bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Binuclear Transport Center of ZupT: A Zinc Transporter from E. coli
Authors: Roberts Cameron , Mitra Bharati 
Published: April 2020

ZupT is a member of the ZIP family of transporters which are responsible for increasing the intracellular concentration of zinc from E. coli to humans. Defects in these transporters are linked to zinc deficiency, cancer, and cardiovascular disease. The mechanism of transport in this family is still not well‐understood, partly due to the bottleneck in the expression and purification of these integral membrane proteins. Recently a structure was solved of a ZIP transporter from B. bronchiseptica. The structure revealed a cadmium binuclear metal transport site, which is relatively uncommon for metal transporters. Using the crystal structure, a homology model was built for ZupT which shares 32% identity and a conserved binuclear center. In the ZupT model the metal ions are separated by roughly 4.2 A and coordinated by a bidentate Glu123 (orange). His148 and Glu152 round out site 1 (red) and Asn120, Asn149, and Glu181 site 2 (yellow). In this work, solution studies including competitive metal titrations were carried out with purified ZupT. Our results show that ZupT binds one equivalent of zinc, its primary substrate, at physiological concentrations. In contrast, it binds two equivalents of iron, a secondary substrate. Site directed mutagenesis was carried out to determine which residues are important in coordinating zinc and iron. Zinc and iron both bind to a ‘primary’ transport site (site 1‐red), while iron also binds to a secondary site (site 2‐yellow). Activity assays were used to determine the relationship between binding and function. ZupT showed transport activity with both iron and zinc; additionally, it was observed that iron positively regulated zinc activity. Our results also indicated that the highly conserved residue His119 (blue) is not essential for binding of either zinc or iron, but is important for overall transport activity, probably by facilitating metal release. Overall these results show that ZIP transporters have a single zinc transport site which can be regulated by iron binding to an adjacent secondary site.Binuclear metal center of ZupT.Utilizing the crystal structure of ZIPB from B. bronchiseptica a homology model was made with the SWISS‐MODEL server. Visualization of the binculear center was carried out with UCSF CHIMERA.Figure 1
Hepatic Tumour Cell Reduction with Quanta Energy – a Technological Exploration and Expanded Mathematical Model.
Authors: Rahman Michael S. , Einstein George P. , Tulp Orien P. , Koynk Carla 
Published: April 2020

Manley (2014) developed several mathematical models based on the CSC concept that not only explain the growth of a tumour, but also its demise based on the applied radiation energy. If a population A of CSCs divide asymmetrically, they will yield one daughter cell of type A and one cancer cell of type B. The speed with which new type B cells emerge depends on the number of A cells – if there are 100 A cells, 100 B cells will emerge in a given unit of time; if the number of A cells is 200, 200 B cells will emerge in the same time window, twice as many as before. Therefore, one can model the growth or increase in numbers of B cells. which allows us to model the growth of B as differential equation. The basic form of this model has the number of CSCs (type A) constant, while the increase in differentiated tumor cells (type B) grows.In other words, the population of cancer cells grows linearly. One can introduce further parameters into this simple equation and distinguish between cancer stem cells A, differentiated tumour cells B and healthy cells H.If we further irradiate cells with energy from X‐rays, proton beams, or light of visible spectrum, photodynamic therapy (PDT) or invisible UV; there will have likely different effects on each cell type A, B and H. Manley (2014) established the following formulas with r1, r2 and r3 as specific effects of radiation or PDT on each of the cell types, the equation in diagram 1 is established.Where S is the desired number of stem cells, and M1 and M2 are the carrying capacity of B and H cells, respectively (Manley, 2014).If we use different assumptions for the parameters in these equations, we can come up with different long term behaviors of the cell lines. The most desirable situation is one in which the number of healthy cells remains constant, while the CSCs and tumor cells are reaching a number near zero. This would mean shrinkage of the tumor without affecting healthy tissue.The different ri values are effects the radiation energy will have on each cell type; one should be able to derive the effect from the wavelength or frequency of the used radiation, but the relationship is not entirely clear. The energy of one quantum of energy can be described using the classic equation of light and quanta.with c as the speed of light (c = 2.998 × 108 m/s) and h as Planck constant (h = 6.626 × 10−34 m2 kg/s. Since the above equations for dA/dt, dB/dt and dH/dt are all describing the total change in cell numbers as influenced or determined by time, r1, r2 and r3 must represent the number of cells that die off by application of energy with radiation of a certain frequency or wavelength in the visible or invisible spectrum of light. Importantly, cells may not be damaged if the radiation that we use to illuminate them with has a frequency that is too low.Simple model of linear tumour growthFigure 1GP Einstein Sigma Quanta tumor growth and development modelFigure 2
Fumarate Reduced Mean Arterial Blood Pressure and Improved Renal Outcomes in DOCA/salt Hypertension
Authors: Edosuyi Osaze , Igbe Ighodaro , Choi Myung , Oyekan Adebayo 
Published: April 2020

The mitochondria, through the tricarboxylic (TCA) acid cycle, converts organic molecules into energy that drives intricate renal blood‐pressure regulatory functions. The TCA cycle intermediaries regulate the metabolic state of the mitochondria and by extension, the redox status of the cell. Recently, fumarase, a major TCA cycle enzyme that converts fumarate to malate, has been linked to L‐arginine metabolism and implicated in the pathophysiology of Dahl‐salt sensitive hypertension. This study aimed to investigate the role of fumarase and L‐arginine in deoxycorticosterone (DOCA)/salt hypertension, a non‐genetic model of hypertension. Hypertension was induced in uninephrectomised rats with DOCA (25 mg/kg) + 1 % NaCL. Mean arterial blood pressure was measured in conscious rats via carotid cannulation. Biochemical and western blot analyses were carried out on homogenized kidney fractions. Fumarate reduced arterial blood pressure (198±5 vs 167±7 mmHg, P<0.01). There were no changes in urinary output and sodium excretion in fumarate‐treated DOCA rats (P>0.05). Superoxide dismutase and catalase activities were higher in the cortex, but not the medulla of DOCA‐hypertensive rats and were exacerbated by fumarate (P<0.01). There was a selective increase in catalase activity in the medulla of fumarate‐treated DOCA‐hypertensive rats (P<0.01). Fumarate also reduced urinary protein excretion in DOCA‐hypertensive rats (23.6 %, P<0.05). Fumarate evoked a selective increase in nitric oxide in the medulla (34.1±2.9 vs 47.3±2.8 nM/μg, P<0.05) of DOCA‐hypertensive rats. Consistent with this, Fumarate also tended to increase arginase activity in the cortex but not the medulla of DOCA‐hypertensive rats. However, there was an increase in expression TGF‐β, an index of fibrosis, in DOCA‐hypertensive rats, which was paradoxically increased in fumarate‐treated rats (P>0.05). In conclusion, fumarate attenuated the hypertension and renal injury but not the fibrosis in DOCA/salt hypertension by mechanisms involving selective reduction of L‐arginine metabolism in the medulla and amelioration of renal oxidative stress.Support or Funding InformationNational Institute of Health grant HL03674 and G12 MD0076056Mean arterial blood changes after 21‐day administration of fumarate to DOCA/salt induced hypertensive rats. Control (n=8), DOCA (n=8), fumarate (n=8). a=P<0.001 when compared to control; b=P<0.01 when compared to DOCA group.Figure 1Nitric oxide produced in homogenized kidneys (cortex and medulla) of DOCA salt induced hypertensive rats after 21‐day fumarate (1 mg/kg) administrationa=P<0.05 when compared to control.Figure 2
Prolotherapy upregulates Fibroblast Growth Factor‐2 (FGF‐2) and Insulin‐like Growth Factor‐1 (IGF‐1) mRNA which may increase chondrocyte proliferation and mediates osteoarthritic inflammation
Authors: Johnston Elisha Daniel , Emani Chandrakanth 
Published: April 2020

Osteoarthritis is a highly prevalent disease that is progressively generated and has no known cure. Current treatments, such as opioids, have highly adverse effects. A more effective treatment can be the form of proliferative therapy (prolotherapy), a complementary medical intervention with promising clinical evidence. The purpose of the present study is to investigate the molecular basis of prolotherapy as a treatment option for osteoarthritis. Specifically, the study investigates whether prolotherapy upregulates proliferation‐enhancing cytokines (FGF‐2 and IGF‐1) in chondroprogenitor cells. In vitro stimulation of chondroprogenitor cells was carried out with prolotherapy treatment followed by qRT‐PCR quantification to determine the mRNA expression of the genes FGF‐2, IGF‐1, CCND‐1, TGF‐β1, AKT, STAT1, and BMP2. Results indicated a likely upregulation of cytokines that enhanced cell proliferation in chondroprogenitor cells with statistically higher levels of expression for FGF‐2 (p<0.001, p<0.05, p<0.01), IGF‐1 (p<0.001), and CCND‐1 (p<0.05) relative to controls. Results in conjunction with prolotherapy and molecular‐focused inflammation theory suggest that prolotherapy has the potential to transform chondroprogenitor cells that are dysregulated by osteoarthritic inflammation. This would contribute to cartilage degradation back into chondroprogenitor cells that proliferate and balance catabolic/anabolic roles to repair and maintain cartilage.P2G (1.5%) upregulates FGF‐2 mRNA expression in chondroprogenitor cells. A Exploratory Trial 1 suggests P2G effects FGF‐2 gene expression (Welch’s one‐way ANOVA, p<0.01). B Confirmatory Trial 2 indicates that relative to time‐specific controls, chondrocytes treated with P2G exhibit increased levels of FGF‐2 at hours 24, 30, and 38 (Welch’s t‐tests yield p<0.001, p<0.05, and p<0.01). qRT‐PCR quantifies mRNA expression. NS p>0.05, * p<0.05, ** p<0.01, *** p<0.001Figure 1P2G (1.5%) upregulates CCND‐1 (Cyclin D) mRNA expression at hour 30 in chondroprogenitor cells in the confirmatory Trial 2 (directional Welch’s t‐test yields p<0.05). CCND‐1 mRNA upregulation at hour 30 suggests subsequent entry chondroprogenitor cell proliferation. qRT‐PCR quantifies mRNA expression. NS p>0.05, * p<0.05, ** p<0.01, *** p<0.001Figure 2
Oligomerization of VDAC1 promotes mtDNA release and lupus‐like disease
Authors: Kim Jeonghan 
Published: April 2020

Mitochondrial stress releases mitochondrial DNA (mtDNA) into the cytosol, triggering immunostimulatory pathways such as the type‐Ι interferon response. Pores formed in the outer membrane of mitochondria (OMM) can promote mtDNA release; however, the identity of such a pore in living cells is not well characterized. Here, we provide genetic and biochemical evidence demonstrating that the oligomerization of voltage‐dependent anion channel 1 (VDAC1) in the OMM promotes mtDNA release and triggers the type‐Ι interferon response in living cells. In addition to forming OMM pores, VDAC1 interacts with mtDNA via its positively charged residues in the N‐terminal domain and increases both VDAC1 oligomerization and type‐Ι interferon response. VBIT‐4, which inhibits VDAC1 oligomerization, decreases mtDNA release, type‐Ι interferon signaling, neutrophil extracellular traps, and disease severity in a mouse model of systemic lupus erythematosus. These findings suggest that VDAC1 oligomerization is a potential therapeutic target for lupus.Support or Funding InformationSupported by the Intramural Research Program of the National Heart Lung and Blood Institute, National Institute of Arthritis and Musculoskeletal and Skin Diseases, and National Institute of Allergy and Infectious Diseases and by a grant from the National Institute for Biotechnology in the Negev (V.S.‐B.) and by a grant of the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health and Welfare, Republic of Korea (grant number HI14C1176) and a grant from the KRIBB Research Initiative Program (Korean Biomedical Scientist Fellowship Program), Korea Research Institute of Bioscience and Biotechnology, Republic of Korea.VDAC oligomers form mitochondrial pores to release mtDNA fragments.Figure 1
Discovery and Quantitative Analysis of Nuclear Factor Erythroid 2‐Related Factor (Nrf2) Activators in Maca (Lepidium meyenii) Using the Synthetic Macamides
Authors: Hahn Dongyup , Lee Mijeong , Jeong Hyeyoon , Kim Jaewon , Kim Minyoul 
Published: April 2020

Maca (Lepidium meyenii) is a biennial herbaceous plant of family Brassicaceae with various bioactivities such as reproductive health enhancement, antifatigue, antioxidation, neuroprotection, anticancer, hepatoprotection, and immunomodulation. But studies on molecular mechanisms beyond the phenomenal physiology are rare because the putative bioactive chemical constituents, macamides, a series of unique N‐benzylamides with long‐chain fatty acids are difficult to isolate from the natural maca specimen. In this study, ten macamides were chemically synthesized to investigate biological mechanisms underlying the various bioactivities. Three of macamides exhibited moderate nuclear factor erythroid 2‐related factor 2 (Nrf2) activation was observed from cell‐based assay (EC50 7.3–16.5 μM), which might account for some of the known bioactivities of maca. Quantitative analysis of macamide compositions in maca specimen using LC‐MS/MS was followed, which revealed the macamides with Nrf2 activation activity consist the majority among macamides contained in maca. The difference in composition of macamides among three cultivars, yellow, red and black macas was also examined.Support or Funding InformationThis research was supported by the National Research Foundation of Korea (NRF) grant funded by the Korean government (2019R1F1A1051041).Chemical synthesis of macamidesFigure 1
Withdrawn: Invisible State of MdmX and Design of its Inhibitors
Authors: Cheng Xiyao , Huang Yongqi , Su Zhengding 
Published: April 2020

Withdrawal: Xiyao Cheng, Yongqi Huang, and Zhengding Su. Invisible State of MdmX and Design of its Inhibitors. The FASEB Journal. 34:s1. doi: 10.1096/fasebj.2020.34.s1.09193.The above abstract, published online on April 20, 2020, in Wiley Online Library (wileyonlinelibrary.com), has been withdrawn by agreement between the authors, FASEB, and Wiley Periodicals, Inc. The abstract has been withdrawn at the request of the authors.
Cyp2j5‐gene Deletion Affects on Acetylcholine and Adenosine‐induced Relaxation in Mice: Role of Angiotensin‐II, CYP‐epoxygenase and Omega‐hydroxylase inhibitors
Authors: Agba Stephanie O. , Hanif Ahmed , Edin Matthew L. , Zeldin Darryl C. , Mohammed Nayeem A. 
Published: April 2020

Previously, we showed vascular endothelial overexpression of human‐CYP2J2 enhances coronary reactive hyperemia in Tie2‐CYP2J2 Tr mice, but we did not see the response in CYP2J‐epoxygenase knockout mice. Therefore, we hypothesized that Cyp2j5‐gene deletion affects acetylcholine‐, NECA (adenosine)‐and CGS 21680 (adenosine A2A receptor)‐induced relaxation and their response is partially inhibited by angiotensin‐II (Ang‐II) and ω‐hydroxylase inhibitor in mice. ACh‐induced response was tested with N‐(methylsulfonyl)‐2‐(2‐propynyloxy)‐benzenehexanamide (MS‐PPOH, CYP‐epoxygenase inhibitor; 10–5M) and Ang‐II (10‐6M). In Cyp2j5−/− mice, ACh‐induced relaxation was different from C57Bl/6 mice, at 10‐5 M (76.1 ± 3.3 vs. 58.3 ± 5.2, P<0.05). However, ACh‐induced relaxation was not blocked by MS‐PPOH in Cyp2j5−/−: 58.5 ± 5.0%, P>0.05, but blocked in C57Bl/6: 52.3 ± 7.5%, P<0.05, and Ang‐II reduces ACh‐induced relaxation in both Cyp2j5−/− and C57Bl/6 mice (38.8 ± 3.9% and 45.9 ± 7.8, P <0.05). In addition, NECA‐induced response tested with Ang‐II and 20‐HETE inhibitor, dibromo‐dodecenyl‐methylsulfimide (DDMS, 10‐5M). In Cyp2j5−/− mice, NECA‐induced response was not different from C57Bl/6 mice at 10‐5M (23.1 ± 2.1 vs. 21.1 ± 3.8, P>0.05). However, NECA‐induced response was reduced by Ang‐II in both Cyp2j5−/− and C57Bl/6 mice (−10.8 ± 2.3% and 3.2 ± 2.7, P <0.05), and the reduced response due to Ang‐II was partially rescued by DDMS in both Cyp2j5−/− and C57Bl/6 mice (at 10‐5M, A2AAR‐agonist, CGS‐21680, 15.8 ± 3.4% and 13.0 ± 2.3, P <0.05). Data suggest that ACh‐induced relaxation in Cyp2j5−/− mice depends on nitric oxide (NO) but not CYP‐epoxygenases, and the NECA‐induced different response in male vs. female Cyp2j5−/− mice when Ang‐II treated.Support or Funding InformationNational Institutes of Health (HL‐114559) to M. A. Nayeem and the Intramural Research Program of the National Institute of Environmental Health Sciences (Z01 ES025034) to D. C. Zeldin supported this work.Flow diagram: Comparison between Cyp2j5−/− vs. C57Bl/6 mice treated with Acetylcholine, Acetylcholine + MS‐PPOH, Acetylcholine + Ang‐II, NECA, NECA + Ang‐II and CGS 21680 + DDMS + Ang‐II.Figure 1
Effects of Jwagwieum on Renal Dysfunction in Ischemia/Reperfusion‐induced Acute Renal Failure in Rats
Authors: Na Se Won , Kim Hye Yoom , Ahn You Mee , Hong Mi Hyeon , Jang Youn Jae , Lee Ho Sub , Kang Dae Gill 
Published: April 2020

Jwagwieum (
, JGE), a Korean traditional medicine, has been used for treatment of kidney‐yin deficient (
) syndrome. However, there is no pharmacological study of JGE on ischemia/reperfusion‐induced acute renal failure (ARF). Therefore, the objective of this study was to evaluate whether JGE ameliorates renal function and its related mechanism in acute kidney injury‐induced by ischemia/reperfusion in rats. In this experiment, 5‐week‐old male Sprague Dawley (SD) rats were used under 12 h light and dark cycle. After a week of adaptation, they were randomly divided into following groups. 1) Cont. (Control), 2) ischemia/reperfusion injury (ARF), 3) ARF + JGE100 (100 mg/kg/day), 4) ARF + JGE200 (200 mg/kg/day). The ARF group was subjected to reperfusion after inserting a clip into the renal artery for 45 min. ARF + JGE100 group was orally administered with JGE 100 mg/kg/day after ischemia/reperfusion surgery, and ARF + JGE200 group was orally administered with JGE 200 mg/kg/day for 4 days, respectively. As a result, JGE suppressed the increase of kidney size in the acute renal failure animal model and alleviated polyuria symptoms. In addition, solute‐free water reabsorption (TcH2O) and creatinine clearance (Ccr), indicators of glomerular filtration rate, were significantly increased in JGE group compared with those of ARF rats. In ARF group, blood urea nitrogen (BUN)/creatinine (Cr) and lactate dehydrogenase (LDH) were significantly increased compared to the control group, respectively, while they were significantly decreased in JGE group. The present results also showed that JGE improved the morphological aspects of renal tissue and ameliorated fibrosis in ARF group. Inflammation is one of the main causes of ischemic acute kidney injury and was inhibited by JGE, which was confirmed by the NLRP3 inflammasome decline.The present results showed that JGE has protective effects against inflammatory process via inhibition of NLRP3 signal pathway, which is associated with improvement of renal function in rats with ARF. These results suggest that JGE has a therapeutic effect on acute kidney injury‐induced by ischemia/reperfusion in rats.Support or Funding InformationThis work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIP) (2017R1A5A2015805)(2015M3A9A5030620)(2017R1A2B4009884).
A MiniG Protein Recruitment Assay for the Histamine Receptor Family to Functionally Characterize Histamine Receptor Ligands
Authors: Hoering Carina , Seibel Ulla , Littmann Timo , Graetz Lukas , Strasser Andrea , Bernhardt Guenther 
Published: April 2020

In addition to the determination of binding affinities, the functional characterization of biologically active compounds is a prerequisite in drug discovery in order to identify lead structures and to optimize potential scaffolds regarding their mode of action and potency. Therefore, our aim was to develop a high‐throughput screening (HTS) compatible assay procedure yielding functional parameters of histamine receptor ligands at a proximal level avoiding signal amplification.For this purpose, we applied the NanoLuc split‐luciferase technology
[1] to the histamine receptor subtypes H1, H2, H3 and H4 and the corresponding G protein surrogates miniGq, miniGs and miniGi
[2]. The small fragment of the NanoLuc (NlucC) was fused to the C‐terminus of the respective receptor to retain the integrity of the receptors regarding binding affinities and intracellular signalling, whereas the large fragment of the NanoLuc (NlucN) was N‐terminally fused to the miniG proteins (see fig.
1). Upon activation by a ligand the corresponding miniG protein is recruited to the receptor which then leads to the formation of a functional NanoLuc enzyme enabling the registration of a concentration dependent luminescence signal (see fig.
1).Compared to reported data, standard ligands for histamine receptors
provide appropriate potencies, intrinsic activities and modes of action.By radioligand saturation binding experiments it could be shown that there was no alteration in the binding affinities of the radioligands due to the minor receptor modification by the C‐terminally fused NlucC. Additionally, this assay principle should be appropriate for HTS assessed by the investigation of the Z′ factor, which was in an excellent range for all histamine receptor subtypes (0.5 < Z′ < 1).From these results we conclude that the application of the presented methodology is an excellent approach to be implemented in routine and detailed functional characterisation of putative histamine receptor ligands synthesized in our group. Moreover, the assay offers an opportunity to draw conclusions on biased signalling of investigated ligands in combination with the results obtained in β‐arrestin recruitment assays
[3].Support or Funding InformationThis project is funded by the DFG research training group GRK1910.Schematic illustration of the assay principle.Two complementing NanoLuc fragments fused to the receptor and the mini G protein form a functional NanoLuc upon activation of the GPCR.Figure 1