bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of a New Formulation of Micronized and Ultramicronized N‐palmitoylethanolamine Treatment in a Tibia Fracture Mouse Model of Complex Regional Pain Syndrome
Authors: Fusco Roberta , Gugliandolo Enrico , Campolo Michela , Evangelista Maurizio , Filippone Alessia , Di Paola Rosanna , Cuzzocrea Salvatore
Published: November 2023

Complex regional pain syndrome type 1 (CRPS‐I) is a disabling and frequently chronic condition. It involves the extremities and is a frequent consequence of distal tibia and radius fractures. The inflamed appearance of the affected CRPS‐I limb suggests that local production of inflammatory mediators may be implicated in the ensuing etiology. A rodent tibia fracture model, characterized by inflammation, chronic unilateral hindlimb warmth, edema, protein extravasation, allodynia and hyperalgesia resembles the clinical features of patients with acute CRPS‐I. N‐palmitoylethanolamine (PEA), a member of the family of naturally‐occurring N‐acylethanolamines, is well‐known for its ability to modulate inflammatory processes and regulate pain sensitivity. However, the large particle size and lipidic nature of PEA may limit its bioavailability and solubility when given orally. Micronized formulations are frequently used to enhance a drug's rate of dissolution and reduce its variability of absorption when orally administered. The aim of this study was to assess the effects of a formulation of micronized and ultramicronized PEA (PEA‐MPS), given orally in a mouse model of CRPS‐I. CD‐1 male mice were subjected to distal tibia fracture and divided into two groups: control and treated with PEA‐MPS (PEA micronized 300 mg/kg and ultramicronized 600 mg/kg). Sensibility to pain was monitored in all mice throughout the course of the experiment. Twenty‐eight days after tibia fracture induction animals were sacrificed and biochemical parameters evaluated. The PEA‐MPS‐treated group showed an improved healing process, fracture recovery and fibrosis score. PEA‐MPS administration decreased mast cell density, nerve growth factor (vehicle 7.666 ± 0.577 treatment 1.666 ± 0.577; p<0.05), matrix metalloproteinase 9 (vehicle 3,333 ± 0,577 treatment 7,111 ± 1,031; p<0.05) and cytokine expression. This treatment also reduced (poly‐ADP)ribose polymerase activation (vehicle7.952 ± 1,526 treatment 2.596 ± 0.624; p<0.05), peroxynitrite formation (vehicle 8.589 ± 0.785 treatment 1.982 ± 0.523; p<0.05) and apoptosis. Our results suggest that PEA‐MPS may be a new therapeutic strategy in the treatment of CRPS‐I.
PMN mediated endothelial cell TRPM2 activation regulates neutrophil trafficking and lung vascular permeability
Authors: Mittal Manish , Nepal Saroj , Tiruppathi Chinnaswamy , Malik Asrar B
Published: November 2023

TRPM2 (Transient receptor potential melastatin‐2) is a cation channel that mediates Ca2+ entry in response to intracellular ADP ribose (ADPR) generated during oxidative stress. We tested the hypothesis that Reactive Oxygen Species (ROS) released by polymorphonuclear leukocytes (PMNs) during sepsis activate endothelial TRPM2 and induce trans‐endothelial PMN migration and cause lung vascular injury. To investigate the role of endothelial TRPM2 in PMN transmigration, we generated tamoxifen inducible endothelial cell‐restricted TRPM2 knockout (Trpm2ΔEC−/−) mice. Lung vascular permeability and PMN transmigration following LPS challenge (10 mg/Kg) did not increase in these mice. Human lung microvascular endothelial cells (HLMVECs) treated with TRPM2‐siRNA or poly ADP ribose polymerase 1 (PARP1)‐siRNA showed significantly reduced Ca2+ entry in response to activated PMNs (PMN‐to‐EC ratio of 5:1). PMNs isolated from gp91phox−/− mice or WT mice treated with DPI (10 μM) significantly reduced Ca2+ entry in ECs as compared to WT PMNs. ECs treated with PARP1‐siRNA also showed significantly reduced ADPR generation in response to H2O2 stimulation. Overexpression of the ADPR insensitive TRPM2 mutant channel (C1008A) in HLMVECs significantly suppressed the H2O2 induced Ca2+entry. In vivo forced expression of the TRPM2 mutant channel or silencing of PARP‐1 protected against LPS‐induced increase in lung vascular permeability and PMN transmigration in mice. We conclude that endothelial Ca2+ entry following activation of TRPM2 in PMNs disrupts endothelial barrier, thereby promotes lung vascular injury.

Support or Funding Information

RO1 HL 077806, RO1 HL 122157
Effects of a Co‐Micronized Composite Containing Palmitoylethanolamide and Polydatin in an Experimental Model of Benign Prostatic Hyperplasia
Authors: Cordaro Marika , Impellizzeri Daniela , Siracusa Rosalba , Gugliandolo Enrico , Fusco Roberta , Esposito Emanuela , Di Paola Rosanna , Cuzzocrea Salvatore
Published: November 2023

Palmitoylethanolamide (PEA), a fatty acid amide‐signalling molecule has well‐known anti‐inflammatory and neuroprotective effects. Nevertheless, PEA does not possess the ability to prevent free radical formation. Polydatin (PLD), a biological precursor of resveratrol, has antioxidant activity. A combination of PEA and PLD could, conceivably, have beneficial effects on oxidative stress produced by inflammatory processes. In the present study we investigated the effects of a co‐micronized composite containing PEA and PLD (m(PEA/PLD)) in a model of testosterone‐induced benign hyperplasia (BPH).

BPH was provoked in rats by daily administration of testosterone propionate (3 mg/kg) for 14 days. This protocol led to alterations in prostate morphology and increased levels of prostaglandin E2 and dihydrotestosterone as well as of 5α‐reductase 1 and 5α‐reductase 2 expression. Moreover, testosterone induced marked inflammation in terms of an increase in nuclear translocation of nuclear factor‐κB p65 and consequently in IκB‐α degradation as well as disregulation of inducible nitric oxide synthase, cyclooxygenase‐2 expression and manganese superoxide dismutase expression and in the apoptosis pathway. Our results show, for the first time, that m(PEA/PLD) is capable of decreasing prostate weight and dihydrotestosterone production in BPH‐induced rats. These effects were most likely correlated to the anti‐inflammatory and apoptotic effects of m(PEA/PLD).

Accordingly, these results support the view that m(PEA/PLD) should be further studied as a potent candidate for the management of BPH.
Changes in DNA Damage Response Proteins During Tamoxifen Treatment and Resistance
Authors: Vergne Ralphdy , Ortiz‐Sanchez Carmen , Godoy‐Munoz Lenin Jose , Encarnación Jarline , Matta Jaime
Published: November 2023

Tamoxifen (TMX) is currently used for the treatment of both early and advanced estrogen receptor positive (ER+) breast cancer in pre‐ and post‐menopausal women. TMX works by blocking the actions of estrogen and preventing proliferation and furthermore, cell death. DNA damage is an alteration of chemical structure of the DNA that activates a network of proteins to start several cellular processes that sense, detect signal and repair DNA lesions. The mechanism of action of several chemotherapeutic drugs is based on damaging the DNA of the cancerous cell. Although TMX's mechanism of action is not through DNA damage, TMX treatment can lead to the production of oxidative stress that in consequence can affect and damage DNA. Our aim is to study the DNA damage induced by TMX resistance in ER+ breast cancer cell line (MCF‐7) cells and assess the expression changes of key proteins involved in this response. Our hypothesis is that there will be more DNA damage in cells treated with TMX in comparison to untreated cells, and the DNA damage contributes to resistance against TMX. To achieve this, we will assess ER expression as an indicator of resistance to TMX, analyze DNA damage during TMX treatment, and study protein expression related to DNA damage using MCF‐7 under TMX. To achieve this, we cultured MCF‐7 cells using complete media without phenol red and 0.5 μM of TMX with 10 μL of insulin. After 1 week, cells were lysed for protein and RNA extraction. Our results shows that ERα expression was similar to untreated cells; therefore, no statistically differences p>0.05 were found. To assess protein expression changes in proteins involved in DNA damage, we assessed PTEN expression by Western blot. Although, no statistically significant differences were found when comparing TMX treated cells with untreated cells (p>0.05) we observed a slight expression change. This preliminary data indicates that PTEN expression might be affected in ER+ breast cancer cells when TMX resistance is developed. Currently, we are developing an MCF‐7 cell line resistant to TMX for additional studies and assessing protein expression changes of other candidates involved in DNA damage response (such as BRCA‐2 and XPF). These results will shed light on the effect of TMX resistance in the DNA damage response of ER+ breast cancer cells. Also, they will allow us to get an inside view and to track the changes in MCF‐7 protein expression during the TMX resistance timeframe.

Support or Funding Information

Funded by NIH‐NIGMS #2R25GM096955 (UPRP‐Rise program)

Supported by grants from the NCI Center to Reduce Health Disparities and NIH‐NIGMS MBRS Program grant #9SC1CA182846‐04 (J. Matta Laboratory)

We also want to acknowledge the support of the Molecular and Genomics Core Laboratory at Ponce Health Sciences University‐Ponce Research Institute (Grant MD00757).
The choline acetyltransferase inhibitor BW813U suppresses the growth of lung adenocarcinoma from smokers
Authors: Akers Austin Tyler , Colclough Kate W , Friedman Jamie R , Bow Eric W , Rimoldi John M , Cutler Stephen J. , Hardmin W. Elaine
Published: November 2023

The clinicopathological profile of lung adenocarcinoma (LAC) in smokers is completely different from those in non‐smokers. One of the unique features of LAC in smokers is that it is relatively resistant to therapies. For example, targeted therapeutic agents like EGFR‐inhibitors (erlotinib and gefitinib) are highly effective in LACs which develop in non‐smokers. However, these EGFR‐inhibitors display much lower anti‐tumor activity in LACs in patients who are active smokers. Similarly, lung cancer patients (who are active smokers) show lower response to chemotherapy than those who are non‐smokers. However, majority of LAC patients are smokers. This underlines the need to identify novel molecular targets relevant for LAC therapy in patients who are exposed to cigarette smoke via active smoking or exposure to secondhand smoke. Several convergent studies show that nicotine (the addictive component of cigarette smoke) accelerates the growth of lung cancers, as well as confers resistance to chemotherapy. One of the mechanisms underlying these biological activity of nicotine is that it promotes the secretion of the neurotransmitter acetylcholine (ACh) from LAC cells. ACh is known to be an autocrine growth factor for LAC cells and is synthesized by the enzyme choline acetyltransferase (ChAT). The present study investigates the feasibility of ChAT as a molecular target for LAC in smokers. We find that ChAT levels are upregulated in human LAC cell lines and tissues in a manner dependent on the smoking history. Finally, the ChAT inhibitor BW813U causes robust apoptosis in human LAC cells. The magnitude of BW813U‐induced apoptosis is similar across LAC cell lines (irrespective of smoking history); however, the concentration of BW813U which causes apoptosis is lower in LAC cell lines belonging to heavy smokers. Our studies validate choline acetyltransferase (CHAT) as a viable drug target for the majority of population of lung cancer patients who are smokers.

Support or Funding Information

Funding for our study was supported by the an NIH R15‐AREA Grant (2R15CA161491‐02), WVU‐MU Health Partnership Grant and by ACTSI Grant from Marshall University. Furthermore, this study was supported in part by an Institutional Development Award (IDeA) Grant number P20GM104932 from the National Institute of General Medical Sciences (NIGMS) and the Research Core B of COBRE, a component of the National Institutes of Health (NIH).
Antiproliferative activity of endocannabinoids in U87‐MG glioblastoma cells
Authors: Gugnani Kuljeet , Rondon‐Ortiz Alejandro N. , Juarez‐Valdez Alejandra , Pardes‐Quiroz Jose M , Pino‐Figueroa Alejandro
Published: November 2023

Anandamide (AEA) and 2‐arachidonoylglycerol (2‐AG) are compounds naturally produced in humans and animals that interact with the cannabinoid system. These compounds activate specific cannabinoid receptors on cells to alter the neurotransmitter release from presynaptic neuron. There are studies which claim that endocannabinoids are selective towards tumor cells without affecting the functioning of normal cells, mainly by regulating different cell‐death pathways and downstream signaling molecules. The purpose of this study is to evaluate the antiproliferative activity of AEA and 2‐AG in U87‐MG glioblastoma cells. A cell viability assay utilizing 3‐(4,5‐dimethylthiazol‐2‐yl)‐5‐(3‐carboxymethoxyphenyl)‐2‐(4‐sulfophenyl)‐2H‐tetrazolium (MTS) was performed to evaluate the antiproliferative effects. The results demonstrated that AEA at concentrations 10 and 30 μM decreased cell viability as compared to control (P≤ 0.001). 2‐arachidonoylglycerol at concentrations 10, 30 and 100 μM decreased cell viability as compared to control (P≤ 0.001). Interestingly, a combination of AEA and 2‐AG at concentrations 10 and 30 μM, respectively reduced cell viability as compared to their individual effects suggesting a synergistic approach. Moreover, a significant reduction in cell viability was observed with a combination of OL‐135, a reversible inhibitor of fatty acid amide hydrolase (FAAH) and AEA as compared to AEA alone. Similarly, a significant reduction in cell viability was observed with a combination of JZL‐184, an irreversible inhibitor for monoacylglycerol lipase (MAGL) and 2‐AG as compared to 2‐AG alone. These results suggest that by inhibiting the enzymes FAAH and MAGL which are involved in the degradation of AEA and 2‐AG, respectively a further reduction in cell viability could be observed, suggesting a strategy to inhibit endocannabinoid degrading enzymes and use of endocannabinoids as potential anti‐cancer agents. Further assays will be performed to determine the mechanism of anti‐cancer activity demonstrated by these compounds.
Anti‐Proliferative Activity of a Highly Potent Fatty Acid Amide Hydrolase Inhibitor: Effects on Non‐Small Cell Lung Cancer
Authors: Alafnan Ahmed , Pino‐Figueroa Alejandro
Published: November 2023

Lung cancer has become the leading cause of cancer deaths in the US and worldwide. The endocannabinoid system has ubiquitous regulatory actions in the body reflecting the importance of a system related with many physio‐pathological processes. It is a promising target for the treatment of several different diseases, one of which might be cancer. Thus, targeting this system is becoming a novel strategy in the search for cancer treatments. It has been demonstrated that fatty acid amide hydrolase (FAAH), which is one of the enzymes responsible for terminating endocannabinoid signaling, is up‐regulated in aggressive human tumors and cancer cells. Since anandamide (AEA) and 2‐arachidonly glycerol (2‐AG) have anti‐proliferative activity, inhibition of FAAH could sustain AEA anti‐proliferative and anti‐invasive properties. Therefore, the aim of this study was to investigate the anti‐proliferative activity of N‐(8Z‐heptadecen‐1‐yl)‐O‐(3‐pyridylmethyl)carbamate (ZYPC), which has shown to possess a potent and irreversible FAAH inhibitory activity. The results revealed that ZYPC inhibits FAAH with an IC50 of 182±18 nM, which is decreased to 42±3 nM following 60 min pre‐incubation. Furthermore, dialysis assay suggested that ZYPC could inhibit FAAH irreversibly. Moreover, ZYPC has significantly decreased cell viability of non‐small lung cancer cells A549 and H460 with IC50 of 4.9 and 0.9 μM, respectively. ZYPC inhibits migration and invasion of A549 cells by 75 and 46%, respectively, and this effects are accompanied with 1.9 fold up‐regulation of tissue inhibitor matrix metalloproteinases‐1 (TIMP‐1) and 0.4 fold down‐regulation of matrix metalloproteinase‐2 (MMP‐2). Further studies will be performed in order to understand the potential mechanism(s) governing ZYPC anti‐tumorigenic effects.
Angiotensin II/NaCl Increases FosB Expression in the Paraventricular and Supraoptic Nucleus
Authors: Franklin Cynthia , Conner Caroline , Biggs Lauren , Tischler Lauren , Rangel Yollanda , Gottlieb Helmut B
Published: November 2023

The purpose of the present study is to examine changes in FosB immunohistochemistry evoked by an in vivo hypertension model, angiotensin II/high salt diet (ANGII), within the supraoptic nucleus (SON) and paraventricular nucleus of the hypothalamus (PVN). Immuno‐detection of Fos, a member of the AP‐1 family of transcription regulatory proteins, has been widely used as an indicator of neural activation. FosB is an inducible member of the Fos family that has been proposed to be a marker of chronic activation in the central nervous system, and increases in Fos expression have been shown to occur under several pathophysiological conditions. Angiotensin II and a high salt diet have been proposed to activate several brain regions involved in autonomic function. As such, we used FosB to determine the changes in neuronal activity in the forebrain of ANGII treated rats as compared to rats treated with high salt diet only, and rats on a normal diet (control group). By examining the changes in FosB, we can determine which hypothalamic brain regions are being chronically modulated by the ANGII model. For further comparison, we also measured changes in FosB in ANGII hypertensive rats treated with a kappa opioid receptor (KOR) agonist, which has been shown to produce effects on cardiovascular and renal function. Male Sprague‐Dawley rats were instrumented with osmotic mini‐pumps filled with angiotensin II for subcutaneous (s.c.) infusion. In addition, rats were implanted with an ICV cannula for delivery of drugs into the lateral cerebral ventricle. After recovering, rats were fed 2% high salt diet for 14 days and at day 15, injected with either U50‐488H, kappa opioid receptor agonist, or saline. Ninety minutes post injection, each rat was anesthetized with Inactin (100 mg/kg i.p.) and perfused with PBS and fixative [4% paraformaldehyde in phosphate buffered saline, PBS, pH =7.4]. The brains were removed and processed for FosB immunocytochemistry using a commercially available antibody (1:1000 rabbit anti‐FosB). Brain sections were examined using light microscope to identify FosB positive cells. ANGII hypertensive model increased FosB expression in the PVN when compared to high salt diet. ANGII also increase FosB expression in the PVN and SON when compared to naïve‐control rats. Although FosB expression increased in the PVN, there were no statistically significant changes in FosB expression between magnocellular and parvocellular region. U50‐488H administration decreased FosB expression in the SON and PVN. The PVN and SON have been shown to be involved in the control of blood pressure, fluid and electrolyte balance. Central KOR receptor inhibition of these forebrain regions may form part of the neuropathways involved in the decreases in water and sodium excretion evoked by KOR activation.
Teaching Radiology/Anatomy to Medical Students: Using Multiple Approaches to Understand 2D/3D Structure of the Heart
Authors: Zill Sasha
Published: November 2023

Training in medical school now requires students to understand and interpret digital images of body structure. Many of these images (MRI, CT) are serial planar sections of body regions and internal organs. Some organs, such as the heart, have both complex internal organization and non‐orthogonal orientation within the body. As a consequence, interpretation of two dimensional sections of the heart is particularly challenging and difficult to relate to its three dimensional structure. As part of training in Gross Anatomy at the Joan C. Edwards School of Medicine, we have developed new multifaceted approaches to teaching the structure of the heart in both two and three dimensions. The format combines independent learning using downloaded videos with cadaver dissection and study of specimens in the Gross Anatomy laboratory. The material for independent learning is based upon study of a single template, a T1 weighted MRI series of the the thorax taken without contrast medium. The image series, chosen for its high definition of vessels and chambers, was imported to PowerPoint and labeled by a medical student under supervision of our department of Radiology. The independent learning exercise first required viewing a short (20 minute) video that incorporated those images. Each image in the series was correlated with other diagrams and photographs that provided cues for orientation to the structure of the heart in three dimensions. Student feedback to date has indicated that the video provided a valuable tool in understanding of the heart in both two and three dimensions, although repeated viewing of images was often necessary. In addition, students were provided prosected specimens of the heart (in the Gross Anatomy laboratory) that were sectioned in the plain of axial MRI or CT. These specimens were prepared by manually cutting hearts perpendicular to an axis determined by the orientations of the Superior and Inferior Venae Cavae. We also plan to develop methods for students to section the hearts they have removed in the course of their own cadaver dissection. In sum, we present methods for teaching students the structure of the heart that provides understanding of the relationship of its three dimensional structure and its appearance in two dimensional sections. We suggest that the combination of approaches (independent learning, guided laboratory instruction) may be beneficial in the future, particularly as schools undergo curricular revisions/compressions that can lead to reduction in the time available for study of Anatomy.
A Multifunctional Platform for Rapid Detection and Post‐Detection Analysis of Circulating Tumor Cells
Authors: Yang Yu‐Ping , Datar Ram , Cote Richard , Daunert Sylvia
Published: November 2023

Circulating tumor cells (CTCs) has been shown a significant prognostic value as a surrogate marker in various cancers. To employ CTC assay into clinical practice, a method requires rapid CTC detection in blood with high sensitivity and specificity. Here, we report a novel bioluminescence‐based assay, due to high sensitivity and extremely low background signal of bioluminescence, by genetically fusing antibody mimetics with bioluminescence proteins for CTC detection. Moreover, multi‐marker combination strategy is employed to increase detection sensitivity due to CTC heterogeneity. In the model of spiking experiments, our approach shows high specificity and sensitivity in detection of viable tumor cells with no cell toxicity observed which subsequently allows expansion of those cells in culture for CTC biobanks. Given the heterogeneous nature of CTCs, molecular characterization of CTCs at single‐cell level offers a precise investigation of patient‐specific cancer‐omics profiles for improved diagnosis and personalized treatment. To further performing post‐detection analysis of CTCs at the single‐cell level, the combination with fluorescence modality enables accurate identification of the individual spiked cells for downstream single‐cell analysis after bioluminescence detection. In conclusion, our technique represents a promising CTC detection approach for cancer management.

Support or Funding Information

National Institutes of Health (NIH)
Linking Nutrition and Molecular Biology Using Data Mining and Graph Theory
Authors: Linchangco Richard , Jay Jeremy J , Brouwer Cory
Published: November 2023

Cardiovascular disease, cancer, and diabetes are among the leading causes of death in the United States. Factors of chronic disease include physical activity, genetics, and diet, but only physical activity and diet can be controlled. Traditionally, diet and human health have been studied through epidemiological studies at the phenotypic and observational level. The molecular mechanisms underlying the health benefits of certain foods remain largely unknown, which creates a disconnect between plant science, nutrition, human health, and molecular biology.

The need to link the disparate domains of nutrition and molecular biology presents a difficult problem. Existing curated resources provide a strong baseline within the distinct domains, but data connecting them does not yet exist in machine‐readable form. This work applies data mining techniques to large collections of both medical and agricultural publication abstracts to generate links between nutrition and biology. This network of integrated data is placed into a graph database, which is the first application of Neo4j to a knowledge‐oriented nutrigenomics graph. This work enables novel queries to discover new evidence in the support of open nutrigenomics questions.

Support or Funding Information

Funding from UNC General Administration and the Plant Pathways Elucidation Project
The Pseudophosphatase MK‐STYX's Effects on Dynein
Authors: Whitaker Adom , Hall‐Mozingo Lauren , Hinton Shanta
Published: November 2023

MK‐STYX [MAPK (mitogen‐activated protein kinase) phosphoserine/threonine/tyrosine‐binding protein] is a pseudophosphatase member of the dual‐specificity family subfamily of protein tyrosine phosphatases. It lacks the essential cysteine in its signature motif required for catalytic activity. We reported that MK‐STYX interacts with G3BP‐1 [Ras‐GAP (GTPAse‐activating protein) SH3 (Src homology 3) domain binding protein‐1], and inhibits stress granule (SG) formation implicating it in the stress response pathway. SGs are large structures in which untranslated mRNAs accumulate and may serve as sites of mRNA sorting when cells are under stress. SGs depend on motor proteins to shuttle silenced messenger ribonucleotides and binding partners. In particular, motor proteins such as dynein and kinesin have a role in the SG pathway. Dynein assists in SG assembly, whereas kinesin assists with disassembly. Recent research identified dynein's heavy chain subunit (DHC) and BicD1 (a dynein adaptor) as key elements in SG assembly. Inhibition of dynein impairs stress granule formation. Perhaps, MK‐STYX inhibits SGs by affecting the dynamics of dynein. Fluorescence microscopy of HEK/293 cells cotransefected with mCherry‐MK‐STYX and GFP‐DCI2 (intermediate chain 2 of dynein) demonstrates that MK‐STYX co‐localizes with DIC2. Furthermore, GFP‐DIC2 aggregates form in cells co‐transfected with mCherry‐MK‐STYX. MK‐STYX causes GFP‐DIC2 aggregates, suggesting that MK‐STYX effects dynein subcellular localization. This work characterizes the relationship between MKSTYX and the motor protein dynein. Our future studies will characterize MK‐STYX's relationship with kinesin; our pilot mass spectrometry data suggests that MK‐STYX interacts with a kinesin motor protein. Therefore, understanding the dynamics of MK‐STYX and motor proteins may help elucidate the mode of action that MK‐STYX uses to prevent stress granule assembly.
Downregulation of X‐linked inhibitor of apoptosis protein by ‘7‐Benzylidenenaltrexone maleate’ sensitizes pancreatic cancer cells to TRAIL‐induced apoptosis
Authors: Kim So‐Young , Park Sojung , Yoo Seona , Rho Jin Kyung , Jun Eun Sung , Chang Suhwan , Kim Kyung Kon , Kim Song Cheol , Kim Inki
Published: November 2023

Tumor necrosis factor‐related apoptosis‐inducing ligand (TRAIL) is a potential biological anticancer agent. However, a wide range of human primary cancers, including pancreatic cancer, display resistance to apoptosis induction by TRAIL. Therefore, this resistance needs to be overcome to allow TRAIL to be successfully used in cancer therapy. In this study, we performed a compound screen to isolate TRAIL sensitizers and found that one of the identified compounds, 7‐benzylidenenaltrexone maleate (BNTX), sensitized pancreatic cancer cells to TRAIL‐induced apoptotic cell death. The combination of BNTX with TRAIL promoted the release of cytochrome c from mitochondria into cytosol with caspase activation and a resulting increase in annexin‐V‐stained cells. From a mechanistic perspective, we found that BNTX downregulated X‐linked inhibitor of apoptosis protein (XIAP) expression when used in combination with TRAIL and BNTX and that TRAIL‐induced apoptosis was augmented by the siRNA‐mediated knockdown of XIAP with caspase activation. We further demonstrated that BNTX promoted the ubiquitin/proteasome‐dependent degradation of XIAP protein via protein kinase C (PKC) alpha/AKT pathway inhibition. Moreover, combined treatment by BNTX with TRAIL suppressed tumor growth in vivo. Therefore, we suggest that inhibitor of apoptosis protein‐mediated resistance of pancreatic cancer cells to anticancer therapeutics can be overcome by inhibiting the PKCα/AKT pathway.

Support or Funding Information

This work was supported (40%) by (1) the Basic Science Research Program (NRF‐2016R1D1A1B03932365) funded by the Ministry of Science, Information & Communication Technology (ICT) and Future Planning, and (2) supported (30%) by a grant of the Korean Health Technology R&D Project, Ministry of Health & Welfare, Republic of Korea (Grant Number: HI14C2640), and (3) supported by a Center for Women in Science, Engineering and Technology (WISET) grant (20%) funded by the Ministry of Science, ICT & Future Planning of Korea (MSIP) under the Program for Returners into R&D (KW‐2015‐PPD‐0134), and (4) supported by internal research program of ASAN Institute for Life Sciences (10%) (16‐512 & 16‐586).
Signaling Pathways Implicated in Butyrate‐Arrested Vascular Smooth Muscle Cell Proliferation
Authors: Mathew Omana P , Ranganna Kasturi , Selvam Chelliah , Yousefipour Zivar
Published: November 2023

Prevalence and relevance of epigenetic components in cardiovascular disease pathways including those associated with atherosclerosis are now being explored to understand their impact on the pathogenesis. Butyrate, an epigenetic histone modifier with histone deacetylase (HDAC) inhibitory activity, inhibits vascular smooth muscle cells (VSMC) proliferation, an important component in the pathogenesis of atherosclerosis and alters cellular size and morphology. Since mTOR signaling pathways are crucial to cell proliferation, cell growth and survival by their multiple downstream pathways, here we investigate some of these pathways to understand the underlying mechanisms associated with butyrate effects on VSMC. Proliferating VSMC were exposed to 5 mM butyrate for different lengths of time and processed for assessing butyrate effects on the status of different signaling proteins by western analysis. Results indicate that butyrate treatment causes inhibition of mTOR expression and accordingly activation of mTOR by phosphorylation of Ser2448 is also inhibited. This inactivation of mTOR is further reflected in the inhibition of activation of its downstream effector p70S6 kinase, and in turn, inhibition of phosphorylation of p70S6 kinase substrate, S6 ribosomal protein, which are involved in the protein synthesis. Furthermore, butyrate treatment causes time‐dependent inhibition of PI3K, Akt/PKB and PDK1 activation by phosphorylation. The results indicate that inhibition of activation of both PI3K/Akt and mTOR pathways appear to be involved in butyrate arrested VSMC proliferation, which suggest that both pathways are interconnected and function as a single pathway in butyrate arrested VSMC proliferation because both pathways are crucial for the cell proliferation, cell growth and survival through the mediation of their downstream effectors.

Support or Funding Information

This study was made possible, in part, by Molecular Biology research infrastructure support from grant number 2G12MD007605 from the NIMHD/NIH”
Comparison of Violacein, Mangosteen and Sidr Extracts as Agents for Inhibiting Breast and Lung Cancer Cell Growth
Authors: Bahannan Alaa Helme , Johnson Terrance , Myles Elbert
Published: November 2023

According to the American Cancer Society, Cancer is the second leading cause of death in the United States. Currently there are a number of effective microbiological and natural treatments of breast and lung cancer that can prevent surgery and removal of damage tissues yet lead to repair of the damaged tissue by the body itself. The purpose of this study was to expose four cell lines, Breast cancer (BT549, BT20, and MCF7) and Lung cancer (A549) to three natural extracts, Violacein, Sidr and Mangosteen. Violacein is a natural purple pigment that has potential medical applications such as antimicrobial, antiviral, and anticancer drugs that is produced by Chromobacterium Violaceum, and it was added to cancer cell cultures at various concentrations. In addition, it has been reported that Mangosteen and Sidr have anti‐tumor, anti‐leukemia, antifungal, antibacterial and antioxidants properties. Xanthones were extracted from the pericarp, whole fruit, heartwood, and leaf of the Mangosteen, and it was added also to the cancer cell cultures at different concentrations. On other hand, the Sidr extract was prepared as a crude methanol extract taken from the leaves of the plant Ziziphus spina‐christi. It was also exposed to cancer cell cultures at a variety of concentrations. The Alamar blue assay was used to view cell death of the breast and lung cancer cell lines. It was found that the Violacein from C. violaceum strain 14N23 inhibited the growth of MCF7. It was demonstrated also that the Mangosteen and Sidr extracts had an effect on the cancer cells A549, MCF7, BT 20, and BT549. The Mangosteen and Sidr extracts were found to be effective in inhibiting the growth of all the cancer cell lines. It was concluded that these extracts have potential for controlling cancer cell growth. Further studies will involve determining the pathways impacted by these extracts and determining the active components of these extracts.
Haspin is a survival factor for H3T3 phosphorylation and survivin recruitment in malignancy of colorectal cancer
Authors: Lee Tsai‐Chia , Chang Yun‐Hsuan , Yu Tzu‐Wei , Shen Tzu‐Keng , Tsai Ming‐Chang , Huang Chi‐Chou , Yang Tzu‐Wei , Lin Chun‐Che , Chiou Guang‐Yuh , Jong Yuh‐Jyh , Chao Jui‐I
Published: November 2023

Colorectal cancer (CRC) is one of the leading causes of cancer‐related morbidity and mortality in the world. Haspin is a histone H3 kinase that mediates the phosphorylation of H3 at threonine 3 (p‐H3T3), which recruits chromosomal passenger complex (CPC) and promotes cell division. However, the precise role of Haspin in CRC is still unclear. Here we show that overexpression of Haspin increased the protein levels of H3T3 and Survivin in CRC cells. Moreover, the Haspin and p‐H3T3 protein levels were highly expressed in CRC tumor tissues but not in normal tissues from clinical CRC patients. Interestingly, we found that Haspin proteins were inhibited but conversely elicited the stemness protein expression of CD133 in CRC tissues using 3‐D tissue culture condition. The similar results were verified by immunofluorescence staining and confocal microscopy. We suggest that Haspin exhibits a crucial role in the regulation of malignant and cancer stemness of CRC; however, the precise mechanism should be further investigated.
A novel synthetic compound overcomes drug resistance on EGFR (T790M) and cancer stemness in human non‐small cell lung cancer
Authors: Pan Pei Ying , Chang Chien‐Jen , Liu Kuang‐Kai , Chan Yu‐Chi , Chen Yan‐Ting , Chao Jui‐I
Published: November 2023

Drug resistance has become bottleneck in cancer therapy of non‐small cell lung carcinoma (NSCLC) patients. The mutations of epidermal growth factor receptor (EGFR) on exon 19 deletion and exon 21 point mutation are respond well to the first and second generation EGFR tyrosine kinase inhibitors (EGFR‐TKIs) such as Gefitinib and Erlotinib for NSCLC therapy; however, patients acquired the T790M mutation of EGFR will induce drug resistance. In addition, cancer stemness properties are also a pivotal factor of drug resistance in NSCLC. Accordingly, development of novel compounds for overcoming drug resistance of NSCLC is still desired. In this study, we show a novel synthetic compound derived from 5,8‐quinolinedione that has potential in overcoming drug resistance of EGFR (T790M) and cancerous stemness in NSCLC. This new compound can induce cell death and apoptosis in drug‐resistant H1975 cells, which contained EGFR (T790M). This compound inhibited Survivin protein expression and conversely increased the active caspase 3 and cleaved PARP for apoptosis induction. In addition, this compound also decreased the cell viability in drug resistance NSCLC cells, which were separated from pleural effusion of NSCLC. Moreover, it can inhibit the stemness protein expression of Oct4 in lung cancer cells. Taken together, we develop a novel compound, which further investigate for overcoming drug resistance and cancer therapy in NSCLC.
EIF5A1 Isoform A Can Modulate the Metabolism in HeLa Cells
Authors: Pereira Karina Danielle , Tamborlin Leticia , Meneguello Leticia , Bomfim Lucas Henrique Montes , de Proença Andre Ricardo Gomes , Melo Carlos Fernando Odir Rodrigues , Catharino Rodrigo Ramos , dos Reis Silveira Leonardo , Luchessi Augusto Ducati
Published: November 2023

The eukaryotic translation initiation factor 5A (eIF5A) is widespread in Archaea and Eukarya and contains the unusual amino acid residue hypusine. Recent studies have shown its involvement in the elongation step of translation. More specifically, acting on the synthesis of proteins containing polyproline stretches. The eIF5A has been involved in different physiological and pathological processes, such as inflammation, cancer, diabetes and virus infection. In humans, different transcript variants related to eIF5A1 are currently registered in databases. Recently, we observed that the transcript variant A is characterized by an additional upstream start codon in‐frame with the canonical initiation codon. Therefore, the product of this transcript, eIF5A1 isoform A, has a N‐terminal extension of 30 amino acid residues, when compared to the canonical isoform B. In addition it was evidenced that the isoform A connects to mitochondria. Thus, the objective of the present study is to evaluate the functionality of eIF5A1 isoform A. HeLa cells were transfected with siRNA‐VA1/2 or siRNA Control and analyzed after 72 and 96 hours. As result, it was observed that the depletion of transcript variant A correlates with reduction in cellular viability and cell cycle arrest. The metabolic profile analysis of these cells revealed that the alternative eIF5A1 isoform has some role in metabolic homeostasis. Indeed, causing negative effects on energy metabolism, as revealed by reduction in lactate production and in the content of transcripts related to mitochondria activity and biogenesis. Thus, it is proposed that eIF5A isoform A protein may modulates the cellular metabolism by acting on oxidative process controlled by mitochondria.

Support or Funding Information

São Paulo Research Foundation (FAPESP)
Metabolic Differences in Nonessential Amino Acid Plasma Concentrations in High‐ and Low‐Active Mice
Authors: Granados Jorge Z. , Ten Have Gabriella A. M. , Letsinger Ayland C. , Deutz Nicolaas E. P. , Lightfoot J. Timothy
Published: November 2023

BACKGROUND

There continues to be a lack of information regarding physiological mechanistic interactions between physical activity and metabolism. This is important in relation to obesity and Type II Diabetes.

PURPOSE

To conduct basic metabolic analysis on mice either high‐ active (HA) (C57L/J) or low‐active (LA) (C3H/HeJ), and determine basic metabolic differences between non‐essential amino acids pathways (NEAA).

METHODS

All procedures were approved by the TAMU IACUC. At 12wks of age, 20 HA male mice (body weight: 27.5±1.2g; lean mass: 22.5±1.3g; fat mass: 2.5±0.7g; food intake: 3.4±0.8g) and 23 LA male mice (body weight: 25.8 ±1.2g; lean mass: 21.0±1.1g; fat mass: 2.5±0.5g; food intake: 3.0 ±1.0g) (Jackson Labs) under anesthesia had a catheter placed in the right jugular vein for blood sampling. Plasma NEAA concentrations were determined by LC‐MS/MS. Data were analyzed using unpaired t‐tests for each NEAA.

RESULTS

LA mice had significantly higher NEAA for GLU, ALA, CIT, ORN and for SER. Remarkable no change in GLN, ARG and GLY (Table).

CONCLUSION

In LA strain mice, specific changes in the GLU/GLN and ARG/CIT/ORN and GLY/SER pathway are observed suggesting modified pathways, linked to their lower activity levels.

Support or Funding Information

Study funded by research development grant from the Vice‐President of Research, Texas A&M University

Amino Acid
Low Active (n=20)
High Active (n=23)
Difference
P

Glutamic Acid (GLU)
36.8 ± 3.0
17.5 ± 2.0
−19.3 ± 3.6
<0.0001

Glutamine (GLN)
641.8 ± 23.9
633.7 ± 20.7
−8.0 ± 31.6
0.8011

Citrulline (CIT)
57.3 ± 2.3
46.4 ±1.5
−10.9 ± 2.8
0.0003

Serine (SER)
185.2 ± 7.1
152.3 ±5.4
−32.9 ± 8.8
0.0006

Glycine (GLY)
401.0 ± 15.4
399.0 ± 12.0
−2.1 ± 19.5
0.9167

Arginine (ARG)
104.6 ± 6.7
106.7 ± 4.2
2.1 ± 7.9
0.7895

Alanine (ALA)
671.5 ± 39.3
453.6 ± 18.7
−217.9 ±43.6
<0.0001

Ornithine (ORN)
55.4 ± 7.3
36.9 ± 2.6
−18.6 ± 7.8
0.0241

SUM NEAA
2153.6 ±105.0
1846.1 ±67.1
−307.6 ± 125.6
0.7632

Plasma concentrations in μM. Data are mean ± SE.
Metabolic Differences in Essential Amino Acid Plasma Concentrations in High‐ and Low‐Active Mice
Authors: Letsinger Ayland C. , Granados Jorge , Ten Have Gabriella A. M. , Deutz Nicolaas E. P. , Lightfoot J. Timothy
Published: November 2023

BACKGROUND

With current health problems such as obesity and Type II Diabetes attributable to both physical inactivity and metabolic disorders, there continues to be a lack of information regarding physiological mechanistic interactions between physical activity and metabolism.

PURPOSE

To conduct basic metabolic analysis on mice either high‐ active (HA) (C57L/J) or low‐active (LA) (C3H/HeJ), and determine basic metabolic differences between essential amino acids (EAA) in these activity types.

METHODS

All procedures were approved by the TAMU IACUC. At 12wks of age, 20 HA male mice (body weight: 27.5±1.2g; lean mass: 22.5±1.3g; fat mass: 2.5±0.7g; food intake: 3.4±0.8g) and 23 LA male mice (body weight: 25.8 ±1.2g; lean mass: 21.0±1.1g; fat mass: 2.5±0.5g; food intake: 3.0±1.0g) (Jackson Labs) under anesthesia had a catheter placed in the right jugular vein for blood sampling. Plasma EAA concentrations were determined by LC‐MS/MS. Data were analyzed using unpaired t‐tests for each EAA.

RESULTS

LA mice had significantly higher EAA for THR, VAL, MET, ILE, LEU, TRP and HIS (Table). LYS was the only essential AA with a significantly lower concentration.

CONCLUSION

We hypothesize that the consistent higher EAA plasma concentrations in LA strain mice indicate a modified essential AA metabolism that is linked to their lower activity levels.

Support or Funding Information

Study funded by research development grant from the Vice‐President of Research, Texas A&M University

Amino Acid
Low Active (n=20)
High Active (n=23)
Difference
P

Threonine (THR)
206.7 ± 9.8
161.4 ± 6.7
−45.3 ± 11.9
0.0005

Valine (VAL)
187.9 ±7.1
156.8 ± 4.9
−31.1 ± 8.6
0.0008

Methionine (MET)
60.9 ± 2.7
47.3 ± 1.5
−13.6 ± 3.1
0.0001

Isoleucine (ILE)
57.7 ± 2.3
48.5 ± 1.9
−9.2 ± 3.0
0.0041

Leucine (LEU)
144.8 ± 6.4
125.2 ± 4.8
−19.6 ± 8.0
0.0184

Tryptophan (TRP)
71.1 ± 3.0
58.2 ± 2.3
−13.0 ±3.8
0.0016

Phenylalanine (PHE)
86.7 ± 4.3
77.5 ± 2.4
−9.2 ± 4.9
0.0724

Histidine (HIS)
109.3 ± 3.9
92.4 ± 2.8
−16.8 ± 4.8
0.0011

Lysine (LYS)
379.9 ± 17.2
481.4 ± 18.2
101.5 ± 25.0
0.0002

Sum EAA
1305.0 ± 56.7
1248.7 ± 45.5
−56.3 ± 73.1
0.9139

Plasma concentrations in μm. Data are mean ± SE.
Ethanol Extract of Phlomis Tuberosa L. Promotes Glucose Uptake in 3T3‐L1 Preadipocytes via Insulin Signaling Pathway
Authors: Baatar Delgerzul , Sukhbaatar Otgonpurev , Oyungerel Baatartsogt , Hwang Seong Gu
Published: November 2023

Phlomis tuberosa L. (Lamiaceae) has been used as traditional medicine for treating diabetes, gastric ulcer, hemorrhoids, inflammation and other wounds in Asian countries. In this study, we investigated anti‐diabetic effect of Phlomis tuberosa L. ethanol extract (PTL) through the glucose transporting mode of action in 3T3‐L1 preadipocytes. Effect of PTL ethanol extract was evaluated in following concentration: 25, 50, 100, 200 μg/mL. Cell viability was measured by CCK‐8 assay. Differentiated 3T3‐L1 cells were stained with Oil Red O and triglyceride (TG) content was measured. Adipocytes differentiation related mRNA and protein expression was analyzed by RT‐PCR and Western Blot (IRS‐1, GLUT‐4, PPARγ and C/EBPα). Our findings showed that PTL had no cytotoxic effect on cell viability. Intracellular lipid accumulation was significantly increased in a dose dependent manner. RT‐PCR and Western Blot analysis showed that PLT promoted glucose uptake by increasing the expression of glucose transporter 4 (GLUT‐4) and insulin receptor substrates‐1 (IRS‐1). PTL also upregulated the expression of peroxisome proliferator‐activated receptor‐γ (PPARγ) and CCAAT‐enhancer‐binding protein‐α (C/EBPα) dose dependently. Our result suggest PTL promotes glucose uptake by activating insulin signaling pathway through the upregulation of PPARγ. Phlomis tuberosa L may be used to develop potential therapeutic agent for Type II Diabetes Mellitus (T2DM).

Support or Funding Information

This work was carried out by the support of the Rural Development Administration Grant (Project No. PJ01268703).
Development of an interdisciplinary undergraduate research training program to improve retention and future success of women in STEM
Authors: Nogaj Luiza A , Roberts Lia , Deprele Sylvine
Published: November 2023

In response to the national need to produce skilled and globally aware STEM students, Mount Saint Mary's University, Los Angeles developed an interdisciplinary undergraduate research program with an international fieldwork experience in the summer. The goals of the program are: (1) to train undergraduates in interdisciplinary research methodologies (cancer laboratory techniques, cross cultural survey research, and policy analysis); (2) to collect real fieldwork data in Peru and the U.S.; (3) to analyze and present the results in professional conferences and publications; and (4) to develop leadership skills and teamwork necessary for student success and retention in STEM fields.

Students enrolled in the program spend two semesters doing course work. In the first semester they learn about the molecular, environmental and nutritional causes of cancer from MSMU's STEM faculty and about the disparities in policy and healthcare access in the U.S. and Peru from political science faculty members. In the second semester, the students develop research skills in both STEM disciplines and the social sciences: laboratory research skills in biology and chemistry, and survey methodology in political science. The students develop an evidence gathering plan, design a survey, and seek an IRB approval in preparation for their global research experience over the summer.

During the summer, the students first interview women in urban and rural areas of California and Peru about factors contributing to their health, such as access to healthcare and nutritional habits. For the remainder of the summer, they analyze their data and study target natural products in the chemistry and biology laboratories.

During the culminating, third semester of the program, the students continue doing research, as they turn their focus to the interpretation of data, presentation of their results at conferences, and future directions for the project. Perhaps most importantly, participants in this program develop leadership skills and examine barriers to success for women in STEM. We expect this combination of regular course work with a summer research component, international field study, and leadership training to be an effective way to increase success and retention rates of undergraduate students in the STEM fields at MSMU.

This program is a model for future research topics thus creating a sustainable and meaningful undergraduate research training.