A Class of Environmental and Endogenous Toxins Induces BRCA2 Haploinsufficiency and Genome Instability.

Mutations truncating a single copy of the tumor suppressor, BRCA2, cause cancer susceptibility. In cells bearing such heterozygous mutations, we find that a cellular metabolite and ubiquitous environmental toxin, formaldehyde, stalls and destabilizes DNA replication forks, engendering structural chromosomal aberrations. Formaldehyde selectively depletes BRCA2 via proteasomal degradation, a mechanism of toxicity that affects very few additional cellular proteins. Heterozygous BRCA2 truncations, by lowering pre-existing BRCA2 expression, sensitize to BRCA2 haploinsufficiency induced by transient exposure to natural concentrations of formaldehyde. Acetaldehyde, an alcohol catabolite detoxified by ALDH2, precipitates similar effects. Ribonuclease H1 ameliorates replication fork instability and chromosomal aberrations provoked by aldehyde-induced BRCA2 haploinsufficiency, suggesting that BRCA2 inactivation triggers spontaneous mutagenesis during DNA replication via aberrant RNA-DNA hybrids (R-loops). These findings suggest a model wherein carcinogenesis in BRCA2 mutation carriers can be incited by compounds found pervasively in the environment and generated endogenously in certain tissues with implications for public health.

Promoted hydrogen and acetaldehyde production from alcohol dehydrogenation enabled by electrochemical hydrogen pumps.

The dehydrogenation reaction of bioderived ethanol is of particular interest for the synthesis of fuels and value-added chemicals. However, this reaction historically suffered from high energy consumption (>260 °C or >0.8 V) and low efficiency. Herein, the efficient conversion of alcohol to hydrogen and aldehyde is achieved by integrating the thermal dehydrogenation reaction with electrochemical hydrogen transfer at low temperature (120 °C) and low voltage (0.06 V), utilizing a bifunctional catalyst (Ru/C) with both thermal-catalytic and electrocatalytic activities. Specifically, the coupled electrochemical hydrogen separation procedure can serve as electrochemical hydrogen pumps, which effectively promote the equilibrium of ethanol dehydrogenation toward hydrogen and acetaldehyde production and simultaneously purifies hydrogen at the cathode. By utilizing this strategy, we achieved boosted hydrogen and acetaldehyde yields of 1,020 mmol g-1 h-1 and 1,185 mmol g-1 h-1, respectively, which are threefold higher than the exclusive ethanol thermal dehydrogenation. This work opens up a prospective route for the high-efficiency production of hydrogen and acetaldehyde via coupled thermal-electrocatalysis.

The role of AdhE on ethanol tolerance and production in Clostridium thermocellum.

Many anaerobic microorganisms use the bifunctional aldehyde and alcohol dehydrogenase enzyme, AdhE, to produce ethanol. One such organism is Clostridium thermocellum, which is of interest for cellulosic biofuel production. In the course of engineering this organism for improved ethanol tolerance and production, we observed that AdhE was a frequent target of mutations. Here, we characterized those mutations to understand their effects on enzymatic activity, as well ethanol tolerance and product formation in the organism. We found that there is a strong correlation between NADH-linked alcohol dehydrogenase (ADH) activity and ethanol tolerance. Mutations that decrease NADH-linked ADH activity increase ethanol tolerance; correspondingly, mutations that increase NADH-linked ADH activity decrease ethanol tolerance. We also found that the magnitude of ADH activity did not play a significant role in determining ethanol titer. Increasing ADH activity had no effect on ethanol titer. Reducing ADH activity had indeterminate effects on ethanol titer, sometimes increasing and sometimes decreasing it. Finally, this study shows that the cofactor specificity of ADH activity was found to be the primary factor affecting ethanol yield. We expect that these results will inform efforts to use AdhE enzymes in metabolic engineering approaches.

Autoreactive B cells against malondialdehyde-induced protein cross-links are present in the joint, lung, and bone marrow of rheumatoid arthritis patients.

Autoantibodies to malondialdehyde (MDA) proteins constitute a subset of anti-modified protein autoantibodies in rheumatoid arthritis (RA), which is distinct from citrulline reactivity. Serum anti-MDA IgG levels are commonly elevated in RA and correlate with disease activity, CRP, IL6, and TNF-α. MDA is an oxidation-associated reactive aldehyde that together with acetaldehyde mediates formation of various immunogenic amino acid adducts including linear MDA-lysine, fluorescent malondialdehyde acetaldehyde (MAA)-lysine, and intramolecular cross-linking. We used single-cell cloning, generation of recombinant antibodies (n = 356 from 25 donors), and antigen-screening to investigate the presence of class-switched MDA/MAA+ B cells in RA synovium, bone marrow, and bronchoalveolar lavage. Anti-MDA/MAA+ B cells were found in bone marrow plasma cells of late disease and in the lung of both early disease and risk-individuals and in different B cell subsets (memory, double negative B cells). These were compared with previously identified anti-MDA/MAA from synovial memory and plasma cells. Seven out of eight clones carried somatic hypermutations and all bound MDA/MAA-lysine independently of protein backbone. However, clones with somatic hypermutations targeted MAA cross-linked structures rather than MDA- or MAA-hapten, while the germline-encoded synovial clone instead bound linear MDA-lysine in proteins and peptides. Binding patterns were maintained in germline converted clones. Affinity purification of polyclonal anti-MDA/MAA from patient serum revealed higher proportion of anti-MAA versus anti-MDA compared to healthy controls. In conclusion, IgG anti-MDA/MAA show distinct targeting of different molecular structures. Anti-MAA IgG has been shown to promote bone loss and osteoclastogenesis in vivo and may contribute to RA pathogenesis.

Efficient Exciton Dissociation on Ceria Chelated Cerium-Based MOF Isogenous S-Scheme Photocatalyst for Acetaldehyde Purification.

Long-term exposure to low concentration indoor VOCs of acetaldehyde (CH3CHO) is harmful to human health. Thus, a novel isogenous heterojunction CeO2/Ce-MOF photocatalyst is synthesized via a one-step hydrothermal method for the effective elimination of CH3CHO in this work. This CeO2/Ce-MOF photocatalyst performs well in CH3CHO removal and achieves an apparent quantum efficiency of 7.15% at 420 nm, which presents ≈6.7 and 3.4 times superior to those generated by CeO2 and Ce-MOF, respectively. The enhanced efficiency is due to two main aspects including i) an effective photocarrier separation ability and the prolonged reaction lifetime of excitons play crucial roles and ii) the formation of an internal electric field (IEF) is sufficient to overcome the considerable exciton binding energy, and increases the exciton dissociation efficiency by up to 50.4%. Moreover, the reasonable pathways and mechanisms of CH3CHO degradation are determined by in situ DRIFTS analysis and simulated DFT calculations. Those results demonstrated that S-scheme heterojunction successfully increases the efficiency of harmful volatile organic compounds elimination, and it offers essential guidance for designing rare earth-based MOF photocatalysts.

Uncovering newly identified aldehyde dehydrogenase 2 genetic variants that lead to acetaldehyde accumulation after an alcohol challenge.

BACKGROUND: Aldehyde dehydrogenase 2 (ALDH2) is critical for alcohol metabolism by converting acetaldehyde to acetic acid. In East Asian descendants, an inactive genetic variant in ALDH2, rs671, triggers an alcohol flushing response due to acetaldehyde accumulation. As alcohol flushing is not exclusive to those of East Asian descent, we questioned whether additional ALDH2 genetic variants can drive facial flushing and inefficient acetaldehyde metabolism using human testing and biochemical assays. METHODS: After IRB approval, human subjects were given an alcohol challenge (0.25 g/kg) while quantifying acetaldehyde levels and the physiological response (heart rate and skin temperature) to alcohol. Further, by employing biochemical techniques including human purified ALDH2 proteins and transiently transfected NIH 3T3 cells, we characterized two newly identified ALDH2 variants for ALDH2 enzymatic activity, ALDH2 dimer/tetramer formation, and reactive oxygen species production after alcohol treatment. RESULTS: Humans heterozygous for rs747096195 (R101G) or rs190764869 (R114W) had facial flushing and a 2-fold increase in acetaldehyde levels, while rs671 (E504K) had facial flushing and a 6-fold increase in acetaldehyde levels relative to wild type ALDH2 carriers. In vitro studies with recombinant R101G and R114W ALDH2 enzyme showed a reduced efficiency in acetaldehyde metabolism that is unique when compared to E504K or wild-type ALDH2. The effect is caused by a lack of functional dimer/tetramer formation for R101G and decreased Vmax for both R101G and R114W. Transiently transfected NIH-3T3 cells with R101G and R114W also had a reduced enzymatic activity by ~ 50% relative to transfected wild-type ALDH2 and when subjected to alcohol, the R101G and R114W variants had a 2-3-fold increase in reactive oxygen species formation with respect to wild type ALDH2. CONCLUSIONS: We identified two additional ALDH2 variants in humans causing facial flushing and acetaldehyde accumulation after alcohol consumption. As alcohol use is associated with a several-fold higher risk for esophageal cancer for the E504K variant, the methodology developed here to characterize ALDH2 genetic variant response to alcohol can lead the way precision medicine strategies to further understand the interplay of alcohol consumption, ALDH2 genetics, and cancer.

Photoionization and Dissociative Photoionization of Acetaldehyde in the 10.0-13.7†eV Range by Synchrotron Photoelectron Photoion Coincidence Spectroscopy.

Photoionization and dissociative photoionization of acetaldehyde (CH3CHO) in the 10.0-13.7 eV energy range are studied by using synchrotron radiation double imaging photoelectron photoion coincidence spectroscopy (i2PEPICO). The X2A' and A2A" electronic states of CH3CHO+ as well as the Franck-Condon gap region between these two states have been populated with several vibrational sequences and assigned in the high-resolution slow photoelectron spectrum (SPES). The adiabatic ionization energies (AIEs) of the X2A' and A2A" states are measured at 10.228±0.006 and 12.52±0.05 eV, respectively. The present results show that the X2A' state is a stable state while the A2A" state is fully dissociative to produce CH3CO+, CHO+ and CH4 + fragment ions. The 0 K appearance energies (AE0K) of CH3CO+ and CHO+ fragment ions are determined through the modeling of the breakdown diagram, i. e., AE0K(CH3CO+)=10.89±0.01 eV (including a reverse barrier of ~0.19 eV) and AE0K(CHO+)=11.54±0.05 eV. In addition, the dissociation mechanisms of CH3CHO+ including statistical dissociation, direct bond breaking and isomerization are discussed with the support of the calculated dissociation limits and transition state energies.

Cyanohydrin Equilibria Implicate Non-Aromatic Aldehydes in Photochemical Production of Oceanic Carbon Monoxide.

Carbonyls have previously been dismissed as significant precursors for carbon monoxide (CO) photoproduction from natural chromophoric dissolved organic matter (CDOM). Here, we used hydrogen cyanide (HCN), which reacts with carbonyls to form photochemically inert cyanohydrins, as a probe to re-examine the role of carbonyls in CO photoproduction. Adding HCN to low-absorbance euphotic zone seawater decreased CO photoproduction. Modeling [HCN] (∼5 to 364 µM) vs the percent decrease in CO photoproduction (%CO↓) yielded carbonyl-cyanohydrin dissociation equilibrium constants, KD, and maximum %CO↓, %CO↓max values. Four Atlantic and Pacific seawater KDs (66.7 ± 19.6 µM) overlap aqueous aliphatic but not aromatic aldehyde KDs. Phenylacetaldehyde (PA) and other ß,γ-unsaturated aldehydes are proposed as prototypical CO precursors. Direct photolysis of ∼10 nM PA can supply the measured daily production of HCN-sensitive CO at an open-ocean site near Bermuda. HCN's %CO↓max was 31 ± 2.5% in North Atlantic seawater vs the 13 ± 2.5% inhibition of CO photoproduction by borohydride, a dilemma since only borohydride affects most ketones. Borohydride also decreased CDOM absorption much more than did HCN. This puzzle probably reflects differing steric and solvation requirements in HCN- and borohydride-CDOM reactions. This study demonstrates cyanophilic aldehydes to be a significant source of open-ocean CO and reveals new clues regarding CDOM photochemistry mechanisms.

Multiple Roles of Anions on the Degradation Efficiency and Mineralization Pathway of Recalcitrant Acetaldehyde by Vacuum Ultraviolet (185 nm) Oxidation.

Vacuum-UV (185 nm, VUV) is widely applied to polish reverse osmosis permeate (ROP), such as the production of electronics-grade ultrapure water. In this study, the VUV oxidation of acetaldehyde, a common carbonyl in ROP, was found to be influenced by anions even at low concentrations. Interestingly, the influencing extent and mechanism varied depending on the anions. Bicarbonate minimally affected the VUV-photon absorption and •OH consumption, but at 5000 µg-C·L-1, it decreased the degradation of acetaldehyde by 58.7% possibly by scavenging organic radicals or other radical chain reactions. Nitrate strongly competed for VUV-photon absorption and •OH scavenging through the formation of nitrite, and at 500 µg-N·L-1, it decreased the removal rate of acetaldehyde degradation by 71.2% and the mineralization rate of dissolved organic carbon by 53.4%. Chloride competed for VUV-photon absorption and also generated reactive chlorine species, which did not affect acetaldehyde degradation but influenced the formation of organic byproducts. The radical chain reactions or activation of anions under VUV irradiation could compensate for the decrease in oxidation performance and need further investigation. In real ROPs, the VUV oxidation of acetaldehyde remained efficient, but mineralization was hindered due to nitrate and chloride anions. This study deepens the understanding of the photochemistry and feasibility of VUV in water with low concentrations of anions.

Significant Biogenic Source of Oxygenated Volatile Organic Compounds and the Impacts on Photochemistry at a Regional Background Site in South China.

Oxygenated volatile organic compounds (OVOCs) significantly modulate atmospheric chemistry, but the sources and air quality impacts of OVOCs in aged urban outflows remain to be elucidated. At a background site in South China, the ozone formation potential of six nonformaldehyde OVOCs studied was equivalent to that of 3.56 ppbv of formaldehyde, more than half of which was contributed by acetaldehyde. Source apportionment incorporating photochemical age revealed that considerable fractions (52.7%-62.6%) of the OVOCs were of biogenic origin, except for ethanol, which was primarily derived from anthropogenic emissions. The oxidation of cis-/trans-2-butene explained 71.1% of the in situ acetaldehyde formation. In contrast, α/ß-pinenes and isoprene contributed 73.8% and 28.4% to acetone and methylglyoxal formation, respectively. An average of 12.4% of net in situ ozone (O3) production rate was attributed to the OVOCs studied, where the biogenic fractions accounted for 59%. The changes in the O3 production rate and hydroxyl radical (OH) concentration caused by OVOCs were mainly affected by ozone formation sensitivity. The effects of primary acetaldehyde and acetaldehyde-led O3 on secondary acetaldehyde formation were weak at this background site; however, they cannot be ignored in polluted areas. This study provides a scientific basis for mitigating O3 pollution driven by biogenic emissions and OVOCs.

Evaluation of quercetin as a potential cytoprotector against acetaldehyde using the cultured hepatocyte model with aldehyde dehydrogenase isozyme deficiency.

Protective effect of quercetin against acetaldehyde was evaluated using the cultured hepatocyte models with aldehyde dehydrogenase (ALDH) isozyme deficiency (aldh2-kd and aldh1a1-kd). The quercetin-induced cytoprotection against acetaldehyde in the ALDH1A1-deficient mutant (aldh1a1-kd) was weaker than that in the wild type. Furthermore, quercetin did not enhance the ALDH activity in aldh1a1-kd cells, suggesting that ALDH1A1 is involved in quercetin-induced cytoprotection.

Genome analysis of a newly isolated Bacillus velezensis-YW01 for biodegrading acetaldehyde.

Acetaldehyde (AL), a primary carcinogen, not only pollutes the environment, but also endangers human health after drinking alcohol. Here a promising bacterial strain was successfully isolated from a white wine cellar pool in the province of Shandong, China, and identified as Bacillus velezensis-YW01 with 16 S rDNA sequence. Using AL as sole carbon source, initial AL of 1 g/L could be completely biodegraded by YW01 within 84 h and the cell-free extracts of YW01 has also been detected to biodegrade the AL, which indicate that YW01 is a high-potential strain for the biodegradation of AL. The optimal culture conditions and the biodegradation of AL of YW01 are at pH 7.0 and 38 °C, respectively. To further analyze the biodegradation mechanism of AL, the whole genome of YW01 was sequenced. Genes ORF1040, ORF1814 and ORF0127 were revealed in KEGG, which encode for acetaldehyde dehydrogenase. Furthermore, ORF0881 and ORF052 encode for ethanol dehydrogenase. This work provides valuable information for exploring metabolic pathway of converting ethanol to AL and subsequently converting AL to carboxylic acid compounds, which opened up potential pathways for the development of microbial catalyst against AL.

Ganshuang granule plays a pharmacological role in anti-alcoholic and anti-hangover via regulating alcohol metabolism and affecting neurotransmitters.

BACKGROUND: To explore the effect of Ganshuang granule on anti-alcoholic and anti-hangover and its potential mechanism. METHODS: SPF SD rats' drunken model and SPF Kunming mice's hangover model were used as models. RESULTS: Ganshuang granule could significantly reduce sleep time, the time to climb in mice, and significantly prolong the tolerance time and shorten sleep time in rats (p < 0.05). The blood ethanol concentration of rats in each administration group was lower than that in the model group at each time point (p < 0.05). Compared with the control group, the activities of ADH and ALDH in the liver of the model group were significantly decreased (p < 0.05); the content of DA and 5-HT in the striatum of the model group was significantly increased (p < 0.05); and the activity of AchE in the hippocampus was significantly decreased (p < 0.05). The above processes could be improved and regulated in the drug administration group. Compared with the control group, there was no significant difference between ADH and ALDH in the serum of the model group (p > 0.05). However, the activities of ADH and ALDH in the liver of drunk rats could be upregulated by Ganshuang granule (p < 0.05). CONCLUSION: Ganshuang granule has the pharmacological effects of anti-alcoholic and anti-hangover, which is related to regulating the activities of ADH and ALDH in the liver, the contents of DA and 5-HT in striatum, and the activity of AchE in the hippocampus.

Quercetin Attenuates Acetaldehyde-Induced Cytotoxicity via the Heme Oxygenase-1-Dependent Antioxidant Mechanism in Hepatocytes.

It is still unclear whether or how quercetin influences the toxic events induced by acetaldehyde in hepatocytes, though quercetin has been reported to mitigate alcohol-induced mouse liver injury. In this study, we evaluated the modulating effect of quercetin on the cytotoxicity induced by acetaldehyde in mouse hepatoma Hepa1c1c7 cells, the frequently used cellular hepatocyte model. The pretreatment with quercetin significantly inhibited the cytotoxicity induced by acetaldehyde. The treatment with quercetin itself had an ability to enhance the total ALDH activity, as well as the ALDH1A1 and ALDH3A1 gene expressions. The acetaldehyde treatment significantly enhanced the intracellular reactive oxygen species (ROS) level, whereas the quercetin pretreatment dose-dependently inhibited it. Accordingly, the treatment with quercetin itself significantly up-regulated the representative intracellular antioxidant-related gene expressions, including heme oxygenase-1 (HO-1), glutamate-cysteine ligase, catalytic subunit (GCLC), and cystine/glutamate exchanger (xCT), that coincided with the enhancement of the total intracellular glutathione (GSH) level. Tin protoporphyrin IX (SNPP), a typical HO-1 inhibitor, restored the quercetin-induced reduction in the intracellular ROS level, whereas buthionine sulphoximine, a representative GSH biosynthesis inhibitor, did not. SNPP also cancelled the quercetin-induced cytoprotection against acetaldehyde. These results suggest that the low-molecular-weight antioxidants produced by the HO-1 enzymatic reaction are mainly attributable to quercetin-induced cytoprotection.

Aldehyde Dehydrogenase 2 (ALDH2) rs671 Polymorphism is a Predictor of Pulmonary Hypertension Due to Left Heart Disease.

AIM: Pulmonary hypertension due to left heart disease (PH-LHD) is commonly seen in patients with heart failure (HF), but there are limited treatment options. Recent studies have shown an association between aldehyde dehydrogenase 2 (ALDH2) rs671 polymorphisms and pulmonary hypertension (PH). Therefore, this study aimed to investigate the occurrence of ALDH2 rs671 polymorphisms, and the association between ALDH2 and risk of PH-LHD in patients with HF. It also investigated different ALDH2 genotypes and examined their association with cardiac structure and function in HF patients with PH-LHD. METHODS: A total of 178 HF patients were consecutively enrolled in this study: 102 without PH-LHD and 76 with PH-LHD. Clinical data, parameters of echocardiography, and relevant biochemical indexes were recorded in both groups. Differences in data obtained between groups were compared, and the risk of variant ALDH2 polymorphisms with PH-LHD in HF patients was analysed using univariate and multivariate logistic regression. RESULTS: The prevalence of ALDH2 rs671 GA/AA polymorphisms (variant ALDH2) was 24 of 102 patients (23.53%) in the HF without PH-LHD group, and 32 of 76 patients (42.10%) in the HF with PH-LHD group, with a statistically significant difference. Univariate and multivariate logistical regression showed that variant ALDH2 is an independent risk factor for HF combined with PH-LHD. A higher proportion of patients with variant ALDH2 in the HF with PH-LHD group had a tricuspid regurgitation velocity >2.8 m/s, and they had higher values of peak early diastolic velocity of the mitral orifice/peak velocity of the early diastolic wave of the mitral orifice, maximum frequency shift of pulmonary valve flow, and pulmonary artery stiffness. CONCLUSIONS: Variant ALDH2 may be an independent risk factor for HF combined with PH-LHD. Variant ALDH2 may also be involved in pulmonary artery remodelling and is a potential new target for clinical treatment of PH-LHD.

Copper (II) Level in Musts Affects Acetaldehyde Concentration, Phenolic Composition, and Chromatic Characteristics of Red and White Wines.

Copper (II), a vital fungicide in organic viticulture, also acts as a wine oxidation catalyst. However, limited data are currently available on the impact that maximum allowed copper (II) ion doses in wine grapes at harvest can have on aged wine quality. This was the focus of the present study. We investigated the copper (II) effects by producing both white and red wines from musts containing three initial metal concentrations according to the limits set for organic farming. In detail, the influence of copper (II) on fermentation evolution, chromatic characteristics, and phenolic compounds was evaluated. Interestingly, the white wine obtained with the highest permitted copper (II) dose initially exceeded the concentration of 1.0 mg/L at fermentation completion. However, after one year of storage, the copper (II) content fell below 0.2 ± 0.01 mg/L. Conversely, red wines showed copper (II) levels below 1.0 mg/L at the end of fermentation, but the initial copper (II) level in musts significantly affected total native anthocyanins, color intensity, hue, and acetaldehyde concentration. After 12-month aging, significant differences were observed in polymeric pigments, thus suggesting a potential long-term effect of copper (II) on red wine color stability.

[Patient and quality characteristics in the treatment with disulfiram (Antabus) in the German "Network for Alcohol Aversive Pharmacotherapy"]. / Patienten- und Qualitätsmerkmale bei der Behandlung mit Disulfiram ("Antabus") im deutschsprachigen "Netzwerk alkoholaversive Pharmakotherapie".

BACKGROUND: More than a decade ago disulfiram lost its approval for use in Germany. Nonetheless, a considerable number of psychiatric hospital outpatient departments as well as practicing physicians continue to prescribe it. These professionals have formed the "Network for Alcohol Aversive Pharmacotherapy" (NAP) to maintain a high quality of this treatment approach. OBJECTIVE: To describe the current use of disulfiram with respect to patient numbers and characteristics, side effects, and use of concomitant multimodal treatment forms. MATERIAL AND METHODS: Since 2019 the NAP has conducted an annual retrospective survey among its members regarding the aforementioned parameters. RESULTS: From 2019 to 2023 a total of 1579 treatment cases were described by 33 centers, 152 patients reported a total of 241 drinking events, 26 of them resulting in hospitalization but none causing complications or permanent harm. The most frequent side effects, in descending order, were unpleasant body odor (2.5%), fatigue, male sexual dysfunction, mildly elevated liver enzymes, allergic skin reactions and polyneuropathy (0.8%). More than one quarter of the patients suffered from comorbid depression, and approximately 5% from ADHD, borderline or other personality disorders, trauma-related disorders and anxiety disorders, respectively. Of the patients 33% were treated with antidepressants and 12% with sedating antipsychotics. Various forms of concomitant group therapy were offered to 66% of the patients. CONCLUSION: Treatment with disulfiram is legally possible, generally well-tolerated and safe. It is offered in most treatment centers as part of a comprehensive treatment plan that includes multimodal treatment of comorbid psychiatric disorders.

[Study based on the acetaldehyde dehydrogenase 2 gene polymorphism and acetaminophen-induced liver injury].

Objective: To explore the association between aldehyde dehydrogenase 2 (ALDH2) gene polymorphisms and abnormal liver function-induced by acetaminophen (APAP) drugs. Methods: An ALDH2 gene knockout mouse model was constructed using CRISPR/Cas9 gene editing technology. The obtained heterozygous mice were mated with opposite sex of heterozygotes. Genomic DNA was extracted from the tail of the offspring mouse. The polymerase chain reaction (PCR) method was used to determine the ALDH2 genotype. APAP was further used to induce acute drug-induced liver injury models in wild-type and ALDH2 knockout mice. Blood and liver tissues of mice were collected for liver function index, HE staining, F4/80 immunohistochemistry, and other detections. The intergroup mean was compared using a one-way ANOVA. The LSD- t test was used for pairwise comparison. Results: ALDH2 knockout mice were bred successfully. The genotyping of the offspring was segregated into the wild-type (ALDH2(+/+)), heterozygous mutant (ALDH2(+/-)), and homozygous mutant (ALDH2(-/-)), respectively. Biochemical and histological results after APAP modeling showed that the level of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and total bilirubin (TBil) was not significantly increased in the blank control group (P < 0.05), while the ALT, AST,ALP, and TBil were all elevated in the APAP experimental group. The levels of ALT (P  = 0.004), AST (P = 0.002), and TBil (P = 0.012) were significantly elevated among the mutant group compared to those in the wild-type group, and the expression levels of these indicators were also significantly elevated among the homozygous mutant group compared to those in the heterozygous mutant group (P = 0.003, 0 and 0.006). In addition, the ALP levels were higher in the heterozygous mutation group than those in the homozygous mutant group (P = 0.085) and wild-type group mice, but the difference was only statistically significant compared to wild-type mice (P = 0.002). HE staining results showed that mice in the APAP experimental group had hepatocyte degeneration, necrosis, and increased inflammatory cell infiltration, which was mostly evident in mutant mice. Simultaneously, the F4/80 immunohistochemical staining results showed that brown granules were visible in the liver tissue of APAP experimental group mice, and its expression levels were significantly enhanced compared to the blank control group. Conclusion: APAP-induced liver function abnormalities were associated with the ALDH2 gene polymorphism. The liver injury symptoms were increased in ALDH2 mutant mice following APAP modeling, and the ALDH2 gene defect may alleviate, to some extent, APAP-induced liver function abnormalities.

Robotic workflows for automated long-term adaptive laboratory evolution: improving ethanol utilization by Corynebacterium glutamicum.

BACKGROUND: Adaptive laboratory evolution (ALE) is known as a powerful tool for untargeted engineering of microbial strains and genomics research. It is particularly well suited for the adaptation of microorganisms to new environmental conditions, such as alternative substrate sources. Since the probability of generating beneficial mutations increases with the frequency of DNA replication, ALE experiments are ideally free of constraints on the required duration of cell proliferation. RESULTS: Here, we present an extended robotic workflow for performing long-term evolution experiments based on fully automated repetitive batch cultures (rbALE) in a well-controlled microbioreactor environment. Using a microtiter plate recycling approach, the number of batches and thus cell generations is technically unlimited. By applying the validated workflow in three parallel rbALE runs, ethanol utilization by Corynebacterium glutamicum ATCC 13032 (WT) was significantly improved. The evolved mutant strain WT_EtOH-Evo showed a specific ethanol uptake rate of 8.45 ± 0.12 mmolEtOH gCDW-1 h-1 and a growth rate of 0.15 ± 0.01 h-1 in lab-scale bioreactors. Genome sequencing of this strain revealed a striking single nucleotide variation (SNV) upstream of the ald gene (NCgl2698, cg3096) encoding acetaldehyde dehydrogenase (ALDH). The mutated basepair was previously predicted to be part of the binding site for the global transcriptional regulator GlxR, and re-engineering demonstrated that the identified SNV is key for enhanced ethanol assimilation. Decreased binding of GlxR leads to increased synthesis of the rate-limiting enzyme ALDH, which was confirmed by proteomics measurements. CONCLUSIONS: The established rbALE technology is generally applicable to any microbial strain and selection pressure that fits the small-scale cultivation format. In addition, our specific results will enable improved production processes with C. glutamicum from ethanol, which is of particular interest for acetyl-CoA-derived products.

Impairment of Endothelial Function by Cigarette Smoke Is Not Caused by a Specific Smoke Constituent, but by Vagal Input From the Airway.

BACKGROUND: Exposure to tobacco or marijuana smoke, or e-cigarette aerosols, causes vascular endothelial dysfunction in humans and rats. We aimed to determine what constituent, or class of constituents, of smoke is responsible for endothelial functional impairment. METHODS: We investigated several smoke constituents that we hypothesized to mediate this effect by exposing rats and measuring arterial flow-mediated dilation (FMD) pre- and post-exposure. We measured FMD before and after inhalation of sidestream smoke from research cigarettes containing normal and reduced nicotine level with and without menthol, as well as 2 of the main aldehyde gases found in both smoke and e-cigarette aerosol (acrolein and acetaldehyde), and inert carbon nanoparticles. RESULTS: FMD was reduced by all 4 kinds of research cigarettes, with extent of reduction ranging from 20% to 46% depending on the cigarette type. While nicotine was not required for the impairment, higher nicotine levels in smoke were associated with a greater percent reduction of FMD (41.1±4.5% reduction versus 19.2±9.5%; P=0.047). Lower menthol levels were also associated with a greater percent reduction of FMD (18.5±9.8% versus 40.5±4.8%; P=0.048). Inhalation of acrolein or acetaldehyde gases at smoke-relevant concentrations impaired FMD by roughly 50% (P=0.001). However, inhalation of inert carbon nanoparticles at smoke-relevant concentrations with no gas phase also impaired FMD by a comparable amount (P<0.001). Bilateral cervical vagotomy blocked the impairment of FMD by tobacco smoke. CONCLUSIONS: There is no single constituent or class of constituents responsible for acute impairment of endothelial function by smoke; rather, we propose that acute endothelial dysfunction by disparate inhaled products is caused by vagus nerve signaling initiated by airway irritation.