bluuebunny/crossref_metadata_2025 huggingface dataset:
Intermittent Normobaric Hypoxic Training Protects the Brain by Regulating Microglia Phenotype
Authors: Joshi Akita , Han Heping , Lewis Sally , Ryou Myoung‐gwi
Published: June 2021

Introduction

Ethanol Withdrawal (EW) syndrome is an etiologically important factor linked to the ischemia and reperfusion injuries, including ischemic stroke. In the US, ischemic stroke is the 4th leading cause of death and the first cause of adult disability. Previously we showed that intermittent normobaric hypoxic training (IHT) protects the brain from EW syndrome by reducing ROS and increasing protective proteins. Pro‐inflammatory microglia has been studied in various CNS diseases. However, the function of microglia in the IHT‐induced brain protective mechanism is not yet thoroughly explored. In here, we focused on how IHT influence on regulating the phenotype of microglia, a pivotal resident phagocytic cell in the brain, in the in vitro IHT model.

Methods

Mouse microglia cells, EOC20 cell, underwent three days of IHT program which composed of 5–7 cycles of hypoxia/normoxia. O2 concentration was gradually decreased from 2.5% to 2 % each daily training cycle. Two sessions of EW treatment was done as following; EtOH (100 mM) for 16 hrs followed by 6 hrs of EtOH cessation. Cells were assigned into four groups; 1) non‐treated control, 2) IHT, 3) EW, 4) IHT+EW. Cell viability was evaluated by Calcein AM assay. Immunocytochemistry and flow cytometry were applied to determine the phenotype of microglia; CD 68 (M1), CD 206 (M2). The expression of mRNA of pro‐ and anti‐inflammatory cytokines were examined with quantitative real‐time rt‐PCR. Additionally, protein contents, which play a role in cerebral protection, were quantified by immunoblot assays. Data were expressed as mean ± SEM. Two factor ANOVA analysis and Tukey's multiple comparison test was used. P value < 0.05 was considered as a significant value.

Results

EW reduced cell viability and IHT was able to salvage the microglia from EW stress. IHT‐induced protection was mediated by increased HIF‐1α and activated Erk, an anti‐apoptotic protein kinase. The expression of CD 68 was not reduced with IHT vs. EW, but CD 206 was significantly increased in the IHT group vs. non‐IHT groups. An increased pro‐inflammatory phenotype of microglia was observed in the flow cytometry. IHT significantly decreased the expression of TNF‐α and IL‐6, pro‐inflammatory cytokines.

Conclusion

IHC drives the microglia toward anti‐inflammatory phenotype, which could be an important factor in the IHC‐induced cerebral protective mechanism.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Placental Ischemia‐Induced Hypertension Is Abolished by Adrenergic Receptor Blockade
Authors: Spradley Frank T.
Published: June 2021

Hypertensive disorders of pregnancy are the number one cause of pregnancy‐related deaths in the United States. Preeclampsia is a disorder of maternal hypertension and vascular dysfunction presenting in the second half of pregnancy along with fetal growth restriction. Currently, there are no steadfast therapies besides early delivery of the fetus and ischemic placenta, which releases pro‐hypertensive factors into the maternal circulation. The sympathetic nervous system might be a novel target in treating preeclampsia, as sympathetic nervous activity is found to be increased in preeclamptic vs. normal pregnant women. However, it is unknown if the sympathetic nervous system mediates the development of placental ischemia‐induced hypertension. In this study, the hypothesis was tested that adrenergic receptor blockade prevents placental ischemia‐induced hypertension. Wistar Hannover rats were randomized to receive reduced uterine perfusion pressure (RUPP, N=15) or Sham RUPP (N=20) surgeries on gestational day 14 and examined at day 19. In RUPP vs. Sham rats, respectively, mean arterial blood pressure (MAP) was increased (121 ± 3 vs. 102 ± 1 mmHg, P<0.05) without a change in heart rate (421 ± 7 vs. 423 ± 4 bpm). Average fetal weights (1.87 ± 0.04 vs. 2.11 ± 0.03 g) and placental weights (0.50 ± 0.02 vs. 0.57 ± 0.01 g) were lower (P<0.05) in RUPP dams. In RUPP over Sham rats, vasoconstriction induced by the alpha‐adrenergic receptor agonist, phenylephrine, was greater in isolated renal interlobar arteries (Emax: 322 ± 19 vs. 251 ± 12 % of KCl response). The Emax response to KCl was not different between RUPP or Sham rats, in sequence: 100 ± 24 vs. 87 ± 14 % increase in force. A subset of RUPP (N=9) and Sham (N=9) rats received combined terazosin and propranolol (3 mg/kg per day of each, subcutaneous osmotic minipump) to block alpha‐ and beta‐adrenergic receptors, respectively, beginning the day of surgery on gestational day 14. By day 19, adrenergic blockade abolished the development of hypertension in RUPP rats (105 ± 2 mmHg, P<0.05) and reduced their heart rate (343 ± 21 bpm, P<0.05) with a lesser reduction detected in treated Sham rats for MAP (97 ± 2 mmHg, P<0.05) and heart rate (387 ± 9 bpm, P<0.05). Treatment did not alter average fetal weights (1.89 ± 0.05 and 2.11 ± 0.03 g) or placental weights (0.47 ± 0.02 and 0.50 ± 0.03 g) in RUPP and Sham rats, in turn, compared to their untreated counterparts. In conclusion, placental ischemia‐induced hypertension depends on activation of the sympathetic nervous system. The mechanisms responsible for this enhanced sympathetic nerve activity and adrenergic receptor signaling remain unknown but are likely to involve factors released from the ischemic placenta.

Support or Funding Information

4R00HL130577‐02

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Effects of Subconcussive Impacts on Heart Rate Variability In Female Youth Soccer Players
Authors: Bonn Marquise , Harriss Alexandra , Dickey James P.
Published: June 2021

Background

ANS function can be measured noninvasively and relatively inexpensively through heart rate variability (HRV). Reduced HRV is representative of an uncoupling between the ANS and cardiovascular system, which may be proportional to the severity of a neurological injury. Reduced HRV has been observed in patients diagnosed with a sport‐related concussion, and therefore may be a useful outcome measure in identifying the chronic effects of heading during a youth soccer season.

Objectives

The purpose of this study was to evaluate the effects of one season of competitive soccer, with the inherent subconcussive header impacts, on autonomic function in female youth soccer players.

Participants

Twenty‐three female youth soccer players (13.4 ± 0.84 years old) were recruited from three competitive soccer teams.

Methods

The number of headers performed during the season were quantified using game video analysis. Players that performed fewer than the average number of headers (20) at the end of the season were classified into the “low header” group (N=13), while players with 20 or more headers at the end of the season were classified into the “high header” group (N=10). A five‐minute resting electrocardiogram data was collected from all players at baseline and post‐season. These data were analyzed using a two‐way mixed ANOVA in order to calculate heart rate variability time‐domain and frequency‐domain measures, which included mean heart rate, standard deviation of the N‐N interval, total power, low frequency, high frequency, and low to high frequency ratio (LF/HF).

Results

There was a statistically significant interaction between time and group for LF/HF from preseason (6.24 ± 5.77; 2.96 ± 1.81) to postseason (6.79 ± 6.75; 1.90 ± 1.02) in the low and high header groups, respectively (p=.024, partial h2=0.313). There were no other statistically significant changes in HRV for any other time or frequency domain outcome measure.

Conclusion

Our preliminary findings indicate that the LF/HF in the high header group differed from the low header group over the course of a soccer season. This may indicate that an increased number of chronic head impacts might result in an imbalance between sympathetic and parasympathetic activity, which would signify an alteration in autonomic function. These preliminary results demonstrate that additional research is needed in order to further understand the effects of heading on ANS function.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Ectonucleotidases in the Hypothalamus of Salt‐Induced Hypertensive Rats: Transcriptional Profile and functional implications
Authors: Sá Renato Willian Martins , Ribeiro Natália , Ribeiro Luciene Maria , Mendonça Johnny Mendes , Santos Karol Martins , Antunes Vagner Roberto
Published: June 2021

Ectonucleotidases consist of families of nucleotide metabolizing enzymes that hydrolyze extracellular nucleotides, such as ATP, to nucleosides. At the CNS level, these enzymes can affect the excitability of hypothalamic neurons by controlling the extracellular availability of ATP and adenosine. ATP, acting on P2 receptor in the PVN neurons, is involved in hyperosmolality‐induced sympathoexcitation and hypertension in rats. Therefore, we hypothesized that elevated sympathetic activity in salt‐induced hypertensive rats occurs in parallel to a suppression of ectonucleotidases gene expression at the PVN that failure to breakdown ATP. To test this hypothesis, we have used 2 cohorts of male Wistar rats: control (C; tap water; n=5) and salt‐loaded with hypertonic solution (HS; NaCl 2%; n=7) in replacement of the tap water for 7 days. The arterial blood pressure (ABP) was monitored 24‐hour by day during the 7 days via telemetry system. Low frequency (LF) component of systolic ABP (SBP) variability was measured by spectral analysis. PVN mRNA levels for E‐NTPDase1–3 and TMPAP were quantified using real time qPCR. All experimental procedures were approved by the ethical committee for animal research of the ICB‐USP (n° 38/2016). After 7 days of high‐salt intake the ABP of the HS group was higher (129±3 mmHg, p<0.05) compared to the C group (104±3 mmHg). The LF of SBP variability was higher in HS compared to C group [(1.33±0.23 mmHg2 (C) and 4.36±1.22 mmHg2 (LF)]. mRNA level for E‐NTPDase3 was higher in HS than in C group (3,09±0,6 ΔΔCt vs 0,99±0,5 ΔΔCt, respectively), and TMPAP expression was very low in HS group. mRNA levels of the E‐NTPDase1,2 did not differ between the groups. The data indicate that high expression of E‐NTPDase3 mRNA may be correlated to a greater ATP breakdown at PVN level of salt‐induced hypertensive rats.

Support or Funding Information

Financial support: FAPESP #2016/21991‐3 #2016/03359‐8; CAPES

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Ionotropic Glutamate Receptors in the Paraventricular Nucleus (PVN) are Required to Trigger, but not Sustain, Sympathoexcitation by TNF alpha
Authors: MOURAO ALINE A. , SHIMOURA CAROLINE G. , PEDRINO GUSTAVO R. , TONEY GLENN M.
Published: June 2021

In the hypothalamic PVN, pro‐inflammatory cytokines (PIC) have been implicated in driving sympathetic nerve activity (SNA) in support of neurogenic hypertension. In other brain regions, tumor necrosis factor alpha (TNF‐a), a prototypical early onset PIC, has been reported to enhance glutamatergic neurotransmission by increasing membrane trafficking and expression of postsynaptic AMPA receptors leading to synaptic long‐term potentiation (LTP). Here, we tested if TNFa triggers such a response in the PVN to increase SNA. In anesthetized male rats (n=5), unilateral PVN injection (50 nl) of aCSF vehicle had no effect on splanchnic SNA (+2 ± 0.6 %), but rat recombinant TNFa (0.6 pmol) injected 10 minutes later elicited a progressive, ramp increase that averaged 52 ± 2 % (p<0.001, n=5). In separate rats (n=3), injection of TNFa after an Emax dose of the AMPA receptor antagonist NBQX (5.2 nmol) increased SNA by only 11 ± 1 %, which was significantly less than the control response (p<0.001). Because AMPA receptor‐mediated membrane potential depolarization can relieve Mg2+ block of NMDA receptors to potentiate postsynaptic responses to AMPA receptor activation, the effect of NMDA receptor blockade with an Emax dose of AP5 (24 nmol) was tested in another group of rats (n=3). TNFα after AP5 increased SNA by 19 ± 3 %, which was significantly less than the response following vehicle and not different from that following NBQX. Results indicate that AMPA and NMDA receptors in the PVN each participate in the TNFα‐induced increase of splanchnic SNA. Of note, neither NBQX or AP5 injection 30 minutes after TNFα altered the ramp‐like increase of SNA. Mechanisms that sustain progressive sympathoexcitation in response to TNFα in the PVN remain to be determined.

Support or Funding Information

Capes (AAM, GRP), NIH HL088052 & AHA 25710176 (GMT)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Augmented Vascular α1‐Adrenergic Receptor Sensitivity in Humans with Chronic Kidney Disease
Authors: Morison Doree Gardner , Li Yunxiao , Fonkoue Ida , Kankam Melanie , DaCosta Dana , Rapista Derick , Downey Ryan M. , Park Jeanie
Published: June 2021

Patients with chronic kidney disease (CKD) have sympathetic overactivity and difficult‐to‐control hypertension. CKD patients also have exercise‐induced hypertension contributing to poor physical capacity. Our previous studies showed that CKD patients have augmented increases in blood pressure in response to the same degree of sympathetic activation during exercise, suggesting heightened neurovascular transduction of sympathetic nerve activity. We hypothesized that augmented vascular α1‐adrenergic receptor (α1‐AR) sensitivity contributes to the exaggerated vasoconstriction and blood pressure responses to sympathetic activation in CKD. In 28 CKD (Stage III and IV) patients and 11 matched Controls, we measured the degree of vasoconstriction of a dorsal hand vein in response to serial infusions of 10 doses (ranging 15–12,000 ng/min) of the selective α1‐AR agonist phenylephrine (PE) using a linear variable differential transformer with a movable central core to detect changes in vascular diameter. Vascular diameters were expressed as a % reduction from baseline maximum dilation, and plotted against PE dose concentrations in individual semi‐logarithmic dose‐response curves, and analyzed using a 5‐variable sigmoid dose‐response model. The natural log (ln) PE dose that produced a 50% maximal constriction (ED50) was calculated to reflect sensitivity to PE (i.e. α1‐AR sensitivity). CKD patients had a significantly lower PE ED50 compared to Controls (ln PE dose 5.15±0.30 vs 6.68±0.64 ng/min, p=0.022), demonstrating that CKD patients have heightened vasoconstriction in response to vascular α1 AR activation. CKD patients also had a significantly greater change in systolic blood pressure (SBP) during rhythmic hand grip exercise performed at 20% of maximum voluntary contraction compared to Controls (SBP +20.18±2.7 vs +10.43±1.74 mmHg, p=0.01), suggesting that augmented vascular α1‐AR sensitivity may contribute to increased neurovascular transduction and heightened exercise pressor responses in CKD. There was also a trend towards a positive linear correlation between estimated glomerular filtration rate (eGFR) and PE ED50. Augmented vascular α1‐AR sensitivity may contribute to increased neurovascular transduction and hypertension in CKD patients.

Support or Funding Information

Supported by NIH HL‐098744, VA Merit I01CX001065, American Heart Association 15CSA24340001

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The caffeine effects on rotenone‐induced Parkinson's disease model in vitro and in vivo
Authors: Liu Tzu‐Hsien , Luo Chi‐Lun , Huang Bin , Lu Tzong‐Shi , Fu Yaw‐Syan
Published: June 2021

Introduction and hypothesis

The degeneration and lost on dopaminergic neurons in midbrain may cause Parkinson's disease, and it is one of the most common neurodegenerative disease in human. Rotenone is a natural lipophilic toxic compound that is easy to cross the blood‐brain barrier, and widely used as insecticide and pesticide. Rotenone may impair mitochondrial oxidative phosphorylation and lead to dopaminergic neuron loss. In this study we investigated the protective effects of caffeine on rotenone‐induced Parkinson's disease models in vitro and in vivo.

Methods

In vitro, the dopaminergic cell line, SH‐SY5Y cells, is pretreated with caffeine (0~100 mM) for 30 mins and followed by treatment with rotenone (0~10 mM) for 1 hr~24 hr respectively. The cell survival rate, proliferation and morphology were analysis by flow cytometry, ELISA, Cell Counting Kit‐8(CCK‐8) and histology studies. Cell apoptosis/necrosis were analyzed by Annexin‐V/PI staining using flow cytometry. In vivo, the SD rats pretreated with caffeine (10 mg/kg daily, P.O.) or saline and followed treated with rotenone (1 mg/kg daily for 1 week, S.C.). The expressions of apoptosis and autophagy related proteins were analyzed by western blotting. The locomotion of rats were carried out by open field tests.

Results

Caffeine treatments (0–100 mM) had rarely toxic effect on SH‐SY5Y cell lines but could promote autophagy. The SH‐SY5Y cells treated with 1 nM rotenone for 8 hr may decrease the mitochondrial activity of around 60%. Pretreated with caffeine (>2 mM) for 30 min and followed treated with 10 nM rotenone for 8 hr could significantly increase total cell numbers and decrease the cell apoptotic ratio, but no effect on the necrosis ratio and did not restore the mitochondrial activity on rotenone‐induced dopaminergic cell toxicity. The locomotion of SD rats that pretreated with caffeine was better than the saline control in rotenone‐induced Parkinson's disease model. After exposure to rotenone, the situation of recovery by bradykinesia in daily caffeine‐treated group is more obviously than saline control.

Conclusions

In rotenone‐induced Parkinson's disease model, caffeine could facilitate autophagy, increase cell viability and animal locomotion. Our results indicated that caffeine has the potential protective effects on rotenone‐induced Parkinson's disease model.

Support or Funding Information

KMU‐TP105E37

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Selective degradation of BET proteins with dBET1 attenuates lipopolysaccharide‐induced pro‐inflammatory response in microglia
Authors: Castro‐Rivera Carolina Isabelle , Yang Changjun Castro , DeMars Kelly M. , Candelario‐Jalil Eduardo Castro
Published: June 2021

Stroke is the fifth leading cause of death in the United States and its most common type, ischemic stroke, is caused when a major blood vessel that carries oxygen and nutrients to the brain is blocked. This blockage initiates tissue death downstream resulting in inflammation and oxidative stress. Several studies have shown that microglia/macrophage neuroinflammatory responses significantly contribute to neuronal death in ischemic stroke. Bromodomain and extraterminal (BET) proteins‐ Brd2, Brd3, Brd4, and Brdt‐ bind to acetylated lysines on histones and act as a scaffold for transcription factors of several pro‐inflammatory genes important for the spread of cell death after stroke. To recreate the inflammatory process that occurs in response to ischemic stroke in vitro we subjected mouse SIM‐A9 microglia to lipopolysaccharide (LPS) stimulation and then treated with dBET1, a proteolysis‐targeting chimera (PROTAC) molecule that produces a targeted degradation of BET proteins. dBET1 is also a hybrid of the highly selective BET inhibitor, JQ1, that blocks the acetylated lysine‐binding pocket of BET proteins, complexed to a ligand for an E3 ubiquitin ligase. The PROTAC approach results in ubiquitination and subsequent degradation of the target protein by the proteasome. In this study, we did a dose‐response and a time course curve and found that 24 hours of treatment with 1 μM of dBET1 optimally decreases Brd2 and Brd4. Using Western blot and qRT‐PCR, we also found that treatment with dBET1 produced a dramatic reduction in the levels of inducible nitric oxide synthase (iNOS), cyclooxygenase‐2 (COX‐2), interleukin (IL)‐1β, tumor necrosis factor‐α (TNFα), IL‐6, chemokine ligand 2 (CCL‐2), and matrix metalloproteinase 9 (MMP‐9). This is the first study showing that dBET1‐mediated targeted degradation of BET proteins prevents pro‐inflammatory responses in LPS‐stimulated microglia. The data collected in vitro show promising results that suggest that in vivo studies using dBET1 may reduce the deleterious effects of stroke.

Support or Funding Information

This project was possible through the funding of the Neuroscience Summer Internship Program from the McKnight Brain Institute at the University of Florida. It also received funding through the University of Puerto Rico at Mayagüez's MARC U*STAR Program and the National Institutes of Health's grant 2T34GM008419‐26.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Differential subregional hippocampal expression patterns of microRNA‐338 and downstream target INSM1 following global cerebral ischemia.
Authors: Griffiths Brian B. , Ouyang Yi‐Bing , Giffard Rona , Stary Creed
Published: June 2021

Cognitive dysfunction remains a devastating outcome in survivors of cardiac arrest. The principle source of cognitive impairment is attributed to delayed death of hippocampal cornu ammonis 1 (CA1) neurons following blood loss to the brain. Treatments that directly target CA1 neurons have not had translational clinical success. Astrocytes regulate the survival of CA1 neurons following injury, and treatments targeting astrocytes have been effective in protection from global cerebral ischemia. MicroRNAs (miRs) are non‐coding RNAs that regulate gene expression at the post‐transcriptional level, and modulation of miRs have been shown to augment survivability of hippocampal astrocytes. We identified miR‐338, a known modulator of astrocyte function, as a potential regulator of the transcriptional repressor INSM1, a critical epigenetic regulator of cell fate. To test the hypothesis that miR‐338 regulates astrocyte‐mediated neuronal survival following global cerebral ischemia by targeting INSM1, we assessed hippocampal subregional expression patterns of INSM1 and miR‐338 following global cerebral ischemia. We observed that prior to injury, INSM1 is highly expressed in the ischemia‐resistant dentate gyrus relative to the ischemia‐sensitive CA1. Following global cerebral ischemia, expression of INSM1 in the CA1 progressively increased, equaling DG expression after 7 days. We confirmed that increased expression in the CA1 was localized to astrocytes. Immediate future experiments will assess CA1 survivability and cognitive behavioral outcomes following miR‐338 inhibition in vivo.

Support or Funding Information

Rona Giffard NIH #5R01NS053898, Creed Stary AHA #14FTF19970029

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
MicroRNA‐181a mediates neuronal differentiation and modulates microtubule stability.
Authors: Rao Anand Nandakumar , Stary Creed
Published: June 2021

Stroke remains among the top causes of death and is the leading cause of persisting neurological disability in the United States. Treatment options remain limited to early reperfusion with thrombolysis or clot retrieval, and therapies capable of repairing damage resulting from ischemic insult are needed. Microtubules (MTs) have been recently recognized as a potential target for post‐injury therapy in the central nervous system. MTs consist of a stable and labile domain, which differ in their resistance to depolymerization, and by modulating intrinsic mechanisms that work to regulate the proportions of stable and labile MT mass in the axon, we can restore MT integrity after injury. MicroRNAs (miRNAs) are ~22‐nucleotide RNAs that regulate gene expression. MicroRNA‐181a (miR‐181a) is enriched in astrocytes, critical regulators of neuronal homeostasis and neurogenesis. Mir‐181a increases in the brain after stroke, and inhibition of miR‐181a improves functional outcomes in animal models. Using primary neuronal stem cell cultures, we demonstrate here that miR‐181a regulates neuronal differentiation. In situ analysis identified miR‐181a as a likely regulator of MT dynamics through the modulation of the MAPT (tau), MAP1B, and FIGN (fidgetin) genes. MAPT (Tau) and MAP1B are microtubule‐associated proteins (MAPs) that have been demonstrated to contribute to MT stability and neurogenesis, while fidgetin is a severing protein thought to target the labile domain of MTs and modulates a cell's ability to migrate and regulates neuronal elaboration. Therefore, we tested the hypothesis that modulation of miR‐181a activity alters MT integrity in primary neuron and astrocyte cultures and N2a cell line cultures. Immunostaining and biochemical analyses revealed injury and miR‐181a dependent changes to MAPs implicated in the modulation of microtubule stability. Overall, these results demonstrate that microRNAs can be a novel and effective target for restoring MT stability after ischemic brain injury, as well as advancing our understanding of the contributions of astrocyte regulation to increase neurogenesis and post‐injury repair after stroke.

Support or Funding Information

American Heart Association Fellow to Faculty Transition award: No. 14FTF19970029

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Intrathecal activation of the lectin pathway in patients with eosinophilic meningitis by Angiostrongylus cantonensis
Authors: González‐Losada Cristobal , Padrón‐González Alexander Ariel , Lumpuy‐Castillo Jairo , Rodriguez‐Pérez Alejandro , Castillo‐González William , Dorta‐Contreras Alberto Juan
Published: June 2021

Eosinophilic meningitis produced by Angiostrongylus cantonensis is an emerging disease in Western hemisphere. MBL and H‐ and M‐ficolins are starters of the lectin pathway.

Objective

To determine if MBL and H‐ and M‐ficolins can be synthesizedin central nervous system in patients suffering from eosinophilic meningitis due to Angiostrongylus cantonensis.

Methods

19 serum and CSF paired samples were obtained from pediatric patients with the diagnosis of eosinophilic meningitis by Angiostrongylus cantonensis. Cerebrospinal fluid was obtained by lumbar puncture and serum from blood was taken by venopuncture. Serum and CSF was stored frozen in aliquots at −80°C until analysis.

The local ethics committee approved this study and all patients' tutors gave informed consent.

CSF and serum albumin were quantified by immunochemical nephelometry with kinetic analysis. MBL and H‐ and M‐ficolins were measured by commercial enzyme‐linked immunosorbent assay (ELISA) kit.

The MBL reibergram was employed in order to determine if MBL can be synthesized inthecentral nervous system or not. The H‐ and M‐ficolins regressions were employed in order to determine if H‐ and M‐ficolins can be synthesized in the central nervous system or not.

Results

8 patients do not have dysfunction of blood/CSF barrier and 4 patients have intratecal synthesis of MBL.

One patient do not have intrathecal synthesis of H‐ficolin (Figure 2A) and all patients have intrathecal synthesis of M‐ficolin (Figure 2B).

Conclusions

Intrathecal activation of the lectin pathway by at least one of the quantified starters in this group of patients with eosinophilic meningitis due to Angiostrongylus cantonensis.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cholera Toxin Decreases SLC26A3 Expression and Function in Intestinal Epithelial Cells
Authors: Anbazhagan Arivarasu Natarajan , Coffing Hayley , Priyamvada Shubha , Kumar Anoop , Alrefai Waddah A. , Saksena Seema , Gill Ravinder K. , Borthakur Alip , Dudeja Pradeep K.
Published: June 2021

Down‐regulated in adenoma (DRA, SLC26A3), the major Cl−/HCO3− exchanger involved in electroneutral NaCl absorption in the mammalian intestine, has emerged as a novel therapeutic target for diarrhea. Diarrhea commonly involves decreased absorption and/or increased secretion of fluid and electrolytes. We and others have shown decreased expression and function of DRA in infectious diarrhea. In this regard, cholera toxin (CTX) secreted by Vibrio cholerae is one of the most potent agents causing infectious diarrhea. Whether DRA function/expression is altered in CTX‐induced diarrhea is not known. Therefore, the current studies were undertaken to examine the effects of CTX on DRA in vitro in intestinal epithelial Caco‐2 cells. DRA function (Cl−/HCO3− exchange activity) was measured as DIDS sensitive 125I− uptake, mRNA and protein levels were assessed by quantitative real‐time PCR and immunoblotting respectively. Our results showed that treatment of Caco‐2 cells with CTX (1, 5 and 10 ng/ml) for 24 h significantly decreased Cl−/HCO3− exchange activity in a dose dependent manner. Concomitant dose‐dependent decrease in DRA mRNA levels was observed in response to CTX [1 ng/ml (46.3 ± 8.8 % p< 0.0001), 5 ng/ml (69.5 ± 3.1 % P < 0.004) and 10 ng/ml (76.5 ± 2.1%, P< 0.02)] compared to untreated control. Similarly, DRA protein levels were also significantly decreased [1 ng/ml (39 ± 15.7 5% p< 0.0001), 5 ng/ml (60.8 ± 6.3 % P < 0.0001) and 10 ng/ml (56.7 ± 12.6%, P< 0.0001)]. In conclusion, our data suggest that CTX‐induced decrease in DRA expression and function could contribute to the pathophysiology of CTX‐induced diarrhea.

Support or Funding Information

(Supported by NIDDK/Department of Veteran Affairs)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Slc4a8 in the Kidney: Expression, Subcellular Localization and Role in Acid Base Homeostasis and Salt Reabsorption
Authors: Xu Jie , Barone Sharon , Zahedi Kamyar , Brooks Marybeth , Soleimani Manoocher
Published: June 2021

Background

The sodium‐dependent bicarbonate transporter Slc4a8 (a.k.a NDCBE) mediates the co‐transport of sodium and bicarbonate in exchange for chloride. It is abundantly detected in the brain, with low expression levels in the kidney. The cell distribution and subcellular localization of Slc4a8 in the kidney and its role in acid/base and electrolyte homeostasis has been the subject of conflicting reports. There are no conclusive localization or functional studies to pinpoint the location and demonstrate the function of Slc4a8 in the kidney.

Methods

Molecular techniques, including in situ hybridization and RT‐PCR, were performed on kidney sections and tagged epitopes were used to examine the membrane targeting of Slc4a8 in polarized kidney cells. Crispr/Cas9 was used to generate and examine Slc4a8 KO mice.

Result

In situ hybridization and Zonal distribution studies showed no expression for Slc4a8 (NDCBE) in the cortex or in cortical collecting ducts (CCD). Slc4a8 was predominantly detected in the outer and inner medullary collecting ducts (OMCD and IMCD), and was targeted to the basolateral membrane of osmotically tolerant MDCK cells when confocal microscopy images were acquired. Slc4a8 KO mice did not show any abnormal salt or bicarbonate wasting under baseline conditions or in response to bicarbonate loading, salt restriction or furosemide‐induced diuresis.

Conclusions

Slc4a8 (NDCBE) is absent in the CCD and is predominantly localized on the basolateral membrane of medullary collecting duct cells. Further, Slc4a8 deletion does not cause any acid base or electrolyte abnormality in pathophysiologic states such as furosemide‐induced salt wasting or salt restriction. We suggest that Slc4a8 is likely involved in intracellular pH and volume regulation in medullary collecting duct cells.

Support or Funding Information

Merit Review 5 I01 BX001000‐06 award from the Department of Veterans Affairs, and funds from the Center on Genetics of Transport and Epithelial Biology at the University of Cincinnati.
A Conserved Pain Syndrome Resulting from the Acute Activation of TrpA1 by Chemotherapy Drugs
Authors: Boiko Nina , Montano Elizabeth , Hargreaves Kenneth M. , Eaton Benjamin A. , Stockand James D.
Published: June 2021

Treatment with anti‐cancer drugs are often compromised by a peripheral pain syndrome referred to as chemotherapy induced peripheral neuropathy (CIPN). Currently there is no treatment for CIPN and the cellular mechanisms underlying the effects of anti‐cancer drugs on sensory neuron function are not well described. Mitochondria dysfunction, axonal degeneration, reduced sensory innervation, and altered gene expression have all been observed in somatosensory neurons from animal models; but it should be noted that these outcomes are the result of prolonged systemic administration of anti‐cancer drugs and do not necessarily represent the early formative events associated with the initiation of CIPN. Here we show that acute exposure to vinca alkaloids and taxanes result in an immediate pain syndrome in both flies and mice that is consistent with the direct activation of peripheral nociceptors. Electrophysiological analyses of polymodal Class IV dendrite arborization (C4da) sensory neurons in Drosophila reveals that vinca alkaloids and taxol specifically activate the transient receptor potential cation channel A1 (TrpA1) resulting in the activation of these sensory neurons. Furthermore, TrpA1 mutant mice and flies have reduced pain in response to vinca alkaloid exposure consistent with the acute activation of TrpA1 by these chemotherapy drugs being sufficient to generate CIPN across phyla. Targeted expression of dTrpA1 in C4da neurons rescues the pain phenotype in dTrpA1 mutants. These findings represent the first demonstration of direct excitation of sensory neurons by CIPN causing chemotherapy drugs, and identify TrpA1 as an important target during the pathogenesis of CIPN.

Support or Funding Information

This work was funded by a CPRIT IIRG grant (RP150408) and a NIH/NCI R21 grant (CA194138) to J. D. Stockand and B. A. Eaton

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
miR429 regulates IL‐17A expression in inflammatory monocytes in Rheumatoid Arthritis
Authors: Alpert Taylor , Rabquer Bradley James
Published: June 2021

Rheumatoid Arthritis (RA) is a chronic, autoimmune disorder that leads to inflammation and degradation of joints. IL17A, a proinflammatory cytokine, has been previously shown to be involved in migratory inflammation pathways in autoimmune disorders like RA. IL17A initiates RA pathogenesis, suggesting that treatments targeted to neutralize IL17A would be effective therapies. Currently, anti‐IL‐17 therapies are among the most efficacious for treating RA. Additionally, studies indicate that monocytes are also causative agents of inflammation in RA. microRNAs (miRNA) have been identified as translational inhibitors in mRNA of eukaryotic cells. The relationship among monocytes, miRNAs and mRNA must be better analyzed in order to depict inflammatory pathways involved in RA. miR429 was chosen in this experiment because previous research in our lab has suggested that it is upregulated in RA monocytes. We hypothesized that the binding of miR429 to the 3′ UTR region of IL17A would down regulate the expression of IL17A and thus inflammation.

Methods and Results

Gene targets involved in monocytes and inflammation were identified for miR429 in the miRNA databases miRanda and mirSVR. miR429 was found to have a relatively high binding score with IL17A. Isolating the 3′ UTR of the IL17A target gene was the first step in our series of experiments. U937 monocytes were grown and cDNA was created from isolated RNA. Primers were then designed using Primer‐BLAST program, and were created based on the hypothetical binding site on the IL17A 3′ UTR. PCR was used to amplify the 3′ UTR and a restriction enzyme digest was performed on the insert DNA and the pMirGlo vector.

Conclusion

Our results suggest that IL17A and miR429 are expressed by monocytes and may interact to modulate inflammation in RA.

Support or Funding Information

Albion College Biology Department and the Foundation for Undergraduate Research, Scholarly, and Creative Activity

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Biliverdin Reductase A (BVRA) Mediates the Gut Microbiome in Alcoholic liver disease
Authors: Hamoud Abdul‐rizaq
Published: June 2021

Alcohol abuse and chronic consumption are a significant medical burden in industrialized countries. Long‐term alcohol intake is associated with changes in the intestinal microbiota that increase intestinal permeability and the release of deleterious metabolites from the gut. Alcoholic liver disease (ALD) starts with the accumulation of fats in hepatocytes that may progress to alcoholic steatohepatitis (ASH), which can further advance to fibrosis, eventually leading to cirrhosis and hepatocellular carcinoma. Plasma bilirubin levels are increased in alcoholic cirrhotic patients, which is reflective of an injurious effect to hepatocytes as a result of liver damage. In the intestine, microflora convert bilirubin to urobilinoids, which are the predominant bile pigments. We have demonstrated that a knockout of biliverdin reductase A (BVRA) in hepatocytes (hsBVRA KO) of mice, which is the enzyme that converts biliverdin to bilirubin, causes steatosis on a high‐fat diet. Recent evidence has shown that bilirubin itself is protective against hepatic lipid accumulation and obesity. We show that BVRA is suppressed in livers of mice fed alcohol. We hypothesized that BVRA is preventive of high alcohol consumption induced steatosis, which protects the liver from damage. Indeed, the hsBVRA KO mice that consume alcohol had exacerbated hepatic steatosis and changes in the gut microbiome. Together, these observations point to an early mechanism whereby high consumption of alcohol may reduce hepatic BVRA and bilirubin excretion to the gut, leading to triglyceride synthesis causing a progression toward ASH. BVRA may be a significant regulator of the gut microbiome. Future studies will investigate BVRA and bilirubin in the regulation of ALD and how they relate to the liver‐gut microbiota signaling axis.

Support or Funding Information

Source of Funding: This research was supported by the National Heart, Lung, and Blood Institute of the National Institutes of Health under Award Number K01HL125445 (T.D.H.).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
High Dietary Salt Intake Drives Sympathetically Mediated Alpha‐1 Adrenoceptor Regulation of NCC to Increase Renal Sodium Retention and Blood Pressure via an OXSR1 Dependent Pathway
Authors: Puleo Franco , Frame Alissa A. , Wainford Richard D.
Published: June 2021

Background

Acute studies have suggested that sympathetic nervous system (SNS) release of norepinephrine (NE) mediates upregulation of the sodium chloride cotransporter (NCC) via synergistic α1 and β adrenergic receptor pathways. Moreover, recent data suggests a key role of oxidative stress responsive 1 (OXSR1), in NCC regulation.

Hypothesis

Norepinephrine (NE) activates an α1 adrenoceptor pathway that is dependent on OXSR1 to mediate increases in NCC activity that drive the development of salt sensitive hypertension (HTN).

Methods

Male Sprague‐Dawley (SD) rats receiving a continuous s.c. saline or NE (600ng/min) infusion were fed a 0.6% (NS) or 4% NaCl (HS) diet for 14 days respectively. Separate groups of NE infused SD rats were given terazosin (10mg/kg/day; s.c.) and a NS or HS diet for 14‐days. An OXSR1 targeted lentivirus was injected in the kidneys of NE infused SD rats prior to a 14 day HS diet. On day 14 basal MAP, NCC activity (peak natriuresis to iv hydrochlorothiazide (HCTZ; 2mg/kg) infusion) and peak MAP change in response to ganglionic blockade (iv hexamethonium; 30mg/kg) were assessed. NCC expression, phosphoNCC, and OXSR1 levels were assayed via immunoblotting (N=4/gp).

Results

SD rats remain normotensive when fed a high salt diet and NCC expression and activity is downregulated. NE infusion however, inhibits the NCC downregulation associated with HS intake and evokes salt sensitive HTN. Use of an α1‐adrenoceptor antagonist in NE infused SD rats abolishes salt sensitive HTN and downregulates NCC activity. Significantly, lentiviral knockdown of OXSR1 (confirmed by immunoblot) also eliminates salt sensitive HTN in a similar fashion. Furthermore, ganglionic blockade did not produce any observable differences in NE infused rats on HS or NS that were treated or not treated with the α1‐adrenoceptor antagonist or lentivirus.

Conclusion

Our data reveals that SNS release of NE can modulate NCC activity via an α1‐adrenoceptor pathway to prevent dietary evoked salt sensitive HTN. Remarkably, both OXSR1 knockdown and α1‐adreoceptor antagonism reverse salt sensitive HTN. As a theoretical model, the SNS release of NE activates a linear α1‐adreoceptor pathway that stimulates OXSR1 mediated regulation of NCC. The PATHWAY‐2 Trail implicates sodium retention as a driver of resistant HTN suggesting that α1‐adreoceptor antagonism and OXSR1 blockade may represent novel therapeutic targets for HTN.

Support or Funding Information

This work was supported by NIH grants R01HL107330 and K02HL112718 and AHA grants 16MM32090001 and AHA17GRNT3367002 to RDW and NIH grant F31DK116501 to AAF.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Recovery From In Vivo Eccentric Skeletal Muscle Damage: Old versus Young
Authors: Godwin Joshua S. , Hodgman Charles F. , Rice Tyler T. , Zwetsloot Kevin A. , Shanely R. Andrew
Published: June 2021

It has been previously reported that contractile function is depressed in old skeletal muscle, compared to young. However, to our knowledge, the recovery response of old skeletal muscle, compared to young, following in vivo eccentric skeletal muscle damage has not been previously reported. Therefore, the purpose of this study was to compare the recovery response of old skeletal muscle to young skeletal muscle following 7‐days of recovery from in vivo eccentric muscle damage. Old male mice (27.3 ± 0.2 mo) and young male mice (5.6 ± 0.1 mo) were anesthetized with isoflurane. In vivo isometric contractions (Aurora Scientific, 1300A) were performed to obtain optimal electrode placement to stimulate the anterior crural muscle group. Following electrode placement, the pre‐injury torque‐frequency relationship was established. Immediately following the pre‐injury torque‐frequency relationship, 150 eccentric contractions were elicited to induce injury. A second torque‐frequency relationship was measured immediately post‐injury to confirm damage. After 7‐days of recovery, mice were anesthetized with isoflurane and the in vivo torque‐frequency relationship was measured again. Repeated measures analysis of variance showed a significant time by group interaction (p=0.035), with no difference between groups in the torque‐frequency relationship pre‐injury (p=0.231) and a difference in the torque‐frequency relationship between groups 7‐days post‐injury (p<0.001). Further analysis revealed that the torque‐frequency relationship recovered to a greater degree in the young (p=0.313) than old (p=0.003). Analysis of a submaximal torque, i.e. 40 Hz, indicated young mice recovered 77% of initial isometric torque (p=0.254 compared to pre‐injury), while old mice only recovered 48% of initial isometric torque (p=0.003 compared to pre‐injury). Analysis of maximal tetanic torque, i.e. 250 Hz, indicated that neither young mice nor old mice recovered to pre‐injury torque output, 46% of initial (p<0.001) and 67% of initial (p=0.003), respectively. In conclusion, isometric torque decrements persist in both young and old male mice after 7‐days of recovery from in vivo eccentric muscle damage. Additionally, these data suggest that old mice exhibit a diminished capacity to recover isometric torque following eccentric damage, compared to young mice.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Knockout of Vascular Smooth Muscle Inward‐Rectifier K+ Channels Causes Symptoms of Overactive Bladder in Mice
Authors: Tykocki Nathan R. , Ross Maxwell S. , Kopec‐Belliveau Gaelin , Klinger‐Lawrence Mary Beth , Nelson Mark T. , Herrera Gerry
Published: June 2021

The urinary bladder is highly vascularized, and chronic decreases in blood flow are linked to bladder dysfunction. Interestingly, bladder arterioles do not constrict in response to changes in intraluminal pressure (“myogenic tone”), due to a maintained membrane hyperpolarization regulated by inward‐rectifier K+ (KIR2.1) channels. We developed a tamoxifen‐inducible, smooth muscle‐specific KIR2.1 channel knockout (KIR2.1SMKO) mouse to provide additional support that vascular smooth muscle KIR2.1 channels are responsible for this lack of myogenic tone (Tykocki et al. AJP Renal Physiol, 2017). It is, however, unknown if increased vascular myogenic tone alone can alter bladder function. Unlike vascular smooth muscle, detrusor smooth muscle normally lacks functional KIR2.1 channels. Thus, we tested the hypothesis that knockout of smooth muscle KIR2.1 channels can cause bladder overactivity solely by increasing vascular tone. Voiding behavior was measured using the UroVoid® System: a novel tool for chronic, non‐invasive measurements of bladder function (void volume and frequency) in mice. At 6 weeks of age, KIR2.1SMKO mice underwent either sham surgery or surgical implantation of a 21‐day tamoxifen‐eluting pellet (10 mg/kg). Bladder function and water intake were measured before (at 4 weeks of age), during (at 8 weeks of age) and after (at 12 weeks of age) tamoxifen treatment. Prior to tamoxifen treatment, void frequency and volume did not differ between groups. At 8 and 12 weeks, sham KIR2.1SMKO mice showed an age‐dependent decrease in void frequency that correlated with an increase in void volume; this was indicative of a normal, age‐dependent increase in bladder size and voiding control. When compared to sham mice, KIR2.1SMKO mice receiving tamoxifen had increased void frequency and decreased void volume during tamoxifen treatment (8 weeks), which remained elevated at 12 weeks. This was not due to increased diuresis, as no differences in water intake were noted between groups. This suggests that increased vascular tone leads to symptoms of overactive bladder in mice, and that bladder blood flow plays an integral role in regulating bladder function.

Support or Funding Information

Supported by NIH K01‐DK103840 (NRT) and R37‐DK053832 (MTN).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.