follicle development, as identified by qPCR and Western blot. The targeting relationship was confirmed by a dual-luciferase assay in 293T cells. Then, the miR-140-y/TCF4 function in dermal fibroblast cells was explored. The results showed that miR-140-y could suppress the proliferation of goose embryonic dermal fibroblast cells (GEDFs) by suppressing the activity of some Wingless-types (Wnt) pathway related genes and proliferation marker genes, while miR-140-y inhibition led to the opposite effect. Similarly, the inhibition of the TCF4 gene results in blocking the proliferation of GEDFs by reducing the activity of some Wnt pathway-related genes. Finally, the co-transfection of miR-140-y inhibitor and siRNA-TCF4 results in a rescue of the TCF4 function and an increase of the Wnt signaling pathway and GEDFs proliferation. In conclusion, these results demonstrated that the miR-140-y-TCF4 axis influences the activity of the Wnt signaling pathway and works as a dynamic regulator during skin and feather follicle development.

Use of double-balloon endoscopy and an endoscopic scoring system to assess endoscopic remission in isolated small bowel Crohn's disease after treatment with infliximab.

Background: It is unclear how clinical and endoscopic factors affect the attainment of endoscopic remission (ER) in patients with small bowel Crohn's disease (SB-CD) who are infliximab-naïve. Objectives: We aimed to identify the effect of different factors on attaining ER using double-balloon endoscopy (DBE) evaluation. Design: A single-center retrospective observational study was conducted from 1 January 2018 to 30 November 2022. Among 262 patients who were screened for isolated SB-CD by baseline DBE, 108 patients were assessed for effectiveness during maintenance infliximab therapy by a second DBE evaluation. Methods: DBE findings before and after infliximab therapy were compared. ER was defined as a simple endoscopic score for CD (SES-CD) below 3, and segmental ER as SES-CD activity of 0. Multivariate regression with calculations of odds ratios (OR) was used to determine the impact of different factors on attaining ER. Results: In all, 41 patients (38.0%) achieved ER. An elevated C-reactive protein at week 6 was independently associated with a decreased probability of ER [OR: 0.86, 95% confidence interval (CI) = 0.75-0.98, p = 0.03]. Segmental ER of the terminal ileum, rather than the proximal ileum, was associated with a higher rate of ER (60.9% versus 38.2%, p = 0.01). High baseline SES-CD (⩾16) was unrelated to overall ER. For patients with disease in the terminal ileum, those with moderate/severe disease were less likely to attain segmental ER than those with mild disease [adjusted odds ratios (aOR): 0.27, 95% CI: 0.09-0.83, p = 0.02]. A large ulcer in the terminal ileum was associated with a lower rate of segmental ER (aOR: 0.18, 95% CI: 0.06-0.56, p = 0.01). Conclusion: For infliximab-naïve patients with SB-CD, the overall severity of the endoscopic score was unrelated to attainment of ER. Patients were less likely to attain segmental ER if they had greater endoscopic inflammation or larger ulcers in the terminal ileum.

Deep learning-based lesion detection and severity grading of small-bowel Crohn's disease ulcers on double-balloon endoscopy images.

BACKGROUND AND AIMS: Double-balloon endoscopy (DBE) is widely used in diagnosing small-bowel Crohn's disease (CD). However, CD misdiagnosis frequently occurs if inexperienced endoscopists cannot accurately detect the lesions. The CD evaluation may also be inaccurate owing to the subjectivity of endoscopists. This study aimed to use artificial intelligence (AI) to accurately detect and objectively assess small-bowel CD for more refined disease management. METHODS: We collected 28,155 small-bowel DBE images from 628 patients from January 2018 to December 2022. Four expert gastroenterologists labeled the images, and at least 2 endoscopists made the final decision with agreement. A state-of-the-art deep learning model, EfficientNet-b5, was trained to detect CD lesions and evaluate CD ulcers. The detection included lesions of ulcer, noninflammatory stenosis, and inflammatory stenosis. Ulcer grading included ulcerated surface, ulcer size, and ulcer depth. A comparison of AI model performance with endoscopists was performed. RESULTS: The EfficientNet-b5 achieved high accuracies of 96.3% (95% confidence interval [CI], 95.7%-96.7%), 95.7% (95% CI, 95.1%-96.2%), and 96.7% (95% CI, 96.2%-97.2%) for the detection of ulcers, noninflammatory stenosis, and inflammatory stenosis, respectively. In ulcer grading, the EfficientNet-b5 exhibited average accuracies of 87.3% (95% CI, 84.6%-89.6%) for grading the ulcerated surface, 87.8% (95% CI, 85.0%-90.2%) for grading the size of ulcers, and 85.2% (95% CI, 83.2%-87.0%) for ulcer depth assessment. CONCLUSIONS: The EfficientNet-b5 achieved high accuracy in detecting CD lesions and grading CD ulcers. The AI model can provide expert-level accuracy and objective evaluation of small-bowel CD to optimize the clinical treatment plans.

A lncRNA from an inflammatory bowel disease risk locus maintains intestinal host-commensal homeostasis.

Inflammatory bowel diseases (IBD) are known to have complex, genetically influenced etiologies, involving dysfunctional interactions between the intestinal immune system and the microbiome. Here, we characterized how the RNA transcript from an IBD-associated long non-coding RNA locus ("CARINH-Colitis Associated IRF1 antisense Regulator of Intestinal Homeostasis") protects against IBD. We show that CARINH and its neighboring gene coding for the transcription factor IRF1 together form a feedforward loop in host myeloid cells. The loop activation is sustained by microbial factors, and functions to maintain the intestinal host-commensal homeostasis via the induction of the anti-inflammatory factor IL-18BP and anti-microbial factors called guanylate-binding proteins (GBPs). Extending these mechanistic insights back to humans, we demonstrate that the function of the CARINH/IRF1 loop is conserved between mice and humans. Genetically, the T allele of rs2188962, the most probable causal variant of IBD within the CARINH locus from the human genetics study, impairs the inducible expression of the CARINH/IRF1 loop and thus increases genetic predisposition to IBD. Our study thus illustrates how an IBD-associated lncRNA maintains intestinal homeostasis and protects the host against colitis.

The Mechanosensitive Ion Channel PIEZO1 in Intestinal Epithelial Cells Mediates Inflammation through the NOD-Like Receptor 3 Pathway in Crohn's Disease.

BACKGROUND: Crohn's disease (CD) is an incurable chronic intestinal inflammatory disease with no recognized cause. It has been reported that the mechanosensitive ion channel PIEZO1 initiates proinflammatory responses. However, little is known about the role of PIEZO1 in CD. METHODS: Ileum biopsies were obtained from 30 patients with CD and 15 healthy volunteers. Clinical data were collected to determine the relationship between CD and PIEZO1. First, HT29 cells were incubated with Yoda1 and GsMTx4 (Grammostola spatulata mechanotoxin 4) to activate and inhibit PIEZO1, respectively. Second, PIEZO1 knockdown was performed using small interfering RNA. Third, calcium imaging, flow cytometry, and immunofluorescence were used to detect intracellular calcium and mitochondrial function. Last, real-time quantitative polymerase chain reaction, immunoblotting, and enzyme-linked immunosorbent assay were used to quantify PIEZO1, proinflammatory cytokines, and NLRP3 (NOD-like receptor 3)-related compounds. RESULTS: PIEZO1 was highly expressed in the ileum of patients with CD and correlated positively with the Crohn's Disease Activity Index, platelet count, and hematocrit and fecal calprotectin levels. In HT29 cells, Yoda1 triggered calcium influx, which was inhibited by GsMTx4 treatment and small interfering RNA-mediated PIEZO1 knockdown. Increased calcium concentrations resulted in increased reactive oxygen species accumulation and decreased mitochondrial membrane potential, whereas decreased calcium concentrations caused by GsMTx4 and PIEZO1 knockdown had the opposite effect. Mechanistically, molecules in the NLRP3 pathway were activated in patients with CD and HT29 cells were stimulated by lipopolysaccharide; these effects were reversed by the knockdown of PIEZO1. Finally, PIEZO1 and NLRP3 knockdown decreased proinflammatory cytokine levels in HT29 cells. CONCLUSIONS: PIEZO1 in intestinal epithelial cells caused calcium influx, which resulted in mitochondrial dysfunction and activated the NLRP3 inflammasome, mediating intestinal inflammation.

PIEZO1 was highly expressed in patients with active Crohn's disease. Its expression was significantly increased in the ileum and was associated with intestinal inflammation. In intestinal epithelial cells, PIEZO1 triggered calcium influx, resulting in mitochondrial dysfunction, and activated the NLRP3 inflammasome, thereby mediating intestinal inflammation.

Vitamin B5 rewires Th17 cell metabolism via impeding PKM2 nuclear translocation.

Metabolic rewiring is essential for Th17 cells' functional identity to sense and interpret environmental cues. However, the environmental metabolic checkpoints with specific regulation of Th17 cells, manifesting potential therapeutic opportunities to autoimmune diseases, remain largely unknown. Here, by screening more than one hundred compounds derived from intestinal microbes or diet, we found that vitamin B5 (VB5) restrains Th17 cell differentiation as well as related autoimmune diseases such as experimental autoimmune encephalomyelitis and colitis. Mechanistically, VB5 is catabolized into coenzyme A (CoA) in a pantothenate kinase (PANK)-dependent manner, and in turn, CoA binds to pyruvate kinase isoform 2 (PKM2) to impede its phosphorylation and nuclear translocation, thus inhibiting glycolysis and STAT3 phosphorylation. In humans, reduced serum VB5 levels are found in both IBD and MS patients. Collectively, our study demonstrates a role of VB5 in Th17 cell metabolic reprograming, thus providing a potential therapeutic intervention for Th17 cell-associated autoimmune diseases.

Detection of a novel antigen for Crohn's disease.

BACKGROUND AND AIMS: Accurate serological assays are desirable for the diagnosis of inflammatory bowel disease (IBD). We identify an antigen-like substance called Crohn's disease (CD) antibody binding polypeptide (CABP). As a serological marker, anti-CABP may contribute to the diagnosis of IBD. The present study aims to evaluate the clinical role of anti-CABP as a serological antibody for IBD. METHODS: Using enzyme-linked immunosorbent assay (ELISA), serum anti-CABP, anti-Saccharomyces cerevisiae antibody (ASCA) and perinuclear anti-neutrophil cytoplasmic antibody (pANCA), titers were tested in 168 CD patients, 123 ulcerative colitis (UC) patients and 170 controls. The correlation between serum antibody and clinical characteristics was investigated. The diagnostic potential of the anti-CABP was evaluated by receiver operating characteristic (ROC) analysis. RESULTS: The titers of anti-CABP (IgA or IgG) and ASCA IgG of CD patients were significantly higher than non-CD group (all p < .01). In the differential diagnosis of CD and non-CD, anti-CABP IgA revealed an area under the curve (AUC) of 0.706 and anti-CABP IgG demonstrated an AUC of 0.788. As an individual antibody, anti-CABP could effectively distinguish CD from non-CD (AUC 0.816), and the diagnostic efficacy was better than that of ASCA (AUC 0.680). The combined use of anti-CABP, ASCA and pANCA significantly improved the diagnostic value (AUC 0.857). Anti-CABP positive rates were associated with perianal lesions and disease location in CD patients (both p < .05). CONCLUSIONS: Our results suggested that anti-CABP could be used as a serological marker to assist the diagnosis of CD. CLINICAL TRIAL REGISTRATION: This trial is registered with clinical trial registration unique identifier ChiCTR2000037094.

Effects of mid-stage nitrogen application timing on the morphological structure and physicochemical properties of japonica rice starch.

BACKGROUND: Nitrogen management of crops, especially when mid-stage nitrogen is applied, is a key factor affecting the yield and grain quality of rice (Oryza sativa). Here, the timing of mid-stage nitrogen application was evaluated for its effect on rice grain quality by assessing the morphological structure and physicochemical properties of starch from two japonica rice cultivars growing in fields (Nangeng 9108 and Nangeng 5055). RESULTS: The experiment was arranged in a split-plot design, with the two rice cultivars as the main plot factor and three timings of mid-stage nitrogen application as the within-plot factor. Briefly, three applications were made: at the emergence of the top-sixth-leaf (ahead), the top-fourth-leaf (normal), and the top-second-leaf (delayed) of the main stem. Delaying mid-stage nitrogen application caused the starch granule surface to become uneven and significantly reduced its particle size, whereas it increased the polished rice rate, chalkiness degree, and protein content. Furthermore, the apparent amylose content decreased with a delay in mid-stage nitrogen application, thereby resulting in higher relative crystallinity, swelling power, water solubility, gelatinization enthalpy, and low retrogradation. Finally, we also found that delaying this nitrogen application lowered the characteristic values of rice flour viscosities, leading to cooking quality deterioration. CONCLUSION: These results therefore suggest that delaying mid-stage nitrogen application enhances the processing and nutritional qualities of japonica rice but evidently has an adverse effect upon its appearance and cooking qualities. © 2020 Society of Chemical Industry.

Two NAD-independent l-lactate dehydrogenases drive l-lactate utilization in Pseudomonas aeruginosa PAO1.

Pseudomonas aeruginosa often establishes a chronic infection in the airways of patients with cystic fibrosis (CF). l-Lactate is the most abundant carbon source in the CF sputum, and l-lactate utilization may be important for P. aeruginosa to survive in the lungs of CF patients. In this study, the key enzymes involved in l-lactate utilization by P. aeruginosa PAO1 were characterized using the synthetic CF sputum medium (SCFM). A highly conserved membrane-bound NAD-independent l-lactate dehydrogenase (l-iLDH) encoded by lldD (PA4771) and a novel flavin-containing membrane-bound l-iLDH encoded by lldA (PA2382) were both found to contribute to l-lactate utilization by P. aeruginosa PAO1. In addition, an lldD and lldA double mutant was incapable of growing in a medium containing l-lactate as the sole carbon source. This study clarifies the mechanism and importance of l-lactate catabolism, and demonstrates the first Pseudomonas spp. expressing two l-lactate-oxidizing enzymes.

2,3-Butanediol catabolism in Pseudomonas aeruginosa PAO1.

2,3-Butanediol (2,3-BD) is a primary microbial metabolite that enhances the virulence of Pseudomonas aeruginosa and alters the lung microbiome. 2,3-BD exists in three stereoisomeric forms: (2R,3R)-2,3-BD, meso-2,3-BD and (2S,3S)-2,3-BD. In this study, we investigated whether and how P. aeruginosa PAO1 utilizes these 2,3-BD stereoisomers and showed that all three stereoisomers were transformed into acetoin by (2R,3R)-2,3-butanediol dehydrogenase (BDH) or (2S,3S)-2,3-BDH. Acetoin was cleaved to form acetyl-CoA and acetaldehyde by acetoin dehydrogenase enzyme system (AoDH ES). Genes encoding (2R,3R)-2,3-BDH, (2S,3S)-2,3-BDH and the E1 and E2 components of AoDH ES were identified as part of a new 2,3-BD utilization operon. In addition, the regulatory protein AcoR promoted the expression of this operon using acetaldehyde, a cleavage product of acetoin, as its direct effector. The results of this study elucidate the integrated catabolic role of 2,3-BD and may provide new insights in P. aeruginosa-related infections.

Coupling between d-3-phosphoglycerate dehydrogenase and d-2-hydroxyglutarate dehydrogenase drives bacterial l-serine synthesis.

l-Serine biosynthesis, a crucial metabolic process in most domains of life, is initiated by d-3-phosphoglycerate (d-3-PG) dehydrogenation, a thermodynamically unfavorable reaction catalyzed by d-3-PG dehydrogenase (SerA). d-2-Hydroxyglutarate (d-2-HG) is traditionally viewed as an abnormal metabolite associated with cancer and neurometabolic disorders. Here, we reveal that bacterial anabolism and catabolism of d-2-HG are involved in l-serine biosynthesis in Pseudomonas stutzeri A1501 and Pseudomonas aeruginosa PAO1. SerA catalyzes the stereospecific reduction of 2-ketoglutarate (2-KG) to d-2-HG, responsible for the major production of d-2-HG in vivo. SerA combines the energetically favorable reaction of d-2-HG production to overcome the thermodynamic barrier of d-3-PG dehydrogenation. We identified a bacterial d-2-HG dehydrogenase (D2HGDH), a flavin adenine dinucleotide (FAD)-dependent enzyme, that converts d-2-HG back to 2-KG. Electron transfer flavoprotein (ETF) and ETF-ubiquinone oxidoreductase (ETFQO) are also essential in d-2-HG metabolism through their capacity to transfer electrons from D2HGDH. Furthermore, while the mutant with D2HGDH deletion displayed decreased growth, the defect was rescued by adding l-serine, suggesting that the D2HGDH is functionally tied to l-serine synthesis. Substantial flux flows through d-2-HG, being produced by SerA and removed by D2HGDH, ETF, and ETFQO, maintaining d-2-HG homeostasis. Overall, our results uncover that d-2-HG-mediated coupling between SerA and D2HGDH drives bacterial l-serine synthesis.

Measurement of red blood cell 6-thioguanine nucleotide is beneficial in azathioprine maintenance therapy of Chinese Crohn's disease patients.

OBJECTIVE: It remains controversial whether 6-thioguanine nucleotide (6-TGN)-based dose adjusting can be beneficial in azathioprine (AZA) therapy. This study is designed to assess the role of 6-TGN concentrations in maintaining clinical remission in Chinese patients with Crohn's disease (CD). MATERIAL AND METHOD: We performed a prospective observational study and collected data of CD patients in the First Affiliated Hospital of Anhui Medical University from June 2013 to April 2014. Demographic material, CD activity index, 6-TGN concentration, and laboratory tests were recorded at baseline and at each visit. In addition, 6-TGN was measured when drug adverse effects occurred. All patients achieved maintenance stage were administered a stable AZA dose at least 3 months before enrollment and were followed up at least 12 months. Thiopurine S-methyltransferase (TPMT) genotype was measured before AZA treatment. RESULTS: Sixty-nine patients receiving maintenance therapy were analyzed. A positive correlation was found between 6-TGN levels and AZA dose (r = 0.258, p = 0.032). The mean 6-TGN concentration was 302.06 ± 115.84 in the remission group vs. 264.94 ± 164.53 pmol/8 × 10(8) RBC in those with active disease (t = 0.847, p = 0.40), and 197.74 ± 66.54 pmol/8 × 10(8) RBC in patients who relapsed vs. 310.26 ± 122.38 pmol/8 × 10(8) RBC for those in sustained remission (t= -2.541, p = 0.013). In the leukopenia group, the 6-TGN concentration was 469.11 ± 115.53 pmol/8 × 10(8) RBC vs. 257.31 ± 83.74 pmol/8 × 10(8) RBC in the non-leukopenia group (t = 7.622, p < 0.001). There was a significant negative correlation between leukocyte count and 6-TGN concentration (r= -0.326, p = 0.006). CONCLUSIONS: 6-TGN measurement is a helpful method of preventing disease relapse and avoiding leukopenia in individual azathioprine maintenance therapy.

Production of diacetyl by metabolically engineered Enterobacter cloacae.

Diacetyl, a high value product that can be extensively used as a food ingredient, could be produced from the non-enzymatic oxidative decarboxylation of α-acetolactate during 2,3-butanediol fermentation. In this study, the 2,3-butanediol biosynthetic pathway in Enterobacter cloacae subsp. dissolvens strain SDM, a good candidate for microbial 2,3-butanediol production, was reconstructed for diacetyl production. To enhance the accumulation of the precursor of diacetyl, the α-acetolactate decarboxylase encoding gene (budA) was knocked out in strain SDM. Subsequently, the two diacetyl reductases DR-I (gdh) and DR-II (budC) encoding genes were inactivated in strain SDM individually or in combination to decrease the reduction of diacetyl. Although the engineered strain E. cloacae SDM (ΔbudAΔbudC) was found to have a good ability for diacetyl production, more α-acetolactate than diacetyl was produced simultaneously. In order to enhance the nonenzymatic oxidative decarboxylation of α-acetolactate to diacetyl, 20 mM Fe(3+) was added to the fermentation broth at the optimal time. In the end, by using the metabolically engineered strain E. cloacae SDM (ΔbudAΔbudC), diacetyl at a concentration of 1.45 g/L was obtained with a high productivity (0.13 g/(L·h)). The method developed here may be a promising process for biotechnological production of diacetyl.