Identification of a cinnamoyl-CoA reductase from Cinnamomum cassia involved in trans-cinnamaldehyde biosynthesis.

MAIN CONCLUSION: The identification of a functional cinnamoyl-CoA reductase enzyme from Cinnamomum cassia involved in trans-cinnamaldehyde biosynthesis offers the potential for enhancing trans-cinnamaldehyde production through genetic engineering. A significant accumulation of trans-cinnamaldehyde has been found in the bark tissues of C. cassia, used in traditional Chinese medicine. trans-Cinnamaldehyde exhibits various pharmacological properties such as anti-inflammatory, analgesic, and protection of the stomach and the digestive tract. However, further elucidation and characterization of the biosynthetic pathway for trans-cinnamaldehyde is required. In this study, we conducted an integrated analysis of trans-cinnamaldehyde accumulation profiles and transcriptomic data from five different C. cassia tissues to identify the genes involved in its biosynthesis. The transcriptome data we obtained included nearly all genes associated with the trans-cinnamaldehyde pathway, with the majority demonstrating high abundance in branch barks and trunk barks. We successfully cloned four C. cassia cinnamoyl-CoA reductases (CcCCRs), a key gene in trans-cinnamaldehyde biosynthesis. We found that the recombinant CcCCR1 protein was the only one that more efficiently converted cinnamoyl-CoA into trans-cinnamaldehyde. CcCCR1 exhibited approximately 14.7-fold higher catalytic efficiency (kcat/Km) compared to the Arabidopsis thaliana cinnamoyl-CoA reductase 1 (AtCCR1); therefore, it can be utilized for engineering higher trans-cinnamaldehyde production as previously reported. Molecular docking studies and mutagenesis experiments also validated the superior catalytic activity of CcCCR1 compared to AtCCR1. These findings provide valuable insights for the functional characterization of enzyme-coding genes and hold potential for future engineering of trans-cinnamaldehyde biosynthetic pathways.

Pharmacologic Targeting of Histone H3K27 Acetylation/BRD4-dependent Induction of ALDH1A3 for Early-phase Drug Tolerance of Gastric Cancer.

Anticancer drug-tolerant persister (DTP) cells at an early phase of chemotherapy reshape refractory tumors. Aldehyde dehydrogenase 1 family member A3 (ALDH1A3) is commonly upregulated by various anticancer drugs in gastric cancer patient-derived cells (PDC) and promotes tumor growth. However, the mechanism underlying the generation of ALDH1A3-positive DTP cells remains elusive. Here, we investigated the mechanism of ALDH1A3 expression and a combination therapy targeting gastric cancer DTP cells. We found that gastric cancer tissues treated with neoadjuvant chemotherapy showed high ALDH1A3 expression. Chromatin immunoprecipitation (ChIP)-PCR and ChIP sequencing analyses revealed that histone H3 lysine 27 acetylation was enriched in the ALDH1A3 promoter in 5-fluorouracil (5-FU)-tolerant persister PDCs. By chemical library screening, we found that the bromodomain and extraterminal (BET) inhibitors OTX015/birabresib and I-BET-762/molibresib suppressed DTP-related ALDH1A3 expression and preferentially inhibited DTP cell growth. In DTP cells, BRD4, but not BRD2/3, was recruited to the ALDH1A3 promoter and BRD4 knockdown decreased drug-induced ALDH1A3 upregulation. Combination therapy with 5-FU and OTX015 significantly suppressed in vivo tumor growth. These observations suggest that BET inhibitors are efficient DTP cell-targeting agents for gastric cancer treatment. SIGNIFICANCE: Drug resistance hampers the cure of patients with cancer. To prevent stable drug resistance, DTP cancer cells are rational therapeutic targets that emerge during the early phase of chemotherapy. This study proposes that the epigenetic regulation by BET inhibitors may be a rational therapeutic strategy to eliminate DTP cells.

Investigating human-derived lactic acid bacteria for alcohol resistance.

BACKGROUND: Excessive alcohol consumption has been consistently linked to serious adverse health effects, particularly affecting the liver. One natural defense against the detrimental impacts of alcohol is provided by alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH), which detoxify harmful alcohol metabolites. Recent studies have shown that certain probiotic strains, notably Lactobacillus spp., possess alcohol resistance and can produce these critical enzymes. Incorporating these probiotics into alcoholic beverages represents a pioneering approach that can potentially mitigate the negative health effects of alcohol while meeting evolving consumer preferences for functional and health-centric products. RESULTS: Five lactic acid bacteria (LAB) isolates were identified: Lactobacillus paracasei Alc1, Lacticaseibacillus rhamnosus AA, Pediococcus acidilactici Alc3, Lactobacillus paracasei Alc4, and Pediococcus acidilactici Alc5. Assessment of their alcohol tolerance, safety, adhesion ability, and immunomodulatory effects identified L. rhamnosus AA as the most promising alcohol-tolerant probiotic strain. This strain also showed high production of ADH and ALDH. Whole genome sequencing analysis revealed that the L. rhamnosus AA genome contained both the adh (encoding for ADH) and the adhE (encoding for ALDH) genes. CONCLUSIONS: L. rhamnosus AA, a novel probiotic candidate, showed notable alcohol resistance and the capability to produce enzymes essential for alcohol metabolism. This strain is a highly promising candidate for integration into commercial alcoholic beverages upon completion of comprehensive safety and functionality evaluations.

A glutathione-independent DJ-1/Pfp1 domain containing glyoxalase III, OsDJ-1C, functions in abiotic stress adaptation in rice.

MAIN CONCLUSION: Overexpression of OsDJ-1C in rice improves root architecture, photosynthesis, yield and abiotic stress tolerance through modulating methylglyoxal levels, antioxidant defense, and redox homeostasis. Exposure to abiotic stresses leads to elevated methylglyoxal (MG) levels in plants, impacting seed germination and root growth. In response, the activation of NADPH-dependent aldo-keto reductase and glutathione (GSH)-dependent glyoxalase enzymes helps to regulate MG levels and reduce its toxic effects. However, detoxification may not be carried out effectively due to the limitation of GSH and NADPH in plants under stress. Recently, a novel enzyme called glyoxalase III (GLY III) has been discovered which can detoxify MG in a single step without needing GSH. To understand the physiological importance of this pathway in rice, we overexpressed the gene encoding GLYIII enzyme (OsDJ-1C) in rice. It was observed that OsDJ-1C overexpression in rice regulated MG levels under stress conditions thus, linked well with plants' abiotic stress tolerance potential. The OsDJ-1C overexpression lines displayed better root architecture, improved photosynthesis, and reduced yield penalty compared to the WT plants under salinity, and drought stress conditions. These plants demonstrated an improved GSH/GSSG ratio, reduced level of reactive oxygen species, increased antioxidant capacity, and higher anti-glycation activity thereby indicating that the GLYIII mediated MG detoxification plays a significant role in plants' ability to reduce the impact of abiotic stress. Furthermore, these findings imply the potential of OsDJ-1C in crop improvement programs.

Identifying a key spot for electron mediator-interaction to tailor CO dehydrogenase's affinity.

Fe‒S cluster-harboring enzymes, such as carbon monoxide dehydrogenases (CODH), employ sophisticated artificial electron mediators like viologens to serve as potent biocatalysts capable of cleaning-up industrial off-gases at stunning reaction rates. Unraveling the interplay between these enzymes and their associated mediators is essential for improving the efficiency of CODHs. Here we show the electron mediator-interaction site on ChCODHs (Ch, Carboxydothermus hydrogenoformans) using a systematic approach that leverages the viologen-reactive characteristics of superficial aromatic residues. By enhancing mediator-interaction (R57G/N59L) near the D-cluster, the strategically tailored variants exhibit a ten-fold increase in ethyl viologen affinity relative to the wild-type without sacrificing the turn-over rate (kcat). Viologen-complexed structures reveal the pivotal positions of surface phenylalanine residues, serving as external conduits for the D-cluster to/from viologen. One variant (R57G/N59L/A559W) can treat a broad spectrum of waste gases (from steel-process and plastic-gasification) containing O2. Decoding mediator interactions will facilitate the development of industrially high-efficient biocatalysts encompassing gas-utilizing enzymes.

Variation of gene expression of fatty acid acyl CoA reductase associated with wax secretion of a scale insect, Ericerus pela, and identification of its regulation factors through the accessible chromatin analyses and yeast one-hybrid.

The Chinese white wax scale insect (CWWSI), Ericerus pela, can secret an amount of wax equivalent to their body weight. Previous studies demonstrated the fatty acyl-CoA reductase (far3) plays a pivotal role in wax secretion of CWWSI. The high expression of far3 is crucial for the massive wax secretion. However, the transcription regulation of far3 was not clear. To identify regulatory factors that control the expression of far3, the assay for transposase-accessible chromatin (ATAC) and yeast one-hybrid (Y1H) were carried out in this study. The ATAC sequencing of the CWWSI at the early wax-secretion stage ATAC-seq resulted in 22.75 GB raw data, generated 75,827,225 clean reads and revealed 142,771 peaks. There was one significant peak in the 3 kb upstream regulation regions. The peak sequence is located between -1000 and -670 bp upstream of the far3 transcription start site, spanning a length of 331 bp. This peak sequence served as bait for creating the pAbAi-peak recombinant vector, used in Y1H screenings to identify proteins interacting with far3 gene. The results indicate a successful CWWSI cDNA library construction with a capacity of 1.2 × 107 colony forming unit, a 95.8% recombination rate, and insert sizes between 1,000 and 2,000 bp. Self-activation tests established that 100 ng/mL of AbA effectively inhibited bait vector self-activation. Finally, a total of 88 positive clones were selected. After sequencing and removal of duplication, 63 unique clones were obtained from these screened colonies. By aligning the clone sequences with full-length transcriptome and genome of CWWSI, the full-length coding sequences of these clones were obtained. BlastX analysis identified a transcription factor, nuclear transcription factor Y beta, and two co-activators, cAMP-response-element-binding-protein-binding protein and WW domain binding protein 2. Reverse transcription quantitative polymerase chain reaction analysis confirmed that their expression patterns were consistent with the developmental stages preceding wax secretion and matched the wax secretion characteristics during ovulation periods. These results are beneficial for further research into the regulatory mechanisms of wax secretion of CWWSI.

[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.

Gehua Jiejiu Dizhi decoction ameliorates alcoholic fatty liver in mice by regulating lipid and bile acid metabolism and with exertion of antioxidant stress based on 4DLabel-free quantitative proteomic study.

OBJECTIVE: To analyze the effect and molecular mechanism of Gehua Jiejiu Dizhi decoction (, GJDD) on alcoholic fatty live disease (AFLD) by using proteomic methods. METHODS: The male C57BL/6J mouse were randomly divided into four groups: control group, model group, GJDD group and resveratrol group. After the AFLD model was successfully prepared by intragastric administration of alcohol once on the basis of the Lieber-DeCarli classical method, the GJDD group and resveratrol group were intragastrically administered with GJDD (4900 mg/kg) and resveratrol (400 mg/kg) respectively, once a day for 9 d. The fat deposition of liver tissue was observed and evaluated by oil red O (ORO) staining. 4DLabel-free quantitative proteome method was used to determine and quantify the protein expression in liver tissue of each experimental group. The differentially expressed proteins were screened according to protein expression differential multiples, and then analyzed by Gene ontology classification and Kyoto Encyclopedia of Genes and Genomes pathway enrichment. Finally, expression validation of the differentially co-expressed proteins from control group, model group and GJDD group were verified by targeted proteomics quantification techniques. RESULTS: In semiquantitative analyses of ORO, all kinds of steatosis (ToS, MaS, and MiS) were evaluated higher in AFLD mice compared to those in GJDD or resveratrol-treated mice. 4DLabel-free proteomics analysis results showed that a total of 4513 proteins were identified, of which 3763 proteins were quantified and 946 differentially expressed proteins were screened. Compared with the control group, 145 proteins were up-regulated and 148 proteins were down-regulated in the liver tissue of model group. In addition, compared with the model group, 92 proteins were up-regulated and 135 proteins were down-regulated in the liver tissue of the GJDD group. 15 differentially co-expressed proteins were found between every two groups (model group vs control group, GJDD group vs model group and GJDD group vs control group), which were involved in many biological processes. Among them, 11 differentially co-expressed key proteins (Aox3, H1-5, Fabp5, Ces3a, Nudt7, Serpinb1a, Fkbp11, Rpl22l1, Keg1, Acss2 and Slco1a1) were further identified by targeted proteomic quantitative technology and their expression patterns were consistent with the results of 4D label-free proteomic analysis. CONCLUSIONS: Our study provided proteomics-based evidence that GJDD alleviated AFLD by modulating liver protein expression, likely through the modulation of lipid metabolism, bile acid metabolism and with exertion of antioxidant stress.

Research Progress on the Correlation between Acetaldehyde Dehydrogenase 2 and Hepatocellular Carcinoma Development.

Hepatocellular carcinoma (HCC) is the predominant pathologic type of primary liver cancer. It is a malignant tumor of liver epithelial cells. There are many ways to treat HCC, but the survival rate for HCC patients remains low. Therefore, understanding the underlying mechanisms by which HCC occurs and develops is critical to explore new therapeutic targets. Aldehyde dehydrogenase 2 (ALDH2) is an important player in the redox reaction of ethanol with endogenous aldehyde products released by lipid peroxidation. Increasing evidence suggests that ALDH2 is a crucial regulator of human tumor development, including HCC. Therefore, clarifying the relationship between ALDH2 and HCC is helpful for formulating rational treatment strategies. This review highlights the regulatory roles of ALDH2 in the development of HCC, elucidates the multiple potential mechanisms by which ALDH2 regulates the development of HCC, and summarizes the progress of research on ALDH2 gene polymorphisms and HCC susceptibility. Meanwhile, we envision viable strategies for targeting ALDH2 in the treatment of HCC SIGNIFICANCE STATEMENT: Numerous studies have aimed to explore novel therapeutic targets for HCC, and ALDH2 has been reported to be a critical regulator of HCC progression. This review discusses the functions, molecular mechanisms, and clinical significance of ALDH2 in the development of HCC and examines the prospects of ALDH2-based therapy for HCC.

Divergent evolution of the alcohol-forming pathway of wax biosynthesis among bryophytes.

The plant cuticle is a hydrophobic barrier, which seals the epidermal surface of most aboveground organs. While the cuticle biosynthesis of angiosperms has been intensively studied, knowledge about its existence and composition in nonvascular plants is scarce. Here, we identified and characterized homologs of Arabidopsis thaliana fatty acyl-CoA reductase (FAR) ECERIFERUM 4 (AtCER4) and bifunctional wax ester synthase/acyl-CoA:diacylglycerol acyltransferase 1 (AtWSD1) in the liverwort Marchantia polymorpha (MpFAR2 and MpWSD1) and the moss Physcomitrium patens (PpFAR2A, PpFAR2B, and PpWSD1). Although bryophyte harbor similar compound classes as described for angiosperm cuticles, their biosynthesis may not be fully conserved between the bryophytes M. polymorpha and P. patens or between these bryophytes and angiosperms. While PpFAR2A and PpFAR2B contribute to the production of primary alcohols in P. patens, loss of MpFAR2 function does not affect the wax profile of M. polymorpha. By contrast, MpWSD1 acts as the major wax ester-producing enzyme in M. polymorpha, whereas mutations of PpWSD1 do not affect the wax ester levels of P. patens. Our results suggest that the biosynthetic enzymes involved in primary alcohol and wax ester formation in land plants have either evolved multiple times independently or undergone pronounced radiation followed by the formation of lineage-specific toolkits.

Development of substituted benzimidazoles as inhibitors of human aldehyde dehydrogenase 1A isoenzymes.

Aldehyde dehydrogenase 1A (ALDH1A) isoforms may be a useful target for overcoming chemotherapy resistance in high-grade serous ovarian cancer (HGSOC) and other solid tumor cancers. However, as different cancers express different ALDH1A isoforms, isoform selective inhibitors may have a limited therapeutic scope. Furthermore, resistance to an ALDH1A isoform selective inhibitor could arise via induction of expression of other ALDH1A isoforms. As such, we have focused on the development of pan-ALDH1A inhibitors, rather than on ALDH1A isoform selective compounds. Herein, we report the development of a new group of pan-ALDH1A inhibitors to assess whether broad spectrum ALDH1A inhibition is an effective adjunct to chemotherapy in HGSOC. Optimization of the CM10 scaffold, aided by ALDH1A1 crystal structures, led to improved biochemical potencies, improved cellular efficacy as demonstrated by reduction in ALDEFLUOR signal in HGSOC cells, and substantial improvements in liver microsomal stability. Based on this work we identified two compounds 17 and 25 suitable for future in vivo proof of concept experiments.

Modulating the pH profile of vanillin dehydrogenase enzyme from extremophile Bacillus ligniniphilus L1 through computational guided site-directed mutagenesis.

Vanillin dehydrogenase (VDH) has recently come forward as an important enzyme for the commercial production of vanillic acid from vanillin in a one-step enzymatic process. However, VDH with high alkaline tolerance and efficiency is desirable to meet the biorefinery requirements. In this study, computationally guided site-directed mutagenesis was performed by increasing the positive and negative charges on the surface and near the active site of the VDH from the alkaliphilic marine bacterium Bacillus ligniniphilus L1, respectively. In total, 20 residues including 15 from surface amino acids and 5 near active sites were selected based on computational analysis and were subjected to site-directed mutations. The optimum pH of the two screened mutants including I132R, and T235E from surface residue and near active site mutant was shifted to 9, and 8.6, with a 2.82- and 2.95-fold increase in their activity compared to wild enzyme at pH 9, respectively. A double mutant containing both these mutations i.e., I132R/T235E was produced which showed a shift in optimum pH of VDH from 7.4 to 9, with an increase of 74.91 % in enzyme activity. Therefore, the double mutant of VDH from the L1 strain (I132R/T235E) produced in this study represents a potential candidate for industrial applications.

Differential S-nitrosylation and characterization of purified S-nitrosoglutathione reductase (GSNOR) from Brassica juncea shows multiple forms of the enzyme.

S-nitrosoglutathione reductase (GSNOR). a master regulator of NO homeostasis, is a single-copy gene in most plants. In Lotus japonicus, two GSNOR isoforms were identified exhibiting similar kinetic properties but differential tissue-specific expressions. Previously, a genome-wide identification in Brassica juncea revealed four copies of GSNOR, each encoding proteins that vary in subunit molecular weights and pI. Here, we report multiple forms of GSNOR using 2D immunoblot which showed 4 immunopositive spots of 41.5 kDa (pl 5.79 and 6.78) and 43 kDa (pl 6.16 and 6.23). To confirm, purification of GSNOR using anion-exchange chromatography yielded 2 distinct pools (GSNOR-A & GSNOR-B) with GSNOR activities. Subsequently, affinity-based purification resulted in 1 polypeptide from GSNOR-A and 2 polypeptides from GSNOR-B. Size exclusion-HPLC confirmed 3 GSNORs with molecular weight of 87.48 ± 2.74 KDa (GSNOR-A); 87.36 ± 3.25 and 82.74 ± 2.75 kDa (GSNOR-B). Kinetic analysis showed Km of 118 ± 11 µM and Vmax of 287 ± 22 nkat/mg for GSNOR-A, whereas Km of 96.4 ± 8 µM and Vmax of 349 ± 15 nkat/mg for GSNOR-B. S-nitrosylation and inhibition by NO showed redox regulation of all BjGSNORs. Both purified GSNORs exhibited variable denitrosylation efficiency as depicted by Biotin Switch assay. To the best of our knowledge, this is the first report confirming multiple isoforms of GSNOR in B. juncea.

S-nitrosothiol homeostasis maintained by ADH5 facilitates STING-dependent host defense against pathogens.

Oxidative (or respiratory) burst confers host defense against pathogens by generating reactive species, including reactive nitrogen species (RNS). The microbial infection-induced excessive RNS damages many biological molecules via S-nitrosothiol (SNO) accumulation. However, the mechanism by which the host enables innate immunity activation during oxidative burst remains largely unknown. Here, we demonstrate that S-nitrosoglutathione (GSNO), the main endogenous SNO, attenuates innate immune responses against herpes simplex virus-1 (HSV-1) and Listeria monocytogenes infections. Mechanistically, GSNO induces the S-nitrosylation of stimulator of interferon genes (STING) at Cys257, inhibiting its binding to the second messenger cyclic guanosine monophosphate-adenosine monophosphate (cGAMP). Alcohol dehydrogenase 5 (ADH5), the key enzyme that metabolizes GSNO to decrease cellular SNOs, facilitates STING activation by inhibiting S-nitrosylation. Concordantly, Adh5 deficiency show defective STING-dependent immune responses upon microbial challenge and facilitates viral replication. Thus, cellular oxidative burst-induced RNS attenuates the STING-mediated innate immune responses to microbial infection, while ADH5 licenses STING activation by maintaining cellular SNO homeostasis.

S-nitrosoglutathione reductase-dependent p65 denitrosation promotes osteoclastogenesis by facilitating recruitment of p65 to NFATc1 promoter.

Osteoclasts, the exclusive bone resorptive cells, are indispensable for bone remodeling. Hence, understanding novel signaling modulators regulating osteoclastogenesis is clinically important. Nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1) is a master transcription factor in osteoclastogenesis, and binding of NF-κB p65 subunit to NFATc1 promoter is required for its expression. It is well-established that DNA binding activity of p65 can be regulated by various post-translational modifications, including S-nitrosation. Recent studies have demonstrated that S-nitrosoglutathione reductase (GSNOR)-mediated protein denitrosation participated in cell fate commitment by regulating gene transcription. However, the role of GSNOR in osteoclastogenesis remains unexplored and enigmatic. Here, we investigated the effect of GSNOR-mediated denitrosation of p65 on osteoclastogenesis. Our results revealed that GSNOR was up-regulated during osteoclastogenesis in vitro. Moreover, GSNOR inhibition with a chemical inhibitor impaired osteoclast differentiation, podosome belt formation, and bone resorption activity. Furthermore, GSNOR inhibition enhanced the S-nitrosation level of p65, precluded the binding of p65 to NFATc1 promoter, and suppressed NFATc1 expression. In addition, mouse model of lipopolysaccharides (LPS)-induced calvarial osteolysis was employed to evaluate the therapeutic effect of GSNOR inhibitor in vivo. Our results indicated that GSNOR inhibitor treatment alleviated the inflammatory bone loss by impairing osteoclast formation in mice. Taken together, these data have shown that GSNOR activity is required for osteoclastogenesis by facilitating binding of p65 to NFATc1 promoter via promoting p65 denitrosation, suggesting that GSNOR may be a potential therapeutic target in the treatment of osteolytic diseases.

Potential clinical significance of ALDH3B1 in auxiliary diagnosis of gastric cancer.

Objective: This research aimed to explore a diagnostic method based on serum ALDH3B1 and to evaluate the clinical diagnostic efficacy in gastric cancer (GC) by comparing it with the traditional GC diagnostic method, the carcinoembryonic protein (CEA) assay. Methods: Serum samples were collected from 70 healthy volunteers and various patients (GC: 76, benign gastric lesions: 20, postoperative: 37, recurrence: 56). The diagnostic efficacy of serum ALDH3B1, CEA and the co-diagnosis were evaluated by receiver operating characteristic curve. ALDH3B1 protein levels were evaluated by western blot. Results: The co-diagnosis of ALDH3B1 and CEA had the highest diagnostic efficacy (area under the curve = 0.841). Conclusion: Serum ALDH3B1 may be used as an auxiliary diagnostic biomarker for GC, and its co-diagnosis with CEA can improve diagnostic efficacy.

Oral Administration of Alcohol-Tolerant Lactic Acid Bacteria Alleviates Blood Alcohol Concentration and Ethanol-Induced Liver Damage in Rodents.

Excessive alcohol consumption can have serious negative consequences on health, including addiction, liver damage, and other long-term effects. The causes of hangovers include dehydration, alcohol and alcohol metabolite toxicity, and nutrient deficiency due to absorption disorders. Additionally, alcohol consumption can slow reaction times, making it more difficult to rapidly respond to situations that require quick thinking. Exposure to a large amount of ethanol can also negatively affect a person's righting reflex and balance. In this study, we evaluated the potential of lactic acid bacteria (LAB) to alleviate alcohol-induced effects and behavioral responses. Two LAB strains isolated from kimchi, Levilactobacillus brevis WiKim0168 and Leuconostoc mesenteroides WiKim0172, were selected for their ethanol tolerance and potential to alleviate hangover symptoms. Enzyme activity assays for alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH) were then conducted to evaluate the role of these bacteria in alcohol metabolism. Through in vitro and in vivo studies, these strains were assessed for their ability to reduce blood alcohol concentrations and protect against alcohol-induced liver damage. The results indicated that these LAB strains possess significant ethanol tolerance and elevate ADH and ALDH activities. LAB administration remarkably reduced blood alcohol levels in rats after excessive alcohol consumption. Moreover, the LAB strains showed hepatoprotective effects and enhanced behavioral outcomes, highlighting their potential as probiotics for counteracting the adverse effects of alcohol consumption. These findings support the development of functional foods incorporating LAB strains that can mediate behavioral improvements following alcohol intake.

A novel tetratricopeptide-repeat protein, TTP1, forms complexes with glutamyl-tRNA reductase and protochlorophyllide oxidoreductase during tetrapyrrole biosynthesis.

The biosynthesis of the tetrapyrrole end-products chlorophyll and heme depends on a multifaceted control mechanism that acts primarily at the post-translational level upon the rate-limiting step of 5-aminolevulinic acid synthesis and upon light-dependent protochlorophyllide oxidoreductase (POR). These regulatory processes require auxiliary factors that modulate the activity, stability, complex formation, and subplastidal localization of the relevant proteins. Together, they ensure optimal metabolic flow during the day and at night. As an Arabidopsis homolog of the POR-interacting tetratricopeptide-repeat protein (Pitt) first reported in Synechocystis, we characterize tetrapyrrole biosynthesis-regulating tetratricopeptide-repeat protein1 (TTP1). TTP1 is a plastid-localized, membrane-bound factor that interacts with POR, the Mg protoporphyrin monomethylester cyclase CHL27, glutamyl-tRNA reductase (GluTR), GluTR-binding protein, and FLUORESCENCE IN BLUE LIGHT. Lack of TTP1 leads to accumulation of GluTR, enhanced 5-aminolevulinic acid synthesis and lower levels of POR. Knockout mutants show enhanced sensitivity to reactive oxygen species and a slower greening of etiolated seedlings. Based on our studies, the interaction of TTP1 with GluTR and POR does not directly inhibit their enzymatic activity and contribute to the control of 5-aminolevulinic acid synthesis. Instead, we propose that TTP1 sequesters a fraction of these proteins on the thylakoid membrane, and contributes to their stability.

RALDH1 Inhibition Shows Immunotherapeutic Efficacy in Hepatocellular Carcinoma.

Globally, hepatocellular carcinoma (HCC) is one of the most commonly diagnosed cancers and a leading cause of cancer-related death. We previously identified an immune evasion pathway whereby tumor cells produce retinoic acid (RA) to promote differentiation of intratumoral monocytes into protumor macrophages. Retinaldehyde dehydrogenase 1 (RALDH1), RALDH2, and RALDH3 are the three isozymes that catalyze RA biosynthesis. In this study, we have identified RALDH1 as the key driver of RA production in HCC and demonstrated the efficacy of RALDH1-selective inhibitors (Raldh1-INH) in suppressing RA production by HCC cells. Raldh1-INH restrained tumor growth in multiple mouse models of HCC by reducing the number and tumor-supporting functions of intratumoral macrophages as well as increasing T-cell infiltration and activation within tumors. Raldh1-INH also displayed favorable pharmacokinetic, pharmacodynamic, and toxicity profiles in mice thereby establishing them as promising new drug candidates for HCC immunotherapy.