bluuebunny/crossref_metadata_2025 huggingface dataset:
Capsaicin-Sensitive Peptidergic Sensory Nerves Are Anti-Inflammatory Gatekeepers in the Hyperacute Phase of a Mouse Rheumatoid Arthritis Model
Authors: Botz Bálint , Kriszta Gábor , Bölcskei Kata , Horváth Ádám István , Mócsai Attila , Helyes Zsuzsanna
Published: February 2021

Capsaicin-sensitive peptidergic sensory nerves play complex, mainly protective regulatory roles in the inflammatory cascade of the joints via neuropeptide mediators, but the mechanisms of the hyperacute arthritis phase has not been investigated. Therefore, we studied the involvement of these afferents in the early, “black box” period of a rheumatoid arthritis (RA) mouse model. Capsaicin-sensitive fibres were defunctionalized by pretreatment with the ultrapotent capsaicin analog resiniferatoxin and arthritis was induced by K/BxN arthritogenic serum. Disease severity was assessed by clinical scoring, reactive oxygen species (ROS) burst by chemiluminescent, vascular permeability by fluorescent in vivo imaging. Contrast-enhanced magnetic resonance imaging was used to correlate the functional and morphological changes. After sensory desensitization, both early phase ROS-burst and vascular leakage were significantly enhanced, which was later followed by the increased clinical severity scores. Furthermore, the early vascular leakage and ROS-burst were found to be good predictors of later arthritis severity. We conclude that the anti-inflammatory role of peptidergic afferents depends on their activity in the hyperacute phase, characterized by decreased cellular and vascular inflammatory components presumably via anti-inflammatory neuropeptide release. Therefore, these fibres might serve as important gatekeepers in RA.
A Novel Triplet-Primed PCR Assay to Detect the Full Range of Trinucleotide CAG Repeats in the Huntingtin Gene (HTT)
Authors: De Luca Alessandro , Morella Annunziata , Consoli Federica , Fanelli Sergio , Thibert Julie R. , Statt Sarah , Latham Gary J. , Squitieri Ferdinando
Published: February 2021

The expanded CAG repeat number in HTT gene causes Huntington disease (HD), which is a severe, dominant neurodegenerative illness. The accurate determination of the expanded allele size is crucial to confirm the genetic status in symptomatic and presymptomatic at-risk subjects and avoid genetic polymorphism-related false-negative diagnoses. Precise CAG repeat number determination is critical to discriminate the cutoff between unexpanded and intermediate mutable alleles (IAs, 27–35 CAG) as well as between IAs and pathological, low-penetrance alleles (i.e., 36–39 CAG repeats), and it is also critical to detect large repeat expansions causing pediatric HD variants. We analyzed the HTT-CAG repeat number of 14 DNA reference materials and of a DNA collection of 43 additional samples carrying unexpanded, IAs, low and complete penetrance alleles, including large (>60 repeats) and very large (>100 repeats) expansions using a novel triplet-primed PCR-based assay, the AmplideX PCR/CE HTT Kit. The results demonstrate that the method accurately genotypes both normal and expanded HTT-CAG repeat numbers and reveals previously undisclosed and very large CAG expansions >200 repeats. We also show that this technique can improve genetic test reliability and accuracy by detecting CAG expansions in samples with sequence variations within or adjacent to the repeat tract that cause allele drop-outs or inaccuracies using other PCR methods.
Pathological Circulating Factors in Moyamoya Disease
Authors: Fang Yao-Ching , Wei Ling-Fei , Hu Chaur-Jong , Tu Yong-Kwang
Published: February 2021

Moyamoya disease (MMD) is a cerebrovascular disease that presents with vascular stenosis and a hazy network of collateral formations in angiography. However, the detailed pathogenic pathway remains unknown. Studies have indicated that in addition to variations in the of genetic factor RNF213, unusual circulating angiogenetic factors observed in patients with MMD may play a critical role in producing “Moyamoya vessels”. Circulating angiogenetic factors, such as growth factors, vascular progenitor cells, cytokines, inflammatory factors, and other circulating proteins, could promote intimal hyperplasia in vessels and excessive collateral formation with defect structures through endothelial hyperplasia, smooth muscle migration, and atypical neovascularization. This study summarizes the hypothesized pathophysiology of how these circulating factors affect MMD and the interactive modulation between them.
Ion and Water Transport in Neutrophil Granulocytes and Its Impairment during Sepsis
Authors: Messerer David Alexander Christian , Schmidt Hanna , Frick Manfred , Huber-Lang Markus
Published: February 2021

Neutrophil granulocytes are the vanguard of innate immunity in response to numerous pathogens. Their activity drives the clearance of microbe- and damage-associated molecular patterns, thereby contributing substantially to the resolution of inflammation. However, excessive stimulation during sepsis leads to cellular unresponsiveness, immunological dysfunction, bacterial expansion, and subsequent multiple organ dysfunction. During the short lifespan of neutrophils, they can become significantly activated by complement factors, cytokines, and other inflammatory mediators. Following stimulation, the cells respond with a defined (electro-)physiological pattern, including depolarization, calcium influx, and alkalization as well as with increased metabolic activity and polarization of the actin cytoskeleton. Activity of ion transport proteins and aquaporins is critical for multiple cellular functions of innate immune cells, including chemotaxis, generation of reactive oxygen species, and phagocytosis of both pathogens and tissue debris. In this review, we first describe the ion transport proteins and aquaporins involved in the neutrophil ion–water fluxes in response to chemoattractants. We then relate ion and water flux to cellular functions with a focus on danger sensing, chemotaxis, phagocytosis, and oxidative burst and approach the role of altered ion transport protein expression and activity in impaired cellular functions and cell death during systemic inflammation as in sepsis.
Anti-Cancer Effects of Glaucarubinone in the Hepatocellular Carcinoma Cell Line Huh7 via Regulation of the Epithelial-To-Mesenchymal Transition-Associated Transcription Factor Twist1
Authors: Seo Jihye , Ha Jain , Kang Eunjeong , Yoon Haelim , Lee Sewoong , Ryu Shi Yong , Kim Kwonseop , Cho Sayeon
Published: February 2021

Hepatocellular carcinoma (HCC), the most common type of liver cancer, is a leading cause of cancer-related deaths. As HCC has a high mortality rate and its incidence is increasing worldwide, understanding and treating HCC are crucial for resolving major public health concerns. In the present study, wound healing screening assays were performed using natural product libraries to identify natural chemicals that can inhibit cancer cell migration. Glaucarubinone (GCB) showed a high potential for inhibiting cell migration. The anti-cancer effects of GCB were evaluated using the HCC cell line, Huh7. GCB showed anti-cancer effects, as verified by wound healing, cell migration, invasion, colony formation, and three-dimensional spheroid invasion assays. In addition, cells treated with GCB showed suppressed matrix metalloproteinase activities. Immunoblotting analyses of intracellular signaling pathways revealed that GCB regulated the levels of Twist1, a crucial transcription factor associated with epithelial-to-mesenchymal transition, and mitogen-activated protein kinase. The invasive ability of cancer cells was found to be decreased by the regulation of Twist1 protein levels. Furthermore, GCB downregulated phosphorylation of extracellular signal-regulated kinase. These results indicate that GCB exhibits anti-metastatic properties in Huh7 cells, suggesting that it could be used to treat HCC.
Glutamine Synthetase as a Therapeutic Target for Cancer Treatment
Authors: Kim Go Woon , Lee Dong Hoon , Jeon Yu Hyun , Yoo Jung , Kim So Yeon , Lee Sang Wu , Cho Ha Young , Kwon So Hee
Published: February 2021

The significance of glutamine in cancer metabolism has been extensively studied. Cancer cells consume an excessive amount of glutamine to facilitate rapid proliferation. Thus, glutamine depletion occurs in various cancer types, especially in poorly vascularized cancers. This makes glutamine synthetase (GS), the only enzyme responsible for de novo synthesizing glutamine, essential in cancer metabolism. In cancer, GS exhibits pro-tumoral features by synthesizing glutamine, supporting nucleotide synthesis. Furthermore, GS is highly expressed in the tumor microenvironment (TME) and provides glutamine to cancer cells, allowing cancer cells to maintain sufficient glutamine level for glutamine catabolism. Glutamine catabolism, the opposite reaction of glutamine synthesis by GS, is well known for supporting cancer cell proliferation via contributing biosynthesis of various essential molecules and energy production. Either glutamine anabolism or catabolism has a critical function in cancer metabolism depending on the complex nature and microenvironment of cancers. In this review, we focus on the role of GS in a variety of cancer types and microenvironments and highlight the mechanism of GS at the transcriptional and post-translational levels. Lastly, we discuss the therapeutic implications of targeting GS in cancer.
Amino Acid Transporter LAT1 (SLC7A5) Mediates MeHg-Induced Oxidative Stress Defense in the Human Placental Cell Line HTR-8/SVneo
Authors: Granitzer Sebastian , Widhalm Raimund , Forsthuber Martin , Ellinger Isabella , Desoye Gernot , Hengstschläger Markus , Zeisler Harald , Salzer Hans , Gundacker Claudia
Published: February 2021

The placental barrier can protect the fetus from contact with harmful substances. The potent neurotoxin methylmercury (MeHg), however, is very efficiently transported across the placenta. Our previous data suggested that L-type amino acid transporter (LAT)1 is involved in placental MeHg uptake, accepting MeHg-L-cysteine conjugates as substrate due to structural similarity to methionine. The aim of the present study was to investigate the antioxidant defense of placental cells to MeHg exposure and the role of LAT1 in this response. When trophoblast-derived HTR-8/SVneo cells were LAT1 depleted by siRNA-mediated knockdown, they accumulated less MeHg. However, they were more susceptible to MeHg-induced toxicity. This was evidenced in decreased cell viability at a usually noncytotoxic concentration of 0.03 µM MeHg (~6 µg/L). Treatment with ≥0.3 µM MeHg increased cytotoxicity, apoptosis rate, and oxidative stress of HTR-8/SVneo cells. These effects were enhanced under LAT1 knockdown. Reduced cell number was seen when MeHg-exposed cells were cultured in medium low in cysteine, a constituent of the tripeptide glutathione (GSH). Because LAT1-deficient HTR-8/SVneo cells have lower GSH levels than control cells (independent of MeHg treatment), we conclude that LAT1 is essential for de novo synthesis of GSH, required to counteract oxidative stress. Genetic predisposition to decreased LAT1 function combined with MeHg exposure could increase the risk of placental damage.
Immunobiology and Application of Aloe vera-Based Scaffolds in Tissue Engineering
Authors: Darzi Saeedeh , Paul Kallyanashis , Leitan Shanilka , Werkmeister Jerome A. , Mukherjee Shayanti
Published: February 2021

Aloe vera (AV), a succulent plant belonging to the Liliaceae family, has been widely used for biomedical and pharmaceutical application. Its popularity stems from several of its bioactive components that have anti-oxidant, anti-microbial, anti-inflammatory and even immunomodulatory effects. Given such unique multi-modal biological impact, AV has been considered as a biomaterial for regenerative medicine and tissue engineering applications, where tissue repair and neo-angiogenesis are vital. This review outlines the growing scientific evidence that demonstrates the advantage of AV as tissue engineering scaffolds. We particularly highlight the recent advances in the application of AV-based scaffolds. From a tissue engineering perspective, it is pivotal that the implanted scaffolds strike an appropriate foreign body response to be well-accepted in the body without complications. Herein, we highlight the key cellular processes that regulate the foreign body response to implanted scaffolds and underline the immunomodulatory effects incurred by AV on the innate and adaptive system. Given that AV has several beneficial components, we discuss the importance of delving deeper into uncovering its action mechanism and thereby improving material design strategies for better tissue engineering constructs for biomedical applications.
Synthesis and Biological Evaluation of Oleanolic Acid Derivatives as Selective Vascular Endothelial Growth Factor Promoter i-Motif Ligands
Authors: Zeng Huang , Kang Shuangshuang , Zhang Yu , Liu Ke , Yu Qian , Li Ding , An Lin-Kun
Published: February 2021

Vascular endothelial growth factor (VEGF) is an angiogenic growth factor and plays a key role in tumor progression. The C-rich DNA sequence of VEGF promoter can form i-motif structure, which is a potential target for the development of novel anticancer agents. However, there is a limited number of chemotypes as the selective ligands of VEGF promoter i-motif, which leaves much room for development. Herein, we report the discovery of the natural oleanolic acid scaffold as a novel chemotype for the development of selective ligands of VEGF i-motif. A series of oleanolic acid derivatives as VEGF promoter i-motif ligands were synthesized. Subsequent evaluations showed that 3c could selectively bind to and stabilize VEGF promoter i-motif without significant binding to G-quadruplex, duplex DNA, and other oncogene i-motifs. Cell-based assays indicated that 3c could effectively downregulate VEGF gene transcription and expression in MCF-7 cells, inhibit tumor cells proliferation and migration, and induce cancer cells apoptosis. This work provides evidence of VEGF promoter i-motif as an anticancer target and will facilitate future efforts for the discovery of oleanolic acid-based selective ligands of VEGF promoter i-motif.
Plasminogen Receptors and Fibrinolysis
Authors: Miles Lindsey A. , Ny Lina , Wilczynska Malgorzata , Shen Yue , Ny Tor , Parmer Robert J.
Published: February 2021

The ability of cells to promote plasminogen activation on their surfaces is now well recognized, and several distinct cell surface proteins have been demonstrated to function as plasminogen receptors. Here, we review studies demonstrating that plasminogen bound to cells, in addition to plasminogen directly bound to fibrin, plays a major role in regulating fibrin surveillance. We focus on the ability of specific plasminogen receptors on eukaryotic cells to promote fibrinolysis in the in vivo setting by reviewing data obtained predominantly in murine models. Roles for distinct plasminogen receptors in fibrin surveillance in intravascular fibrinolysis, immune cell recruitment in the inflammatory response, wound healing, and lactational development are discussed.
The Effect of Neddylation Inhibition on Inflammation-Induced MMP9 Gene Expression in Esophageal Squamous Cell Carcinoma
Authors: Hryniewicz-Jankowska Anita , Wierzbicki Jaroslaw , Tabola Renata , Stach Kamilla , Sossey-Alaoui Khalid , Augoff Katarzyna
Published: February 2021

Inhibition of the protein neddylation process by the small-molecule inhibitor MLN4924 has been recently indicated as a promising direction for cancer treatment. However, the knowledge of all biological consequences of MLN4924 for cancer cells is still incomplete. Here, we report that MLN4924 inhibits tumor necrosis factor-alpha (TNF-α)-induced matrix metalloproteinase 9 (MMP9)-driven cell migration. Using real-time polymerase chain reaction (PCR) and gelatin zymography, we found that MLN4924 inhibited expression and activity of MMP9 at the messenger RNA (mRNA) and protein levels in both resting cells and cells stimulated with TNF-α, and this inhibition was closely related to impaired cell migration. We also revealed that MLN4924, similar to TNF-α, induced phosphorylation of inhibitor of nuclear factor kappa B-alpha (IκB-α). However, contrary to TNF-α, MLN4924 did not induce IκB-α degradation in treated cells. In coimmunoprecipitation experiments, nuclear IκB-α which formed complexes with nuclear factor kappa B p65 subunit (NFκB/p65) was found to be highly phosphorylated at Ser32 in the cells treated with MLN4924, but not in the cells treated with TNF-α alone. Moreover, in the presence of MLN4924, nuclear NFκB/p65 complexes were found to be enriched in c-Jun and cyclin dependent kinase inhibitor 1 A (CDKN1A/p21) proteins. In these cells, NFκB/p65 was unable to bind to the MMP9 gene promoter, which was confirmed by the chromatin immunoprecipitation (ChIP) assay. Taken together, our findings identified MLN4924 as a suppressor of TNF-α-induced MMP9-driven cell migration in esophageal squamous cell carcinoma (ESCC), likely acting by affecting the nuclear ubiquitin–proteasome system that governs NFκB/p65 complex formation and its DNA binding activity in regard to the MMP9 promoter, suggesting that inhibition of neddylation might be a new therapeutic strategy to prevent invasion/metastasis in ESCC patients.
Long, Noncoding RNA SRA Induces Apoptosis of β-Cells by Promoting the IRAK1/LDHA/Lactate Pathway
Authors: Huang Yu-Nan , Chiang Shang-Lun , Lin Yu-Jung , Liu Su-Ching , Li Yen-Hsien , Liao Yu-Chen , Lee Maw-Rong , Su Pen-Hua , Tsai Fuu-Jen , Hung Hui-Chih , Wang Chung-Hsing
Published: February 2021

Long non-coding RNA steroid receptor RNA activators (LncRNA SRAs) are implicated in the β-cell destruction of Type 1 diabetes mellitus (T1D), but functional association remains poorly understood. Here, we aimed to verify the role of LncRNA SRA regulation in β-cells. LncRNA SRAs were highly expressed in plasma samples and peripheral blood mononuclear cells (PBMCs) from T1D patients. LncRNA SRA was strongly upregulated by high-glucose treatment. LncRNA SRA acts as a microRNA (miR)-146b sponge through direct sequence–structure interactions. Silencing of lncRNA SRA increased the functional genes of Tregs, resulting in metabolic reprogramming, such as decreased lactate levels, repressed lactate dehydrogenase A (LDHA)/phosphorylated LDHA (pLDHA at Tyr10) expression, decreased reactive oxygen species (ROS) production, increased ATP production, and finally, decreased β-cell apoptosis in vitro. There was a positive association between lactate level and hemoglobin A1c (HbA1c) level in the plasma from patients with T1D. Recombinant human interleukin (IL)-2 treatment repressed lncRNA SRA expression and activity in β-cells. Higher levels of lncRNA-SRA/lactate in the plasma are associated with poor regulation in T1D patients. LncRNA SRA contributed to T1D pathogenesis through the inhibition of miR-146b in β-cells, with activating signaling transduction of interleukin-1 receptor-associated kinase 1 (IRAK1)/LDHA/pLDHA. Taken together, LncRNA SRA plays a critical role in the function of β-cells.
Effects of Platelet-Rich Plasma on Cellular Populations of the Central Nervous System: The Influence of Donor Age
Authors: Delgado Diego , Bilbao Ane Miren , Beitia Maider , Garate Ane , Sánchez Pello , González-Burguera Imanol , Isasti Amaia , López De Jesús Maider , Zuazo-Ibarra Jone , Montilla Alejandro , Domercq María , Capetillo-Zarate Estibaliz , García del Caño Gontzal , Sallés Joan , Matute Carlos , Sánchez Mikel
Published: February 2021

Platelet-rich plasma (PRP) is a biologic therapy that promotes healing responses across multiple medical fields, including the central nervous system (CNS). The efficacy of this therapy depends on several factors such as the donor’s health status and age. This work aims to prove the effect of PRP on cellular models of the CNS, considering the differences between PRP from young and elderly donors. Two different PRP pools were prepared from donors 65–85 and 20–25 years old. The cellular and molecular composition of both PRPs were analyzed. Subsequently, the cellular response was evaluated in CNS in vitro models, studying proliferation, neurogenesis, synaptogenesis, and inflammation. While no differences in the cellular composition of PRPs were found, the molecular composition of the Young PRP showed lower levels of inflammatory molecules such as CCL-11, as well as the presence of other factors not found in Aged PRP (GDF-11). Although both PRPs had effects in terms of reducing neural progenitor cell apoptosis, stabilizing neuronal synapses, and decreasing inflammation in the microglia, the effect of the Young PRP was more pronounced. In conclusion, the molecular composition of the PRP, conditioned by the age of the donors, affects the magnitude of the biological response.
A Narrative Review of Genetic Alterations in Primary Thyroid Epithelial Cancer
Authors: Romei Cristina , Elisei Rossella
Published: February 2021

Thyroid carcinoma is the most frequent endocrine neoplasia. Different types of thyroid carcinoma are described: well-differentiated papillary thyroid carcinoma (PTC), poorly differentiated thyroid carcinoma (PDTC), follicular thyroid carcinoma (FTC), anaplastic thyroid carcinoma (ATC), and medullary thyroid carcinoma (MTC). MTC is inherited as an autosomal dominant trait in 25% of cases. The genetic landscape of thyroid carcinoma has been largely deciphered. In PTC, genetic alterations have been found in about 95% of tumors: BRAF mutations and RET rearrangements are the main genetic alterations. BRAF and RAS mutations have been confirmed to play an important role also in PDTC and ATC, together with TP53 mutations that are fundamental in tumor progression. It has also been clearly demonstrated that telomerase reverse transcriptase (TERT) promoter mutations and TP53 mutations are present with a high-frequency in more advanced tumors, frequently associated with other mutations, and their presence, especially if simultaneous, is a signature of aggressiveness. In MTC, next-generation sequencing confirmed that mutations in the RET gene are the most common molecular events followed by H-RAS and K-RAS mutations. The comprehensive knowledge of the genetic events responsible for thyroid tumorigenesis is important to better predict the biological behavior and better plan the therapeutic strategy for specific treatment of the malignancy based on its molecular profile.
The Role of Low Complexity Regions in Protein Interaction Modes: An Illustration in Huntingtin
Authors: Kastano Kristina , Mier Pablo , Andrade-Navarro Miguel A.
Published: February 2021

Low complexity regions (LCRs) are very frequent in protein sequences, generally having a lower propensity to form structured domains and tending to be much less evolutionarily conserved than globular domains. Their higher abundance in eukaryotes and in species with more cellular types agrees with a growing number of reports on their function in protein interactions regulated by post-translational modifications. LCRs facilitate the increase of regulatory and network complexity required with the emergence of organisms with more complex tissue distribution and development. Although the low conservation and structural flexibility of LCRs complicate their study, evolutionary studies of proteins across species have been used to evaluate their significance and function. To investigate how to apply this evolutionary approach to the study of LCR function in protein–protein interactions, we performed a detailed analysis for Huntingtin (HTT), a large protein that is a hub for interaction with hundreds of proteins, has a variety of LCRs, and for which partial structural information (in complex with HAP40) is available. We hypothesize that proteins RASA1, SYN2, and KAT2B may compete with HAP40 for their attachment to the core of HTT using similar LCRs. Our results illustrate how evolution might favor the interplay of LCRs with domains, and the possibility of detecting multiple modes of LCR-mediated protein–protein interactions with a large hub such as HTT when enough protein interaction data is available.
Treatment of Oxidative Stress with Exosomes in Myocardial Ischemia
Authors: Liu Yun , Wang Mengxue , Liang Yin , Wang Chen , Naruse Keiji , Takahashi Ken
Published: February 2021

A thrombus in a coronary artery causes ischemia, which eventually leads to myocardial infarction (MI) if not removed. However, removal generates reactive oxygen species (ROS), which causes ischemia–reperfusion (I/R) injury that damages the tissue and exacerbates the resulting MI. The mechanism of I/R injury is currently extensively understood. However, supplementation of exogenous antioxidants is ineffective against oxidative stress (OS). Enhancing the ability of endogenous antioxidants may be a more effective way to treat OS, and exosomes may play a role as targeted carriers. Exosomes are nanosized vesicles wrapped in biofilms which contain various complex RNAs and proteins. They are important intermediate carriers of intercellular communication and material exchange. In recent years, diagnosis and treatment with exosomes in cardiovascular diseases have gained considerable attention. Herein, we review the new findings of exosomes in the regulation of OS in coronary heart disease, discuss the possibility of exosomes as carriers for the targeted regulation of endogenous ROS generation, and compare the advantages of exosome therapy with those of stem-cell therapy. Finally, we explore several miRNAs found in exosomes against OS.
The Role of the Thalamus in Post-Traumatic Stress Disorder
Authors: Yoshii Takanobu
Published: February 2021

Post-traumatic stress disorder (PTSD) has a high lifetime prevalence and is one of the more serious challenges in mental health care. Fear-conditioned learning involving the amygdala has been thought to be one of the main causative factors; however, recent studies have reported abnormalities in the thalamus of PTSD patients, which may explain the mechanism of interventions such as eye movement desensitization and reprocessing (EMDR). Therefore, I conducted a miniature literature review on the potential contribution of the thalamus to the pathogenesis of PTSD and the validation of therapeutic approaches. As a result, we noticed the importance of the retinotectal pathway (superior colliculus−pulvinar−amygdala connection) and discussed therapeutic indicators.
Development of Atropa belladonna L. Plants with High-Yield Hyoscyamine and without Its Derivatives Using the CRISPR/Cas9 System
Authors: Zeng Lingjiang , Zhang Qiaozhuo , Jiang Chunxue , Zheng Yueyue , Zuo Youwei , Qin Jianbo , Liao Zhihua , Deng Hongping
Published: February 2021

Atropa belladonna L. is one of the most important herbal plants that produces hyoscyamine or atropine, and it also produces anisodamine and scopolamine. However, the in planta hyoscyamine content is very low, and it is difficult and expensive to independently separate hyoscyamine from the tropane alkaloids in A. belladonna. Therefore, it is vital to develop A. belladonna plants with high yields of hyoscyamine, and without anisodamine and scopolamine. In this study, we generated A. belladonna plants without anisodamine and scopolamine, via the CRISPR/Cas9-based disruption of hyoscyamine 6β-hydroxylase (AbH6H), for the first time. Hyoscyamine production was significantly elevated, while neither anisodamine nor scopolamine were produced, in the A. belladonna plants with homozygous mutations in AbH6H. In summary, new varieties of A. belladonna with high yields of hyoscyamine and without anisodamine and scopolamine have great potential applicability in producing hyoscyamine at a low cost.
Maternal Low-Grade Chronic Inflammation and Intrauterine Programming of Health and Disease
Authors: Parisi Francesca , Milazzo Roberta , Savasi Valeria M. , Cetin Irene
Published: February 2021

Overweight and obesity during pregnancy have been associated with increased birth weight, childhood obesity, and noncommunicable diseases in the offspring, leading to a vicious transgenerational perpetuating of metabolic derangements. Key components in intrauterine developmental programming still remain to be identified. Obesity involves chronic low-grade systemic inflammation that, in addition to physiological adaptations to pregnancy, may potentially expand to the placental interface and lead to intrauterine derangements with a threshold effect. Animal models, where maternal inflammation is mimicked by single injections with lipopolysaccharide (LPS) resembling the obesity-induced immune profile, showed increased adiposity and impaired metabolic homeostasis in the offspring, similar to the phenotype observed after exposure to maternal obesity. Cytokine levels might be specifically important for the metabolic imprinting, as cytokines are transferable from maternal to fetal circulation and have the capability to modulate placental nutrient transfer. Maternal inflammation may induce metabolic reprogramming at several levels, starting from the periconceptional period with effects on the oocyte going through early stages of embryonic and placental development. Given the potential to reduce inflammation through inexpensive, widely available therapies, examinations of the impact of chronic inflammation on reproductive and pregnancy outcomes, as well as preventive interventions, are now needed.
Voluntary Wheel Running Partially Compensates for the Effects of Global Estrogen Receptor-α Knockout on Cortical Bone in Young Male Mice
Authors: Dirkes Rebecca K. , Winn Nathan C. , Jurrissen Thomas J. , Lubahn Dennis B. , Vieira-Potter Victoria J. , Padilla Jaume , Hinton Pamela S.
Published: February 2021

Estrogen receptor-α knockout (ERKO) in female, but not male, mice results in an impaired osteogenic response to exercise, but the mechanisms behind this ability in males are unknown. We explored the main and interactive effects of ERKO and exercise on cortical geometry, trabecular microarchitecture, biomechanical strength, and sclerostin expression in male mice. At 12 weeks of age, male C57BL/6J ERKO and WT animals were randomized into two groups: exercise treatment (EX) and sedentary (SED) controls, until 22 weeks of age. Cortical geometry and trabecular microarchitecture were measured via μCT; biomechanical strength was assessed via three-point bending; sclerostin expression was measured via immunohistochemistry. Two-way ANOVA was used to assess sclerostin expression and trabecular microarchitecture; two-way ANCOVA with body weight was used to assess cortical geometry and biomechanical strength. ERKO positively impacted trabecular microarchitecture, and exercise had little effect on these outcomes. ERKO significantly impaired cortical geometry, but exercise was able to partially reverse these negative alterations. EX increased cortical thickness regardless of genotype. There were no effects of genotype or exercise on sclerostin expression. In conclusion, male ERKO mice retain the ability to build bone in response to exercise, but altering sclerostin expression is not one of the mechanisms involved.
Understanding Viral Infection Mechanisms and Patient Symptoms for the Development of COVID-19 Therapeutics
Authors: Choi Hyung Muk , Moon Soo Youn , Yang Hyung In , Kim Kyoung Soo
Published: February 2021

Coronavirus disease 2019 (COVID-19), caused by the SARS-CoV-2 virus, has become a worldwide pandemic. Symptoms range from mild fever to cough, fatigue, severe pneumonia, acute respiratory distress syndrome (ARDS), and organ failure, with a mortality rate of 2.2%. However, there are no licensed drugs or definitive treatment strategies for patients with severe COVID-19. Only antiviral or anti-inflammatory drugs are used as symptomatic treatments based on clinician experience. Basic medical researchers are also trying to develop COVID-19 therapeutics. However, there is limited systematic information about the pathogenesis of COVID-19 symptoms that cause tissue damage or death and the mechanisms by which the virus infects and replicates in cells. Here, we introduce recent knowledge of time course changes in viral titers, delayed virus clearance, and persistent systemic inflammation in patients with severe COVID-19. Based on the concept of drug reposition, we review which antiviral or anti-inflammatory drugs can effectively treat COVID-19 patients based on progressive symptoms and the mechanisms inhibiting virus infection and replication.
Overview of MMP-13 as a Promising Target for the Treatment of Osteoarthritis
Authors: Hu Qichan , Ecker Melanie
Published: February 2021

Osteoarthritis (OA) is a common degenerative disease characterized by the destruction of articular cartilage and chronic inflammation of surrounding tissues. Matrix metalloproteinase-13 (MMP-13) is the primary MMP involved in cartilage degradation through its particular ability to cleave type II collagen. Hence, it is an attractive target for the treatment of OA. However, the detailed molecular mechanisms of OA initiation and progression remain elusive, and, currently, there are no interventions available to restore degraded cartilage. This review fully illustrates the involvement of MMP-13 in the initiation and progression of OA through the regulation of MMP-13 activity at the molecular and epigenetic levels, as well as the strategies that have been employed against MMP-13. The aim of this review is to identify MMP-13 as an attractive target for inhibitor development in the treatment of OA.
Disabled 1 Is Part of a Signaling Pathway Activated by Epidermal Growth Factor Receptor
Authors: Dlugosz Paula , Teufl Magdalena , Schwab Maximilian , Kohl Katharina Eva , Nimpf Johannes
Published: February 2021

Disabled 1 (Dab1) is an adapter protein for very low density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2) and an integral component of the Reelin pathway which orchestrates neuronal layering during embryonic brain development. Activation of Dab1 is induced by binding of Reelin to ApoER2 and VLDLR and phosphorylation of Dab1 mediated by Src family kinases. Here we show that Dab1 also acts as an adaptor for epidermal growth factor receptor (EGFR) and can be phosphorylated by epidermal growth factor (EGF) binding to EGFR. Phosphorylation of Dab1 depends on the kinase activity of EGFR constituting a signal pathway independent of Reelin and its receptors.
In Vitro and In Vivo Efficacy of a Novel Glucose–Methotrexate Conjugate in Targeted Cancer Treatment
Authors: Woźniak Marta , Pastuch-Gawołek Gabriela , Makuch Sebastian , Wiśniewski Jerzy , Krenács Tibor , Hamar Peter , Gamian Andrzej , Szeja Wiesław , Szkudlarek Danuta , Krawczyk Monika , Agrawal Siddarth
Published: February 2021

Methotrexate (MTX) is a commonly used antimetabolite, which inhibits folate and DNA synthesis to be effective in the treatment of various malignancies. However, MTX therapy is hindered by the lack of target tumor selectivity. We have designed, synthesized and evaluated a novel glucose–methotrexate conjugate (GLU–MTX) both in vitro and in vivo, in which a cleavable linkage allows intracellular MTX release after selective uptake through glucose transporter−1 (GLUT1). GLU–MTX inhibited the growth of colorectal (DLD-1), breast (MCF-7) and lung (A427) adenocarcinomas, squamous cell carcinoma (SCC-25), osteosarcoma (MG63) cell lines, but not in WI-38 healthy fibroblasts. In tumor cells, GLU–MTX uptake increased 17-fold compared to unconjugated MTX. 4,6-O-ethylidene-α-D-glucose (EDG), a GLUT1 inhibitor, significantly interfered with GLU–MTX induced growth inhibition, suggesting a glucose-mediated drug uptake. Glu-MTX also caused significant tumor growth delay in vivo in breast cancer-bearing mice. These results show that our GLUT-MTX conjugate can be selectively uptake by a range of tumor cells to cause their significant growth inhibition in vitro, which was also confirmed in a breast cancer model in vivo. GLUT1 inhibitor EDG interfered with these effects verifying the selective drug uptake. Accordingly, GLU–MTX offers a considerable tumor selectivity and may offer cancer growth inhibition at reduced toxicity.
UAS-Based Hyperspectral Environmental Monitoring of Acid Mine Drainage Affected Waters
Authors: Flores Hernan , Lorenz Sandra , Jackisch Robert , Tusa Laura , Contreras I. , Zimmermann Robert , Gloaguen Richard
Published: February 2021

The exposure of metal sulfides to air or water, either produced naturally or due to mining activities, can result in environmentally damaging acid mine drainage (AMD). This needs to be accurately monitored and remediated. In this study, we apply high-resolution unmanned aerial system (UAS)-based hyperspectral mapping tools to provide a useful, fast, and non-invasive method for the monitoring aspect. Specifically, we propose a machine learning framework to integrate visible to near-infrared (VNIR) hyperspectral data with physicochemical field data from water and sediments, together with laboratory analyses to precisely map the extent of acid mine drainage in the Tintillo River (Spain). This river collects the drainage from the western part of the Rio Tinto massive sulfide deposit and discharges large quantities of acidic water with significant amounts of dissolved metals (Fe, Al, Cu, Zn, amongst others) into the Odiel River. At the confluence of these rivers, different geochemical and mineralogical processes occur due to the interaction of very acidic water (pH 2.5–3.0) with neutral water (pH 7.0–8.0). This complexity makes the area an ideal test site for the application of hyperspectral mapping to characterize both rivers and better evaluate contaminated water bodies with remote sensing imagery. Our approach makes use of a supervised random forest (RF) regression for the extended mapping of water properties, using the samples collected in the field as ground-truth and training data. The resulting maps successfully estimate the concentration of dissolved metals and related physicochemical properties in water, and trace associated iron species (e.g., jarosite, goethite) within sediments. These results highlight the capabilities of UAS-based hyperspectral data to monitor water bodies in mining environments, by mapping their hydrogeochemical properties, using few field samples. Hence, we have demonstrated that our workflow allows the rapid discrimination and mapping of AMD contamination in water, providing an essential basis for monitoring and subsequent remediation.
The Effect of Myristica fragrans on Texture Properties and Shelf-Life of Innovative Chewable Gel Tablets
Authors: Matulyte Inga , Mataraite Akvile , Velziene Saule , Bernatoniene Jurga
Published: February 2021

Chewable gel tablets are an underdeveloped subject, even though there are many simple chewable tablets and gummy candies in the food and pharmaceutical industries. Chewable gel tablets are not as sweet, they can have an active substance, pharmacological effect, and a value of nutrition. The aim of this study was to prepare gelatin-based chewable tablets with Myristica fragrans as a preservative and to determine the shelf-life variability depending on storage conditions, and to evaluate texture changes. Firmness and springiness of gel tablets were measured by a texture analyzer and compared between different storage conditions and the shelf-life of tablets was established by mold growing time. Chewable gel tablets were prepared by using silicone form. Mold was most likely to grow on tablets that have been packaged in squeezable bags (after 14 days 60% of all formulations had a mold, p < 0.05). The most stable tablets (over 180 days) were in sealed boxes and contained nutmeg essential oil or its solution, or ethanolic nutmeg extract. The gel tablets’ firmness increased about 4 times when they were stored in opened plastic boxes and their springiness decreased about 1.65 times after 28 days in the mentioned conditions, p < 0.05. Nutmeg hydrolat had the highest influence on texture variation (p < 0.05).
Prevalence and Clinical Significance of Drug–Drug and Drug–Dietary Supplement Interactions among Patients Admitted for Cardiothoracic Surgery in Greece
Authors: Spanakis Marios , Melissourgaki Maria , Lazopoulos George , Patelarou Athina E. , Patelarou Evridiki
Published: February 2021

Background: Drug interactions represent a major issue in clinical settings, especially for critically ill patients such as those with cardiovascular disease (CVD) who require cardiothoracic surgery (CTS) and receive a high number of different medications. Methods: A cross-sectional study aimed at evaluating the exposure and clinical significance of drug–drug (DDIs) and drug–dietary supplement interactions (DDSIs) in patients admitted for CTS in the University Hospital of Crete Greece. DDIs were evaluated regarding underlying pharmacological mechanisms upon admission, preoperation, postoperation, and discharge from CTS clinic. Additionally, upon admission, the use of dietary supplements (DSs) and if patients had informed their treating physician that they were using these were recorded with subsequent analysis of potential DDSIs with prescribed medications. Results: The study employed 76 patients who were admitted for CTS and accepted to participate. Overall, 166 unique DDIs were identified, with 32% of them being related to pharmacokinetic (PK) processes and the rest (68%) were related to possible alterations of pharmacodynamic (PD) action. CVD medications and drugs for central nervous system disorders were the most frequently interacting medications. In total, 12% of the identified DDIs were of serious clinical significance. The frequency of PK-DDIs was higher during admission and discharge, whereas PD-DDIs were mainly recorded during pre- and postoperation periods. Regarding DS usage, 60% of patients were using DSs and perceived them as safe, and the majority had not informed their treating physician of this or sought out medical advice. Analysis of medical records showed 30 potential combinations with prescribed medications that could lead in DDSIs due to modulation of PK or PD processes, and grapefruit juice consumption was involved in 38% of them. Conclusions: An increased burden of DDIs and DDSIs was identified mostly upon admission for patients in CTS clinics in Greece. Healthcare providers, especially prescribing physicians in Greece, should always take into consideration the possibility of DDIs and the likely use of DS products by patients to promote their well-being; this should only be undertaken after receiving medical advice and an evidenced-based evaluation.
Conjugation of Mannans to Enhance the Potency of Liposome Nanoparticles for the Delivery of RNA Vaccines
Authors: Goswami Roshan , O’Hagan Derek T. , Adamo Roberto , Baudner Barbara C.
Published: February 2021

Recent approval of mRNA vaccines to combat COVID-19 have highlighted the potential of this platform. Lipid nanoparticles (LNP) is the delivery vehicle of choice for mRNA as they prevent its enzymatic degradation by encapsulation. We have recently shown that surface exposition of mannose, incorporated in LNPs as stable cholesterol-amine conjugate, enhances the potency of self-amplifying RNA (SAM) replicon vaccines through augmented uptake by antigen presenting cells (APCs). Here, we generated a new set of LNPs whose surface was modified with mannans of different length (from mono to tetrasaccharide), in order to study the effect on antibody response of model SAM replicon encoding for the respiratory syncytial virus fusion F protein. Furthermore, the impact of the mannosylated liposomal delivery through intradermal as well as intramuscular routes was investigated. The vaccine priming response showed to improve consistently with increase in the chain length of mannoses; however, the booster dose response plateaued above the length of disaccharide. An increase in levels of IgG1 and IgG2a was observed for mannnosylated lipid nanoparticles (MLNPs) as compared to LNPs. This work confirms the potential of mannosylated SAM LNPs for both intramuscular and intradermal delivery, and highlights a disaccharide length as sufficient to ensure improved immunogenicity compared to the un-glycosylated delivery system.
Evaluation of Binders in Twin-Screw Wet Granulation
Authors: Köster Claudia , Pohl Sebastian , Kleinebudde Peter
Published: February 2021

The binders povidone (Kollidon 30), copovidone (Kollidon VA64), hypromellose (Pharmacoat 606), and three types of hyprolose (HPC SSL-SFP, HPC SSL, and HPC SL-FP) were evaluated regarding their suitability in twin-screw wet granulation. Six mixtures of lactose and binder as well as lactose without binder were twin-screw granulated with demineralized water at different barrel fill levels and subsequently tableted. A screening run with HPC SSL determined the amount of water as an influential parameter for oversized agglomerates. Subsequent examination of different binders, especially Kollidon 30 and Kollidon VA64 resulted in large granules. All binders, except Pharmacoat 606, led to a reduction of fines compared to granulation without a binder. The molecular weight of applied hyproloses did not appear as influential. Tableting required an upstream sieving step to remove overlarge granules. Tableting was possible for all formulations at sufficient compression pressure. Most binders resulted in comparable tensile strengths, while Pharmacoat 606 led to lower and lactose without a binder to the lowest tensile strength. Tablets without a binder disintegrated easily, whereas binder containing tablets of sufficient tensile strength often nearly failed or failed the disintegration test. Especially tablets containing Pharmacoat 606 and HPC SL-FP disintegrated too slowly.
Physiological and Biochemical Responses of Ungrafted and Grafted Bell Pepper Plants (Capsicum annuum L. var. grossum (L.) Sendtn.) Grown under Moderate Salt Stress
Authors: Kacjan Maršić Nina , Štolfa Petra , Vodnik Dominik , Košmelj Katarina , Mikulič-Petkovšek Maja , Kump Bojka , Vidrih Rajko , Kokalj Doris , Piskernik Saša , Ferjančič Blaz , Dragutinović Maja , Veberič Robert , Hudina Metka , Šircelj Helena
Published: February 2021

The response of grafted bell pepper plants (Capsicum annuum L. var. grossum (L.) Sendtn.) to salt stress was investigated by analyzing the photosynthetic traits and mineral content of the plants and the metabolic composition of the fruit. The bell pepper variety “Vedrana” was grafted onto the salt-tolerant rootstock “Rocal F1” and grown at two salinities (20 mM and 40 mM NaCl) and control (0 mM NaCl) during the spring–summer period. On a physiological level, similar stomatal restriction of photosynthesis in grafted and ungrafted plants indicated that grafting did not alleviate water balance disturbances under increased salt exposure. Measurements of midday water potential did not show improved water status of grafted plants. The similar metabolic changes in grafted and ungrafted plants were also reflected in similarly reduced fruit yields. Thus, this grafting did not reduce the risk of ionic and osmotic imbalance in pepper plants grown under moderate salt treatment. Changes in the biochemical profiles of the pepper fruit were seen for both added-salt treatments. The fruit phenolic compounds were affected by rootstock mediation, although only for the July harvest, where total phenolics content increased with 40 mM NaCl treatment. Fruit ascorbic acid content increased with the duration of salt stress, without the mediation of the rootstock. The high salt dependence of this quality trait in pepper fruit appears to lead to more limited rootstock mediation effects.