bluuebunny/crossref_metadata_2025 huggingface dataset:
POSSIBLE PROTECTIVE EFFICACY OF ETHANOL EXTRACT OF GARCINIA KOLA SEED ON CANDIDIASIS COMPLICATION IN DIABETES MELLITUS
Authors: Adeyeoluwa Temitayo Esther , Okeleye Benjamin Ifeoluwa , McGaw Lyndy , Ashafa Anofi Omotayo
Published: June 2021

Management of the complications of diabetes mellitus (DM) like candidiasis, a prevalent condition in DM patients, is important in the survival and longevity of life of the patient. In this study, Garcinia Kola nut, a medicinal seed with established anti‐diabetic and anti‐inflammatory activity, was investigated for its anti‐candida properties. Three (3) different extracts of G. kola, viz; the aqueous, hydro‐ethanol and ethanol extracts were prepared and in vitro assays were carried out to determine the anti‐ candida activity and minimum inhibitory concentration (MIC) of the seed extracts using agar well and microdilution techniques, while routine, in vitro analyses of the phytochemical constituents, anti‐oxidative, anti‐diabetic and anti‐inflammatory assays were also carried out on the seed extracts. Ethanol extract showed high anti‐oxidative and anti‐diabetic activities compared to the aqueous and hydro‐ethanol extracts. It also had higher phenolic, flavonoid and flavonol contents. The ethanol extract also showed low α‐amylase and high α‐glucosidase inhibitory activity. Similarly, the ethanol extract had a zone of inhibition of 12–15 mm in the agar‐gel diffusion assay against all the strains of Candida albicans. It had an MIC of 0.39 mg/ml and MFC of 1.563 mg/mL compared with the MIC and MFC of 0.781 mg/mL and 3.125 mg/mL for both aqueous and hydro‐ethanol extracts respectively. The cytotoxicity assay also showed that the ethanol extract has LC 50 of 90 μg/mL. The results of this study show that Garcinia kola seed can be an effective anti‐candida agent suitable for use in diabetic patients.

Support or Funding Information

Directorate of research development. University of the Free state, South Africa. Internal fund

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Low‐Density Lipoprotein Receptor‐Related Protein 1 (LRP1) Is Degraded Primarily by The Lysosomal Degradation System in Hepatocellular Carcinoma (HepG2) Cells
Authors: Tenglin Karin , Aragòn Brian Aranibar , Aguirre Stefany Huaynasi , Rondon‐Ortiz Alejandro Nico , Pino‐Figueroa Alejandro
Published: June 2021

LRP1 is a transmembrane protein responsible for binding and internalizing at least 50 different ligands, one of which is amyloid beta (Aβ). LRP1 has a role in clearance of brain and systemic Aβ. Understanding LRP1's role in Aβ degradation could have important implications for understanding Alzheimer's Disease (AD) pathogenesis. Since LRP1 in the liver is primarily responsible for systemic clearance of Aβ, its degradation could have important implications for AD pathogenesis.

Normal degradation of proteins typically occurs through the ubiquitin proteasome system (UPS) and/or the lysosomal degradation system. To study LRP1 degradation through either of these two processes, one system was inhibited, then subsequent LRP1 output was analyzed. By inhibiting one degradation system at a time, we can analyze the protein output to quantify how much protein is degraded by each system. Bafilomycin A1 was used to inhibit the lysosomal degradation system, and MG132 was used to inhibit UPS. Finally, cycloheximide, a protein synthesis inhibitor, was used to determine LRP1 half‐life and to determine whether protein output is significantly affected by protein synthesis during inhibition of the protein degradation system (cycloheximide chase analysis).

HepG2 cells were used to study LRP1 degradation in these experiments. The cells were treated with Bafilomycin A1, MG132, or dimethyl sulfoxide (DMSO; control group) and lysed at time points 0, 8, 16, and 24 hours to analyze LRP1 protein levels over time. Experiments with Bafilomycin A1 showed increasing levels of LRP1 over time, whereas experiments with MG132 showed unchanged, then decreasing levels of LRP1 over time. Cycloheximide was tested to determine the optimal concentration for HepG2 cells, which was found to be 30 μM. Then, experiments were performed with either cycloheximide and DMSO (as control), cycloheximide and Bafilomycin A1, or cycloheximide and MG132 at time points 0, 8, 16, and 24 hours to determine the half‐life of LRP1. Our results showed an extension of LRP1 half‐life by inhibition of lysosomal degradation system when using Bafilomycin A1, but half‐life was not extended when UPS was inhibited by MG132. Due to its significant increase in half‐life with Bafilomycin A1, but not with MG132, it's concluded that LRP1's degradation is primarily produced through the lysosomal degradation system.

Support or Funding Information

MCPHS 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.
Silencing of MAP Kinase Phosphatase 1 Blocks the Renal Inflammatory Response Induced by Chronic Stress
Authors: Roland Miranda M. , Langreck Cory , Duric Vanja , Babich Victor , Di Sole Francesca
Published: June 2021

Chronic stress can lead to disruption of the body systemic equilibrium which may correspond to dysfunction of both nervous and peripheral organ systems. A relationship between chronic stress, mood disorders (e.g. major depression) and systemic disease (e.g. kidney disease) is proposed via an immune system reaction and increased level of proinflammatory cytokines. This study aims to determine whether: 1. chronic stress induced by exposure of rats to corticosterone (CORT) in drinking water for 3 weeks (i.e., 50 μg/mL CORT ad libitum for 2 weeks; 25 μg/mL for 3 days and, then, 12.5 μg/mL for another 3 days) associates with a renal immune reaction and, possibly, kidney dysfunction and 2. silencing of MAP Kinase Phosphatase 1 (MKP‐1) affects the renal immune reaction mediated by CORT. MKP‐1 is a negative regulator of the MAP kinase signaling that is overexpressed within multiple human hippocampal sub‐regions of the brain of depressed subjects and is fundamental for the development of depressive‐like behaviors in rodent model of chronic stress. The neutrophil gelatinase‐associated lipocalin (NGAL) and IL‐18 are early diagnostic biomarkers. They accumulate in the kidney in response to inflammation, kidney injury, and decreased kidney function. Protein levels of NGAL and IL18 were analyzed by immunocytochemistry. NGAL and IL18 protein levels were significantly increased in rats treated with CORT compared to their controls (over 70% increase in NGAL and IL18 protein levels in the glomeruli, 40% increase in NGAL and 6% increase in IL18 protein levels in the tubules). The increase in NGAL and IL18 protein levels was blocked by micro‐infusion of recombinant adeno‐associated virus encoding Cas9 with guide RNA targeting MKP‐1 used to silence MKP‐1 protein expression prior to exposure to CORT. In summary, renal inflammation induced by chronic stress was blocked by MKP1 silencing. This study supports the therapeutic potential of MKP‐1 as target for signaling pathways between chronic stress, mood disorders and kidney dysfunction.

Support or Funding Information

Iowa Osteopathic Education and Research Funds

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Angiotensin AT2 Receptor Stimulation is Protective in Lipopolysaccharide‐Induced Inflammation and Renal Injury
Authors: Hussain Tahir , Dhande Isha , Patel Sanket Niranjanbhai , Ali Quaisar
Published: June 2021

Recently angiotensin type‐2 receptor (AT2R) activation has been shown to exert anti‐inflammatory and reno‐protective response against chronic injury. Our prior in vitro studies using the proximal tubule epithelial cells (PTECs) and macrophages revealed that prior activation of angiotensin‐II type 2 receptor (AT2R) reduces pro‐inflammatory cytokine production in response to lipopolysaccharide (LPS). However, the in vivo effects of AT2R stimulation on LPS‐mediated inflammatory renal injury are not known. The present study tests the hypothesis that AT2R activation attenuates inflammation and preserves renal structure and function in the early stages of LPS‐mediated renal injury. Mice were pretreated with vehicle or AT2R agonist C21 (300 μg/kg, i.p.) prior to LPS (5 mg/kg, i.p) challenge. Pretreatment with C21 significantly prevented the LPS‐induced increase in the levels of renal pro‐inflammatory cytokines (TNF‐α, IL‐6), TLR4 expression, and markers of renal injury (blood urea nitrogen, plasma creatinine and albuminuria). Moreover, C21 treatment also protected against LPS‐induced tubular vacuolization and tubular degeneration. Renal and circulating anti‐inflammatory IL‐10 levels were higher with C21 pretreatment at time of LPS challenge. In vitro studies in HK‐2 cells (human PTECs) demonstrate that AT2R activation directly stimulates IL‐10 production. Further, conditioned‐media derived from the C21‐treated HK‐2 cells reduced LPS‐induced TNF‐α production via IL‐10 originating from HK‐2 cells. Our findings suggest that prior activation of AT2R prevents LPS‐mediated inflammatory renal injury, possibly via an increase in renal IL‐10 production.

Support or Funding Information

This study was supported by NIH R01 grant DK61578

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Gonadectomy Affects Water‐Deprivation‐Induced Urinary Concentration in Female and Male Mice
Authors: Delaney Dalton , Waturuocha Chiedozie , Fan Liming , Rouch Al
Published: June 2021

The urinary concentrating mechanism is critical for maintaining whole‐body fluid balance and plasma osmolarity. Estrogen has been shown to downregulate vasopressin‐sensitive aquaporin‐2 (Aqp2) expression (Cheema, MU, et al. Am J Physiol, Renal Physiol, 309, F305–F317, 2015). While sex steroids have been reported to influence urinary concentration, additional studies are clearly needed. The purpose of this study was to determine if gonadectomy affects urine osmolarity (Uosm) during water deprivation (WD) in mice. Intact female and male (IF, IM), ovariectomized (OVX), and castrated (CA) CD‐1 mice at four weeks of age were obtained from Envigo, Inc. (Indianapolis, IN). Two studies were conducted: the first served as a control study and the second involved WD. The control study consisted of a four‐day period where IF, OVX, IM, and CA mice (3/group), in metabolic cages, consumed, ad libitum, standard rodent chow (0.25% salt) and water for four days. The WD study also consisted of four days where water was removed from 7 pm to 7 am in the final three days. Daily measurements included food and water intake, urine flow rate, Uosm, and sodium and potassium excretion. Urine samples were collected after 7 am just before replacing water for the mice. Uosm was measured using a vapor‐pressure osmometer. In the control study no differences between any groups were measured. ANOVA revealed that in the WD study, the IF mice had significantly lower Uosm (mOsm/L), averaged from the three WD days, compared to OVX mice (5,924 ± 431 vs 8,400 ± 691, respectively, p < 0.01). The IM mice had significantly higher Uosm compared to CA mice (8,683 ± 691 vs 3,590 ± 382, respectively, p<0.0001). Other Uosm comparisons: IM > IF (p<0.005) and IF > CA (p<0.01). We conclude that gonadectomy augments urinary concentration in female mice while it reduces urinary concentration in male mice during WD. Results suggest significant roles for the sex steroids, estrogen and testosterone, in urinary concentration. RT‐qPCR studies are underway to determine potential expression differences in Aqp2 in all groups from this study.

Support or Funding Information

OSU‐CHS

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Submaximal Level Single Twitch Kinetics Dependent on Disease State in Duchenne Muscular Dystrophy Mouse Model
Authors: Peczkowski Kyra K. , Rastogi Neha , Schultz Eric J. , Rafael‐Fortney Jill A. , Janssen Paul M. L.
Published: June 2021

In muscular dystrophy research, it is pertinent to analyze the force of the muscles affected by the disease to assess pathology and to assess potential effectiveness of interventions. Maximal tetanic contractions are the most commonly used protocol to assess muscle function. However, the vast majority of in vivo muscle function is at submaximal contraction. Moreover, although force of contraction is assessed, the kinetics of the contraction are rarely reported. To investigate the effect of muscular dystrophy on twitch contraction kinetics, isolated diaphragm and extensor digitorum longus (EDL) of 20‐week untreated “het” (dystrophin deficient and utrophin haploinsufficient) and wild‐type C57 mice were analyzed. In isolated diaphragm strips, Time‐to‐Peak (TTP) is slower in “het” mice (14.2 ± 0.3 ms) than in C57 (11.5 ± 0.3 ms) mice, while no significant difference was identified for Time for 50% Relaxation (RT50) of the twitch contraction. In isolated EDL muscles, TTP was also slower in “het” mice (24.0 ± 0.4 ms) compared to C57 mice (20.7 ± 0.3 ms). Unlike the diaphragm, it was found that RT50 of EDL muscles is also disease‐dependent. The RT50 of “het” mice (15.3 ± 0.6 ms) was significantly slower than the C57 model (11.8 ± 0.2 ms). These findings suggest that not only is the maximal force of the muscles important, but also the kinetics of the muscles are impacted by disease.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Isometric Handgrip Echocardiography is a More Powerful Diastolic Discriminator than Conventional Cycle Echocardiography
Authors: Samuel T. Jake , Beaudry Rhys , Haykowsky Mark , Sarma Satyam , Nelson Michael Douglas
Published: June 2021

Cycle echocardiography is currently recommended for assessing cardiac involvement (i.e. left ventricular diastolic dysfunction) in patients with unexplained dyspnea on exertion. However, cycle echocardiography is often limited by movement and respiratory artifact, particularly in obese patients with dyspnea. Our group has recently shown that isometric handgrip echocardiography is a powerful diastolic discriminator, capable of unmasking subclinical stress induced diastolic dysfunction. How isometric handgrip echocardiography compares to conventional cycle echocardiography however, remains unclear and was the focus of the present investigation. We hypothesized that isometric handgrip echocardiography would unmask more diastolic dysfunction, given its markedly different hemodynamic load compared to cycle echocardiography. To test this hypothesis, we recruited 24 individuals from the community (9 male, 15 female, age range: 18 – 80), who all performed 3 minutes of isometric handgrip echocardiography followed by 3 minutes of dynamic cycle exercise (20 W). At rest and during the final minute of each exercise protocol heart rate (HR), mean arterial pressure (MAP) and Doppler derived E/e′ were recorded. Consistent with our previous work, and that of others, responders were defined as ΔE/e′ > 1.5 while non‐responders were defined as ΔE/e′ < 1.5. Both isometric handgrip and low‐intensity cycle exercise resulted in a similar rise in HR (ΔHR: 22 ± 13 vs. 25 ± 7, handgrip vs. cycle exercise, P > 0.05), while isometric handgrip resulted in a larger increase in MAP (ΔMAP: 28 ± 14 vs. 16 ± 12, handgrip vs. cycle exercise, P = 0.0003). Remarkably, the isolated increase in afterload experienced by the myocardium during isometric handgrip exercise was more sensitive at unmasking sub‐clinical diastolic dysfunction in community dwelling individuals compared to cycle exercise (handgrip: n = 14 vs. n = 10; and cycle: n = 10 vs. n = 14, responders vs. non‐responders, respectively). Taken together, these data highlight the usefulness of isometric handgrip echocardiography at isolating myocardial diastolic relaxation abnormalities in community dwelling individuals, beyond that of dynamic cycle exercise. Future work should focus on confirming the sensitivity of this method in individuals at risk for or with diagnosed heart failure.

Support or Funding Information

AHA 16SDG27260115 and the Harry S. Moss Heart Trust.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Respiratory Muscle Training Positively Affects Vasomotor Response in Young Healthy Women
Authors: Esposito Fabio , Bisconti Angela Valentina , Devoto Michela , Venturelli Massimo , Bryner Randall , Olfert Mark Ivan , Chantler Paul
Published: June 2021

Vasomotor response is related to the capacity of the vessel to maintain the homeostasis of the vascular tone. Two main control mechanisms are involved: the autonomic control of the sympathetic neural drive (global control) and the endothelial smooth cells capacity to respond to mechanical stress by releasing vasoactive factors (local control). The aim of the study was to evaluate the effects of respiratory muscle training (RMT) on vasomotor response, assessed by flow‐mediated dilation and heart rate variability, in young healthy females. The hypothesis was that RMT may enhance the balance between sympathetic and parasympathetic neural drive and reduce vessels shear stress. To this purpose, twenty‐four women were assigned randomly to RMT or SHAM group. Maximal inspiratory mouth pressure and maximum voluntary ventilation were utilized to assess the effectiveness of the RMT program, which consisted of three sessions of isocapnic hyperventilation per week, eight weeks, for a total of twenty‐four training sessions. Heart rate variability was utilized to evaluate the autonomic balance, a global factor regulating vasomotor response. Endothelial function was determined by measuring brachial artery vasodilation normalized by the shear rate (%FMD/SR). After RMT, maximal inspiratory mouth pressure and maximum voluntary ventilation increased significantly (+31% and +16%, respectively) only in RMT. Changes in heart rate variability were negligible in both groups. Only RMT exhibited a significant increase in endothelial function (%FMD/SR +45%; p<0.05). These data indicate a positive effect of RMT on vasomotor response likely via a reduction in shear stress in the vessels, and not through a modulation of sympatho‐vagal balance.

Support or Funding Information

Grant from the Department of Biomedical Sciences for Health, Università degli Studi di Milano.

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 High‐intensity Interval Training and Moderate‐intensity Continuous Training on Circulating MicroRNAs in Sedentary Males
Authors: Chen Kuan Ming , Huang Ching‐Hui , Wang Jong‐Shyan
Published: June 2021

Circulating microRNAs (c‐miRs) serve as the reliable biomarkers for exercise adaption. The present study aimed to clarify the relationships between changes of c‐miRs expressions and aerobic capacity/hemodynamic functions caused by various exercise regimens in men. Twenty‐four healthy sedentary males were randomized to engage in either high‐intensity interval training (HIIT, 3‐minute intervals at 40% and 80% of VO2max, n=12) or moderate‐intensity continuous training (MICT, sustained 60% of VO2max, n=12) for 30 minutes/day, 5 days/week for 6 weeks. Various c‐miRs involved in angiogenesis (miR‐126), cardiac hypertrophy (miR‐222, miR‐221), and hypoxia/ischemia adaptation (miR‐21, miR‐146a) were measured by qRT‐PCR, respectively. The analyzed results showed that 6‐week HIIT and MICT significantly enhanced VO2max, which were accompanied by increased stroke volume and cardiac output during exercise. Moreover, both HIIT and MICT increased expressions of miR‐126 (1.49±0.03 and 1.27±0.09 folds) and miR‐222 (1.32±0.10 and 1.25±0.15 folds), respectively. Additionally, HIIT elevated miR‐146a expression (1.33±0.09 folds), whereas MICT upregulated miR‐21 (1.32±0.08 folds). However, no significant change in plasma miR‐221 level was occurred after both HIIT and MCT. Therefore, we conclude that the exercise‐induced cardiovascular adaptions are associated with the changes of angiogenesis/hypertrophy‐related c‐miRs following either HIIT or MCT.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
SPECC1L: A Novel Cytoskeletal Regulator of Epithelial Organization in Craniofacial Morphogenesis
Authors: Saadi Irfan
Published: June 2021

Craniofacial anomalies are extremely common among birth defects, with clefts of the lip and palate affecting approximately 1/700 live‐births. While many contributory genes have been identified, a large proportion of the genetic component of this complex disease is unknown. Thus, there is a continued need to identify additional genes and to understand underlying pathogenetic mechanisms. We first identified heterozygous SPECC1L mutations in two patients with atypical facial clefts and cleft palate. New heterozygous missense SPECC1L mutations have been identified in two autosomal dominant Opitz G/BBB syndrome families with hypertelorism and cleft palate, and in two patients with Teebi hypertelorism syndrome, underscoring the importance of SPECC1L in craniofacial development. To understand the role of SPECC1L, we have generated several mouse models with null and hypomorphic (truncation, in‐frame deletion, missense) mutations. Mutants for these alleles manifest a range of structural malformations, including exencephaly, cleft palate and ventral body wall defects. The homozygous null mutants die by embryonic day 9.5 (E9.5) with an open neural tube and failure of cranial neural crest cells to delaminate from the neuroectoderm. We have shown that the latter is due to increased expression of adherens junction markers in the neuroectoderm and reduced PI3K‐AKT signaling. Homozygous mutants for a hypomorphic allele that results in SPECC1L truncation die perinatally and show midline subepidermal blebbing with no cleft palate. Specc1lnull/hypo compound mutants with moderate SPECC1L deficiency show palate elevation defects at E14.5, which resolve within one embryonic day such that only a few mutants are born with cleft palate. However, these compound mutants show abnormal organization of the oral periderm of palatal shelves and occasional oral adhesions at E13.5. Consistent with these periderm anomalies, SPECC1L expression in the palate requires IRF6, thus making it a novel member of the IRF6/GRHL3 pathway in periderm development. Lastly, homozygous mouse mutants carrying human Opitz G/BBB patient mutation (T397P) show highly penetrant cleft palate, indicating a gain‐of‐function that specifically affects palatogenesis. Given the broad phenotypic spectrum of Specc1l allelic series mutants, our studies indicate that SPECC1L regulates epithelial organization at various stages and in multiple tissues throughout embryonic development.

Support or Funding Information

This research is supported by NIH/NIDCR R01DE026172.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Protective Effects of Human Milk Oligosaccharides on Intestinal Epithelial Function Assessed in Enteroid‐Derived Monolayers
Authors: Drobny Alice , Ibeawuchi Stella Rita , Das Soumita , Bode Lars , Barrett Kim Elaine
Published: June 2021

Human milk oligosaccharides (HMOs) are structurally diverse carbohydrates that are unique to human milk and have beneficial effects on the growth and development of the intestine and immune system in breastfed infants. Infectious diarrheal diseases account for significant morbidity and mortality in young children and breastfeeding is protective against non‐typhoidal Salmonella infections and associated diarrhea in infants. Several studies suggest that HMOs may prevent enteric infections in breastfed infants and/or improve barrier function in transformed cell lines. Some of the beneficial effects of HMOs may be attributable to changes in the microbiota, but they likely have direct effects on the epithelium as well. The purpose of this study was therefore to evaluate mechanisms whereby HMOs may improve intestinal integrity and any protective effects of HMOs on Salmonella infection in a more clinically‐relevant model. We previously showed that Salmonella infection decreases epithelial barrier function in murine colon and in murine enteroid‐derived monolayers (EDM). In the present study, EDM were developed on permeable supports and, after differentiation, were exposed to pooled HMOs and then subsequently infected with Salmonella typhimurium at a multiplicity of infection of 10:1. Barrier integrity was determined by measuring transepithelial electrical resistance (TEER). We also measured bacterial internalization using a gentamicin protection assay followed by colony counts. Exposure of EDM to 10 mg/mL pooled HMOs for 2–8 h progressively increased TEER. Prior exposure to pooled HMOs prevented the decrease in TEER that was otherwise produced by Salmonella infection at 1 h after infection, but this effect was lost at later time‐points. Pooled HMOs had no effect on numbers of intracellular bacteria and did not alter bacterial growth when added to bacteria directly. Our results suggest that HMOs act directly on intestinal epithelial cells to strengthen intestinal barrier function both at baseline and when compromised by a pathogen. These effects may contribute to protection against the adverse outcomes of neonatal enteric infections.

Support or Funding Information

Larsson Rosenquist Foundation Mother Milk Infant Center of Research Excellence

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
OVARIECTOMIZED RATS EXHIBIT ANXIETY‐LIKE BEHAVIOR AND INCREASED NEURONAL ACTIVATION IN THE PERIVENTRICULAR NUCLEUS
Authors: Campos Glenda Siqueira Viggiano , Noronha Sylvana , De Souza Aline Viggiano , Lima Paulo , Chianca‐Jr Deoclecio , De Menezes Rodrigo
Published: June 2021

Introduction

Ovarian follicle failure and subsequent estradiol deficiency as a result of menopause leads to impairments in the cardiovascular system and brain. Reduction of estradiol levels is associated with the development of hypertension and anxiety, suggesting estradiol exerts cardioprotective and neuroprotective effects. Furthermore, studies have shown that estradiol deficiency increases circulating and tissues levels of the vasoconstrictor angiotensin II, which itself can increase anxiety responses. The paraventricular nucleus (PVN) is a brain area that has a high density of angiotensin type 1 receptors (AT1R) and beta‐estrogen receptors (ERβ). These observations suggest estradiol modulates anxiety responses in part through regulation of the renin angiotensin system (RAS). The aim of this study was to evaluate the impact of estradiol on anxiety‐like behavior and the neuronal activation on the PVN.

Methods

Three month old (mo) female Fischer rats were divided into Intact (INT), ovariectomized (OVX) and Sham‐operated (SHAM) animal groups. Three weeks after surgery, the animals were submitted to a battery of anxiety tests including the elevated T‐maze (ETM), open field (OF) and light/dark box (LDB). Ninety minutes later, the animals were euthanized and the brains were collected to assess c‐Fos activity. Anxiety tests in INT animals were conducted during proestrus or diestrus phase while Sham animals were tested in the diestrus phase. A handling group (H), which was not exposed to behavioral tests, was added as a control for the c‐Fos protocol.

Results

Our results found that the different phases of the estrus cycle did not influence the behavior of the INT group on the ETM, OF and LDB behavioral tests (p>0.05, Mann‐Whitney test). In contrast, the OVX group showed increased inhibitory avoidance and reduced escape latencies in the ETM (p<0,05, Mann‐Whitney test) and also reduced numbers of entries on the dark side of the LDB test (p<0.05, Mann‐Whitney test) in comparison to the Sham control, indicating anxiety‐like behavior. Moreover, the OVX group showed higher neuronal activation in the PVN compared to the other animal groups including OVX‐H (p < 0.001), INT (p < 0.001), and Sham (p < 0.01) by one‐way ANOVA.

Conclusion

Estradiol deprivation caused by ovariectomy induces anxiety‐like behavior in a myriad of behavioral tests under conditions in which neuronal activation of the PVN occurs.

Support or Funding Information

CAPES, CNPq, FAPEMIG, UFOP

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Neurobiological Correlates of Pain Avoidance Behavior in Opioid vs. Alcohol Dependence
Authors: Pahng Amanda R. , Paulsen Rod I. , McGinn M. Adrienne , Schindler Mackenzie E. , Edwards Kim N. , Edwards Scott
Published: June 2021

In contrast to their analgesic properties, excessive use of either alcohol or opioids produces a paradoxical emergence of heightened pain sensitivity to noxious stimuli, termed hyperalgesia, which may promote further drinking or use of opioids to manage worsening pain symptoms. Hyperalgesia has traditionally been measured in rodents via reflex‐based assays, including the von Frey method. To better model the cognitive/motivational dimension of pain in a state of alcohol/opioid dependence and withdrawal, we employed a recently developed non‐reflex‐based method for measuring pain avoidance‐like behavior in animals (Mechanical Conflict Avoidance test). Adult male Wistar rats were administered an escalating dose regimen of morphine (opioid‐dependent group) or repeated saline (control group). Morphine‐dependent rats exhibited significantly greater avoidance of noxious stimuli during withdrawal. Such pain avoidance‐like behavior exhibited a modest correlation with mechanical paw withdrawal thresholds as measured via the von Frey method (r=−0.4413, p=0.0452), indicating that the two behavioral assays measure overlapping but not entirely similar pain‐like conditions. We next investigated individual relationships between pain avoidance‐like behavior and alterations in protein phosphorylation in central motivation‐related brain areas. We discovered that pain avoidance‐like behavior was significantly correlated with alterations in phosphorylation status of protein kinases (ERK, CaMKII), transcription factors (CREB), presynaptic markers of neurotransmitter release (Synapsin), and the rate‐limiting enzyme for dopamine synthesis (TH) across specific brain regions. Separate groups of male Wistar rats were made dependent on alcohol via exposure to chronic, intermittent ethanol vapor and compared to air‐exposed (non‐dependent) controls. Here, alcohol‐dependent animals exhibited enhanced pain avoidance‐like behavior of shorter probe heights that were innocuous (i.e., not avoided) in non‐dependent animals, indicative of a state of allodynia. We are currently examining the neural correlates of pain avoidance in these groups to compare to our results in opioid‐dependent animals. Our findings suggest that alterations in phosphorylation events in specific brain centers may support cognitive/motivational responses to avoid pain, and that qualitatively distinct dimensions of pain avoidance‐like behavior (hyperalgesia vs. allodynia) may exist in opioid‐ vs. alcohol‐dependent animals.

Support or Funding Information

National Institute on Alcohol Abuse and Alcoholism (T32AA007577, ARP; R00AA020839, SE), LSUHSC School of Medicine start‐up funds.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Comparison of effects of serine racemase knockout on nociceptive behavior among different models of neuropathic and inflammatory pain
Authors: Kato Eiko , Saotome Toshiya , Fukushima Teruyuki , Takasusuki Toshifumi , Yamaguchi Shigeki , Hori Yuuichi
Published: June 2021

Peripheral nerve injury and tissue inflammation often induce abnormal pain. The activation of N‐methyl‐D‐aspartate (NMDA) receptors mediates the development and maintenance of such abnormal pain. D‐serine co‐localizes with NMDA receptors and acts as an endogenous co‐agonist of the NMDA receptor at the glycine binding site. D‐serine is synthesized from L‐serine by serine racemase (SR), which is an enzyme that is abundant in the central nervous system. It remains unclear how alterations in SR activity are involved in modulation of neuropathic and inflammatory pain behavior. In the present investigation, we compared pain behaviors in SR knockout (KO) mice and wild‐type (WT) mice after subjection to a chronic constriction injury (CCI model of Bennett), partial sciatic nerve ligation (PSL model of Seltzer), spinal nerve ligation (SNL model of Chung), and an inflammatory pain model that was induced by subcutaneous formalin.

Seven‐ to nine‐week‐old male WT and SR‐KO mice were used for the experiments. Under halothane anesthesia, the mice were subjected to neuropathic surgery. Three types of nerve ligation were performed, as previously described by Bennett et al. (1988), Kim and Chung (1992), and Seltzer et al. (1990). The left sciatic nerve was tied loosely with four ligatures of chromic gut at the mid‐thigh level (Bennett model), the left L5 and L6 spinal nerves were isolated and ligated tightly with a 4‐0 silk thread (Chung model), and the dorsal one‐third to one‐half of the left sciatic nerve was tightly ligated (Seltzer model). Mechanical allodynia was assessed by measuring the frequency of withdrawal responses using von Frey filaments both before and after ligation. To induce inflammatory pain, paraformaldehyde (50 μL, 25% in distilled water) was injected subcutaneously in the dorsal surface of the left hind paw. The mice were continuously observed for 60 min after the formalin injection, and the number of flinches of the injected paw was measured during the observation period.

Behavioral observations showed the following results: 1) The second phase of the formalin‐induced licking response was augmented in SR‐KO mice, while the first phase was not; 2) mechanical allodynia in the Seltzer model was quite similar between SR‐KO mice and WT mice; 3) mechanical allodynia in both the Chung and Bennett models was significantly augmented in SR‐KO mice compared with WT mice.

The mechanisms of the presently observed different effects of SR knockout on different pain models are unclear. We are currently investigating how differently synaptic circuit activity in the spinal dorsal horn is affected by SR knockout in different pain models.

Support or Funding Information

Dokkyo Med Univ Mutual Fund

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
An Examination of Virtual Reality Interventions as a Resource for the Treatment of Phantom Limb Pain
Authors: Johnson Ashley , Todd Tyler , Block Helen , Preston Ashton , Raynes Edilberto
Published: June 2021

Virtual reality is a new and developing technique entering the medical community that may be effective in relieving pain. A specific type of pain termed phantom limb pain (PLP), is the occurrence of pain in an area that has been amputated and is reported to affect more than 50% of patients with amputations. Currently, the treatment of phantom limb pain is highly variable in the strategies used and their effectiveness. Based on the theory of maladaptive cortical reorganization causing PLP, we hypothesized that the use of a virtual reality intervention would elicit a reduction in PLP. Using the Oxford Centre of Evidence‐based Medicine (OCEBM) classification, a systematic review of the literature, was conducted to determine the pathophysiological mechanisms contributing to PLP and how virtual reality techniques could beneficially impact cortical reorganization. Based on OCEBM, the majority of the articles reviewed were level 2 evidence and supported virtual reality as a resource for alleviating PLP. We concluded that the available evidence supports that the use of virtual reality could be an alternative treatment for PLP and could decrease PLP.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effects of Roux‐en‐Y Gastric Bypass Surgery on Mitochondrial Quality Control Proteins in Human Myotubes Derived from Severely Obese Humans
Authors: Kugler Benjamin N. , Gona Philimon N. , Saunders Sasha N. , Park Sanghee , Dubis Gabe , Houmard Joseph A. , Zou Kai
Published: June 2021

Impaired skeletal muscle glucose oxidation in response to insulin is a hallmark characteristic of humans with severe obesity and Type 2 Diabetes. Roux‐en‐Y gastric bypass (RYGB) surgery is an effective treatment of these metabolic diseases by markedly improving metabolic health, including enhanced glucose oxidation and insulin action in skeletal muscle. However, the underlying mechanisms responsible for these improvements are not fully understood. Skeletal muscle mitochondria are critical in preserving cellular health, but exhibit defective functions in severely obese humans. Recent studies have suggested that dysregulated mitochondrial dynamics are associated with mitochondrial dysfunction and insulin resistance. This study aims to determine the effects of severe obesity and RYGB surgery on mitochondrial dynamic proteins and its association with insulin sensitivity and glucose oxidation in human skeletal muscle. Human skeletal muscle cell culture model was utilized by isolating human skeletal muscle cells from muscle biopsies obtained from lean subjects (n=6, BMI = 22.7 ± 1.2 kg/m2, HOMA‐IR = 1.9 ± 0.4) and RYGB patients prior to, 1‐month and 7‐months following surgery (n=6, BMI = 47.3 ± 2.8 kg/m2, 41.0 ± 2.9 kg/m2, 33.7 ± 2.2 kg/m2, HOMA‐IR = 3.4 ± 0.4, 2.9 ± 0.9, 1.7 ± 0.2, respectively). Human skeletal muscle cells were differentiated to myotubes. On day 7 of differentiation, myotubes were harvested for immunoblot analysis in order to assess the expressions of mitochondria dynamics proteins. Myotubes derived from severely obese patients had a 35% increase in mitochondrial fusion protein Optic Atrophy 1 (Opa1) expression before surgery when compared to lean controls (p <0.05). In addition, there was a trend towards a decrease of fusion proteins Opa1 and Mitofusin 1 (Mfn1) expressions in myotubes derived from the same patients at 7‐months following surgery (26% and 18%, p=0.065, respectively) in comparison to pre‐surgery state, and virtually no difference from lean controls. Furthermore, Opa1 expression negatively correlated with insulin‐stimulated glucose oxidation in myotubes (r = −0.614, p <0.05), while Mfn1 is positively correlated with HOMA‐IR (r = 0.495, p <0.05). Mitochondrial fission and mitophagy proteins observed no differences between lean, pre‐surgery, 1‐month and 7‐months following RYGB surgery. In conclusion, these data suggest that RYGB surgery corrected severe obesity‐induced dysregulation in mitochondrial fusion process in human myotubes, which may contribute to improved insulin sensitivity and skeletal muscle glucose oxidation in RYGB patients.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Tendon developmental plasticity and functional regeneration
Authors: Galloway Jenna L. , Niu Xubo , Shah Rishita R. , Nerurkar Nandan L. , Noedl Marie
Published: June 2021

Tendons enable movement by transferring force from muscle to bone. They are comprised of a highly organized collagen‐rich extracellular matrix. Disruption to the development and maintenance of tendons can lead to birth defects and result in injury in adults. We currently have an incomplete understanding of how tendons interact with their neighboring tissues to regulate the development and morphogenesis of the musculoskeletal system. In addition, once mature mammalian tendon tissue is formed, it has limited regenerative potential after injury. Central to this challenge is to uncover new mechanisms underlying tendon cell regulation during musculoskeletal development and regeneration. To accomplish this, we have turned to the zebrafish system because it is highly regenerative and amenable to genetic manipulations and live imaging. We have previously demonstrated that zebrafish tendons have molecular, developmental, morphological and biomechanical similarities to mammalian tendons, thereby making them an excellent vertebrate model to study tendon biology. However, no study has investigated their capacity to regenerate their tendons. Using the zebrafish, we developed methods to perform genetic tendon cell ablation, lineage tracing, live imaging, and mechanical testing to determine if developing and adult zebrafish can regenerate their tendon tissues. Following tendon cell ablation in developing zebrafish embryos, we observe defects in tendon gene expression and ECM composition. Cartilage and muscle tissue morphology was also disrupted following tendon cell loss. Tendon cells and the collagen organization returned after 1–2 weeks and 1–2 months, respectively. Interestingly, the pattern of the attachment sites was also restored, indicating the presence of regulatory programs that recruit and/or promote tendon cell proliferation and differentiation at the previous attachment sites. We are currently focused on identifying the developmental origins of the regenerating tendon cells and the pathways restoring the attachment site pattern. In addition to embryonic tendon regeneration, we also found that adult zebrafish appear to regenerate their collagen organization and mechanical properties after an acute physical injury. Together, our work demonstrates robust tendon regeneration at embryonic and adult stages in the zebrafish. These studies also provide a system to examine the morphogenetic and molecular events underlying tendon regeneration and musculoskeletal tissue‐tissue interactions, which has significance for regenerative biology approaches to musculoskeletal injuries.

Support or Funding Information

NICHD HD06953, NIDCR DE024771, Charles H. Hood Foundation, HSCI Seed Grant

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Renal sympathetic nerve contribute to high blood pressure in male PCOS offspring
Authors: Maranon Rodrigo Oscar
Published: June 2021

Prenatal factors may program elevated blood pressure in offspring of women with polycystic ovary syndrome (PCOS). We recently determined in an experimental model of PCOS that their male offspring have elevated blood pressure compared to offspring of controls. Existing evidence shows that PCOS women may have increased renal sympathetic nerve activity (RSNA) which contributes to their elevated blood pressure. However, whether the elevated blood pressure in PCOS offspring is due to activation of the renal sympathetic nervous system (RSNS) remains undetermined. We hypothesized that activation of the RSNS contributes to the increase in blood pressure in male PCOS offspring. PCOS dams were treated with DHT pellets (DHT pellet 7.5mg/90d) beginning at 4 wks of age and continuing through pregnancy and lactation. Controls were given placebo pellets. Male PCOS offspring (mPCOS‐off) and male control offspring (mCon‐off), 20 weeks of age, were subjected to renal denervation (RD) (cutting renal nerves bilaterally and painting with 10% phenol in ethanol) or sham surgery (movement of the kidneys) (n=9/grp): mPCOS‐off‐sham, mPCOS‐off‐RD, mCon‐off‐sham, and mCon‐off‐RD, respectively. Baseline mean arterial pressure (MAP, telemetry) was significantly higher in mPCOS‐off‐sham than mCon‐off‐sham (124±1 mmHg vs. 119±1 mmHg; p<0.05). After recovery from renal denervation or sham and telemetry implantation for 14 days, MAP was decreased with renal denervation in mPCOS‐off‐RD (mPCOS‐sham: 124±1 vs mPCOS‐RD: 111±2 mmHg, p<0.05) but not in control offspring (mCon‐off‐sham: 119±1 vs mCon‐RD: 118±3 mmHg; p=NS). Thus, male PCOS offspring have an increase in MAP compared to the male control offspring, and renal denervation reduced their blood pressure, suggesting that exposure to hyperandrogenemia in utero may activate the RSNS later in life, which could contribute to high blood pressure. These data suggest that children of women with PCOS should closely monitor their blood pressure as they age to maturity.

Support or Funding Information

Supported by AHA14POST18640015 and P20GM121334 (ROM).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Uncoupling of Feedforward Control on Blood Pressure by Heart Rate in Patients with Recent Myocardial Infarction
Authors: Badhwar Smriti , Chandran Dinu , Jaryal Ashok , Narang Rajiv , Patel Chetan , Deepak Kishore Kumar
Published: June 2021

Background

Mechanically induced respiratory synchronous thoracic pressure fluctuations produce High Frequency(HF) BP oscillations. In addition, HF oscillations in BP are also modulated through feedforward coupling with Heart Rate(HR). Reduced baroreflex sensitivity, heart rate variability (HRV) and blood pressure variability (BPV) is known to be associated with increased mortality after Myocardial Infarction(MI).

Objective

The objective of the study was to assess the LF and HF oscillations in HR and BP and their associations in patients of recent MI with and without scar.

Methods

Non‐invasive beat‐to‐beat BP measurement using Portapres® with simultaneous recording of Lead II ECG was used to assess short term (5min) HRV and BPV. Thirty‐two patients of recent MI (≤6 months) were divided into those with scar (S) (n = 21, age 54.3±10.7years) and those without scar (NS) (n = 11, age 54.7±9.22years) on the basis of myocardial perfusion imaging (MPI) using single photon emission computed tomography. Transfer function analysis was done using BP as the input variable and HR as the output variable. The signals were linearly interpolated and resampled at 1Hz. The series was subdivided into 256‐point segments with 50% overlap for spectral estimation. The coherence, gain and phase was calculated for the LF (0.04–0.15) and HF (0.15–0.4) band. LF and HF gain and phase was evaluated only if coherence between the two signals was >0.3.

Results

LF and HF HRV were similar between the two groups (LF normalized units (nu) S vs NS 53.6±22.4 vs 54.7±18.1, HF nu S vs NS 35.9±17.9 vs 36.0±12.0). Similarly, LF nu Systolic BPV was comparable between the two groups (S vs NS 66.7±17.2 vs 57.9±15.9). HF nu systolic BPV was significantly lower in the S group compared to the NS group [S vs NS 28.3(13.4–39.3) vs 39.1(19.8–46.5) p = 0.04]. LF and HF gain was also similar between the two groups [LF gain S vs NS 5.24±2.25 vs 5.86±3.09, HF gain S vs NS 7.19(3.26–10.4) vs 8.40(5.94–13.5) ms/mmHg]. There was no significant difference in the LF phase between the two groups (S vs NS 1.55±0.39 vs 1.13±0.61 radians). However, HF phase coupling of BP with HR of the S group was significantly more positive than the NS group (S vs NS 0.39±0.92 vs −0.61±1.0 radians p = 0.02).

Conclusion

HRV was comparable between the scar and no scar group however HF oscillations in BPV are attenuated in patients with irreversible damage to cardiac tissue when compared to those in whom it is preserved. The positive phase in HF in the scar group could possibly be indicative of a feedforward uncoupling between HR and BP in the scar group while the same is preserved in the no scar group.