bluuebunny/crossref_metadata_2025 huggingface dataset:
Brassinazole Resistant 1 Activity Is Organ-Specific and Genotype-Dependent in Barley Seedlings
Authors: Groszyk Jolanta , Szechyńska-Hebda Magdalena
Published: December 2021

Brassinosteroids (BRs) control many plant developmental processes by regulating different groups of transcription factors, and consequently gene expressions. The most known is BZR1, the main member of the BES1 family. However, to date, it is poorly characterized in crop species. The main goal of the presented study was to identify HvBZR1 and determine its activity in 5-day-old barley (the stage is related to one leaf on the main shoot and a few seminal roots) using two cultivars with different sensitivities to BRs. Using the anti-OsBZR1 antibody, we identified the forms of HvBZR1 transcription factor with different molecular weights, which can be related to different phosphorylated forms of serine/threonine residues. Two phosphorylated forms in the shoots and one dephosphorylated form in the roots were determined. A minor amount of the dephosphorylated form of the HvBZR1 in the Haruna Nijo shoots was also found. The phosphorylated forms gave a higher band intensity for Golden Promise than Haruna Nijo. The bands were similar in their intensity, when two different phosphorylated forms were compared in Golden Promise, while a reduced intensity was detected for the phosphorylated form with a lower molecular weight for Haruna Nijo. Degradation of the phosphorylated forms in the shoots (complete degradation in Golden Promise and significant but not complete in Haruna Nijo) and the presence of the dephosphorylated form in the roots were proven for the etiolated barley. In the case of Haruna Nijo, a wider range of the regulators of the BR biosynthesis and signaling pathways induced the expected effects, 24-EBL (0.001 µM) and bikinin (10 and 50 µM) caused low amount of the phosphorylated forms, and at the same time, a tiny band of dephosphorylated form was detected. However, the expression of genes related to the BR biosynthesis and signaling pathways was not a determinant for the protein amount.
Glial-Neuronal Interactions in Pathogenesis and Treatment of Spinal Cord Injury
Authors: Lukacova Nadezda , Kisucka Alexandra , Kiss Bimbova Katarina , Bacova Maria , Ileninova Maria , Kuruc Tomas , Galik Jan
Published: December 2021

Traumatic spinal cord injury (SCI) elicits an acute inflammatory response which comprises numerous cell populations. It is driven by the immediate response of macrophages and microglia, which triggers activation of genes responsible for the dysregulated microenvironment within the lesion site and in the spinal cord parenchyma immediately adjacent to the lesion. Recently published data indicate that microglia induces astrocyte activation and determines the fate of astrocytes. Conversely, astrocytes have the potency to trigger microglial activation and control their cellular functions. Here we review current information about the release of diverse signaling molecules (pro-inflammatory vs. anti-inflammatory) in individual cell phenotypes (microglia, astrocytes, blood inflammatory cells) in acute and subacute SCI stages, and how they contribute to delayed neuronal death in the surrounding spinal cord tissue which is spared and functional but reactive. In addition, temporal correlation in progressive degeneration of neurons and astrocytes and their functional interactions after SCI are discussed. Finally, the review highlights the time-dependent transformation of reactive microglia and astrocytes into their neuroprotective phenotypes (M2a, M2c and A2) which are crucial for spontaneous post-SCI locomotor recovery. We also provide suggestions on how to modulate the inflammation and discuss key therapeutic approaches leading to better functional outcome after SCI.
Long Noncoding RNAs: Recent Insights into Their Role in Male Infertility and Their Potential as Biomarkers and Therapeutic Targets
Authors: Zhao Shanjiang , Heng Nuo , Weldegebriall Sahlu Bahlibi , Wang Huan , Zhu Huabin
Published: December 2021

Long noncoding RNAs (lncRNAs) are composed of nucleotides located in the nucleus and cytoplasm; these are transcribed by RNA polymerase II and are greater than 200 nt in length. LncRNAs fulfill important functions in a variety of biological processes, including genome imprinting, cell differentiation, apoptosis, stem cell pluripotency, X chromosome inactivation and nuclear transport. As high throughput sequencing technology develops, a substantial number of lncRNAs have been found to be related to a variety of biological processes, such as development of the testes, maintaining the self-renewal and differentiation of spermatogonial stem cells, and regulating spermatocyte meiosis. These indicate that lncRNAs can be used as biomarkers and potential therapeutic targets for male infertility. However, only a few comprehensive reviews have described the role of lncRNAs in male reproduction. In this paper, we summarize recent findings relating to the role of lncRNAs in spermatogenesis, their potential as biomarkers for male infertility and the relationship between reproductive arrest and transgenerational effects. Finally, we suggest specific targets for the treatment of male infertility from the perspective of lncRNAs.
Augmented KCa2.3 Channel Feedback Regulation of Oxytocin Stimulated Uterine Strips from Nonpregnant Mice
Authors: Zak Megan , Kestler Bri , Cornwell Trudy , Taylor Mark S.
Published: December 2021

Uterine contractions prior to 37 weeks gestation can result in preterm labor with significant risk to the infant. Current tocolytic therapies aimed at suppressing premature uterine contractions are largely ineffective and cause serious side effects. Calcium (Ca2+) dependent contractions of uterine smooth muscle are physiologically limited by the opening of membrane potassium (K+) channels. Exploiting such inherent negative feedback mechanisms may offer new strategies to delay labor and reduce risk. Positive modulation of small conductance Ca2+-activated K+ (KCa2.3) channels with cyclohexyl-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-pyrimidin-4-yl]-amine (CyPPA), effectively decreases uterine contractions. This study investigates whether the receptor agonist oxytocin might solicit KCa2.3 channel feedback that facilitates CyPPA suppression of uterine contractions. Using isometric force myography, we found that spontaneous phasic contractions of myometrial tissue from nonpregnant mice were suppressed by CyPPA and, in the presence of CyPPA, oxytocin failed to augment contractions. In tissues exposed to oxytocin, depletion of internal Ca2+ stores with cyclopiazonic acid (CPA) impaired CyPPA relaxation, whereas blockade of nonselective cation channels (NSCC) using gadolinium (Gd3+) had no significant effect. Immunofluorescence revealed close proximity of KCa2.3 channels and ER inositol trisphosphate receptors (IP3Rs) within myometrial smooth muscle cells. The findings suggest internal Ca2+ stores play a role in KCa2.3-dependent feedback control of uterine contraction and offer new insights for tocolytic therapies.
Dynamic Coupling of Tyrosine 185 with the Bacteriorhodopsin Photocycle, as Revealed by Chemical Shifts, Assisted AF-QM/MM Calculations and Molecular Dynamic Simulations
Authors: Chen Sijin , Ding Xiaoyan , Sun Chao , Watts Anthony , He Xiao , Zhao Xin
Published: December 2021

Aromatic residues are highly conserved in microbial photoreceptors and play crucial roles in the dynamic regulation of receptor functions. However, little is known about the dynamic mechanism of the functional role of those highly conserved aromatic residues during the receptor photocycle. Tyrosine 185 (Y185) is a highly conserved aromatic residue within the retinal binding pocket of bacteriorhodopsin (bR). In this study, we explored the molecular mechanism of the dynamic coupling of Y185 with the bR photocycle by automated fragmentation quantum mechanics/molecular mechanics (AF-QM/MM) calculations and molecular dynamic (MD) simulations based on chemical shifts obtained by 2D solid-state NMR correlation experiments. We observed that Y185 plays a significant role in regulating the retinal cis–trans thermal equilibrium, stabilizing the pentagonal H-bond network, participating in the orientation switch of Schiff Base (SB) nitrogen, and opening the F42 gate by interacting with the retinal and several key residues along the proton translocation channel. Our findings provide a detailed molecular mechanism of the dynamic couplings of Y185 and the bR photocycle from a structural perspective. The method used in this paper may be applied to the study of other microbial photoreceptors.
Exogenous Application of Low-Concentration Sugar Enhances Brassinosteroid Signaling for Skotomorphogenesis by Promoting BIN2 Degradation
Authors: Sheng Huachun , Zhang Shuangxi , Wei Yanping , Chen Shaolin
Published: December 2021

In plants, seedling growth is subtly controlled by multiple environmental factors and endogenous phytohormones. The cross-talk between sugars and brassinosteroid (BR) signaling is known to regulate plant growth; however, the molecular mechanisms that coordinate hormone-dependent growth responses with exogenous sucrose in plants are incompletely understood. Skotomorphogenesis is a plant growth stage with rapid elongation of the hypocotyls. In the present study, we found that low-concentration sugars could improve skotomorphogenesis in a manner dependent on BR biosynthesis and TOR activation. However, accumulation of BZR1 in bzr1-1D mutant plants partially rescued the defects of skotomorphogenesis induced by the TOR inhibitor AZD, and these etiolated seedlings displayed a normal phenotype like that of wild-type seedlings in response to both sucrose and non-sucrose treatments, thereby indicating that accumulated BZR1 sustained, at least partially, the sucrose-promoted growth of etiolated seedlings (skotomorphogenesis). Moreover, genetic evidence based on a phenotypic analysis of bin2-3bil1bil2 triple-mutant and gain-of-function bin2–1 mutant plant indicated that BIN2 inactivation was conducive to skotomorphogenesis in the dark. Subsequent biochemical and molecular analyses enabled us to confirm that sucrose reduced BIN2 levels via the TOR–S6K2 pathway in etiolated seedlings. Combined with a determination of the cellulose content, our results indicated that sucrose-induced BIN2 degradation led to the accumulation of BZR1 and the enhancement of cellulose synthesis, thereby promoting skotomorphogenesis, and that BIN2 is the converging node that integrates sugar and BR signaling.
Plasma Membrane Calcium ATPase-Neuroplastin Complexes Are Selectively Stabilized in GM1-Containing Lipid Rafts
Authors: Ilic Katarina , Lin Xiao , Malci Ayse , Stojanović Mario , Puljko Borna , Rožman Marko , Vukelić Željka , Heffer Marija , Montag Dirk , Schnaar Ronald L. , Kalanj-Bognar Svjetlana , Herrera-Molina Rodrigo , Mlinac-Jerkovic Kristina
Published: December 2021

The recent identification of plasma membrane (Ca2+)-ATPase (PMCA)-Neuroplastin (Np) complexes has renewed attention on cell regulation of cytosolic calcium extrusion, which is of particular relevance in neurons. Here, we tested the hypothesis that PMCA-Neuroplastin complexes exist in specific ganglioside-containing rafts, which could affect calcium homeostasis. We analyzed the abundance of all four PMCA paralogs (PMCA1-4) and Neuroplastin isoforms (Np65 and Np55) in lipid rafts and bulk membrane fractions from GM2/GD2 synthase-deficient mouse brains. In these fractions, we found altered distribution of Np65/Np55 and selected PMCA isoforms, namely PMCA1 and 2. Cell surface staining and confocal microscopy identified GM1 as the main complex ganglioside co-localizing with Neuroplastin in cultured hippocampal neurons. Furthermore, blocking GM1 with a specific antibody resulted in delayed calcium restoration of electrically evoked calcium transients in the soma of hippocampal neurons. The content and composition of all ganglioside species were unchanged in Neuroplastin-deficient mouse brains. Therefore, we conclude that altered composition or disorganization of ganglioside-containing rafts results in changed regulation of calcium signals in neurons. We propose that GM1 could be a key sphingolipid for ensuring proper location of the PMCA-Neuroplastin complexes into rafts in order to participate in the regulation of neuronal calcium homeostasis.
Entrectinib—A SARS-CoV-2 Inhibitor in Human Lung Tissue (HLT) Cells
Authors: Peralta-Garcia Alejandro , Torrens-Fontanals Mariona , Stepniewski Tomasz Maciej , Grau-Expósito Judith , Perea David , Ayinampudi Vikram , Waldhoer Maria , Zimmermann Mirjam , Buzón María J. , Genescà Meritxell , Selent Jana
Published: December 2021

Since the start of the COVID-19 outbreak, pharmaceutical companies and research groups have focused on the development of vaccines and antiviral drugs against SARS-CoV-2. Here, we apply a drug repurposing strategy to identify drug candidates that are able to block the entrance of the virus into human cells. By combining virtual screening with in vitro pseudovirus assays and antiviral assays in Human Lung Tissue (HLT) cells, we identify entrectinib as a potential antiviral drug.
Characterizing the Interaction between the HTLV-1 Transactivator Tax-1 with Transcription Elongation Factor ELL2 and Its Impact on Viral Transactivation
Authors: Kohrt Stephan , Strobel Sarah , Mann Melanie , Sticht Heinrich , Fleckenstein Bernhard , Thoma-Kress Andrea
Published: December 2021

The human T-cell leukemia virus type 1 (HTLV-1)-encoded transactivator and oncoprotein Tax-1 is essential for HTLV-1 replication. We recently found that Tax-1 interacts with transcription elongation factor for RNA polymerase II 2, ELL2, which enhances Tax-1-mediated transactivation of the HTLV-1 promotor. Here, we characterize the Tax-1:ELL2 interaction and its impact on viral transactivation by confocal imaging, co-immunoprecipitation, and luciferase assays. We found that Tax-1 and ELL2 not only co-precipitate, but also co-localize in dot-like structures in the nucleus. Tax-1:ELL2 complex formation occurred independently of Tax-1 point mutations, which are crucial for post translational modifications (PTMs) of Tax-1, suggesting that these PTMs are irrelevant for Tax-1:ELL2 interaction. In contrast, Tax-1 deletion mutants lacking either N-terminal (aa 1–37) or C-terminal regions (aa 150–353) of Tax-1 were impaired in interacting with ELL2. Contrary to Tax-1, the related, non-oncogenic Tax-2B from HTLV-2B did not interact with ELL2. Finally, we found that ELL2-R1 (aa 1–353), which carries an RNA polymerase II binding domain, and ELL2-R3 (aa 515–640) are sufficient to interact with Tax-1; however, only ELL2-truncations expressing R1 could enhance Tax-1-mediated transactivation of the HTLV-1 promoter. Together, this study identifies domains in Tax-1 and ELL2 being required for Tax-1:ELL2 complex formation and for viral transactivation.
Duodenal Dysbiosis and Relation to the Efficacy of Proton Pump Inhibitors in Functional Dyspepsia
Authors: Wauters Lucas , Tito Raúl Y. , Ceulemans Matthias , Lambaerts Maarten , Accarie Alison , Rymenans Leen , Verspecht Chloë , Toth Joran , Mols Raf , Augustijns Patrick , Tack Jan , Vanuytsel Tim , Raes Jeroen
Published: December 2021

Proton pump inhibitors (PPI) may improve symptoms in functional dyspepsia (FD) through duodenal eosinophil-reducing effects. However, the contribution of the microbiome to FD symptoms and its interaction with PPI remains elusive. Aseptic duodenal brushings and biopsies were performed before and after PPI intake (4 weeks Pantoprazole 40 mg daily, FD-starters and controls) or withdrawal (2 months, FD-stoppers) for 16S-rRNA sequencing. Between- and within-group changes in genera or diversity and associations with symptoms or duodenal factors were analyzed. In total, 30 controls, 28 FD-starters and 19 FD-stoppers were followed. Mucus-associated Porphyromonas was lower in FD-starters vs. controls and correlated with symptoms in FD and duodenal eosinophils in both groups, while Streptococcus correlated with eosinophils in controls. Although clinical and eosinophil-reducing effects of PPI therapy were unrelated to microbiota changes in FD-starters, increased Streptococcus was associated with duodenal PPI effects in controls and remained higher despite withdrawal of long-term PPI therapy in FD-stoppers. Thus, duodenal microbiome analysis demonstrated differential mucus-associated genera, with a potential role of Porphyromonas in FD pathophysiology. While beneficial effects of short-term PPI therapy were not associated with microbial changes in FD-starters, increased Streptococcus and its association with PPIeffects in controls suggest a role for duodenal dysbiosis after long-term PPI therapy.
Lytic Polysaccharide Monooxygenase from Talaromyces amestolkiae with an Enigmatic Linker-like Region: The Role of This Enzyme on Cellulose Saccharification
Authors: Méndez-Líter Juan Antonio , Ayuso-Fernández Iván , Csarman Florian , de Eugenio Laura Isabel , Míguez Noa , Plou Francisco J. , Prieto Alicia , Ludwig Roland , Martínez María Jesús
Published: December 2021

The first lytic polysaccharide monooxygenase (LPMO) detected in the genome of the widespread ascomycete Talaromyces amestolkiae (TamAA9A) has been successfully expressed in Pichia pastoris and characterized. Molecular modeling of TamAA9A showed a structure similar to those from other AA9 LPMOs. Although fungal LPMOs belonging to the genera Penicillium or Talaromyces have not been analyzed in terms of regioselectivity, phylogenetic analyses suggested C1/C4 oxidation which was confirmed by HPAEC. To ascertain the function of a C-terminal linker-like region present in the wild-type sequence of the LPMO, two variants of the wild-type enzyme, one without this sequence and one with an additional C-terminal carbohydrate binding domain (CBM), were designed. The three enzymes (native, without linker and chimeric variant with a CBM) were purified in two chromatographic steps and were thermostable and active in the presence of H2O2. The transition midpoint temperature of the wild-type LPMO (Tm = 67.7 °C) and its variant with only the catalytic domain (Tm = 67.6 °C) showed the highest thermostability, whereas the presence of a CBM reduced it (Tm = 57.8 °C) and indicates an adverse effect on the enzyme structure. Besides, the potential of the different T. amestolkiae LPMO variants for their application in the saccharification of cellulosic and lignocellulosic materials was corroborated.
Single-Dose P2 X4R Single-Chain Fragment Variable Antibody Permanently Reverses Chronic Pain in Male Mice
Authors: Westlund Karin N. , Montera Marena A. , Goins Aleyah E. , Alles Sascha R. A. , Suri Nikita , McIlwrath Sabrina L. , Bartel Robyn , Durvasula Ravi V. , Kunamneni Adinarayana
Published: December 2021

Non-opioid single-chain variable fragment (scFv) small antibodies were generated as pain-reducing block of P2X4R receptor (P2X4R). A panel of scFvs targeting an extracellular peptide sequence of P2X4R was generated followed by cell-free ribosome display for recombinant antibody selection. After three rounds of bio-panning, a panel of recombinant antibodies was isolated and characterized by ELISA, cross-reactivity analysis, and immunoblotting/immunostaining. Generated scFv antibodies feature binding activity similar to monoclonal antibodies but with stronger affinity and increased tissue penetrability due to their ~30% smaller size. Two anti-P2X4R scFv clones (95, 12) with high specificity and affinity binding were selected for in vivo testing in male and female mice with trigeminal nerve chronic neuropathic pain (FRICT-ION model) persisting for several months in untreated BALBc mice. A single dose of P2X4R scFv (4 mg/kg, i.p.) successfully, completely, and permanently reversed chronic neuropathic pain-like measures in male mice only, providing retention of baseline behaviors indefinitely. Untreated mice retained hypersensitivity, and developed anxiety- and depression-like behaviors within 5 weeks. In vitro P2X4R scFv 95 treatment significantly increased the rheobase of larger-diameter (>25 µm) trigeminal ganglia (TG) neurons from FRICT-ION mice compared to controls. The data support use of engineered scFv antibodies as non-opioid biotherapeutic interventions for chronic pain.
Investigating Pathogenicity and Virulence of Staphylococcus pettenkoferi: An Emerging Pathogen
Authors: Ahmad-Mansour Nour , Plumet Lucile , Huc-Brandt Sylvaine , Magnan Chloé , Yahiaoui-Martinez Alex , Kissa Karima , Pantel Alix , Lavigne Jean-Philippe , Molle Virginie
Published: December 2021

Staphylococcus pettenkoferi is a coagulase-negative Staphylococcus identified in 2002 that has been implicated in human diseases as an opportunistic pathogenic bacterium. Its multiresistant character is becoming a major health problem, yet the pathogenicity of S. pettenkoferi is poorly characterized. In this study, the pathogenicity of a S. pettenkoferi clinical isolate from diabetic foot osteomyelitis was compared with a Staphylococcus aureus strain in various in vitro and in vivo experiments. Growth kinetics were compared against S. aureus, and bacteria survival was assessed in the RAW 264.7 murine macrophage cell line, the THP-1 human leukemia monocytic cell line, and the HaCaT human keratinocyte cell line. Ex vivo analysis was performed in whole blood survival assays and in vivo assays via the infection model of zebrafish embryos. Moreover, whole-genome analysis was performed. Our results show that S. pettenkoferi was able to survive in human blood, human keratinocytes, murine macrophages, and human macrophages. S. pettenkoferi demonstrated its virulence by causing substantial embryo mortality in the zebrafish model. Genomic analysis revealed virulence factors such as biofilm-encoding genes (e.g., icaABCD; rsbUVW) and regulator-encoding genes (e.g., agr, mgrA, sarA, saeS) well characterized in S. aureus. This study thus advances the knowledge of this under-investigated pathogen and validates the zebrafish infection model for this bacterium.
Tumor-Specific miRNA Signatures in Combination with CA19-9 for Liquid Biopsy-Based Detection of PDAC
Authors: Kim Min Woo , Koh Hani , Kim Jee Ye , Lee Suji , Lee Hyojung , Kim Young , Hwang Ho Kyoung , Kim Seung Il
Published: December 2021

Pancreatic ductal adenocarcinoma (PDAC) is considered one of the most aggressive malignancies and has high mortality and poor survival rates. Therefore, there is an urgent need to discover non-invasive biomarkers for early detection before PDAC reaches the incurable stage. We hypothesized that liquid biopsy of PDAC-derived extracellular vesicles (PDEs) containing abundant microRNAs (miRNAs) could be used for early diagnosis of PDAC because they can be selectively enriched and because they are biologically stable. We isolated PDEs by immunocapture using magnetic beads, and we identified 13 miRNA candidates in 20 pancreatic cancer patients and 20 normal controls. We found that expression of five miRNAs, including miR-10b, miR-16, miR-155, miR-429, and miR-1290, was markedly higher in PDEs. Furthermore, the miRNA signatures along with serum carbohydrate antigen 19-9 (CA19-9) were optimized by logistic regression, and the miRNA signature and CA19-9 combination markers (CMs) were effective at differentiating PDAC patients from normal controls. As a result, the CMs represented a high sensitivity (AUC, 0.964; sensitivity, 100%; specificity, 80%) and a high specificity (AUC, 0.962; sensitivity, 85.71%; specificity, 100%). These findings suggest that five miRNAs expressed in PDEs and CA19-9 are valuable biomarkers for screening and diagnosis of pancreatic cancer by liquid biopsy.
Polyurethane Foam Incorporated with Nanosized Copper-Based Metal-Organic Framework: Its Antibacterial Properties and Biocompatibility
Authors: Lee Do Nam , Gwon Kihak , Nam Yunhee , Lee Su Jung , Tran Ngoc Minh , Yoo Hyojong
Published: December 2021

Polyurethane foams (PUFs) have attracted attention as biomaterials because of their low adhesion to the wound area and suitability as biodegradable or bioactive materials. The composition of the building blocks for PUFs can be controlled with additives, which provide excellent anti-drug resistance and biocompatibility. Herein, nanosized Cu-BTC (copper(II)-benzene-1,3,5-tricarboxylate) was incorporated into a PUF via the crosslinking reaction of castor oil and chitosan with toluene-2,4-diisocyanate, to enhance therapeutic efficiency through the modification of the surface of PUF. The physical and thermal properties of the nanosized Cu-BTC-incorporated PUF (PUF@Cu-BTC), e.g., swelling ratio, phase transition, thermal gravity loss, and cell morphology, were compared with those of the control PUF. The bactericidal activities of PUF@Cu-BTC and control PUF were evaluated against Pseudomonas aeruginosa, Klebsiella pneumoniae, and methicillin-resistant Staphylococcus aureus. PUF@Cu-BTC exhibited selective and significant antibacterial activity toward the tested bacteria and lower cytotoxicity for mouse embryonic fibroblasts compared with the control PUF at a dose of 2 mg mL−1. The Cu(II) ions release test showed that PUF@Cu-BTC was stable in phosphate buffered saline (PBS) for 24 h. The selective bactericidal activity and low cytotoxicity of PUF@Cu-BTC ensure it is a candidate for therapeutic applications for the drug delivery, treatment of skin disease, and wound healing.
Enhanced Anti-Atherosclerotic Efficacy of pH-Responsively Releasable Ganglioside GM3 Delivered by Reconstituted High-Density Lipoprotein
Authors: Rong Tong , Wei Bo , Ao Meiying , Zhao Haonan , Li Yuanfang , Zhang Yang , Qin Ying , Zhou Jinhua , Zhou Fenfen , Chen Yong
Published: December 2021

Recently, the atheroprotective role of endogenous GM3 and an atherogenesis-inhibiting effect of exogenous GM3 suggested a possibility of exogenous GM3 being recruited as an anti-atherosclerotic drug. This study seeks to endow exogenous GM3 with atherosclerotic targetability via reconstituted high-density lipoprotein (rHDL), an atherosclerotic targeting drug nanocarrier. Unloaded rHDL, rHDL loaded with exogenous GM3 at a low concentration (GM3L-rHDL), and rHDL carrying GM3 at a relatively high concentration (GM3H-rHDL) were prepared and characterized. The inhibitory effect of GM3-rHDL on lipid deposition in macrophages was confirmed, and GM3-rHDL did not affect the survival of red blood cells. In vivo experiments using ApoE−/− mice fed a high fat diet further confirmed the anti-atherosclerotic efficacy of exogenous GM3 and demonstrated that GM3 packed in HDL nanoparticles (GM3-rHDL) has an enhanced anti-atherosclerotic efficacy and a reduced effective dose of GM3. Then, the macrophage- and atherosclerotic plaque-targeting abilities of GM3-rHD, most likely via the interaction of ApoA-I on GM3-rHDL with its receptors (e.g., SR-B1) on cells, were certified via a microsphere-based method and an aortic fragment-based method, respectively. Moreover, we found that solution acidification enhanced GM3 release from GM3-rHDL nanoparticles, implying the pH-responsive GM3 release when GM3-rHDL enters the acidic atherosclerotic plaques from the neutral blood. The rHDL-mediated atherosclerotic targetability and pH-responsive GM3 release of GM3-rHDL enhanced the anti-atherosclerotic efficacy of exogenous GM3. The development of the GM3-rHDL nanoparticle may help with the application of exogenous GM3 as a clinical drug. Moreover, the data imply that the GM3-rHDL nanoparticle has the potential of being recruited as a drug nanocarrier with atherosclerotic targetability and enhanced anti-atherosclerotic efficacy.
Maternal N-Acetyl-Cysteine Prevents Neonatal Hypoxia-Induced Brain Injury in a Rat Model
Authors: Gutziet Ola , Iluz Roee , Ben Asher Hila , Segal Linoy , Ben Zvi Dikla , Ginsberg Yuval , Khatib Nizar , Zmora Osnat , Ross Michael G. , Weiner Zeev , Beloosesky Ron
Published: December 2021

Perinatal hypoxia is a major cause of infant brain damage, lifelong neurological disability, and infant mortality. N-Acetyl-Cysteine (NAC) is a powerful antioxidant that acts directly as a scavenger of free radicals. We hypothesized that maternal-antenatal and offspring-postnatal NAC can protect offspring brains from hypoxic brain damage.Sixty six newborn rats were randomized into four study groups. Group 1: Control (CON) received no hypoxic intervention. Group 2: Hypoxia (HYP)-received hypoxia protocol. Group 3: Hypoxia-NAC (HYP-NAC). received hypoxia protocol and treated with NAC following each hypoxia episode. Group 4: NAC Hypoxia (NAC-HYP) treated with NAC during pregnancy, pups subject to hypoxia protocol. Each group was evaluated for: neurological function (Righting reflex), serum proinflammatory IL-6 protein levels (ELISA), brain protein levels: NF-κB p65, neuronal nitric oxide synthase (nNOS), TNF-α, and IL-6 (Western blot) and neuronal apoptosis (histology evaluation with TUNEL stain). Hypoxia significantly increased pups brain protein levels compared to controls. NAC administration to dams or offspring demonstrated lower brain NF-κB p65, nNOS, TNF-α and IL-6 protein levels compared to hypoxia alone. Hypoxia significantly increased brain apoptosis as evidenced by higher grade of brain TUNEL reaction. NAC administration to dams or offspring significantly reduce this effect. Hypoxia induced acute sensorimotor dysfunction. NAC treatment to dams significantly attenuated hypoxia-induced acute sensorimotor dysfunction. Prophylactic NAC treatment of dams during pregnancy confers long-term protection to offspring with hypoxia associated brain injury, measured by several pathways of injury and correlated markers with pathology and behavior. This implies we may consider prophylactic NAC treatment for patients at risk for hypoxia during labor.
Multi-Omics Reveal the Immunological Role and the Theragnostic Value of miR-216a/GDF15 Axis in Human Colon Adenocarcinoma
Authors: Tung Chun-Bin , Li Chia-Ying , Lin Hung-Yu
Published: December 2021

Colon adenocarcinoma (COAD) is the most common type of gastrointestinal cancer and is still the third leading cause of cancer-related mortality worldwide. Accurate screening tools for early diagnosis and prediction of prognosis and precision treatment strategies are urgently required to accommodate the unmet medical needs of COAD management. We herein aimed to explore the significance of the microRNA (miR)-216a/growth differentiation factor 15 (GDF15) axis in terms of clinical value, tumor immunity, and potential mechanisms in COAD by using multi-omic analysis. The gene expression levels of miR-216a and GDF15 showed an increase in the COAD group compared to those of the normal group. The expression of miR-216a presented a negative correlation with GDF15 in COAD tumor tissue. The use of an in vitro luciferase reporter assay and bioinformatic prediction revealed that miR-216a-3p acted toward translational inhibition on GDF15 by targeting its 3′untranslated region (UTR) site. High miR-216a expression was associated with decreased overall survival (OS), while the high expression of GDF15 was associated with increased OS. Enriched type 1 T-helper (Th1), enriched regulatory T (Treg), enriched eosinophils, and decreased nature killer T-cells (NKTs) in COAD tumor tissue may play counteracting factors on the tumor-regulatory effects of miR-216a and GDF15. In addition, high GDF15 expression had associations with suppressed immunoinhibitory genes and negative correlations with the infiltration of macrophages and endothelial cells. The enrichment analysis revealed that GDF15 and its co-expression network may be implicated in mitochondrial organization, apoptosis signaling, and endoplasmic reticulum (ER) stress response. The Genomics of Drug Sensitivity in Cancer (GDSC) and Cancer Therapeutics Response Portal (CTRP) analysis identified that Gemcitabine acted as a precision treatment for COAD when GDF15 expression was low. This study supports the miR-216a/GDF15 axis as a diagnostic/prognostic panel for COAD, identifies Th1, Treg, eosinophils, and NKTs as counteracting factors, indicates potential relationships underlying immunomodulation, mitochondrial organization, apoptotic signaling, and ER stress and unveil Gemcitabine as a potential drug for the development of treatment strategy when combined with targeting GDF15.
The Effect of a Sustained High-Fat Diet on the Metabolism of White and Brown Adipose Tissue and Its Impact on Insulin Resistance: A Selected Time Point Cross-Sectional Study
Authors: Maharjan Babu Raja , McLennan Susan V. , Yee Christine , Twigg Stephen M. , Williams Paul F.
Published: December 2021

(1) Background: studies on the long-term dynamic changes in fat depot metabolism in response to a high-fat diet (HFD) on hepatic lipid deposition and insulin resistance are sparse. This study investigated the dynamic changes produced by HFD and the production of dysfunctional fat depots on insulin resistance and liver lipid metabolism. (2) Methods: mice fed a chow or HFD (45% kcal fat) diet had three fat depots, liver, and blood collected at 6, 10, 20, and 30 weeks. Anthropometric changes and gene markers for adipogenesis, thermogenesis, ECM remodeling, inflammation, and tissue insulin resistance were measured. (3) Results: early responses to the HFD were increased body weight, minor deposition of lipid in liver, increased adipocyte size, and adipogenesis. Later changes were dysfunctional adipose depots, increased liver fat, insulin resistance (shown by changes in ITT) accompanied by increased inflammatory markers, increased fibrosis (fibrosis > 2-fold, p < 0.05 from week 6), and the presence of crown cells in white fat depots. Later, changes did not increase thermogenic markers in response to the increased calories and decreased UCP1 and PRDM16 proteins in WAT. (4) Conclusions: HFD feeding initially increased adipocyte diameter and number, but later changes caused adipose depots to become dysfunctional, restricting adipose tissue expansion, changing the brown/beige ratios in adipose depots, and causing ectopic lipid deposition and insulin resistance.
In Situ Cell Signalling of the Hippo-YAP/TAZ Pathway in Reaction to Complex Dynamic Loading in an Intervertebral Disc Organ Culture
Authors: Croft Andreas S. , Roth Ysaline , Oswald Katharina A. C. , Ćorluka Slavko , Bermudez-Lekerika Paola , Gantenbein Benjamin
Published: December 2021

Recently, a dysregulation of the Hippo-YAP/TAZ pathway has been correlated with intervertebral disc (IVD) degeneration (IDD), as it plays a key role in cell survival, tissue regeneration, and mechanical stress. We aimed to investigate the influence of different mechanical loading regimes, i.e., under compression and torsion, on the induction and progression of IDD and its association with the Hippo-YAP/TAZ pathway. Therefore, bovine IVDs were assigned to one of four different static or complex dynamic loading regimes: (i) static, (ii) “low-stress”, (iii) “intermediate-stress”, and (iv) “high-stress” regime using a bioreactor. After one week of loading, a significant loss of relative IVD height was observed in the intermediate- and high-stress regimes. Furthermore, the high-stress regime showed a significantly lower cell viability and a significant decrease in glycosaminoglycan content in the tissue. Finally, the mechanosensitive gene CILP was significantly downregulated overall, and the Hippo-pathway gene MST1 was significantly upregulated in the high-stress regime. This study demonstrates that excessive torsion combined with compression leads to key features of IDD. However, the results indicated no clear correlation between the degree of IDD and a subsequent inactivation of the Hippo-YAP/TAZ pathway as a means of regenerating the IVD.
Mucins Dynamics in Physiological and Pathological Conditions
Authors: Melhem Hassan , Regan-Komito Daniel , Niess Jan Hendrik
Published: December 2021

Maintaining intestinal health requires clear segregation between epithelial cells and luminal microbes. The intestinal mucus layer, produced by goblet cells (GCs), is a key element in maintaining the functional protection of the epithelium. The importance of the gut mucus barrier is highlighted in mice lacking Muc2, the major form of secreted mucins. These mice show closer bacterial residence to epithelial cells, develop spontaneous colitis and became moribund when infected with the attaching and effacing pathogen, Citrobacter rodentium. Furthermore, numerous observations have associated GCs and mucus layer dysfunction to the pathogenesis of inflammatory bowel disease (IBD). However, the molecular mechanisms that regulate the physiology of GCs and the mucus layer remain obscured. In this review, we consider novel findings describing divergent functionality and expression profiles of GCs subtypes within intestinal crypts. We also discuss internal (host) and external (diets and bacteria) factors that modulate different aspects of the mucus layer as well as the contribution of an altered mucus barrier to the onset of IBD.
Role of Circulating Biomarkers in Platinum-Resistant Ovarian Cancer
Authors: Sassu Carolina Maria , Palaia Innocenza , Boccia Serena Maria , Caruso Giuseppe , Perniola Giorgia , Tomao Federica , Di Donato Violante , Musella Angela , Muzii Ludovico
Published: December 2021

Ovarian cancer (OC) is the second most common cause of death in women with gynecological cancer. Considering the poor prognosis, particularly in the case of platinum-resistant (PtR) disease, a huge effort was made to define new biomarkers able to help physicians in approaching and treating these challenging patients. Currently, most data can be obtained from tumor biopsy samples, but this is not always available and implies a surgical procedure. On the other hand, circulating biomarkers are detected with non-invasive methods, although this might require expensive techniques. Given the fervent hope in their value, here we focused on the most studied circulating biomarkers that could play a role in PtR OC.
Partial Agonistic Actions of Sex Hormone Steroids on TRPM3 Function
Authors: Persoons Eleonora , Kerselaers Sara , Voets Thomas , Vriens Joris , Held Katharina
Published: December 2021

Sex hormone steroidal drugs were reported to have modulating actions on the ion channel TRPM3. Pregnenolone sulphate (PS) presents the most potent known endogenous chemical agonist of TRPM3 and affects several gating modes of the channel. These includes a synergistic action of PS and high temperatures on channel opening and the PS-induced opening of a noncanonical pore in the presence of other TRPM3 modulators. Moreover, human TRPM3 variants associated with neurodevelopmental disease exhibit an increased sensitivity for PS. However, other steroidal sex hormones were reported to influence TRPM3 functions with activating or inhibiting capacity. Here, we aimed to answer how DHEAS, estradiol, progesterone and testosterone act on the various modes of TRPM3 function in the wild-type channel and two-channel variants associated with human disease. By means of calcium imaging and whole-cell patch clamp experiments, we revealed that all four drugs are weak TRPM3 agonists that share a common steroidal interaction site. Furthermore, they exhibit increased activity on TRPM3 at physiological temperatures and in channels that carry disease-associated mutations. Finally, all steroids are able to open the noncanonical pore in wild-type and DHEAS also in mutant TRPM3. Collectively, our data provide new valuable insights in TRPM3 gating, structure-function relationships and ligand sensitivity.
Recent Advances in the Inhibition of the IL-4 Cytokine Pathway for the Treatment of Allergen-Induced Asthma
Authors: Massey Oliver , Suphioglu Cenk
Published: December 2021

The IL-4 and IL-13 cytokine pathways play integral roles in stimulating IgE inflammation, with the IL-4 cytokine being a major cytokine in the etiology of thunderstorm asthma, atopic dermatitis, and allergic rhinitis. The increasing prevalence of thunderstorm asthma in the younger population and the lessening efficacy of corticosteroids and other anti-inflammatories has created a need for more effective pharmaceuticals. This review summarizes the IL-4 and IL-13 pathways while highlighting and discussing the current pathway inhibitors aimed at treating thunderstorm asthma and atopic dermatitis, as well as the potential efficacy of peptide therapeutics in this field.
Regulatory Peptides in Asthma
Authors: Kaczyńska Katarzyna , Zając Dominika , Wojciechowski Piotr , Jampolska Monika
Published: December 2021

Numerous regulatory peptides play a critical role in the pathogenesis of airway inflammation, airflow obstruction and hyperresponsiveness, which are hallmarks of asthma. Some of them exacerbate asthma symptoms, such as neuropeptide Y and tachykinins, while others have ameliorating properties, such as nociception, neurotensin or β-defensin 2. Interacting with peptide receptors located in the lungs or on immune cells opens up new therapeutic possibilities for the treatment of asthma, especially when it is resistant to available therapies. This article provides a concise review of the most important and current findings regarding the involvement of regulatory peptides in asthma pathology.
Molecular Characterization of U-box E3 Ubiquitin Ligases (TaPUB2 and TaPUB3) Involved in the Positive Regulation of Drought Stress Response in Arabidopsis
Authors: Kim Jae Ho , Kim Moon Seok , Kim Dae Yeon , Amoah Joseph Noble , Seo Yong Weon
Published: December 2021

Plant U-box E3 ubiquitin ligase (PUB) is involved in various environmental stress conditions. However, the molecular mechanism of U-box proteins in response to abiotic stress in wheat remains unknown. In this study, two U-box E3 ligase genes (TaPUB2 and TaPUB3), which are highly expressed in response to adverse abiotic stresses, were isolated from common wheat, and their cellular functions were characterized under drought stress. Transient expression assay revealed that TaPUB2 was localized in the cytoplasm and Golgi apparatus, whereas TaPUB3 was expressed only in the Golgi apparatus in wheat protoplasts. Additionally, TaPUB2 and TaPUB3 underwent self-ubiquitination. Moreover, TaPUB2/TaPUB3 heterodimer was identified in yeast and the cytoplasm of wheat protoplasts using a pull-down assay and bimolecular fluorescence complementation analysis. Heterogeneous overexpression of TaPUB2 and TaPUB3 conferred tolerance to drought stress. Taken together, these results implied that the heterodimeric form of U-box E3 ubiquitin ligases (TaPUB2/TaPUB3) responded to abiotic stress and roles as a positive regulator of drought stress tolerance.
New Inhibitors of Laccase and Tyrosinase by Examination of Cross-Inhibition between Copper-Containing Enzymes
Authors: Chaudhary Dinesh , Chong Fangchen , Neupane Trilok , Choi Joonhyeok , Jee Jun-Goo
Published: December 2021

Coppers play crucial roles in the maintenance homeostasis in living species. Approximately 20 enzyme families of eukaryotes and prokaryotes are known to utilize copper atoms for catalytic activities. However, small-molecule inhibitors directly targeting catalytic centers are rare, except for those that act against tyrosinase and dopamine-β-hydroxylase (DBH). This study tested whether known tyrosinase inhibitors can inhibit the copper-containing enzymes, ceruloplasmin, DBH, and laccase. While most small molecules minimally reduced the activities of ceruloplasmin and DBH, aside from known inhibitors, 5 of 28 tested molecules significantly inhibited the function of laccase, with the Ki values in the range of 15 to 48 µM. Enzyme inhibitory kinetics classified the molecules as competitive inhibitors, whereas differential scanning fluorimetry and fluorescence quenching supported direct bindings. To the best of our knowledge, this is the first report on organic small-molecule inhibitors for laccase. Comparison of tyrosinase and DBH inhibitors using cheminformatics predicted that the presence of thione moiety would suffice to inhibit tyrosinase. Enzyme assays confirmed this prediction, leading to the discovery of two new dual tyrosinase and DBH inhibitors.
Effects of Microvesicles Derived from NK Cells Stimulated with IL-1β on the Phenotype and Functional Activity of Endothelial Cells
Authors: Markova Kseniia , Mikhailova Valentina , Milyutina Yulia , Korenevsky Andrey , Sirotskaya Anastasia , Rodygina Veronika , Tyshchuk Elizaveta , Grebenkina Polina , Simbirtsev Andrey , Selkov Sergey , Sokolov Dmitry
Published: December 2021

Microvesicles (MVs) are plasma extracellular vesicles ranging from 100 (150) to 1000 nm in diameter. These are generally produced by different cells through their vital activity and are a source of various protein and non-protein molecules. It is assumed that MVs can mediate intercellular communication and modulate cell functions. The interaction between natural killer cells (NK cells) and endothelial cells underlies multiple pathological conditions. The ability of MVs derived from NK cells to influence the functional state of endothelial cells in inflammatory conditions has yet to be studied well. In this regard, we aimed to study the effects of MVs derived from NK cells of the NK-92 cell line stimulated with IL-1β on the phenotype, caspase activity, proliferation and migration of endothelial cells of the EA.hy926 cell line. Endothelial cells were cultured with MVs derived from cells of the NK-92 cell line after their stimulation with IL-1β. Using flow cytometry, we evaluated changes in the expression of endothelial cell surface molecules and endothelial cell death. We evaluated the effect of MVs derived from stimulated NK cells on the proliferative and migratory activity of endothelial cells, as well as the activation of caspase-3 and caspase-9 therein. It was established that the incubation of endothelial cells with MVs derived from cells of the NK-92 cell line stimulated with IL-1β and with MVs derived from unstimulated NK cells, leads to the decrease in the proliferative activity of endothelial cells, appearance of the pan leukocyte marker CD45 on them, caspase-3 activation and partial endothelial cell death, and reduced CD105 expression. However, compared with MVs derived from unstimulated NK cells, a more pronounced effect of MVs derived from cells of the NK-92 cell line stimulated with IL-1β was found in relation to the decrease in the endothelial cell migratory activity and the intensity of the CD54 molecule expression on them. The functional activity of MVs is therefore mediated by the conditions they are produced under, as well as their internal contents.
Gene-Expression Programming-Based Model for Estimating the Compressive Strength of Cement-Fly Ash Stabilized Soil and Parametric Study
Authors: Pham Van-Ngoc , Oh Erwin , Ong Dominic E. L.
Published: December 2021

The study aims to develop a reliable model using gene-expression programming (GEP) technique for estimating the unconfined compressive strength (UCS) of soil stabilization by cement and fly ash. The model considered the effects of several parameters, including the fly ash characteristics such as calcium oxide (CaO) content, CaO/SiO2 ratio, and loss of ignition. The research results show that the proposed model demonstrates superior performance with a high correlation coefficient (R > 0.955) and low errors. Therefore, the model could be confidently applied in practice for a variety of fly ash qualities. Besides, the parametric study was conducted to examine the effect of fly ash characteristics on the strength of soil stabilization. The study indicates that if the fly ash contains a high amount of calcium oxide, the strength of fly ash stabilized soil is significant. In addition, fly ash could be used in combination with cement to increase the strength of the mixture. A fly ash replacement ratio is suggested from 0.19 to 0.35, corresponding to the total binder used from 10% to 30%. The research findings could help engineers in optimizing the fly ash proportion and estimating the UCS of soil stabilization by cement and fly ash.