bluuebunny/crossref_metadata_2025 huggingface dataset:
Therapeutic Strategies for Disseminated Intravascular Coagulation Associated with Aortic Aneurysm
Authors: Yamada Shinya , Asakura Hidesaku
Published: January 2022

Aortic aneurysms are sometimes associated with enhanced-fibrinolytic-type disseminated intravascular coagulation (DIC). In enhanced-fibrinolytic-type DIC, both coagulation and fibrinolysis are markedly activated. Typical cases show decreased platelet counts and fibrinogen levels, increased concentrations of fibrin/fibrinogen degradation products (FDP) and D-dimer, and increased FDP/D-dimer ratios. Thrombin-antithrombin complex or prothrombin fragment 1 + 2, as markers of coagulation activation, and plasmin-α2 plasmin inhibitor complex, a marker of fibrinolytic activation, are all markedly increased. Prolongation of prothrombin time (PT) is not so obvious, and the activated partial thromboplastin time (APTT) is rather shortened in some cases. As a result, DIC can be neither diagnosed nor excluded based on PT and APTT alone. Many of the factors involved in coagulation and fibrinolysis activation are serine proteases. Treatment of enhanced-fibrinolytic-type DIC requires consideration of how to control the function of these serine proteases. The cornerstone of DIC treatment is treatment of the underlying pathology. However, in some cases surgery is either not possible or exacerbates the DIC associated with aortic aneurysm. In such cases, pharmacotherapy becomes even more important. Unfractionated heparin, other heparins, synthetic protease inhibitors, recombinant thrombomodulin, and direct oral anticoagulants (DOACs) are agents that inhibit serine proteases, and all are effective against DIC. Inhibition of activated coagulation factors by anticoagulants is key to the treatment of DIC. Among them, DOACs can be taken orally and is useful for outpatient treatment. Combination therapy of heparin and nafamostat allows fine-adjustment of anticoagulant and antifibrinolytic effects. While warfarin is an anticoagulant, this agent is ineffective in the treatment of DIC because it inhibits the production of coagulation factors as substrates without inhibiting activated coagulation factors. In addition, monotherapy using tranexamic acid in cases of enhanced-fibrinolytic-type DIC may induce fatal thrombosis. If tranexamic acid is needed for DIC, combination with anticoagulant therapy is of critical importance.
Mitochondrial Processing Peptidases—Structure, Function and the Role in Human Diseases
Authors: Kunová Nina , Havalová Henrieta , Ondrovičová Gabriela , Stojkovičová Barbora , Bauer Jacob A. , Bauerová-Hlinková Vladena , Pevala Vladimir , Kutejová Eva
Published: January 2022

Mitochondrial proteins are encoded by both nuclear and mitochondrial DNA. While some of the essential subunits of the oxidative phosphorylation (OXPHOS) complexes responsible for cellular ATP production are synthesized directly in the mitochondria, most mitochondrial proteins are first translated in the cytosol and then imported into the organelle using a sophisticated transport system. These proteins are directed mainly by targeting presequences at their N-termini. These presequences need to be cleaved to allow the proper folding and assembly of the pre-proteins into functional protein complexes. In the mitochondria, the presequences are removed by several processing peptidases, including the mitochondrial processing peptidase (MPP), the inner membrane processing peptidase (IMP), the inter-membrane processing peptidase (MIP), and the mitochondrial rhomboid protease (Pcp1/PARL). Their proper functioning is essential for mitochondrial homeostasis as the disruption of any of them is lethal in yeast and severely impacts the lifespan and survival in humans. In this review, we focus on characterizing the structure, function, and substrate specificities of mitochondrial processing peptidases, as well as the connection of their malfunctions to severe human diseases.
Genome-Wide Survey Indicates Diverse Physiological Roles of Dendrobium officinale Calcium-Dependent Protein Kinase Genes
Authors: Yang Xingyu , Chen Zhiyu , Yin Xin , Wang Yuhua , Yang Yunqiang , Yang Yongping
Published: January 2022

Calcium-dependent protein kinases (CDPKs) are crucial calcium ions (Ca2+) sensors in plants with important roles in signal transduction, plant growth, development, and stress responses. Here, we identified 24 genes encoding CDPKs in Dendrobium officinale using genome-wide analysis. The phylogenetic analysis revealed that these genes formed four groups, with similar structures in the same group. The gene expression patterns following hormone treatments and yeast two-hybrid of homologous CDPK gene pairs with Rbohs showed differences, indicating functional divergence between homologous genes. In addition, the rapid accumulation of hydrogen peroxide (H2O2) and stomatal closure was observed in response to salicylic acid (SA)/jasmonic acid (JA) stress. Our data showed that CDPK9-2 and CDPK20-4 interacted with Rboh D and Rboh H, respectively, and were implicated in the generation of H2O2 and regulation of the stomatal aperture in response to salicylic acid/jasmonic acid treatment. We believe these results can provide a foundation for the functional divergence of homologous genes in D. officinale.
Aberrant Methylation of 20 miRNA Genes Specifically Involved in Various Steps of Ovarian Carcinoma Spread: From Primary Tumors to Peritoneal Macroscopic Metastases
Authors: Loginov Vitaly I. , Pronina Irina V. , Filippova Elena A. , Burdennyy Alexey M. , Lukina Svetlana S. , Kazubskaya Tatiana P. , Uroshlev Leonid A. , Fridman Marina V. , Brovkina Olga I. , Apanovich Natalya V. , Karpukhin Alexander V. , Stilidi Ivan S. , Kushlinskii Nikolay E. , Dmitriev Alexey A. , Braga Eleonora A.
Published: January 2022

Our work aimed to differentiate 20 aberrantly methylated miRNA genes that participate at different stages of development and metastasis of ovarian carcinoma (OvCa) using methylation-specific qPCR in a representative set of clinical samples: 102 primary tumors without and with metastases (to lymph nodes, peritoneum, or distant organs) and 30 peritoneal macroscopic metastases (PMM). Thirteen miRNA genes (MIR107, MIR124-2, MIR124-3, MIR125B-1, MIR127, MIR129-2, MIR130B, MIR132, MIR193A, MIR339, MIR34B/C, MIR9-1, and MIR9-3) were hypermethylated already at the early stages of OvCa, while hypermethylation of MIR1258, MIR137, MIR203A, and MIR375 was pronounced in metastatic tumors, and MIR148A showed high methylation levels specifically in PMM. We confirmed the significant relationship between methylation and expression levels for 11 out of 12 miRNAs analyzed by qRT-PCR. Moreover, expression levels of six miRNAs were significantly decreased in metastatic tumors in comparison with nonmetastatic ones, and downregulation of miR-203a-3p was the most significant. We revealed an inverse relationship between expression levels of miR-203a-3p and those of ZEB1 and ZEB2 genes, which are EMT drivers. We also identified three miRNA genes (MIR148A, MIR9-1, and MIR193A) that likely regulate EMT–MET reversion in the colonization of PMM. According to the Kaplan–Meier analysis, hypermethylation of several examined miRNA genes was associated with poorer overall survival of OvCa patients, and high methylation levels of MIR130B and MIR9-1 were related to the greatest relative risk of death.
SEC23B Loss-of-Function Suppresses Hepcidin Expression by Impairing Glycosylation Pathway in Human Hepatic Cells
Authors: Rosato Barbara Eleni , Marra Roberta , D’Onofrio Vanessa , Del Giudice Federica , Della Monica Simone , Iolascon Achille , Andolfo Immacolata , Russo Roberta
Published: January 2022

Biallelic pathogenic variants in the SEC23B gene cause congenital dyserythropoietic anemia type II (CDA II), a rare hereditary disorder hallmarked by ineffective erythropoiesis, hemolysis, erythroblast morphological abnormalities, and hypo-glycosylation of some red blood cell membrane proteins. Abnormalities in SEC23B, which encodes the homonymous cytoplasmic COPII (coat protein complex II) component, disturb the endoplasmic reticulum to Golgi trafficking and affect different glycosylation pathways. The most harmful complication of CDA II is the severe iron overload. Within our case series (28 CDA II patients), approximately 36% of them exhibit severe iron overload despite mild degree of anemia and slightly increased levels of ERFE (the only erythroid regulator of hepcidin suppression). Thus, we hypothesized a direct role of SEC23B loss-of-function in the pathomechanism of hepatic iron overload. We established a hepatic cell line, HuH7, stably silenced for SEC23B. In silenced cells, we observed significant alterations of the iron status, due to both the alteration in BMP/SMADs pathway effectors and a reduced capability to sense BMP6 stimulus. We demonstrated that the loss-of-function of SEC23B is responsible of the impairment in glycosylation of the membrane proteins involved in the activation of the BMP/SMADs pathway with subsequent hepcidin suppression. Most of these data were confirmed in another hepatic cell line, HepG2, stably silenced for SEC23B. Our findings suggested that the pathogenic mechanism of iron overload in CDA II is associated to both ineffective erythropoiesis and to a specific involvement of SEC23B pathogenic variants at hepatic level. Finally, we demonstrated the ability of SEC23B paralog, i.e., SEC23A, to rescue the hepcidin suppression, highlighting the functional overlap between the two SEC23 paralogs in human hepatic cells.
Plant Cytogenetics in the Micronuclei Investigation—The Past, Current Status, and Perspectives
Authors: Kwasniewska Jolanta , Bara Adrianna Wiktoria
Published: January 2022

Cytogenetic approaches play an essential role as a quick evaluation of the first genetic effects after mutagenic treatment. Although labor-intensive and time-consuming, they are essential for the analyses of cytotoxic and genotoxic effects in mutagenesis and environmental monitoring. Over the years, conventional cytogenetic analyses were a part of routine laboratory testing in plant genotoxicity. Among the methods that are used to study genotoxicity in plants, the micronucleus test particularly represents a significant force. Currently, cytogenetic techniques go beyond the simple detection of chromosome aberrations. The intensive development of molecular biology and the significantly improved microscopic visualization and evaluation methods constituted significant support to traditional cytogenetics. Over the past years, distinct approaches have allowed an understanding the mechanisms of formation, structure, and genetic activity of the micronuclei. Although there are many studies on this topic in humans and animals, knowledge in plants is significantly limited. This article provides a comprehensive overview of the current knowledge on micronuclei characteristics in plants. We pay particular attention to how the recent contemporary achievements have influenced the understanding of micronuclei in plant cells. Together with the current progress, we present the latest applications of the micronucleus test in mutagenesis and assess the state of the environment.
Identification of New KRAS G12D Inhibitors through Computer-Aided Drug Discovery Methods
Authors: Kulkarni Apoorva M. , Kumar Vikas , Parate Shraddha , Lee Gihwan , Yoon Sanghwa , Lee Keun Woo
Published: January 2022

Owing to several mutations, the oncogene Kirsten rat sarcoma 2 viral oncogene homolog (KRAS) is activated in the majority of cancers, and targeting it has been pharmacologically challenging. In this study, using an in silico approach comprised of pharmacophore modeling, molecular docking, and molecular dynamics simulations, potential KRAS G12D inhibitors were investigated. A ligand-based common feature pharmacophore model was generated to identify the framework necessary for effective KRAS inhibition. The chemical features in the selected pharmacophore model comprised two hydrogen bond donors, one hydrogen bond acceptor, two aromatic rings and one hydrophobic feature. This model was used for screening in excess of 214,000 compounds from InterBioScreen (IBS) and ZINC databases. Eighteen compounds from the IBS and ten from the ZINC database mapped onto the pharmacophore model and were subjected to molecular docking. Molecular docking results highlighted a higher affinity of four hit compounds towards KRAS G12D in comparison to the reference inhibitor, BI-2852. Sequential molecular dynamics (MD) simulation studies revealed all four hit compounds them possess higher KRAS G12D binding free energy and demonstrate stable polar interaction with key residues. Further, Principal Component Analysis (PCA) analysis of the hit compounds in complex with KRAS G12D also indicated stability. Overall, the research undertaken provides strong support for further in vitro testing of these newly identified KRAS G12D inhibitors, particularly Hit1 and Hit2.
Toward Xeno-Free Differentiation of Human Induced Pluripotent Stem Cell-Derived Small Intestinal Epithelial Cells
Authors: Saari Jaakko , Siddique Fatima , Korpela Sanna , Mäntylä Elina , Ihalainen Teemu O. , Kaukinen Katri , Aalto-Setälä Katriina , Lindfors Katri , Juuti-Uusitalo Kati
Published: January 2022

The small intestinal epithelium has an important role in nutrition, but also in drug absorption and metabolism. There are a few two-dimensional (2D) patient-derived induced pluripotent stem cell (iPSC)-based intestinal models enabling easy evaluation of transcellular transport. It is known that animal-derived components induce variation in the experimental outcomes. Therefore, we aimed to refine the differentiation protocol by using animal-free components. More specifically, we compared maturation of 2D-cultured iPCSs toward small intestinal epithelial cells when cultured either with or without serum, and either on Geltrex or on animal-free, recombinant laminin-based substrata. Differentiation status was characterized by qPCR, immunofluorescence imaging, and functionality assays. Our data suggest that differentiation toward definitive endoderm is more efficient without serum. Both collagen- and recombinant laminin-based coating supported differentiation of definitive endoderm, posterior definitive endoderm, and small intestinal epithelial cells from iPS-cells equally well. Small intestinal epithelial cells differentiated on recombinant laminin exhibited slightly more enterocyte specific cellular functionality than cells differentiated on Geltrex. Our data suggest that functional small intestinal epithelial cells can be generated from iPSCs in serum-free method on xeno-free substrata. This method is easily converted to an entirely xeno-free method.
Optical Study of Solvatochromic Isocyanoaminoanthracene Dyes and 1,5-Diaminoanthracene
Authors: Nagy Miklós , Fiser Béla , Szőri Milán , Vanyorek László , Viskolcz Béla
Published: January 2022

Isocyanoaminoarenes (ICAAr-s) are a novel and versatile group of solvatochromic fluorophores. Despite their versatile applicability, such as antifungals, cancer drugs and analytical probes, they still represent a mostly unchartered territory among intramolecular charge-transfer (ICT) dyes. The current paper describes the preparation and detailed optical study of novel 1-isocyano-5-aminoanthrace (ICAA) and its N-methylated derivatives along with the starting 1,5-diaminoanthracene. The conversion of one of the amino groups of the diamine into an isocyano group significantly increased the polar character of the dyes, which resulted in a significant 50–70 nm (2077–2609 cm−1) redshift of the emission maximum and a broadened solvatochromic range. The fluorescence quantum yield of ICAAs is strongly influenced by the polarity of the solvent. The starting anthracene-diamine is highly fluorescent in every solvent (√f = 12–53%), while the isocyano derivatives are practically nonfluorescent in solvents more polar than dioxane. This phenomenon implies the potential application of ICAAs to probe the polarity of the medium and is favorable in practical applications, such as cell-staining, resulting in a reduced background fluorescence. The ICT character of the emission states of ICAAs are in good agreement with the computational findings presented in TD-DFT calculations and molecular electrostatic potential (MESP) isosurfaces.
Changing Temperature Conditions during Somatic Embryo Maturation Result in Pinus pinaster Plants with Altered Response to Heat Stress
Authors: Sales Ester , Cañizares Eva , Pereira Catia , Pérez-Oliver María Amparo , Nebauer Sergio G. , Pavlović Iva , Novák Ondřej , Segura Juan , Arrillaga Isabel
Published: January 2022

Under the global warming scenario, obtaining plant material with improved tolerance to abiotic stresses is a challenge for afforestation programs. In this work, maritime pine (Pinus pinaster Aiton) plants were produced from somatic embryos matured at different temperatures (18, 23, or 28 °C, named after M18, M23, and M28, respectively) and after 2 years in the greenhouse a heat stress treatment (45 °C for 3 h/day for 10 days) was applied. Temperature variation during embryo development resulted in altered phenotypes (leaf histology, proline content, photosynthetic rates, and hormone profile) before and after stress. The thickness of chlorenchyma was initially larger in M28 plants, but was significantly reduced after heat stress, while increased in M18 plants. Irrespective of their origin, when these plants were subjected to a heat treatment, relative water content (RWC) and photosynthetic carbon assimilation rates were not significantly affected, although M18 plants increased net photosynthesis rate after 10 days recovery (tR). M18 plants showed proline contents that increased dramatically (2.4-fold) when subjected to heat stress, while proline contents remained unaffected in M23 and M28 plants. Heat stress significantly increased abscisic acid (ABA) content in the needles of maritime pine plants (1.4-, 3.6- and 1.9-fold in M18, M23, and M28 plants, respectively), while indole-3-acetic acid content only increased in needles from M23 plants. After the heat treatment, the total cytokinin contents of needles decreased significantly, particularly in M18 and M28 plants, although levels of active forms (cytokinin bases) did not change in M18 plants. In conclusion, our results suggest that maturation of maritime pine somatic embryos at lower temperature resulted in plants with better performance when subjected to subsequent high temperature stress, as demonstrated by faster and higher proline increase, lower increases in ABA levels, no reduction in active cytokinin, and a better net photosynthesis rate recovery.
Impaired Intracellular Ca2+ Dynamics, M-Band and Sarcomere Fragility in Skeletal Muscles of Obscurin KO Mice
Authors: Pierantozzi Enrico , Szentesi Péter , Paolini Cecilia , Dienes Beatrix , Fodor János , Oláh Tamás , Colombini Barbara , Rassier Dilson E. , Rubino Egidio Maria , Lange Stephan , Rossi Daniela , Csernoch László , Bagni Maria Angela , Reggiani Carlo , Sorrentino Vincenzo
Published: January 2022

Obscurin is a giant sarcomeric protein expressed in striated muscles known to establish several interactions with other proteins of the sarcomere, but also with proteins of the sarcoplasmic reticulum and costameres. Here, we report experiments aiming to better understand the contribution of obscurin to skeletal muscle fibers, starting with a detailed characterization of the diaphragm muscle function, which we previously reported to be the most affected muscle in obscurin (Obscn) KO mice. Twitch and tetanus tension were not significantly different in the diaphragm of WT and Obscn KO mice, while the time to peak (TTP) and half relaxation time (HRT) were prolonged. Differences in force-frequency and force-velocity relationships and an enhanced fatigability are observed in an Obscn KO diaphragm with respect to WT controls. Voltage clamp experiments show that a sarcoplasmic reticulum’s Ca2+ release and SERCA reuptake rates were decreased in muscle fibers from Obscn KO mice, suggesting that an impairment in intracellular Ca2+ dynamics could explain the observed differences in the TTP and HRT in the diaphragm. In partial contrast with previous observations, Obscn KO mice show a normal exercise tolerance, but fiber damage, the altered sarcomere ultrastructure and M-band disarray are still observed after intense exercise.
Sp1 Plays a Key Role in Vasculogenic Mimicry of Human Prostate Cancer Cells
Authors: Han Deok-Soo , Lee Eun-Ok
Published: January 2022

Sp1 transcription factor regulates genes involved in various phenomena of tumor progression. Vasculogenic mimicry (VM) is the alternative neovascularization by aggressive tumor cells. However, there is no evidence of the relationship between Sp1 and VM. This study investigated whether and how Sp1 plays a crucial role in the process of VM in human prostate cancer (PCa) cell lines, PC-3 and DU145. A cell viability assay and three-dimensional culture VM tube formation assay were performed. Protein and mRNA expression levels were detected by Western blot and reverse transcriptase-polymerase chain reaction, respectively. The nuclear twist expression was observed by immunofluorescence assay. A co-immunoprecipitation assay was performed. Mithramycin A (MiA) and Sp1 siRNA significantly decreased serum-induced VM, whereas Sp1 overexpression caused a significant induction of VM. Serum-upregulated vascular endothelial cadherin (VE-cadherin) protein and mRNA expression levels were decreased after MiA treatment or Sp1 silencing. The protein expression and the nuclear localization of twist were increased by serum, which was effectively inhibited after MiA treatment or Sp1 silencing. The interaction between Sp1 and twist was reduced by MiA. On the contrary, Sp1 overexpression enhanced VE-cadherin and twist expressions. Serum phosphorylated AKT and raised matrix metalloproteinase-2 (MMP-2) and laminin subunit 5 gamma-2 (LAMC2) expressions. MiA or Sp1 silencing impaired these effects. However, Sp1 overexpression upregulated phosphor-AKT, MMP-2 and LAMC2 expressions. Serum-upregulated Sp1 was significantly reduced by an AKT inhibitor, wortmannin. These results demonstrate that Sp1 mediates VM formation through interacting with the twist/VE-cadherin/AKT pathway in human PCa cells.
Role of the mtDNA Mutations and Mitophagy in Inflammaging
Authors: Dabravolski Siarhei A. , Nikiforov Nikita G. , Zhuravlev Alexander D. , Orekhov Nikolay A. , Grechko Andrey V. , Orekhov Alexander N.
Published: January 2022

Ageing is an unavoidable multi-factorial process, characterised by a gradual decrease in physiological functionality and increasing vulnerability of the organism to environmental factors and pathogens, ending, eventually, in death. One of the most elaborated ageing theories implies a direct connection between ROS-mediated mtDNA damage and mutations. In this review, we focus on the role of mitochondrial metabolism, mitochondria generated ROS, mitochondrial dynamics and mitophagy in normal ageing and pathological conditions, such as inflammation. Also, a chronic form of inflammation, which could change the long-term status of the immune system in an age-dependent way, is discussed. Finally, the role of inflammaging in the most common neurodegenerative diseases, such as Alzheimer’s and Parkinson’s, is also discussed.
A Polymorphism of Bactericidal/Permeability-Increasing Protein Affects Its Neutralization Efficiency towards Lipopolysaccharide
Authors: Ederer Katharina U. , Holzinger Jonas M. , Maier Katharina T. , Zeller Lisa , Werner Maren , Toelge Martina , Gessner André , Bülow Sigrid
Published: January 2022

Gram-negative sepsis driven by lipopolysaccharide (LPS) has detrimental outcomes, especially in neonates. The neutrophil-derived bactericidal/permeability-increasing protein (BPI) potently neutralizes LPS. Interestingly, polymorphism of the BPI gene at position 645 (rs4358188) corresponds to a favorable survival rate of these patients in the presence of at least one allele 645 A as opposed to 645 G. When we exploited the existing X-ray crystal structure, the corresponding amino acid at position 216 was revealed as surface exposed and proximal to the lipid-binding pocket in the N-terminal domain of BPI. Our further analysis predicted a shift in surface electrostatics by a positively charged lysine (BPI216K) exchanging a negatively charged glutamic acid (BPI216E). To investigate differences in interaction with LPS, we expressed both BPI variants recombinantly. The amino acid exchange neither affected affinity towards LPS nor altered bactericidal activity. However, when stimulating human peripheral blood mononuclear cells, BPI216K exhibited a superior LPS-neutralizing capacity (IC50 12.0 ± 2.5 pM) as compared to BPI216E (IC50 152.9 ± 113.4 pM, p = 0.0081) in respect to IL-6 secretion. In conclusion, we provide a functional correlate to a favorable outcome of sepsis in the presence of BPI216K.
Improving Protein Quantity and Quality—The Next Level of Plant Molecular Farming
Authors: Liu Hai , Timko Michael P.
Published: January 2022

Plants offer several unique advantages in the production of recombinant pharmaceuticals for humans and animals. Although numerous recombinant proteins have been expressed in plants, only a small fraction have been successfully put into use. The hugely distinct expression systems between plant and animal cells frequently cause insufficient yield of the recombinant proteins with poor or undesired activity. To overcome the issues that greatly constrain the development of plant-produced pharmaceuticals, great efforts have been made to improve expression systems and develop alternative strategies to increase both the quantity and quality of the recombinant proteins. Recent technological revolutions, such as targeted genome editing, deconstructed vectors, virus-like particles, and humanized glycosylation, have led to great advances in plant molecular farming to meet the industrial manufacturing and clinical application standards. In this review, we discuss the technological advances made in various plant expression platforms, with special focus on the upstream designs and milestone achievements in improving the yield and glycosylation of the plant-produced pharmaceutical proteins.
Genome Editing Using CRISPR-Cas9 and Autoimmune Diseases: A Comprehensive Review
Authors: Lee Min Ho , Shin Jae Il , Yang Jae Won , Lee Keum Hwa , Cha Do Hyeon , Hong Jun Beom , Park Yeoeun , Choi Eugene , Tizaoui Kalthoum , Koyanagi Ai , Jacob Louis , Park Seoyeon , Kim Ji Hong , Smith Lee
Published: January 2022

Autoimmune diseases are disorders that destruct or disrupt the body’s own tissues by its own immune system. Several studies have revealed that polymorphisms of multiple genes are involved in autoimmune diseases. Meanwhile, gene therapy has become a promising approach in autoimmune diseases, and clustered regularly interspaced palindromic repeats and CRISPR-associated protein 9 (CRISPR-Cas9) has become one of the most prominent methods. It has been shown that CRISPR-Cas9 can be applied to knock out proprotein convertase subtilisin/kexin type 9 (PCSK9) or block PCSK9, resulting in lowering low-density lipoprotein cholesterol. In other studies, it can be used to treat rare diseases such as ornithine transcarbamylase (OTC) deficiency and hereditary tyrosinemia. However, few studies on the treatment of autoimmune disease using CRISPR-Cas9 have been reported so far. In this review, we highlight the current and potential use of CRISPR-Cas9 in the management of autoimmune diseases. We summarize the potential target genes for immunomodulation using CRISPR-Cas9 in autoimmune diseases including rheumatoid arthritis (RA), inflammatory bowel diseases (IBD), systemic lupus erythematosus (SLE), multiple sclerosis (MS), type 1 diabetes mellitus (DM), psoriasis, and type 1 coeliac disease. This article will give a new perspective on understanding the use of CRISPR-Cas9 in autoimmune diseases not only through animal models but also in human models. Emerging approaches to investigate the potential target genes for CRISPR-Cas9 treatment may be promising for the tailored immunomodulation of some autoimmune diseases in the near future.
The Role of Matrix Proteins in Cardiac Pathology
Authors: Trinh Katie , Julovi Sohel M. , Rogers Natasha M.
Published: January 2022

The extracellular matrix (ECM) and ECM-regulatory proteins mediate structural and cell-cell interactions that are crucial for embryonic cardiac development and postnatal homeostasis, as well as organ remodeling and repair in response to injury. These proteins possess a broad functionality that is regulated by multiple structural domains and dependent on their ability to interact with extracellular substrates and/or cell surface receptors. Several different cell types (cardiomyocytes, fibroblasts, endothelial and inflammatory cells) within the myocardium elaborate ECM proteins, and their role in cardiovascular (patho)physiology has been increasingly recognized. This has stimulated robust research dissecting the ECM protein function in human health and disease and replicating the genetic proof-of-principle. This review summarizes recent developments regarding the contribution of ECM to cardiovascular disease. The clear importance of this heterogeneous group of proteins in attenuating maladaptive repair responses provides an impetus for further investigation into these proteins as potential pharmacological targets in cardiac diseases and beyond.
DNA Polymerase-Parental DNA Interaction Is Essential for Helicase-Polymerase Coupling during Bacteriophage T7 DNA Replication
Authors: Lo Chen-Yu , Gao Yang
Published: January 2022

DNA helicase and polymerase work cooperatively at the replication fork to perform leading-strand DNA synthesis. It was believed that the helicase migrates to the forefront of the replication fork where it unwinds the duplex to provide templates for DNA polymerases. However, the molecular basis of the helicase-polymerase coupling is not fully understood. The recently elucidated T7 replisome structure suggests that the helicase and polymerase sandwich parental DNA and each enzyme pulls a daughter strand in opposite directions. Interestingly, the T7 polymerase, but not the helicase, carries the parental DNA with a positively charged cleft and stacks at the fork opening using a β-hairpin loop. Here, we created and characterized T7 polymerases each with a perturbed β-hairpin loop and positively charged cleft. Mutations on both structural elements significantly reduced the strand-displacement synthesis by T7 polymerase but had only a minor effect on DNA synthesis performed against a linear DNA substrate. Moreover, the aforementioned mutations eliminated synergistic helicase-polymerase binding and unwinding at the DNA fork and processive fork progressions. Thus, our data suggested that T7 polymerase plays a dominant role in helicase-polymerase coupling and replisome progression.
Cortical Dysplasia and the mTOR Pathway: How the Study of Human Brain Tissue Has Led to Insights into Epileptogenesis
Authors: Lee Wei Shern , Baldassari Sara , Stephenson Sarah E. M. , Lockhart Paul J. , Baulac Stéphanie , Leventer Richard J.
Published: January 2022

Type II focal cortical dysplasia (FCD) is a neuropathological entity characterised by cortical dyslamination with the presence of dysmorphic neurons only (FCDIIA) or the presence of both dysmorphic neurons and balloon cells (FCDIIB). The year 2021 marks the 50th anniversary of the recognition of FCD as a cause of drug resistant epilepsy, and it is now the most common reason for epilepsy surgery. The causes of FCD remained unknown until relatively recently. The study of resected human FCD tissue using novel genomic technologies has led to remarkable advances in understanding the genetic basis of FCD. Mechanistic parallels have emerged between these non-neoplastic lesions and neoplastic disorders of cell growth and differentiation, especially through perturbations of the mammalian target of rapamycin (mTOR) signalling pathway. This narrative review presents the advances through which the aetiology of FCDII has been elucidated in chronological order, from recognition of an association between FCD and the mTOR pathway to the identification of somatic mosaicism within FCD tissue. We discuss the role of a two-hit mechanism, highlight current challenges and future directions in detecting somatic mosaicism in brain and discuss how knowledge of FCD may inform novel precision treatments of these focal epileptogenic malformations of human cortical development.
The Transmembrane Protease TMPRSS2 as a Therapeutic Target for COVID-19 Treatment
Authors: Wettstein Lukas , Kirchhoff Frank , Münch Jan
Published: January 2022

TMPRSS2 is a type II transmembrane protease with broad expression in epithelial cells of the respiratory and gastrointestinal tract, the prostate, and other organs. Although the physiological role of TMPRSS2 remains largely elusive, several endogenous substrates have been identified. TMPRSS2 serves as a major cofactor in SARS-CoV-2 entry, and primes glycoproteins of other respiratory viruses as well. Consequently, inhibiting TMPRSS2 activity is a promising strategy to block viral infection. In this review, we provide an overview of the role of TMPRSS2 in the entry processes of different respiratory viruses. We then review the different classes of TMPRSS2 inhibitors and their clinical development, with a focus on COVID-19 treatment.
Investigating the Molecular Mechanisms of Renal Hepcidin Induction and Protection upon Hemoglobin-Induced Acute Kidney Injury
Authors: Diepeveen Laura E. , Stegemann Gaby , Wiegerinck Erwin T. , Roelofs Rian , Naber Myrthe , Lóreal Olivier , Smeets Bart , Thévenod Frank , Swinkels Dorine W. , van Swelm Rachel P. L.
Published: January 2022

Hemolysis is known to cause acute kidney injury (AKI). The iron regulatory hormone hepcidin, produced by renal distal tubules, is suggested to exert a renoprotective role during this pathology. We aimed to elucidate the molecular mechanisms of renal hepcidin synthesis and its protection against hemoglobin-induced AKI. In contrast to known hepatic hepcidin induction, incubation of mouse cortical collecting duct (mCCDcl1) cells with IL-6 or LPS did not induce Hamp1 mRNA expression, whereas iron (FeS) and hemin significantly induced hepcidin synthesis (p < 0.05). Moreover, iron/heme-mediated hepcidin induction in mCCDcl1 cells was caused by the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway, as indicated by increased nuclear Nrf2 translocation and induced expression of Nrf2 downstream targets GCLM (p < 0.001), NQO1 (p < 0.001), and TXNRD1 (p < 0.005), which could be prevented by the known Nrf2 inhibitor trigonelline. Newly created inducible kidney-specific hepcidin KO mice demonstrated a significant reduction in renal Hamp1 mRNA expression. Phenylhydrazine (PHZ)-induced hemolysis caused renal iron loading and oxidative stress in both wildtype (Wt) and KO mice. PHZ treatment in Wt induced inflammatory markers (IL-6, TNFα) but not Hamp1. However, since PHZ treatment also significantly reduced systemic hepcidin levels in both Wt and KO mice (both p < 0.001), a dissection between the roles of systemic and renal hepcidin could not be made. Combined, the results of our study indicate that there are kidney-specific mechanisms in hepcidin regulation, as indicated by the dominant role of iron and not inflammation as an inducer of renal hepcidin, but also emphasize the complex interplay of various iron regulatory mechanisms during AKI on a local and systemic level.
Strain Echocardiography to Predict Postoperative Atrial Fibrillation
Authors: Sánchez Francisco Javier , Pueyo Esther , Diez Emiliano Raúl
Published: January 2022

Postoperative atrial fibrillation (POAF) complicates 15% to 40% of cardiovascular surgeries. Its incidence progressively increases with aging, reaching 50% in octogenarians. This arrhythmia is usually transient but it increases the risk of embolic stroke, prolonged hospital stay, and cardiovascular mortality. Though many pathophysiological mechanisms are known, POAF prediction is still a hot topic of discussion. Doppler echocardiogram and, lately, strain echocardiography have shown significant capacity to predict POAF. Alterations in oxidative stress, calcium handling, mitochondrial dysfunction, inflammation, fibrosis, and tissue aging are among the mechanisms that predispose patients to the perfect “atrial storm”. Manifestations of these mechanisms have been related to enlarged atria and impaired function, which can be detected prior to surgery. Specific alterations in the atrial reservoir and pump function, as well as atrial dyssynchrony determined by echocardiographic atrial strain, can predict POAF and help to shed light on which patients could benefit from preventive therapy.
Overexpression of a Cotton Aquaporin Gene GhTIP1;1-like Confers Cold Tolerance in Transgenic Arabidopsis
Authors: Cheng Gongmin , Wang Mengdi , Zhang Longyan , Wei Hengling , Wang Hantao , Lu Jianhua , Yu Shuxun
Published: January 2022

Cold stress can significantly affect the development, yield, and quality of crops and restrict the geographical distribution and growing seasons of plants. Aquaporins are the main channels for water transport in plant cells. Abiotic stresses such as cold and drought dehydrate cells by changing the water potential. In this study, we cloned a gene GhTIP1;1-like encodes tonoplast aquaporin from the transcriptome database of cotton seedlings after cold stress. Expression analysis showed that GhTIP1;1-like not only responds to cold stress but was also induced by heat, drought and salt stress. Subcellular localization showed that the protein was anchored to the vacuole membrane. Promoter deletion analysis revealed that a MYC motif within the promoter region of GhTIP1;1-like were the core cis-elements in response to low temperature. Virus-induced gene silencing (VIGS) and histochemical staining indicate that GhTIP1;1-like plays a positive role in plant cold tolerance. Overexpression of GhTIP1;1-like in Arabidopsis delayed the senescence process and enhanced the cold tolerance of transgenic plants. Compared with the wild type, the soluble protein concentration and peroxidase activity of the transgenic lines under cold stress were higher, while the malondialdehyde content was lower. In addition, the expression levels of cold-responsive genes were significantly increased in transgenic plants under cold stress. Our results indicate that GhTIP1;1-like could respond to different abiotic stresses and be positively involved in regulating the cold tolerance of cotton.
Functional Characterization of Mechanosensitive Piezo1 Channels in Trigeminal and Somatic Nerves in a Neuron-on-Chip Model
Authors: Mikhailov Nikita , Plotnikova Lidiia , Singh Prateek , Giniatullin Rashid , Hämäläinen Riikka H.
Published: January 2022

Mechanosensitive ion channels, Piezo1 and 2, are activated by pressure and involved in diverse physiological functions, including senses of touch and pain, proprioception and many more. Understanding their function is important for elucidating the mechanosensitive mechanisms of a range of human diseases. Recently, Piezo channels were suggested to be contributors to migraine pain generation. Migraine is typically characterized by allodynia and mechanical hyperalgesia associated with the activation and sensitization of trigeminal ganglion (TG) nerve fibers. Notably, migraine specific medicines are ineffective for other types of pain, suggesting a distinct underlying mechanism. To address, in a straightforward manner, the specificity of the mechanosensitivity of trigeminal vs. somatic nerves, we compared the activity of Piezo1 channels in mouse TG neurons vs. dorsal root ganglia (DRG) neurons. We assessed the functional expression of Piezo1 receptors using a conventional live calcium imaging setup equipped with a multibarrel application system and utilizing a microfluidic chip-based setup. Surprisingly, the TG neurons, despite higher expression of the Piezo1 gene, were less responsive to Piezo1 agonist Yoda1 than the DRG neurons. This difference was more prominent in the chip-based setup, suggesting that certain limitations of the conventional approach, such as turbulence, can be overcome by utilizing microfluidic devices with laminar solution flow.
Extracellular Vesicles Derived from Young Neural Cultures Attenuate Astrocytic Reactivity In Vitro
Authors: Almansa Daniel , Peinado Héctor , García-Rodríguez Raquel , Casadomé-Perales Álvaro , Dotti Carlos G. , Guix Francesc X.
Published: January 2022

Extracellular vesicles (EVs) play an important role in intercellular communication and are involved in both physiological and pathological processes. In the central nervous system (CNS), EVs secreted from different brain cell types exert a sundry of functions, from modulation of astrocytic proliferation and microglial activation to neuronal protection and regeneration. However, the effect of aging on the biological functions of neural EVs is poorly understood. In this work, we studied the biological effects of small EVs (sEVs) isolated from neural cells maintained for 14 or 21 days in vitro (DIV). We found that EVs isolated from 14 DIV cultures reduced the extracellular levels of lactate dehydrogenase (LDH), the expression levels of the astrocytic protein GFAP, and the complexity of astrocyte architecture suggesting a role in lowering the reactivity of astrocytes, while EVs produced by 21 DIV cells did not show any of the above effects. These results in an in vitro model pave the way to evaluate whether similar results occur in vivo and through what mechanisms.
Topical Melatonin Exerts Immunomodulatory Effect and Improves Dermatitis Severity in a Mouse Model of Atopic Dermatitis
Authors: Chang Yung-Sen , Tsai Chih-Chen , Yang Pang-Yan , Tang Chih-Yu , Chiang Bor-Luen
Published: January 2022

Oral melatonin supplement has been shown to improve dermatitis severity in children with AD, but the mechanism of the effect is unclear, and it is uncertain whether melatonin has a direct immunomodulatory effect on the dermatitis. Topical melatonin treatment was applied to DNCB-stimulated Balb/c mice, and gross and pathological skin findings, serum IgE, and cytokine levels in superficial lymph nodes were analyzed. Secretion of chemokines and cell proliferative response after melatonin treatment in human keratinocyte HaCaT cells were also studied. We found that in DNCB-stimulated Balb/c mice, topical melatonin treatment improved gross dermatitis severity, reduced epidermal hyperplasia and lymphocyte infiltration in the skin, and decreased IP-10, CCL27, IL-4, and IL-17 levels in superficial skin-draining lymph nodes. Melatonin also reduced cytokine-induced secretion of AD-related chemokines IP-10 and MCP-1 and decreased IL-4-induced cell proliferation in HaCaT cells. Melatonin seems to have an immunomodulatory effect on AD, with IP-10 as a possible target, and topical melatonin treatment is a potentially useful treatment for patients with AD.
Nischarin Deletion Reduces Oxidative Metabolism and Overall ATP: A Study Using a Novel NISCHΔ5-6 Knockout Mouse Model
Authors: Nguyen Tina H. , Yousefi Hassan , Okpechi Samuel C. , Lauterboeck Lothar , Dong Shengli , Yang Qinglin , Alahari Suresh K.
Published: January 2022

Nischarin (Nisch) is a cytosolic scaffolding protein that harbors tumor-suppressor-like characteristics. Previous studies have shown that Nisch functions as a scaffolding protein and regulates multiple biological activities. In the current study, we prepared a complete Nisch knockout model, for the first time, by deletion of exons 5 and 6. This knockout model was confirmed by Qrt–PCR and Western blotting with products from mouse embryonic fibroblast (MEF) cells. Embryos and adult mice of knockouts are significantly smaller than their wild-type counterparts. Deletion of Nisch enhanced cell migration, as demonstrated by wound type and transwell migration assays. Since the animals were small in size, we investigated Nisch’s effect on metabolism by conducting several assays using the Seahorse analyzer system. These data indicate that Nisch null cells have lower oxygen consumption rates, lower ATP production, and lower levels of proton leak. We examined the expression of 15 genes involved in lipid and fat metabolism, as well as cell growth, and noted a significant increase in expression for many genes in Nischarin null animals. In summary, our results show that Nischarin plays an important physiological role in metabolic homeostasis.
Histamine H4 Receptor Agonism Induces Antitumor Effects in Human T-Cell Lymphoma
Authors: Clauzure Mariángeles , Táquez Delgado Mónica A. , Phillip Jude M. , Revuelta Maria V. , Cerchietti Leandro , Medina Vanina A.
Published: January 2022

The discovery of the human histamine H4 receptor (H4R) has contributed to our understanding of the role of histamine in numerous physiological and pathological conditions, including tumor development and progression. The lymph nodes of patients with malignant lymphomas have shown to contain high levels of histamine, however, less is known regarding the expression and function of the H4R in T-cell lymphoma (TCL). In this work we demonstrate the expression of H4R isoforms (mRNA and protein) in three human aggressive TCL (OCI-Ly12, Karpas 299, and HuT78). Histamine and specific H4R agonists (VUF8430 and JNJ28610244) significantly reduced cell viability in a dose-dependent manner (p < 0.05). The combined treatment with the H4R antagonist (JNJ7777120, 10 µM) reversed the effects of the H4R ligands. Importantly, we screened a drug repurposing library of 433 FDA-approved compounds (1 μM) in combination with histamine (10 μM) in Hut78 cells. Histamine produced a favorable antitumor effect with 18 of these compounds, including the histone deacetylase inhibitor panobinostat. Apoptosis, proliferation, and oxidative stress studies confirmed the antitumoral effects of the combination. We conclude that the H4R is expressed in TCL, and it is involved in histamine-mediated responses.
The Role of Small Extracellular Vesicles in the Progression of Colorectal Cancer and Its Clinical Applications
Authors: Chang Li-Chun , Chiu Han-Mo , Wu Ming-Shiang , Shen Tang-Long
Published: January 2022

Colorectal cancer (CRC) is one of the most common cancers worldwide and a longstanding critical challenge for public health. Screening has been suggested to effectively reduce both the incidence and mortality of CRC. However, the drawback of the current screening modalities, both stool-based tests and colonoscopies, is limited screening adherence, which reduces the effectiveness of CRC screening. Blood tests are more acceptable than stool tests or colonoscopy as a first-line screening approach. Therefore, identifying blood biomarkers for detecting CRC and its precancerous neoplasms is urgently needed to fulfill the unmet clinical need. Currently, many kinds of blood contents, such as circulating tumor cells, circulating tumor nucleic acids, and extracellular vesicles, have been investigated as biomarkers for CRC detection. Among these, small extracellular vesicles (sEVs) have been demonstrated to detect CRC effectively in recent reports. sEVs enable intercellular shuttling—for instance, trafficking between recipient cancer cells and stromal cells—which can affect tumor initiation, proliferation, angiogenesis, immune regulation; metastasis, the cancer-specific molecules, such as proteins, microRNAs, long noncoding RNAs, and circular RNAs, loaded into cancer-derived sEVs may serve as biomarkers for the detection of cancers, including CRC. Indeed, accumulating evidence has shown that nucleic acids and proteins contained in CRC-derived sEVs are effective as blood biomarkers for CRC detection. However, investigations of the performance of sEVs for diagnosing CRC in clinical trials remains limited. Thus, the effectiveness of sEV biomarkers for diagnosing CRC needs further validation in clinical trials.