bluuebunny/crossref_metadata_2025_split huggingface dataset:
Muscle function recovery after acute lung injury is impaired in aged mice, linked in part, to metabolic alterations attributed to MuRF1 by quantitative targeted metabolomics analysis of acyl‐carnitines
Authors: Ilaiwy Amro , Parry Traci L. , Bain James R. , Muehlbauer Michael J. , Willis Monte S. , Files Clark 
Published: November 2023

Most survivors of acute lung injury suffer from substantial physical complications months after the initial lung injury resolves, and persistent muscle weakness is a very common sequelae of lung injury. Using a mouse model of LPS‐induced lung injury [acute lung injury (ALI) mice], we found that old (20 month) ALI mice failed to recover skeletal muscle function 10 days after injury compared to young (6 month) ALI control mice. Non‐targeted GC‐MS analysis of old ALI gastrocnemius muscle vs. vehicle treated controls identified 18 significantly different metabolites, including the long chain fatty acid linoleic acid (1.75 fold increased), indicative of altered linoleic acid metabolism. Our laboratory recently identified that muscle ring finger protein 1 (MuRF1) inhibits fatty acid oxidation in vivo via mono‐ubiquitination of peroxisome proliferator‐activated receptor alpha. Moreover, we've identified that MuRF1−/− mice are resistant to LPS‐induced muscle atrophy. Western analysis of old and young ALI muscles for MuRF1 found significantly greater expression of MuRF1 3 days following lung injury in old ALI mice, leading us to hypothesize that old ALI muscles had inhibited fatty acid oxidation. We next performed targeted metabolomics analysis of fatty acid intermediates (acyl‐carnitines) and amino acids of the affected muscles. At day 10, we identified 13 significantly altered acyl carnitines (by t‐test, p<0.05) compared to vehicle treated controls (0.27–1.14 fold). We also identified that total long chain acyl‐carnitines were almost decreased by half, paralleling the acyl‐carnitine profile recently published in MuRF1−/− hearts. Together, these studies demonstrate for the first time age‐associated susceptibility of ALI‐induced muscle wasting and characterize the metabolic alterations in fatty acid metabolism, suggesting MuRF1's role in regulating fatty acid metabolism, which may contribute to impaired skeletal muscle functional recovery.Support or Funding InformationThis work was supported by the National Institutes of Health (R01HL104129 to M.W.), the Leducq Foundation Transatlantic Networks of Excellence (to M.W.), the Claude D. Pepper Older Americans Independence Center (P30AG21332 to D.C.F and O.D.), and the American Thoracic Society Foundation (D.C.F.).
Novel lymphoid targeted prodrugs of the glutamine antagonist DON for the treatment of hematological malignancies
Authors: Jancarik Andrej , Rais Rana , Alt Jesse , Tenora Lukáš , Krecmerova Marcela , Cheng Chih , Pommier Elie , Rojas Camilo , Powell Jonathan , Majer Pavel , Slusher Barbara 
Published: November 2023

ObjectiveThe glutamine antagonist 6‐diazo‐5‐oxo‐l‐norleucine (DON) has shown robust efficacy in both lymphoma and leukemia animal models and exploratory clinical studies. DON broadly blocks glutamine utilizing reactions critical for the synthesis of nucleic acids/proteins and the generation of alpha‐ketoglutarate for energy metabolism. Unfortunately its severe toxicity hampered its development. Using a prodrug approach, we aimed to synthesize lymphoid‐targeted prodrugs of DON designed to circulate intact in blood and preferentially activate inside PBMCs affording reduced systemic exposure and improved tolerability, while maintaining its robust efficacy.methodsSynthetic efforts for DON prodrugs were particularly challenging due to DON's inherent instability. We found it was imperative to mask both the carboxylate as well as the amine functionality of DON to achieve sufficient chemical stability. We concealed the carboxylate and amine with various alkyl esters (e.g. methyl, ethyl and isopropyl), amino acids (e.g. leucine, alanine), and other bulkier promoeities used in FDA approved drugs (e.g. POC, POM, ODOL). We then evaluated the prodrugs for in vitro plasma stability across multiple species (mouse, dog, monkey, pig, human). Ex vivo whole blood experiments were further conducted to determine selective conversion in PBMCs. Lastly in vivo experiments using pigs were carried out and PBMC to plasma ratios were calculated.resultsOur novel DON prodrugs showed species specific plasma stability with rodents metabolizing the prodrug more rapidly compared to higher species. Of the various modifications on the carboxylate, the isopropyl ester afforded significant lymphoid‐targeting in pigs, monkeys, dog, and humans. In human whole blood ex vivo studies several of the isopropyl ester containing prodrugs delivered DON into PBMCs versus plasma with a >30‐fold differential, in contrast to DON which delivered a small percentage (<10%) of drug to PBMCs. Lastly in vivo studies in pigs confirm this PBMC selectivity with favorable pharmacokinetics.conclusionsWe have successfully synthesized lymphoid‐targeted DON prodrugs which deliver >30‐fold more DON to lymphoid cells in human whole blood versus plasma. Comparative efficacy and toxicity studies are ongoing in lymphoma animal models. These novel prodrugs may pave a path for the development of a new and safer therapeutic for hematological malignancies.
Challenges and solutions to developing an automated high‐throughput/high‐content screening platform for the ‘neglected’ schistosome parasite
Authors: Caffrey Conor , Chen Steven , Suzuki Brian , Singh Rahul , Arkin Michelle 
Published: November 2023

Automated high‐throughput (HT) and/or high‐content (HC) screening platforms are now well‐established in early drug discovery, including for many parasitic organisms causing ‘neglected tropical diseases.’ However, for the helminth (worm) parasites responsible for a range of globally prevalent and debilitating infections, such systems are still in their infancy. Using the schistosome, a flatworm parasite that infects over 200 million people, as a particularly challenging example, we developed an HTS/HCS platform to quantify the parasite's responses to chemical insult. I will highlight the hurdles encountered to standardize the preparation and handling of the parasite, and how we interfaced the parasite with an automated instrumentation environment. I will then discuss the approaches taken regarding image acquisition, object segmentation and tracking, and feature extraction. We tested the platform with seven anti‐schistosomal drugs to measure a range of static and dynamic parasite responses as a function of time and concentration. We designed a user interface to visualize and interrogate the data, which are maintained in a customized database. We combined the high‐dimensional data into a single metric output suitable for primary first‐pass screening: data from screening campaigns of high‐value small molecule collections will be presented. The new platform increases throughput, improves rigor and will support the identification of drugs, drug targets and mechanisms of action.Support or Funding InformationSupported by NIH R01AI089896 and R21AI107390.
Modulation of GRP78‐CHOP‐JNK‐Caspase12 pathway by Chinese herb medicine in atrophied soleus muscle induced by tail suspension
Authors: Zhang Shu , Wu Shiwen , Yuan Ming , Wang Jingyu , Yang Honghui 
Published: November 2023

ObjectiveMicrogravity could induce a series of changes on the physiological state in humans and animals, among which skeletal muscle atrophy is striking. The countermeasure of skeletal muscle atrophy mainly relies on the physical exercise which need expensive and heavy special apparatus in space. However, complementary medicine such as Chinese herb medicine may have distinctive effects on the countermeasure of skeletal muscle atrophy induced by microgravity or simulated microgravity. This study was carried out to test the effects of Xuefuzhuyu capsules(XFZY), an Chinese medicine known to improve the blood supply of tissue, on the skeletal muscle atrophy induced by simulated microgravity.MethodsRats fed the XFZY were tail‐suspended for 30 days(XFZY group), and the muscle mass and cross‐sectional area (CSA) were then compared with those of the suspended rats(TS group) and control rats(Con group). The protein expression and phosphorylation level in soleus muscle were measured by Western blot.ResultsTreatment with the XFZY attenuated the loss of muscle mass and CSA induced by tail suspension in rats. Furthermore, western blot analysis showed that the phosphatidyl‐inositol 3‐kinase/Phospho‐AKT were both down‐regulated after 30d tail suspension, which were reversed by treatment of XFZY. Meanwhile, Tai suspension increased the expression of GRP78, CHOP, phospho‐JNK level and Caspase 12 in soleus muscle, and XFZY alleviated the increased protein expression in soleus muscle induced by tail suspension.ConclusionOur results suggested that the XFZY can partially reverse the effects of tail suspension on soleus muscle and maybe target on the endoplasmic reticulum stress (ERS) system by regulating GRP78‐CHOP‐JNK‐Caspase12 pathway.
Structure‐based ligand discovery for nutrient transporters
Authors: Schlessinger Avner 
Published: November 2023

Alterations in cell metabolism support rapid growth and proliferation of cells ‐ key hallmarks of cancer. Solute Carrier (SLC) transporters are membrane proteins that transport solutes such as metabolites and drugs across membranes, and play a major role in mediating nutrient delivery in reprogrammed cancer metabolism networks. Here, we describe a structure‐based discovery approach to identify small molecule modulators for key nutrient SLC transporters. In particular, we use homology modeling, virtual screening, and experimental testing to identify small molecule ligands including drugs, metabolites, and lead‐like molecules. We target a variety of nutrient transporters such as the amino acid transporters LAT‐1 and ASCT2, and the glucose transporter GLUT1, which function cooperatively in cancer metabolism and are highly expressed in the blood‐brain‐barrier (BBB). Our results may explain some of the pharmacological effects (i.e., efficacy and/or side effects) of known drugs via polypharmacology, and rationalize the enhanced brain permeability of drug‐like molecules. Finally, our top hits inhibited proliferation of various cancer cell lines via distinct molecular mechanisms, providing useful chemical tools to characterize reprogrammed metabolic networks, as well as a framework for developing efficacious drugs against these key targets and other SLC transporters.Support or Funding InformationR01 GM108911
Phorbol Myristate Acetate Suppresses Breast Cancer Cell Growth via Down‐regulation of P‐Rex1 Expression
Authors: Jiang Haihong , Wong Chuu‐Yun A. , Abel Peter W. , Scofield Margaret A. , Xie Yan , Wei Taotao , Tu Yaping 
Published: November 2023

Phosphatidylinositol‐3,4,5‐trisphosphate‐dependent Rac exchange factor 1 (P‐Rex1) is highly overexpressed in estrogen receptor‐positive and ErbB2‐overexpressing human luminal breast tumors. The present study was to investigate the role and mechanism underlying phorbol‐12‐myristate‐13‐acetate (PMA), a protein kinase C (PKC) activator, down‐regulation of P‐Rex1 in breast cancer cells. P‐Rex1 mRNA and protein expression were determined by real‐time PCR and western blot analysis, respectively. Cell growth was analyzed by counting cells number and 5‐bromo‐2′‐deoxyuridine (BrdU) cell proliferation assay. The translocation of PKC was examined by immunofluorescence staining. Our results showed that treatment with PMA caused a time‐ and concentration‐dependent decrease in P‐Rex1 protein levels in breast cancer MCF‐7 and BT‐474 cells with a maximum reduction of 87.2 ± 1.1 and 57.0 ± 8.6%, respectively, at a concentration of 10 ng/ml. Interestingly, either PMA treatment or silence of P‐Rex1 by siRNA inhibits MCF‐7 and BT‐474 cell growth by 60–80%, which can be attenuated by restoring P‐Rex1 expression. MCF‐7 and BT‐474 cells express PMA‐sensitive conventional (α and β) and novel (δ, ɛ and η) PKC isoforms. Immunofluorescence staining analysis of breast cancer cells showed translocation of PKCs from cytosol to the plasma membrane and/or nucleus, an indication of their activation upon PMA stimulation. The PKC general inhibitor Gö6983, but not the conventional PKC isoform inhibitor Gö6976, blocked PMA inhibition of P‐Rex1 expression and breast cancer cell growth. Expression of a constitutively active PKCɛ mutant suppressed P‐Rex1 expression and breast cancer cell growth, which was blocked by Gö6983 treatment. In contrast, expression of wild‐type PKCɛ, wild‐type or constitutively active PKCδ and PKCη had no effect. Together, our data suggest that PMA suppresses breast cancer cell growth via P‐Rex1 down‐regulation and PKCɛ activation plays a key role in this regulation. Our study may provide a novel strategy for development of chemotherapeutic agents for P‐Rex1‐overexpressing breast cancer patients that develop resistance to anti‐estrogen and/or anti‐ErbB2 therapies.Support or Funding InformationSupported by grants from NIH (5P20GM103489 and R01HL116849), Nebraska State LB595 research program and the National Natural Science Foundation of China (31100595 and 31300683).
Differential Specificity of the Human and Mouse Multidrug Resistance Protein 4 (MRP4) Orthologs for Arsenic Metabolites
Authors: Whitlock Brayden D. , Swanlund Diane P. , Le X. Chris , Schuetz John D. , Cole Susan P. C. , Leslie Elaine M. 
Published: November 2023

An estimated 160 million people world‐wide are exposed to levels of arsenic in their drinking water higher than the limit of 10 ppb (10 μg/L) set by the World Health Organization. Arsenic is a Group 1 (proven) human carcinogen which causes skin, lung, and bladder tumors, and is associated with numerous other adverse health effects, including neuropathy and cardiovascular dysfunctions. Multidrug resistance proteins (MRPs/ABCCs) mediate the cellular efflux of a chemically diverse array of endogenous and xenobiotic metabolites. Substrates include glutathione (GSH) conjugates of inorganic and methylated arsenic species and dimethylarsinic acid (DMAV). Unlike in humans, arsenic is a poor carcinogen in rodents, requiring at least 100‐fold higher doses for tumour induction. Differences in toxicokinetics between humans and other species are also well documented. We have shown that human MRP4 (hMRP4) reduces the cytotoxicity and cellular accumulation of inorganic and methylated arsenic species, and hMRP4‐enriched membrane vesicle transport studies identified the GSH conjugate of the highly toxic methylated arsenic metabolite monomethylarsonous acid (MMAIII), MMA(GS)2, and the major human urinary metabolite DMAV as the transported forms. The objective of the current study was to determine if mouse Mrp4 (mMrp4) conferred resistance to and/or transported the same arsenic species as hMRP4. HEK293 clonal cell lines stably expressing mMrp4 were established and used in cytotoxicity assays. Our results showed that mMrp4 did not confer resistance to any of the arsenic species tested [arsenite (AsIII), arsenate (AsV), MMAIII, monomethylarsonic acid (MMAV), dimethylarsinous acid (DMAIII) or DMAV]. Studies with mMrp4‐enriched membrane vesicles showed that unlike hMRP4, mMrp4 did not transport MMA(GS)2 or DMAV. Thus, under the conditions tested, arsenicals are not substrates for mMrp4. These results suggest that hMRP4/mMrp4 could contribute to differences in arsenic toxicokinetics between humans and mice. Furthermore, Mrp4/Abcc4(−/−) mice are unlikely to be a relevant model for understanding the in vivo contribution of hMRP4 to arsenic detoxification and elimination.Support or Funding InformationCanadian Institutes of Health Research, Alberta Innovates Health Solutions, Faculty of Medicine and Dentistry, University of Alberta
Diethyldithiophosphate (DEDTP) Role in Proportion of Macrophages and Natural Killer Cells in Lymph Vessel and 4T1 Murine Breast Tumor Model
Authors: BUELVAS DUNIA MARGARITA MEDINA , Muñiz Elizabeth Estrada , Loyo Libia Vega 
Published: November 2023

The diethyldithiophosphate (DEDTP) is a metabolite produced by the degradation of organophosphorus pesticides (POFs), and a dialkylphosphate (DAP) which is also chemically synthesized with a widespread commercial use and so the world population is chronically exposed to this compound. It is suggested that organophosphorus compounds produce deleterious effects on the immune system of the exposed population, since it has been demonstrated that DAPs affect peripheral blood lymphocytes activation and proliferation of T lymphocytes, furthermore, in vivo assays using C57 mice, the exposure to DEDTP enlarges skin tumors of BMK (Baby Mouse Kidney) cells, these results suggest that DEDTP exposure in vivo may modify the immune response and enhance the growth of tumor cells. Therefore, our objective is to determine the effect of DEDTP in the proportion of macrophages and natural killer cells (NKs) in lymph nodes and 4T1 breast cancer tumors in mice. To this end, we used Balb/c female mice of 6–8 weeks old, exposed for 8 days to 0.01 g/kg i.p. of DEDTP and inoculated with 1×105 4T1 cells. Tumors and lymph nodes were excised and measured. The percentage of total (F4/80+) and M2 (CD206+) macrophages and natural killer (NK) cells (CD355+) were assessed by flow cytometry. Exposure to DEDTP for 8 days increased the proportion of M2 macrophages. After 15 days of exposure to DEDTP, NK cells proportion was reduced but increased after 20 days. In mice challenged with 4T1 cells during DEDTP exposure, the number of total macrophages increased while the proportion of CD355+ cells in the lymph node proximal to the tumor decreased after 5 days; 10 days after, the number of M2 macrophages increased. Finally, after 20 days the number of M2 macrophages and NK cells in the proximal lymph node increased. In tumors, the number of infiltrating NK cells and total macrophages were reduced, followed by a reduction of M2 macrophages. Subsequently, after 15 days we observed an increase of infiltrating NK cells and M2 macrophages; after 20 days of tumor, M2 macrophages were increased. None of these changes had a significant impact on the establishment and development of tumors. With these results we conclude that exposure to DEDTP, in the absence of a particular antigen challenge, exerts immunomodulatory effects on individuals, apparently by inducing alternative activation of macrophages, however, this effect had no significant effect in the growth of 4T1 tumor cells. This could probably be because the more aggressive behavior of 4T1 tumor cells than BMK cells (previously tested on C57 mice) and the pronounced Th2 immune response profile of Balb/c mice (contrary to C57 animals). These results suggest that the effect of DEDTP on the immune system is not decisive against an aggressive tumor challenge, but may have relevance against a bacterial or parasitic infectious challenge.Support or Funding InformationCONACyT grant 153468
Oxidative and Inflammatory Responses of Monocytes or Macrophages to Ambient Particulate Matter
Authors: Bliss Bishop Charles , Solaimani Parissa , Campbell Arezoo 
Published: November 2023

Oxidative stress and inflammation are two major responses to ambient particulate matter (PM) and are thought to be responsible for the majority of its adverse health impacts. The hypothesis of this study is that monocytes and macrophages will have different responses to PM and that the oxidative and inflammatory responses will be linked. A human monocytic cell line (THP‐1) was used as a model for monocytes. These cells were also differentiated to macrophages via exposure to phorbol ester (PMA) for 48 hours. These cells were then exposed to three different size fractions of particulate matter: PM10 (coarse), PM2.5 (fine), and PM0.1 (ultrafine) for 24 hours. Oxidative stress was measured using the DCFH‐DA assay. Reactive oxygen species (ROS) increased compared to the control, but the increase was not dependent on PM size. There was no significant difference in ROS levels in monocytes compared to macrophages. Inflammatory cytokines TNF‐α and IL‐1β were measured using ELISA. In both monocytes and macrophages, fine and coarse particles elicited a significant increase. No significance occurred after exposure to ultrafine particles and there was no difference in the response to monocytes compared to macrophages. This study suggests that monocytes and macrophages respond similarly to particulate matter. Also, PM‐induced oxidative and inflammatory responses may not be directly linked. The mechanism by which PM causes these observed responses in immune competent cells is the subject of future studies in the laboratory.Support or Funding InformationBritish Petroleum/ Southern California Air Quality Management District (BP/SCAQMD Award #11527) and Western University of Health Sciences
Inhibitory Effects of Green Coffee Bean Extract on Serotonin Uptake in the Rat Brain Cortex
Authors: Ohia Sunny Edet , Bush Leah , Robinson Jenaye , Njie‐Mbye Ya Fatou , Bagchi Debasis 
Published: November 2023

Caffeine/Coffee consumption has been reported to increase the thermic effects of food and fat oxidation (Acheson et al., Am. J. Clin. Nutr. 33: 989, 1980) and can cause weight loss in obese subjects (Westerterp‐Plantenga et al., Physiol. Behav. 89:85, 2006).PurposeSince serotonin (5‐hydroxytryptamine, 5‐HT) has been implicated in the regulation of feeding behavior, the aim of the present study was to determine the effect of extract of green coffee bean (GCBE) on the uptake of radiolabeled serotonin ([3H]‐5‐HT) in the rat brain cortex.MethodsIsolated rat brain cortex slices were incubated in oxygenated Krebs solution maintained at 37°C for 20 minutes and transferred to warm oxygenated buffer solutions containing [3H]‐5‐HT (80 nM) for different time intervals. In some experiments, tissues were exposed to GCBE (300 μM ‐ 3 mM) and the selective serotonin reuptake inhibitors (SSRI) fluoxetine (100 μM) plus clomipramine (10 μM).ResultsA time‐dependent uptake of [3H]‐5‐HT occurred in control brain cortex slices reaching a maximum at 90 minutes. GCBE (300 μM – 3 mM) and the well‐known serotonin reuptake inhibitors fluoxetine plus clomipramine inhibited the time‐dependent uptake of [3H]‐5‐HT. For example, at 30 minutes, GCBE (1 mM) caused a 23 ± 5.4 % (n = 6) decrease in [3H]‐5‐HT uptake, whereas fluoxetine plus clomipramine inhibited [3H]‐5‐HT uptake by 52 ± 3.1 % (n = 6). At 90 minutes, GCBE (1 mM) reduced uptake by 23 ± 4.16% (n = 6), while fluoxetine plus clomipramine inhibited [3H]‐5‐HT uptake by 25 ± 10.2 % (n = 6).ConclusionGCBE can inhibit [3H]‐5‐HT uptake and increase 5‐HT availability in rat isolated brain cortex slices in a manner similar to that of the well‐established SSRIs suggesting a potential role of this compound in regulation of feeding behavior.Support or Funding InformationCepham Inc. 144 12th Street Piscataway, NJ 08854