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1.
Front Cardiovasc Med ; 11: 1342686, 2024.
Article in English | MEDLINE | ID: mdl-38562191

ABSTRACT

Background: Iron deficiency (ID) is the most common nutritional deficiency, with little research on its prevalence and long-term outcomes in the general population and those with heart failure (HF). Both the relationships between dietary iron and ID, as well as dietary folate and ID, are understudied. Methods: We used data from the National Health and Nutrition Examination Survey from 1999 to 2002 to investigate the prevalence, prognosis, and relationship between dietary and ID defined by different criteria in the general population (n = 6,660) and those with HF (n = 182). Results: There was no significant difference in the prevalence of ID between HF patients and the general population after propensity score matching. Transferrin saturation (TSAT) <20% was associated with higher 5-year all-cause mortality (HR: 3.49, CI: 1.40-8.72, P = 0.007), while ferritin <30 ng/ml was associated with higher 10-year (HR: 2.70, CI: 1.10-6.67, P = 0.031) and 15-year all-cause mortality (HR: 2.64, CI: 1.40-5.00, P = 0.003) in HF patients. Higher dietary total folate but dietary iron reduced the risk of ID (defined as ferritin <100 ng/ml) in HF patients (OR: 0.80; 95% CI: 0.65-1.00; P = 0.047). Conclusions: The prevalence of ID was identical in HF and non-HF individuals. Ferritin <30 ng/ml was associated with long-term outcomes whereas TSAT <20% was associated with short-term prognosis in both the general population and HF patients. A diet rich in folate might have the potential for prevention and treatment of ID in HF patients.

2.
Sheng Wu Gong Cheng Xue Bao ; 40(3): 621-643, 2024 Mar 25.
Article in Chinese | MEDLINE | ID: mdl-38545968

ABSTRACT

L-tryptophan is an essential amino acid that is widely used in food, medicine and feed sectors. L-tryptophan can be produced through fermentation, and the main producing strains are engineered Escherichia coli and Corynebacterium glutamicum, which are constructed by rational design methods based on metabolic engineering and synthetic biology. However, due to the long metabolic pathway, complex and unclear regulatory mechanism for L-tryptophan production in microbial cells, the production efficiency and robustness of L-tryptophan producing strains are still low. In this connection, irrational design methods such as laboratory adaptive evolution, are often applied to improve the performance of L-tryptophan producing strains. This review summarizes the recent progress on biosynthesis metabolism of L-tryptophan and its regulation, the construction and optimization of L-tryptophan producing strains, and fermentative production of L-tryptophan, and prospects future development perspective. This review may facilitate research and development for fermentative production of L-tryptophan.


Subject(s)
Corynebacterium glutamicum , Tryptophan , Fermentation , Metabolic Engineering , Metabolic Networks and Pathways , Corynebacterium glutamicum/genetics , Corynebacterium glutamicum/metabolism , Escherichia coli/genetics , Escherichia coli/metabolism
3.
Foods ; 13(5)2024 Feb 23.
Article in English | MEDLINE | ID: mdl-38472784

ABSTRACT

Research accumulated over the past decades has shown that mycoprotein could serve as a healthy and safe alternative protein source, offering a viable substitute for animal- and plant-derived proteins. This study evaluated the impact of substituting whey protein with fungal-derived mycoprotein at different levels (10%, 20%, and 30%) on the quality of high-protein nutrition bars (HPNBs). It focused on nutritional content, textural changes over storage, and sensory properties. Initially, all bars displayed similar hardness, but storage time significantly affected textural properties. In the early storage period (0-5 days), hardness increased at a modest rate of 0.206 N/day to 0.403 N/day. This rate dramatically escalated from 1.13 N/day to 1.36 N/day after 5 days, indicating a substantial textural deterioration over time. Bars with lower mycoprotein levels (10%) exhibited slower hardening rates compared with those with higher substitution levels (20% and 30%), pointing to a correlation between mycoprotein content and increased bar hardness during storage. Protein digestibility was assessed through in vitro gastric and intestinal phases. Bars with no or low-to-medium levels of mycoprotein substitution (PB00, PB10, and PB20) showed significantly higher digestibility (40.3~43.8%) compared with those with the highest mycoprotein content (PB30, 32.9%). However, digestibility rates for all mycoprotein-enriched bars were lower than those observed for whey-protein-only bars (PB00, 84.5%), especially by the end of the intestinal digestion phase. The introduction of mycoprotein enriched the bars' dietary fiber content and improved their odor, attributing a fresh mushroom-like smell. These findings suggest that modest levels of mycoprotein can enhance nutritional value and maintain sensory quality, although higher substitution levels adversely affect texture and protein digestibility. This study underscores the potential of mycoprotein as a functional ingredient in HPNBs, balancing nutritional enhancement with sensory acceptability, while also highlighting the challenges of textural deterioration and reduced protein digestibility at higher substitution levels.

4.
Metab Eng ; 83: 52-60, 2024 May.
Article in English | MEDLINE | ID: mdl-38521489

ABSTRACT

2-Pyrone-4,6-dicarboxylic acid (PDC), a chemically stable pseudo-aromatic dicarboxylic acid, is a promising building block compound for manufacturing biodegradable polyesters. This study aimed to construct high-performance cell factories enabling the efficient production of PDC from glucose. Firstly, the effective enzymes of the PDC biosynthetic pathway were overexpressed on the chromosome of the 3-dehydroshikimate overproducing strain. Consequently, the one-step biosynthesis of PDC from glucose was achieved. Further, the PDC production was enhanced by multi-copy integration of the key gene PsligC encoding 4-carboxy-2-hydroxymuconate-6-semialdehyde dehydrogenase and co-expression of Vitreoscilla hemoglobin. Subsequently, the PDC production was substantially improved by redistributing the metabolic flux for cell growth and PDC biosynthesis based on dynamically downregulating the expression of pyruvate kinase. The resultant strain PDC50 produced 129.37 g/L PDC from glucose within 78 h under fed-batch fermentation conditions, with a yield of 0.528 mol/mol and an average productivity of 1.65 g/L/h. The findings of this study lay the foundation for the potential industrial production of PDC.


Subject(s)
Escherichia coli , Metabolic Engineering , Polyesters , Pyrones , Escherichia coli/genetics , Escherichia coli/metabolism , Polyesters/metabolism , Pyrones/metabolism , Glucose/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Escherichia coli Proteins/genetics , Escherichia coli Proteins/metabolism , Dicarboxylic Acids/metabolism
5.
Front Pharmacol ; 15: 1267585, 2024.
Article in English | MEDLINE | ID: mdl-38414735

ABSTRACT

Myocardial infarction (MI) imposes a huge medical and economic burden on society, and cardiac repair after MI involves a complex series of processes. Understanding the key mechanisms (such as apoptosis, autophagy, inflammation, and fibrosis) will facilitate further drug development and patient treatment. Presently, a substantial body of evidence suggests that the regulation of epigenetic processes contributes to cardiac repair following MI, with DNA methylation being among the notable epigenetic factors involved. This article will review the research on the mechanism of DNA methylation regulation after MI to provide some insights for future research and development of related drugs.

6.
Nature ; 625(7995): 585-592, 2024 Jan.
Article in English | MEDLINE | ID: mdl-38200309

ABSTRACT

Oncogene-induced replication stress generates endogenous DNA damage that activates cGAS-STING-mediated signalling and tumour suppression1-3. However, the precise mechanism of cGAS activation by endogenous DNA damage remains enigmatic, particularly given that high-affinity histone acidic patch (AP) binding constitutively inhibits cGAS by sterically hindering its activation by double-stranded DNA (dsDNA)4-10. Here we report that the DNA double-strand break sensor MRE11 suppresses mammary tumorigenesis through a pivotal role in regulating cGAS activation. We demonstrate that binding of the MRE11-RAD50-NBN complex to nucleosome fragments is necessary to displace cGAS from acidic-patch-mediated sequestration, which enables its mobilization and activation by dsDNA. MRE11 is therefore essential for cGAS activation in response to oncogenic stress, cytosolic dsDNA and ionizing radiation. Furthermore, MRE11-dependent cGAS activation promotes ZBP1-RIPK3-MLKL-mediated necroptosis, which is essential to suppress oncogenic proliferation and breast tumorigenesis. Notably, downregulation of ZBP1 in human triple-negative breast cancer is associated with increased genome instability, immune suppression and poor patient prognosis. These findings establish MRE11 as a crucial mediator that links DNA damage and cGAS activation, resulting in tumour suppression through ZBP1-dependent necroptosis.


Subject(s)
Cell Transformation, Neoplastic , MRE11 Homologue Protein , Nucleosomes , Nucleotidyltransferases , Humans , Cell Proliferation , Cell Transformation, Neoplastic/metabolism , Cell Transformation, Neoplastic/pathology , DNA Damage , MRE11 Homologue Protein/metabolism , Necroptosis , Nucleosomes/metabolism , Nucleotidyltransferases/metabolism , Radiation, Ionizing , Triple Negative Breast Neoplasms/genetics , Triple Negative Breast Neoplasms/metabolism , Triple Negative Breast Neoplasms/pathology , Genomic Instability
7.
Crit Rev Biotechnol ; : 1-19, 2024 Jan 17.
Article in English | MEDLINE | ID: mdl-38232958

ABSTRACT

The market size of biosurfactants (BSs) has been expanding at an extremely fast pace due to their broad application scope. Therefore, the re-construction of cell factories with modified genomic and metabolic profiles for desired industrial performance has been an intriguing aspect. Typical mutagenesis approaches generate huge mutant libraries, whereas a battery of specific, robust, and cost-effective high-throughput screening (HTS) methods is requisite to screen target strains for desired phenotypes. So far, only a few specialized HTS assays have been developed for BSs that were successfully applied to obtain anticipated mutants. The most important milestones to reach, however, continue to be: specificity, sensitivity, throughput, and the potential for automation. Here, we discuss important colorimetric and fluorometric HTS approaches for possible intervention on automated HTS platforms. Moreover, we explain current bottlenecks in developing specialized HTS platforms for screening high-yielding producers and discuss possible perspectives for addressing such challenges.

8.
J Agric Food Chem ; 72(1): 604-612, 2024 Jan 10.
Article in English | MEDLINE | ID: mdl-38153978

ABSTRACT

The global protein shortage is intensifying, and promising means to ensure daily protein supply are desperately needed. The mycoprotein produced by Fusarium venenatum is a good alternative to animal/plant-derived protein. To comprehensively improve the mycoprotein synthesis, a stepwise strategy by blocking the byproduct ethanol synthesis and the gluconeogenesis pathway and by optimizing the fermentation medium was herein employed. Ultimately, compared to the wild-type strain, the synthesis rate, carbon conversion ratio, and protein content of mycoprotein produced from the engineered strain were increased by 57% (0.212 vs 0.135 g/L·h), 62% (0.351 vs 0.217 g/g), and 57% (61.9 vs 39.4%), respectively, accompanied by significant reductions in CO2 emissions. These results provide a referential strategy that could be useful for improving mycoprotein synthesis in other fungi; more importantly, the obtained high-mycoprotein-producing strain has the potential to promote the development of the edible protein industry and compensate for the gap in protein resources.


Subject(s)
Carbon Dioxide , Fusarium , Animals , Fermentation , Metabolic Engineering
9.
Clin Pharmacol ; 15: 113-123, 2023.
Article in English | MEDLINE | ID: mdl-38090388

ABSTRACT

Background: Dronedarone is an effective drug for maintaining the sinus rhythm in patients with atrial fibrillation (AF). The efficacy and safety of dronedarone versus amiodarone in patients with AF after catheter ablation (CA) needs more evidence. We retrospectively compared the efficacy and safety of dronedarone and amiodarone in our hospital. Methods: Patients who underwent CA from January 2021 to January 2022 and used dronedarone (n=229) or amiodarone (n=202) during the blind period were enrolled. The recurrence of AF in post-and during the blanking period was compared between the groups; the rehospitalization for re-ablation and adverse drug events (ADE) were also calculated. Results: During an average follow-up period of 14.28 months, the long-term recurrence rate of AF did not differ significantly between the amiodarone group and dronedarone group (22.71% vs 21.29%, hazard ratio [HR], 1.033, 95% confidence interval [CI], 0.661-1.614; p=0.888). The recurrence rate in the blanking period also showed no statistically significant differences between the amiodarone group and dronedarone group (9.90% vs 14.41%, HR, 0.851; 95% CI, 0.463-1.564; p=0.604). The re-hospitalization rates for re-ablation between two groups did not differ between the amiodarone group and dronedarone group (4.65% vs 13.46%; p =0.144). The incidence of ADE was higher in the dronedarone groups than that in the amiodarone group (16.59% vs 5.45%, p <0.001). The main adverse drug events in the dronedarone and amiodarone groups were gastrointestinal (6.99%) and bradycardia (2.48%), respectively. Conclusion: Compared to the amiodarone group, the dronedarone group had a similar blank-period and long-term recurrence rate of AF and a higher incidence of ADE.

10.
J Med Chem ; 66(20): 14133-14149, 2023 10 26.
Article in English | MEDLINE | ID: mdl-37782247

ABSTRACT

Methyl-lysine reader p53 binding protein 1 (53BP1) is a central mediator of DNA break repair and is associated with various human diseases, including cancer. Thus, high-quality 53BP1 chemical probes can aid in further understanding the role of 53BP1 in genome repair pathways. Herein, we utilized focused DNA-encoded library screening to identify the novel hit compound UNC8531, which binds the 53BP1 tandem Tudor domain (TTD) with an IC50 of 0.47 ± 0.09 µM in a TR-FRET assay and Kd values of 0.85 ± 0.17 and 0.79 ± 0.52 µM in ITC and SPR, respectively. UNC8531 was cocrystallized with the 53BP1 TTD to guide further optimization efforts, leading to UNC9512. NanoBRET and 53BP1-dependent foci formation experiments confirmed cellular target engagement. These results show that UNC9512 is a best-in-class small molecule 53BP1 antagonist that can aid further studies investigating the role of 53BP1 in DNA repair, gene editing, and oncogenesis.


Subject(s)
DNA Repair , Intracellular Signaling Peptides and Proteins , Humans , DNA , Intracellular Signaling Peptides and Proteins/metabolism , Tumor Suppressor p53-Binding Protein 1/chemistry , Tumor Suppressor p53-Binding Protein 1/genetics , Tumor Suppressor p53-Binding Protein 1/metabolism , Tudor Domain
11.
Microb Cell Fact ; 22(1): 172, 2023 Sep 04.
Article in English | MEDLINE | ID: mdl-37667234

ABSTRACT

BACKGROUND: Pyruvate is a widely used value-added chemical which also serves as a hub of various metabolic pathways. The fastest-growing bacterium Vibrio natriegens is a promising chassis for synthetic biology applications with high substrate uptake rates. The aim of this study was to investigate if the high substrate uptake rates of V. natriegens enable pyruvate production at high productivities. RESULTS: Two prophage gene clusters and several essential genes for the biosynthesis of byproducts were first deleted. In order to promote pyruvate accumulation, the key gene aceE encoding pyruvate dehydrogenase complex E1 component was down-regulated to reduce the carbon flux into the tricarboxylic acid cycle. Afterwards, the expression of ppc gene encoding phosphoenolpyruvate carboxylase was fine-tuned to balance the cell growth and pyruvate synthesis. The resulting strain PYR32 was able to produce 54.22 g/L pyruvate from glucose within 16 h, with a yield of 1.17 mol/mol and an average productivity of 3.39 g/L/h. In addition, this strain was also able to efficiently convert sucrose or gluconate into pyruvate at high titers. CONCLUSION: A novel strain of V. natriegens was engineered which was capable to provide higher productivity in pyruvate synthesis. This study lays the foundation for the biosynthesis of pyruvate and its derivatives in fast-growing V. natriegens.


Subject(s)
Pyruvic Acid , Vibrio , Metabolic Engineering , Vibrio/genetics , Biological Transport
12.
Sheng Wu Gong Cheng Xue Bao ; 39(8): 3290-3301, 2023 Aug 25.
Article in Chinese | MEDLINE | ID: mdl-37622361

ABSTRACT

Salicylate 2-O-ß-d-glucoside (SAG) is a derivative of salicylate in plants. Recent reports showed that SAG could be considered as a potential anti-inflammatory substance due to its anti-inflammatory and analgesic effects, and less irritation compared with salicylic acid and aspirin. The biological method uses renewable resources to produce salicylic acid compounds, which is more environmentally friendly than traditional industry methods. In this study, Escherichia coli Tyr002 was used as the starting strain, and a salicylic acid producing strain of E. coli was constructed by introducing the isochorismate pyruvate lyase gene pchB from Pseudomonas aeruginosa. By regulating the expression of the key genes in the downstream aromatic amino acid metabolic pathways, the titer of salicylic acid reached 1.05 g/L in shake flask fermentation. Subsequently, an exogenous salicylic acid glycosyltransferase was introduced into the salicylic acid producing strain to glycosylate the salicylic acid. The newly engineered strain produced 5.7 g/L SAG in shake flask fermentation. In the subsequent batch fed fermentation in a 5 L fermentation tank, the titer of SAG reached 36.5 g/L, which is the highest titer reported to date. This work provides a new route for biosynthesis of salicylate and its derivatives.


Subject(s)
Escherichia coli , Glucosides , Escherichia coli/genetics , Metabolic Engineering , Salicylic Acid , Pyruvic Acid
13.
Biotechnol Biofuels Bioprod ; 16(1): 92, 2023 Jun 01.
Article in English | MEDLINE | ID: mdl-37264438

ABSTRACT

BACKGROUND: 2-Pyrone-4,6-dicarboxylic acid (PDC), a chemically stable pseudoaromatic dicarboxylic acid, represents a promising building block for the manufacture of biodegradable polyesters. Microbial production of PDC has been extensively investigated, but low titers and yields have limited industrial applications. RESULTS: In this study, a multi-step biosynthesis strategy for the microbial production of PDC was demonstrated using engineered Escherichia coli whole-cell biocatalysts. The PDC biosynthetic pathway was first divided into three synthetic modules, namely the 3-dehydroshikimic acid (DHS) module, the protocatechuic acid (PCA) module and the PDC module. Several effective enzymes, including 3-dehydroshikimate dehydratase for the PCA module as well as protocatechuate 4,5-dioxygenase and 4-carboxy-2-hydroxymuconate-6-semialdehyde dehydrogenase for the PDC module were isolated and characterized. Then, the highly efficient whole-cell bioconversion systems for producing PCA and PDC were constructed and optimized, respectively. Finally, the efficient multi-step biosynthesis of PDC from glucose was achieved by smoothly integrating the above three biosynthetic modules, resulting in a final titer of 49.18 g/L with an overall 27.2% molar yield, which represented the highest titer for PDC production from glucose reported to date. CONCLUSIONS: This study lays the foundation for the microbial production of PDC, including one-step de novo biosynthesis from glucose as well as the microbial transformation of monoaromatics.

14.
Biotechnol Biofuels Bioprod ; 16(1): 37, 2023 Mar 04.
Article in English | MEDLINE | ID: mdl-36870984

ABSTRACT

BACKGROUND: Owing to the Crabtree effect, Saccharomyces cerevisiae produces a large amount of ethanol in the presence of oxygen and excess glucose, leading to a loss of carbon for the biosynthesis of non-ethanol chemicals. In the present study, the potential of a newly constructed Crabtree negative S. cerevisiae, as a chassis cell, was explored for the biosynthesis of various non-ethanol compounds. RESULTS: To understand the metabolic characteristics of Crabtree negative S. cerevisiae sZJD-28, its transcriptional profile was compared with that of Crabtree positive S. cerevisiae CEN.PK113-11C. The reporter GO term analysis showed that, in sZJD-28, genes associated with translational processes were down-regulated, while those related to carbon metabolism were significantly up-regulated. To verify a potential increase in carbon metabolism for the Crabtree negative strain, the production of non-ethanol chemicals, derived from different metabolic nodes, was then undertaken for both sZJD-28 and CEN.PK113-11C. At the pyruvate node, production of 2,3-butanediol and lactate in sZJD-28-based strains was remarkably higher than that of CEN.PK113-11C-based ones, representing 16.8- and 1.65-fold increase in titer, as well as 4.5-fold and 0.65-fold increase in specific titer (mg/L/OD), respectively. Similarly, for shikimate derived p-coumaric acid, the titer of sZJD-28-based strain was 0.68-fold higher than for CEN.PK113-11C-based one, with a 0.98-fold increase in specific titer. While farnesene and lycopene, two acetoacetyl-CoA derivatives, showed 0.21- and 1.88-fold increases in titer, respectively. From malonyl-CoA, the titer of 3-hydroxypropionate and fatty acids in sZJD-28-based strains were 0.19- and 0.76-fold higher than that of CEN.PK113-11C-based ones, respectively. In fact, yields of products also improved by the same fold due to the absence of residual glucose. Fed-batch fermentation further showed that the titer of free fatty acids in sZJD-28-based strain 28-FFA-E reached 6295.6 mg/L with a highest reported specific titer of 247.7 mg/L/OD in S. cerevisiae. CONCLUSIONS: Compared with CEN.PK113-11C, the Crabtree negative sZJD-28 strain displayed a significantly different transcriptional profile and obvious advantages in the biosynthesis of non-ethanol chemicals due to redirected carbon and energy sources towards metabolite biosynthesis. The findings, therefore, suggest that a Crabtree negative S. cerevisiae strain could be a promising chassis cell for the biosynthesis of various chemicals.

15.
Front Bioeng Biotechnol ; 11: 1108653, 2023.
Article in English | MEDLINE | ID: mdl-36845173

ABSTRACT

Yarrowia lipolytica has been widely used in the food biotech-related industry, where it plays the host's role in producing erythritol. Nevertheless, a temperature of about 28°C-30°C has been estimated as the yeast's optimal growth temperature, leading to the consumption of a considerable quantity of cooling water, especially in summer, which is obligatory for fermentation. Herein is described a method for improving the thermotolerance and erythritol production efficiency at high temperatures of Y. lipolytica. Through screening and testing different heat resistant devices, eight refactored engineered strains showed better growth at higher temperature and the antioxidant properties of the eight engineered strains were also improved. In addition, the erythritol titer, yield and productivity of the strain FOS11-Ctt1 represented the best among the eight strains, reaching at 39.25 g/L, 0.348 g/g glucose, and 0.55 g/L/h respectively, which were increased by 156%, 86% and 161% compared with the control strain, respectively. This study provides insight into an effective heat-resistant device that could enhance the thermotolerance and erythritol production of Y. lipolytica, which might be considered a valued scientific reference for other resistant strains' construction.

16.
Biotechnol Adv ; 64: 108105, 2023.
Article in English | MEDLINE | ID: mdl-36736865

ABSTRACT

Sugar alcohols are polyols that are widely employed in the production of chemicals, pharmaceuticals, and food products. Chemical synthesis of polyols, however, is complex and necessitates the use of hazardous compounds. Therefore, the use of microbes to produce polyols has been proposed as an alternative to traditional synthesis strategies. Many biotechnological approaches have been described to enhancing sugar alcohols production and microbe-mediated sugar alcohol production has the potential to benefit from the availability of inexpensive substrate inputs. Among of them, microbe-mediated erythritol production has been implemented in an industrial scale, but microbial growth and substrate conversion rates are often limited by harsh environmental conditions. In this review, we focused on xylitol, mannitol, sorbitol, and erythritol, the four representative sugar alcohols. The main metabolic engineering strategies, such as regulation of key genes and cofactor balancing, for improving the production of these sugar alcohols were reviewed. The feasible strategies to enhance the stress tolerance of chassis cells, especially thermotolerance, were also summarized. Different low-cost substrates like glycerol, molasses, cellulose hydrolysate, and CO2 employed for producing these sugar alcohols were presented. Given the value of polyols as precursor platform chemicals that can be leveraged to produce a diverse array of chemical products, we not only discuss the challenges encountered in the above parts, but also envisioned the development of their derivatives for broadening the application of sugar alcohols.


Subject(s)
Sugar Alcohols , Sugars , Sugar Alcohols/metabolism , Xylitol/metabolism , Mannitol/metabolism , Erythritol/metabolism
17.
Synth Syst Biotechnol ; 8(1): 141-147, 2023 Mar.
Article in English | MEDLINE | ID: mdl-36687472

ABSTRACT

CRISPR/Cas9-mediated homology-directed recombination is an efficient method to express target genes. Based on the above method, providing ideal neutral integration sites can ensure the reliable, stable, and high expression of target genes. In this study, we obtained a fluorescent transformant with neutral integration and high expression of the GFP expression cassette from the constructed GFP expression library and named strain FS. The integration site mapped at 4886 bp upstream of the gene FVRRES_00686 was identified in strain FS based on a Y-shaped adaptor-dependent extension, and the sequence containing 600 bp upstream and downstream of this site was selected as the candidate region for designing sgRNAs (Sites) for CRISPR/Cas9-mediated homology-directed recombination. PCR analysis showed that the integration efficiency of CRISPR/Cas9-mediated integration of target genes in designed sites reached 100%. Further expression stability and applicability analysis revealed that the integration of the target gene into the above designed sites can be stably inherited and expressed and has no negative effect on the growth of F. venenatum TB01. These results indicate the above designed neutral sites have the potential to accelerate the development of F. venenatum TB01 through overexpression of target genes in metabolic engineering.

18.
Electrophoresis ; 44(9-10): 854-863, 2023 05.
Article in English | MEDLINE | ID: mdl-36645159

ABSTRACT

Recent research has questioned the validity of housekeeping proteins in Western blot. Our present study proposed new ideas for Western blot normalization that improved the reproducibility of scientific research. We used the Gene Expression Omnibus (GEO) database and the web tool GEO2R to exclude unstable housekeeping genes quickly. In ischemic heart tissues, actin and tubulin changed significantly, whereas no statistically significant changes were observed in the expression of genes relative to glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Besides, the reliability of GAPDH was further examined by Western blot. Additionally, unstable housekeeping genes were found in other animal models of cardiovascular medicine. We also found that sodium dodecyl sulfate and temperature significantly impacted the results of Ponceau S staining. Membranes stained with Ponceau S after immunodetection could avoid this interference, and the coefficients of variation for post-immunodetection staining are lower than those produced by GAPDH immunodetection. Overall, we described a new use of differential gene expression analysis and proposed a modified Ponceau S staining method, which provided researchers with a proper loading control for Western blot and hence could improve reproducibility in research.


Subject(s)
Actins , Glyceraldehyde-3-Phosphate Dehydrogenases , Animals , Reproducibility of Results , Actins/analysis , Glyceraldehyde-3-Phosphate Dehydrogenases/genetics , Glyceraldehyde-3-Phosphate Dehydrogenases/analysis , Glyceraldehyde-3-Phosphate Dehydrogenases/metabolism , Staining and Labeling , Blotting, Western
19.
Front Cardiovasc Med ; 9: 1037176, 2022.
Article in English | MEDLINE | ID: mdl-36386380

ABSTRACT

Over the last few decades, catheter ablation has emerged as the first-line treatment for ventricular arrhythmias. However, detailed knowledge of cardiac anatomy during the surgery remains the prerequisite for successful ablation. Intracardiac echocardiography (ICE) is a unique imaging technique, which provides real-time visualization of cardiac structures, and is superior to other imaging modalities in terms of precise display of cardiac tissue characteristics as well as the orientation of anatomical landmarks. This article aimed to introduce the various advantages and limitations of ICE in the ablation of ventricular arrhythmias.

20.
Front Cardiovasc Med ; 9: 1027000, 2022.
Article in English | MEDLINE | ID: mdl-36426220

ABSTRACT

Background: Acetaldehyde dehydrogenase 2 (ALDH2) is an essential enzyme in alcohol metabolism, playing a vital function in resisting oxidative stress. Lots of gene variants have been associated with atrial fibrillation (AF), among which the association between ALDH2 rs671 polymorphism and AF is variable. This study aimed to investigate the relationship between ALDH2 rs671 polymorphism and AF occurrence or progression and AF recurrence after catheter ablation. Methods: A total of 924 subjects were enrolled in the study. The ALDH2 genotypes are composed of wild-type homozygotes (ALDH2*1/*1), heterozygotes (ALDH2*1/*2), and mutant homozygotes (ALDH2*2/*2), in which the genotypes ALDH2*1/*2 and ALDH2*2/*2 are combined into the ALDH2*2. Univariate and multivariate logistic regression analyses were performed to investigate the association between ALDH2*2 and AF occurrence and progression. COX regression analysis was used to explore the association of ALDH2*2 with AF recurrence after catheter ablation. Results: The prevalence of AF differed significantly between the ALDH2*2 group (102/251) and ALDH2*1/*1 group (330/673) (P = 0.023). For AF occurrence, in the univariate analysis, alcohol consumption was a risk factors (OR: 1.503, P = 0.003), whereas ALDH2*2 was a protective factor (OR: 0.712, P = 0.023). In the multivariate analysis, alcohol consumption (P = 0.156) and ALDH2*2 (P = 0.096) were no longer independent factors. ALDH2*2 with non-drinking was associated with a decreased AF occurrence (OR: 0.65, P = 0.021), whereas ALDH2*2 with drinking was not (P = 0.365). For AF progression, multivariate analysis revealed ALDH2*2 could promote persistent AF in female AF patients (OR: 2.643, P = 0.008). Cox regression analysis suggested that ALDH2*2 (P = 0.752) was not a risk factor for AF recurrence after catheter ablation during a median 6 months follow-up. Conclusion: While ALDH2*2 was not directly related to AF, ALDH2*2 with non-drinking was associated with a decreased incidence of AF. ALDH2*2 may accelerate AF progression in female patients, increasing the likelihood of developing persistent AF. Therefore, individuals with ALDH2*2 should refrain from consuming alcohol to decrease the onset and progression of AF.

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