bluuebunny/crossref_metadata_2025 huggingface dataset:
Possible Role of Chitin‐Like Proteins in the Etiology of Alzheimer's Disease
Authors: Prancan Arthur , Lomiguen Christine , Vidal Luis , Kozlowski Piotr
Published: June 2021

Chitin is a β‐linked straight chain carbohydrate matrix monopolymer prominent in invertebrates, from fungi to arthropods. Surprisingly, chitin is now documented in vertebrates, including humans, a component of vertebrate physiology that has been neglected until now. Chitin levels are elevated in Alzheimer's disease (AD) patients, not only in the central nervous system but also in the cerebrospinal fluid and plasma. Elevated levels of chitin lectin have been reported in patients with AD. Chitinase activity varies widely in the human population. Chitin levels can increase in individuals with intrinsically low chitinase activity. Elevated amounts of chitin can reflect accumulation of the small chitin fragments that remain wherever rapid hyaluronan synthesis occurs. Another source of chitin may be from remote fungal infections. Chitin can be toxic for neurons, and its accumulation may lead to the development of AD. We present new suggestions for animal models and treatment modalities that could prove useful in future research endeavors. An unexpected connection with Gaucher's disease patients and their heterozygote relatives is also identified. These chitin‐related mechanisms are novel approaches to AD whose etiology until now has defied explication.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Icariin ameliorates learning and memory function via improving cerebral glucose metabolism disorder in APP/PS1/Tau triple transgenic Alzheimer's disease mice
Authors: Jin Feng , Zhang Ying , Nie Jing , Shi Jingshan
Published: June 2021

OBJECTIVE

To investigate whether icariin (ICA), a flavonoid extracted from the Chinese herb (Berberidaceae epimedium L), can ameliorate learning and memory function via improving cerebral glucose metabolism disorder in APP/PS1/Tau triple transgenic Alzheimer's disease mice (3×Tg‐AD mice).

METHODS

Three‐month‐old male 3×Tg‐AD mice were randomly divided into three groups (n=10 per group): 3×Tg group, 3×Tg + ICA 30 mg/kg group, 3×Tg + ICA 60 mg/kg group, and age‐matched male wild type (WT) mice were randomly divided into two groups (n=10 per group): WT control group, WT+ ICA 60 mg/kg group. ICA in vehicle (0.5% Tween‐80 in distilled water) was given orally once a day for five months in the 3×Tg+ICA groups, 3×Tg and WT control groups were given an equal volume vehicle. Morris water maze was used to detect the learning and memory function of mice. Brain glucose metabolism in 3×Tg‐AD mice was observed by 18F‐FDG microPET imaging technique. Nissl staining and HE staining were used to evaluate the survival neurons in hippocampus of mice. Glucose oxidase assay was used to detect glucose contents in cortex of mice. The protein expression of APP, Aβ1‐40, Aβ1‐42 , glucose transporter 1 (GLUT1) and GLUT3, and the phosphorylation level of tau protein at multiple sites in hippocampus were detected by Western Blot.

RESULTS

Behavioral examination revealed a profound decrease learning and memory function, accompanied by a decrease in number of neuronal cells in 3×Tg‐AD mice. Moreover, the cerebral 18F‐FDG uptake rate per gram tissue was reduced, the glucose contents in the hippocampus and cortex were increased in 3×Tg‐AD mice. In addition, Western blot analysis showed that the expression of APP, Aβ1‐40, Aβ1‐42 proteins and the levels of tau protein phosphorylation at Ser199/202 and PHF‐1 (Ser396/404) sites were increased significantly, followed by a decrease of GLUT1 and GLUT3 expression in hippocampus of 3×Tg‐AD mice. All of these changes in behavioral functions, 18F‐FDG uptake rate, glucose contents, neuronal loss and the related protein expression were reversed when mice were treated with ICA.

CONCLUSION

ICA can improve the learning and memory ability of AD model mice, the mechanism may be related to the improvement of cerebral glucose metabolism dysfunction by increasing the expression of GLUT.

Support or Funding Information

This work was supported by the National Natural Science Foundation of China (81660599), the Foundation of Zunyi Medical University (2013F‐686, F‐738), The Postgraduate Education Foundation of Guizhou Province (KYJJ2017008).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
RNA‐Seq Reveals Potential Mechanisms of Baicalein in Improving Cognition in SAMP8 Mice
Authors: Li Jiaqi , Du Guanhua , Gao Li , Qin Xuemei
Published: June 2021

Objective

Although baicalein has shown benefits for an array of human diseases including dementia, the effects and mechanisms underlying the neuroprotective effect of baicalein have not been fully explored in senescence‐accelerated mouse prone 8 (SAMP8).

Methods

In the present work, 8‐month‐old SAMP8 mice were intragastrically administrated with 200 mg/kg baicalein once a day for eight weeks, and age‐matched SAMR1 mice were used as control group. The effects of baicalein on hypomnesis in SAMP8 mice were evaluated by behavioral tests including Morris water maze, Novel object recognition test, and Olfactory memory test, and the mechanisms were explored by RNA‐seq technology combined with network pharmacology and quantitative real‐time PCR (qRT‐PCR).

Results

The results showed that treatment with baicalein improved the cognitive functions of the SAMP8 mice, including spatial learning and memory, object recognition memory, and olfactory memory. The RNA‐seq results showed that 1035 differentially expressed genes (DEGs) were found in 10‐month‐old SAMP8 mice versus the age‐matched SAMR1 mice. Treatment of baicalein altered 654 DEGs in the SAMP8 mice and among them, 214 DEGs could be reversely regulated by baicalein resulting in similar RNA levels to SAMR1 mice. Based on the DEGs, a gene network was constructed containing 908 nodes and 3771 edges. Fifty hub DEGs and eight signaling pathways including Jak‐STAT signaling pathway and PI3K‐Akt signaling pathway were regulated by baicalein through network pharmacology analysis and KEGG pathway analysis. The hub DEGs in Jak‐STAT signaling pathway and PI3K‐Akt signaling pathway including Prlr, Gh, Il7, Il20rb, Il5, Comp, Ibsp, Gm5741 were verified through qRT‐PCR.

Conclusion

Our results indicate that the effects of baicalein on improving cognitive impairments of SAMP8 mice might be attributed to modulation of Jak‐STAT signaling pathway and PI3K‐Akt signaling pathway.

Support or Funding Information

This project was supported by the National Natural Science Foundation of China (81603319), and partly supported by Science and Technology Innovation Team of Shanxi Province (201605D131045‐18), and Key laboratory of Effective Substances Research and Utilization in TCM of Shanxi province (201705D111008‐21).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Klotho Improves Alzheimer's Disease‐like Pathology and Cognitive Deficits in APP/PS1 Mice
Authors: Du Junrong , Zeng Chenye , Yang Tingting , Zhou Hongjing
Published: June 2021

Alzheimer's disease (AD) is the most common type of senile dementia. The anti‐aging gene Klotho is reported to decline in the brain of AD patients and animals. However, the role of Klotho in the progression of AD remains elusive. The present study explored the effects and underlying mechanism of Klotho in a mouse model of AD. The upregulation of cerebral Klotho expression was mediated by an intracerebroventricular injection of a lentiviral vector that encoded Klotho (LV‐KL) in 7‐month‐old APP/PS1 transgenic mice. Three months later, LV‐KL significantly induced Klotho overexpression in the brain and effectively ameliorated cognitive deficit and Alzheimer's disease‐like pathology in APP/PS1 mice. LV‐KL induced autophagy activation both in AD mice and BV2 murine microglia. These results suggest that the upregulation of Klotho expression in the brain may promote the autophagic clearance of Aβ and protect against cognitive deficits in AD mice. These findings highlight the preventive and therapeutic potential of Klotho for the treatment of AD.

Support or Funding Information

This work was supported by the National Science Foundation of China (81473219), and partly by targeting drug delivery system of Sichuan Province Youth Science and Technology Innovation Team (2016TD0001) and 111 Project of the National Ministry of Education (B18035).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Delta‐9‐tetrahydrocannabinol (THC) Enhances Heroin‐Induced Antinociception and Disruptions of Learning and Performance in Rhesus Macaques
Authors: Nilges Mark R , Grace Joseph A , Winsauer Peter J
Published: June 2021

Cannabinoid administration prior to opioid administration can produce opioid‐sparing antinociceptive effects in several species by reducing the dosage of opioid needed to produce effective antinociception. These potentially valuable clinical effects could be undermined, however, if the cannabinoids also enhance the disruptive effects of opioids on neurobehavioral function. To address this question, the combined effects of THC and heroin were examined in nonhuman primates (rhesus macaques, n=4) responding in a learning and performance task and concurrently exposed to a tail‐withdrawal antinociception procedure. Testing in this case was facilitated by examining 4 discrete cycles of acquisition and performance that were separated by timeout periods to facilitate tail‐withdrawal tests and cumulative dosing. Each cycle included a 10‐min time out period for drug onset, a 15‐min acquisition component, and a 5‐min performance component. In the learning and performance task, subjects acquired a new 5‐response key‐sequence, or emitted a well‐rehearsed 5‐response key‐sequence, respectively. Responding in both components of the task was reinforced with a food pellet after every three completions of the sequence (i.e., a second‐order fixed‐ratio [FR3] schedule of food reinforcement). Warm‐water tail‐withdrawal antinociception was measured using three water temperatures (40°, 50°, and 55°C). Heroin alone dose‐dependently decreased response rates in the acquisition and performance components without markedly increasing percent errors. When the opioid antagonist naltrexone (1 mg/kg) preceded the cumulative heroin injections, the heroin dose‐effect curves for both the behavioral disruptions and antinociception were shifted rightward indicating both effects were mediated by mu‐opioid receptors. In contrast, THC alone disrupted both behaviors at doses that had no antinociceptive effect, but it shifted the heroin dose‐effect curves for learning, performance, and antinociception leftward. Overall, these data demonstrate that THC produces its opioid‐sparing antinociceptive effects at doses that are behaviorally disruptive alone and in combination with heroin.

Support or Funding Information

This work was supported by a National Institute of Drug Abuse Fellowship (F32DA47013‐01)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Comparing the Neuroprotective Efficacy of P7C3‐A20 to Novel P7C3 Derivatives in a Rodent Model of Paclitaxel‐Induced Peripheral Neuropathy
Authors: Smith Hudson R. , LoCoco Peter M. , Chavera Teresa A. , Zamora Joshua C. , Jennings Elaine M. , Pando Miryam M. , Berg Kelly A. , Clarke William P.
Published: June 2021

Chemotherapy‐induced peripheral neuropathy (CIPN) is a prevalent adverse effect of anti‐cancer chemotherapeutics like paclitaxel (PTX), a microtubule targeting agent known to induce severe and persistent neuropathic pain. The development of CIPN not only diminishes the quality of life of cancer patients, but also downgrades their cancer prognosis as dose reduction or cessation of their anti‐cancer regimen are the only clinical interventions currently available for CIPN. Thus, the development of a pharmacological intervention that limits the neuropathic side‐effects of anti‐cancer chemotherapy is urgently needed. Here we tested the neuroprotective effects of the aminopropyl carbazole, P7C3‐A20 (A20), and several A20 derivatives in a rat model of paclitaxel‐induced peripheral neuropathy. All experiments were done by investigators blinded to the drug treatments. Racemic A20 (2.2, 6.6, 20 mg/kg, i.p., q.d.), or vehicle, was administered daily to male Sprague‐Dawley rats over a 14‐day period beginning 2 days before PTX treatment (11.7 mg/kg/day, i.p.; every other day for 3 injections). A20 dose‐dependently attenuated PTX‐induced mechanical allodynia in rats. Evaluation of intraepidermal nerve fiber (IENF) density in rat paw biopsies using immunohistochemical staining revealed that treatment with PTX produced a profound loss in IENF density that was prevented by treatment with A20. Further, statistical analysis indicated strong correlations between IENF density, a diagnostic marker of CIPN in humans, and nociceptive threshold to mechanical stimuli. Enantiomer specific mixtures of A20 (20 mg/kg, i.p., q.d.), racemic A20 (20 mg/kg, i.p., q.d.), or vehicle were then administered utilizing the same behavioral paradigm and PTX dosing schedule as above. Consistent with our previous data, racemic A20 attenuated PTX‐induced mechanical allodynia and the S‐enantiomer was identified as the active component of the racemic mixture. We have also tested the neuroprotective efficacy of four novel A20 derivatives coded as compounds A through D. The A20 derivatives were administered daily at 20 mg/kg by i.p. injection and mechanical allodynia was assayed as described above. One compound, Drug C, provided robust protection against PTX‐induced mechanical allodynia, whereas the compounds A, B, and D demonstrated limited neuroprotective efficacy. Taken together, A20 and perhaps its derivatives are an exciting new class of pharmacological agents that may have clinical utility for alleviating peripheral neuropathies caused by various anti‐cancer chemotherapeutics.

Support or Funding Information

Supported in part by Calico Life Sciences and Proneurotech.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hydrogen‐Deuterium Exchange Reveals Distinct Activation States of PLCβ2 BY G‐Proteins
Authors: Fisher Isaac , Jenkins Meredith , Tall Greg , Burke John , Smrcka Alan
Published: June 2021

G‐protein coupled receptors (GPCRs) are an important class of cell surface receptors that transduce extracellular stimuli to intracellular signaling events via G‐proteins. One major effector of G proteins is phospholipase C (PLC), which hydrolyzes Phosphatidylinositol 4,5 bisphosphate (PIP2) into Diacylglycerol (DAG) and Inositol 1,4,5, trisphosphate (IP3). PLCβ2 and PLCβ3 isoforms are directly activated by binding Gαq, Gβγ or both. The mechanism of activation of PLCβ2 by Gβγ is unknown, and the binding site(s) for Gβγ on PLCβ remain controversial.

Here we use hydrogen‐deuterium exchange mass spectrometry (HDXMS) to understand the conformational dynamics of PLCβ and to answer open questions in the field concerning the mechanism of activation of PLC by Gβγ and Gαq. For these experiments, several complexes were examined with HDXMS: PLCβ2 alone, PLCβ2+ PIP2/Phosphoethanolamine (PE)/Phosphatidylserine (PS) vesicles, or PLCβ2+ PIP2/PE/PS vesicles plus Gβγ or Gαq‐AlF4. Incubation of PLCβ2 with PIP2/PE/PS vesicles strongly protects regions in the C‐terminus, confirming that this region is involved in membrane binding. In the presence of Gαq‐AlF4, PLCβ2 shows protection from deuterium exchange in the helix‐loop‐helix region of the proximal C‐terminal domain (CTD) and the EF‐hand domain on PLCβ2 that exactly correspond to regions shown to interact with Gαq in the Gαq‐PLCβ3 crystal structure. These data validate that our approach accurately reports bona fide protein‐protein and protein‐lipid interactions. Incubation of Gβγ with PLCβ2 does not reveal any regions of strong protection, but rather causes strong increases in deuterium exchange in the of the CTD of PLCβ2, indicating that the CTD of PLC undergoes a large conformational rearrangement upon Gβγ binding. This increase in deuterium exchange of the CTD of PLC is not seen upon Gαq activation. This indicates that there are distinct conformational states of PLCβ induced by Gαq and Gβγ interactions, suggesting that Gαq and Gβγ use different mechanisms for activation of PLCβ. To determine the mechanism of activation of PLCβ, we used a purified reconstitution system of PLC activity. In vitro reconstitution experiments show that CTD‐deleted PLCβ2 exhibits increased activity in response to Gβγ. Additionally, the distal CTD of PLCβ inhibits CTD‐deleted PLCβ2 activity in both lipid‐dependent and lipid‐independent reconstitution system. Taken together, these results suggest that Gβγ activates PLCβ by breaking an autoinhibitory interaction of the distal CTD with the core enzyme of PLC.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
How Do Taste Buds EAT?: Defining the Embryo‐to‐Adult Transition in Mouse Taste Bud Development and Regeneration
Authors: Barlow Linda A , Golden Erin J , Fellin Tim J
Published: June 2021

Taste buds are collections of heterogeneous taste receptor cells (TRCs) that detect sweet, sour, salt, bitter and umami. TRCs are relatively short‐lived and continually replaced by proliferative progenitor cells situated adjacent to taste buds. In mouse embryos, taste bud precursors are first evident as small clusters of columnar epithelial cells or placodes on the developing tongue surface. Each taste placode comprises 10–20 cells that express both cytokeratin (K) 8 and the secreted protein Sonic Hedgehog (SHH). Previously we showed that SHH+ placode cells differentiate into TRCs in the first postnatal week, but do not give rise to the K14+ progenitor population that supports adult TRC renewal (Thirumangalathu et al., 2009 Development). This raised the questions of how and when TRC turnover commences. Using 3D image analysis, we find that taste placode and taste bud cell number are static during embryonic development and within 1–2 days of birth, respectively; instead EdU incorporation studies and K14 genetic lineage tracing reveal that the progenitor contribution to taste buds begins by postnatal day (P) 2, steadily increasing through P14. Work from our lab and others has shown that SHH is a negative regulator of taste cell fate in embryos but paradoxically SHH promotes TRC differentiation in adults. Thus, we hypothesized that the switch in SHH function coincides with the onset of taste cell renewal. We are currently testing this idea using K14CreER to drive SHH overexpression at specific postnatal time points. In cultured embryonic tongues, excess SHH represses formation of taste placodes, whereas in adult mice, this genetic manipulation induces formation of ectopic taste buds throughout the anterior lingual epithelium (Castillo et al., 2014 Development). Additionally we are using RNAseq analysis to identify candidate regulators of the embryo‐to‐adult transition, and more specifically to define the emerging taste bud progenitor pool in postnatal animals.

Support or Funding Information

Supported by R01 DC012383 to LAB and F32 DC015958 to EJG

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
b1‐Adrenergic Receptors in the Golgi Apparatus are Activated by Cell Permeable Agonists and Stimulate PLC‐mediated PI4P Hydrolysis in Cardiac Myocytes
Authors: Wei Wenhui , Nash Craig , Smrcka Alan
Published: June 2021

Previously, our laboratory identified a phospholipase C (PLC)‐mediated PI4P hydrolysis pathway at the Golgi apparatus that is important for regulation of cardiac hypertrophy, a pathology associated with development of heart failure. cAMP stimulates this pathway via Epac‐mediated activation of Rap1 which directly binds to PLCɛ. We recently found that a cell membrane permeable β‐adrenergic receptor (βAR) agonist, dobutamine (dob), induces Golgi PI4P hydrolysis in neonatal rat ventricular myocytes (NRVMs) while a cell membrane impermeable β‐adrenergic (βAR) agonist, Isoproterenol, does not. We used a YFP‐tagged mini Gs protein2 which is recruited to Gs‐coupled receptors upon their activation, to monitor the membrane location of these activated receptors. Dobutamine induced robust and rapid recruitment of mini Gs to both the plasma membrane, and the perinuclear region corresponding to the Golgi apparatus. In contrast, Isoproterenol, induced a strong recruitment to the plasma membrane but not to the Golgi. Treatment with brefeldin A significantly reversed mini Gs recruitment to the internal membrane induced by dobutamine, supporting the idea that active β1‐ARs are localized to the Golgi membrane. A membrane permeable βAR antagonist, metoprolol, fully reversed dob‐mediated PI4P hydrolysis and mini Gs recruitment to the Golgi membrane while a membrane impermeable antagonist, sotalol had no effects on dob‐mediated PI4P hydrolysis and only partially reversed the mini Gs recruitment to the Golgi membrane. In addition, an Epac inhibitor HJC0726 completely inhibited dob‐mediated PI4P hydrolysis whereas gallein, a Gβγ inhibitor had no effect. Over expression of the RA1 domain of PLCɛ, which competes PLCɛ away from the mAKAPβ scaffold, also inhibited dobutamine stimulation of PI4P hydrolysis. These data indicate that activation of endogenous intracellular β1ARs stimulates production of a local pool of cAMP that activates the perinuclear Epac‐PLCɛ‐PI4P hydrolysis pathway involved in heart failure.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Roles of GPR39 in Regulating Keratinocyte Proliferation and Inflammation
Authors: Satianrapapong Wilasinee , Pongkorpsakol Pawin , Muanprasat Chatchai
Published: June 2021

Zinc is an important trace element that plays key roles in the human body. Previous reports showed that zinc supplement could relieve dermatitis and improve wound healing process with unknown molecular target. In this study, we investigated the roles of G‐protein coupled receptor 39 (GPR39), a zinc‐sensing receptor, on keratinocyte proliferation and inflammation using a GPR39 agonist TC‐G 1008. Immortalized human keratinocyte cell line (HaCaT cells) was used as an in vitro model. We first verified protein expression and function of GPR39 in HaCaT cells by Western blot and Indo‐1 calcium flux assays. Importantly, at 48 h post‐treatment, Bromodeoxyuridine (BrdU) incorporation assay showed that TC‐G 1008 significantly increased keratinocyte proliferation in a concentration‐dependent manner with a maximal effect being observed at 1 μM. Moreover, TC‐G 1008 enhanced mammalian target of rapamycin (mTOR) phosphorylation in a time‐dependent manner, indicating that GPR39 activation led to keratinocyte proliferation. Next, immunofluorescence staining demonstrated that TC‐G 1008 significantly reduced tumor necrosis factor‐alpha (TNF‐α)‐induced nuclear factor‐kappa B (NF‐κB) nuclear translocation. Interestingly, this effect was abolished by co‐treatment with compound C, AMP‐activated protein kinase (AMPK) inhibitor. These results supported that GPR39 activation promotes wound healing and exerts anti‐inflammatory effect in human keratinocytes, at least in part, by stimulating mTOR and AMPK, respectively.

Support or Funding Information

This work was supported by Thailand Research Fund and Mahidol University (BRG5980008), Faculty of Science, Mahidol University and the Medical Scholars Program (Ph.D.‐M.D. program), Mahidol University.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
In Vitro Evaluation of Excipients as Inhibitors of Human Intestinal P‐glycoprotein
Authors: Bajaj Ruchika , Chong Lisa B , Zou Ling , Tsakalozou Eleftheria , Ni Zhanglin , Giacomini Kathleen M , Kroetz Deanna L
Published: June 2021

P‐glycoprotein (P‐gp) is one of the major barriers to oral bioavailability and contributes to the interindividual variability observed in intestinal absorption of P‐gp substrates. A well characterized source of variable P‐gp function in the intestine is inhibition by co‐administered drugs. However, the potential interplay between intestinal P‐gp and pharmaceutical excipients used in orally administered drug products has not been systematically studied and is the focus of this investigation. A fluorescence assay using calcein AM (5 mM), which served as a model substrate for P‐gp, was used to assess inhibition of P‐gp transport function by 130 excipients in HEK293 cells stably expressing human P‐gp or transfected with an empty vector (control). Specific inhibition of P‐gp‐mediated calcein AM efflux by excipients was expressed as a ratio of inhibition observed in P‐gp overexpressing cells over control cells. An inhibition screen was performed at an excipient concentration of 200 mM where possible, or at 50 mM where excipient solubility was limiting. Six potential inhibitors (inhibition ratio >1.3) were identified, including five dyes (D&C Red #6, D&C Brown#1, Naphthol blue black, Acid blue 9, Light green CF yellowish) and one suspending agent (Butylparaben). None of these potential inhibitors achieved >30% inhibition of calcein‐AM efflux at concentrations up to 300 mM. Intestinal concentrations of these potential inhibitors estimated from the maximum amount of each excipient allowed per unit dose (http://excipients.ucsf.bkslab.org/) ranged from 0.50–2.3 mM, with the exception of light green CF yellowish (214 mM). While these in vitro findings indicate minimal inhibition of human P‐gp at intestinal excipient concentrations typically expected from marketed drugs, further studies are needed to assess the impact of these excipients on the oral bioavailability of P‐gp substrates.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Caveolin scaffolding domain plays an important role in cancer cell migration
Authors: Okerblom Jonathan , Okada Sunaho , Raja Sadaf , Ray Supriyo , Okada Hideshi , Kawamura Itta , Murray Fiona , Murofushi Yoshiteru , Patel Hemal
Published: June 2021

Caveolin‐1 is the integral membrane protein that plays an important role in the structure and function of terminally differentiated cells. As a structural component of caveolae, the caveolin scaffolding domain (CSD) has been associated with many cellular signaling processes and recent reports have implicated Cav‐1 as a negative regulator in tumor metastasis. We hypothesized that Cav‐1 may be inhibiting cell migration through its CSD. To assess this, HCT‐116 cells (which contain high endogenous Cav1) and HT‐29 cells (which contain low endogenous Cav1) were transduced with a lentiviral construct expressing either Cav‐1 or Cav‐1 without a functional CSD (ΔCSD). HeLa cells were also genetically engineered to stably overexpress Cav‐1(Cav‐1 OE), ΔCSD, or enhanced green fluorescent protein (EGFP) as a comparative control. In all cases, migration was significantly greater in ΔCSD cells compared to Cav‐1 OE and control cells. Phosphorylated STAT‐3 was also decreased in Cav‐1 OE cells compared to control and ΔCSD cells. G2/M phase was significantly increased and G1 was decreased in ΔCSD cells compared to control and CAV1 OE cells. Cdc2 phosphorylation was also increased in ΔCSD cells. To further confirm that CSD affects cell migration, we performed migration assays in Hela cells expressing either a pEGFP tagged CSD or a scrambled sequence. CSD overexpressing cells showed a lower migration rate comparison to control cells. These findings suggest that Cav‐1 CSD affects cell migration, possibly by participating in a G2/M checkpoint. These findings may represent a new approach to the treatment of cancer metastasis.

Support or Funding Information

NIH R01 HL091071

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Extracts of the medicinal plants Pao Pereira and Rauwolfia vomitoria inhibit ovarian cancer stem cells in vitro
Authors: Chen Qi , Dong Ruochen , Chen Ping
Published: June 2021

Objectives

Ovarian cancer has an enrichment of stem‐like cancer cells (CSCs) which contribute to the treatment‐resistant tumor's high rate of recurrence and metastasis. Here we investigated two extracts from the medicinal plants Pao Pereira (Pao) and Rauwolfia vomitoria (Rau) each for their activities against ovarian CSCs.

Methods

The Pao and Rau extracts were provided by Natural Source Inc. Cell viability was detected by MTT assay. Ovarian CSC population was examined by tumorsphere formation assays. Ovarian CSC specific cell surface markers were examined by flow cytometry. Western blot and RT‐qPCR was used for protein and mRNA expression.

Results

Both Pao and Rau inhibited the proliferation of multiple human ovarian cancer cell lines with IC50 values ranging from 210 to 420 μg/mL for 48 hours of treatment, and had limited cytotoxicity to normal ovarian epithelial cells. Pao and Rau had similar potency in inhibiting ovarian CSCs, with IC50s of ~120 μg/mL for 24 hours treatment in reducing CD44+CD117+ sub‐population, and IC50s of ~50 μg/mL for long‐term tumorsphere formation. Nuclear β‐catenin levels were decreased, suggesting suppression of Wnt/β‐catenin signaling pathway.

Conclusions

Data here showed that Pao and Rau both inhibited ovarian cancer stem cells in preference to the bulk of tumor cells. These data provide the first evidence in the activities of Pao and Rau in inhibiting ovarian cancer stem cells. Further mechanistic studies and in vivo investigation validating these findings are a warrant, giving that inhibition of cancer stem cells holds the promise of comprehensively inhibiting cancer metastasis, drug resistance, and recurrence.

Support or Funding Information

The study was supported by the Beljanski Foundation. The Beljanski Foundation has no influence on the study design, performance of experiments, data collection and analysis.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Pharmacologic Inhibition or Genetic Deletion of Soluble Epoxide Hydrolase Improves Survival Following Myocardial Infarction in Aged Mice
Authors: Jamieson Lockhart , Sosnowski Deanna K , Darweshe Ahmed M , Wang Wang , Zhabyeyev Pavel , Edin Matthew , Zeldin Darryl , Kassiri Zamaneh , Oudit Gavin , Seubert John M
Published: June 2021

Objective

Myocardial infarction (MI) accounts for a significant proportion of death and disability in aged individuals. CYP450 metabolism of n−6 PUFA arachidonic acid results in formation of numerous metabolites, called eicosanoids, that exhibit a wide range of cellular effects. Previous studies demonstrate eicosanoids can induce alterations to the mitochondria resulting in preserved cardiac function following ischemic injury. These metabolites are further metabolised by the enzyme soluble epoxide hydrolase (sEH) reducing their endogenous activity. This study investigated post‐ischemic cardiac function in aged mice with either genetic deletion or pharmacologic inhibition of sEH. Furthermore, a primary component of this study was to elucidate potential sex differences between male and female mice undergoing the experimental protocol.

Methods

Male and female WT and sEH null mice averaging 15 months old underwent permanent occlusion of the left anterior descending coronary artery (LAD). On the day of surgery, WT mice were given either vehicle (0.1% DMSO) or sEH inhibitor tAUCB (10 mg/ml dissolved in 0.1% DMSO) in drinking water for 28 days ad libitum. Cardiac function was assessed at baseline, 7 days and 28 days post‐MI by echocardiography and electrocardiogram. Protein expression was determined using immunoblotting techniques while mitochondrial enzymatic activities were assessed by spectrophotometry. Mitochondrial respiration in cardiac fibres was measured using a Clark‐type electrode. A full metabolite profile was determined in control and 28‐day post‐MI hearts by LCMS/MS.

Results

Pharmacological inhibition or genetic deletion of sEH was associated with fundamental changes in the n−3 and n−6 PUFA metabolite profile in cardiac tissue, correlating with better diastolic function, preserved mitochondrial respiration and overall improved survival at 28‐days post‐MI.

Conclusions

These data suggest that pharmacologic inhibition or genetic deletion of sEH elicits fundamental changes in the aged myocardium following long‐term ischemic injury.

Support or Funding Information

This research is supported by grants from the Canadian Heart and Stroke Foundation and Canadian Institute of Health Research. LJ is supported by a graduate studentship from Alberta Innovates‐Health Solutions.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Prenatal exposure to Bisphenol‐F followed by chronic estrogen exposure in adulthood increases systolic blood pressure in female rats
Authors: Chuang Yen‐Jun , Jeyaraj Ninitha A. , Jacob Elyssa B. , MohanKumar Puliyur S. , MohanKumar Sheba M. J.
Published: June 2021

Endocrine disrupting chemicals (EDCs) such as bisphenols and phthalates are produced in high volume in the US and all over the world. They are capable of leaching out of plastics and entering the food chain. They produce a variety of effects on the body, but not all effects are understood. Epidemiological evidence indicates that there is a marked increase in the incidence of hypertension and pre‐hypertension in young children. We hypothesized that exposure to these EDCs during pregnancy can affect brain circuits and cause high blood pressure in offspring. The risk is even greater for female offspring who are subsequently exposed to an environmental estrogen or oral contraceptives in adulthood. To test this, we exposed pregnant Sprague Dawley dams to saline, or 1μg/kg BW of bisphenol‐F (BPF). Two female offspring from each dam were either sham implanted or implanted with estradiol‐17β (E2) pellets that were capable of releasing 20 ng of E2/day for 90 days. Animals were also implanted with a telemeter (Data Sciences International; HD‐S10) in the femoral artery and were allowed to recover. Blood pressure recordings were obtained for once a week for 12 weeks over a 24‐hour time period. In control rats that were treated with E2, there was a modest increase in systolic BP. When prenatal BPF exposure was followed by exposure to E2 in adulthood, it produced a greater increase in systolic BP (p = 0.0031 during the day; p= 0.0081 at night). In contrast, prenatal BPF exposure followed by sham implantation resulted in marked increases in diastolic BP. These results indicate that prenatal exposure to BPF can increase BP in female rats. This risk is augmented when females are subjected to chronic low dose E2 exposure in adulthood. Further studies are needed to understand the underlying mechanisms.

Support or Funding Information

Supported by UGA research foundation and start‐up funds.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
DL0108 suppresses the development of pulmonary arterial hypertension in rats through its protective effect on pulmonary arteries
Authors: Yuan Tianyi , Chen Di , Lyu Yang , Fang Lianhua , Du Guanhua
Published: June 2021

Background

Pulmonary arterial hypertension (PAH) is a severe disease but in short of an effective drug. DL0108 is a flavonoid discovered by our lab, and is recently found useful in the treatment of PAH. The present study aims to evaluate the potency of DL0108 in PAH rats induced by monocrotaline (MCT).

Method

The PAH rat model was established by subcutaneous injection of 50 mg/kg MCT on day 0. From the second day, 3, 10, or 30 mg/kg DL0108 was respectively administrated (i.p.) daily for 4 weeks in three tested groups, while the model and control group were treated with normal saline. During the experiment, the survival rate and body weight were recorded. At the end of the study, the right ventricular systolic pressure (RVSP) was measured through right cardiac catheterization. The heart ventricle was isolated to calculate the Fulton index (RV/LV+S). The contraction and relaxation of pulmonary artery rings were tested in a classical isolated vessel experiment.

Result

On day 28, the survival rate of the model group was 66.7%. Treatment with 3, 10, or 30 mg/kg DL0108 increased the survival rate to 73.3%, 86.7% and 86.7%, respectively. However, the decrease of body weight induced by PAH was not improved by DL0108. The RVSP of PAH rats in the model group increased from 21.73 to 49.61 mmHg, while the treatment with DL0108 reduced the RVSP to 30.56, 27.42 or 24.9 mmHg, respectively. Meanwhile, DL0108 significantly reduced the right ventricular hypertrophy in a dose‐dependent manner. In the isolated vessel experiment, it was found that 3 and 10 mg/kg DL0108 improved the contraction response of pulmonary artery rings to 10−6 M phenylephrine and the relaxant effect induced by the accumulation of acetylcholine (10−8–10−4 M Ach).

Conclusion

Early intervention of PAH rats with DL0108 can delay the progression of the disease and increase the survival rate. The preliminary study indicates that DL0108 protects the function of pulmonary arteries, relieves the remodeling of vessels to control the pressure, and in turn attenuates the injury of heart ventricle. This study contributes to the further development of DL0108 for the treatment of PAH.

Support or Funding Information

This project was supported by grants from the Nation Natural Science Foundation of China (No. 81603101, 81773935, 81573645), Natural Science Foundation of Beijing (No. 7174322) and CAMS Innovation Fund for Medical Sciences (2016‐I2M‐1‐010, 2017‐I2M‐1‐010).

The survival rates of PAH rats treated by DL0108.

image

The survival rates of PAH rats treated by DL0108.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Spinal Instability in Rats Counteracted by Pentadecapeptide BPC 157
Authors: Dokuzovic Stjepan , Bebek Igor , Perovic Darko , Bilic Vide , Corluka Stipe , Krezic Ivan , Durasin Tajana , Drmic Domagoj , Seiwerth Sven , Sikiric Predrag
Published: June 2021

To induce spinal instability, we focused on bilateral facetectomy in rats and possible therapeutic benefit with the stable gastric pentadecapeptide BPC 157 given in the drinking water. Recently, using a well‐designed rat model (L2‐L3 compression), we demonstrated that the stable gastric pentadecapeptide BPC 157 improved the spinal cord injury (FASEB J 2015, 29, 617.5), concluding that under the conditions of one single application, given soon after spinal cord injury, motor function was markedly recovered, axonal and neuronal necrosis was counteracted, as was demyelination and cyst formation. Previously shown to counteract the consequence of peripheral (sciatic) nerve transection/anastomosis, improve nerve healing (Regul Pept. 2010 Feb 25;160(1–3):33–41.), brain trauma (Regul Pept. 2010;160(1–3):26–32.), and various encephalopathies (Curr Pharm Des. 2018;24(18):1990–2001), BPC 157 (LD1 not achieved) was implemented as an anti‐ulcer peptide in inflammatory bowel disease trials and now in a multiple sclerosis trial(Curr Pharm Des. 2018;24(18):1990–2001). In this study, the bilateral paravertebral muscles attached to the L3–L4 segment were peeled from the lumbar spine to expose the posterior bony elements. The rats then underwent complete resection of bilateral L3–L4 facet joints without neural tissue injuries. The medication was given in drinking water (BPC 157 10 ng/kg, 0.16 ng/mL, 12 ml/rat/day) through next 8 weeks, while controls received drinking water only. Radiological assesment of rat spines acquired at 1 week or 8 weeks was conducted (Figure 1). At 1 week, controls and BPC 157 animals exhibited no gross deformity observed in any plane, disk spaces seem unaffected, neural foramen at operated level is slightly widened, though BPC 157 drinking animals had higher bone density overall. At 8 weeks, both groups exhibited no gross deformity observed in any plane, unaffected disk spaces, widened neural foramen at the operated level is slightly widened, though there was an additional discrepancy in bone density overall between groups (BPC 157 having markedly more sclerotic and better defined bones). Also worth noting is the lack of abundant callus formation in the BPC 157 rats whereas the controls have visible callus formation in seemingly random patterns near the affected level. In addition, a significant motor disability which appears in controls immediately upon injury induction, was completely counteracted in BPC 157 drinking rats.

Support or Funding Information

This work was supported by the Ministry of Science, Education and Sports, Republic of Croatia [grant number 108‐1083570‐3635].

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Stable Gastric Pentadecapeptide BPC 157 Recovers Motor Function After Rat Spinal Cord Injury
Authors: Perovic Darko , Krezic Ivan , Dokuzovic Stjepan , Bilic Vide , Gojkovic Slaven , Drmic Domagoj , Seiwerth Sven , Sikiric Predrag
Published: June 2021

Using a well‐designed rat model, we demonstrated that the stable gastric pentadecapeptide BPC 157 improved the spinal cord injury (FASEB J 2015, 29, 617.5). Previously shown to counteract the consequence of peripheral (sciatic) nerve transection/anastomosis, and improve nerve healing (Regul Pept. 2010 Feb 25;160(1–3):33–41.), and brain trauma (Regul Pept. 2010 Feb 25;160(1–3):26–32.), and various encephalopathies (Curr Pharm Des. 2018;24(18):1990–2001), BPC 157, LD1 not achieved, was implemented as an anti‐ulcer peptide in inflammatory bowel disease trials and now in a multiple sclerosis trial(Curr Pharm Des. 2018;24(18):1990–2001). In a study from the day 1 to 1 year, rats underwent laminectomy (level L2–L3), 60 second compression (neurosurgical piston (60–66 g) on exposed dural sac with sacrocaudal spinal cord) received one medication of BPC 157 (200 or 2 μg/kg) or saline (5 ml/kg)) intraperitoneally at 10 min after injury (FASEB J 2015, 29, 617.5). All injured rats underwent BPC 157 exhibit consistent improvement, clinically, constantly better tail motor function, no autotomy; resolved spasticity already by the day 15; microscopically (since day 7), vacuolas and loss of axons in white matter, edema and loss of motoneurons in gray matter, and decrease of the number of large myelinated axons in rat caudal nerve were largely counteracted in BPC 157 rats. EMG recording evidences markedly lower MUP in tail muscle. Concluding, under the conditions of one single application, given soon after spinal cord injury, counteracted were axonal and neuronal necrosis, demyelination, cyst formation. Now, to create, however, a more reliable situation and therapy effect, the therapy with BPC 157 was started after 4 days. The medication was given in drinking water (BPC 157 10 μg/kg, 0.16 μg/mL, 12 ml/rat/day) through next 4 weeks, while controls received drinking water only. Specifically, tail motor function was scored as follows: 1 ‐ complete loss of tail function; 2 ‐ elevation maximum of 1/4 of tail length; 3 ‐ elevation maximum of 1/2 of tail; 4 ‐ elevation maximum of 3/4 of tail length; 5 – normal function. During evaluation, tail motor function score remained debilitated in rats underwent spinal cord injury that received post‐injury. Contrarily, BPC 157 treated rats exhibited huge improvement, which was continued to full recovery.

Conclusion

We documented that even given later after injury, the neuroprotective effect of BPC 157 quickly appears, and, BPC 157 given in drinking water, accordingly recovers the failed motor function in rats with spinal cord injury.

Support or Funding Information

This work was supported by the Ministry of Science, Education and Sports, Republic of Croatia [grant number 108‐1083570‐3635] .

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Ataxia in the ASIC5 Knockout Mouse Associated with Decreased Excitability of Vestibulocerebellum Unipolar Brush Cells
Authors: Boiko Nina Y. , Kreko‐Pierce Tabita , Pugh Jason , Stockand James D.
Published: June 2021

Acid‐sensing ion channels (ASICs) are neuronal ligand‐gated sodium channels. They are the members of the Degenerin/Epithelial Na+ channel (Deg/ENaC) family. In mammalian neurons ASICs play an important role in a variety of complex behaviors such as fear, pain, anxiety, learning, and memory. Among all five members of ASIC1‐5 channels, ASIC5 is least known. To reveal the expression and function of this channel we generated ASIC5 reporter and knockout mice. We found that ASIC5 is restrictively expressed in a subset of interneurons in the granular layer of nodulus of the vestibulocerebellum. Immunohistochemical and morphological analysis identified these interneurons as unipolar brush cells (UBC) that also express the metabotropic glutamate receptor 1α (mGluR1α). Such observations are consistent with ASIC5 playing a key role in the physiology of UBCs and in the function of the vestibulocerebellum. Consistent with this, knockout of ASIC5 results in ataxia and uncoordinated bodily movements. Electrophysiological analysis of UBC revealed a decrease in excitability of these neurons in the ASIC5 knockout mice that underlies these behavioral dysfunctions.

Support or Funding Information

This work was funded by a NIH Ro1 grant (DK113816)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.