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International Journal of Molecular... Jan 2023Hepatitis C virus (HCV) frequently causes chronic infection in the human liver, which may progress to advanced hepatic fibrosis, cirrhosis, and hepatocellular carcinoma.... (Review)
Review
Hepatitis C virus (HCV) frequently causes chronic infection in the human liver, which may progress to advanced hepatic fibrosis, cirrhosis, and hepatocellular carcinoma. HCV primarily infects highly differentiated quiescent hepatocytes and can modulate cell cycle-regulatory genes and proliferation pathways, which ultimately contribute to persistent infection and pathogenesis. On the other hand, several studies have shown differential regulation of HCV RNA and viral protein expression levels, depending on the proliferation state of hepatocytes and the phase of the cell cycle. HCV typically requires factors provided by host cells for efficient and persistent viral replication. Previously, we found that HCV infection upregulates the expression of ribonucleotide reductase subunit M2 (RRM2) in quiescent hepatocytes. RRM2 is a rate-limiting protein that catalyzes de novo synthesis of deoxyribonucleotide triphosphates, and its expression is highly regulated during various phases of the cell cycle. RRM2 functions as a pro-viral factor essential for HCV RNA synthesis, but its functional role in HCV-induced liver diseases remains unknown. Here, we present a comprehensive review of the role of the hepatocyte cell cycle, in correlation with RRM2 expression, in the regulation of HCV replication. We also discuss the potential relevance of this protein in the pathogenesis of HCV, particularly in the development of hepatocellular carcinoma.
Topics: Humans; Hepacivirus; Carcinoma, Hepatocellular; Hepatitis C; Hepatocytes; Liver Neoplasms; Virus Replication; Liver Cirrhosis; RNA
PubMed: 36768940
DOI: 10.3390/ijms24032619 -
Biomedicine & Pharmacotherapy =... Apr 2023Elevated myocardial intracellular sodium ([Na]) was shown to decrease mitochondrial calcium ([Ca]) via mitochondrial sodium/calcium exchanger (NCX), resulting in...
BACKGROUND
Elevated myocardial intracellular sodium ([Na]) was shown to decrease mitochondrial calcium ([Ca]) via mitochondrial sodium/calcium exchanger (NCX), resulting in decreased mitochondrial ATP synthesis. The sodium-glucose co-transporter 2 inhibitor (SGLT2i) ertugliflozin (ERTU) improved energetic deficit and contractile dysfunction in a mouse model of high fat, high sucrose (HFHS) diet-induced diabetic cardiomyopathy (DCMP). As SGLT2is were shown to lower [Na] in isolated cardiomyocytes, we hypothesized that energetic improvement in DCMP is at least partially mediated by a decrease in abnormally elevated myocardial [Na].
METHODS
Forty-two eight-week-old male C57BL/6J mice were fed a control or HFHS diet for six months. In the last month, a subgroup of HFHS-fed mice was treated with ERTU. At the end of the study, left ventricular contractile function and energetics were measured simultaneously in isolated beating hearts by P NMR (Nuclear Magnetic Resonance) spectroscopy. A subset of untreated HFHS hearts was perfused with vehicle vs. CGP 37157, an NCX inhibitor. Myocardial [Na] was measured by Na NMR spectroscopy.
RESULTS
HFHS hearts showed diastolic dysfunction, decreased contractile reserve, and impaired energetics as reflected by decreased phosphocreatine (PCr) and PCr/ATP ratio. Myocardial [Na] was elevated > 2-fold in HFHS (vs. control diet). ERTU reversed the impairments in HFHS hearts to levels similar to or better than control diet and decreased myocardial [Na] to control levels. CGP 37157 normalized the PCr/ATP ratio in HFHS hearts.
CONCLUSIONS
Elevated myocardial [Na] contributes to mitochondrial and contractile dysfunction in DCMP. Targeting myocardial [Na] and/or NCX may be an effective strategy in DCMP and other forms of heart disease associated with elevated myocardial [Na].
Topics: Mice; Male; Animals; Diabetic Cardiomyopathies; Sodium-Glucose Transporter 2 Inhibitors; Sodium; Calcium; Deoxycytidine Monophosphate; Myocardial Contraction; Mice, Inbred C57BL; Myocardium; Adenosine Triphosphate; Diabetes Mellitus
PubMed: 36731341
DOI: 10.1016/j.biopha.2023.114310 -
Nucleosides, Nucleotides & Nucleic Acids 2023We describe a new demethylation method for dimethyl phosphonate esters using sodium ethanethiolate. The new procedure allows demethylation of nucleoside dimethyl...
We describe a new demethylation method for dimethyl phosphonate esters using sodium ethanethiolate. The new procedure allows demethylation of nucleoside dimethyl phosphonate esters without 1'-α-anomerization, providing an improved synthesis of 5'-methylene substituted 2',5'-deoxynucleotides.
Topics: Esters; Organophosphonates; Deoxyribonucleotides; Demethylation
PubMed: 36629008
DOI: 10.1080/15257770.2023.2166064 -
Medicine Dec 2022This prospective cohort study explored whether body constitution (BC) independently predicts new-onset albuminuria in persons with type 2 diabetes mellitus (T2DM)...
Traditional Chinese medicine body constitution predicts new-onset diabetic albuminuria in patients with type 2 diabetes: Taichung diabetic body constitution prospective cohort study.
This prospective cohort study explored whether body constitution (BC) independently predicts new-onset albuminuria in persons with type 2 diabetes mellitus (T2DM) enrolled in the diabetes care management program (DCMP) of a medical center, providing evidence of integrating traditional Chinese medicine into DCMP for improving care quality. Persons with T2DM (n = 426) originally without albuminuria enrolled in DCMP were recruited in 2010 and were then followed up to 2015 for detecting new-onset albuminuria. The participants received urinalysis and blood test annually. Albuminuria was determined by an elevated urinary albumin/creatinine ratio (≥ 30 µg/mg), and poor glucose control was defined as Glycosylated hemoglobin above or equal to 7%. BC type (Yin deficiency, Yang deficiency, and phlegm stasis) was assessed using a well-validated body constitution questionnaire at baseline. Risk factors for albuminuria (sociodemographic factors, diabetes history, lifestyle behaviors, lipid profile, blood pressure, and kidney function) were also recorded. Hazard ratios (HR) of albuminuria for BC were estimated using multivariate Cox proportional hazards model. During the 4-year follow-up period, albuminuria occurred in 30.5% of participants (n = 130). The HR indicated that Yin deficiency was significantly associated with an increased risk of new-onset albuminuria in persons with T2DM and good glucose control after adjustment for other risk factors (HR = 2.09; 95% confidence interval = 1.05-4.17, P = .04), but not in those with poor glucose control. In persons with T2DM and poor glucose control, phlegm stasis was also significantly associated with a higher risk of albuminuria (2.26; 1.03-4.94, P = .04) after multivariate adjustment, but not in those with good glucose control. In addition to already-known risk factors, BC is an independent and significant factor associated with new-onset albuminuria in persons with T2DM. Our results imply Yin deficiency and phlegm stasis interacting with glucose control status may affect new-onset albuminuria in persons with T2DM.
Topics: Humans; Albuminuria; Blood Glucose; Body Constitution; Deoxycytidine Monophosphate; Diabetes Mellitus, Type 2; Diabetic Nephropathies; Medicine, Chinese Traditional; Prospective Studies; Yin Deficiency
PubMed: 36550881
DOI: 10.1097/MD.0000000000032342 -
ELife Dec 2022Replication of the genome must be coordinated with gene transcription and cellular metabolism, especially following replication stress in the presence of limiting...
Replication of the genome must be coordinated with gene transcription and cellular metabolism, especially following replication stress in the presence of limiting deoxyribonucleotides. The Rad53 (CHEK2 in mammals) checkpoint kinase plays a major role in cellular responses to DNA replication stress. Cell cycle regulated, genome-wide binding of Rad53 to chromatin was examined. Under replication stress, the kinase bound to sites of active DNA replication initiation and fork progression, but unexpectedly to the promoters of about 20% of genes encoding proteins involved in multiple cellular functions. Rad53 promoter binding correlated with changes in expression of a subset of genes. Rad53 promoter binding to certain genes was influenced by sequence-specific transcription factors and less by checkpoint signaling. However, in checkpoint mutants, untimely activation of late-replicating origins reduces the transcription of nearby genes, with concomitant localization of Rad53 to their gene bodies. We suggest that the Rad53 checkpoint kinase coordinates genome-wide replication and transcription under replication stress conditions.
Topics: Protein Serine-Threonine Kinases; Cell Cycle Proteins; Saccharomyces cerevisiae Proteins; Checkpoint Kinase 2; DNA Replication; Saccharomyces cerevisiae; Cell Cycle; Cell Cycle Checkpoints; DNA Damage; Phosphorylation
PubMed: 36520028
DOI: 10.7554/eLife.84320 -
Langmuir : the ACS Journal of Surfaces... Dec 2022The fate of biomolecules in the environment depends in part on understanding the surface chemistry occurring at the biological-geochemical (bio-geo) interface. Little is...
The fate of biomolecules in the environment depends in part on understanding the surface chemistry occurring at the biological-geochemical (bio-geo) interface. Little is known about how environmental DNA (eDNA) or smaller components, like nucleotides and oligonucleotides, persist in aquatic environments and the role of surface interactions. This study aims to probe surface interactions and adsorption behavior of nucleotides on oxide surfaces. We have investigated the interactions of individual nucleotides (dGMP, dCMP, dAMP, and dTMP) on TiO particle surfaces as a function of pH and in the presence of complementary and noncomplementary base pairs. Using attenuated total reflectance-Fourier transform infrared spectroscopy, there is an increased number of adsorbed nucleotides at lower pH with a preferential interaction of the phosphate group with the oxide surface. Additionally, differential adsorption behavior is seen where purine nucleotides are preferentially adsorbed, with higher surface saturation coverage, over their pyrimidine derivatives. These differences may be a result of intermolecular interactions between coadsorbed nucleotides. When the TiO surface was exposed to two-component solutions of nucleotides, there was preferential adsorption of dGMP compared to dCMP and dTMP, and dAMP compared to dTMP and dCMP. Complementary nucleotide base pairs showed hydrogen-bond interactions between a strongly adsorbed purine nucleotide layer and a weaker interacting hydrogen-bonded pyrimidine second layer. Noncomplementary base pairs did not form a second layer. These results highlight several important findings: (i) there is differential adsorption of nucleotides; (ii) complementary coadsorbed nucleotides show base pairing with a second layer, and the stability depends on the strength of the hydrogen bonding interactions and; (iii) the first layer coverage strongly depends on pH. Overall, the importance of surface interactions in the adsorption of nucleotides and the templating of specific interactions between nucleotides are discussed.
Topics: Thymidine Monophosphate; Deoxycytidine Monophosphate; Oxides; Hydrogen Bonding; Hydrogen
PubMed: 36445255
DOI: 10.1021/acs.langmuir.2c01604 -
Scientific Reports Nov 2022Immunotherapy has a number of advantages over traditional anti-tumor therapy but can cause severe adverse reactions due to an overactive immune system. In contrast, a...
Immunotherapy has a number of advantages over traditional anti-tumor therapy but can cause severe adverse reactions due to an overactive immune system. In contrast, a novel metabolic treatment approach can induce metabolic vulnerability through multiple cancer cell targets. Here, we show a therapeutic effect by inducing nucleotide imbalance and apoptosis in triple negative breast cancer cells (TNBC), by treating with cytosolic thymidylate 5'-phosphohydrolase (CT). We show that a sustained consumption of dTMP by CT could induce dNTP imbalance, leading to apoptosis as tricarboxylic acid cycle intermediates were depleted to mitigate this imbalance. These cytotoxic effects appeared to be different, depending on substrate specificity of the 5' nucleotide or metabolic dependency of the cancer cell lines. Using representative TNBC cell lines, we reveal how the TNBC cells were affected by CT-transfection through extracellular acidification rate (ECAR)/oxygen consumption rate (OCR) analysis and differential transcription/expression levels. We suggest a novel approach for treating refractory TNBC by an mRNA drug that can exploit metabolic dependencies to exacerbate cell metabolic vulnerability.
Topics: Humans; Triple Negative Breast Neoplasms; Thymidine Monophosphate; Cell Line, Tumor; Apoptosis; Phosphoric Monoester Hydrolases
PubMed: 36414668
DOI: 10.1038/s41598-022-24706-4 -
International Journal of Molecular... Nov 2022The genome editing approach using the components of the CRISPR/Cas system has found wide application in molecular biology, fundamental medicine and genetic engineering....
The genome editing approach using the components of the CRISPR/Cas system has found wide application in molecular biology, fundamental medicine and genetic engineering. A promising method is to increase the efficacy and specificity of CRISPR/Cas-based genome editing systems by modifying their components. Here, we designed and chemically synthesized guide RNAs (crRNA, tracrRNA and sgRNA) containing modified nucleotides (2'-O-methyl, 2'-fluoro, LNA-locked nucleic acid) or deoxyribonucleotides in certain positions. We compared their resistance to nuclease digestion and examined the DNA cleavage efficacy of the CRISPR/Cas9 system guided by these modified guide RNAs. The replacement of ribonucleotides with 2'-fluoro modified or LNA nucleotides increased the lifetime of the crRNAs, while other types of modification did not change their nuclease resistance. Modification of crRNA or tracrRNA preserved the efficacy of the CRISPR/Cas9 system. Otherwise, the CRISPR/Cas9 systems with modified sgRNA showed a remarkable loss of DNA cleavage efficacy. The kinetic constant of DNA cleavage was higher for the system with 2'-fluoro modified crRNA. The 2'-modification of crRNA also decreased the off-target effect upon in vitro dsDNA cleavage.
Topics: CRISPR-Cas Systems; Endonucleases; Gene Editing; Nucleotides; RNA, Small Untranslated
PubMed: 36362256
DOI: 10.3390/ijms232113460 -
Current Opinion in Structural Biology Dec 2022Ribonucleotide reductases (RNRs) use radical-based chemistry to convert ribonucleotides into deoxyribonucleotides, an essential step in DNA biosynthesis and repair.... (Review)
Review
Ribonucleotide reductases (RNRs) use radical-based chemistry to convert ribonucleotides into deoxyribonucleotides, an essential step in DNA biosynthesis and repair. There are multiple RNR classes, the best studied of which is the class Ia RNR that is found in Escherichia coli, eukaryotes including humans, and many pathogenic and nonpathogenic prokaryotes. This review covers recent advances in our understanding of class Ia RNRs, including a recent reporting of a structure of the active state of the E. coli enzyme and the impacts that the structure has had on spurring research into the mechanism of long-range radical transfer. Additionally, the review considers other recent structural and biochemical research on class Ia RNRs and the potential of that work for the development of anticancer and antibiotic therapeutics.
Topics: Humans; Ribonucleotide Reductases; Escherichia coli
PubMed: 36272229
DOI: 10.1016/j.sbi.2022.102489 -
Molecules (Basel, Switzerland) Sep 2022In 2002, a new class of thymidylate synthase (TS) involved in the de novo synthesis of dTMP named Flavin-Dependent Thymidylate Synthase (FDTS) encoded by the thyX gene...
In 2002, a new class of thymidylate synthase (TS) involved in the de novo synthesis of dTMP named Flavin-Dependent Thymidylate Synthase (FDTS) encoded by the thyX gene was discovered; FDTS is present only in 30% of prokaryote pathogens and not in human pathogens, which makes it an attractive target for the development of new antibacterial agents, especially against multi-resistant pathogens. We report herein the synthesis and structure-activity relationship of a novel series of hitherto unknown pyrido[1,2-e]purine-2,4(1H,3H)-dione analogues. Several synthetics efforts were done to optimize regioselective N1-alkylation through organopalladium cross-coupling. Modelling of potential hits were performed to generate a model of interaction into the active pocket of FDTS to understand and guide further synthetic modification. All those compounds were evaluated on an in-house in vitro NADPH oxidase assays screening as well as against Mycobacterium tuberculosis ThyX. The highest inhibition was obtained for compound 23a with 84.3% at 200 µM without significant cytotoxicity (CC50 > 100 μM) on PBM cells.
Topics: Anti-Bacterial Agents; Dinitrocresols; Flavins; Humans; Mycobacterium tuberculosis; NADPH Oxidases; Purines; Structure-Activity Relationship; Thymidine Monophosphate; Thymidylate Synthase
PubMed: 36234754
DOI: 10.3390/molecules27196216