bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genetically encoded multivalent liquid glycan array displayed on M13 bacteriophage
Authors: Sojitra Mirat , Sarkar Susmita , Maghera Jasmine , Schmidt Edward N. , Rodrigues Emily , Carpenter Eric J. , Seth Shaurya , Vinals Daniel F. , Bennett Nicholas J. , Reddy Revathi , Khalil Amira , Xue Xiaochao , Bell Michael R. , Zheng Ruixiang B. , Zhang Ping , Nycholat Corwin , Bailey Justin J. , Ling Chang‐Chun , Lowary Todd L. , Paulson James C. , Macauley Matthew S. , Derda Ratmir 
Published: May 2022

Abnormal cell surface glycosylation plays a major role in disease processes such as immune evasion. However, the underlying role of glycans is yet to be fully understood. Binding information obtained from glycan arrays can provide critical starting points for downstream applications such as the development of carbohydrate‐based inhibitors, vaccines, and other therapeutics. However, it is challenging to use powerful techniques like DNA deep sequencing to analyze glycan recognition due to the lack of 1:1 correspondence between DNA and glycan structures. Therefore, we have developed Liquid Glycan Array (LiGA), a technology that allows for genetic encoding of glycans. LiGA provides a 1:1 correspondence between the glycan displayed in multiple copies on a bacteriophage carrier and the phage genetic material. LiGA is generated by acylation of phage pVIII protein with a dibenzocyclooctyne, followed by ligation of azido‐modified glycans. The display of glycans on each phage virion can be controlled from 30‐1500 copies to probe the critical variables in glycan recognition: valency and density. A simple pulldown of the LiGA along with lectins followed by deep sequencing of the DNA in the bound phage decodes the recognized glycans. LiGA is target agnostic and measures binding profile of lectins expressed on intact cells, such as hCD22 (Siglec‐2) and DC‐SIGN (Dendritic Cell‐Specific Intercellular adhesion molecule‐3‐Grabbing Non‐integrin), and in live mice (Nat. Chem. Bio. 17, 806–816, 2021). From a mixture of 50‐100 multivalent glycan probes, LiGA identifies the glycan‐phage conjugates with optimal valency and density for binding to antibodies and lectins on cells in vitro and in vivo.Sialic acid‐binding immunoglobulin‐type lectins (Siglecs) expressed on the surface of immune cells are exploited by cancer to evade immune response. We applied LiGA to study the binding specificity of Siglec‐7, a cell surface receptor that cancer cells use to evade immune response from natural killer (NK) cells. Additionally, we explored the roles of valency and density in ganglioside interaction with Siglec‐1 using a cell‐based assay. Building on these successes, we plan to use LiGA to identify the valency and affinity required by trans‐ glycan to overcome the cis‐ masking on the surface of immune cells.
Proteins Interacting with S‐Nitrosoglutathione reductase to Regulate Nitric Oxide Homeostasis
Authors: Kaur Yadwinder , Treffon Patrick , Vierling Elizabeth 
Published: May 2022

In plants, nitric oxide (NO) is a crucial signaling molecule involved in many physiological processes and stress pathways. NO can influence plant germination, flowering, and root development1. NO is usually found in an adduct with glutathione, forming S‐nitrosoglutathione (GSNO), which can affect protein S‐nitrosation, an important post‐translational modification that can alter protein activity and stability. The enzyme S‐nitrosoglutathione reductase (GSNOR) catalyzes the irreversible NADH‐dependent reduction of GSNO. GSNOR’s ability to regulate GSNO levels and subsequently affect S‐nitrosation activity makes it a critical enzyme in NO signaling2. The objective of this project is to identify proteins that interact with GSNOR to regulate its activity and stability, thereby impacting NO homeostasis. Based on previous experiments, we hypothesize that thioredoxins and catalase may be recovered as interacting candidates. We are utilizing enzyme‐catalyzed proximity labeling with the biotin ligase, TurboID, coupled to mass spectrometry to identify interacting partners of GSNOR in the plant model Arabidopsis thaliana. Currently, we have shown that a GSNOR‐TurboID fusion protein that has been transformed into a GSNOR null mutant can rescue the mutant phenotype without displaying any adverse effects on plant growth. As controls, A. thaliana lines carrying TurboID fused to YFP and targeted to the nucleus or cytosol have also been characterized. Importantly, upon treatment with biotin, TurboID produces strong biotinylation signals in these lines. These findings indicate that TurboID is suitable for use in A. thalianaand that biotinylated proteins can be recovered for identification by mass spectrometry. These experiments will advance the current understanding of GSNOR’s regulation, leading to an improved model of NO signaling.1. Xu, S., D. Guerra, U. Lee, & E. Vierling. S‐Nitrosoglutathione reductases are low‐copy number, cysteine‐rich proteins in plants that control multiple developmental and defense responses in Arabidopsis. Front. Plant Sci. 4: 1‐13 (2013). doi: 10.3389/fpls.2013.00430. 2. Lee, U., Wie, C., Fernandez, B. O., Feelisch, M. & Vierling, E. Modulation of nitrosative stress by S‐nitrosoglutathione reductase is critical for thermotolerance and plant growth in Arabidopsis. Plant Cell. 20: 786–802 (2008). doi: 10.1105/tpc/107.052647.
Anti‐hypertensive effect of BPM4 on spontaneously hypertensive rats that antagonizes Dopamine βeta hydroxylase
Authors: Saini Manisha , Dey Sanjay K. , Kundu Suman 
Published: May 2022

BackgroundHypertension, which is described as chronically increased blood pressure above 140/90 mmHg, has emerged as a starring cause of mortality and morbidity worldwide. It is a leading risk factor for major cardiovascular events (such as heart attack and stroke), chronic kidney disease, heart failure, cognitive impairment and dementia. The dire consequences of uncontrolled hypertension indicate that controlling it will have a direct preventive effect on the vital organs. Despite the availability of a large number of classical anti‐hypertensive drugs it still affects humans adversely and poses a global disease burden thus necessitating new drugs. A non‐conventional target whose inhibition can regulate hypertension is Dopamine βeta hydroxylase (DBH). This enzyme, which plays a significant role in the catecholamine biosynthetic pathway, utilizes the body’s blood pressure (BP) lowering mechanism and thus can combat resistant hypertension.MethodologyIn the study, we have screened thousands of small molecules against the active site of DBH (PDB ID:4ZEL). Hits identified in silico were assessed for their inhibitory potential and strength of binding (Kd) in vitro. The ex vivo toxicity parameters were evaluated to get rid of toxic molecules before going for in vivostudies. Selected leads were screened in spontaneously hypertensive rats (SHRs) to evaluate their efficacy in reducing blood pressure as well as their ability to improve other cardiac parameters. Thereafter, acute as well as genotoxicity study was performed to minimize the rate of attrition in later phases of clinical development.ResultsOur best lead molecule “BPM4” inhibited DBH efficiently with an IC50 value of 18 μM and dissociation constant of 22 nM. BPM4 when administered through IP at a dose of 10 mg/kg BW significantly reduced BP with a p‐value of 0.001 as compared to the untreated group. Additionally, a significant reduction was observed in IVSs, IVSd, LVPWs and LVPWd, whereas increment was observed in internal diameter in the case of end systole and diastole (LVIDs and LVIDd). A high oral lethal dose (LD50) > 2000 mg/kg was observed in the acute toxicity test. Ames test showed no positive mutagenic effect +‐S9. No statistically significant decrease in % mitotic index nor any aberrant cells were observed in lymphocytes and bone marrow of rodents.ConclusionBPM4 (at 10 mg/kg BW) statistically reduced the BP as well as improved the left ventricular hypertrophy parameters in SHRS. The area of fibrosis was also decreased post treatment (cardiac fibrosis disrupts the normal myocardial structure and has a direct impact on heart function and survival). Subsequently, the compound successfully passed the three battery genotoxicity tests as it did not cause any structural damage to DNA nor did it induce any ploidy changes, holds scope for being a potent anti‐hypertensive drug candidate.
Molecular, Biochemical and Physiological Studies on the Potential Influence of the Beta vulgaris Extract to Mitigate the Effect of Chemical Colorants of Food on Male Rats
Authors: Okab Aly B. , Elbanna Sabah G. , Khalil Khaled A. Z. 
Published: May 2022

Synthetic food colorings as carmoisine (E122) contains the azo group (–N=N) that linked to the aromatic rings in the chemical molecular structures used as red color. Red beetroot (Beta vulgaris) ‐ a natural antioxidant‐ is a rich plant source of nutrients that give tremendous health benefits, red beetroot includes anthocyanins and betalains, which are natural antioxidants and can protect against age‐related diseases. The objective of this study was to investigate the potential protective effects of Beta vulgaris extracts against Carmoisine induced oxidative damage among male rats. In this study animals were divided randomly into 6 groups (18 animals each). Hemoglobin significantly increased after 15 days in Red beetroot 10 mg/kg (B), Carmoisine low dose 50 mg/kg (CLD), and beetroot + carmoisine high dose 10 mg/kg + 100 mg/kg (BCHD), while decreased after 30 days in CHD and increased after 45 days in CHD, BCLD, and BCHD. While WBCs increased after 15 and 45 days but decreased after 30 days in all groups. Thiobarbituric acid reactive substances (TBARS) in blood plasma increased after 15 days in all groups, while, after 30 days, TBARS in blood plasma decrease in B, CLD, BCHD, whereas, TBARS in liver increased in all groups, and TBARS in testes increased in all groups except B. After 45 days TBARS in testes decreased in B, increased in CLD, CHD, and BCHD. Cytochrome b5 in liver increased after 15 days in all groups, except BCLD. Cytochrome P450 increased in all groups, while, after 30 days cytochrome b5 and cytochrome P450 decreased in BCLD. After 45 days cytochrome b5 increased in CHD, and decreased in BCLD. While, cytochrome P450 increased in CLD, CHD, and BCHD, and decreased in BCLD. Superoxide dismutase (SOD), catalase in liver male rats after 15 days increased in all groups, while DNA breakage decreased in BCLD and BCHD, whereas, after 30 days catalase decreased in CLD and BCHD, and DNA breakage increased in CLD and decreased in BCHD, however, after 45 days SOD decreased in BCHD, catalase decreased in BCLD and BCHD, however DNA breakage increased in all groups except B. Catalase in testes of male rats after 15 days decreased in all groups except BCHD, however, testes DNA breakage decreased in B and CHD, while, after 30 days SOD increased in all groups, while catalase decreased in all groups except CHD, while, DNA breakage increased in CHD, but decreased in BCLD and BCHD, whereas, after 45 days catalase increased in all groups, DNA breakage decreased in B, BCLD, and BCHD, except, CLD and CHD.From the present results it was concluded that Co‐administration of beetroot decreased the oxidative stress in liver and testes and also prevent liver damage, also decreased DNA breakage.
Hepatoprotective Effects of Astragaloside IV and Formononetin Against Oxidative Injury
Authors: Tran Quoc‐Anh , Duong Nhi , Luu Vy T. , Xiong Hou M. , Ho Linh 
Published: May 2022

The mitochondrial SIRT3 is a major deacetylase protein that regulates important mitochondrial biology, including metabolic pathways, reactive oxygen species detoxification, mitochondrial dynamics, and the mitochondrial unfolded protein response. We found that astragalus aqueous extract increased the activity of SIRT3 in vitro. We further explore the potential antioxidative effects of Astragaloside IV (AST‐IV) and Formononetin (FMR) on induced oxidative stressed AML12 (alpha mouse liver 12) cells. The AML12 cell line was established from healthy hepatocytes from a mouse (CD1 Strain, Line MT42) transgenic for human TFG alpha.We first tested whether AST‐IV or formononetin increases SIRT3 expression in cell model. We cultured AML12 and treated with various concentrations of AST‐IV or formononetin in parallel with untreated control, negative control, and positive control. We found that AST‐IV and formononetin induced SIRT3 expression through Western Blotting (WB) analysis.In addition, we assessed rescue effects of AST‐IV and formononetin on t‐BHP‐induced oxidative injury in AML12 hepatocytes under three separate conditions: pre‐treatment with AST‐IV or formononetin, post‐treatment with AST‐IV or formononetin, and both pre‐treatment and post‐treatment with AST‐IV and formononetin. Through CCK‐8 analysis, we identified that both pre‐treatment and post‐treatment significantly increased cell proliferation on induced‐oxidative stress AML12 cells compared to no treatment and control. Pre‐treatment of AML12 cells before induction of oxidative stress with AST‐IV and formononetin also showed cell proliferation but was less significant compared to AML12 cells that received both pre‐treatment and post‐treatment with AST‐IV or formononetin. Post‐treatment AST‐IV or formononetin after induction of oxidative stress on AML12 cells did not significantly show any difference in cell proliferation compared to non‐treatment and control. Significantly AST‐IV and formononetin showed induction of the SIRT3 protein on tert‐butyl hydroperoxide (t‐BHP) induced oxidative stressed AML12 hepatocytes by WB.Taken together, the increase in cell proliferation is indicative of the antioxidative effects of AST‐IV and formononetin through induction of SIRT3 protein. Further investigation is currently conducted to provide evidence of AST‐IV and formononetin's mechanistic effect on inducing the expression of the SIRT3 protein‐linked downstream pathways for their antioxidative benefit to hepatocytes. The ultimate goal is to utilize these SIRT3 modulators as pharmacologic agents in treating oxidative injury hepatocytes.
Identification of protein markers for skeletal muscle‐derived extracellular vesicles (SkM‐EVs) by quantitative proteomics reveals how SkM‐EVs function in vivo
Authors: Watanabe Sho , Sudo Yuri , Sakurai Hidetoshi , Sato Ryuichiro , Yamauchi Yoshio 
Published: May 2022

BackgroundSkeletal muscle serves as a secretory organ; it releases various soluble factors called myokine. They act as mediators for cell‐cell communications in autocrine, paracrine, and endocrine fashions. Extracellular vesicles (EVs) have been proposed to be a partial paracrine and/or endocrine effectors by facilitating the exchange of biological cargoes among cells and tissue. Some in vitro study showed that skeletal muscle released EVs and skeletal muscle‐derived EVs (SkM‐EVs) could induce biological effects in recipient cells. However, whether the skeletal muscle actively releases EVs in vivo, how much proportion of plasma EVs are derived from this tissue, and where SkM‐EVs are delivered and exert their roles remain largely unknown. Here, we aimed to identify SkM‐EV marker proteins by quantitative proteomics on human and mouse myocyte‐derived EVs to evaluate SkM‐EVs abundance in vivo. In addition, we explored whether SkM‐EVs isolated from mice could exert biological functions.MethodsProteomic analysis of EVs released from mouse C2C12 myotubes and human iPS‐derived myocytes was performed to identify potential SkM‐EVs markers. The levels of SkM‐EVs in plasma and muscle interstitium were evaluated by the abundance of identified SkM‐EVs protein markers. Based on previous methods, we developed a modified protocol to isolated SkM‐EVs from muscle interstitium. We confirmed that muscle interstitium‐EVs were incorporated into C2C12 myoblast and investigated the expression levels of myogenesis‐related genes.Results& DiscussionWe first determined proteomic profiles of EVs released from C2C12 myoblasts, C2C12 myotubes, and hiPSC‐derived myocytes and identified several proteins that serve as potential markers for SkM‐EVs. Among them, we identified ATP2A1, β‐enolase, and desmin as reliable SkM‐EV marker proteins. Using these protein markers, we confirmed that SkM‐EVs were present in muscle interstitium but little in the blood, which were supported by our electron microscopy. Unexpectedly, the abundance of SkM‐EVs present in plasma and muscle interstitium was not changed in response to acute exercise. Furthermore, we showed that SkM‐EVs accumulated in muscle interstitium space were incorporated into C2C12 myoblast and promoted muscle differentiation by regulating gene expression of Pax7 and myosin heavy chain at least partially through myomiRs (miRs‐1, ‐206, ‐431, and ‐486). We expect that molecular characterization and in vivo distribution of SkM‐EVs could provide a conceptually new basis how skeletal muscle EVs exert their functions.
Type‐2 Diabetic/Obese Patients with Associated Vascular Disease Have Increased Circulating Acetylated HMGB1
Authors: Liu Zeyu , Raviprakash Srishti , Williamson Kelly , Mota Roberto I. 
Published: May 2022

IntroductionType‐2 Diabetes (T2D) is highly associated with peripheral artery disease (PAD), with both PAD and T2D highly associated with chronic inflammation. High mobility group box 1 (HMGB1) is key in regulating both inflammation and diabetes. Acetylated HMGB1 is the extracellularly released form of HMGB1 from post translational modifications (PTM) that occur after nuclear HMGB1 turnover. Acetylated HMGB1 has significant cytokine/chemokine activity. Plasma HMGB1 high molecular weight (HMW) complexes have been identified in the literature as PTM in chronic inflammation. Glycoprotein cytokine Osteoprotegerin (OPG) is a known marker for vascular disease, with research showing that high levels of OPG correlate tightly with increased levels of HMGB1 in T2D patients with PAD. We hypothesize that obese, T2D patients will display elevated levels of circulating acetylated HMGB1 in HMW complexes and that we will find associated high circulating OPG in same patients.MethodsPlasma was collected from 25 male/female adult patients diagnosed with chronic T2D sorted by sex, obese or lean, w/ and w/o vascular disease. Healthy and trauma patients plasma was used as experimental controls. Diluted plasma (1:25) were denatured in SDS page gels and ran for immunoblots with luminescence using the primary antibodies HMGB1 (abcam ab18256) and acetyl‐HMGB1 (Aviva OASG03545). OPG quantification was measured in same plasma samples as above by ELISA (Thermofisher TNFRSF11B) following the manufacturer protocol. Data is represented as mean±SEM. A total of 5‐10 samples per experimental group were analyzed, representative images of immunoblots are shown. Statistical analysis was performed using t‐test and ANOVA with Tukey’s correction.ResultsImmunoblot analysis showed that there is an increase in plasma HMGB1 in plasma samples from T2D, obese patients with vascular disease; predominantly acetylated HMGB1 in HMW complexes (>100kDa) were present within obese‐T2D patients compared to both healthy patients and trauma patients. Since we wanted to understand if vascular disease was associated in plasma samples from these patients, we performed OPG quantification. When we analyzed our cohort, results showed increased OPG in obsese‐T2D patients compared to lean T2D patients with a clear difference by sex (males>females). Obese T2D males with arterial injury have significantly higher circulating OPG compared to lean T2D males with arterial injury.ConclusionsThe drastically higher HMW complexes found uniquely in obese/T2D patients identifies acetylated HMGB1 as the predominant circulating form of HMGB1 in obese/T2D patients. Findings show there might be some association of elevated circulating OPG and acetylated HMGB1 as potential biomarkers. Future studies will focus on understanding the role of acetyl‐HMGB1 in the disease progression of T2D/obese patients with vascular disease.
Craniofacial Bone Mineral Density at Muscle Attachment Sites in Mice with Osteogenesis Imperfecta
Authors: Murabito Lauren , Miller Courtney A. , McBride Alexandra H. , Organ Jason M. , Menegaz Rachel 
Published: May 2022

Osteogenesis imperfecta (OI) is a genetic disease commonly caused by mutations in the COL1A1 or COL1A2 gene affecting the production of type I collagen, a major component of bone that determines the toughness of bone, or its resistance to fracture. Thus, patients with OI present with brittle bones and higher rates of skeletal fractures. Additionally, defects in the collagen framework of developing bones impair their biomineralization, further compromising bone mechanical properties. Previous work in our lab investigated BMD at cranial locations which experience compressive forces during feeding (e.g. joints). Muscles also contribute to cranial growth through tensile forces exerted on bone during contraction, but the relationship between muscle function and bone quality in OI is unclear. The aim of this study is to quantify the BMD in a mouse model of OI, with a focus on skeletal sites associated with cranial musculature. We hypothesize that the mice with OI will have lower BMD in skeletal areas associated with muscle attachment compared to their unaffected littermates.To test our hypothesis, we used the homozygous recessive OI murine (OIM) mouse (B6C3Fe a/a‐Col1a2oim/J), a model for the severe type III OI seen in humans. Adult (16 weeks old) OIM mice (n=11) and wildtype (WT, n=13) littermates were CT scanned, and BMD was measured using the Bruker CTAnalyzer software in 7 regions of interest encompassing muscle attachment sites on the mandible, zygomatic arch, and parietal bone. For each region, BMD values were compared between genotypes by a Kruskal‐Wallis test.OI mice had significantly lower BMD at the coronoid process (p=0.007), parietal bone (p=0.031), and posterior mylohyoid line (p=0.007) compared to WT mice. No significant differences were found in BMD at the gonial angle or zygomatic arch. This suggests a disproportionate weakness of the temporalis and mylohyoid muscles in mice with OI, but not in the masseter muscle. While type I collagen defects impact the structural organization of skeletal muscle, it is possible that masseter muscle function is preserved in OIM mice because of its key roles in rodent feeding and postural control of the jaw. The imbalance between temporalis and masseter muscles may contribute to the prevalence of dental malocclusions in human patients with type III OI.The findings of this study support further exploration of specific muscle groups (e.g. jaw adductors/temporalis, hyoid muscles/mylohyoid) and efforts to improve their functions during feeding, as a way of recovering facial growth and preventing dental malocclusions in OI. A better appreciation of muscle‐bone interactions in the skull and their effect on bone quality can potentially be used to inform oro‐motor therapies to improve feeding function, muscle strength, and facial bone quality for patients with OI.