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International Journal For Parasitology.... Jun 2024Cryptosporidium parvum is a waterborne and foodborne zoonotic protozoan parasite, a causative agent of moderate to severe diarrheal diseases in humans and animals....
Lower micromolar activity of the antifungal imidazoles on the bacterial-type bifunctional aldehyde/alcohol dehydrogenase (AdhE) in Cryptosporidium parvum and in vitro efficacy against the zoonotic parasite.
Cryptosporidium parvum is a waterborne and foodborne zoonotic protozoan parasite, a causative agent of moderate to severe diarrheal diseases in humans and animals. However, fully effective treatments are unavailable for medical and veterinary uses. There is a need to explore new drug targets for potential development of new therapeutics. Because C. parvum relies on anaerobic metabolism to produce ATP, fermentative enzymes in this parasite are attractive targets for exploration. In this study, we investigated the ethanol-fermentation in the parasite and characterized the basic biochemical features of a bacterial-type bifunctional aldehyde/alcohol dehydrogenase, namely CpAdhE. We also screened 3892 chemical entries from three libraries and identified 14 compounds showing >50% inhibition on the enzyme activity of CpAdhE. Intriguingly, antifungal imidazoles and unsaturated fatty acids are the two major chemical groups among the top hits. We further characterized the inhibitory kinetics of selected imidazoles and unsaturated fatty acids on CpAdhE. These compounds displayed lower micromolar activities on CpAdhE (i.e., IC values ranging from 0.88 to 11.02 μM for imidazoles and 8.93 to 35.33 μM for unsaturated fatty acids). Finally, we evaluated the in vitro anti-cryptosporidial efficacies and cytotoxicity of three imidazoles (i.e., tioconazole, miconazole and isoconazole). The three antifungal imidazoles exhibited lower micromolar efficacies against the growth of C. parvum in vitro (EC values ranging from 4.85 to 10.41 μM and selectivity indices ranging from 5.19 to 10.95). The results provide a proof-of-concept data to support that imidazoles are worth being further investigated for potential development of anti-cryptosporidial therapeutics.
PubMed: 38875756
DOI: 10.1016/j.ijpddr.2024.100551 -
Frontiers in Microbiology 2024Extremophilic proteins are valuable in various fields, but their expression can be challenging in traditional hosts like due to misfolding and aggregation. (), a...
Extremophilic proteins are valuable in various fields, but their expression can be challenging in traditional hosts like due to misfolding and aggregation. (), a halophilic expression system, offers a solution. This study examined cleavable and non-cleavable purification tags at both the N- and C-termini when fused with the superfolder green fluorescent protein (sfGFP) in . Our findings reveal that an N-terminal 8xHis-tag or Strep-tagII significantly enhances protein production, purity, and yield in . Further experiments with mCherry and halophilic alcohol dehydrogenase (ADH) showed improved expression and purification yields when the 8xHis-tag or Strep-tagII was positioned at the C-terminus for mCherry and at the N-terminus for ADH. Co-positioning 8xHis-tag and Twin-Strep-tag at the N-terminus of sfGFP, mCherry, and ADH yielded significantly enhanced results. These findings highlight the importance of thoughtful purification tag design and selection in , providing valuable insights for improving protein production and purification with the potential to advance biotechnological applications.
PubMed: 38873150
DOI: 10.3389/fmicb.2024.1403623 -
Proceedings of the National Academy of... Jun 2024Alcohol dehydrogenase 1B (ADH1B) is a primate-specific enzyme which, uniquely among the ADH class 1 family, is highly expressed both in adipose tissue and liver. Its...
Alcohol dehydrogenase 1B (ADH1B) is a primate-specific enzyme which, uniquely among the ADH class 1 family, is highly expressed both in adipose tissue and liver. Its expression in adipose tissue is reduced in obesity and increased by insulin stimulation. Interference with expression has also been reported to impair adipocyte function. To better understand the role of ADH1B in adipocytes, we used CRISPR/Cas9 to delete in human adipose stem cells (ASC). Cells lacking ADH1B failed to differentiate into mature adipocytes manifested by minimal triglyceride accumulation and a marked reduction in expression of established adipocyte markers. As ADH1B is capable of converting retinol to retinoic acid (RA), we conducted rescue experiments. Incubation of ADH1B-deficient preadipocytes with 9-cis-RA, but not with all-transretinol, significantly rescued their ability to accumulate lipids and express markers of adipocyte differentiation. A homozygous missense variant in (p.Arg313Cys) was found in a patient with congenital lipodystrophy of unknown cause. This variant significantly impaired the protein's dimerization, enzymatic activity, and its ability to rescue differentiation in ADH1B-deficient ASC. The allele frequency of this variant in the Middle Eastern population suggests that it is unlikely to be a fully penetrant cause of severe lipodystrophy. In conclusion, ADH1B appears to play an unexpected, crucial and cell-autonomous role in human adipocyte differentiation by serving as a necessary source of endogenous retinoic acid.
Topics: Humans; Alcohol Dehydrogenase; Adipogenesis; Adipocytes; Tretinoin; Cell Differentiation; CRISPR-Cas Systems; Mutation, Missense; Adipose Tissue
PubMed: 38838011
DOI: 10.1073/pnas.2319301121 -
Frontiers in Plant Science 2024Abscisic acid (ABA) is a key phytohormone involved in wound healing in fruits and vegetables, while fluridone (FLD) is its synthetic inhibitor. However, it is unknown...
Abscisic acid (ABA) is a key phytohormone involved in wound healing in fruits and vegetables, while fluridone (FLD) is its synthetic inhibitor. However, it is unknown whether ABA signaling and downstream transcription factors are involved in the synthesis of phenolic acids and lignin monomers in muskmelon wounds, and the underlying mechanisms. In our study, exogenous ABA promoted endogenous ABA synthesis by increasing the levels of β-carotenoid and zeaxanthin, activating 9-cis-epoxycarotenoid dioxygenase (NCED) and zeaxanthin epoxidase (ZEP), facilitated ABA signaling by increasing the expression levels of protein phosphatases type 2C () and ABA-responsive element binding factors (), upregulated the expression levels of and , and ABA induced phenylpropanoid metabolism by activating phenylalanine ammonia-lyase (PAL), 4-coenzyme A ligase (4CL), and cinnamyl alcohol dehydrogenase (CAD), which further increased the synthesis of phenolic acids and lignin monomers in muskmelon wounds during healing. Taken together, exogenous ABA induced phenylpropanoid metabolism and increased the synthesis of phenolic acid and lignin monomer in muskmelon wounds during healing, and may be involved in endogenous ABA synthesis and signaling and related transcription factors.
PubMed: 38835857
DOI: 10.3389/fpls.2024.1404477 -
Arthritis Research & Therapy Jun 2024Gout is a prevalent manifestation of metabolic osteoarthritis induced by elevated blood uric acid levels. The purpose of this study was to investigate the mechanisms of...
BACKGROUND
Gout is a prevalent manifestation of metabolic osteoarthritis induced by elevated blood uric acid levels. The purpose of this study was to investigate the mechanisms of gene expression regulation in gout disease and elucidate its pathogenesis.
METHODS
The study integrated gout genome-wide association study (GWAS) data, single-cell transcriptomics (scRNA-seq), expression quantitative trait loci (eQTL), and methylation quantitative trait loci (mQTL) data for analysis, and utilized two-sample Mendelian randomization study to comprehend the causal relationship between proteins and gout.
RESULTS
We identified 17 association signals for gout at unique genetic loci, including four genes related by protein-protein interaction network (PPI) analysis: TRIM46, THBS3, MTX1, and KRTCAP2. Additionally, we discerned 22 methylation sites in relation to gout. The study also found that genes such as TRIM46, MAP3K11, KRTCAP2, and TM7SF2 could potentially elevate the risk of gout. Through a Mendelian randomization (MR) analysis, we identified three proteins causally associated with gout: ADH1B, BMP1, and HIST1H3A.
CONCLUSION
According to our findings, gout is linked with the expression and function of particular genes and proteins. These genes and proteins have the potential to function as novel diagnostic and therapeutic targets for gout. These discoveries shed new light on the pathological mechanisms of gout and clear the way for future research on this condition.
Topics: Gout; Humans; Mendelian Randomization Analysis; Genome-Wide Association Study; Genetic Predisposition to Disease; Quantitative Trait Loci; Single-Cell Analysis; DNA Methylation; Polymorphism, Single Nucleotide; Protein Interaction Maps; Alcohol Dehydrogenase
PubMed: 38831441
DOI: 10.1186/s13075-024-03348-z -
International Journal of Nanomedicine 2024, a dual-purpose food and medicine, displays limited efficacy in alcohol detoxification and liver protection, with previous research primarily focused on puerarin in its...
PURPOSE
, a dual-purpose food and medicine, displays limited efficacy in alcohol detoxification and liver protection, with previous research primarily focused on puerarin in its dried roots. In this study, we investigated the potential effects and mechanisms of fresh root-derived exosome-like nanovesicles (P-ELNs) for mitigating alcoholic intoxication, promoting alcohol metabolism effects and protecting the liver in C57BL/6J mice.
METHODS
We isolated P-ELNs from fresh root using differential centrifugation and characterized them via transmission electron microscopy, nanoscale particle sizing, ζ potential analysis, and biochemical assays. In Acute Alcoholism (AAI) mice pre-treated with P-ELNs, we evaluated their effects on the timing and duration of the loss of the righting reflex (LORR), liver alcohol metabolism enzymes activity, liver and serum alcohol content, and ferroptosis-related markers.
RESULTS
P-ELNs, enriched in proteins, lipids, and small RNAs, exhibited an ideal size (150.7 ± 82.8 nm) and negative surface charge (-31 mV). Pre-treatment with 10 mg/(kg.bw) P-ELNs in both male and female mice significantly prolonged ebriety time, shortened sobriety time, enhanced acetaldehyde dehydrogenase (ALDH) activity while concurrently inhibited alcohol dehydrogenase (ADH) activity, and reduced alcohol content in the liver and serum. Notably, P-ELNs demonstrated more efficacy compared to P-ELNs supernatant fluid (abundant puerarin content), suggesting alternative active components beyond puerarin. Additionally, P-ELNs prevented ferroptosis by inhibiting the reduction of glutathione peroxidase 4 (GPX4) and reduced glutathione (GSH), and suppressing acyl-CoA synthetase long-chain family member 4 (ACSL4) elevation, thereby mitigating pathological liver lipid accumulation.
CONCLUSION
P-ELNs exhibit distinct exosomal characteristics and effectively alleviate alcoholic intoxication, improve alcohol metabolism, suppress ferroptosis, and protect the liver from alcoholic injury. Consequently, P-ELNs hold promise as a therapeutic agent for detoxification, sobriety promotion, and prevention of alcoholic liver injury.
Topics: Animals; Pueraria; Mice, Inbred C57BL; Exosomes; Mice; Male; Alcoholic Intoxication; Plant Roots; Liver; Ethanol; Plant Extracts; Alcoholism; Isoflavones
PubMed: 38828197
DOI: 10.2147/IJN.S462602 -
Scientific Reports Jun 2024Observational studies have shown controversial associations between alcohol intake and radiographic osteoarthritis (OA). This study investigated whether this association...
Observational studies have shown controversial associations between alcohol intake and radiographic osteoarthritis (OA). This study investigated whether this association was causal using a Mendelian randomization (MR) study in a population-based cohort in Korean. The study enrolled 2429 subjects (1058 men, 1371 women) from the Dong-gu Study. X-rays of the hand and knee joints were scored using a semi-quantitative grading system to calculate the total score of the hand and knee joints. ALDH2 rs671 genotyping was performed by high-resolution melting analysis. MR instrumental variable analysis and observational multivariable regression analysis were used to estimate the association between genetically predicted alcohol intake and the radiographic severity of OA. Subjects with the G/G genotype had a higher current alcohol intake than those with the G/A and A/A genotypes in both men and women (all P < 0.001). Men with the G/G genotype had higher total knee (P < 0.001) and hand scores (P = 0.042) compared to those with the G/A and A/A genotypes after adjusting for age and body mass index, but not in women. In the observational multivariable regression analysis, each alcohol drink per day in men was associated with increased knee (P = 0.001) and hand joint scores (P = 0.013) after adjustment, but not in women. In our MR analysis, utilizing ALDH2 rs671 genotypes as instrumental variables for alcohol consumption, has shown a significant link between each additional daily alcohol drink and increased radiographic joint severity in men.
Topics: Humans; Male; Alcohol Drinking; Female; Middle Aged; Osteoarthritis, Knee; Aldehyde Dehydrogenase, Mitochondrial; Osteoarthritis; Aged; Radiography; Severity of Illness Index; Hand Joints; Genotype; Mendelian Randomization Analysis; Polymorphism, Single Nucleotide; Knee Joint
PubMed: 38825629
DOI: 10.1038/s41598-024-63559-x -
BMC Genomics May 2024The citral-type is the most common chemotype in Cinnamomum bodinieri Levl (C. bodinieri), which has been widely used in the daily necessities, cosmetics, biomedicine,...
The citral-type is the most common chemotype in Cinnamomum bodinieri Levl (C. bodinieri), which has been widely used in the daily necessities, cosmetics, biomedicine, and aromatic areas due to their high citral content. Despite of this economic prospect, the possible gene-regulatory roles of citral biosynthesis in the same geographic environment remains unknown. In this study, the essential oils (EOs) of three citral type (B1, B2, B3) and one non-citral type (B0) varieties of C. bodinieri were identified by GC-MS after hydrodistillation extraction in July. 43 components more than 0.10% were identified in the EOs, mainly composed of monoterpenes (75.8-91.84%), and high content citral (80.63-86.33%) were identified in citral-type. Combined transcriptome and metabolite profiling analysis, plant-pathogen interaction(ko04626), MAPK signaling pathway-plant(ko04016), starch and sucrose metabolism(ko00500), plant hormone signal transduction(ko04075), terpenoid backbone biosynthesis (ko00900) and monoterpenoid biosynthesis (ko00902) pathways were enriched significantly. The gene expression of differential genes were linked to the monoterpene content, and the geraniol synthase (CbGES), alcohol dehydrogenase (CbADH), geraniol 8-hydroxylase-like (CbCYP76B6-like) and 8-hydroxygeraniol dehydrogenase (Cb10HGO) were upregulated in the citral-type, indicating that they were associated with high content of geraniol and citral. The activities of CbGES and CbADH in citral type were higher than in non-citral type, which was corroborated by enzyme-linked immunosorbent assay (ELISA). This study on the accumulation mechanism of citral provides a theoretical basis for the development of essential oil of C. bodinieri.
Topics: Cinnamomum; Acyclic Monoterpenes; Monoterpenes; Gene Expression Profiling; Transcriptome; Oils, Volatile; Gene Expression Regulation, Plant; Genes, Plant
PubMed: 38822238
DOI: 10.1186/s12864-024-10419-7 -
Rice (New York, N.Y.) May 2024Rice is one of the most important food crops in the world, and with the development of direct seeding methods for rice, exposure to anaerobic stress has become a major...
Identification of Key Genes and Pathways for Anaerobic Germination Tolerance in Rice Using Weighted Gene Co-Expression Network Analysis (WGCNA) in Association with Quantitative Trait Locus (QTL) Mapping.
BACKGROUND
Rice is one of the most important food crops in the world, and with the development of direct seeding methods for rice, exposure to anaerobic stress has become a major factor limiting its growth.
RESULTS
In this experiment, we tested the tolerance to anaerobic germination of rice varieties NIP and HD84, and they were used as parents to construct a DH (doubled-haploid) population. The transcriptomes of NIP (highly tolerant) and HD86 (intolerant), and their progeny HR (highly tolerant) and NHR (intolerant) were sequenced from normal and anaerobic environments. The differentially-expressed genes (DEGs) were subjected to GO (Gene ontology), KEGG (Kyoto Encyclopedia of Genes and Genomes), and WGCNA analyses. QTL mapping of the DH population identified tolerance to anaerobic germination-related chromosomal segments. The transcriptome results from 24 samples were combined with the anaerobic stress QTL results for 159 DH population lines to construct a metabolic network to identify key pathways and a gene interaction network to study the key genes. Essential genes were initially subjected to rigorous functional validation, followed by a comprehensive analysis aimed at elucidating their potential utility in domestication and breeding practices, particularly focusing on the exploitation of dominant haplotypes.
CONCLUSION
The results show that pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH) are the starting signals of energy metabolism for coleoptile length growth, the auxin transporter EXPA is the determining signal for coleoptile length growth. The pivotal genes Os05g0498700 and Os01g0866100 exert a negative regulatory influence on coleoptile length, ultimately enhancing tolerance to anaerobic germination in rice. Analyses of breeding potential underscore the additional value of Os05g0498700-hyp2 and Os01g0866100-hyp2, highlighting their potential utility in further improving rice through breeding programs. The results of our study will provide a theoretical basis for breeding anaerobic-tolerant rice varieties.
PubMed: 38819744
DOI: 10.1186/s12284-024-00714-y -
Heliyon May 2024It is important to accurately determine the blood ethanol concentration (BEC) to ensure appropriate diagnosis and treatment of patients in the emergency department (ED)...
BACKGROUND
It is important to accurately determine the blood ethanol concentration (BEC) to ensure appropriate diagnosis and treatment of patients in the emergency department (ED) and protect their legal rights. This study aimed to determine whether sterilization of venipuncture site with ethanol, which is frequently used in practice in the ED would affect BEC.
METHODS
Venous blood samples were collected by two consecutive techniques from 94 individuals who were admitted to the ED, had an indication for BEC measurement, and volunteered to participate in the study. The reference technique involved applying 3 cc of 10 % povidone-iodine solution to a gauze pad, cleaning the right arm antecubital region, and performing phlebotomy. The index technique used 3 cc of alcohol-based antiseptic (89 % ethanol) on another gauze for cleaning the left arm antecubital region. Both techniques allowed the antiseptic to air-dry for 30 s before phlebotomy. Two blood sample tubes per patient were sent to the laboratory, and BEC were measured using the alcohol dehydrogenase enzymatic method.
RESULTS
94 patients were included in the study. The mean age was 37.8 years (±15.7), with 77 % (n = 72) of them were male. The median BEC levels measured by both the reference and index techniques were 2 mg/dL (IQR: 0.97-16.25) and 2 mg/dL (IQR: 0.90-15.22), respectively, with no significant statistical difference (p = 0.536). 72 (77 %) of the patients had a BEC level below the legal driving limit of 20 mg/dL. Bland-Altman analysis, performed on these patients, revealed a small negative bias, -0.116 mg/dL with a standard deviation of 1.13 mg/dL. The upper and lower limit of the agreement was 2.092 and -2.323 respectively.
CONCLUSION
In patients with a BEC level of less than 20 mg/dL, using ethanol-containing antiseptics before blood sampling does not lead to erroneously elevated BEC levels.
PubMed: 38813189
DOI: 10.1016/j.heliyon.2024.e31517