Low-Dose Colchicine Ameliorates Doxorubicin Cardiotoxicity Via Promoting Autolysosome Degradation.

BACKGROUND: The only clinically approved drug that reduces doxorubicin cardiotoxicity is dexrazoxane, but its application is limited due to the risk of secondary malignancies. So, exploring alternative effective molecules to attenuate its cardiotoxicity is crucial. Colchicine is a safe and well-tolerated drug that helps reduce the production of reactive oxygen species. High doses of colchicine have been reported to block the fusion of autophagosomes and lysosomes in cancer cells. However, the impact of colchicine on the autophagy activity within cardiomyocytes remains inadequately elucidated. Recent studies have highlighted the beneficial effects of colchicine on patients with pericarditis, postprocedural atrial fibrillation, and coronary artery disease. It remains ambiguous how colchicine regulates autophagic flux in doxorubicin-induced heart failure. METHODS AND RESULTS: Doxorubicin was administered to establish models of heart failure both in vivo and in vitro. Prior studies have reported that doxorubicin impeded the breakdown of autophagic vacuoles, resulting in damaged mitochondria and the accumulation of reactive oxygen species. Following the administration of a low dose of colchicine (0.1 mg/kg, daily), significant improvements were observed in heart function (left ventricular ejection fraction: doxorubicin group versus treatment group=43.75%±3.614% versus 57.07%±2.968%, P=0.0373). In terms of mechanism, a low dose of colchicine facilitated the degradation of autolysosomes, thereby mitigating doxorubicin-induced cardiotoxicity. CONCLUSIONS: Our research has shown that a low dose of colchicine is pivotal in restoring the autophagy activity, thereby attenuating the cardiotoxicity induced by doxorubicin. Consequently, colchicine emerges as a promising therapeutic candidate to improve doxorubicin cardiotoxicity.

Ginsenoside Rb1 attenuates doxorubicin induced cardiotoxicity by suppressing autophagy and ferroptosis.

Ginsenoside Rb1 (Rb1), an active component isolated from traditional Chinese medicine Ginseng, is beneficial to many cardiovascular diseases. However, whether it can protect against doxorubicin induced cardiotoxicity (DIC) is not clear yet. In this study, we aimed to investigate the role of Rb1 in DIC. Mice were injected with a single dose of doxorubicin (20 mg/kg) to induce acute cardiotoxicity. Rb1 was given daily gavage to mice for 7 days. Changes in cardiac function, myocardium histopathology, oxidative stress, cardiomyocyte mitochondrion morphology were studied to evaluate Rb1's function on DIC. Meanwhile, RNA-seq analysis was performed to explore the potential underline molecular mechanism involved in Rb1's function on DIC. We found that Rb1 treatment can improve survival rate and body weight in Dox treated mice group. Rb1 can attenuate Dox induced cardiac dysfunction and myocardium hypertrophy and interstitial fibrosis. The oxidative stress increase and cardiomyocyte mitochondrion injury were improved by Rb1 treatment. Mechanism study found that Rb1's beneficial role in DIC is through suppressing of autophagy and ferroptosis. This study shown that Ginsenoside Rb1 can protect against DIC by regulating autophagy and ferroptosis.

PLEKHM2 deficiency induces impaired mitochondrial clearance and elevated ROS levels in human iPSC-derived cardiomyocytes.

Pleckstrin homology domain-containing family M member 2 (PLEKHM2) is an essential adaptor for lysosomal trafficking and its homozygous truncation have been reported to cause early onset dilated cardiomyopathy (DCM). However, the molecular mechanism of PLEKHM2 deficiency in DCM pathogenesis and progression is poorly understood. Here, we generated an in vitro model of PLEKHM2 knockout (KO) induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) to elucidate the potential pathogenic mechanism of PLEKHM2-deficient cardiomyopathy. PLEKHM2-KO hiPSC-CMs developed disease phenotypes with reduced contractility and impaired calcium handling. Subsequent RNA sequencing (RNA-seq) analysis revealed altered expression of genes involved in mitochondrial function, autophagy and apoptosis in PLEKHM2-KO hiPSC-CMs. Further molecular experiments confirmed PLEKHM2 deficiency impaired autophagy and resulted in accumulation of damaged mitochondria, which triggered increased reactive oxygen species (ROS) levels and decreased mitochondrial membrane potential (Δψm). Importantly, the elevated ROS levels caused oxidative stress-induced damage to nearby healthy mitochondria, resulting in extensive Δψm destabilization, and ultimately leading to impaired mitochondrial function and myocardial contractility. Moreover, ROS inhibition attenuated oxidative stress-induced mitochondrial damage, thereby partially rescued PLEKHM2 deficiency-induced disease phenotypes. Remarkably, PLEKHM2-WT overexpression restored autophagic flux and rescued mitochondrial function and myocardial contractility in PLEKHM2-KO hiPSC-CMs. Taken together, these results suggested that impaired mitochondrial clearance and increased ROS levels play important roles in PLEKHM2-deficient cardiomyopathy, and PLEKHM2-WT overexpression can improve mitochondrial function and rescue PLEKHM2-deficient cardiomyopathy.

Clinical features of Danon disease and insights gained from LAMP-2 deficiency models.

Danon disease (DD) is an X-linked multisystem disorder with clinical features characterized by the triad of hypertrophic cardiomyopathy, skeletal muscle weakness, and mental retardation. Cardiac involvement can be fatal in the absence of an effective treatment option such as heart transplantation. Molecular studies have proved that LAMP-2 protein deficiency, mainly LAMP-2B isoform, resulting from LAMP2 gene mutation, is the culprit for DD. Autophagy impairment due to LAMP-2 deficiency mediated the accumulation of abnormal autophagic vacuoles in cells. While it is not ideal for mimicking DD phenotypes in humans, the emergence of LAMP-2-deficient animal models and induced pluripotent stem cells from DD patients provided powerful tools for exploring DD mechanism. In both in vitro and in vivo studies, much evidence has demonstrated that mitochondria dysfunction and fragmentation can result in DD pathology. Fundamental research contributes to the therapeutic transformation. By targeting the molecular core, several potential therapies have demonstrated promising results in partial phenotypes improvement. Among them, gene therapies anticipate inaugurate a class of symptom control and prevention drugs as their in vivo effects are promising, and one clinical trial is currently underway.

Impact of advanced liver fibrosis on atrial fibrillation recurrence after ablation in non-alcoholic fatty liver disease patients.

Aim: Advanced liver fibrosis is independently associated with new onset of atrial fibrillation (AF). Non-invasive liver fibrosis scores are considered an effective strategy for assessing liver fibrosis. This study aimed to investigate the association between advanced liver fibrosis and AF recurrence after ablation in patients with non-alcoholic fatty liver disease (NAFLD). Materials and methods: A total of 345 AF patients with NAFLD who underwent de novo ablation between 2019 and 2020 at two large hospitals in China were included in this study. AF recurrence was defined as the occurrence of atrial arrhythmia for more than 30 s by electrocardiogram or 24 h Holter monitoring after the first 3 months of ablation. Predictive values of non-alcoholic fatty liver disease fibrosis score (NFS) and Fibrosis-4 (FIB-4) scores for AF burden and recurrence after ablation were assessed. Results: At the 1 year follow-up after ablation, 38.8% of patients showed recurrence. Patients with recurrence who had higher FIB-4 and NFS scores were more likely to have persistent AF and a duration of AF ≥ 3 years. In Kaplan-Meier analysis, patients with intermediate and high NFS and FIB-4 risk categories had a higher risk of AF recurrence. Compared to patients with the low risk, intermediate and high NFS, and FIB-4 risk were independently associated with AF recurrence in multivariate Cox regression analysis (high risk: NFS, hazard ratio (HR): 3.11, 95% confidence interval (CI): 1.68∼5.76, p < 0.001; FIB-4, HR: 3.91, 95% CI: 2.19∼6.98, p < 0.001; intermediate risk: NFS, HR: 1.85, 95% CI: 1.10∼3.10, p = 0.020; FIB-4, HR: 2.08, 95% CI: 1.27∼3.41, p = 0.003). Conclusion: NFS and FIB-4 scores for advanced liver fibrosis are associated with AF burden. Advanced liver fibrosis is independently associated with AF recurrence following ablation. Advanced liver fibrosis might be meaningful in risk classification for patients after AF ablation.

Generation of an iPSC line (ZZUNEUi021-A) from a hypertrophic cardiomyopathy patient with TNNT2 mutation.

A 25-years-old hypertrophic cardiomyopathy male patient donated his peripheral blood mononuclear cells (PBMCs) with heterozygote mutation in theTNNT2 gene. We generated induced pluripotent stem cell (iPSC) with normal karyotypic and expressing NANOG, Lin28, GDF3 and DNMT3. The iPSC line has demonstrated pluripotency by differentiating into three germ layers in vitro. The ZZUNEUi021-A would serve as an in vitro model for loss of TNNT2 function.

Effects of ultraviolet-C light-emitting diodes at 275 nm on inactivation of Alicyclobacillusacidoterrestris vegetative cells and its spores as well as the quality attributes of orange juice.

Alicyclobacillus acidoterrestris is a thermoacidophilic, spore-forming bacillus. A. acidoterrestris and its spores can survive in pasteurized juices and cause microbial spoilage. In this work, the effects of ultraviolet-C light-emitting diodes at 275 nm on the inactivation of A. acidoterrestris vegetative cells and its spores in commercial pasteurized orange juice were studied. Meanwhile, the effects of ultraviolet-C light-emitting diodes on the quality attributes of the orange juice were also investigated. The quantities of A. acidoterrestris vegetative cells and its spores inoculated in orange juice were reduced by 6.04 and 2.49 log10 CFU/mL after ultraviolet-C light-emitting diode treatment at 220 mJ/cm2, respectively. The Weibull and Weibull plus tail models were satisfactorily fitted to estimate the reductions of A. acidoterrestris vegetative cells and its spores in orange juice, respectively. Physicochemical properties (pH, titratable acidity, total soluble solids, and clarity) of orange juice did not change significantly after exposure to ultraviolet-C light-emitting diodes. However, the total phenolic content of orange juice decreased with increasing fluence. In addition, ultraviolet-C light-emitting diode treatment at a higher fluence led to a noticeable color difference. These results indicate that ultraviolet-C light-emitting diode treatment has a potential application in the juice processing industry.

Effects of atmospheric pressure plasma jet on the physicochemical, functional, and antioxidant properties of flaxseed protein.

The aim of this study was to explore the effect of atmospheric pressure plasma jet (APPJ) on the physicochemical, functional, and antioxidant properties of flaxseed protein following APPJ treatment (0 to 240 s). The results showed that the pH value continuously dropped with the minimum value of 3.45 ± 0.15 after 240 s of APPJ treatment (-61.7%, P < 0.05). The relative protein solubility significantly declined after 15 s of APPJ treatment (-43.1%, P < 0.05), which was accompanied by the evident increase in mean particle size of flaxseed protein in aqueous solution (+157%, P < 0.05). Moreover, the surface hydrophobicity and contents of disulfide bonds gradually raised when the APPJ exposure time extended from 30 to 240 s. Notably, the foaming, emulsifying, and in vitro antioxidant properties of flaxseed protein were significantly improved following short time of APPJ treatment (5 to 15 s), which was paralleled with the changes of spatial conformation, mild protein oxidation, as well as the release of phenolic acids and flavonoids from naturally occurring protein-phenolic complex. Our findings elucidated that APPJ may be considered as an effective strategy to improve the functionality and antioxidant activities of flaxseed protein. PRACTICAL APPLICATION: We had evaluated the effect of APPJ treatment on the physicochemical, functional, and antioxidant properties of flaxseed protein, which was conducive to tailor flaxseed protein with the optimal techno-functionality and antioxidant activities as a potential nano-delivery vehicle.

Effects of thermal processing on the structural and functional properties of soluble dietary fiber from whole grain oats.

Normal pressure steaming, high pressure steaming, microwave, and frying are widely used to deactivate enzyme in the oats, but these thermal processing methods may affect the structural and functional properties of soluble dietary fiber, which contribute greatly to the health benefits of oat foods. The objective of this study was to evaluate the effects of four different thermal processing methods on the structural and functional properties of soluble dietary fiber from whole grain oats. The results showed that the thermal processing resulted in changes on nutritional components of whole grain oats. Especially dietary fiber components, the total dietary fiber, insoluble dietary fiber, and soluble dietary fiber content of heat-treated oats were significantly increased ( p < 0.05). Moreover, thermal processing can not only result in an increase in molecular weight and particle size, but also cause molecular aggregation and different functional properties of soluble dietary fiber. High pressure steaming-treated oat soluble dietary fiber displayed significantly higher swelling and emulsifying ( p < 0.05), but microwave-treated oat soluble dietary fiber exhibited the highest glucose, cholesterol, and sodium cholate adsorption capacities. These results might provide basic information to help to better understand the functionality of oat soluble dietary fiber and improve the process efficiency of oat foods with high nutritional qualities.

Inactivation of soybean trypsin inhibitor by dielectric-barrier discharge (DBD) plasma.

Soybean trypsin inhibitor (STI) is considered as one of the most important anti-nutritional factors in soybeans. The objective of this study was to investigate the impacts and underling mechanisms of dielectric-barrier discharge (DBD) plasma on STI activities. The results shown that DBD plasma treatment significantly induced the inactivation of STI in soymilk and Kunitz-type trypsin inhibitor from soybean (SKTI) in a model system. After exposure to DBD plasma at 51.4W for 21min, the STI activities of soymilk were reduced by 86.1%. Affter being treated by DBD plasma, the intrinsic fluorescence and surface hydrophobicity of SKTI were significantly decreased, while the sulfhydryl contents were increased. It is assumed that DBD plasma-induced conformational changes and oxidative modification might contribute to the inactivation of SKTI. In summary, DBD plasma technology is a potential alternative to heat treatment for the inactivation of anti-nutritional substances in food legumes.

Production of human blood group B antigen epitope conjugated protein in Escherichia coli and utilization of the adsorption blood group B antibody.

BACKGROUND: In the process of ABO-incompatible (ABOi) organ transplantation, removal of anti-A and/or B antibodies from blood plasma is a promising method to overcome hyperacute rejection and allograft loss caused by the immune response between anti-A and/or B antibodies and the A and/or B antigens in the recipient. Although there are commercial columns to do this work, the application is still limited because of the high production cost. RESULTS: In this study, the PglB glycosylation pathway from Campylobacter jejuni was exploited to produce glycoprotein conjugated with Escherichia coli O86:B7 O-antigen, which bears the blood group B antigen epitope to absorb blood group B antibody in blood. The titers of blood group B antibody were reduced to a safe level without changing the clotting function of plasma after glycoprotein absorption of B antibodies in the plasma. CONCLUSIONS: We developed a feasible strategy for the specific adsorption/removal of blood group antibodies. This method will be useful in ABOi organ transplantation and universal blood transfusion.

Enhancing GDP-fucose production in recombinant Escherichia coli by metabolic pathway engineering.

Guanosine 5'-diphosphate (GDP)-fucose is the indispensible donor substrate for fucosyltransferase-catalyzed synthesis of fucose-containing biomolecules, which have been found involving in various biological functions. In this work, the salvage pathway for GDP-fucose biosynthesis from Bacterioides fragilis was introduced into Escherichia coli. Besides, the biosynthesis of guanosine 5'-triphosphate (GTP), an essential substrate for GDP-fucose biosynthesis, was enhanced via overexpression of enzymes involved in the salvage pathway of GTP biosynthesis. The production capacities of metabolically engineered strains bearing different combinations of recombinant enzymes were compared. The shake flask fermentation of the strain expressing Fkp, Gpt, Gmk and Ndk obtained the maximum GDP-fucose content of 4.6 ± 0.22 µmol/g (dry cell mass), which is 4.2 fold that of the strain only expressing Fkp. Through fed-batch fermentation, the GDP-fucose content further rose to 6.6 ± 0.14 µmol/g (dry cell mass). In addition to a better productivity than previous fermentation processes based on the de novo pathway for GDP-fucose biosynthesis, the established schemes in this work also have the advantage to be a potential avenue to GDP-fucose analogs encompassing chemical modification on the fucose residue.

Biosynthesis of nucleotide sugars by a promiscuous UDP-sugar pyrophosphorylase from Arabidopsis thaliana (AtUSP).

Nucleotide sugars are activated forms of monosaccharides and key intermediates of carbohydrate metabolism in all organisms. The availability of structurally diverse nucleotide sugars is particularly important for the characterization of glycosyltransferases. Given that limited methods are available for preparation of nucleotide sugars, especially their useful non-natural derivatives, we introduced herein an efficient one-step three-enzyme catalytic system for the synthesis of nucleotide sugars from monosaccharides. In this study, a promiscuous UDP-sugar pyrophosphorylase (USP) from Arabidopsis thaliana (AtUSP) was used with a galactokinase from Streptococcus pneumoniae TIGR4 (SpGalK) and an inorganic pyrophosphatase (PPase) to effectively synthesize four UDP-sugars. AtUSP has better tolerance for C4-derivatives of Gal-1-P compared to UDP-glucose pyrophosphorylase from S. pneumoniae TIGR4 (SpGalU). Besides, the nucleotide substrate specificity and kinetic parameters of AtUSP were systematically studied. AtUSP exhibited considerable activity toward UTP, dUTP and dTTP, the yield of which was 87%, 85% and 84%, respectively. These results provide abundant information for better understanding of the relationship between substrate specificity and structural features of AtUSP.

NahK/GlmU fusion enzyme: characterization and one-step enzymatic synthesis of UDP-N-acetylglucosamine.

The availability of uridine 5'-diphosphate N-acetylglucosamine (UDP-GlcNAc) is a prerequisite for the GlcNAc-transferase-catalyzed glycosylation reaction. UDP-GlcNAc has already been synthesized using an N-acetylhexosamine 1-kinase (NahK) and a GlcNAc-1-P uridyltransferase (truncated GlmU) and here, a fusion enzyme was constructed with truncated GlmU and NahK. After determination of the optimum catalytic condition (pH 8.0 at 40 °C), the fusion enzyme was used to synthesize UDP-GlcNAc in a single step with a yield of 88 % from GlcNAc, ATP and UTP. Furthermore, a simplified purification method was demonstrated using separation by gel filtration after by-product digestion with alkaline phosphatase. An overall yield of 77 % and a purity of over 90 % were achieved.

Bioconversion of D-galactose to D-tagatose: continuous packed bed reaction with an immobilized thermostable L-arabinose isomerase and efficient purification by selective microbial degradation.

The continuous enzymatic conversion of D-galactose to D-tagatose with an immobilized thermostable L-arabinose isomerase in packed-bed reactor and a novel method for D-tagatose purification were studied. L-arabinose isomerase from Thermoanaerobacter mathranii (TMAI) was recombinantly overexpressed and immobilized in calcium alginate. The effects of pH and temperature on D-tagatose production reaction catalyzed by free and immobilized TMAI were investigated. The optimal condition for free enzyme was pH 8.0, 60°C, 5 mM MnCl(2). However, that for immobilized enzyme was pH 7.5, 75°C, 5 mM MnCl(2). In addition, the catalytic activity of immobilized enzyme at high temperature and low pH was significantly improved compared with free enzyme. The optimum reaction yield with immobilized TMAI increased by four percentage points to 43.9% compared with that of free TMAI. The highest productivity of 10 g/L h was achieved with the yield of 23.3%. Continuous production was performed at 70°C; after 168 h, the reaction yield was still above 30%. The resultant syrup was then incubated with Saccharomyces cerevisiae L1 cells. The selective degradation of D-galactose was achieved, obtaining D-tagatose with the purity above 95%. The established production and separation methods further potentiate the industrial production of D-tagatose via bioconversion and biopurification processes.