bluuebunny/crossref_metadata_2025 huggingface dataset:
Diacylglycerol Kinase alpha in X Linked Lymphoproliferative Disease Type 1
Authors: Velnati Suresh , Centonze Sara , Girivetto Federico , Baldanzi Gianluca
Published: May 2021

Diacylglycerol kinases are intracellular enzymes that control the balance between the secondary messengers diacylglycerol and phosphatidic acid. DGKα and DGKζ are the prominent isoforms that restrain the intensity of T cell receptor signalling by metabolizing PLCγ generated diacylglycerol. Thus, their activity must be tightly controlled to grant cellular homeostasis and refine immune responses. DGKα is specifically inhibited by strong T cell activating signals to allow for full diacylglycerol signalling which mediates T cell response. In X-linked lymphoproliferative disease 1, deficiency of the adaptor protein SAP results in altered T cell receptor signalling, due in part to persistent DGKα activity. This activity constrains diacylglycerol levels, attenuating downstream pathways such as PKCθ and Ras/MAPK and decreasing T cell restimulation induced cell death. This is a form of apoptosis triggered by prolonged T cell activation that is indeed defective in CD8+ cells of X-linked lymphoproliferative disease type 1 patients. Accordingly, inhibition or downregulation of DGKα activity restores in vitro a correct diacylglycerol dependent signal transduction, cytokines production and restimulation induced apoptosis. In animal disease models, DGKα inhibitors limit CD8+ expansion and immune-mediated tissue damage, suggesting the possibility of using inhibitors of diacylglycerol kinase as a new therapeutic approach.
Trypsin-Like Proteases and Their Role in Muco-Obstructive Lung Diseases
Authors: Carroll Emma L. , Bailo Mariarca , Reihill James A. , Crilly Anne , Lockhart John C. , Litherland Gary J. , Lundy Fionnuala T. , McGarvey Lorcan P. , Hollywood Mark A. , Martin S. Lorraine
Published: May 2021

Trypsin-like proteases (TLPs) belong to a family of serine enzymes with primary substrate specificities for the basic residues, lysine and arginine, in the P1 position. Whilst initially perceived as soluble enzymes that are extracellularly secreted, a number of novel TLPs that are anchored in the cell membrane have since been discovered. Muco-obstructive lung diseases (MucOLDs) are characterised by the accumulation of hyper-concentrated mucus in the small airways, leading to persistent inflammation, infection and dysregulated protease activity. Although neutrophilic serine proteases, particularly neutrophil elastase, have been implicated in the propagation of inflammation and local tissue destruction, it is likely that the serine TLPs also contribute to various disease-relevant processes given the roles that a number of these enzymes play in the activation of both the epithelial sodium channel (ENaC) and protease-activated receptor 2 (PAR2). More recently, significant attention has focused on the activation of viruses such as SARS-CoV-2 by host TLPs. The purpose of this review was to highlight key TLPs linked to the activation of ENaC and PAR2 and their association with airway dehydration and inflammatory signalling pathways, respectively. The role of TLPs in viral infectivity will also be discussed in the context of the inhibition of TLP activities and the potential of these proteases as therapeutic targets.
A Computational Model of Kidney Function in a Patient with Diabetes
Authors: Hu Rui , Layton Anita
Published: May 2021

At the onset of diabetes, the kidney grows large and the glomerular filtration rate becomes abnormally high. These structural and hemodynamics changes affect kidney function and may contribute to the development of chronic kidney disease. The goal of this study is to analyze how kidney function is altered in patients with diabetes and the renal effects of an anti-hyperglyceamic therapy that inhibits the sodium-glucose cotransporter 2 (SGLT2) in the proximal convoluted tubules. To accomplish that goal, we have developed a computational model of kidney function in a patient with diabetes and conducted simulations to study the effects of diabetes and SGLT2 inhibition on solute and water transport along the nephrons. Simulation results indicate that diabetes-induced hyperfiltration and tubular hypertrophy enhances Na+ transport, especially along the proximal tubules and thick ascending limbs. These simulations suggest that SGLT2 inhibition may attenuate glomerular hyperfiltration by limiting Na+-glucose transport, raising luminal [Cl−] at the macula densa, restoring the tubuloglomerular feedback signal, thereby reducing single-nephron glomerular filtration rate.
Cyclovirobuxine D Induced-Mitophagy through the p65/BNIP3/LC3 Axis Potentiates Its Apoptosis-Inducing Effects in Lung Cancer Cells
Authors: Zeng Cheng , Zou Tingting , Qu Junyan , Chen Xu , Zhang Suping , Lin Zhenghong
Published: May 2021

Mitophagy plays a pro-survival or pro-death role that is cellular-context- and stress-condition-dependent. In this study, we revealed that cyclovirobuxine D (CVB-D), a natural compound derived from Buxus microphylla, was able to provoke mitophagy in lung cancer cells. CVB-D-induced mitophagy potentiates apoptosis by promoting mitochondrial dysfunction. Mechanistically, CVB-D initiates mitophagy by enhancing the expression of the mitophagy receptor BNIP3 and strengthening its interaction with LC3 to provoke mitophagy. Our results further showed that p65, a transcriptional suppressor of BNIP3, is downregulated upon CVB-D treatment. The ectopic expression of p65 inhibits BNIP3 expression, while its knockdown significantly abolishes its transcriptional repression on BNIP3 upon CVB-D treatment. Importantly, nude mice bearing subcutaneous xenograft tumors presented retarded growth upon CVB-D treatment. Overall, we demonstrated that CVB-D treatment can provoke mitophagy and further revealed that the p65/BNIP3/LC3 axis is one potential mechanism involved in CVB-D-induced mitophagy in lung cancer cells, thus providing an effective antitumor therapeutic strategy for the treatment of lung cancer patients
Molecular Topology for the Search of New Anti-MRSA Compounds
Authors: Bueso-Bordils Jose I. , Alemán-López Pedro A. , Martín-Algarra Rafael , Duart Maria J. , Falcó Antonio , Antón-Fos Gerardo M.
Published: May 2021

The variability of methicillin-resistant Staphylococcus aureus (MRSA), its rapid adaptive response against environmental changes, and its continued acquisition of antibiotic resistance determinants have made it commonplace in hospitals, where it causes the problem of multidrug resistance. In this study, we used molecular topology to develop several discriminant equations capable of classifying compounds according to their anti-MRSA activity. Topological indices were used as structural descriptors and their relationship with anti-MRSA activity was determined by applying linear discriminant analysis (LDA) on a group of quinolones and quinolone-like compounds. Four extra equations were constructed, named DFMRSA1, DFMRSA2, DFMRSA3 and DFMRSA4 (DFMRSA was built in a previous study), all with good statistical parameters, such as Fisher–Snedecor F (>68 in all cases), Wilk’s lambda (<0.13 in all cases), and percentage of correct classification (>94% in all cases), which allows a reliable extrapolation prediction of antibacterial activity in any organic compound. The results obtained clearly reveal the high efficiency of combining molecular topology with LDA for the prediction of anti-MRSA activity.
The Microbiota and the Gut–Brain Axis in Controlling Food Intake and Energy Homeostasis
Authors: Romaní-Pérez Marina , Bullich-Vilarrubias Clara , López-Almela Inmaculada , Liébana-García Rebeca , Olivares Marta , Sanz Yolanda
Published: May 2021

Obesity currently represents a major societal and health challenge worldwide. Its prevalence has reached epidemic proportions and trends continue to rise, reflecting the need for more effective preventive measures. Hypothalamic circuits that control energy homeostasis in response to food intake are interesting targets for body-weight management, for example, through interventions that reinforce the gut-to-brain nutrient signalling, whose malfunction contributes to obesity. Gut microbiota–diet interactions might interfere in nutrient sensing and signalling from the gut to the brain, where the information is processed to control energy homeostasis. This gut microbiota–brain crosstalk is mediated by metabolites, mainly short chain fatty acids, secondary bile acids or amino acids-derived metabolites and subcellular bacterial components. These activate gut–endocrine and/or neural-mediated pathways or pass to systemic circulation and then reach the brain. Feeding time and dietary composition are the main drivers of the gut microbiota structure and function. Therefore, aberrant feeding patterns or unhealthy diets might alter gut microbiota–diet interactions and modify nutrient availability and/or microbial ligands transmitting information from the gut to the brain in response to food intake, thus impairing energy homeostasis. Herein, we update the scientific evidence supporting that gut microbiota is a source of novel dietary and non-dietary biological products that may beneficially regulate gut-to-brain communication and, thus, improve metabolic health. Additionally, we evaluate how the feeding time and dietary composition modulate the gut microbiota and, thereby, the intraluminal availability of these biological products with potential effects on energy homeostasis. The review also identifies knowledge gaps and the advances required to clinically apply microbiome-based strategies to improve the gut–brain axis function and, thus, combat obesity.
Structural Basis for the Diminished Ligand Binding and Catalytic Ability of Human Fetal-Specific CYP3A7
Authors: Sevrioukova Irina F.
Published: May 2021

Cytochrome P450 3A7 (CYP3A7) is a fetal/neonatal liver enzyme that participates in estriol synthesis, clearance of all-trans retinoic acid, and xenobiotic metabolism. Compared to the closely related major drug-metabolizing enzyme in adult liver, CYP3A4, the ligand binding and catalytic capacity of CYP3A7 are substantially reduced. To better understand the structural basis for these functional differences, the 2.15 Å crystal structure of CYP3A7 has been solved. Comparative analysis of CYP3A enzymes shows that decreased structural plasticity rather than the active site microenvironment defines the ligand binding ability of CYP3A7. In particular, a rotameric switch in the gatekeeping amino acid F304 triggers local and long-range rearrangements that transmit to the F-G fragment and alter its interactions with the I-E-D-helical core, resulting in a more rigid structure. Elongation of the β3-β4 strands, H-bond linkage in the substrate channel, and steric constraints in the C-terminal loop further increase the active site rigidity and limit conformational ensemble. Collectively, these structural distinctions lower protein plasticity and change the heme environment, which, in turn, could impede the spin-state transition essential for optimal reactivity and oxidation of substrates.
β-Hexachlorocyclohexane Drives Carcinogenesis in the Human Normal Bronchial Epithelium Cell Line BEAS-2B
Authors: Rubini Elisabetta , Minacori Marco , Paglia Giuliano , Altieri Fabio , Chichiarelli Silvia , Romaniello Donatella , Eufemi Margherita
Published: May 2021

Organochlorine pesticides constitute the majority of the total environmental pollutants, and a wide range of compounds have been found to be carcinogenic to humans. Among all, growing interest has been focused on β-hexachlorocyclohexane (β-HCH), virtually the most hazardous and, at the same time, the most poorly investigated member of the hexachlorocyclohexane family. Considering the multifaceted biochemical activities of β-HCH, already established in our previous studies, the aim of this work is to assess whether β-HCH could also trigger cellular malignant transformation toward cancer development. For this purpose, experiments were performed on the human normal bronchial epithelium cell line BEAS-2B exposed to 10 µM β-HCH. The obtained results strongly support the carcinogenic potential of β-HCH, which is achieved through both non-genotoxic (activation of oncogenic signaling pathways and proliferative activity) and indirect genotoxic (ROS production and DNA damage) mechanisms that significantly affect cellular macroscopic characteristics and functions such as cell morphology, cell cycle profile, and apoptosis. Taking all these elements into account, the presented study provides important elements to further characterize β-HCH, which appears to be a full-fledged carcinogenic agent.
Antimicrobial Effect of Chitosan Films on Food Spoilage Bacteria
Authors: Wrońska Natalia , Katir Nadia , Miłowska Katarzyna , Hammi Nisrine , Nowak Marta , Kędzierska Marta , Anouar Aicha , Zawadzka Katarzyna , Bryszewska Maria , El Kadib Abdelkrim , Lisowska Katarzyna
Published: May 2021

Synthetic materials commonly used in the packaging industry generate a considerable amount of waste each year. Chitosan is a promising feedstock for the production of functional biomaterials. From a biological point of view, chitosan is very attractive for food packaging. The purposes of this study were to evaluate the antibacterial activity of a set of chitosan-metal oxide films and different chitosan-modified graphene (oxide) films against two foodborne pathogens: Campylobacter jejuni ATCC 33560 and Listeria monocytogenes 19115. Moreover, we wanted to check whether the incorporation of antimicrobial constituents such as TiO2, ZnO, Fe2O3, Ag, and graphene oxide (GO) into the polymer matrices can improve the antibacterial properties of these nanocomposite films. Finally, this research helps elucidate the interactions of these materials with eukaryotic cells. All chitosan-metal oxide films and chitosan-modified graphene (oxide) films displayed improved antibacterial (C. jejuni ATCC 33560 and L. monocytogenes 19115) properties compared to native chitosan films. The CS-ZnO films had excellent antibacterial activity towards L. monocytogenes (90% growth inhibition). Moreover, graphene-based chitosan films caused high inhibition of both tested strains. Chitosan films with graphene (GO, GOP, GOP-HMDS, rGO, GO-HMDS, rGOP), titanium dioxide (CS-TiO2 20:1a, CS-TiO2 20:1b, CS-TiO2 2:1, CS-TiO2 1:1a, CS-TiO2 1:1b) and zinc oxide (CS-ZnO 20:1a, CS-ZnO 20:1b) may be considered as a safe, non-cytotoxic packaging materials in the future.
DNA Damage Response in the Adaptive Arm of the Immune System: Implications for Autoimmunity
Authors: Manolakou Theodora , Verginis Panayotis , Boumpas Dimitrios T.
Published: May 2021

In complex environments, cells have developed molecular responses to confront threats against the genome and achieve the maintenance of genomic stability assuring the transfer of undamaged DNA to their progeny. DNA damage response (DDR) mechanisms may be activated upon genotoxic or environmental agents, such as cytotoxic drugs or ultraviolet (UV) light, and during physiological processes requiring DNA transactions, to restore DNA alterations that may cause cellular malfunction and affect viability. In addition to the DDR, multicellular organisms have evolved specialized immune cells to respond and defend against infections. Both adaptive and innate immune cells are subjected to DDR processes, either as a prerequisite to the immune response, or as a result of random endogenous and exogenous insults. Aberrant DDR activities have been extensively studied in the immune cells of the innate arm, but not in adaptive immune cells. Here, we discuss how the aberrant DDR may lead to autoimmunity, with emphasis on the adaptive immune cells and the potential of therapeutic targeting.
Extracellular Vesicles: Potential Mediators of Psychosocial Stress Contribution to Osteoporosis?
Authors: He Yangyang , Wuertz-Kozak Karin , Kuehl Linn K. , Wippert Pia-Maria
Published: May 2021

Osteoporosis is characterized by low bone mass and damage to the bone tissue’s microarchitecture, leading to increased fracture risk. Several studies have provided evidence for associations between psychosocial stress and osteoporosis through various pathways, including the hypothalamic-pituitary-adrenocortical axis, the sympathetic nervous system, and other endocrine factors. As psychosocial stress provokes oxidative cellular stress with consequences for mitochondrial function and cell signaling (e.g., gene expression, inflammation), it is of interest whether extracellular vesicles (EVs) may be a relevant biomarker in this context or act by transporting substances. EVs are intercellular communicators, transfer substances encapsulated in them, modify the phenotype and function of target cells, mediate cell-cell communication, and, therefore, have critical applications in disease progression and clinical diagnosis and therapy. This review summarizes the characteristics of EVs, their role in stress and osteoporosis, and their benefit as biological markers. We demonstrate that EVs are potential mediators of psychosocial stress and osteoporosis and may be beneficial in innovative research settings.
One Week of CDAHFD Induces Steatohepatitis and Mitochondrial Dysfunction with Oxidative Stress in Liver
Authors: Sugasawa Takehito , Ono Seiko , Yonamine Masato , Fujita Shin-ichiro , Matsumoto Yuki , Aoki Kai , Nakano Takuro , Tamai Shinsuke , Yoshida Yasuko , Kawakami Yasushi , Takekoshi Kazuhiro
Published: May 2021

The prevalence of nonalcoholic fatty liver disease (NAFLD) has been rapidly increasing worldwide. A choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) has been used to create a mouse model of nonalcoholic steatohepatitis (NASH). There are some reports on the effects on mice of being fed a CDAHFD for long periods of 1 to 3 months. However, the effect of this diet over a short period is unknown. Therefore, we examined the effect of 1-week CDAHFD feeding on the mouse liver. Feeding a CDAHFD diet for only 1-week induced lipid droplet deposition in the liver with increasing activity of liver-derived enzymes in the plasma. On the other hand, it did not induce fibrosis or cirrhosis. Additionally, it was demonstrated that CDAHFD significantly impaired mitochondrial respiration with severe oxidative stress to the liver, which is associated with a decreasing mitochondrial DNA copy number and complex proteins. In the gene expression analysis of the liver, inflammatory and oxidative stress markers were significantly increased by CDAHFD. These results demonstrated that 1 week of feeding CDAHFD to mice induces steatohepatitis with mitochondrial dysfunction and severe oxidative stress, without fibrosis, which can partially mimic the early stage of NASH in humans.
The Potential of Asiatic Acid in the Reversion of Cyclophosphamide-Induced Hemorrhagic Cystitis in Rats
Authors: Wróbel Andrzej , Zapała Łukasz , Kluz Tomasz , Rogowski Artur , Misiek Marcin , Juszczak Kajetan , Sieńko Jacek , Gold Daniela , Stangel-Wójcikiewicz Klaudia , Poleszak Ewa , Radziszewski Piotr
Published: May 2021

The purpose of this study was to determine if asiatic acid may act efficiently in the model of cyclophosphamide (CYP)-induced cystitis in rats. We performed experiments after administration of CYP (single dose 200 mg/kg, intraperitoneally), asiatic acid (30 mg/kg/day for 14 consecutive days, by oral gavage), or CYP plus asiatic acid, during which conscious cystometry, measurements of urothelium thickness and bladder edema, as well as selected biomarkers analyses were conducted. In rats that received asiatic acid together with CYP, a drop in bladder basal pressure, detrusor overactivity index, non-voiding contraction amplitude, non-voiding contraction frequency, and the area under the pressure curve were observed, when compared to the CYP group. Furthermore, a significant increase in threshold pressure, voided volume, intercontraction interval, bladder compliance, and volume threshold to elicit NVC were found in that group accordingly. Administration of the asiatic acid successfully restored concentrations of biomarkers both in bladder urothelium (BDNF, CGRP, OCT-3, IL-1β, IL-6, NGF, nitrotyrosine, malondialdehyde, TNF-α, SV2A, SNAP23, SNAP25, PAC-1, ORM1, occludin, IGFBP-3, HB-EGF, T–H protein, Z01, and HPX) and detrusor muscle (Rho kinase and VAChT) in CYP-treated rats. Finally, asiatic acid significantly decreased urothelium thickness and bladder oedema. Asiatic acid proved to be a potent and effective drug in the rat model of CYP-induced cystitis.
The BAG2 and BAG6 Genes Are Involved in Multiple Abiotic Stress Tolerances in Arabidopsis Thaliana
Authors: Arif Muhammad , Li Zitong , Luo Qiong , Li Luhua , Shen Yuequan , Men Shuzhen
Published: May 2021

The BAG proteins are a family of multi-functional co-chaperones. In plants, BAG proteins were found to play roles both in abiotic and biotic stress tolerance. However, the function of Arabidopsis BAG2 remains largely unknown, whereas BAG6 is required for plants’ defense to pathogens, although it remains unknown whether BAG6 is involved in plants’ tolerance to abiotic stresses. Here, we show that both BAG2 and BAG6 are expressed in various tissues and are upregulated by salt, mannitol, and heat treatments and by stress-related hormones including ABA, ethylene, and SA. Germination of bag2, bag6 and bag2 bag6 seeds is less sensitive to ABA compared to the wild type (WT), whereas BAG2 and BAG6 overexpression lines are hypersensitive to ABA. bag2, bag6, and bag2 bag6 plants show higher survival rates than WT in drought treatment but display lower survival rates in heat-stress treatment. Consistently, these mutants showed differential expression of several stress- and ABA-related genes such as RD29A, RD29B, NCED3 and ABI4 compared to the WT. Furthermore, these mutants exhibit lower levels of ROS after drought and ABA treatment but higher ROS accumulation after heat treatment than the WT. These results suggest that BAG2 and BAG6 are negatively involved in drought stress but play a positive role in heat stress in Arabidopsis.
Dysregulation of the Acrosome Formation Network by 8-oxoguanine (8-oxoG) in Infertile Sperm: A Case Report with Advanced Techniques
Authors: Kim Sung Woo , Kim Bongki , Mok Jongsoo , Kim Eun Seo , Park Joonghoon
Published: May 2021

8-Hydroxyguanine (8-oxoG) is the most common oxidative DNA lesion and unrepaired 8-oxoG is associated with DNA fragmentation in sperm. However, the molecular effects of 8-oxoG on spermatogenesis are not entirely understood. Here, we identified one infertile bull (C14) due to asthenoteratozoospermia. We compared the global concentration of 8-oxoG by reverse-phase liquid chromatography/mass spectrometry (RP-LC/MS), the genomic distribution of 8-oxoG by next-generation sequencing (OG-seq), and the expression of sperm proteins by 2-dimensional polyacrylamide gel electrophoresis followed by peptide mass fingerprinting (2D-PAGE/PMF) in the sperm of C14 with those of a fertile bull (C13). We found that the average levels of 8-oxoG in C13 and C14 sperm were 0.027% and 0.044% of the total dG and it was significantly greater in infertile sperm DNA (p = 0.0028). Over 81% of the 8-oxoG loci were distributed around the transcription start site (TSS) and 165 genes harboring 8-oxoG were exclusive to infertile sperm. Functional enrichment and network analysis revealed that the Golgi apparatus was significantly enriched with the products from 8-oxoG genes of infertile sperm (q = 2.2 × 10−7). Proteomic analysis verified that acrosome-related proteins, including acrosin-binding protein (ACRBP), were downregulated in infertile sperm. These preliminary results suggest that 8-oxoG formation during spermatogenesis dysregulated the acrosome-related gene network, causing structural and functional defects of sperm and leading to infertility.
A Dual-Color Tyr-FISH Method for Visualizing Genes/Markers on Plant Chromosomes to Create Integrated Genetic and Cytogenetic Maps
Authors: Kudryavtseva Natalya , Ermolaev Aleksey , Karlov Gennady , Kirov Ilya , Shigyo Masayoshi , Sato Shusei , Khrustaleva Ludmila
Published: May 2021

In situ imaging of molecular markers on a physical chromosome is an indispensable tool for refining genetic maps and validation genome assembly at the chromosomal level. Despite the tremendous progress in genome sequencing, the plant genome assembly at the chromosome level remains a challenge. Recently developed optical and Hi-C mapping are aimed at assistance in genome assembly. For high confidence in the genome assembly at chromosome level, more independent approaches are required. The present study is aimed at refining an ultrasensitive Tyr-FISH technique and developing a reliable and simple method of in situ mapping of a short unique DNA sequences on plant chromosomes. We have carefully analyzed the critical steps of the Tyr-FISH to find out the reasons behind the flaws of this technique. The accurate visualization of markers/genes appeared to be significantly dependent on the means of chromosome slide preparation, probe design and labeling, and high stringency washing. Appropriate adjustment of these steps allowed us to detect a short DNA sequence of 1.6 Kb with a frequency of 51.6%. Based on our results, we developed a more reliable and simple protocol for dual-color Tyr-FISH visualization of unique short DNA sequences on plant chromosomes. This new protocol can allow for more accurate determination of the physical distance between markers and can be applied for faster integration of genetic and cytogenetic maps.
Diminazene Aceturate Stabilizes Atherosclerotic Plaque and Attenuates Hepatic Steatosis in apoE-Knockout Mice by Influencing Macrophages Polarization and Taurine Biosynthesis
Authors: Stachowicz Aneta , Wiśniewska Anna , Kuś Katarzyna , Białas Magdalena , Łomnicka Magdalena , Totoń-Żurańska Justyna , Kiepura Anna , Stachyra Kamila , Suski Maciej , Bujak-Giżycka Beata , Jawień Jacek , Olszanecki Rafał
Published: May 2021

Atherosclerosis and nonalcoholic fatty liver disease are leading causes of morbidity and mortality in the Western countries. The renin–angiotensin system (RAS) with its two main opposing effectors, i.e., angiotensin II (Ang II) and Ang-(1–7), is widely recognized as a major regulator of cardiovascular function and body metabolic processes. Angiotensin-converting enzyme 2 (ACE2) by breaking-down Ang II forms Ang-(1–7) and thus favors Ang-(1–7) actions. Therefore, the aim of our study was to comprehensively evaluate the influence of prolonged treatment with ACE2 activator, diminazene aceturate (DIZE) on the development of atherosclerotic lesions and hepatic steatosis in apoE−/− mice fed a high-fat diet (HFD). We have shown that DIZE stabilized atherosclerotic lesions and attenuated hepatic steatosis in apoE−/− mice fed an HFD. Such effects were associated with decreased total macrophages content and increased α-smooth muscle actin levels in atherosclerotic plaques. Moreover, DIZE changed polarization of macrophages towards increased amount of anti-inflammatory M2 macrophages in the atherosclerotic lesions. Interestingly, the anti-steatotic action of DIZE in the liver was related to the elevated levels of HDL in the plasma, decreased levels of triglycerides, and increased biosynthesis and concentration of taurine in the liver of apoE−/− mice. However, exact molecular mechanisms of both anti-atherosclerotic and anti-steatotic actions of DIZE require further investigations.
Tumor Extracellular Matrix Stiffness Promptly Modulates the Phenotype and Gene Expression of Infiltrating T Lymphocytes
Authors: Chirivì Maila , Maiullari Fabio , Milan Marika , Presutti Dario , Cordiglieri Chiara , Crosti Mariacristina , Sarnicola Maria Lucia , Soluri Andrea , Volpi Marina , Święszkowski Wojciech , Prati Daniele , Rizzi Marta , Costantini Marco , Seliktar Dror , Parisi Chiara , Bearzi Claudia , Rizzi Roberto
Published: May 2021

The immune system is a fine modulator of the tumor biology supporting or inhibiting its progression, growth, invasion and conveys the pharmacological treatment effect. Tumors, on their side, have developed escaping mechanisms from the immune system action ranging from the direct secretion of biochemical signals to an indirect reaction, in which the cellular actors of the tumor microenvironment (TME) collaborate to mechanically condition the extracellular matrix (ECM) making it inhospitable to immune cells. TME is composed of several cell lines besides cancer cells, including tumor-associated macrophages, cancer-associated fibroblasts, CD4+ and CD8+ lymphocytes, and innate immunity cells. These populations interface with each other to prepare a conservative response, capable of evading the defense mechanisms implemented by the host’s immune system. The presence or absence, in particular, of cytotoxic CD8+ cells in the vicinity of the main tumor mass, is able to predict, respectively, the success or failure of drug therapy. Among various mechanisms of immunescaping, in this study, we characterized the modulation of the phenotypic profile of CD4+ and CD8+ cells in resting and activated states, in response to the mechanical pressure exerted by a three-dimensional in vitro system, able to recapitulate the rheological and stiffness properties of the tumor ECM.
The Density of Group I mGlu5 Receptors Is Reduced along the Neuronal Surface of Hippocampal Cells in a Mouse Model of Alzheimer’s Disease
Authors: Martín-Belmonte Alejandro , Aguado Carolina , Alfaro-Ruiz Rocío , Albasanz José Luis , Martín Mairena , Moreno-Martínez Ana Esther , Fukazawa Yugo , Luján Rafael
Published: May 2021

Metabotropic glutamate receptor subtype 5 (mGlu5) is implicated in the pathophysiology of Alzheimer’s disease (AD). However, its alteration at the subcellular level in neurons is still unexplored. Here, we provide a quantitative description on the expression and localisation patterns of mGlu5 in the APP/PS1 model of AD at 12 months of age, combining immunoblots, histoblots and high-resolution immunoelectron microscopic approaches. Immunoblots revealed that the total amount of mGlu5 protein in the hippocampus, in addition to downstream molecules, i.e., Gq/11 and PLCβ1, was similar in both APP/PS1 mice and age-matched wild type mice. Histoblots revealed that mGlu5 expression in the brain and its laminar expression in the hippocampus was also unaltered. However, the ultrastructural techniques of SDS-FRL and pre-embedding immunogold demonstrated that the subcellular localisation of mGlu5 was significantly reduced along the neuronal surface of hippocampal principal cells, including CA1 pyramidal cells and DG granule cells, in APP/PS1 mice at 12 months of age. The decrease in the surface localisation of mGlu5 was accompanied by an increase in its frequency at intracellular sites in the two neuronal populations. Together, these data demonstrate, for the first time, a loss of mGlu5 at the plasma membrane and accumulation at intracellular sites in different principal cells of the hippocampus in APP/PS1 mice, suggesting an alteration of the excitability and synaptic transmission that could contribute to the cognitive dysfunctions in this AD animal model. Further studies are required to elucidate the specificity of mGlu5-associated molecules and downstream signalling pathways in the progression of the pathology.
CCNE1 Is a Putative Therapeutic Target for ARID1A-Mutated Ovarian Clear Cell Carcinoma
Authors: Kawahara Naoki , Yamada Yuki , Kobayashi Hiroshi
Published: May 2021

Background: Ovarian clear cell carcinoma (OCCC) is resistant to platinum chemotherapy and is characterized by poor prognosis. Today, the use of poly (ADP-ribose) polymerase (PARP) inhibitor, which is based on synthetic lethality strategy and characterized by cancer selectivity, is widely used for new types of molecular-targeted treatment of relapsed platinum-sensitive ovarian cancer. However, it is less effective against OCCC. Methods: We conducted siRNA screening to identify synthetic lethal candidates for the ARID1A mutation; as a result, we identified Cyclin-E1 (CCNE1) as a potential target that affects cell viability. To further clarify the effects of CCNE1, human OCCC cell lines, namely TOV-21G and KOC7c (ARID1A mutant lines), and RMG-I and ES2 (ARID1A wild type lines) were transfected with siRNA targeting CCNE1 or a control vector. Results: Loss of CCNE1 reduced proliferation of the TOV-21G and KOC7c cells but not of the RMG-I and ES2 cells. Furthermore, in vivo interference of CCNE1 effectively inhibited tumor cell proliferation in a xenograft mouse model. Conclusion: This study showed for the first time that CCNE1 is a synthetic lethal target gene to ARID1A-mutated OCCC. Targeting this gene may represent a putative, novel, anticancer strategy in OCCC treatment.
The Odd Faces of Oligomers: The Case of TRAF2-C, A Trimeric C-Terminal Domain of TNF Receptor-Associated Factor
Authors: Di Venere Almerinda , Nicolai Eleonora , Minicozzi Velia , Caccuri Anna Maria , Di Paola Luisa , Mei Giampiero
Published: May 2021

TNF Receptor Associated Factor 2 (TRAF2) is a trimeric protein that belongs to the TNF receptor associated factor family (TRAFs). The TRAF2 oligomeric state is crucial for receptor binding and for its interaction with other proteins involved in the TNFR signaling. The monomer-trimer equilibrium of a C- terminal domain truncated form of TRAF2 (TRAF2-C), plays also a relevant role in binding the membrane, causing inward vesiculation. In this study, we have investigated the conformational dynamics of TRAF2-C through circular dichroism, fluorescence, and dynamic light scattering, performing temperature-dependent measurements. The data indicate that the protein retains its oligomeric state and most of its secondary structure, while displaying a significative increase in the heterogeneity of the tyrosines signal, increasing the temperature from ≈15 to ≈35 °C. The peculiar crowding of tyrosine residues (12 out of 18) at the three subunit interfaces and the strong dependence on the trimer concentration indicate that such conformational changes mainly involve the contact areas between each pair of monomers, affecting the oligomeric state. Molecular dynamic simulations in this temperature range suggest that the interfaces heterogeneity is an intrinsic property of the trimer that arises from the continuous, asymmetric approaching and distancing of its subunits. Such dynamics affect the results of molecular docking on the external protein surface using receptor peptides, indicating that the TRAF2-receptor interaction in the solution might not involve three subunits at the same time, as suggested by the static analysis obtainable from the crystal structure. These findings shed new light on the role that the TRAF2 oligomeric state might have in regulating the protein binding activity in vivo.
Development of Real-Time and Conventional PCR Assays for Identifying a Newly Named Species of Root-Lesion Nematode (Pratylenchus dakotaensis) on Soybean
Authors: Chowdhury Intiaz Amin , Yan Guiping
Published: May 2021

A rapid and accurate PCR-based method was developed in this study for detecting and identifying a new species of root-lesion nematode (Pratylenchus dakotaensis) recently discovered in a soybean field in North Dakota, USA. Species-specific primers, targeting the internal transcribed spacer region of ribosomal DNA, were designed to be used in both conventional and quantitative real-time PCR assays for identification of P. dakotaensis. The specificity of the primers was evaluated in silico analysis and laboratory PCR experiments. Results showed that only P. dakotaensis DNA was exclusively amplified in conventional and real-time PCR assays but none of the DNA from other control species were amplified. Detection sensitivity analysis revealed that the conventional PCR was able to detect an equivalent to 1/8 of the DNA of a single nematode whereas real-time PCR detected an equivalent to 1/32 of the DNA of a single nematode. According to the generated standard curve the amplification efficiency of the primers in real-time PCR was 94% with a R2 value of 0.95 between quantification cycle number and log number of P. dakotaensis. To validate the assays to distinguish P. dakotaensis from other Pratylenchus spp. commonly detected in North Dakota soybean fields, 20 soil samples collected from seven counties were tested. The PCR assays amplified the DNA of P. dakotaensis and discriminated it from other Pratylenchus spp. present in North Dakota soybean fields. This is the first report of a species-specific and rapid PCR detection method suitable for use in diagnostic and research laboratories for the detection of P. dakotaensis.
Oral Microbiome in Relation to Periodontitis Severity and Systemic Inflammation
Authors: Plachokova Adelina S. , Andreu-Sánchez Sergio , Noz Marlies P. , Fu Jingyuan , Riksen Niels P.
Published: May 2021

Systemic inflammation induced by periodontitis is suggested to be the link between periodontitis and cardiovascular disease. The aim of this work was to explore the oral microbiome in periodontitis in relation to disease severity and systemic inflammation. The saliva and subgingival microbiome from periodontal pocket samples of patients with severe (n = 12) and mild periodontitis (n = 13) were analyzed using metagenomic shotgun sequencing. The taxa and pathways abundances were quantified. The diversity was assessed and the abundances to phenotype associations were performed using ANCOM and linear regression. A panel of inflammatory markers was measured in blood and was associated with taxa abundance. The microbial diversity and species richness did not differ between severe and mild periodontitis in either saliva or periodontal pockets. However, there were significant differences in the microbial composition between severe and mild periodontitis in the subgingival microbiome (i.e., pocket samples) and, in a lower grade, in saliva, and this is positively associated with systemic inflammatory markers. The “red complex” and “cluster B” abundances in periodontal pockets were strongly associated with inflammatory markers interleukin-6 and the white blood cell count. Our data suggest that systemic inflammation in severe periodontitis may be driven by the oral microbiome and may support the indirect (inflammatory) mechanism for the association between periodontitis and cardiovascular disease.
Magnesium Isoglycyrrhizinate Reduces Hepatic Lipotoxicity through Regulating Metabolic Abnormalities
Authors: Lu Li , Hao Kun , Hong Yu , Liu Jie , Zhu Jinwei , Jiang Wenjiao , Zhu Zheying , Wang Guangji , Peng Ying
Published: May 2021

The excessive accumulation of lipids in hepatocytes induces a type of cytotoxicity called hepatic lipotoxicity, which is a fundamental contributor to liver metabolic diseases (such as NAFLD). Magnesium isoglycyrrhizinate (MGIG), a magnesium salt of the stereoisomer of natural glycyrrhizic acid, is widely used as a safe and effective liver protectant. However, the mechanism by which MGIG protects against NAFLD remains unknown. Based on the significant correlation between NAFLD and the reprogramming of liver metabolism, we aimed to explore the beneficial effects of MGIG from a metabolic viewpoint in this paper. We treated HepaRG cells with palmitic acid (PA, a saturated fatty acid of C16:0) to induce lipotoxicity and then evaluated the antagonistic effect of MGIG on lipotoxicity by investigating the cell survival rate, DNA proliferation rate, organelle damage, and endoplasmic reticulum stress (ERS). Metabolomics, lipidomics, and isotope tracing were used to investigate changes in the metabolite profile, lipid profile, and lipid flux in HepaRG cells under different intervention conditions. The results showed that MGIG can indeed protect hepatocytes against PA-induced cytotoxicity and ERS. In response to the metabolic abnormality of lipotoxicity, MGIG curtailed the metabolic activation of lipids induced by PA. The content of total lipids and saturated lipids containing C16:0 chains increased significantly after PA stimulation and then decreased significantly or even returned to normal levels after MGIG intervention. Lipidomic data show that glycerides and glycerophospholipids were the two most affected lipids. For excessive lipid accumulation in hepatocytes, MGIG can downregulate the expression of the metabolic enzymes (GPATs and DAGTs) involved in triglyceride biosynthesis. In conclusion, MGIG has a positive regulatory effect on the metabolic disorders that occur in hepatocytes under lipotoxicity, and the main mechanisms of this effect are in lipid metabolism, including reducing the total lipid content, reducing lipid saturation, inhibiting glyceride and glycerophospholipid metabolism, and downregulating the expression of metabolic enzymes in lipid synthesis.
Deficits in Prenatal Serine Biosynthesis Underlie the Mitochondrial Dysfunction Associated with the Autism-Linked FMR1 Gene
Authors: Nolin Sarah L. , Napoli Eleonora , Flores Amanda , Hagerman Randi J. , Giulivi Cecilia
Published: May 2021

Fifty-five to two hundred CGG repeats (called a premutation, or PM) in the 5′-UTR of the FMR1 gene are generally unstable, often expanding to a full mutation (>200) in one generation through maternal inheritance, leading to fragile X syndrome, a condition associated with autism and other intellectual disabilities. To uncover the early mechanisms of pathogenesis, we performed metabolomics and proteomics on amniotic fluids from PM carriers, pregnant with male fetuses, who had undergone amniocentesis for fragile X prenatal diagnosis. The prenatal metabolic footprint identified mitochondrial deficits, which were further validated by using internal and external cohorts. Deficits in the anaplerosis of the Krebs cycle were noted at the level of serine biosynthesis, which was confirmed by rescuing the mitochondrial dysfunction in the carriers’ umbilical cord fibroblasts using alpha-ketoglutarate precursors. Maternal administration of serine and its precursors has the potential to decrease the risk of developing energy shortages associated with mitochondrial dysfunction and linked comorbidities.
Glucose Metabolic Dysfunction in Neurodegenerative Diseases—New Mechanistic Insights and the Potential of Hypoxia as a Prospective Therapy Targeting Metabolic Reprogramming
Authors: Han Rongrong , Liang Jing , Zhou Bing
Published: May 2021

Glucose is the main circulating energy substrate for the adult brain. Owing to the high energy demand of nerve cells, glucose is actively oxidized to produce ATP and has a synergistic effect with mitochondria in metabolic pathways. The dysfunction of glucose metabolism inevitably disturbs the normal functioning of neurons, which is widely observed in neurodegenerative disease. Understanding the mechanisms of metabolic adaptation during disease progression has become a major focus of research, and interventions in these processes may relieve the neurons from degenerative stress. In this review, we highlight evidence of mitochondrial dysfunction, decreased glucose uptake, and diminished glucose metabolism in different neurodegeneration models such as Alzheimer’s disease (AD), Parkinson’s disease (PD), amyotrophic lateral sclerosis (ALS), and Huntington’s disease (HD). We also discuss how hypoxia, a metabolic reprogramming strategy linked to glucose metabolism in tumor cells and normal brain cells, and summarize the evidence for hypoxia as a putative therapy for general neurodegenerative disease.
The Evolution of Rag Gene Enhancers and Transcription Factor E and Id Proteins in the Adaptive Immune System
Authors: Yoshikawa Genki , Miyazaki Kazuko , Ogata Hiroyuki , Miyazaki Masaki
Published: May 2021

Adaptive immunity relies on the V(D)J DNA recombination of immunoglobulin (Ig) and T cell receptor (TCR) genes, which enables the recognition of highly diverse antigens and the elicitation of antigen-specific immune responses. This process is mediated by recombination-activating gene (Rag) 1 and Rag2 (Rag1/2), whose expression is strictly controlled in a cell type-specific manner; the expression of Rag1/2 genes represents a hallmark of lymphoid lineage commitment. Although Rag genes are known to be evolutionally conserved among jawed vertebrates, how Rag genes are regulated by lineage-specific transcription factors (TFs) and how their regulatory system evolved among vertebrates have not been fully elucidated. Here, we reviewed the current body of knowledge concerning the cis-regulatory elements (CREs) of Rag genes and the evolution of the basic helix-loop-helix TF E protein regulating Rag gene CREs, as well as the evolution of the antagonist of this protein, the Id protein. This may help to understand how the adaptive immune system develops along with the evolution of responsible TFs and enhancers.
Dual Mechanisms of Metabolism and Gene Expression of the CCRF-CEM Leukemia Cells under Glucocorticoid Treatment
Authors: Lambrou George , Karakonstantakis Theodoros , Vlahopoulos Spiros , Zaravinos Apostolos
Published: May 2021

Background: Glucocorticoids play an essential part in anti-leukemic therapies, but resistance is a crucial event for the prognosis of the disease. Glucocorticoids influence the metabolic properties of leukemic cells. The inherent plasticity of clinically evolving cancer cells justifies the characterization of drug-induced early oncogenic pathways, which represent a likely source of detrimental secondary effects. Aim: The present work aims to investigate the effect of glucocorticoids in metabolic pathways in the CCRF-CEM leukemic cells. Metabolic factors and gene expression profiles were examined in order to unravel the possible mechanisms of the CCRF-CEM leukemic cell growth dynamics. Methods: CCRF-CEM cells were used as a model. Cells were treated with prednisolone with concentrations 0–700 μM. Cell culture supernatants were used for glucose, lactic acid, LDH, Na+, K+ and Ca++ measurements. Cytotoxicity was determined with flow cytometry. Microarray analysis was performed using two different chips of 1.2 k and 4.8 k genes. Gene Ontology enrichment analysis was applied to find metabolism- and GC-related genes. Results: Higher prednisolone concentrations inhibited glucose uptake, without exhibiting any cytotoxic effects. Glucose consumption did not correlate with the total cell population, or the viable population, indicating that growth is not directly proportional to glucose consumption. Neither of the subpopulations, i.e., viable, necrotic, or apoptotic cells, contributed to this. Conclusions: Different types of leukemic cells seem to exhibit different patterns of glucose metabolism. Both resistant and sensitive CCRF-CEM cells followed the aerobic pathway of glycolysis. There is probably a rapid change in membrane permeability, causing a general shutdown towards everything that is outside the cell. This could in part also explain the observed resistance. Glucocorticoids do not enter the cell passively anymore and therefore no effects are observed. Based on our observations, ion concentrations are measurable factors both in vitro and in vivo, which makes them possible markers of glucocorticoid cytotoxic action.