bluuebunny/crossref_metadata_2025 huggingface dataset:
Female Spontaneous Hypertensive Rats (SHR) Have Better Recovery In Response To Renal Ischemia Reperfusion Injury Than Males
Authors: Mohamed Riyaz , Crislip Ryan G. , Ray Sarah , Wei Qingqing , Sullivan Jennifer C.
Published: June 2021

Hypertension is a known risk factor for acute kidney injury (AKI) but there is little research on the link between the two. Normotensive males are considered to be more susceptible to AKI compared with females. However, how hypersensitive males and females respond to AKI is not well understood. The purpose of the present study is to test the hypothesis that male SHR have worse outcomes than females following renal ischemia reperfusion (IR). 13 week old male and female SHR were subjected to sham‐operated or 25‐minute warm bilateral ischemia followed by 7 days of reperfusion (n=5–6). Renal injury and function were assessed by the following methods: 1) plasma creatinine quantified by Quantichrome creatinine kit, 2) peritubular capillary congestion by Masson's trichrome staining, and 3) tubular damage by hematoxylin and eosin staining of kidney sections and Western blot analysis of kidney injury molecule‐1 (KIM‐1). 24 hours following IR, both male and female SHR showed a significant increase in plasma creatinine compared to respective sham‐operated controls, although the increase in creatinine was greater in males (males: 0.55±0.22 vs. 0.25 ±0.06 mg/dl; females: 0.45±0.06 vs. 0.26±0.06 mg/dl; PIR=0.02; Psex=0.3; Psex*IR=0.03). After 7 days of reperfusion, plasma creatinine remained elevated in male SHR compared to baseline (0.46±0.21 mg/dl vs 0.26±0.065), whereas plasma creatinine in female SHR returned to baseline levels (0.27±0.06 vs 0.26±0.06 mg/dl; PIR=0.03; Psex=0.05; Psex*IR=0.04). Further, histological examination of kidneys following 7 days of reperfusion showed a significant increase in the percentage of peritubular capillary congestion (male: 20±6% increase; female: 0% increase: PIR=0.642; Psex=0.018; Psex*IR=0.002) and damaged tubules (male: 42±13% increase; female: 6±3% increase; PIR=0.0001; Psex=0.0019; Psex*IR=0.0019) compared to sham controls only in males. KIM‐1 protein expression also remained significantly elevated in the outer medulla of both male and female SHR following 7 days of reperfusion, although levels were higher in males compared to female SHR (male: 0.779±0.012 vs 0.073±0.006; female: 0.240±0.147 vs 0.0374±0.038; PIR=0.0002; Psex=0.0092; Psex*IR=0.019). In conclusion, our data support the hypothesis that there is a sex difference in recovery from IR where female SHR recovered faster than male SHR. Further studies are ongoing in SHR to dissect the molecular mechanism(s) driving sex differences in IR outcomes.

Support or Funding Information

This work was supported by National Institute of Health (1P01HL134604‐01 to J.C. Sullivan)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Beneficial effects of Tetrahydrobiopterin on intracellular PO2 and skeletal muscle metabolic demand in patients with COPD
Authors: Layec Gwenael , Decker Stephen , Kwon Oh‐Sung , Thurston Taylor , Le Fur Yann , Trinity Joel , Jeong Eun‐Kee
Published: June 2021

Poor exercise tolerance and peripheral muscle dysfunction are frequent complications associated with chronic obstructive pulmonary disease (COPD). A key peripheral abnormality in these patients is the greater metabolic demand from the skeletal muscle, which occurs in the face of a compromised O2 transport to the limb and therefore exacerbates the development of muscle fatigue. While the etiology of limb muscle dysfunction is still unclear, increasing evidence suggest that chronic oxidative stress and diminished nitric oxide (NO) bioavailablity might play a central role. Therefore, using an integrative approach, the goal of the present study was to determine the effects of acute tetrahydrobiopterin (BH4) supplementation on O2 availability and muscle metabolic demand during exercise in patients with COPD. Five patients with COPD performed dynamic plantar flexion exercise at 40% of maximal work rate with proton/phosphorus magnetic resonance spectroscopy (1H/31P‐MRS), near‐infrared spectroscopy (NIRS), and vascular Doppler ultrasound assessments following oral supplementation of BH4 (10 mg.kg−1) or placebo (PL). BH4 supplementation attenuated the exercise‐induced changes in phosphocreatine (PL: Δ10 ± 4 mM; BH4: Δ 8 ± 4 mM, P<0.05) and pH (PL: 6.89 ± 0.11; BH4: 6.96 ± 0.06, P=0.06). In addition, the ATP synthesis rate was reduced by ~30% (PL: 32 ± 13 mM.min−1; BH4: 22 ± 18 mM.min−1, P=0.11). In contrast, end‐exercise limb blood flow was not significantly different between conditions (PL: 758 ± 255 ml.min−1; BH4: 796 ± 227 ml.min−1, P=0.77), which resulted in a non‐significant, but substantial increase in the intracellular partial O2 pressure (iPO2) measured by 1H‐MRS of myoglobin (PL: 9 ± 4 mmHg; BH4: 14 ± 12 ml.min−1, P=0.3). Together, these preliminary results suggest that NO synthase uncoupling, induced by a deficit in BH4 bioavailability, plays a central role in the development of peripheral dysfunction in O2 transport and muscle metabolism in patients with COPD. From a clinical standpoint, oral BH4 supplementation appears to be a promising therapy to improve the exquisite matching between O2 supply and metabolic demand in the skeletal muscle of patients with COPD, with the prospective that BH4 supplementation may be able to restore functional capacity and improve the quality of life of these patients.

Support or Funding Information

This study was funded, in part by NIH Heart, Lung, and Blood Institute (K99HL125756), the Flight Attendant Medical Research Institute (FAMRI), the Veterans Affairs Rehabilitation Research and Development Service (Merit Awards: E6910‐R, E1697‐R), SPiRe Awards (E1433‐P), Career Development award (IK2RX001215), the American Heart Association (1850039),

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effect of Electrical Stimulation Frequency and Pattern on Muscle Fatigue in Spinal Cord Injured Individuals
Authors: Draghici Adina E. , Qiu Shuang , Picard Glen , Taylor J. Andrew
Published: June 2021

Spinal cord injury (SCI) leads to an extensive skeletal muscle atrophy in the regions below the level of injury. Moreover, the paralyzed skeletal muscles shift towards predominantly Type II (fast‐twitch and highly fatigable) fibers. Functional electrical stimulation (FES) has been used to produce purposeful and controlled muscle contractions to restore functional movements, but rapid muscle fatigue remains a problem. Both pattern and frequency of stimulation can influence muscle fatigue rate, but optimization of stimulation parameters has rarely been investigated for those with SCI. FES has traditionally used constant‐frequency trains (CFT; brief stimulation pulses separated by regular interpulse intervals). However, it has been suggested that doublet‐frequency trains (DFT; close‐spaced pulse trains separated by longer intervals) can result in lesser fatigue. Also, stimulation frequency may determine muscle fatigue, with high‐frequency trains resulting in lesser fatigue compared to low‐frequency trains. However, most studies comparing muscle fatigue across stimulation parameters have been only examined in able‐bodied individuals and it is unknown whether paralyzed muscles respond similarly. There are a few studies suggesting that low‐frequency trains produce less fatigue than high‐frequency trains in those with SCI, in direct contrast to the able‐bodied. Therefore, we examined differences in muscle fatigue across and within able‐bodied individuals (N=8) and those with SCI (N=6) during CFT and DFT stimulation patterns at high‐ (40 Hz) and low‐ (20 Hz) frequencies. Fatigue was defined as a sustained reduction in force of >20% at a constant stimulation intensity. Differences were considered significant at p<0.05 and values are mean ± SEM. For both groups, low‐frequency trains delayed fatigue onset compared to high‐frequency trains, on average by 30 ± 3 %. In the able‐bodied, CFT resulted in slower fatigue onset (#contractions to fatigue CFT vs DFT: 20 Hz 39 ± 4 vs. 34 ± 4, 40 Hz 26 ± 4 vs. 23 ± 3). In those with SCI, DFT resulted in slower fatigue onset (#contractions to fatigue CFT vs DFT: 20 Hz 26 ± 2 vs. 35 ± 4, 40 Hz 20 ± 2 vs. 25 ± 2). In fact, DFT resulted in similar #contractions to fatigue in both populations, whereas CFT induced muscle fatigue significantly faster in those with SCI. These results may indicate that DFT preferentially recruit fast‐twitch fibers, leading to similar fatigue in both able‐bodied and those with SCI. DFT has been previously shown to limit fatigue potentially by increasing calcium release from the sarcoplasmic reticulum and/or by increasing muscle stiffness. These data indicate that DFT trains appear to be preferable for fatigue reduction in the SCI population.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Do sprinters and distance runners have the same autonomic nervous system activity and hemodynamic responses?
Authors: SUN XINRU , Saito Sayaka , Yang Hongxia , Hoshi Junko , Tanno Hiromasa , Kanno Emi , Maruyama Ryoko
Published: June 2021

The autonomic nervous system (ANS) has an important role in physical performance. However, the cardiac ANS activity between sprinters (SPR) and distance runners (DR) has been poorly explored. Not much research has been conducted to determine the differences between sprinters (SPR) and distance runners (DR). We aimed to investigate the influences of ANS activity and circulatory dynamics on different types of exercise training. For this study, five male SPR and seven male DR, aged 18–24 years old (21.2±2.0 years), were recruited from Tohoku University sports club. They had trained using the same exercise type for more than 3 years. We measured weight, height, body mass index, muscle mass, bone mass, visceral fat rating, basal metabolic rate, and part fat percentage and part muscle mass. We also measured blood pressure (BP) and heart rate variability (HRV) at rest and during the postural change from the supine position to the sitting position. Spectral analysis of HRV was performed to evaluate low‐frequency power (LF), high‐frequency power (HF), and the LF/HF ratio to observe the changes in cardiac sympathetic and parasympathetic nerve activity. Data regarding type, duration, and content of the training were obtained using questionnaires after the measurements. Significant differences were observed between both groups with regards to bone mass and left‐hand muscle mass. SPR had significantly higher HRV in supine position compared with DR. The HF of SPR was significant higher than DR in all positions. During postural change from the supine to the sitting position, LF/HF was significantly higher in DR compared with the SPR. Systolic BP in SPR was observed to significantly increase during postural change from the supine position to sitting position, while the systolic BP during postural change was reduced in the DR group. We thus observed different responses in both groups suggesting significant differences in ANS activity and circulatory dynamics between SPR and DR.

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‐intensity Interval Training Enhanced Platelet Mitochondrial Bioenergetics in Patients with Heart Failure
Authors: Wang Jong‐Shyan
Published: June 2021

Mitochondrial functionalities in platelets involve the cellular bioenergetics and thrombogenesis. This study elucidates how high‐intensity interval training (HIIT) affects systemic aerobic capacity and platelet mitochondrial bioenergetics in the patients with heart failure (HF). Thirty‐four HF patients with were randomized to engage HIIT (3‐minute intervals at 40% and 80% VO2peak, n=17) for 30 minutes/day, 3 days/week for 12 weeks, or to a general health care group (GHC, n=17) that did not received exercise intervention. Systemic aerobic capacity (i.e., peak O2 consumption, VO2peak) and platelet mitochondrial O2 consumption rate (OCR) in the HF patients were measured using automatic gas analysis and high‐resolution respirometry, respectively. The experimental results demonstrated that the HIIT group exhibited higher VO2peak and O2 uptake efficiency slope, as well as, lower VE‐VCO2 slope following the 12‐week intervention, compared to those of the GHC group. Moreover, the HIIT, but not the GHC, regimen (i) increased the capacities for the maximal and reserve of OCR, (ii) enhanced the Complex I‐ and II‐mediated OCRs, and (iii) elevated the bioenergetics health index in platelets. Additionally, the VO2max levels were positively correlated with the maximal and reserve capacities of OCR and Complex I‐ and II‐mediated OCRs in platelets. Hence, the HIIT regimen improves aerobic fitness and ventilatory efficiency, which is accompanied by enhanced capacity for platelet mitochondrial bioenergetics in patients with HF.

Support or Funding Information

National Science Council of Taiwan (grant number NSC 106‐2314‐B‐182 ‐048 ‐MY3).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Overexpression of SOD2 in the diaphragm provides partial protection against ventilator‐induced diaphragm atrophy and contractile dysfunction.
Authors: Morton Aaron B. , Smuder Ashley J. , Hyatt Hayden W. , Hinkley J Matthew , Ichinoseki‐Sekine Noriko , Mor Andres , Powers Scott K.
Published: June 2021

Mechanical ventilation (MV) is used clinically to maintain adequate pulmonary ventilation in patients that are unable to on their own. Unfortunately, an undesired side‐effect of prolonged MV is the rapid development of diaphragmatic atrophy and contractile dysfunction, termed collectively as ventilator‐induced diaphragm dysfunction (VIDD). The development of VIDD is clinically important because diaphragmatic weakness is a primary risk factor for difficult weaning from the ventilator. While the mechanism(s) leading to VIDD are not fully understood, increased mitochondrial production of reactive oxygen species (ROS) is a requirement for the development of VIDD. In this regard, superoxide dismutase 2 (SOD2) resides in the mitochondrial matrix and operates as the first line of defense to eliminate superoxide radicals. Therefore, we tested the hypothesis that overexpression of SOD2 in diaphragm fibers will protect against VIDD. Compared to control, viral transfection (rAAV9) of the SOD2 transgene in the diaphragm resulted in a ~70% increase in SOD2 protein abundance in diaphragm fibers. This overexpression of SOD2 in the diaphragm resulted in partial protection against VIDD by diminishing MV‐induced diaphragmatic contractile dysfunction. Nonetheless, overexpression of SOD2 did not protect against MV‐induced diaphragmatic atrophy. These findings demonstrate that increased expression of SOD2 alone in the diaphragm is not sufficient to prevent MV‐induced diaphragm atrophy but does provide partial protection against MV‐induced diaphragmatic contractile dysfunction.

Support or Funding Information

Supported by NIH R01 AR064189 awarded to SKP

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Identification and Characterization of Protein Phosphatase Methylesterase 1 (Ppme1) in Skeletal Muscle
Authors: Labuzan Sydney , Waddell David
Published: June 2021

Skeletal muscle atrophy results from a variety of physiological conditions, including immobilization, denervation, glucocorticoid treatment, cancer, and aging. In order to elucidate a better understanding of the molecular and genetic mechanisms of skeletal muscle atrophy, a microarray study was performed using muscle tissue isolated from mice undergoing neurogenic atrophy in response to denervation. The data generated from this microarray identified a suite of several hundred predicted genes that displayed differential expression under conditions of muscle wasting, including protein phosphatase methylesterase 1 (Ppme1). The Ppme1 gene was observed to be transcriptionally upregulated in response to neurogenic skeletal muscle atrophy (i.e. denervation). To confirm that Ppme1 is expressed in muscle, we successfully cloned the predicted Ppme1 cDNA from mouse muscle cells. Additionally, Ppme1 mRNA expression levels were analyzed by quantitative polymerase chain reaction (qPCR) and found to be highly expressed in both proliferating and differentiated muscle cells. Furthermore, Ppme1 protein levels were analyzed by Western blot using protein homogenates isolated from muscle cells and Ppme1 was found to be ubiquitously and uniformly expressed at all time points throughout muscle cell proliferation and differentiation. In order to assess the transcriptional activity of the Ppme1 gene, a fragment of the promoter was cloned, fused to a reporter gene, transfected into C2C12 cells, and found to be transcriptionally active. Additionally, the Ppme1 reporter gene was transfected in combination with MyoD1 and myogenin expression plasmids and differential effects on the Ppme1 reporter gene activity were observed in response to myogenic regulatory factor ectopic expression. The results presented here help contribute new insight into the molecular and genetic mechanisms of muscle atrophy and help provide a better understanding of the genome‐wide transcriptional changes that occur during neurogenic atrophy‐induced muscle wasting.

Support or Funding Information

The work was support by University of North Florida Transformational Learning Opportunity grants to D.W.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Identification and Characterization of FGGY Carbohydrate Kinase Domain Containing (Fggy) in Skeletal Muscle
Authors: Novo Karla , Waddell David
Published: June 2021

Skeletal muscle atrophy can result from a range of physiological conditions, including denervation, corticosteroid exposure, immobilization and aging. In order to better characterize the molecular genetic events of atrophy, skeletal muscle was isolated from mice following 3 days and 14 days of denervation. The gene expression profile of the denervated muscle tissue was analyzed by microarray and compared to control muscle in order to identify novel atrophy‐induced genes. The microarray revealed that FGGY carbohydrate kinase domain containing (Fggy) is expressed in skeletal muscle and is induced in response to denervation. Bioinformatic analysis of the Fggy gene locus revealed two validated alternative isoforms, which we have termed Fggy‐001 and Fggy‐003, that have distinct transcription initiation sites. In order to confirm that Fggy is expressed in muscle cells, the cDNAs of the two validated alternative splice variants were cloned from mouse myoblast cells. Interestingly, a novel alternative spice variant for each of the validated alternative isoforms was also cloned from mouse muscle cells, suggesting that at least four Fggy variants are expressed in skeletal muscle. In order to better characterize the expression profile of the Fggy gene, quantitative RT‐PCR (RT‐qPCR) was performed using RNA isolated from muscle cells and primer pairs that distinguish the four alternative Fggy transcripts. The RT‐qPCR data reveals that the Fggy‐001 transcripts are more highly expressed in proliferating muscle cells, while the Fggy‐003 transcripts are more evenly expressed in muscle cells that are both proliferating and differentiated. Western Blot analysis of protein homogenates isolated from muscle cells confirmed that the Fggy‐001 isoform is more highly expressed in proliferating muscle cells, while the Fggy‐003 isoform is more ubiquitously expressed. Finally, confocal fluorescent microscopy analysis revealed that the Fggy‐001 full length transcript is localized throughout the cell, while the Fggy‐001 novel alternative transcript only appears to be localized to the cytoplasm. Moreover, the Fggy‐003 novel alternative transcript produces a punctuate localization pattern throughout the cytoplasm of proliferating muscle cells. The characterization of novel genes that are activated during neurogenic atrophy helps improve our understanding of the molecular and cellular events that lead to muscle atrophy and could eventually lead to new therapeutic targets for the treatment of muscle wasting.

Support or Funding Information

The work was support by University of North Florida Transformational Learning Opportunity grants to D.W.

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 role of Nrf2 in exercise training‐mediated regulation of antioxidant enzymes in skeletal muscle
Authors: Okutsu Mitsuharu , Yamada Mami , Iwata Masahiro
Published: June 2021

Oxidative stress has been implicated in the pathogenesis of skeletal muscle wasting. Regular exercise training prevents oxidative stress‐induced muscle wasting by at least partially improving the antioxidant defense system. However, the underlying molecular mechanisms involved remain elusive. In this study, we assessed whether the nuclear factor erythroid 2‐related factor (Nrf2), the master regulator of antioxidant gene transcription, is essential for the exercise training‐mediated increase in expression of antioxidant enzymes. We performed 3 complementary studies. First, C2C12 myotubes were subjected to 110% cyclic uniaxial stretch for 1 and 6 hours at a frequency of 1 Hz to assess whether stretching regulates mRNA expression of Nrf2. Second, C57BL/6 mice were subjected to either voluntary wheel running for 4 weeks or no exercise to examine whether exercise training regulates Nrf2 and antioxidant enzyme protein expression in muscle. Third, muscle‐specific Nrf2 knockout (Nrf2MKO) and wild‐type littermates (WT) mice were assigned to either exercise training or sedentary groups, and the muscle content of antioxidant enzymes was determined after 4 weeks of exercise training. Our results revealed a significant elevation in Nrf2 mRNA expression in C2C12 myotubes at 6 hours of mechanical stretch. Voluntary running in C57BL/6 mice increased antioxidant enzyme protein (i.e., Cu/ZnSOD, EcSOD) in soleus muscle. Moreover, expression of the Nrf2 target antioxidant gene NQO1 in Nrf2MKO skeletal muscle was abolished compared to WT mice. Loss of Nrf2 in skeletal muscle also blunted the exercise training‐mediated increase in antioxidant enzymes in soleus muscles. These findings indicate that Nrf2 is required for exercise training‐mediated increase of antioxidant enzymes in skeletal muscle.

Support or Funding Information

This study was funded by Grant‐in‐Aid for Challenging Exploratory Research (16K13019) and Grant‐in‐Aid for Scientific Research (B) (15H03080) to M. Okutsu.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
p62/SQSTM1 is required for exercise training‐mediated regulation of antioxidant enzymes in skeletal muscle
Authors: Yamada Mami , Iwata Masahiro , Okutsu Mitsuharu
Published: June 2021

Oxidative stress has been implicated in the pathogenesis of skeletal muscle wasting. Regular exercise training prevents oxidative stress‐induced muscle wasting at least partially by improving the antioxidant defense system. It has been shown that p62/sequestosome1 (SQSTM1) competitively binds to Keap1 activating the nuclear factor erythroid 2‐related factor 2 (Nrf2), which stimulates transcription of antioxidant enzymes. However, it remains to be determined if this process is active in skeletal muscle. Therefore, this study aims to determine whether p62 is required for the exercise training‐mediated increase in antioxidant enzymes in skeletal muscle. To do so, we used 3 complementary studies: i) C2C12 myotubes were subjected to 110% cyclic uniaxial stretch for 1 and 6 hours at a frequency of 1 Hz to assess whether stretching regulates p62 mRNA expression; ii) C57BL/6 mice were assigned to either voluntary wheel running for 4 weeks or no exercise to examine whether exercise training regulates p62 protein expression and phosphorylation (Ser351), both of which increase p62 affinity for Keap1 binding; and iii) muscle‐specific p62 knockout (p62MKO) and wild‐type littermates (WT) mice were assigned to either exercise training or sedentary groups, and the muscle content of antioxidant enzymes was determined after 4 weeks of exercise training. Our results demonstrated that p62 mRNA levels were significantly increased at 6 hours of mechanical stretch in C2C12 myotubes. Voluntary running in C57BL/6 mice increased p62 protein and p62 phosphorylation in soleus muscle. Moreover, expression of the Nrf2 target antioxidant gene NQO1 in p62MKO skeletal muscle was abolished compared to WT mice. Loss of p62 in skeletal muscle also blunted the exercise training‐mediated increase in antioxidant enzymes (i.e. Cu/ZnSOD, EcSOD) in soleus muscle. These findings indicate that p62 is required for exercise training‐mediated increase of antioxidant enzymes in skeletal muscle likely via modulation of Nrf2 function.

Support or Funding Information

This study was funded by Grant‐in‐Aid for Challenging Exploratory Research (16K13019) and Grant‐in‐Aid for Scientific Research (B) (15H03080) to M. Okutsu.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Mechanisms of neuroprotection against oxidative stress in the anoxia tolerant turtle Trachemys scripta
Authors: Reiterer Melissa , Milton Sarah L.
Published: June 2021

The detrimental effects of oxidative stress caused by the accumulation of Reactive Oxygen Species (ROS) have been acknowledged as major factors in aging, senescence, and several neurodegenerative conditions including Parkinson's Disease and stroke (ischemia/reperfusion). Mammalian models are extremely susceptible to these stresses and the restoration of oxygen after anoxia causes oxidative damage; however some organisms including the freshwater turtle Trachemys scripta can withstand such conditions without any apparent pathology. T. scripta thus provides us with a model in which we can investigate physiological mechanisms of neuroprotection in anoxia and reoxygenation without the damaging effects that come with oxidative stress.

The ability of the turtle to survive these stressors is thought to be related to its high innate levels of antioxidants and its capacity to suppress ROS production. One potential antioxidant mechanism is the Methionine Sulfoxide Reductase System (Msr) that catalyzes the reduction of oxidized Methionine (Met) residues. Methionine residues are easily oxidized amino acids and oxidized Met is associated with decreased protein activity. Msr may also restore function to damaged proteins and it has been shown to scavenge ROS and thus ameliorate cellular damage.

The present study investigated the role and regulation of MsrA in turtle primary neuronal cell cultures exposed to 4 hours of anoxia and 4 hours of anoxia followed by 4 hours of reoxygenation. Anoxia and anoxia/reoxygenation increase MsrA mRNA and protein levels, which suggests a protective role under these conditions. MsrA expression was then altered with target specific siRNA; we found that the knock down of MsrA expression resulted in decreased cell viability. As the factors that control MsrA expression in this model are still unknown, we also induced FOXO3a expression by transfecting a FOXO3a plasmid into the cells, as it has been shown to regulate MsrA levels in other animal models. The induction of FOXO3a protected neurons against cell death. Furthermore, treatment of cells with the green tea extract Epigallocatechin gallate (EGCG), which has antioxidant properties, results in increased expression of FOXO3a when the cells are exposed to oxidative stress. Together these results suggest that MsrA may play an important role in protection against anoxia and oxidative stress in the turtle brain, and may prove to be a target to treat diseases of ischemia and oxidative stress.

Support or Funding Information

NIH‐NIA Project # 1R15AG033374‐01,

FAU Foundation

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Seal Endothelial Cells: a Comparative Model to Study Natural Tolerance to Ischemia/Reperfusion
Authors: Vazquez‐Medina Jose Pablo , Allen Kaitlin , Hindle Allyson G.
Published: June 2021

Reperfusion injury is a common complication derived from ischemia/reperfusion events that occur in the course of myocardial infarction, stroke, embolism and other peripheral vascular diseases. Lack of blood during ischemic episodes activates cellular pathways that upregulate pro‐inflammatory signaling and increase oxidant generation. Reperfusion after ischemia leads to the recruitment of inflammatory cells further exacerbating oxidant production and ultimately resulting in cell death, tissue injury and organ dysfunction. Seals experience routine, repetitive episodes of peripheral ischemia/reperfusion during and after dives. These ischemia/reperfusion events are a direct consequence of the cardiovascular adjustments that allow seals to maximize the use of their oxygen stores while diving but result in selectively reduced perfusion to peripheral tissues. Remarkably, seals show no apparent detrimental effects associated with these ischemia/reperfusion events. The cellular mechanisms that underlie this natural tolerance to a potentially pathological condition, however, remain unknown. We isolated and cryopreserved primary endothelial cells from Weddell seal placental arteries to study the mechanisms that confer ischemic tolerance to seal cells. Seal endothelial cells were evaluated for dil‐acetylated low‐density lipoprotein (dil‐AcLDL) uptake and immunostained with endothelial markers to confirm the endothelial phenotype of the preparation. Seal cells were also stimulated with phorbol ester to test whether or not they are capable of producing oxidants. Weddell seal endothelial cell monolayers exhibit a cobblestone‐like morphology characteristic of human and murine endothelial cells and stained positive for vascular endothelial cadherin by immunofluorescence. Live seal cells in culture were functionally capable of incorporating labeled dil‐AcLDL and generated hydrogen peroxide in a concentration‐dependent manner when stimulated with phorbol ester. These results suggest that primary seal endothelial cells in culture are a viable, novel and potentially useful model to study cellular mechanisms associated with natural tolerance to ischemia/reperfusion.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Novel Regulators of Epithelial Migration and Differentiation
Authors: Dunnwald Martine
Published: June 2021

Epithelial migration and differentiation are key cellular processes required for wound closure. We recently identified Interferon Regulatory Factor 6 (IRF6) as a transcription factor that regulates both of these processes. We previously demonstrated that Irf6‐deficient embryos and Irf6‐deficient keratinocytes displayed impaired wound healing and cellular migration. Furthermore, individuals with IRF6 mutations (van der Woude syndrome, the most common form of syndromic orofacial cleft) exhibited increased likelihood of wound healing complications following surgical repair. Therefore, we hypothesized that altered expression of Irf6 and its downstream effectors would lead to impaired wound healing in adults. Using murine models with different Irf6 levels, we showed that reduced levels of Irf6 are more detrimental in early stages of wound healing (day 4) and increased levels are more detrimental in late stages of wound healing (day 7). Furthermore, keratinocytes from syndromic orofacial cleft patients were slower at closing an in vitro scratch compared to non‐syndromic keratinocytes. One of the downstream targets of Irf6 is Arhgap29 (Rho GTPase Activating Protein 29). Arhgap29 is decreased in the absence of Irf6, and could mediate the Irf6‐dependent migration. We are currently testing the role of Arhgap29 in vivo using a novel Arhgap29 allele containing a genetic variant identified in a patient with cleft lip and palate. Together, these results demonstrate that the positive and negative perturbation of the levels of Irf6 lead to defects throughout the entire period of wound healing. Ultimately, these data suggest the existence of a critical therapeutic window for IRF6 that should be considered in designing new clinical treatments for wound healing.

Support or Funding Information

NIH/MIAMS AR067739

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Consumption of a Refined Carbohydrate Diet Does Not Impair Vasodilation of Cranial Tibial Arteries in the Mourning Dove, Zenaida macroura
Authors: Clark William , Basile Anthony J. , Deviche Pierre , Sweazea Karen L.
Published: June 2021

Chronic hyperglycemia in mammals correlates positively with the production of reactive oxygen species (ROS) and is associated with impaired vasodilation. Plasma glucose (Pglu) in birds including Mourning Doves (MODO) is generally higher than in mammals, yet vascular ROS in MODO are similar to levels measured in healthy rodents. This difference may be due to abundant endogenous antioxidants such as uric acid and vitamin C in birds. We hypothesized that feeding doves a refined carbohydrate diet would not impair vasodilation. To examine this, we captured MODO (n = 11) in Tempe, AZ, and housed them for five weeks. Birds were acclimated for one week during which they transitioned either to a nutritionally‐balanced dove seed diet (controls, CONT) or to a refined carbohydrate (WB; white sandwich bread) diet. We collected a blood sample from the brachial vein of each bird upon arrival and at the end of the 4‐week treatment for the measurement of serum metabolites. At the end of the study, we euthanized birds and cranial tibial arteries were isolated, cannulated with glass pipettes, and pressurized in a vessel chamber. Isolated arteries were pre‐constricted to 50% of the initial diameter by adding increasing doses of phenylephrine to the superfusate. We then measured vasodilation by adding increasing doses of the endothelium‐dependent vasodilator acetylcholine (10−9 M to 10−5 M). Vasodilation, serum glucose, uric acid, triglycerides, and free glycerol were not significantly different between groups (p>0.05). To examine the effects of captivity, we compared data to those from previous studies on isolated arteries from wild MODO (WILD). Endothelium‐dependent vasodilation was greater in captive than wild doves (CONT vs. WILD, p<0.01; WB vs. WILD, p<0.05). These findings are consistent with our hypothesis as refined carbohydrate consumption did not impair vasodilation. Moreover, captivity improved vasodilation when compared to wild doves regardless of diet type. The findings contrast with those in mammals, in which high carbohydrate consumption results in impaired endothelium‐dependent vasodilation.

Support or Funding Information

This study was funded in part by an IOSP APS summer research fellowship (WC).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hemoglobin In An Ancestral Hibernator, The Common Tenrec, Tenrec ecaudatus
Authors: York Sarah E. , van Breukelen Frank
Published: June 2021

Common tenrecs, Tenrec ecaudatus, are medium‐sized mammalian tropical hibernators endemic to Madagascar. Tenrecs are capable of effectively hibernating at temperatures between 12 and 28°C and do not undergo periodic arousals during hibernation, a feature common to all other known hibernators. Interestingly, these tenrecs can also experience spontaneous bouts of torpor during the active season known as facultative torpor (FT). During hibernation and FT, oxygen consumption rates are 1.1–14% of active rates depending on ambient temperature. We have observed marked variation in hematocrit (between 15 and 50%). We asked what happens to hemoglobin concentration and type across the annual cycle and state. A colorimetric assay was performed for hemoglobin concentration in whole blood. Isoelectric focusing was used to assay for changes in hemoglobin type (e.g. fetal vs adult types) across the physiological states. These data will serve as an initial step in helping to determine how tenrecs are able to achieve low metabolic rates without concomitant decreases in body temperature.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Immune Function In The Basoendothermic Common Tenrec, Tenrec ecaudatus
Authors: Smith Catlene J. , van Breukelen Frank
Published: June 2021

Common tenrecs, Tenrec ecaudatus, are small hibernating mammals endemic to Madagascar and demonstrate extreme variability in body temperature and metabolism. When housed at ambient temperatures of 12–28 °C, tenrecs may hibernate or be active with body temperatures of ~13–34 °C depending on ambient temperature. During hibernation, tenrecs are consistently lethargic and do not exhibit the typical euthermic periods of rewarming that other hibernators experience. In classic hibernators, injection of lipopolysaccharide (LPS) during torpor results in an elevated temperature during the subsequent periodic arousal between torpor bouts. In tenrecs, injection of LPS does not result in increased temperature suggesting no immune recognition of LPS. Complement activity in tenrecs will be determined through the ability to lyse foreign red blood cells to determine the status of immune function in various states and as a function of body temperature. These data will help define the immune status across the annual cycle in a mammal with ancestral features.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effect of Incubation Temperature on Survival, Growth and Oxygen Consumption of Developing Brine shrimp (Artemia franciscana)
Authors: Melendez Christopher L. , Mueller Casey A.
Published: June 2021

Brine shrimp (Artemia franciscana) are commercially harvested aquatic crustaceans that naturally inhabit inland and costal hypersaline environments. Brine shrimp are highly adaptable to such abiotic environmental fluctuations and exhibit the ability to survive in salinity concentrations ranging from 10–200 parts per thousand (ppt), and water temperatures ranging from 10–35°C. To assess the effect of the environment on development, we examined whether various incubation temperatures (20°C, 25°C and 30°C) directly affect morphological and physiological phenotypes of developing brine shrimp during a 40 day period from hatch to late juvenile/adult stages. Morphology was assessed throughout development using measurements of brine shrimp total body length (top of the median eye to the posterior of the caudal furca) and third appendage length (body's midline to the tip of the third swimming appendage). Developmental instar stage and survival were also recorded. Oxygen consumption rate (Vo2) was used as a functional measure of energy use throughout development. We found survival significantly decreased throughout development (P < 0.001). Survival to 40 days post hatch (dph) was similar at 20°C and 25°C, but no brine shrimp survived at 30°C beyond 30 dph. While a two‐way ANOVA indicated temperature had a significant effect on total body length (P = 0.002) and length of the third appendage (P < 0.001), temperature was not significant when examined on each developmental day. Therefore, the effect of developmental day was examined for all temperatures combined. Both total body length and length of the third appendage displayed trends of increasing length with development for all temperatures combined (P < 0.001 for both). Instar stage significantly increased with developmental day (P < 0.0001), but temperature only had a significant effect on development until 11 dph. The interaction between incubation temperature and developmental day for whole animal Vo2 was not significant (P=0.074), and thus the slope of whole animal Vo2 against developmental day did not differ between temperatures. Mass‐specific Vo2 was also not strongly affected by temperature, but brine shrimp showed a peak in mass‐specific Vo2 around 15 dph. The minimal temperature effect on development, morphology, growth, and Vo2 suggest that developmental brine shrimp metabolically compensate for temperatures within the range of 20–30°C, which likely reflects the variable thermal environment they live in.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Defining Functional Human Variants in the HMG Box of SOX Genes of >60,000 Human Individuals for Potential Cardiovascular and Cancer Genetic Risk
Authors: Rasicci Daniel , Underwood Adam C. , Freeland Thomas , Milsted Amy , Jacob Howard J. , Prokop Jeremy
Published: June 2021

Our group and others have previously linked SOX (SRY‐related HMG‐box) genes to cardiovascular regulation and cancer. SOX proteins are transcription factors containing a highly conserved HMG (high mobility group) box DNA binding domain that enables sequence specific DNA interaction, bending, and gene regulation. Sequence variation in the HMG box of SOX genes has not been fully explored in humans; therefore, we have evaluated the variants within the 20 human SOX genes from ~60,000 exomes/genomes of gnomAD and also the cancer COSMIC database for somatic variations. Each variant was analyzed using codon selection and molecular modeling. A total of 3,049 variants were identified from gnomAD and 1,566 from COSMIC, with an enrichment of variants that falls within the HMG domain in cancer patients. Uniquely we highlighted an enrichment of HMG box variants in cancers of the large intestine within SOX genes with the highest enrichment in the SOX9 gene. Our most conserved and potentially impactful variant was identified as SOX18 E137K, found within 0.8% of Latino individuals. By comparing stability differences of the E137K variant against wild type SOX18 structures, using molecular dynamics simulations, SNP analysis at amino acid 137 (Glutamic acid to Lysine) was shown to disrupt the structure of the third α‐helix of the HMG box. Since other variants that fall within the HMG box of SOX18 are associated with disorders of the vasculature and lymphatics, we suspect SOX18 E137K to potentially affect vascular function through the alteration of its interaction with DNA. Work is underway to begin to assess the physiologic implications of this novel variant.

Support or Funding Information

Support for this study was from NIH‐K01ES025435 (to JWP) and Walsh University

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
White light emitting diode suppresses proliferation and induces apoptosis in hippocampal neuron cells through mitochondrial cytochrome c oxydase‐mediated IGF‐1 and TNF‐α pathways
Authors: Yang Yang
Published: June 2021

Light emitting diode (LED) light has been used in the treatment of traumatic brain injury, neural degenerative diseases and psychiatric disorders. A growing number of studies indicate that blue LED light affects cell proliferation and apoptosis in photosensitive cells and cancer cells, yet it remains unknown whether white LED light exposure impacts hippocampal neural cell proliferation and apoptosis. The present study was aimed to demonstrate the effect of white LED light exposure on viability of hippocampal neural cells and to reveal the underlying mechanisms. Cells exposed to white LED light for 24 h showed reduced viability, with higher apoptotic rate and a cell cycle arrest at G0/G1 phase. White LED light activated mitochondrial cytochrome c oxidase (Cco), in association with enhanced ATP synthase activity and elevated intracellular ATP concentration. Also, reactive oxygen species (ROS) and nitric oxide (NO) production were increased, accompanied by higher calcium concentration and lower mitochondrial membrane potential. Concurrently, the mRNA expression and the concentration of IGF‐1 were decreased, while that of TNF‐α were increased, in light‐exposed cells, which was supported by the luciferase activity of both gene promoters. The down‐stream mitogen‐activated protein kinase (MAPK), AKT/mTOR pathways were inhibited, in association with an activation of apoptotic caspase 3. N‐Acetylcysteine, a ROS scavenger, protected the cells from LED light‐induced cellular damage, with rescued cell viability and restored mRNA expression of IGF‐1 and TNF‐α. Our data demonstrate that white LED light suppresses proliferation and induces apoptosis in hippocampal neuron cells through mitochondrial Cco/ROS‐mediated IGF‐1 and TNF‐α pathways.

Support or Funding Information

The National Basic Research Program of China (2012CB124703), the Special Fund for Agro‐scientific Research in the Public Interest (201003011), the Fundamental Research Funds for the Central Universities (KYZ201212) and the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Determining the Effect of MRP4 Knockout on Pigment Granule Position in Mouse RPE
Authors: Ibarra Dora Evelyn , Mireles Thomas , Pattillo Amanda , Przybylski Connor , Roberts Tasha , Wood Tiffany S. , Schuetz John , García Dana M.
Published: June 2021

In the retinal pigment epithelium (RPE), pigment granules of wild type mice aggregate in darkness and disperse in light. In fish, pigment aggregation involves transport of cAMP into the RPE in vitro. We speculate that multidrug resistance protein 4 (MRP4) is responsible in vivo for the export of cAMP from the photoreceptors into the sub‐retinal space, making it available for uptake into the RPE in both fish and mice. We hypothesize if the MRP4 gene is knocked out in mice, we will not see a significant difference in pigment position between light and dark adapted mice. To test the hypothesis, we have quantified pigment position in RPE from wild type mice and knockout mice by estimating pigment granule density in the basal part of the cells examined by TEM. Bruch's membrane was used as a marker to determine the location of the base of the RPE. To measure pigment position, 2 μm × 2 μm boxes were aligned along Bruch's membrane, the pigment granules within each box were counted, and the average pigment density was calculated. The average pigment granule density for light adapted wildtype (LAWT; n = 3 mice) was 0.7 pigment granules/μm2 (± 0.2 pigment granules/μm2) and for dark adapted wildtype (DAWT; n = 3 mice) was 1.4 pigment granules/μm2 (± 0.3 pigment granules/μm2); pigment density was statistically significantly different (P < 0.02) between LAWT and DAWT. Preliminary data suggests the average pigment granule density for dark adapted MRP4 knockout is 0.57 pigment granules/μm2 (DAKO; n = 2). Our results with WT indicate basal pigment density increases in the dark, consistent with pigment aggregation and suggests dark adaptive pigment aggregation may be impaired in knockout mice.

Support or Funding Information

This project was supported by an Undergraduate Research Fellowship to AP and funds from the Office of the Provost awarded to DG in recognition of her service as Presidential Fellow. DEI received salary support from HSI STEM Program (84.031c) Award #P031C160035.

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