bluuebunny/crossref_metadata_2025 huggingface dataset:
Treatment with Supercritical CO2 Reduces Off-Flavour of White Alfalfa Protein Concentrate
Authors: Hansen Mikkel , Hobley Timothy John , Jensen Peter Ruhdal
Published: February 2023

White alfalfa protein concentrate from alfalfa (Medicago sativa) is a promising substitute for milk and egg protein due to its functionality. However, it contains many unwanted flavours that limits the amount that can be added to a food without affecting its taste negatively. In this paper, we have demonstrated a simple method for the extraction of white alfalfa protein concentrate followed by a treatment with supercritical CO2. Two concentrates were produced at lab scale and pilot scale, with yields of 0.012 g (lab scale) and 0.08 g (pilot scale), of protein per g of total protein introduced into the process. The solubility of the protein produced at lab scale and pilot scale was approximately 30% and 15%, respectively. By treating the protein concentrate at 220 bar and 45 °C for 75 min with supercritical CO2, off-flavours were lowered. The treatment did not decrease the digestibility or alter the functionality of white alfalfa protein concentrate when it was used to substitute egg in chocolate muffins and egg white in meringues.
Effects of Ultra-High Pressure Synergistic Enzymatic Hydrolysis on Flavor of Stropharia rugoso-annulata
Authors: Bao Chenligen , Xin Minghang , Su Keyu , Guan Chunbo , Wang Dawei
Published: February 2023

In this study, using gas chromatography-mass spectrometry (HS-SPME-GC-MS), electronic nose (E-nose), high performance liquid chromatography (HPLC), and electronic tongue (E-tongue) to analyze the effect of ultra-high pressure (UHP) synergistic enzymatic hydrolysis on the flavor compounds of enzymatic hydrolysates of S. rugoso-annulata. The results demonstrated that 38 volatile flavor substances were identified in the enzymatic hydrolysates of S. rugoso-annulata treated at atmospheric pressure and 100, 200, 300, 400, and 500 MPa, mainly 6 esters, 4 aldehydes, 10 alcohols, 5 acids, and 13 other volatile flavor substances, and the most kinds of flavor substances reached 32 kinds when the pressure was 400 MPa. E-nose can effectively distinguish the overall changes of enzymatic hydrolysates of S. rugoso-annulata treated with atmospheric pressure and different pressures. There was 1.09 times more umami amino acids in the enzymatic hydrolysates at 400 MPa than in the atmospheric pressure enzymatic hydrolysates and 1.11 times more sweet amino acids at 500 MPa than in the atmospheric pressure enzymatic hydrolysates. The results of the E-tongue indicate that the UHP treatment increased umami and sweetness and reduced bitterness, which was also confirmed by the results of amino acid and 5′-nucleotide analysis. In conclusion, the UHP synergistic enzymatic hydrolysis can effectively improve the overall flavor of the enzymatic hydrolysates of S. rugoso-annulata; this study also lays the theoretical foundation for the deep processing and comprehensive utilization of S. rugoso-annulata.
Evaluation of Vaporizing Diesel Spray with High-Speed Laser Absorption Scattering Technique for Measuring Vapor and Liquid Phase Concentration Distributions
Authors: Ray Samir Chandra , Safiullah  , Naito Shinichiro , Andersson Mats , Nishida Keiya , Ogata Yoichi
Published: February 2023

The Conventional Laser Absorption Scattering (C-LAS) technique is used to measure the mixture concentration and visualize the vapor phase. The former is determined by the attenuation of visible and ultraviolet light whereas the latter is achieved via light absorption and scattering theory. The C-LAS uses the Nd: YAG pulsed laser and CCD cameras to provide one spray shot at a particular instance which requires time and effort. However, the temporal measurement of a single spray shot is not possible. To record the distribution of the whole vapor phase in an injection event and measure liquid and vapor concentrations inside the spray, a High-Speed Laser Absorption Scattering (HS-LAS) technique was developed. The HS-LAS consists of continuous diode light sources, high-speed video cameras, and an image intensifier for UV light, which can provide the temporal variation of a single-shot spray. In the experiment, a commercial seven-hole injector with a hole diameter of 0.123 mm allowing high injection pressure of up to 100 MPa was used to avoid the potential inconsistencies with a single-hole test injector. The diesel surrogate fuel which consists of 97.5% n-tridecane and 2.5% of volume-based 1-methylnaphthalene was used. The injection amount of 5.0 mg/hole was selected to investigate the structure and mixture formation process of the spray. The findings of the experiments show that this imaging approach is a promising diagnostic technique for concurrently obtaining quantitative information on the quantity of vapor and droplets in a fuel spray. Furthermore, the turbulent/vortex fluid dynamics’ temporal development/variation can be investigated.
Fear God, Not COVID-19: Is Conservative Protestantism Associated with Risky Pandemic Lifestyles?
Authors: Bartkowski John P. , Klee Katherine , Hill Terrence D. , Garcia-Alexander Ginny , Ellison Christopher G. , Burdette Amy M.
Published: February 2023

Previous research has established attitudinal and behavioral health variations in relation to the COVID-19 pandemic, but scholarship on the religious antecedents associated with these outcomes has only recently gained momentum. Rhetoric from some leading conservative Protestants in the U.S. has underplayed the threat of the pandemic and may have contributed to unhealthy pandemic behaviors within this faith tradition. Moreover, previous inquiries have revealed that conservative Protestantism’s otherworldly focus can thwart personal and community health. We use nationally representative data to test the hypotheses that, compared with other religious groups and the non-religious, conservative Protestants will tend to (1) perceive the pandemic as less threatening and (2) engage in riskier pandemic lifestyles. These hypotheses are generally supported net of confounding factors. We conclude that affiliation with a conservative Protestant denomination can undermine public health among this faith tradition’s adherents and may therefore compromise general health and well-being during a pandemic. We discuss the implications of these findings, offer recommendations for pandemic health promotion among conservative Protestants, and delineate promising avenues for future research on this important topic.
Exploring the Impact of Storytelling for Hospitalized Patients Recovering from COVID-19
Authors: Gurney Lara , Chung Vincci , MacPhee Maura , Chan Evelyn , Snyman Claire , Robinson Jaclyn , Bertoli-Haley Serena , Baron Elizabeth
Published: February 2023

There are mental and physical deficits associated with COVID-19 infection, particularly among individuals requiring hospitalization. Storytelling is a relational intervention that has been used to help patients make sense of their illness experiences and to share their experiences with others, including other patients, families and healthcare providers. Relational interventions strive to create positive, healing stories versus negative ones. In one urban acute care hospital, an initiative called the Patient Stories Project (PSP) uses storytelling as a relational intervention to promote patient healing, including the development of healthier relationships among themselves, with families and with healthcare providers. This qualitative study employed a series of interview questions that were collaboratively developed with patient partners and COVID-19 survivors. The questions asked consenting COVID-19 survivors about why they chose to tell their stories and to flesh out more about their recovery process. Thematic analyses of six participant interviews resulted in the identification of key themes along a COVID-19 recovery pathway. Patients’ stories revealed how survivors progress from being overwhelmed by their symptoms to making sense of what is happening to them, providing feedback to their care providers, feeling gratitude for care received, becoming aware of a new state of normal, regaining control of their lives, and ultimately discovering meaning and an important lesson behind their illness experience. Our study’s findings suggest that the PSP storytelling approach holds potential as a relational intervention to support COVID-19 survivors along a recovery journey. This study also adds knowledge about survivors beyond the first few months of recovery.
Making London Porcelain—A Multidisciplinary Project Connecting Local Communities with the Technological and Innovation Histories of London’s Early Porcelain Manufacturers
Authors: Burgio Lucia , Domoney Kelly , Haseldine Georgia , McCaffrey-Howarth Caroline
Published: February 2023

This collaborative multidisciplinary pilot project involving the Victoria and Albert Museum (V&A), the Ashmolean Museum, and Newham Borough of London, examined the composition of a selection of eighteenth-century porcelain objects by two of London’s first porcelain manufacturers, Bow and Chelsea. As the first science-based public engagement project to be piloted by the V&A, it succeeded in bringing together young Londoners and their communities to investigate local histories of scientific and artistic innovation through the analysis and remaking of eighteenth-century porcelain. Scientific object analysis informed activities with local sixth-form students, revealing the intimate link between art and science, and showcasing the V&A Science Lab as a national hub for heritage science. Public outreach activities, including an exhibition at Stratford Library and workshops for Newham Heritage Month also provided hands-on learning, including curatorial and object-handling experience, and the embodied practices of remaking. Ultimately, this project stimulated new ways of engaging with ceramics collections and explored how the creativity and ingenuity of eighteenth-century ceramics pioneers can provide inspiration for the next generation of makers.
Datasets for Material Culture Studies: A Protocol for the Systematic Compilation of Items Held in Private Hands
Authors: Spennemann Dirk H. R.
Published: February 2023

Crowdsourcing has been widely used in cultural heritage research, but mainly from an institutional perspective. Research into items of material culture often requires the researcher to examine specimens held in private hands. The dispersed nature of such holdings, primarily as collectable and, thus, tradeable objects, requires different techniques for identification and access than that for items held in museum collections. Crowdsourcing data from online discussion groups and online marketplaces, coupled with snowballing, represents a very powerful tool. This paper discusses the relevant parameters and provides a protocol for the systematic compilation of items held in private hands.
Inside the Scriptorium: Non-Invasive In Situ Identification of Dyes in Illuminated Manuscripts by Microspectrofluorimetry and Multivariate Analysis: The Crescenzago Choir Books (End 15th Century) as a Case Study
Authors: Longoni Margherita , Hopes Noemi , Rossi Federico , Tessera Miriam Rita , Petoletti Marco , Bruni Silvia
Published: February 2023

In the present work, a method is proposed that combines spectrofluorimetry with multivariate analysis (cluster and principal components) to non-invasively investigate natural dyes used, mainly in the form of lakes, in illuminations. First, a consistent number of mock-up samples of red and purple lakes (from brazilwood, madder, kermes, cochineal, lac dye, folium and orcein) were prepared following the recipes reported in ancient treatises, then the experimental conditions for their non-invasive investigation were optimized by benchtop instrumentation and transferred to a portable microprobe suitable for in situ analyses. For all the coloring materials examined, it was thus possible to obtain a satisfactory grouping of samples containing the same dye. The analytical protocol was finally extended to the characterization of dyes in the Crescenzago choir books, illuminated manuscripts dating back to the end of the 15th century.
Rapid and Efficient Regeneration of Rhododendron decorum from Flower Buds
Authors: Wu Hairong , Ao Qian , Li Huie , Long Fenfang
Published: February 2023

Rhododendron decorum is a woody species with high ornamental and medical value. Herein, we introduce a novel in vitro regeneration method for R. decorum. We used flower buds to develop an efficient and rapid plant regeneration protocol. Sterile flower buds of R. decorum of a 2 cm size were used as explants to study the effects of the culture medium and plant growth regulators on the callus induction and adventitious shoot differentiation, proliferation, and rooting. According to the results, the optimal medium combination for callus induction was WPM + 1 mg/L TDZ + 0.2 mg/L NAA, and its induction rate reached 95.08%. The optimal medium combination for adventitious shoot differentiation from the callus was WPM + 0.5 mg/L TDZ + 0.1 mg/L NAA, and its differentiation rate reached 91.32%. The optimal medium combination for adventitious shoot proliferation was WPM + 2 mg/L ZT + 0.5 mg/L NAA, for which the proliferation rate reached 95.32% and the proliferation coefficient reached 9.45. The optimal medium combination for rooting from adventitious shoots was WPM + 0.1 mg/L NAA + 1 mg/L IBA, and its rooting rate reached 86.90%. The survival rates of the rooted regenerated plantlets exceeded 90% after acclimatization and transplantation. This regeneration system has the advantages of being simple and highly efficient, and it causes little damage to the shoots of the mother plants, laying a foundation for the plantlet propagation, genetic transformation, and new-variety breeding of R. decorum.
The Coordination of Environmental Protection and Female Discrimination Based on the Concept of Affirmative Action
Authors: Ling Xia , Liu Yanhong
Published: February 2023

With the development of society, the chemical industry is expanding, and the hazy weather everywhere is becoming increasingly frequent, already affecting people’s lives and causing them to pay more attention to environmental issues. Therefore, this paper highlights the role of women in environmental protection by studying the coordination of environmental protection and female discrimination based on the concept of affirmative action. Through this study and a survey, we found that China has not yet realized that women’s participation in environmental protection plays a key role in improving the quality of our environment and the development of ecological civilization. However, we should clearly understand that environmental issues are not only personal, they are related to the survival and development of a country, and as a member of that country, both women and men should have the right and obligation to protect the environment. Therefore, this article discusses the concept and meaning of affirmative action and gender discrimination in the context of research on these concepts, discussing the problems and phenomena that women encounter in environmental protection. These include the system of women’s environmental protection, gender issues for women in society, and the unequal treatment from the Government based on some studies. Through the study and analysis of the system of women’s environmental protection, the role and position of women in this regard is summarized. Finally, it is suggested that, for the construction and development of ecological civilization in China, it is necessary to fully integrate ecological civilization into all aspects of society and pay attention to environmental protection issues. Therefore, we should pay attention to the role of women in environmental protection, provide corresponding policies, and actively encourage women to partake in environmental protection in order to build an environmentally friendly and resource-saving society together.
Quantification of Xanthone and Anthocyanin in Mangosteen Peel by UPLC-MS/MS and Preparation of Nanoemulsions for Studying Their Inhibition Effects on Liver Cancer Cells
Authors: Li Rui , Inbaraj Baskaran Stephen , Chen Bing-Huei
Published: February 2023

Mangosteen peel, a waste produced during mangosteen processing, has been reported to be rich in xanthone and anthocyanin, both of which possess vital biological activities such as anti-cancer properties. The objectives of this study were to analyze various xanthones and anthocyanins in mangosteen peel by UPLC-MS/MS for the subsequent preparation of both xanthone and anthocyanin nanoemulsions to study their inhibition effects on liver cancer cells HepG2. Results showed that methanol was the optimal solvent for the extraction of xanthones and anthocyanins, with a total amount of 68,543.39 and 2909.57 μg/g, respectively. A total of seven xanthones, including garcinone C (513.06 μg/g), garcinone D (469.82 μg/g), γ-mangostin (11,100.72 μg/g), 8-desoxygartanin (1490.61 μg/g), gartanin (2398.96 μg/g), α-mangostin (51,062.21 μg/g) and β-mangostin (1508.01 μg/g), as well as two anthocyanins including cyanidin-3-sophoroside (2889.95 μg/g) and cyanidin-3-glucoside (19.72 μg/g), were present in mangosteen peel. The xanthone nanoemulsion was prepared by mixing an appropriate portion of soybean oil, CITREM, Tween 80 and deionized water, while the anthocyanin nanoemulsion composed of soybean oil, ethanol, PEG400, lecithin, Tween 80, glycerol and deionized water was prepared as well. The mean particle size of the xanthone extract and nanoemulsion were, respectively, 22.1 and 14.0 nm as determined by DLS, while the zeta potential was −87.7 and −61.5 mV. Comparatively, xanthone nanoemulsion was more effective than xanthone extract in inhibiting the growth of HepG2 cells, with the IC50 being 5.78 μg/mL for the former and 6.23 μg/mL for the latter. However, the anthocyanin nanoemulsion failed to inhibit growth of HepG2 cells. Cell cycle analysis revealed that the proportion of the sub-G1 phase followed a dose-dependent increase, while that of the G0/G1 phase showed a dose-dependent decline for both xanthone extracts and nanoemulsions, with the cell cycle being possibly arrested at the S phase. The proportion of late apoptosis cells also followed a dose-dependent rise for both xanthone extracts and nanoemulsions, with the latter resulting in a much higher proportion at the same dose. Similarly, the activities of caspase-3, caspase-8 and caspase-9 followed a dose-dependent increase for both xanthone extracts and nanoemulsions, with the latter exhibiting a higher activity at the same dose. Collectively, xanthone nanoemulsion was more effective than xanthone extract in inhibiting the growth of HepG2 cells. Further research is needed to study the anti-tumor effect in vivo.
Adrenomedullin Improves Hypertension and Vascular Remodeling partly through the Receptor-Mediated AMPK Pathway in Rats with Obesity-Related Hypertension
Authors: Wang Hong-Yu , Wang Fang-Zheng , Chang Rui , Wang Qian , Liu Si-Yu , Cheng Ze-Xiong , Gao Qing , Zhou Hong , Zhou Ye-Bo
Published: February 2023

Adrenomedullin (ADM) is a novel cardiovascular peptide with anti-inflammatory and antioxidant properties. Chronic inflammation, oxidative stress and calcification play pivotal roles in the pathogenesis of vascular dysfunction in obesity-related hypertension (OH). Our study aimed to explore the effects of ADM on the vascular inflammation, oxidative stress and calcification in rats with OH. Eight-week-old Sprague Dawley male rats were fed with either a Control diet or a high fat diet (HFD) for 28 weeks. Next, the OH rats were randomly subdivided into two groups as follows: (1) HFD control group, and (2) HFD with ADM. A 4-week treatment with ADM (7.2 μg/kg/day, ip) not only improved hypertension and vascular remodeling, but also inhibited vascular inflammation, oxidative stress and calcification in aorta of rats with OH. In vitro experiments, ADM (10 nM) in A7r5 cells (rat thoracic aorta smooth muscle cells) attenuated palmitic acid (PA, 200 μM) or angiotensin II (Ang II, 10 nM) alone or their combination treatment-induced inflammation, oxidative stress and calcification, which were effectively inhibited by the ADM receptor antagonist ADM22-52 and AMP-activated protein kinase (AMPK) inhibitor Compound C, respectively. Moreover, ADM treatment significantly inhibited Ang II type 1 receptor (AT1R) protein expression in aorta of rats with OH or in PA-treated A7r5 cells. ADM improved hypertension, vascular remodeling and arterial stiffness, and attenuated inflammation, oxidative stress and calcification in OH state partially via receptor-mediated AMPK pathway. The results also raise the possibility that ADM will be considered for improving hypertension and vascular damage in patients with OH.
Starch and Sucrose Metabolism and Plant Hormone Signaling Pathways Play Crucial Roles in Aquilegia Salt Stress Adaption
Authors: Chen Lifei , Meng Yuan , Bai Yun , Yu Haihang , Qian Ying , Zhang Dongyang , Zhou Yunwei
Published: February 2023

Salt stress is one of the main abiotic stresses that strongly affects plant growth. Clarifying the molecular regulatory mechanism in ornamental plants under salt stress is of great significance for the ecological development of saline soil areas. Aquilegia vulgaris is a perennial with a high ornamental and commercial value. To narrow down the key responsive pathways and regulatory genes, we analyzed the transcriptome of A. vulgaris under a 200 mM NaCl treatment. A total of 5600 differentially expressed genes were identified. The Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis pointed out that starch and sucrose metabolism and plant hormone signal transduction were significantly improved. The above pathways played crucial roles when A. vulgaris was coping with salt stress, and their protein–protein interactions (PPIs) were predicted. This research provides new insights into the molecular regulatory mechanism, which could be the theoretical basis for screening candidate genes in Aquilegia.
5-FU-miR-15a Inhibits Activation of Pancreatic Stellate Cells by Reducing YAP1 and BCL-2 Levels In Vitro
Authors: Almanzar Vanessa M. Diaz , Shah Kunal , LaComb Joseph F. , Mojumdar Aisharja , Patel Hetvi R. , Cheung Jacky , Tang Meiyi , Ju Jingfang , Bialkowska Agnieszka B.
Published: February 2023

Chronic pancreatitis is characterized by chronic inflammation and fibrosis, processes heightened by activated pancreatic stellate cells (PSCs). Recent publications have demonstrated that miR-15a, which targets YAP1 and BCL-2, is significantly downregulated in patients with chronic pancreatitis compared to healthy controls. We have utilized a miRNA modification strategy to enhance the therapeutic efficacy of miR-15a by replacing uracil with 5-fluorouracil (5-FU). We demonstrated increased levels of YAP1 and BCL-2 (both targets of miR-15a) in pancreatic tissues obtained from Ptf1aCreERTM and Ptf1aCreERTM;LSL-KrasG12D mice after chronic pancreatitis induction as compared to controls. In vitro studies showed that delivery of 5-FU-miR-15a significantly decreased viability, proliferation, and migration of PSCs over six days compared to 5-FU, TGFβ1, control miR, and miR-15a. In addition, treatment of PSCs with 5-FU-miR-15a in the context of TGFβ1 treatment exerted a more substantial effect than TGFβ1 alone or when combined with other miRs. Conditioned medium obtained from PSC cells treated with 5-FU-miR-15a significantly inhibits the invasion of pancreatic cancer cells compared to controls. Importantly, we demonstrated that treatment with 5-FU-miR-15a reduced the levels of YAP1 and BCL-2 observed in PSCs. Our results strongly suggest that ectopic delivery of miR mimetics is a promising therapeutic approach for pancreatic fibrosis and that 5-FU-miR-15a shows specific promise.
90K/Mac-2 BP Is a New Predictive Biomarker of Response to Infliximab Therapy in IBD Patients
Authors: Pesole Pasqua Letizia , Liso Marina , Donghia Rossella , Guerra Vito , Lippolis Antonio , Mastronardi Mauro , Iacovazzi Palma Aurelia
Published: February 2023

Inflammatory bowel diseases (IBD), comprising Crohn’s disease (CD) and Ulcerative Colitis (UC), are multifactorial disorders characterized by a chronic inflammatory status with the secretion of cytokines and immune mediators. Biologic drugs targeting pro-inflammatory cytokines, such as infliximab, are broadly used in the treatment of IBD patients, but some patients lose responsiveness after an initial success. The research into new biomarkers is crucial for advancing personalized therapies and monitoring the response to biologics. The aim of this single center, observational study is to analyze the relationship between serum levels of 90K/Mac-2 BP and the response to infliximab, in a cohort of 48 IBD patients (30 CD and 18 UC), enrolled from February 2017 to December 2018. In our IBD cohort, high 90K serum levels were found at baseline in patients who then developed anti-infliximab antibodies at the fifth infusion (22 weeks after the first), becoming non-responders (9.76 ± 4.65 µg/mL compared to 6.53 ± 3.29 µg/mL in responder patients, p = 0.005). This difference was significant in the total cohort and in CD, but not significant in UC. We then analyzed the relationship between serum levels of 90K, C-reactive protein (CRP), and Fecal calprotectin. A significant positive correlation was found at baseline between 90K and CRP, the most common serum inflammation marker (R = 0.42, p = 0.0032). We concluded that circulating 90K could be considered a new non-invasive biomarker for monitoring the response to infliximab. Furthermore, 90K serum level determination, before the first infliximab infusion, in association with other inflammatory markers such as CRP, could assist in the choice of biologics for the treatment of IBD patients, thereby obviating the need for a drug switch due to loss of response, and so improving clinical practice and patient care.
Dermal PapillaCell-Derived Exosomes Regulate Hair Follicle Stem Cell Proliferation via LEF1
Authors: Li Jiali , Zhao Bohao , Yao Shuyu , Dai Yingying , Zhang Xiyu , Yang Naisu , Bao Zhiyuan , Cai Jiawei , Chen Yang , Wu Xinsheng
Published: February 2023

Hair follicle (HF) growth and development are controlled by various cell types, including hair follicle stem cells (HFSCs) and dermal papilla cells (DPCs). Exosomes are nanostructures that participate in many biological processes. Accumulating evidence indicates that DPC-derived exosomes (DPC-Exos) mediate HFSC proliferation and differentiation during the cyclical growth of hair follicles. In this study, we found that DPC-Exos increase ki67 expression and CCK8 cell viability readouts in HFSCs but reduce annexin staining of apoptotic cells. RNA sequencing of DPC-Exos-treated HFSCs identified 3702 significantly differentially expressed genes (DEGs), including BMP4, LEF1, IGF1R, TGFβ3, TGFα, and KRT17. These DEGs were enriched in HF growth- and development-related pathways. We further verified the function of LEF1 and showed that overexpression of LEF1 increased the expression of HF development-related genes and proteins, enhanced HFSC proliferation, and reduced HFSC apoptosis, while knockdown of LEF1 reversed these effects. DPC-Exos could also rescue the siRNA-LEF1 effect in HFSCs. In conclusion, this study demonstrates that DPC-Exos mediated cell-to-cell communication can regulate HFSCs proliferation by stimulating LEF1 and provide novel insights into HF growth and development regulatory mechanisms.
Clonal Hematopoiesis Mutations Are Present in Atherosclerotic Lesions in Peripheral Artery Disease
Authors: Büttner Petra , Böttner Julia , Krohn Knut , Baber Ronny , Platzbecker Uwe , Cross Michael , Desch Steffen , Thiele Holger , Steiner Sabine , Scheinert Dierk , Metzeler Klaus H. , Branzan Daniela
Published: February 2023

Clonal hematopoiesis (CH)-associated mutations increase the risk of atherosclerotic cardiovascular diseases. However, it is unclear whether the mutations detected in circulating blood cells can also be detected in tissues associated with atherosclerosis, where they could affect physiology locally. To address this, the presence of CH mutations in peripheral blood, atherosclerotic lesions and associated tissues was assessed in a pilot study of 31 consecutive patients with peripheral vascular disease (PAD) who underwent open surgical procedures. Next-generation sequencing was used to screen the most commonly mutated loci (DNMT3A, TET2, ASXL1 and JAK2). Twenty CH mutations were detected in peripheral blood of 14 (45%) patients, 5 of whom had more than one mutation. TET2 (11 mutations, 55%) and DNMT3A (8 mutations, 40%) were the most frequently affected genes. Altogether, 88% of the mutations detectable in peripheral blood were also present in the atherosclerotic lesions. Twelve patients also had mutations in perivascular fat or subcutaneous tissue. The presence of CH mutations in PAD-associated tissues as well as in blood suggests that CH mutations may make a previously unknown contribution to PAD disease biology.
Specific Mutations near the Amyloid Precursor Protein Cleavage Site Increase γ-Secretase Sensitivity and Modulate Amyloid-β Production
Authors: Suzuki Ryota , Takahashi Haruka , Yoshida Chika , Hidaka Masafumi , Ogawa Tomohisa , Futai Eugene
Published: February 2023

Amyloid-β peptides (Aβs) are produced via cleavage of the transmembrane region of the amyloid precursor protein (APP) by γ-secretase and are responsible for Alzheimer’s disease. Familial Alzheimer’s disease (FAD) is associated with APP mutations that disrupt the cleavage reaction and increase the production of neurotoxic Aβs, i.e., Aβ42 and Aβ43. Study of the mutations that activate and restore the cleavage of FAD mutants is necessary to understand the mechanism of Aβ production. In this study, using a yeast reconstruction system, we revealed that one of the APP FAD mutations, T714I, severely reduced the cleavage, and identified secondary APP mutations that restored the cleavage of APP T714I. Some mutants were able to modulate Aβ production by changing the proportions of Aβ species when introduced into mammalian cells. Secondary mutations include proline and aspartate residues; proline mutations are thought to act through helical structural destabilization, while aspartate mutations are thought to promote interactions in the substrate binding pocket. Our results elucidate the APP cleavage mechanism and could facilitate drug discovery.
Conformational Dynamics of Phytoglobin BvPgb1.2 from Beta vulgaris ssp. vulgaris
Authors: Christensen Simon , Stenström Olof , Akke Mikael , Bülow Leif
Published: February 2023

Plant hemoglobins, often referred to as phytoglobins, play important roles in abiotic stress tolerance. Several essential small physiological metabolites can be bound to these heme proteins. In addition, phytoglobins can catalyze a range of different oxidative reactions in vivo. These proteins are often oligomeric, but the degree and relevance of subunit interactions are largely unknown. In this study, we delineate which residues are involved in dimer formation of a sugar beet phytoglobin type 1.2 (BvPgb1.2) using NMR relaxation experiments. E. coli cells harboring a phytoglobin expression vector were cultivated in isotope-labeled (2H, 13C and 15N) M9 medium. The triple-labeled protein was purified to homogeneity using two chromatographic steps. Two forms of BvPgb1.2 were examined, the oxy-form and the more stable cyanide-form. Using three-dimensional triple-resonance NMR experiments, sequence-specific assignments for CN-bound BvPgb1.2 were achieved for 137 backbone amide cross-peaks in the 1H-15N TROSY spectrum, which amounts to 83% of the total number of 165 expected cross-peaks. A large proportion of the non-assigned residues are located in α-helixes G and H, which are proposed to be involved in protein dimerization. Such knowledge around dimer formation will be instrumental for developing a better understanding of phytoglobins’ roles in planta.
Pathophysiology and Treatment of Chronic Thromboembolic Pulmonary Hypertension
Authors: Otani Naoyuki , Watanabe Ryo , Tomoe Takashi , Toyoda Shigeru , Yasu Takanori , Nakamoto Takaaki
Published: February 2023

Chronic thromboembolic pulmonary hypertension (CTEPH) is a condition in which an organic thrombus remains in the pulmonary artery (PA) even after receiving anticoagulation therapy for more than 3 months and is complicated by pulmonary hypertension (PH), leading to right-sided heart failure and death. CTEPH is a progressive pulmonary vascular disease with a poor prognosis if left untreated. The standard treatment for CTEPH is pulmonary endarterectomy (PEA), which is usually performed only in specialized centers. In recent years, balloon pulmonary angioplasty (BPA) and drug therapy for CTEPH have also shown good results. This review discusses the complex pathogenesis of CTEPH and presents the standard of care, PEA, as well as a new device called BPA, which is showing remarkable progress in efficacy and safety. Additionally, several drugs are now demonstrating established evidence of efficacy in treating CTEPH.
Calpain Regulates Reactive Oxygen Species Production during Capacitation through the Activation of NOX2 and NOX4
Authors: Ortiz-García César I. , Salgado-Lucio Monica L. , Roa-Espitia Ana L. , Muñoz-Sánchez Aidé A. , Cordero-Martínez Joaquín , Hernández-González Enrique O.
Published: February 2023

Capacitation is a series of physiological, biochemical, and metabolic changes experienced by mammalian spermatozoa. These changes enable them to fertilize eggs. The capacitation prepares the spermatozoa to undergo the acrosomal reaction and hyperactivated motility. Several mechanisms that regulate capacitation are known, although they have not been fully disclosed; among them, reactive oxygen species (ROS) play an essential role in the normal development of capacitation. NADPH oxidases (NOXs) are a family of enzymes responsible for ROS production. Although their presence in mammalian sperm is known, little is known about their participation in sperm physiology. This work aimed to identify the NOXs related to the production of ROS in guinea pig and mouse spermatozoa and define their participation in capacitation, acrosomal reaction, and motility. Additionally, a mechanism for NOXs’ activation during capacitation was established. The results show that guinea pig and mouse spermatozoa express NOX2 and NOX4, which initiate ROS production during capacitation. NOXs inhibition by VAS2870 led to an early increase in the capacitation and intracellular concentration of Ca2+ in such a way that the spermatozoa also presented an early acrosome reaction. In addition, the inhibition of NOX2 and NOX4 reduced progressive motility and hyperactive motility. NOX2 and NOX4 were found to interact with each other prior to capacitation. This interaction was interrupted during capacitation and correlated with the increase in ROS. Interestingly, the association between NOX2-NOX4 and their activation depends on calpain activation, since the inhibition of this Ca2+-dependent protease prevents NOX2-NOX4 from dissociating and ROS production. The results indicate that NOX2 and NOX4 could be the most important ROS producers during guinea pig and mouse sperm capacitation and that their activation depends on calpain.
Oxidative-Stress-Mediated ER Stress Is Involved in Regulating Manoalide-Induced Antiproliferation in Oral Cancer Cells
Authors: Peng Sheng-Yao , Tang Jen-Yang , Lan Ting-Hsun , Shiau Jun-Ping , Chen Kuan-Liang , Jeng Jiiang-Huei , Yen Ching-Yu , Chang Hsueh-Wei
Published: February 2023

Manoalide provides preferential antiproliferation of oral cancer but is non-cytotoxic to normal cells by modulating reactive oxygen species (ROS) and apoptosis. Although ROS interplays with endoplasmic reticulum (ER) stress and apoptosis, the influence of ER stress on manoalide-triggered apoptosis has not been reported. The role of ER stress in manoalide-induced preferential antiproliferation and apoptosis was assessed in this study. Manoalide induces a higher ER expansion and aggresome accumulation of oral cancer than normal cells. Generally, manoalide differentially influences higher mRNA and protein expressions of ER-stress-associated genes (PERK, IRE1α, ATF6, and BIP) in oral cancer cells than in normal cells. Subsequently, the contribution of ER stress on manoalide-treated oral cancer cells was further examined. ER stress inducer, thapsigargin, enhances the manoalide-induced antiproliferation, caspase 3/7 activation, and autophagy of oral cancer cells rather than normal cells. Moreover, N-acetylcysteine, an ROS inhibitor, reverses the responses of ER stress, aggresome formation, and the antiproliferation of oral cancer cells. Consequently, the preferential ER stress of manoalide-treated oral cancer cells is crucial for its antiproliferative effect.
Fat Loss in Patients with Metastatic Clear Cell Renal Cell Carcinoma Treated with Immune Checkpoint Inhibitors
Authors: Lee Ji Hyun , Hwang Soohyun , Jee ByulA , Kim Jae-Hun , Lee Jihwan , Chung Jae Hoon , Song Wan , Sung Hyun Hwan , Jeon Hwang Gyun , Jeong Byong Chang , Seo Seong Il , Jeon Seong Soo , Lee Hyun Moo , Park Se Hoon , Kwon Ghee Young , Kang Minyong
Published: February 2023

The purpose of this study was to determine the prognostic impact of fat loss after immune checkpoint inhibitor (ICI) treatment in patients with metastatic clear cell renal cell carcinoma (ccRCC). Data from 60 patients treated with ICI therapy for metastatic ccRCC were retrospectively analyzed. Changes in cross-sectional areas of subcutaneous fat (SF) between the pre-treatment and post-treatment abdominal computed tomography (CT) images were expressed as percentages and were divided by the interval between the CT scans to calculate ΔSF (%/month). SF loss was defined as ΔSF < −5%/month. Survival analyses for overall survival (OS) and progression-free survival (PFS) were performed. Patients with SF loss had shorter OS (median, 9.5 months vs. not reached; p < 0.001) and PFS (median, 2.6 months vs. 33.5 months; p < 0.001) than patients without SF loss. ΔSF was independently associated with OS (adjusted hazard ratio (HR), 1.49; 95% confidence interval (CI), 1.07–2.07; p = 0.020) and PFS (adjusted HR, 1.57; 95% CI, 1.17–2.12; p = 0.003), with a 5%/month decrease in SF increasing the risk of death and progression by 49% and 57%, respectively. In conclusion, Loss of SF after treatment initiation is a significant and independent poor prognostic factor for OS and PFS in patients with metastatic ccRCC who receive ICI therapy.
Managing the Dual Nature of Iron to Preserve Health
Authors: Silvestri Laura , Pettinato Mariateresa , Furiosi Valeria , Bavuso Volpe Letizia , Nai Antonella , Pagani Alessia
Published: February 2023

Because of its peculiar redox properties, iron is an essential element in living organisms, being involved in crucial biochemical processes such as oxygen transport, energy production, DNA metabolism, and many others. However, its propensity to accept or donate electrons makes it potentially highly toxic when present in excess and inadequately buffered, as it can generate reactive oxygen species. For this reason, several mechanisms evolved to prevent both iron overload and iron deficiency. At the cellular level, iron regulatory proteins, sensors of intracellular iron levels, and post-transcriptional modifications regulate the expression and translation of genes encoding proteins that modulate the uptake, storage, utilization, and export of iron. At the systemic level, the liver controls body iron levels by producing hepcidin, a peptide hormone that reduces the amount of iron entering the bloodstream by blocking the function of ferroportin, the sole iron exporter in mammals. The regulation of hepcidin occurs through the integration of multiple signals, primarily iron, inflammation and infection, and erythropoiesis. These signals modulate hepcidin levels by accessory proteins such as the hemochromatosis proteins hemojuvelin, HFE, and transferrin receptor 2, the serine protease TMPRSS6, the proinflammatory cytokine IL6, and the erythroid regulator Erythroferrone. The deregulation of the hepcidin/ferroportin axis is the central pathogenic mechanism of diseases characterized by iron overload, such as hemochromatosis and iron-loading anemias, or by iron deficiency, such as IRIDA and anemia of inflammation. Understanding the basic mechanisms involved in the regulation of hepcidin will help in identifying new therapeutic targets to treat these disorders.
Chemistry towards Biology
Authors: Hricovini Milos , Jampilek Josef
Published: February 2023

Although it may not seem like it, chemical biology has existed for a long time from today’s perspective [...]
The Alpha-1 Subunit of the Na+/K+-ATPase (ATP1A1) Is a Host Factor Involved in the Attachment of Porcine Epidemic Diarrhea Virus
Authors: Xiong Moukang , Liu Xianhui , Liang Tairun , Ban Yanfang , Liu Yanling , Zhang Leyi , Xu Zheng , Song Changxu
Published: February 2023

Porcine epidemic diarrhea (PED) is an acute and severe atrophic enteritis caused by porcine epidemic diarrhea virus (PEDV) that infects pigs and makes huge economic losses to the global swine industry. Previously, researchers have believed that porcine aminopeptidase-N (pAPN) was the primary receptor for PEDV, but it has been found that PEDV can infect pAPN knockout pigs. Currently, the functional receptor for PEDV remains unspecified. In the present study, we performed virus overlay protein binding assay (VOPBA), found that ATP1A1 was the highest scoring protein in the mass spectrometry results, and confirmed that the CT structural domain of ATP1A1 interacts with PEDV S1. First, we investigated the effect of ATP1A1 on PEDV replication. Inhibition of hosts ATP1A1 protein expression using small interfering RNA (siRNAs) significantly reduced the cells susceptibility to PEDV. The ATP1A1-specific inhibitors Ouabain (a cardiac steroid) and PST2238 (a digitalis toxin derivative), which specifically bind ATP1A1, could block the ATP1A1 protein internalization and degradation, and consequently reduce the infection rate of host cells by PEDV significantly. Additionally, as expected, overexpression of ATP1A1 notably enhanced PEDV infection. Next, we observed that PEDV infection of target cells resulted in upregulation of ATP1A1 at the mRNA and protein levels. Furthermore, we found that the host protein ATP1A1 was involved in PEDV attachment and co-localized with PEDV S1 protein in the early stage of infection. In addition, pretreatment of IPEC-J2 and Vero-E6 cells with ATP1A1 mAb significantly reduced PEDV attachment. Our observations provided a perspective on identifying key factors in PEDV infection, and may provide valuable targets for PEDV infection, PEDV functional receptor, related pathogenesis, and the development of new antiviral drugs.
De Novo Assembly and Characterization of the Transcriptome of an Omnivorous Camel Cricket (Tachycines meditationis)
Authors: Lu Jun-Hui , Guan De-Long , Xu Sheng-Quan , Huang Huateng
Published: February 2023

Tachycines meditationis (Orthoptera: Rhaphidophoridae: Tachycines) is a widely distributed insect in eastern Asia. This species is common in urban environments, and its unique omnivorous diet may contribute to its success in various habitats. However, molecular studies on the species are scarce. Here, we obtained the first transcriptome sequence of T. meditationis and performed preliminary analyses to test whether the evolution of coding sequences fits the expectations based on the species’ ecology. We retrieved 476,495 effective transcripts and annotated 46,593 coding sequences (CDS). We analysed the codon usage and found that directional mutation pressure was the leading cause of codon usage bias in this species. This genome-wide relaxed codon usage pattern in T. meditationis is surprising, given the potentially large population size of this species. Moreover, despite the omnivorous diet, the chemosensory genes of this species do not exhibit codon usage deviating significantly from the genome-level pattern. They also do not seem to experience more gene family expansion than other cave cricket species do. A thorough search for rapidly evolved genes using the dN/dS value showed that genes associated with substance synthesis and metabolic pathways, such as retinol metabolism, aminoacyl-tRNA biosynthesis, and fatty acid metabolism, underwent species-specific positive selection. While some results seem to contradict the species ecology, our transcriptome assembly provides a valuable molecular resource for future studies on camel cricket evolution and molecular genetics for feeding ecology in insects, in general.
Development of a Novel Anti-CD44 Variant 6 Monoclonal Antibody C44Mab-9 for Multiple Applications against Colorectal Carcinomas
Authors: Ejima Ryo , Suzuki Hiroyuki , Tanaka Tomohiro , Asano Teizo , Kaneko Mika K. , Kato Yukinari
Published: February 2023

CD44 is a cell surface glycoprotein, and its isoforms are produced by the alternative splicing with the standard and variant exons. The CD44 variant exon-containing isoforms (CD44v) are overexpressed in carcinomas. CD44v6 is one of the CD44v, and its overexpression predicts poor prognosis in colorectal cancer (CRC) patients. CD44v6 plays critical roles in CRC adhesion, proliferation, stemness, invasiveness, and chemoresistance. Therefore, CD44v6 is a promising target for cancer diagnosis and therapy for CRC. In this study, we established anti-CD44 monoclonal antibodies (mAbs) by immunizing mice with CD44v3-10-overexpressed Chinese hamster ovary (CHO)-K1 cells. We then characterized them using enzyme-linked immunosorbent assay, flow cytometry, western blotting, and immunohistochemistry. One of the established clones (C44Mab-9; IgG1, kappa) reacted with a peptide of the variant 6-encoded region, indicating that C44Mab-9 recognizes CD44v6. Furthermore, C44Mab-9 reacted with CHO/CD44v3-10 cells or CRC cell lines (COLO201 and COLO205) by flow cytometry. The apparent dissociation constant (KD) of C44Mab-9 for CHO/CD44v3-10, COLO201, and COLO205 was 8.1 × 10−9 M, 1.7 × 10−8 M, and 2.3 × 10−8 M, respectively. C44Mab-9 detected the CD44v3-10 in western blotting, and partially stained the formalin-fixed paraffin-embedded CRC tissues in immunohistochemistry. Collectively, C44Mab-9 is useful for detecting CD44v6 in various applications.
Investigation of Molecular Mechanisms Involved in Sensitivity to the Anti-Cancer Activity of Costunolide in Breast Cancer Cells
Authors: Choi Yu-Jeong , Choi Youn Kyung , Ko Seong-Gyu , Cheon Chunhoo , Kim Tai Young
Published: February 2023

Costunolide (CTL), an active compound isolated from Saussurea lappa Clarke and Laurus nobilis L, has been shown to induce apoptosis via reactive oxygen species (ROS) generation in various types of cancer cells. However, details of molecular mechanisms underlying the difference in sensitivity of cancer cells to CTL are still largely unknown. Here, we tested the effect of CTL on the viability of breast cancer cells and found that CTL had a more efficient cytotoxic effect against SK-BR-3 cells than MCF-7 cells. Mechanically, ROS levels were significantly increased upon CTL treatment only in SK-BR-3 cells, which leads to lysosomal membrane permeabilization (LMP) and cathepsin D release, and subsequent activation of the mitochondrial-dependent intrinsic apoptotic pathway by inducing mitochondrial outer membrane permeabilization (MOMP). In contrast, treatment of MCF-7 cells with CTL activated PINK1/Parkin-dependent mitophagy to remove damaged mitochondria, which prevented the elevation of ROS levels, thereby contributing to their reduced sensitivity to CTL. These results suggest that CTL is a potent anti-cancer agent, and its combination with the inhibition of mitophagy could be an effective method for treating breast cancer cells that are less sensitive to CTL.