bluuebunny/crossref_metadata_2025 huggingface dataset:
Acidity suppresses T cell function and increases memory T cell development
Authors: Wu Hao , Estrella Veronica , Enriquez‐Navas Pedro , Abrahams Dominique , Ibrahim‐Hashim Arig , Luddy Kimberly , Damaghi Mehdi , RavindranadhanPillai Smitha , Pilon‐Thomas Shari , Swietach Pawel , Gillies Robert
Published: June 2021

Acidity is a common microenvironmental factor in solid tumors. We and others have observed that mild acidosis (pH ~6.6) can inhibit the interferon‐γ (IFN‐γ) production by CD8+ T cells and suppress the T‐cell mediated anti‐tumor cell cytolysis. However, the mechanism by which acidosis inhibits effector fuction of T‐cells is not fully understood. Here we show the inhibition of IFN‐γ production by acidity is titratable and is not transduced through the inhibition of major proton‐sensing transporters, such as TDAG8, OGR1, ASIC‐3, or ADIC‐4, all of which we show are expressed in T cells. Neither pharmacologically blockade nor genetic knockout of these transporters could precent the inhibition of IFN‐γ or IL‐2 production by acidosis. Acidosis also inhibited the secretion of other important cytokines by T cells. However, secretion of MDC, MIG, and IP‐10, was increased under acidic pH, indicating that acidosis is not working through a general inhibition of secretion. Acid treated T cells showed the same kinetics of cytokine production compared with non‐treated cells when re‐stimulated under normal physiological pH. These together suggest the inhibition is not due to general failure of cell activity. Furthermore, we found that T cell can be successfully activated under low pH, evidenced by cell surface marker profile change, but more cells were directed to a memory T cell phenotype, suggesting that the acidity directs T cells to differentiate to memory status. Mechanically, extracellular acidification potentaly inhibited the monocarboxylated transporter, MCT1, blocking lactate and H+ efflux leading to an inhibition of glycolysis, which is required for effector T cell function.

Support or Funding Information

This work is supported by Team Science project of Moffitt Cancer Center

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hypoxia‐Induced Inactivation of 26S Proteasome Increases Immunogenicity of Allogeneic Mesenchymal Stem Cells
Authors: Dhingra Sanjiv , Abu‐El‐Rub Ejlal , Sequiera Glen Lester , Sareen Niketa , Yan Weiang , Rafieerad Alireza
Published: June 2021

Allogeneic (donor derived) mesenchymal stem cells (MSCs) derived from bone marrow are in phase I and II clinical trials for cardiac regeneration. Even though the outcome of allogeneic MSCs based animal studies and initial clinical trials was encouraging, the overall enthusiasm of lately has come down. This is due to poor survival of transplanted cells in the recipient heart. The recent reports on allogeneic MSCs based studies found a switch in the phenotype of transplanted cells from immunoprivileged to immunogenic state that led to rejection of cells by host immune system. In the current study, we discovered a novel mechanism of immune switch in MSCs. Our studies demonstrate that hypoxia/or ischemic environment induces an immune shift in MSCs from immunoprivileged to immunogenic state. The immunoprivilege of MSCs is preserved by absence of major histocompatibility complex class II (MHC‐II) molecules. We found that 26S proteasome‐mediated degradation of MHC‐II prevents its expression on cell surface in MSCs and preserves their immunoprivilege. The exposure to hypoxia leads to dissociation of 19S and 20S subunits, and inactivation of 26S proteasome. This prevented the degradation of MHC‐II, and increased immunogenicity of MSCs. Furthermore, hypoxia‐induced downregulation of a chaperon protein HSP90α is responsible for inactivation of 26S proteasome. Maintaining HSP90α levels in hypoxic MSCs prevented 26S inactivation and preserved the immunoprivilege of MSCs. Therefore, hypoxia‐induced defects in 26S proteasome assembly causes loss of immunoprivilege of allogeneic MSCs. Maintaining 26S proteasome activity in MSCs preserves immunoprivilege and prevents rejection of allogeneic stem cells in the heart.

Support or Funding Information

Canadian Institute of Health Research

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effects of Polystyrene Nanoplastics on Goldfish Olfactory Function
Authors: Stoeber Hayden , Huertas Mar
Published: June 2021

Increased plastic pollution has begun to affect aquatic environments from oceans to freshwater streams with little signs of it slowing down since plastics are a cheap and durable resource to use for manufacturing. Due to increased plastic material waste in natural aquatic ecosystems, fish are threatened by the natural break down of these plastics into sizes and molecules that can be easily ingested and absorbed by tissue. When plastics are broken down and are present in the water, the absorption of these molecules becomes less regulated by tissue. Most of the studies regarding plastic effects at this point in time on aquatic organisms are focused on the transfer of plastic particles through generation. We hypothesized that polystyrene nanoplastic (PS NP) particles on Carassius auratus are incorporated through the nose to the brain, dampening olfaction and compromising several physiological responses associated with the sense of smell. We analyzed the physiological effect PS NPs have on the brain and olfactory system of C. auratus. After the fish were exposed to Nanocs 20 nm Fluorescein isothiocyanate (FITC) labeled PS NPs in water for 24 hours in a dark aquarium, or injected directly in the nose, several organs were dissected. The brain, nose, intestines, kidney, liver, and gills were all snap frozen, mounted in OCT and immediately cut at 20 μm on a cryotome before organs were viewed with fluorescent microscopy. PS NP fluorescence was observed mainly in the nose, brain and intestines while the other organs are still in the process of being analyzed. Accumulation of nanoparticles in the brain was stronger in hypothalamic and olfactory bulb areas. Behavioral and electrophysiology studies of olfactory function after nanoplastic exposure are under development to more closely look at the direct possible effects that nanoplastic materials in the brain might have on behavior. The observed fluorescence supported the existence and ability of PS NPs to infiltrate nervous tissues in C. auratus and points to a possible interference with the sense of smell.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Interleukin‐22, a Pro‐inflammatory Cytokine, is Increased in the Serum of Canines with Inflammatory Bowel Disorder
Authors: Panickar Kiran S , Hollar Regina L
Published: June 2021

Inflammatory Bowel Disease (IBD) is characterized by a chronic and/or recurrent inflammatory condition associated with adverse gastrointestinal signs including diarrhea, vomiting, and weight loss. Inflammation plays a key role in the initiation and continual progression of IBD. Interleukin 22 (IL‐22) is a pro‐inflammatory cytokine that is expressed by various cells of the immune system including dendritic cells, natural killer cells, and T cells. IL‐22 is a cytokine that can have either a helpful or a deleterious role in the intestine. The protective role of IL‐22 includes promoting mucosal barrier integrity and epithelial cell regeneration. Its deleterious actions may depend on the presence of other cytokines as well including IL‐23 and IL‐22 binding proteins. Regardless, a continually increased level in IL‐22 can increase associated gastrointestinal inflammatory processes. We tested the circulating levels of IL‐22 in the serum of dogs that were clinically diagnosed as having IBD following an endoscopy conducted by a veterinarian. Blood was collected from dogs with IBD (n=6; 2.2–13.1 yr) and control dogs (n=5; 2.2–11.3 yr) which did not show any signs of IBD. Blood was collected at two different time points (~1 year apart) from the same dogs. Using an ELISA technique, we found that the mean levels of serum IL‐22 from both time points were significantly higher in IBD (286.5 ± 69.32 pg/ml) when compared to controls (104.2 ± 17.8; p<0.05). The levels of IL‐22 in IBD dogs were higher than controls at both individual time points assessed as well. Our results identify IL‐22 as a promising biomarker of inflammatory bowel disorder in dogs and strategies to decrease a continual and elevated level of IL‐22 may be an effective way to reduce the adverse conditions associated with IBD. Further, an optimal anti‐inflammatory nutritional therapy may be a promising option to pursue as a therapeutic measure for IBD in canines

Support or Funding Information

Funded by Hills PNC

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Effects of Acute Hyperglycemia on Cardiac Autonomic Balance Between Sexes
Authors: Skalk Alexandra , Saran Gary , Rampuri Zahrah , Soriano Jan Elaine , Lorida Jeena , Joki Tysen , Hewitt Sarah , Day Trevor A
Published: June 2021

Males and females have differential cardiac autonomic functioning, possibly contributing to high rates of cardiovascular disease in men with aging. In addition, there is still a general lack of understanding regarding normal physiology and disease risk in women. In young, healthy populations, a sex difference in cardiovascular autonomic function has not been established. Acute glucose ingestion has been shown to stimulate a cardiovascular sympathetic response, likely through carotid body activation. We hypothesized that young men would have a greater cardiovascular sympathetic response following glucose loading than women, measured through heart rate variability (HRV) metrics. We aimed to test the effects of acute relative hyperglycemia on HRV metrics to study cardiovascular autonomic function between young, healthy men and women. We recruited 40 healthy participants (21.6±1.7 yrs; BMI 24.1±4.2 kg/m2; 19 females), and instrumented them with electrocardiogram (ECG) to measure heart rate (HR) and finger photoplethysmography to measure mean arterial pressure (MAP). Blood glucose was tested using a glucometer and capillary draw via sterile lancets. Cardiovascular measures were recorded at rest in a fasted state and 30‐min after an acute relative hyperglycemic load (75g glucose, 300ml; 4.8±0.4 vs. 7.5±1.3 mmol/L; P<0.0001). HRV was quantified in two ways: frequency domain, with low frequency (LF; sympathetic activity), and high frequency (HF; parasympathetic activity), and with Poincaré plots, deriving SD1 (parasympathetic activity) and SD2 (sympathetic activity). Frequency domain metrics were minimally affected by glucose loading, increasing HF (P=0.03) and decreasing LF (P=0.02). In both fasted and glucose loaded states, LF and HF domains were different between males and females, with males having increased LF values (P<0.001) and females having increased HF domain (P<0.001). SD1 was also significantly increased in females in both fasted and glucose loaded sates (P=0.013). Our data demonstrates a sex difference in cardiovascular autonomic functioning using HRV metrics. Specifically, we observed increased resting sympathetic activity in males, possibly contributing to high rates of cardiovascular disease later in life. Females had increased resting parasympathetic activity, possibly offering clinical relevance in understanding reduced cardiovascular disease risk in women over the lifespan.

Support or Funding Information

NSERC Discovery and MRU Faculty of Science and Technology

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effect of Beta‐hydroxybutyrate on Myoblast Proliferation and Differentiation
Authors: Melanson Joshua R. , Olsen Zachary E , Walton Chase M. , Bikman Benjamin T. , Thomson David M.
Published: June 2021

Ketogenic diets (KDs) and fasting have been promoted for weight loss and health. KDs cause an increase in circulating levels of ketone bodies, including beta‐hydroxybutyrate (βHB). The effects of amplified levels of βHB on skeletal muscle proliferation and differentiation are largely unknown. Therefore, we tested whether βHB has an effect on proliferation and differentiation of C2C12 myoblast cells in vitro. Experiment 1: to assess proliferation, myoblasts were plated sparsely and incubated in growth media for 6 hours. They were then incubated overnight (~12 hours) with 0 – 10 mM bHB followed by the addition of BrDU to the media for an additional 2 hours to label proliferating cells. The cells were fixed and stained for BrDU and DAPI and proliferation index was calculated as the percentage of BrDU+ nuclei/total nuclei. We found that myoblast proliferation did not increase significantly with βHB. Experiment 2: to assess differentiation, myoblasts were grown to 60% confluence in growth media, followed by differentiation media for 4 days with 0–10 mM bHB. The myotubes were then fixed and stained with DAPI and antibodies against myosin heavy chain. Differentiation index was calculated as the percentage of nuclei associated with MHC+ myotubes. We found that myoblast differentiation increased 22% with 0.6125 mM βHB, and tended to increase (p = 0.12) at 1.25 mM bHB. This suggests that moderate levels of bHB (similar to those achieved in those on KDs or fasting) may be beneficial to muscle repair and/or regeneration.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
What Is Known About Users Of Medical Cannabis Against Chronic Musculoskeletal Pain? A Scoping Review Of The Literature
Authors: Soliz Daniela Furrer , Dionne Clermont E , Marcotte Martine , Jauvin Nathalie , Bélanger Richard , Ware Mark , Foldes‐Busque Guillaume , Aubin Michèle , Pluye Pierre , Kröger Edeltraut
Published: June 2021

Introduction

Chronic musculoskeletal pain (CMP) is one of the most common causes of chronic pain that leads to reduced physical function. Currently, the pharmacotherapeutic options against CMP are limited and consist mainly in pain management with gabapentinoids or with opioids, which may have major adverse effects. Although the effectiveness of cannabis for medical purposes (or “medical cannabis”, MC) in the treatment of chronic pain still lacks evidence, a number of patients suffering from CMP are exploring this new therapeutic option. Knowledge regarding MC users suffering from CMP is quite limited but necessary to guide prescribers and policy makers considering this treatment as well as researchers planning rigorous randomized clinical trials on the effectiveness of MC.

Methods

We conducted a scoping literature review to describe the views and perceptions of patients who had consumed MC to relieve CMP, including demographic characteristics, patterns of MC use and perceived positive, negative or adverse effects.

Results

Forty‐six studies were included in the present scoping review: 21 were conducted in the US, nine in Canada, ten in Europe, two in Australia, one in Israel, and three included data from several countries (up to 43) in Europe and North America. Our review shows that currently typical MC users are mainly young or middle‐aged men (mean age: 28–61), and the preferred form of use is smoking. A majority of the participants of included studies reported that MC use was helpful in reducing chronic pain with only minor adverse effects; in addition, they reported improved psychological well‐being.

Conclusions

Although the information described in the included studies is still exploratory, MC use seems, from the perspective of persistent MC users suffering from chronic pain, to produce more benefits than harms.

Support or Funding Information

This study was supported by the Canadian Institutes of Health Research (CIHR).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Protective Role of the Somatostatin Receptor Subtype 4 in the Indomethacin‐Induced Gastrointestinal Mucosal Injury Model
Authors: Csekő Kata , Kormos Viktória , Horváth Ádám , Sudalina Maria , Iarushkina Natalia , Zelena Dóra , Filaretova Ludmila , Helyes Zsuzsanna
Published: June 2021

Since gastrointestinal ulceration is still associated with high mortality despite advancements in the endoscopic and pharmacological management, precise understanding of the mucosal defensive and protective mechanisms is important. The neuropeptide somatostatin is present in the endocrine cells and sensory nerves, is analogues were shown to reduce mucosal blood flow, pepsin and gastric acid secretion. Its endocrine functions are mediated by the receptors sst2,3,5, and our group discovered that the anti‐inflammatory and analgesic actions are exerted via the sst4 receptor. Sst4 agonists are preclinically tested as novel drug candidates in theses areas. Therefore, we investigated the role of sst4 in gastrointestinal injury.

Sst4 gene‐deleted (sst4 −/−) mice and wildtype counterparts (sst4 +/+) were treated s.c. with 35 mg/kg indomethacine (IDM) or its vehicle. Macroscopic and microscopic mucosal lesions in the stomach and small intestine were evaluated by the ImageJ program and semiquantitative histopathological scoring after 4 and 48 h, respectively. Small intestinal length, thymus, adrenal glands, spleen weight and plasma glucose levels were measured. Neutrophil myeloperoxidase (MPO) activity and plasma extravasation were measured by bioluminescent/fluorescent ex vivo imaging after 24 h.

Macroscopic lesion area and MPO activity in the gastric mucosa were significantly greater in IDM‐treated sst4−/− mice than in wildtypes 4 h and 24 h after IDM administration, respectively. Similar tendency was observed in the duodenum and jejunum even after 48 h. However, IDM did not induce mucosal plasma extravasation in either group at the 24 h timepoint. Thymus weight was significantly less in sst4−/− mice than in sst4+/+ ones. Histopathological scores, small intestinal length, adrenal and spleen weight, and plasma glucose level were not altered by the sst4 deletion.

These results provide evidence for a protective role of the sst4 receptor against chemical injury‐induced gastrointestinal mucosal damage. This shows the safety of sst4 agonist drug candidates in the gastrointestinal tract, and even highlights their potential use for gastroprotective indication.

Support or Funding Information

GINOP‐2.3.2‐15‐2016‐00050 – PEPSYS, GINOP‐2.3.2 STAY ALIVE, EFOP‐3.6.2‐16‐2017‐00008, ÚNKP‐18‐3 New National Excellence Program of the Ministry of Human Capacities

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Allosteric and orthosteric activation of mGlu2 receptors to alleviate dyskinesia and psychosis in the parkinsonian marmoset
Authors: Hamadjida Adjia , Nuara Stephen , Gourdon Jim , Huot Philippe
Published: June 2021

Objective

To determine the effect of combined allosteric and orthosteric activation of metabotropic glutamate 2 (mGlu2) receptors on L‐3,4‐dihydroxyphenylalanine (L‐DOPA)‐induced psychosis‐like behaviours (PLBs) and dyskinesia in the 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐lesioned marmoset model of Parkinson's disease (PD).

Background

We have previously demonstrated that activation of mGlu2 receptors via positive allosteric modulation with LY‐487,379 or orthosteric stimulation with LY‐354,740 alleviates PLBs and dyskinesia in experimental parkinsonism. Here, we seek to determine if synergy ensues when mGlu2 positive allosteric modulation and orthosteric stimulation are combined, in the MPTP‐lesioned marmoset.

Methods

Six common marmosets were rendered parkinsonian by MPTP injection. Following repeated administration of L‐DOPA/benserazide (L‐DOPA) to elicit stable PLBs and dyskinesia, they were administered acute challenges of LY‐487,379 (1 mg/kg), LY‐354,740 (1 mg/kg), LY‐487,379/LY‐354,740 (each 1 mg/kg) or vehicle, in combination with L‐DOPA, after which the severity of each of PLBs, dyskinesia and parkinsonian disability was rated.

Results

LY‐487,379, LY‐354,740 and LY‐487,379/LY‐354,740 each significantly reduced the severity of the global dyskinesia score, by ≈ 42%, 55% and 64% (each P < 0.001), when compared to L‐DOPA/vehicle. The combination LY‐487,379/LY‐354,740 was significantly more effective than either treatment alone (both P < 0.05). The severity of the global PLB score was also significantly reduced by each of LY‐487,379, LY‐354,740 and LY‐487,379/LY‐354,740, by ≈ 51%, 44% and 56% (each P < 0.001), when compared to L‐DOPA/vehicle. The combination LY‐487,379/LY‐354,740 was significantly more effective than LY‐487,379 (P < 0.01), but not LY‐354,740 (P > 0.05). The benefits on dyskinesia and PLBs were achieved without compromising the therapeutic effect of L‐DOPA on parkinsonism.

Conclusions

Our results confirm mGlu2 activation, via both positive allosteric modulation and orthosteric stimulation, is a promising strategy to reduce dyskinesia and psychosis in PD. Moreover, they suggest that the combination of a positive allosteric modulator and an orthosteric agonist may lead to a synergistic effect, thereby providing greater anti‐dyskinetic and anti‐psychotic effects.

Support or Funding Information

Fonds de Recherche Québec ‐ Santé, Natural Sciences and Engineering Research Council of Canada, Parkinson Canada, Weston Brain Institute, Michael J Fox Foundation for Parkinson's Research, Healthy Brains for Healthy Lives.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Selective Sensitization of Transduction pathways in Carotid Body Glomus Cells of Spontaneously Hypertensive Rats (SHR)
Authors: Tan Zhiyong , Lu Yongjun , Whiteis Carol , Chapleau Mark , Abboud Francois
Published: June 2021

We have reported that the non‐voltage gated Acid‐Sensing Ion (Na+) Channels (ASICs) and the Tandem pore‐domain Acid‐Sensitive K+ channels (TASK) are low pH sensors in isolated rat glomus cells causing an initial rapid depolarization, and a sustained (Sust.) depolarization, respectively (Circ. Res. 2007; 101:1009‐19). TASK also mediate glomus cells' responses to hypoxia and to the mitochondrial inhibitor cyanide (Cn) (J Physiol. 2004; 556:175‐91).

In the present study we tested whether hypersensitivity to low pH (6.0), which we had noted in glomus cells of young (4‐6 weeks) prehypertensive SHR (Hypertension, 2008; see table below), would also be evident with Cn‐induced mitochondrial inhibition. Our hypothesis was that the responses would be equally enhanced with Cn since both low pH and Cn inhibit TASK channels.

Using real‐time PCR and western blot analyses we found significant increases in mRNA and protein expression of both ASIC3 and TASK1 in carotid bodies of SHR compared to WKY. However, the enhanced depolarizations (ΔmV) seen at low pH (6.0) in SHR compared to WKY (*P<0.05) were not seen with Cn (2 mM).

We conclude that the non‐voltage gated Na+ (ASICs) and K+ (TASK) channels are significantly more sensitive to low pH than to inhibition of mitochondrial electron transport in SHR. The possibility of alternative transduction pathways for low pH vs. hypoxia or Cn remains to be determined.
Angiotensin II induces upregulation of AT1 receptors via the sequential activation of transcription factors NFkB, Elk‐1 and AP‐1 in Cath.a cells
Authors: Mitra Amit Kumar , Gao Lie , Zucker Irving H
Published: June 2021

It has been clearly established that increased circulating Angiotensin II (Ang II) with concurrent upregulation of brain and peripheral Angiotensin 1‐receptors (AT1R) are important mediators in the pathophysiology of heart failure. In this study we aim to determine the role of sequential activation of transcription factors NFκB, AP‐1 and Elk‐1 in AT1R upregulation. We used Cath.a cells as our neuronal cell model which was treated with Ang II (100nM) over a time course. Western blotting was done for protein expression. Following Ang II activation, there was a temporal increase of the p65 subunit of NFkB. There was a concomitant decrease of IκB and increased IκK expression and AT1R expression. The activation of NFκB was blocked by using the inhibitor Parthenolide leading to a decrease in AT1R expression. The expression of Elk‐1 was upregulated following Ang II activation and was decreased following NFκB inhibition. Gene silencing using p65‐siRNA had similar effects as Parthenolide. The data are presented in the table. Our results suggest a combined role of NFκB, Elk‐1 and AP‐1 in the upregulation AT‐1R in Cath.a cells.

Table showing fold change in protein expression compared to control. *p<0.01 vs. control.
High resolution duplex visualization details development of normal and exaggerated luminal stenosis following endarterectomy
Authors: Brown Aliza , Luo Shaoke , Howard Carol , Eidt John , Moursi Mohammed
Published: June 2021

Reparative surgical procedures in the carotid are limited by intimal hyperplasia. To understand initial patterns of hyperplasia development, we studied circumferential measurements of luminal narrowing, arterial peak systolic velocity (PSV) and end diastolic velocity (EDV) measurements in a rat carotid endarterectomy (CEA).

Three male Sprague Dawley rats (450 g BW) served as a normal hyperplasia development group (CON) and N=4 rats had dietary 4.5 g/kg DL‐homocystine (HCYS) added for exaggerated proliferation. Carotids were visualized in real time and duplexed pre‐operatively and 4, 9 and 14 days post‐CEA (Pre‐OP or D4, D9 and D14). PSV/EDV measurements and lesion development was noted longitudinally and circumferential measurements were taken across the suture line. Volume calculations measured lumenal narrowing and were compared to Pre‐Op.

Post‐Op PSV/EDV measurements were increased above Pre‐Op levels at D4 through D14 at p<0.05. In HCYS both PSV/EDV were higher at D14 vs. CON at p=0.051 and p=0.018, respectively. Along the anastomosis lumenal narrowing was indicated for both CON and HCYS in Pre‐OP vs. Post‐Op beginning at D4 (p=0.0241 and p=0.0450, respectively). At D14, there was more luminal narrowing in HCYS rats vs. CON at p=0.0001.

We observed by duplex that lumenal stenosis is dynamic and changes occur as early as D4 with concomitant influences on arterial pressure. Funded by US Gov, VA Merit Review.
Regulation of RIP140 expression by microRNA
Authors: Lin Ya‐Lun , Tsai Nien‐Pei , Wei Li‐Na
Published: June 2021

Receptor interacting protein 140 (RIP140) is a versatile transcription co‐regulator that plays critical roles in gene regulation. Its expression and regulation are less understood. In this study we examined the expression of RIP140 and identified its 5¡¦ un‐translated region (5¡¦ UTR). We found that its 5¡¦‐UTR could enhance RIP140 translation and be targeted by microRNA‐346 (miR‐346). By manipulating the endogenous miR‐346 level in P19 embryonic carcinoma cells, a functional role for miR‐346 in up‐regulation of RIP140 protein was revealed. This is the first study demonstrating the expression and regulation of endogenous RIP140 protein in animal tissues and cell lines, and revealing the unique regulation of RIP140 expression at the post‐transcriptional level by miR‐346 that targets the 5¡¦ UTR of RIP140 transcript.
Oxidative stress contributes to increased susceptibility to renal injury induced by mild ischemia‐reperfusion in low birth weight
Authors: Ojeda Norma B
Published: June 2021

The risk for renal disease is increased in low birth weight (LBW). Susceptibility to renal injury is increased in a rat model of LBW; however, the mechanism(s) involved is not clear. We hypothesize that oxidative stress contributes to LBW susceptibility to renal injury. To investigate rats were treated with vehicle (UT) or superoxide dismutase mimetic (Tempol, 30mg/k, T) starting 15 min prior to mild ischemia (15 min) and during reperfusion (2 hr). Treatment ameliorated the increase in mean arterial pressure (MAP) and renal vascular resistance (RVR), and the decrease in glomerular filtration rate (GFR), and renal blood flow (RBF), and the increase in renal oxidative stress in LBW (Table). Therefore, this data suggest that oxidative stress may contribute to increased susceptibility to renal injury in LBW.

* P < 0.05 vs. Control

† P < 0.05 vs. UT Counterpart
Pheromones Produced by the Intact Male Mouse causes a Hypertensive Response during Restrained and Free‐Roaming conditions
Authors: Wilson Justin L. , Lee Dexter L.
Published: June 2021

Pheromones regulate aggression in sexually mature male mice and castrated males no longer produce these pheromones. We tested the hypothesis that pheromones from the urine of sexually mature‐intact (SMI) males placed in a restrainer or in cage bedding would elicit a hypertensive response in male mice. Sexually mature male C57BL/6 mice were implanted with a biotelemetry device to measure mean arterial pressure (MAP), heart rate (HR) and locomotor activity. All groups were exposed to 200 μl of urine from SMI males, castrated mice and saline. Urine from SMI mice placed in the cages caused a peak change in MAP (24 + 5 mmHg), HR (151 + 25 bpm) and activity (11 + 3 counts). Urine from castrated mice caused a peak change in MAP (12 + 5 mmHg), HR (93 + 15 bpm), and activity (7 + 1 counts). Saline did not increase MAP, HR or activity in any group. During restraint, urine from SMI mice caused a peak change in MAP (19 + 3 mmHg), HR (159 + 27 bpm). Urine from castrated mice caused a peak change in MAP (9 + 2 mmHg), HR (75 + bpm). Saline elicited a peak change in MAP (4 + 1 mmHg) and HR (20 + 7 bpm). These results indicate that a significant pressor response occurs when male mice are exposed to urine from SMI males. Therefore, the SMI male pheromones excreted in the cage bedding and mouse restrainers are responsible for significant increases in the pressor response.
Paraventricular nucleus (PVN) neurons projecting to the rostral ventrolateral medulla (RVLM) contain both oxytocin and glutamate
Authors: Shi Peng , Jiang Nan , Li Hongwei , Mecca Adam P , Raizada Mohan K , Sumners Colin
Published: June 2021

The PVN is crucial to the CNS regulation of sympathetic outflow. Glutamate is the prevalent excitatory neurotransmitter in the CNS, including the PVN. Oxytocin (OT) is specifically produced by PVN and supraoptic nucleus neurons, and microinjection of OT into the RVLM induces a pressor response. The current study is aimed at determining whether glutamate PVN neurons projecting to the RVLM contain OT as a co‐transmitter. Real time RT‐PCR analyses of PVN tissues from 4‐6 week‐old Sprague‐Dawley (SD) rats revealed the presence of OT and phosphate‐activated glutaminase (PAG; specific marker of glutamate neurons) mRNAs. Microinjection of AAV2‐PAG‐eGFP (200 nL; 1.2 x 1012 gc/mL) bilaterally into PVN of 5 week‐old SD rats produced glutamate neuron‐specific expression of GFP within two weeks. Isolation and ex‐vivo culture of PVN cells from these rats produced GFP labeled neurons, and single‐cell PCR analysis of these revealed that 8 out of 13 (61.53%) were OT‐positive. To identify the phenotype of the GFP expressing glutamate neurons that project to RVLM, fluorescent retrobeads were microinjected bilaterally into the RVLM of SD rats that had received injections of AAV2‐PAG‐eGFP into the PVN. Immunocytochemistry indicated that GFP‐ and retrobead‐labeled PVN neurons contained immunoreactive OT. Thus we conclude that RVLM‐projecting PVN neurons contain both OT and glutamate as major excitatory neurotransmitters.
Influence of salt and estrogen on inner medullary NOS expression in female spontaneously hypertensive rats (SHR)
Authors: Brinson Krystal N. , Sullivan Jennifer C.
Published: June 2021

Estradiol has been suggested to protect against salt‐sensitive hypertension. The renal inner medulla, the final site for Na+ reabsorption regulation, has an important role in blood pressure (BP) control. Nitric oxide (NO) regulates medullary water and Na+ excretion, and estradiol is associated with increased NO production. Thus, the aim of this study was to determine the effect of ovariectomy (OVX, at 11‐weeks) on salt sensitivity and renal medullary NOS protein expression in spontaneously hypertensive rats (SHR.) 14‐week old intact and OVX female SHR were placed on a normal salt (NS; 0.1% salt; N=6) or high salt (HS; 4% salt; N=8) diet for 2 weeks. BP was measured by telemetry. Inner medullary NOS protein expression was determined by Western blot. HS diet did not change BP in intact females (change in BP from baseline: week 1, 0.4±1 mmHg; week 2, 0.3±1 mmHg). HS increased BP in OVX at week 1 (9±2 mmHg), yet this effect was lost by week 2 (2±2 mmHg). NOS1 expression was similar in intact and OVX on NS (relative densitometric units (rdu) 0.5±0.2 vs. 0.5±0.03), however HS tended to increase expression (intact: 0.9±0.08; OVX: 1.7±0.5 rdu). NOS3 expression was significantly decreased by OVX on NS (1.3±0.2 vs. 0.5±0.2 rdu, p=0.4) and this effect was lost with HS (intact: 1.8±0.5; OVX: 1.6±0.3 rdu). We conclude that in female SHR, inner medullary NOS expression is regulated by both estrogen status and salt.
Intra‐Renal Delivery of Progenitor Cells Improves Systemic Inflammation, Oxidative Stress, and Myocardial Microvascular Function in Experimental Renovascular Disease
Authors: Urbieta Victor Hugo , Zhu Xiangyang , Chade Alejandro , Krier James D , Lerman Amir , Lerman Lilach O
Published: June 2021

Renovascular disease (RVD) and hypertension (HT) are cardiovascular risk factors, but if recovery of renal function can reverse cardiac dysfunction remained unclear.

We hypothesized that improved stenotic kidney function would rescue myocardial microvascular (MMV) function in renovascular HT by reducing oxidative stress and inflammation.

Single‐kidney hemodynamics and function and MMV permeability were assessed in pigs in vivo using CT after 6 weeks of RVD. Autologous endothelial progenitor cells (EPC) were then infused intra‐renally, and studies repeated 4 weeks later (RVD+EPC, n=7). Normal and untreated RVD (n=7 each) served as controls.

Blood pressure was similarly increased in RVD and RVD+EPC pigs. Compared to normal, RVD showed elevated systemic TNF‐alpha (142.3± 3.7 vs 43.3 ± 3.5 pg/ml, p<0.001) and oxidized‐LDL, blunted glomerular filtration rate (47.9±10.1 vs 70.8 ± 4.3 ml/min, p<0.05), decreased MMV volume fraction, increased MMV permeability, and myocardial fibrosis.

EPC were rarely observed in myocardium. However, RVD+EPC showed a decrease in TNF‐alpha (90.5± 2.9, p<0.01 vs RVD) and oxidized‐LDL, implying decreased inflammation and oxidative stress. Furthermore, not only renal, but also MMV function were significantly improved in RVD+EPC.

In conclusion, selective improvement in renal function reduces oxidative stress and inflammation and preserves remote MMV function, underscoring functionally important cardiorenal crosstalk.
Vascular Endothelial Growth Factor Improves Renal and Endothelial Function, and Normalizes Blood Pressure in Hypertensive Pregnant Rats.
Authors: Verzwyvelt Joseph D , Gilbert Jeffrey S , Colson Drew , Arany Marietta , Ryan Michael J , Granger Joey P
Published: June 2021

We have recently shown that chronic VEGF administration during the last trimester of pregnancy partially decreases the hypertension associated with placental ischemia. While we have shown that placental ischemia results in endothelial dysfunction, whether VEGF can restore endothelial cell function remains unknown. We hypothesized that restoration of VEGF levels to that seen in normal pregnant rats would further attenuate the hypertension, normalize glomerular filtration rate (GFR), and reverse the endothelial dysfunction associated with placental ischemia. VEGF (180ug/kg/day) was administered for five days via osmotic mini‐pump placed intraperitoneal. Arterial pressure (AP) and tissues were obtained on day 19 of pregnancy from RUPP+VEGF, RUPP and normal pregnant (NP) dams. Endothelial function was assessed by acetylcholine (ACh) induced relaxation of U‐46619 preconstricted carotid artery rings via wire myography. AP (mm Hg) was increased in the RUPP (131±3) compared to the NP (102±1) and infusion of VEGF resolved the hypertension (105±5). GFR (ml/min), decreased in the RUPP dams (1.5±0.3) returned to NP levels (3.1±0.5) by VEGF treatment (3.1±0.4). Relaxation to ACh was inhibited in the RUPP dams compared to the NP but was enhanced by the VEGF treatment (P<0.05). These data demonstrate that VEGF restores GFR and endothelial function, and reduces high blood pressure associated with placental ischemia.
ETA receptor dependent increases in glomerular permeability in experimentally induced diabetic rats
Authors: Saleh Mohamed A. , Pollock Jennifer S. , Savin Virginia J. , Pollock David M.
Published: June 2021

A glomerular permeability defect occurs early in type 1 diabetes and precedes the onset of albuminuria and renal morphological changes. The aim of the current study was to test the hypothesis that the ETA receptor contributes to increased glomerular albumin permeability (Palb) in a model of type 1 diabetes. Male SD rats (250‐275 g) were given streptozotocin (diabetic, D; i.v., 65 mg/kg) or saline (S). Half of each group received the ETA‐selective antagonist, ABT‐627 (ABT), in drinking water at 5 mg/kg/d (n=6 in all groups). Treatment was begun immediately after confirming plasma glucose >300 mg/dl. Six weeks later, glomeruli were isolated by sieving techniques and Palb determined from the change in glomerular volume induced by exposing glomeruli to oncotic gradients of albumin. Palb was increased in D (0.91 ±0.09 vs. S, 0.05 ±0.01, P<0.01); ABT significantly reduced Palb when compared with D (D+ABT, 0.40 ±0.10, P<0.05). ABT had no effect on Palb in S rats. Acute in vitro incubation of glomeruli isolated from D with ABT (10−6 M, 15 min at 37°C), significantly reduced Palb (0.89±0.11 to 0.41±0.09, P<0.05) while selective ETB blockade, A192621 (10−3 to 10−7 M), had no effect. In conclusion, these data support the hypothesis that ETA but not ETB receptors play an important role in maintaining elevated Palb in type 1 diabetes. Mechanisms responsible for additional changes not affected by ABT remain to be determined.
THE PATHOGENESIS OF HEREDITARY HEMOCHROMATOSIS
Authors: Ramos Emilio , Ganz Tomas , Nemeth Elizabeta
Published: June 2021

Hereditary hemochromatosis (HH) is an iron overload disease which can result from mutations in HFE, transferrin receptor 2 (TfR2), hemojuvelin (HJV) or hepcidin, all of which lead to inadequate synthesis of the iron‐regulatory hormone hepcidin. Hepcidin deficiency in HH results in excessive iron absorption. In normal homeostasis, hepcidin production is increased by iron loading. In HH, however, hepcidin is inappropriately low for the degree of iron load suggesting that HFE, TfR2 and HJV may be involved in hepcidin regulation by iron.

We explored hepcidin regulation by iron in three mouse models of HH: TfR2 mutants, HFE ‐/‐ and HJV‐/‐. HFE−/− and TfR2m/m had a dramatically blunted response to 1 d iron feeding. HFE‐/‐ also failed to increase hepcidin even after 21 d on iron diet but TfR2m/m increased hepcidin comparable to wt mice. This suggests there are two types of iron signals regulating hepcidin, one likely involving circulating iron and depending on both HFE and TfR2 (acute signal) and the other involving HFE but not TfR2 (chronic signal). HJV−/− mice, which had extremely low hepcidin mRNA expression, were not able to increase hepcidin after 1, 7 or 21 d of iron diet, indicating that apart from setting hepcidin baseline, HJV may also be necessary for hepcidin regulation by iron. HH can thus result from both decreased hepcidin baseline and the loss of hepcidin response to acute or chronic changes in iron levels.
HuR and AUF1 Competitively Bind to the JunD mRNA and Regulate its Stability in Intestinal Epithelial Cells
Authors: Zou Tongtong , Rao Jaladanki N , Liu Lan , Xiao Lan , Yu Tingxi , Wang Pengyuan , Zhang Xian , Gorospe Myriam , Wang Jian‐Ying
Published: June 2021

JunD is an AP‐1 transcription factor and plays an important role in regulating intestinal epithelial integrity, but the mechanism underlying JunD expression remains unclear. RNA‐binding proteins HuR and AUF1 associate with labile mRNAs bearing AU‐rich elements (AREs) and modulate their turnover and translation. This study determined if HuR and AUF1 regulate JunD expression by interacting with its 3′‐untranslated region (3'UTR).

Methods

Studies were conducted in intestinal crypt cells. RNA‐binding was examined by biotin pull‐down and RNP‐IP assays. JunD mRNA stability was examined by its half‐life and the chimeric Luc‐JunD‐3'UTR reporter assays. HuR and AUF1 functions were tested by their silencing and overexpression.

Results

HuR and AUF bound to JunD mRNA via AREs in its 3'UTR. Studies mapping JunD 3'UTR revealed that HuR specifically bound to the fragments (F) 3 and 5 that contain high ARE levels, whereas AUF1 interacted with the F5 only. HuR competitively inhibited AUF1 binding to JunD mRNA; HuR silencing not only decreased its binding to JunD 3'UTR but also induced AUF1 affinity for JunD mRNA. Increased HuR stabilized JunD mRNA by both increasing its interaction with the JunD 3'UTR and decreasing AUF1 binding to JunD mRNA.

Conclusions

These results indicate that HuR and AUF1 competitively bind to the JunD 3'UTR and that the ratios of HuR and AUF1 bound to JunD mRNA permit dynamic control of JunD mRNA stability.
In vitro effects of lactic acid on bioelectric properties of equine nonglandular squamous mucosa
Authors: Andrews Frank Michael , Buchanan Benjamin R , Elliott Sarah B , Jassim Rafat Al , McGowan Catherine , Saxton Arnold
Published: June 2021

Equine gastric ulcers are common in the nonglandular (NG) region and volatile fatty acids (VFAs), byproducts of carbohydrate fermentation by resident gastric bacteria, have been implicated. Lactic acid (LA), also produced by bacteria, may cause ulcers.

Our objective was to investigate the in vitro effects of LA on NG mucosal bioelectric properties. Equine gastric tissues (n=13) were studied in Ussing chambers to measure short‐circuit current (Isc) and potential difference (PD). Electrical resistance (R) and conductance (G) were calculated after addition of HCl and LA (5, 10, 20, or 40 mM) in normal Ringer's Solution. Tissues were examined for histopathology.

Mucosa exposed to HCl in NRS at pH 1.5 and 4.0 had a significant decrease in Isc and PD; whereas tissues exposed to LA (5, 10, and 20 mM) did not demonstrate significant decreases in Isc and PD. However, mucosa exposed to LA (40 mM), at pH 1.5 demonstrated increased G and decreased R. Histologic changes were consistent with HCl‐induced damage.

HCl induced alterations in bioelectric properties of equine NG mucosa, whereas addition of LA (5, 10, 20 mM) did not. However, 40 mM LA at pH 1.5 increased tissue permeability. Lactic acid, produced by resident gastric bacteria, may increase equine NG mucosal permeability by a different mechanism than VFAs which may be important in gastric ulcer pathogenesis.

Funded by the Comparative Gastroenterology Society.
Increased BNP during Myocardial Injury Decreases Gastric Emptying and Gastrointestinal Absorption
Authors: Addisu Anteneh , Gower William R. , Dietz John R.
Published: June 2021

We have shown that B‐type Natriuretic peptide (BNP) decreases gastric emptying and absorption in mice. We examined whether acute myocardial injury which releases BNP also affects gastric emptying and absorption. Wild‐type (WT) and NPR‐A receptor knockout (KO) mice were anesthetized with isoflurane. A 2 mm incision was made between the 4th and 5th intercostal space and a probe cooled in liquid nitrogen applied directly to the left ventricle to induce cryo‐infarction. In WT mice, gastric emptying (%) one week after MI was 68 ± 6 vs. 89 ± 3 in sham (n=5, P<0.05). Two weeks after MI, emptying was 82 ± 1 vs. 98 ± 1, MI vs. sham (n=5, P<0.05). Absorption, (relative plasma fluorescence) was 350 ± 79 vs. 632 ± 121; (n=6, P< 0.05) MI vs. sham. KO mice showed no significant difference in emptying or absorption. Echocardiography showed left ventricular (LV) ejection fraction of 48 ± 3 vs. 64 ± 2 % (P<0.05), fractional shortening of 24 ± 2 vs. 35 ± 2 % (P<0.01), LV systolic volume (μl) of 51 ± 7 vs. 26 ± 3 (P<0.05), LV posterior wall thickness (mm) of 0.76 ± 0.10 vs. 1.15 ± 0.10 (P<0.05), MI vs. sham respectively. BNP levels (pg/ml) were 4292 ± 276 one week after MI vs. 105 ± 11 in sham (n=5, p<0.05). BNP levels were 1965 ± 755 two weeks after MI, (n=5, p<0.05).

Conclusions

Elevated plasma BNP following acute myocardial injury decreases gastric emptying and absorption in mice. (Support: VA Merit Award to William R. Gower Jr. & NSF/IGERT, Grant to A. Addisu)
Age‐dependent diabetes induces endothelial dysfunction: the role of VEGF and TGF‐beta
Authors: Eatman Danita , Stokley Cyrenah , McKenzie India , Bayorh Mohamed
Published: June 2021

Transforming growth factor (TGF) ‐ beta and vascular endothelial growth factor (VEGF) have been implicated in the endothelial dysfunction and the vasculopathy of diabetes; however, the specific mechanisms involved in the vasculopathic effect of these growth factors have not been clearly elucidated. Therefore, we hypothesized that diabetes induced by streptozotocin (STZ) leads to a reduction in endothelium‐derived relaxing agents, which may be caused by upregulation of VEGF and/or TGF‐beta in an age‐dependent manner. To test the hypothesis, young (4‐5 weeks old) and mature (>7 months old) male Sprague‐Dawley rats were given a single injection of STZ to induce diabetes. Survival rate was age‐dependent [young (100%) vs. old (50%)]. The increase in blood glucose levels had no effect on blood pressure. Plasma nitric oxide and prostacyclin levels were reduced by 20% and 45% respectively in the young diabetic rats. STZ increased plasma thromboxane A2 levels in both groups. Furthermore, STZ amplified VEGF expression in both groups; however, only the mature rats showed a significant change in TGF‐beta expression. Taken together, these findings indicate that diabetes induced endothelial dysfunction/mortality is greater in older rats and due to the over expression of both TGF‐beta and VEGF. This research was funded, in part, by NIDDK grant 1SC1DK082385‐01.
Central Activation of Melanocortin System Modulates Pancreatic Function Independent of Leptin
Authors: Mansour Mahmoud M , Collins Robert , White Doug , Wernette Catherine , Dennis John , Morrison Edward , Tao Ya‐Xiong
Published: June 2021

The hypothalamic melanocortin system (MCs) is part of a leptin regulated cascade that regulates feeding and energy expenditure. Recent observations suggest that hypothalamic MCs neuronal circuits' activation regulates pancreatic secretion of insulin. We hypothesized that hypothalamic infusion with the alpha MSH analog NDP‐MSH (a melanocortin receptor 4 [Mc4r] agonist) would modulate Mc4r expression and insulin levels via a leptin‐independent pathway. We infused the third brain ventricle of lean (with intact leptin signaling) and obese diabetic Zucker rats (with leptin receptor deficiency) with NDP‐MSH to test our hypothesis. NDP‐MSH infusion significantly decreased serum insulin and upregulated Mc4r mRNA message in the pancreas. Immunohistochemistry detected Mc4r expression in pancreatic islet‐like structures that also expressed neuron‐specific beta tubulin. This finding suggests that autonomic neurons are involved in a communication pathway between central MCs and peripheral pancreatic islets.