bluuebunny/crossref_metadata_2025 huggingface dataset:
Acute prorenin infusion mediates angiotensin II‐dependent pressor response via binding to (pro)renin receptor in the central nervous system
Authors: Li Wencehng , Peng Hua , Ichihara Atsuhiro , Feng Yumei
Published: June 2021

The brain renin‐angiotensin system plays an important role in blood pressure (BP) regulation. However, the origin of brain Ang II remains unknown due to the low brain renin activity. We previously reported that prorenin and (pro)renin receptor (PRR) is present in the brain. To test our hypothesis that prorenin mediates Ang II formation in the brain via binding to PRR, the neuron‐specific PRR knockout (Nefh‐PRRKO) and wild type (WT) mice (n=5/group) were implanted with telemetric probes and intracerebroventricular (ICV) cannulas. BP was recorded in conscious mice during ICV infusion (0.3μl/min) for ten minutes. ICV infusion of Ang II (100ng/μl) increased BP (ΔMAP, mmHg) in both WT (48±5) and Nefh‐PRRKO (40±2)mice; and this effect was blocked by AT1 receptor blocker (Losartan) indicating that the AT1R is functionally intact in Nefh‐PRRKO mice. ICV infusion of mouse prorenin (100ng/μl) induced a pressor response in WT mice (42.3±5.7), and this response was completely blocked by Losartan or Captopril suggesting that prorenin induced Ang II‐dependent pressor response in WT mice. Interestingly, the pressor response to mouse prorenin was abolished in Nefh‐PRRKO mice (5±1; P<0.05), indicating that prorenin mediates pressor response via binding to PRR. In summary, prorenin via binding to PRR mediates Ang II‐dependent pressor response and the activation of prorenin/PRR might be the key source of Ang II in the brain.
Forearm vasodilator response to isoproterenol in premenopausal and postmenopausal women
Authors: Harvey Ronee , Barnes Jill N. , Charkoudian Nisha , Curry Timothy B. , Eisenach John H. , Hart Emma C. J. , Joyner Michael J.
Published: June 2021

Vascular function in older women versus young women may differ based on the β‐adrenergic receptor‐mediated contribution to basal vascular tone. Thus, we tested the hypothesis that forearm blood flow (FBF) responses to the β‐receptor agonist isoproterenol (ISO) are blunted in older, postmenopausal women compared to young, premenopausal women. We used venous occlusion plethysmography to measure FBF in young (26 ± 3 yr; n = 13) and older (61 ± 6 yr; n = 12) women. FBF and mean arterial pressure (MAP) were measured at baseline and during ISO infusion at 1.0, 3.0, 6.0, and 12.0 ng/100 ml tissue/min. The two groups did not differ in body mass index or MAP. Baseline FBF (3.7 ± 1.7 vs. 2.9 ± 1.3 ml/100 ml/min, respectively) and forearm vascular conductance (FVC) (0.04 ± 0.02 vs. 0.03 ± 0.01 AU, respectively) were similar between young and older women. FBF and FVC remained similar between the groups for all ISO doses, including the 12.0 ng/100 ml tissue/min dose of ISO, for which the greatest difference between the groups would be expected; FBF was 10.7 ± 2.3 vs. 9.4 ± 2.1 ml/100 ml/min (young vs. older, respectively) and FVC was 0.11 ± 0.02 vs. 0.10 ± 0.02 AU (young vs. older, respectively). In conclusion, β‐adrenergic receptor‐mediated forearm responses to stimulation are similar between young and older women, suggesting that β‐adrenergic receptor response and contribution to vascular tone are preserved with aging in women. Funded by NIH
Alterations of calcium channels and cell excitability in intracardiac and stellate ganglion neurons from chronic heart failure rats
Authors: Liu Jinxu , Tu Huiyin , Zhang Dongze , Li Yu‐Long
Published: June 2021

Chronic heart failure (CHF) is characterized by increased cardiac sympathetic and decreased cardiac parasympathetic nerve activity. This autonomic imbalance increases the risk of arrhythmias and sudden death in CHF patients. As the final pathways for the autonomic control of the cardiac function, sympathetic postganglionic neurons from the stellate ganglia (SG) release norepinephrine and parasympathetic postganglionic neurons of the intracardiac ganglia (ICG) release acetylcholine upon neuronal excitation. In the present study, we measured the protein expression and current density of Ca2+ channels in the SG and ICG neurons and investigated the role of Ca2+ channels in neuronal excitability and also recorded the cardiac sympathetic nerve activity in sham and coronary artery ligation‐induced CHF rats. Immunofluorescent data showed that L, P/Q, N, and R types of Ca2+ channels were expressed in the SG and ICG neurons, but only protein expression of N‐type Ca2+ channels was decreased in the ICG neurons from CHF rats. Using whole‐cell patch clamp technique, we found that CHF significantly enhanced the Ca2+ currents and cell excitability in SG neurons and reduced the Ca2+ currents and cell excitability in ICG neurons. N‐type Ca2+ channel blocker (1 μM ω‐Conotoxin GVIA) lowered the Ca2+ currents and cell excitability to the same level in SG and ICG neurons from sham and CHF rats. In addition, the cardiac sympathetic nerve activity was augmented in CHF rats. Our results indicate that CHF caused changes of Ca2+ channels and cell excitability in SG and ICG neurons may contribute to the cardiac autonomic imbalance in CHF.
Optogenetic stimulation of the locus coeruleus noradrenergic neurons increase inhibitory neurotransmission to cardiac vagal neurons in the nucleus ambiguus
Authors: Wang Xin , Pinol Ramon A , Mendelowitz David
Published: June 2021

Locus coeruleus (LC) neurons play an essential role in wakefulness, analgesia and autonomic regulation, but the descending projections to caudal medulla that alter autonomic function remains unclear. Cardiac vagal neurons (CVNs) located in nucleus ambiguus control heart rate. Since heart rate changes in awake‐sleep cycle and anesthesia status we hypothesized that these changes may be mediated by LC modulation of CVNs. Mice that express Cre recombinase selectively in tyrosine hydroxylase neurons were crossbred with a channelrhodopsin‐2/EYFP Cre‐dependent mouse strain. CVNs were identified by the retrograde tracer rhodamine. Photostimulation evoked depolarization and induced an average inward current of 150pA in ChR2 LC expressing neurons. Laser stimulation of LC neurons did not alter excitatory neurotransmission to CVNs but evoked a significant increase in postsynaptic inhibitory neurotransmission. The IPSCs frequency in CVNs was facilitated by 73.7±21.8% (p<0.01) upon LC photoexcitation. This increase was blocked by á‐1 receptor inverse agonist, prazosin (3μm), á‐2 receptor antagonist, Yohimbine (5 μm), and â‐1 receptor antagonist, atenolol (100 μm). This LC induced modulation is likely a mechanism for heart rate changes that occur in awake/sleep cycles and during anesthesia mediated by LC induced inhibition of CVNs.
The effects of melatonin combined with exercise on anatomical changes and reorganization in the brain after spinal cord injury
Authors: Park Kanghui , Lee Seunghoon , Hong Yunkyung , Hong Yonggeun
Published: June 2021

We have confirmed that melatonin combined with exercise is effective for functional recovery after spinal cord injury (SCI); however, the mechanisms have not been well described. The purpose of this study was to elucidate the effects of melatonin combined with exercise on recovery of motor function and neural regeneration in the brain after SCI. 8‐week‐old male Sprague‐Dawley (SD, 250–280g) rats were used for contusion SCI surgery. Melatonin (10mg/kg) was administrated injected subcutaneously, twice a day (07:00, 19:00) and exercise (10m/min) was performed on the treadmill for 15 min, twice a day (17:00, 20:00) 6 days per week. There was a significant increased in motor function in SCI+MT+Ex group from 7 to 28 days after injury (p<0.001), also NeuN positive cells were increased in the primary motor cortex of the SCI+MT+Ex compared with the other groups at day 7 after injur (P<0.01, P<0.01, P<0.01). In the pyramid of medulla, SCI+MT+Ex and SCI+MT significantly increased the Oligo2 positive cell compared with SCI at 7 days after injury (P<0.01). These results suggest that synergistic effect of melatonin combined exercise improves motor function after SCI, and induces a positive effect for preventive approaches on neuronal cell death of secondary damage in the cerebral cortex, through the proliferation and differentiation of neuronal cells.
Neurotrophic growth factor response to lower body resistance training in older adults
Authors: Walsh Jeremy James , Scribbans Trisha , Bentley Robert Francis , Kellawan Jeremy Mikhail , Gurd Brendon , Tschakovsky Michael
Published: June 2021

Multimodal cognitive interventions (resistance training + cognitive training; RT + CT) may synergistically improve cognitive function in older adults. RT stimulates the release of neurotrophic growth factors (NGF) which orchestrate structural and functional brain plasticity in the aging brain, while CT increases regional cerebral blood flow. Timing the administration of CT following a bout of RT, when NGF levels are at their highest, may maximize the blood‐borne dose of NGF delivered to active brain tissue. We refer to this as the ‘anabolic window’. Currently, when and how much NGF levels in the blood increase following a bout of RT is poorly understood. The purpose of this work was to characterize the timing and magnitude of insulin like growth factor‐1 (IGF‐1) and brain derived neurotrophic factor (BDNF) levels in venous blood for 2 hours following an acute bout of RT.

Methods

10 older adults (ages 60 – 77) performed 1 hour of lower‐body RT and rested for 2 hours post‐exercise. Blood was taken before and for 2 hours after RT at set time points. This procedure was repeated after 8‐weeks of regular RT.

Results

BDNF increased immediately post‐exercise then returned to resting levels before and after 8 weeks of RT. IGF‐1 levels did not change.

Conclusions

these data are the first to show an increase in BDNF immediately following RT in older adults. This may represent the anabolic window for performing CT. NSERC
Flying before diving: induction of neuroprotection against CNS‐oxygen toxicity (CNS‐OT) seizures while breathing 100% oxygen at 5 atmospheres absolute (ATA)
Authors: Pilla Raffaele , Held Heather E. , Fiorelli Tina , Landon Carol S. , Dean Jay B.
Published: June 2021

In clinical and military procedures, excesses of hyperbaric oxygen (HBO) can cause CNS‐OT. We tested the hypothesis that oxidative preconditioning (PC), using a sub‐lethal dose of HBO, induces neuroprotection against seizures.

99 male rats were implanted with radio‐telemetry transmitters to record diaphragmatic electromyogram, ECG, EEG, body temperature and activity. After 1 week, rats underwent PC for 1hr, every other day for 3 times over 5 days using: a‐b) air at 1ATA (control) or 2ATA; c‐d) 100%O2 at 1 or 2ATA; e) hyperbaric normoxia, f) 2ATA using 90% helium (Heliox) and g) nitrogen (Nitrox); h) normobaric hypoxia (10%O2); and i) hypobaric (0.5ATA) normoxia and j) hypobaric hypoxia (10%O2 at 0.5ATA). 24h after last PC treatment, rats were placed in a hyperbaric chamber and exposed to 5ATA O2, until CNS‐OT. Data were collected and latency to seizure (LS) determined.

Results showed:

1) hyperoxic PC did not provide neuroprotection against seizure; 2) inert gases PC (2ATA Nitrox/Heliox) decreased LS by 57/78%; 3) hypobaric PC significantly extended LS by 56% and 38%; and 4) hippocampi western blot suggested that hyperbaric pressure per se accelerates the CNS‐OT via downregulation of Early Growth Response‐1, involved in neuroprotection against oxidative stress.

Exposing rats to intermittent flights at~16.000 feet before diving at 5ATA O2 decreases CNS‐OT risks, whereas air dives exacerbate the risk of CNS‐OT. (ONR)
Gene expression of appetite regulatory factors and glucose transporters in hypothalamus, pectoralis major, abdominal fat and liver after insulin injection in chicken lines selected for high or low body weight
Authors: Zhang Wei , Sumners Lindsay , Siegel Paul , Cline Mark , Gilbert Elizabeth
Published: June 2021

Chickens selected for high (HWS) and low (LWS) body weight for 55 generations differ by 12‐fold in body weight at 8 weeks old and display differences in appetite and body composition. We hypothesized that there are differences between the lines in appetite regulatory factor and glucose transporter (GLUT) mRNA that are accentuated by exogenous insulin. Female 87–92 d old HWS and LWS were injected with insulin or vehicle and at 60 min mRNA was measured in hypothalamus, pectoralis major skeletal muscle, abdominal fat and liver. In hypothalamus, neuropeptide Y (NPY) mRNA was lower (P < 0.05) in HWS than LWS, while Forkhead box protein O1 (FOXO1) was greater (P < 0.05) in HWS. Abundance of GLUT1 and GLUT9 mRNA were greater (P < 0.05) in HWS than LWS in all tissues. The GLUT1 mRNA was greater in liver (P < 0.05) than in hypothalamus or fat. Although GLUT3 mRNA was not detected in hypothalamus or muscle, expression was greater (P < 0.05) in fat than liver. For GLUT2, GLUT8 and GLUT9, mRNA was lowest in fat (P < 0.05). In liver, insulin was associated with reduced (P < 0.05) GLUT3 mRNA as compared with vehicle. These data show differential expression of NPY, FOXO1 and glucose transporters in HWS and LWS.
P2X7 Receptors Mediate Hormone Release in nerve terminals of the Neurohypophysis (NH)
Authors: Cuadra Adolfo Ernesto , Custer Edward , Bosworth Elizabeth , Raizada Mohan K. , Lemos Jose R
Published: June 2021

P2X7R expression in neurons is controversial; however, studies suggest that it may play a role in hormone release in the hypothalamic NH system (HNS). We examined this in isolated NH terminals (NHT). Application of the P2X7R agonist, BzATP, to NHT resulted in increased levels of sustained AVP release, compared with ATP. Further experiments showed that the sustained release was inhibited by the P2X7R antagonist brilliant blue‐G (BBG). Confocal microscopy showed P2X7R‐immunoreactivity (IR) in slice sections and in isolated terminals of the NH. However, the cell bodies in the hypothalamus that project to the NH were devoid of P2X7R‐IR. Patch clamp studies revealed that subpopulations of NHT displayed differential sensitivity to ATP in evoking ion currents. One population showed a pEC50 of 5.2 ± 0.1 M and the second showed a pEC50 of 3.9 ± 0.2 M (T‐test, p< 0.0002, n=5–6), the latter being consistent with P2X7R. In low divalent cationic media the BzATP, gave a pEC50 of 5.6 ± 0.15 and pretreatment with 10 and 300 nM BBG inhibited currents by 51.8 ± 13.1 and 12.3 ± 4.2 percent of control, respectively (1 way ANOVA, P < 0.0001, n=5). The poor sensitivity to ATP and the marked inhibition of BzATP‐mediated current by BBG are hallmarks of P2X7R function. These results lead us to conclude that P2X7R is present in NHT and play a functional role in hormone release in the HNS. (Study supported by NIH grant NS29470 to JRL)
Intermittent Hypoxia‐induced hERG degradation involves ROS Activated Calpains
Authors: Wang Ning , Khan Shakil A , Ahmmed Gias A , Prabhakar Nanduri R , Nanduri Jayasri
Published: June 2021

Human ether‐go‐go‐related gene (hERG) protein, the alpha subunit of voltage gated potassium channel, mediates a variety of cellular functions including the cell excitability. Intermittent hypoxia (IH) is a hallmark feature of recurrent apnea. The present study examined the effects of IH on hERG protein expression in neuroblastoma (SH‐SY5Y) cells. IH decreased hERG current density by ~60%. The decreased current density was associated with stimulus‐dependent down‐regulation of hERG protein, which could not be rescued with proteasomal and lysosomal inhibitors. IH increased calpain activity and ALLM a calpain protease inhibitor prevented IH‐evoked degradation of hERG protein. Further analysis showed that IH‐induced calpain activation involves elevation of [Ca2+]I by reactive oxygen species. Similar downregulation of hERG was also observed in adrenal chromaffin cells from IH exposed neonatal rats, and anti‐oxidant treatment abolished this effect. The IH evoked hERG degradation might contribute to increased cell excitability and augmented catecholamine release from neonatal adrenal chrmoaffin cells.
miRNAs Regulate Skeletal Muscle Development via Down‐regulation of Myosin Heavy Chain
Authors: Chen Xin , Zhu Minsheng
Published: June 2021

MicroRNAs (miRNAs) are a class of small non‐coding RNAs which suppress gene expression through posttranscriptional binding 3′‐untranslated region (UTR). Some miRNAs have been found mediate muscle differentiation via repressing the transcription factor or signal molecule in myogenesis. However, the functional role and molecular mechanism by which miRNAs regulate muscle differentiation via contractile protein are not understood. Myosin II is a family of actin‐binding proteins and it¡s the important structure protein for muscle contraction. Using luciferase reporter assay and western blot, we revealed miR‐23a directly inhibited muscle myosin heavy chain (MyHC) expression, while miR‐199a‐5p depressed level of non‐muscle myosin heavy chain IIA (NMHC IIA) protein. Augmentation or reduction of the two miRNAs either inhibited or promoted myogenic differentiation in C2C12 cell line, respectively. Furthermore, knockdown of muscle MyHC or NMHC IIA with siRNA also impaired muscle differentiation. These data suggest a novel microRNA‐myosin mechanism during muscle differentiation.
A comparative study of the effect of hindlimb unloading (HLS) on male and female Sprague Dawley rats
Authors: Chowdhury Parimal , Long Ashley , Harris Gabriella , Soulsby Michael E. , Dobretsov Maxim
Published: June 2021

The aim of this study is to test the gender‐specific effects in a hind‐limb suspended rat model of microgravity.

Methods

Adult male and female rats were suspended using tail and pelvic suspension techniques for two weeks. A control non‐suspended groups of male and female rats were also used. Body weights, food and water consumption, soleus muscle weights and hind limb pressure‐pain thresholds (PPT) were measured.

Results

For a given gender, none of measured characteristics differed between tail‐ and pelvic suspended rats. Therefore the results with different suspension techniques were pooled together for further analysis. Food and water intakes were not different between genders and between control and suspended groups. Compared to control males, control female rats had the same PPT value, but weighted less, had lower soleus muscle mass and gained weight slower. HLS nearly canceled the weight gain and resulted in the development of pressure hyperalgesia in both gender groups. However, HLS‐associated soleus muscle atrophy was detected in male rats only. Furthermore, in a sub‐group of experiments in which both PPT and plasma insulin were measured, HLS was found to decrease insulin concentrations and affect pain thresholds in male rats only.

Conclusions

Overall HLS appear to influence growth, postural muscles and deep pressure sensitivity of males and it is stronger and consistent than those of female rats
Bradykinin 2 receptor blockade attenuates the responses of group III afferents to contraction in decerebrate cats
Authors: Leal Anna Katherine , Stone Audrey , Yamauchi Katsuya , Kaufman Marc
Published: June 2021

Exercise‐induced stimulation of group III and IV muscle afferents raises blood pressure, heart rate, and ventilation. Bradykinin has been shown to stimulate and sensitize group III and IV afferents, but the effect of bradykinin 2 (B2) receptor blockade on the responses of group III afferents during exercise was unknown. Using decerebrate cats, we examined the effect of carotid arterial injections of the B2 receptor antagonist, HOE‐140 (40 μg/kg) on the responses of group III afferents (conduction velocity range: 2.8 – 26.2 m/s; mean: 16.1 ± 2.3 m/s) to both 45 second intermittent and static contraction. Before HOE‐140, static contraction increased the impulse activity of ten group III afferents from 0.3 ± 0.1 imp/s at rest to 6.7 ± 3.0 imp/s during contraction (p<0.05). After HOE‐140, impulse activity went from 0.7 ± 0.3 imp/s at rest to 2.9 ± 1.2 imp/s during contraction (p>;0.05). There was a similar trend during intermittent contraction. Before HOE‐140, intermittent contraction increased the activity of seven group III afferents from 0.2 ± 0.04 imp/s at rest to 9.1 ± 5.2 imp/s during contraction (p>;0.05). After HOE‐140, impulse activity increased from 0.5 ± 0.2 imp/s at rest to 4.0 ± 2.3 imp/s during contraction (p>;0.05). Our data suggests B2 receptors contribute to group III afferent impulse activity during exercise.
Limb Movement‐Induced Central and Peripheral Hemodynamics in Heart Failure: The Role of Afferent Feedback
Authors: Ives Stephen , Amann Markus , Venturelli Massimo , Groot H. Jonathan , Witman Melissa A.H. , Morgan David E. , Wray D. Walter , Richardson Russell S.
Published: June 2021

The mechanoreflex has previously been reported to be sensitized in patients with heart failure (HF). Therefore, this study sought to better understand the role of afferent signaling on movement‐induced changes in central and peripheral hemodynamics.

METHODS

In 10 NYHA class II HF patients we performed measurements of femoral blood flow (FBF), mean arterial pressure (MAP), femoral vascular conductance (FVC), heart rate (HR), stroke volume (SV), and cardiac output (CO) at baseline and during 3 min of passive knee extension. This was repeated after intrathecal injection of the μ‐opioid antagonist, fentanyl, to pharmacologically blunt afferent feedback.

RESULTS

Fentanyl had no effect on baseline measures, but increased (p < 0.05, control vs. fentanyl) the movement‐induced FBF (493±155 vs. 804±198 ΔmL/min), and FVC (4.7±2 vs. 8.5±3 ΔmL/min/mmHg), while NE spillover (42 ± 8 vs. 27 ± 8 Δpg/L), and retrograde FBF (371±115 vs. 260±68 ΔmL/min) tended to be reduced (p <0.10). There was no effect of fentanyl on MAP, HR, SV, or CO in response to passive movement.

CONCLUSION

As movement‐induced central hemodynamics were unchanged while peripheral hemodynamics were augmented with fentanyl blockade, this study suggests that the heightened mechanoreflex, associated with HF, plays a selective role likely affecting peripheral sympathetic vasoconstriction and subsequently exercise tolerance. Support: NIH P01HL1091830.
Contributions of central command and exercise pressor reflex on myocardial perfusion during exercise in rats
Authors: Tsuchimochi Hirotsugu , Sonobe Takashi , Inagaki Tadakatsu , Shirai Mikiyasu
Published: June 2021

To examine how myocardial perfusion is controlled by a feedforward (termed “central command”) and feedback (exercise pressor reflex) neural mechanisms during exercise, left ventricular (LV) myocardial perfusion was measured during either electrical stimulation of subthalamic locomotor region (SLR), the electrical or chemical stimulation of which is known to elicit coordinated locomotion and concomitant cardiorespiratory responses, or electrical stimulation of tibial nerve in either anesthetized or decerebrated rats. Changes in LV myocardial perfusion and muscle blood flow to the right gastrocnemius muscle were measured by a Laser‐Doppler flowmetry. Arterial pressure and heart rate (HR) were also measured. Stimulation of SLR rapidly increased both LV myocardial perfusion and mean arterial pressure (MAP) in a stimulus intensity‐dependent manner. Stimulation of SLR also increased gastrocnemius muscle blood flow gradually throughout the stimulus period. Beta‐adrenergic blockade attenuated the rapid increase in myocardial perfusion evoked by SLR stimulation. Tibial nerve stimulation also rapidly increased LV myocardial perfusion and MAP. These results suggest that both feedforward and feedback control of the myocardial perfusion play an important role in meeting myocardial demands for adequate blood supply.
Role of Calcium‐Sensing Receptor in Esophageal Epithelium
Authors: Abdulnour‐Nakhoul Solange , Brown Karen L , Nakhoul Nazih
Published: June 2021

The calcium sensing receptor (CaSR), a G‐protein coupled receptor, senses changes in extracellular Ca+2 and mediates its effects on cellular functions. In cells not directly involved in Ca+2 homeostasis, CaSR plays a role in many cellular processes including differentiation, proliferation and secretion. We investigated the role of CaSR in modulating esophageal barrier function. We used immunohistochemistry, Western blotting and PCR to identify CaSR and cell‐cell adhesion molecules in pig esophageal stratified squamous epithelium (SSE). In primary cultures of SSE, we investigated the effect of CaSR agonist (cinacalcet, CCT) and antagonist, calhex, on cell growth and cell‐cell junction formation. We used MTT assay to monitor proliferation under different conditions that affect CaSR activity. SSE cells exposed to CCT (2.5μM, 24 hours), changed from polygonal to spindle‐shaped cells, a transformation proportional to CCT concentration. Staining with E‐cadherin and β‐catenin antibodies, showed altered distribution of these proteins in CCT treated cells. Our results indicate: 1) CaSR is expressed in supra‐basal cells of esophageal epithelium but not in differentiated apical layers; 2) CaSR activation decreases the rate of growth of SSE in culture; 3) CaSR activity alters expression and distribution of intercellular adhesion proteins and could modulate barrier properties of esophageal epithelium. Support: VA Merit grant and Tulane Enhancement Funds
Divalent cation‐dependent 3H‐L‐leucine transport in lobster intestine
Authors: Abdel‐Malak Rania , Ahearn Gregory A
Published: June 2021

The crustacean intestine displays a diversity of transport mechanisms for the transport of dietary substances, including metal ions and amino acids. The intestine of the American lobster Homarus americanus was isolated and perfused in vitro with a physiological saline. Transmural 20 μM mucosa to serosa (MS) 3H‐L‐leucine fluxes across the intestine were a hyperbolic function of luminal Ca concentration. Ca‐dependent 3H‐L‐leucine transport exhibited low Km (high affinity) and low Jmax compared to kinetic constants when both Ca and Zn were simultaneously present, and under these conditions amino acid transport Jmax was doubled. MS flux of 3H‐L‐leucine was a hyperbolic function of L‐leucine and was stimulated by the addition of 20 μM luminal Mn. In the presence of Mn alone, 3H‐L‐leucine transport showed an increase in Jmax, but no effect on L‐leucine influx Km. With the addition of Ca, the Jmax decreased, but Ca had no effect on L‐leucine influx Km, suggesting that Ca may be acting as an allosteric inhibitor of metal‐stimulated amino acid transport. In other experiments calcium was a non‐competitive inhibitor of manganese activation of 3H‐L‐leucine transport, apparently binding to a site other than the metal binding site. Ongoing experiments are researching the nature of Ca inhibition of metal stimulated amino acid transport.

Agriculture and Food Research Initiative Competitive USDA Grant no. 2010–65206‐20617.
Physician assistant anatomy education: does prior anatomy experience predict performance?
Authors: Terhune Claire E. , Melcher Betsy Q. , Taylor Andrea B.
Published: June 2021

Anatomy education in the Duke Physician Assistant (PA) program has been historically prosection‐based and heavily clinically oriented. Based on alumni feedback, in 2009 this course was transformed into a foundational anatomy course with studentperformed dissection. As part of this modification, data were collected describing incoming students’ prior anatomy experience. These data were used to combine experienced and non‐experienced students into lab groups to optimize group learning and maximize lab resource use. At present we have three years of data for students in this new anatomy curriculum. These data also allow us to evaluate whether past experience predicts course performance and/or performance on the Physician Assistant Clinical Knowledge Rating and Assessment Tool (PACKRAT). Relationships between experience (e.g., number of semesters, dissection vs. prosection) and performance in the course and on the PACKRAT were examined using linear regression analysis and ANOVA. Contrary to our initial expectations, results indicate that prior anatomy experience is not a significant (P>;0.05) predictor of anatomy performance or PACKRAT scores. However, anatomy performance was significantly related to performance on the PACKRAT. We suggest there may multiple potential confounding factors that could be related to these outcomes, which will be incorporated into future analyses.

Grant Funding Source: n/a

Submitted to PAEA 5/25/2012 (bqmelcher)
E‐testing in anatomical teaching
Authors: Streicher Johannes
Published: June 2021

In modern curricula on the one hand the time frame for anatomy teaching is progressively reduced, and on the other hand the successful integration with clinical applications must be confirmed. Taking into account the high number of 720 students/year at the Medical University of Vienna, and the parallel education of second‐ and third‐year students, more than 4000 individual assessments have to be plannend, held and evaluated each term. For this reasons and in order to avoid biases by different examiners, two e‐testing modalities were implemented. In settings where students have no access to individual PCs (e.g. in the dissection halls), auditory response systems were employed, whereas assessments held in a PC‐room used the moodle Course Management System. In both cases most of the questions were image‐based multiple choice questions. Due to the information about their grades immediately after the assessment, the equal level of difficulty for all students, and the chance to instantly clear up misunderstandings, the students’ compliance was continously encouraging during the past four years. Both types are technically mature and – once an adequate question library is established – cost‐ and time‐saving. Immediate statistical analyses of answer patterns provide teachers with valuable, objective feedback and enable teachers to rapidly accommodate contents and objectives to the students actual level of competence.
Varying Effects of Prosection and Dissection Anatomy Curriculums on Confidence in the Clinical Setting
Authors: Young Stephanie , Guzman Carlos R. , Wimmers Paul F. , Byus Craig V. , Wisco Jonathan J.
Published: June 2021

The David Geffen School of Medicine at UCLA has a prosectionbased anatomy course (PRO), and the Thomas Haider Program in Biomedical Sciences at UC Riverside has a dissection‐based anatomy course (DIS). Students from both programs converge for 3rd and 4th year clinical clerkships. We conducted the first of several annual surveys in 2010 of medical students in both PRO and DIS programs (59% response rate out of 185 surveys). 80% of DIS students and 62% of PRO students said their anatomy curriculum increased confidence in physical exam skills. However, 40% of DIS students and 86% of PRO students said their anatomy curriculum helped prepare them for the USMLE Board Exams. When asked if the anatomy curriculum helped prepare students for 3rd and 4th year clinical rotations, 96% and 91% of PRO students responded positively, respectively, while 100% of DIS students responded positively for both clerkship years. Both cohorts regarded the anatomy curriculums highly, but expressed that prosection allowed for learning of semantics while dissection was a better preamble for surgical training. This study was approved with an IRB exempt protocol.

Grant Funding Source: AAA
Establishing the validity and reliability of computer‐based simulation for cerebral angiography using the ANGIO Mentor Express
Authors: Nguyen Ngan , Eagleson Roy , Deribaupierre Sandrine
Published: June 2021

Computer‐based simulation (CBS) is increasingly used in medical education, but evidence for its efficacy has been limited and inconsistent. Deficiencies and heterogeneity in validity and reliability study methodologies have been cited as major limitations to drawing conclusions regarding the effectiveness of CBS training. Since untested assessment methodologies can lead to reporting inaccurate performance data, it is crucial to validate CBS systems and their assessment instruments before examining the effectiveness of CBS training. This study establishes the validity (face, content and construct) and reliability (test‐retest) of a CBS system (ANGIO Mentor Express) for left and right cerebral angiography (CA). Face and content validities were established by asking experienced catheter‐based physicians to judge the overall realism of the simulated CA procedures and evaluate the appropriateness of the assessment instruments. Construct validity was assessed by comparing the performance of an expert cohort (experienced physicians) to a novice cohort (medical students and residents) of subjects. Test‐retest reliability was established by correlating performances on the simulated left CA with the right CA. Confirmation of the validity and reliability of the CBS system and its assessment instruments assures the integrity of future studies evaluating the efficacy of CBS training for CA.

Grant Funding Source: None
Use of an audience response (ARS) in an interactive histology laboratory
Authors: Feasel Patrick C , Xanthos Evan , Borges Nicole , Ream Larry
Published: June 2021

The realization that medical students may fail to integrate normal histology, histopathology and resulting clinical manifestations with the advent of virtual‐histology has led us to identify the need to introduce methods that promote active learning in an interactive laboratory setting. Little research has been done on the potential value of introducing interactive learning methods in the histology laboratory using an audience response system (ARS). Many studies have shown that the use of an ARS can facilitate interactivity in large group settings and promote active learning. Therefore, in an effort to maximize the effectiveness of the combined laboratory‐based use of microscopes and glass slides with computer‐based instructional technologies, we plan to introduce a post‐laboratory exercise that utilizes an ARS. Our intended purpose of using the ARS will be to promote individual students’ responsibility for their own learning and allowing for practical application of learned material, and to become less reliant on memorization and overly focused on short‐term objectives.
Medical student perceptions of a radiology tutorial in anatomy education
Authors: Hoover Alison , Smith Lindsay , Brian Pamela , Lazarus Michelle
Published: June 2021

Students’ perceptions of their education influence their learning. We hypothesized that medical students would value an interactive radiology computer tutorial and that this tutorial would benefit their anatomy education. Tutorials were developed using radiologic images from two different anatomic regions and tested by first year medical students through qualitative surveys. Students overwhelmingly agreed (100%, n=22) that the tutorial was beneficial in understanding anatomic concepts and that they would recommend this tutorial to future students. All students agreed that early radiology exposure is beneficial to their medical education, and 95% agreed it would benefit their careers. In citing the source of the student's radiology knowledge, 73% of students credited the tutorial versus 38% of students who credited radiology lectures. A majority of students (86%) agreed that multiple types of imaging techniques were beneficial in learning anatomy (CT, plain film, etc.). Most students (91%) felt that they would use the tutorial later in medical training, while all agreed they would use it at least once a week during their formal anatomy course. Given the trend in decreasing anatomy instruction and vertical integration, our findings suggest that a self‐guided radiology tutorial is a promising solution for learning anatomy in a flexible environment throughout the entirety of medical school.

Grant Funding Source: n/a
A Learning Strategy for the Muscles and Fascial Planes of the head and Neck
Authors: Sanders Luke E , Morton David A. , Nielsen Mark T.
Published: June 2021

Learning the relationships of the muscles and fascial planes of the head and neck is a difficult task for medical students, which is further complicated by the plethora of terminology that anatomists use to define the fascial sheets and compartments of these regions. This complexity in the musculofascial anatomy arises at the interfaced formed between pharyngeal arch anatomy and somitic body wall anatomy at the head‐trunk junction. Migrating muscles from cervical and occipital somites and from the pharyngeal arches establish a number of muscular and fascial layers in the head and neck that become a challenge to understand. However, a simple pattern of four musculofascial planes emerges during development as the muscles migrate from their sites of origin in the pharyngeal arches and cervical and occipital somites to their definitive positions in the head and neck. This pattern introduces a clear logic to the layers of muscles in the head and neck, while also clarifying the fascial layers and compartments of the region. Understanding the fascial planes and compartments is of significant clinical interest regarding the spread of inflammation and disease.

Grant Funding Source: N/A
Structural characterization of the major DNA‐DNA cross‐link formed by epichlorohydrin
Authors: Sirka Katarina , Millard Julie T.
Published: June 2021

Epichlorohydrin (ECH) is a bifunctional alkylating agent used extensively as an industrial intermediate, a laboratory reagent, and as an insecticide. ECH is classified as a probable human carcinogen since occupational exposure to ECH has been linked to elevated cancer risk for industrial workers. The ability of ECH to form DNA‐DNA cross‐links by consecutive alkylation of two nucleobases within a DNA molecule leads to the possibility of both cytotoxic and promutagenic properties. Previous work provided evidence for the formation of interstrand ECH‐DEB cross‐links involving deoxyguanosine residues within 5′‐GC and 5′‐GGC sites. However, the covalent structure of ECH‐induced DNA cross‐links has not been determined. In the present study, ECH interstrand cross‐links within synthetic DNA oligonucleotides containing a central 5′‐GGC site will be purified and characterized via electrospray ionization mass spectroscopy. This project was supported by grants from the National Center for Research Resources (5P20RR016463–12) and the National Institute of General Medical Sciences (8 P20 GM103423–12) from the National Institutes of Health.
Crystallographic snapshots of metalloenzyme complexes involved in biological carbon dioxide sequestration
Authors: Drennan Catherine L. , Kung Yan , Ando Nozomi , Doukov Tzanko , Bender Gunes , Ragsdale Stephen W.
Published: June 2021

The Wood‐Ljungdahl pathway in acetogenic bacteria involves nine enzymes working together to convert two molecules of carbon dioxide and coenzyme A into acetyl‐CoA, which can subsequently be assimilated as cell carbon or converted to other products such as acetate or ethanol or butanol, depending on the acetogenic strain. Harnessing the ability of acetogens to produce commercially useful multicarbon compounds from carbon dioxide has long been of interest to the biotechnology community. The Drennan laboratory, in collaboration with the Ragsdale group, has solved crystal structures of metalloenzymes of this pathway, both alone and in complex, in order to gain mechanistic insight into the chemistry involved in these remarkable transformations, as well as to understand the molecular movements that allow these enzymes to act in concert. From structures of carbon monoxide dehydrogenase in complex with acetyl‐CoA synthase, we have obtained molecular pictures indicating how a unique Ni‐Fe‐S cluster can carry out reduction of carbon dioxide. From structures of our methyltransferase complexes, we have visualized the molecular movements that must occur for one‐carbon units to be passed from enzyme to enzyme.
The QA repeat domain of TCERG1 plays a role in inhibiting C/EBPα and in its ability to undergo redistribution in the nucleus.
Authors: Roesler William J , Miller Nick , Timchenko Nikolaj , Moazed Banafsheh
Published: June 2021

TCERG1 is a nuclear protein that functions in transcriptional elongation and the integration of transcription and mRNA processing. Our laboratory has shown that it also inhibits the various biochemical activities of C/EBPα, through a mechanism that involves its redistribution from its normal nuclear speckle compartment to pericentromeric regions which is where C/EBPα resides. We undertook structure/function analysis of TCERG1 to identify the domain that mediates this inhibitory effect. Previous studies from our lab indicated that the domain resided in the amino terminus portion of the protein, which contains three WW domains as well as a domain consisting of 38 QA repeats. While mutation of one or more of the WW domains had no effect on the inhibitory activity of TCERG1 or its ability to be redistributed, deletion of the QA domain abrogated both of these activities. Reducing the number of QA repeats to 20 had no measurable effect, while a further reduction to 10 repeats eliminated these activities of TCERG1. This study not only suggests a role for this unique domain, but also indicates that there is a minimal number of repeats necessary for it to function.
MeCP2_e2 does not contribute Rett syndrome phenotypes but essential for placenta
Authors: ITOH MASAYUKI , Tahimic Candice G.T. , Goto Yu‐ichi , Kurimasa Akihiro
Published: June 2021

Methyl CpG‐binding protein 2 (MeCP2) mutations are implicated in Rett syndrome (RTT). Two MeCP2 isoforms have been reported as MeCP2_e2 and MeCP2_e1. Their expression levels vary among tissues, with MeCP2_e1 in adult brain and with MeCP2_e2 in placenta, liver, and skeletal muscle.

We performed specific disruption of the MeCP2_e2 and examined the consequences of selective loss of MeCP2_e2 function in vivo. MeCP2_e2 null mice showed no RTT‐associated neurological phenotypes but confers a survival disadvantage. In addition, selective loss of MeCP2_e2 resulted in placenta defects and upregulation of peg‐1 as determined by the parental origin of the mutant allele. It is indicated a novel role for MeCP2 in normal placenta development and illustrate how X chromosome inactivation in extraembryonic tissues confers a survival disadvantage for carriers of a mutant MeCP2_e2 allele. Moreover, our findings provide an explanation for the absence of reports on MeCP2_e2‐specific exon 2 mutations in RTT. MeCP2_e2 mutations in humans may result in a phenotype that evades a diagnosis of RTT.
Primary and alternative S18 ribosomal protein in Mycobacterium tuberculosis
Authors: Prisic Sladjana , Husson Robert N.
Published: June 2021

Protein synthesis is a tightly regulated process in every living organism and is likely to be a point of control in Mycobacterium tuberculosis (Mtb) during the course of infection. Mtb has four alternative ribosomal proteins organized in a single operon, whose function is not known, but may play an important role in translation regulation. Intriguingly, one of these proteins, S18–2, and its primary ribosomal protein paralog, S18–1, are both phosphorylated in vivo. Measured by their ability to bind another protein in a course of a ribosome biogenesis, their activities are not affected by phosphorylation, but it changes their stability in vitro. This possibly helps the exchange in response to zinc concentration, which regulates (represses) only expression of S18–2. In addition, presence of zinc had a profound and opposed effect on in vitro activities of S18–1 and S18–2 proteins, i.e., zinc activates S18–1 and inhibits S18–2. These data together suggest a model in which S18–1 is able to be a dominant ribosome constituent in high zinc conditions, but that it may be replaced by S18–2 when zinc is deficient. Therefore, S18–2 and other alternative ribosomal proteins may play a role in Mtb viability in necrotizing granulomas, which are thought to be a low zinc environment and are important for TB transmission.
Failure of autophagy induction makes multidrug resistant cells vulnerable to BH3‐mimetic gossypol
Authors: Ahn Jun‐Ho , Jang Gun‐Hee , Lee Michael
Published: June 2021

Developing potential therapeutic strategies to overcome multidrug resistance (MDR) is a major challenge in cancer research. Here, we explored the use of gossypol as a chemotherapeutic agent that is effective against both v‐Ha‐ras‐transformed NIH 3T3 cells (Ras‐NIH 3T3) and their mdr equivalents (Ras‐NIH 3T3/Mdr). We found that Ras‐NIH 3T3/Mdr cells, which exhibit a strong cross‐resistance to many chemotherapeutic agents, exhibited essentially no resistance to gossypol. This study also revealed that gossypol significantly decreased mdr‐1 and mdr‐3 mRNA expression in Ras‐NIH 3T3/Mdr cells. However, the rhodamine assay indicated that P‐glycoprotein is not likely to be a major mechanism of the chemotherapy sensitization by gossypol. While both cell lines can be growth arrested, cell death is apoptotic in the Ras‐NIH 3T3 cells but mixed necrotic and apoptotic in the Ras‐NIH 3T3/Mdr cells. More notably, gossypol preferentially induced autophagy in Ras‐NIH 3T3 cells but not in Ras‐NIH 3T3/Mdr cells, suggesting that defective autophagy makes MDR cells vulnerable to gossypol despite being resistant to G2/M block. Taken together, these results suggest that gossypol could be an effective anticancer agent to treat malignancies that are resistant to conventional therapies, either used alone or in association with other autophagy‐blocking treatments.
The role of enhanced autophagy in acquired resistance to new B‐Raf inhibitor UI‐152
Authors: Jang Gun‐Hee , Ahn Jun‐Ho , Lee Michael
Published: June 2021

B‐Raf mutant melanomas have limited duration of clinical response with selective B‐Raf inhibitors because of rapidly acquired resistance. To investigate if chronic B‐Raf inhibition could lead to acquired drug resistance, we used chronic selection to establish B‐Raf(V600E) melanoma clones with acquired resistance to new B‐Raf inhibitor UI‐152. The A375P cells were highly sensitive to B‐Raf inhibition by UI‐152 (IC50 ¡Ü 0.5 ¦ÌM), whereas A375P/Mdr cells that had been chronically treated with UI‐152 required higher doses of the drug for partial growth inhibition (IC50 >; 20 ¦ÌM). The selected clones did not show re‐activation of the MAPK pathway, indicating that input from alternative, MAPK‐independent pathways induce B‐Raf inhibitor resistance mechanisms. Flow cytometric analysis showed that UI‐152 treatment induced G0/G1 cell‐cycle arrest and decreased the percentage of cells in the S and G2/M phases. Few cells with GFP‐LC3 punctate dots were detected in A375 cells treated with UI‐152. Most surprisingly, the fraction of A375P/Mdr cells that underwent autophagy was significantly increased after treatment with UI‐152, suggesting that functional autophagy in response to UI‐152 may lead to cell survival. Vice versa, too little apoptosis was observed in A375P/Mdr cells. Taken together, our results suggest that the acquisition of resistance may be multi‐factorial and mediated by pathways other than the MAPK.