bluuebunny/crossref_metadata_2025 huggingface dataset:
Thiol reducing agents provide protection and rescue for gastric glands following monochloramine‐induced injury
Authors: Tai Kaniza , Kohler Jonathan E. , Blass Amy L. , Liu Jingjing , Mathew Jeff , Soybel David I.
Published: June 2021

Aim: Monochloramine (NH2Cl) is a thiol‐directed oxidant produced in inflamed, H. pylori‐infested gastric mucosa. We tested the hypothesis that NH2Cl‐induced death of isolated gastric glands can be reversed by thiol reducing agents such as dithiothreitol (DTT) or Lipoic Acid (LA) but not oxidant scavengers such as Vitamin C (VitC).

Methods: Isolated rabbit gastric glands were incubated with Ringers solutions containing NH2Cl for 1 hour, then Ringers solution for 2 hours. Cell viability was monitored, using calcein‐AM uptake and conversion to fluorescent calcein, during exposure to NH2Cl. In some studies, DTT, LA or VitC were present throughout the entire 3 hr experiment; in others, they were present only following the 1 hr exposure to NH2Cl.

Results: Exposure to NH2Cl resulted in significant (p<0.05) and dose‐dependent decreases in cell viability. Both DTT and Vitamin C were significantly protective when co‐administered with NH2Cl. Only DTT provided partial reversal of injury when given after NH2Cl. No protection was observed in response to LA, given concurrently or after exposure to NH2Cl.

Conclusion: Some, but not all, thiol reducing agents protect gastric glands from NH2Cl‐induced injury by concurrent administration and can arrest and reverse injury after exposure to the oxidant. Generic oxidant scavengers can prevent, but do not reverse, injury. Funding: APS Summer Research Fellowship, NIH R01DK069929‐03
Regulation of Cl−/HCO3− exchanger SLC26A3 by intracellular pH
Authors: Hayashi Hisayoshi
Published: June 2021

Electroneutral NaCl absorption in the colon is mediated by functional coupling of the Na+/H+ exchanger and Cl−/HCO3− exchanger. SLC26A3 is highly expressed in the colon, and mutations in the SLC26A3 cause congenital chloride diarrhea. It is proposed that the coupling of these exchangers may occur as a result of indirect linkage by changes of intracellular pH (pHi). We therefore investigate whether SLC26A3 is regulated by intracellular pH. We generated an HA epitope‐tagged human SLC26A3 construct and expressed in CHO cells. Transport activities were measured with a chloride‐sensitive dye, MEQ. pHi was clamped at a range of value from 6.0 to 7.4 with varying K+ concentrations of medium that contained the K+/H+ exchanging ionophore, nigericin. Since SLC26A3 transports nitrate, we monitored changes of intracellular Cl− as chloride exchanges for nitrate and also as chloride exchanges for bicarbonate at varying pHi. Whereas Cl−/HCO3− exchange was steeply inhibited by acidic pHi, Cl−/NO3− exchange was essentially insensitive to pHi. We also characterized pHi dependency with the entire C‐terminal truncation mutant. Removal of C‐terminal decreased activity, but did not noticeably affect the inhibition by intracellular acidification. These results suggest that cytoplasmic domain of C‐terminus does not contribute to regulation of SLC26A3 by pHi.
The conductance properties of Asp555 in the electrogenic Na/HCO3 cotransporter SLC4A4/NBCe1
Authors: Kim Eunjin , Choi Inyeong
Published: June 2021

The electrogenic Na/HCO3 cotransporter SLC4A4 (NBCe1) is important for HCO−3 reabsorption in the proximal tubules of the kidney. By sequence comparison between electrogenic transporters and electroneutral transporters, we previously identified Asp555 in NBCe1 as a residue that affects electrogenic HCO−3 current. In this study, we further characterized the HCO−3 conductance of the mutant transporter D555E (substitution of Asp555 with a Glu) using two‐electrode voltage clamp. In oocytes expressing D555E, the application of CO2/HCO−3 caused a negative shift of the reversal potential for HCO−3 current. However, the current progressively decreased at positive voltages that otherwise should be favorable for HCO−3 conductance. The current was larger at negative voltages. No inward current was produced at low [Na+]o that should reverse the direction of Na+/HCO−3 movement as in wild type NBCe1. In the steady state condition, which was achieved by superfusing oocytes with Na+‐free CO2/HCO−3 for 15 min, the application of a series of [Na+]o produced HCO−3 currents. The half maximum effective concentration of external Na+ was 2 mM for D555E, significantly smaller than 12 mM for wild type NBCe1. The substitution of Asp555 with an Asn (D555N) produced a progressive decrease in HCO−3 current at positive voltages and a negative shift of the reversal potential for the current. Reciprocal substitutions of Glu742 in the electroneutral Na/HCO3 cotransporter SLC4A7 (NBCn1) with Asp or Gln failed to produce electrogenic HCO−3 current. We conclude that the steric properties near Asp555 in NBCe1 likely contribute to ion translocation of the transporter.
Effect of basolateral CO2 on the luminal ANG II sensitivity of HCO3 reabsorption by rabbit S2 proximal tubules
Authors: Zhou Yuehan , Boron Walter F
Published: June 2021

Our previous work showed that: (1) Using out‐of‐equilibrium (OOE) solutions to raise basolateral (BL) [CO2] from 0% to 20% causes graded increases of HCO3 reabsorption (JHCO3) by the proximal tubule (PT). (2) Adding ANG II to the lumen (L) produces dose‐dependent effects on JHCO3. (3) Blockade or knockout of the apical AT1A receptor prevents the response to BL CO2, consistent with the observation of others that the PT secretes an ANG II‐related product into the lumen. And (4) BL but not L lisinopril (apparently by blocking intracellular ACE activity) prevents the action of BL CO2, suggesting that the PT secretes preformed ANG II. To obtain a “true” dose‐response curve for the effect of L ANG II on CO2‐evoked HCO3 reabsorption, we blocked secretion of preformed ANG II by exposing isolated perfused PTs to 240 nM BL lisinopril. Adding 10−11 M exogenous ANG II to L increased JHCO3 by 43% at [CO2]BL = 5% (P = 0.01, n = 5) and by 61% at [CO2]BL = 20% (P = 0.02, n = 5), but had no effect at [CO2]BL = 0% (P = 0.37, n = 6). Further, adding 10−9 M ANG II to L raised JHCO3 by 60% at [CO2]BL = 20% (P = 0.01, n = 6) but had no effect at [CO2]BL = 5% (P = 0.33, n = 5). Our data indicate that the PT's ability to raise JHCO3 in response to increases in [ANG II]L absolutely requires basolateral CO2, and that the response rises with increases in [CO2]BL. Thus, CO2 plays an important role in the signal‐transduction pathway initiated by luminal ANG II.
ERK Activation Regulates Acid Base Balance in Proximal Tubule‐like LLC‐PK1 Cells
Authors: Oliver Robert E. , Friday Ellen , Turturro Francesco , Welbourne Tomas
Published: June 2021

Both metabolic acidosis and glitazones (rosiglitazone, RGZ, and troglitazone, TRO) activate ERK in LLC‐PK1 cells. To determine the effect of p‐ERK on acid base balance without the complication of an extracellular acidosis, TRO was used to elevate p‐ERK with acid extrusion and total production monitored over a 12min time course. After 12min, the p‐ERK to total ERK ratio increased (p<0.01) 4.8fold (0.7± 0.2 to3.35±0.7) while the spontaneous pHi dropped (7.10±0.02 to 6.78±0.04, p<0.001). NHE3 activity assayed at the end of the 12min by acid (NH4+) pulsing was reduced by 54% with the recovery pHi less (p<0.01) than the control recovery pHi (6.67± 0.07 vs. 7.04± 0.05). Activation of p‐ERK was associated with a 2fold increase (p<0.001) in acid production (402±34 vs. 207± 37nmol/12min) despite the fall in lactate released to the media with ERK activation (38 ± 3 vs. 57± 6nmol/12min, p<0.05). EIPA (100uM) reduced NHE3 activity by 63% but reduced pHi only minimally and failed to activate ERK or enhance acid production. Replacing glucose with 2‐deoyglucose abrogated the cellular acidosis (6.94± 0.03 vs. 6.71± 0.02, p<0.02) and prevented enhanced acid production (187± 9 vs. 423± 17nmoles/12min, p<0.001) with TRO but did not prevent the ERK activation. MEKK inhibitors PD98059 (30uM)+UO126 (10uM) completely prevented TRO‐induced ERK activation (0.3± 0.2 vs. 3.35± 1.1, p<0.01) and the fall in spontaneous pHi (7.02±0.03 vs. 6.78±0.04, p<0.001); total acid production and NHE3 activity in the presence of TRO returned to the control levels. RGZ gave similar effects but was less potent than TRO. These results are consonant with ERK activation playing a key role in PT cell acid base balance through regulation of both acid production and extrusion and suggest that the former may modulate extrusion.
Blood Oxygen Level Dependent Magnetic Resonance Imaging and Renal Tissue Oxygenation with Successive Inhibition of Renal Sodium Transport
Authors: Warner Lizette , Haas John A. , Bolterman Rodney , Gomez Sabas , Joyner Michael J. , Romero Juan Carlos
Published: June 2021

Functional renal oxygen consumption, VrO2 is due to tubular reabsorption of sodium (TNa) in the proximal and distal nephrons, resulting in compartments (cortex/medulla) with distinct VrO2/TNa relationships. Compartmental divisions promote non‐invasive imaging strategies like MRI, to indirectly assess tissue oxygenation, (PtO2). However, the extent to which these regional PtO2 differences are affected by successive TNa are not fully understood. The objective is to determine VrO2 and intra‐renal regional alterations in PtO2 directly and indirectly during successive TNa inhibition. It is hypothesized successive TNa inhibition is accompanied by intra‐renal regional changes in PtO2. Pigs (n=6) underwent MRI scans at baseline, furosemide and acetazolamide. Five days later, the right kidney was instrumented with PtO2 sensors (cortex/medulla). PtO2 was measured during drug infusions (identical to MRI exam). There was a decrease in VrO2, and a 40% and 30% increase in medullary and cortical PtO2 following drug infusions. There was ~25% medullary and ~20–30% cortical decrease in BOLD signal with furosemide and acetazolamide administration, respectively. This study suggests successive inhibition of TNa is accompanied by intra‐renal regional PtO2 changes and these changes are detectable with BOLD‐MRI, a non‐invasive imaging modality.

Funding provided by NIH Grant (HL16496‐32), 2007 APS Porter Physiology Fellowship and the Mayo Foundation.
RAGE targeting leads to protection from hyperoxia‐induced lung injury
Authors: Reynolds Paul R , Kasteler Stephen D , Sturrock Anne , Sanders Karl , Kennedy Thomas P , Hoidal John R
Published: June 2021

Critically ill patients often require supplemental oxygen during treatment; however the impact of hyperoxia on pulmonary biology is not well established. Receptors for advanced glycation end‐products (RAGE) are multiligand Type I cell surface receptors that influence development and cigarette smoke‐induced inflammation (Reynolds et al, 2007), but studies that address the role of RAGE in acute lung injury are insufficient. RAGE null mice exposed to hyperoxia survived 3 days longer than age‐matched wild‐type mice (9 vs. 6 days). After four days in hyperoxia, RAGE null mice had lower total cell infiltration into the airway, including PMNs and macrophages, and decreased total protein leak. Additionally, an inflammatory cytokine antibody array (Chemicon) and Western blotting revealed decreased secretion of several pro‐inflammtory molecules in lavage fluid obtained from RAGE knock out mice when compared to wild‐type. These data reveal that RAGE targeting leads to a diminished hyperoxia‐induced pulmonary inflammatory response. Further research into the role of RAGE signaling in the lung should identify novel targets likely to be important in the therapeutic alleviation of lung injury and associated persistent inflammation. Supported by the Flight Attendants Medical Research Institute (FAMRI, PRR) and the Parker B. Francis Foundation (PRR).
Lamellar body secretion in cyclically stretched rat alveolar type II cells depends on the geometry of cell‐cell interaction
Authors: Felder Edward , Appelt Daniel , Zheng Xing , Dietl Paul
Published: June 2021

Alveolar type II cells respond to mechanical stimulation with a fusion of surfactant containing lamellar bodies (LBs) with the cell membrane. Single fusion events of LBs can be detected with the fluorescent dye FM1‐43 in vitro which was used to investigate the effect of cyclic stretch on LB exocytosis in primary rat type II cells. After cultivating the cells on an elastic growth support we used a micro‐robotic arm (kleindiek nanotechnik) that allowed us to locally stretch the underlying growth substrate and hence to apply unidirectional stretch to individual clusters of cells. Stretch of different frequencies (1, 2 and 4hz) revealed that the fusion response was inversely related to the frequency but only slighly lower (1.84Hz) in responders as compared to non‐responders (2.0Hz). Also the stretch amplitude (from 1 to 25%) did barely correlate with the number of LB fusions. Cells that responded with fusions were generally attached to more cells than the ones without response indicating cell‐cell interaction of some kind to account for the process. In cases were the position of the adjacent cells with respect to the stretch axis could precisely be defined, we found that an unsymmetric deformation rather than simple elongation of a cell cluster lead to an increased number of fusions. Latter shows that the mechanosensitive machinery of the ATII cell is able to distinguish between force patterns and we suggest cytoskeletal elements to be a key player in mediating this process.

Supported by the FWF, grant P15743, the DFG, grant D1402, the Landesstiftung Baden‐Württemberg and the 6th framework of the EU, Pulmo‐Net.
Model simulation of expired CO2 from its alveolar distribution in parallel and series models
Authors: Caucha Luis Jhony , Cruz Julio Cesar , Rueda Luis Antonio
Published: June 2021

The CO2 model of Scherer (J. Appl. Physiol. 64:1022, 1988) is here applied to the 7 regions model of Jeng et al (Ann. Biomed. Eng. 28:453, 2000). While Scherer et al ended up with an alveolar CO2 concentration of 3.1% and used a pulmonary blood flow of 100 mL/s, we applied West concept of less CO2 and blood flow at the apex. The alveolar CO2 concentrations and the pulmonary blood flow at the apex and base of the lung were: 2.6% and 4.4%; 1.2 mL/s and 41mL/s, respectively. The anatomical model of Weibel is scaled for each region The results are on the figure. Although the curves are similar, our curve shows a larger dead space and less slope of phase 3. The base of the lung contributes more to the onset of CO2 in the capnogram.

Supported in part by ONG‐Centro de Enseñanza, Investigación y Servicios.
Measuring Collateral Pathways in Health via 3He MRI
Authors: Woods Jason C , Bartel Seth‐Emil , Conradi Susan H , Chang Yulin V , Yablonskiy Dmitriy A , Gierada David S , Conradi Mark S
Published: June 2021

Introduction

There has been recent interest in the use of 3He diffusion MRI because of the close correlations of the imaging measure with tissue morphometry. The long‐range diffusion coefficient (LRDC) has been shown to be a measure of collateral pathways in emphysema, but little is known of the role of collaterals in the LRDC in health. A recent publication attempted to explain the LRDC by simulations within single lung acini, without collateral pathways.

Methods

We measured the LRDC of 3He ex‐vivo in 4 human donor lungs that were rejected for transplantation and in 3 pig lungs, which are known to have greatly reduced collateral ventilation. We also simulated the interacinar diffusion of 3He (at 2‐cm diffusion distances) via the diffusion equation in a strictly branching model of human lung with no collateral pathways. (Airway radii, lengths, and branching angles were taken from detailed, morphometric literature.)

Results

LRDC in pig lungs (0.004 cm2/s) was 4x lower than in the human lungs (0.022 cm2/s), implying that collateral pathways can play a significant role in the measurements. The robustly tested simulations revealed a surprisingly low LRDC (24x lower than the measured value), implying that diffusion via the airway tree alone (no collaterals) cannot account for the measured value in healthy lung. A surprisingly small number of 5‐μ pores is shown to account for the discrepancy.
Acute effects of nerve growth factor (NGF) on membrane properties of mouse lower airway parasympathetic ganglionic neurons
Authors: Myers Allen C , Rohde Holly K
Published: June 2021

The neurotrophin, NGF, is located in lower airway smooth muscle and glands, tissue innervated by local parasympathetic ganglionic neurons, and NGF is increased by inflammation in this tissue. However, the acute effect of NGF release on parasympathetic nerves is unknown. Using single cell current clamp recording of tracheal and bronchial ganglionic neurons, NGF (30 ng/ml) decreased the action potential afterhyperpolarization amplitude and duration from 17±2mV and 52±8ms, respectively, in control neurons to 11±3 mV and 34±14ms after treatment (n=7); NGF (1–30 ng/ml) had no effect on accommodation in phasic (n=4) or tonic (n=3) neurons. NGF (1–30 ng/ml) had no effect on the resting membrane potential or input resistance (n=7). Using intracellular QX314, vagus nerve stimulation evoked 16±2mV (range of 13–19 mV; n=6) nicotinic fast excitatory postsynaptic potentials (fEPSPs) in control neurons which were increased to 21±3mV in the presence of 30ng/ml NGF. Using whole cell patch clamp recording, DMPP (1μM, 30s) evoked a 213pA inward current in a control neuron and 258pA in the presence of NGF (10ng/ml). In this study, we determined that NGF affects action potential and synaptic membrane properties of mouse ganglionic neurons; the effect on fEPSPs is possibly postsynaptic as NGF increased the nicotinic current. This study provides additional support that NGF has short‐term, as well as long‐term, effects on adult mammalian neurons. This work was supported by NIH‐NHLBI (ACM).
Estrogens modulate intracellular Ca2+ in human airway smooth muscle
Authors: Townsend Elizabeth A , Thompson Michael A , Pabelick Christina M , Prakash YS
Published: June 2021

Asthma is a disease characterized by airway hyperresponsiveness and inflammation. One‐third of pregnant women with pre‐existing disease experience a decrease in symptoms during the third trimester, when estrogen levels are highest. Previous studies have shown that estrogens relax vascular smooth muscle, however, little is known about estrogen effects on airway smooth muscle (ASM). We tested the hypothesis that estrogen modulates human ASM contractility. Western blot analysis and immunohistochemical studies showed the presence of both full‐length and truncated isoforms of ERα and β. Furthermore, ER isoforms were localized to caveolae. Caveolin‐1 siRNA downregulated ERα isoform in human ASM. Exposure to the pro‐inflammatory cytokine TNFα substantially increased ERα expression in ASM. In fura‐2 loaded human ASM cells, acute exposure to biologically‐active 17β‐estradiol decreased the intracellular Ca2+ response to contractile agonists such as histamine and bradykinin; this effect was blunted by the ER antagonist ICI 182, 780. Overall, these data suggest that estrogens may have a bronchodilatory effect. Accordingly, increased ER expression with airway inflammation may actually be protective during pregnancy in that bronchodilation resulting from higher estrogen levels (acting via ERs) may alleviate the increased ASM contractility of asthma.

Supported by the Mayo Graduate School and NIH grant UL1RR024150‐01(Mayo Clinic CTSA awards to YSP and CMP)
TLR3 activation stimulates TLR2 expression and innate immune responses to TLR2 ligands in airway epithelial cells
Authors: Melkamu Tamene , O’Grady Scott M.
Published: June 2021

Toll‐like receptor (TLR1‐6) mRNAs are expressed in normal human bronchial epithelial (NHBE) cells with higher basal levels of TLR3. Expression of TLR2 mRNA was enhanced by treatment with Poly IC, a TLR3 ligand but not by the TLR2 ligand, PAM3CSK4, suggesting that viral infection can alter epithelial sensitivity and response to microbial components sensed by TLR2. Western blot analysis revealed that Poly IC pretreatment (24 hrs) produced up‐regulation of TLR2 protein. Activation of TLR3 also up‐regulated mRNA expression of MyD88 and TIRAP, adaptor molecules that are involved in TLR2 signaling. To assess consequences of TLR3 activation on TLR2 function, monolayers of NHBE cells were pretreated with Poly IC for 24 hrs and then stimulated with PAM3CSK4 for another 24 hrs. ELISA analysis of media harvested from the apical and basolateral compartments revealed significantly higher secretion of IL‐6 and GM‐CSF after Poly IC stimulation compared with PAM3CSK4 or flagellin (TLR5 ligand) and that apical secretion was significantly greater than basolateral secretion. Additionally, PAM3CSK4 stimulation of IL‐6 and GM‐CSF secretion following Poly IC pretreatment was significantly greater than the effects of PAM3CSK4 without Poly IC pretreatment. Our findings indicate that TLR3 activation enhances expression of the TLR2 signaling pathway and potentiates the functional response of the airway epithelium to TLR2 ligands.
Transepithelial glucose transport and metabolism in H441 human airway epithelial cells
Authors: Kalsi Kameljit Kaur , Fraser Owen , Chung Yuen‐Li , Tarelli Edward , Philips Barbara , Baker Emma , Baines Deborah L
Published: June 2021

Increased airway glucose is associated with increased risk of lung infection. The aim of this study was to characterise transepithelial glucose transport and metabolism in a model of human airway epithelial cells. Transepithelial transport of glucose from the basolateral compartment (B) to the apical compartment (A) or from A to B was traced using 3H‐D‐glucose or non‐metabolizable 14C‐L‐glucose. More apical radiolabel accumulated (B to A) than basolateral (A to B) with 5mM 3H‐D‐glucose (0.103 ± 0.013 μmoles and 0.04 ± 0.01 μmoles per hour cm−2 p < 0.05, n = 4 respectively). Apical (B to A) and basolateral (A to B) radiolabel accumulation was similar with 5mM 14C‐L‐glucose (0.03 ± 0.01 μmoles per hour cm‐2 and 0.03 ± 0.02 μmoles per hour cm‐2 respectively). However, HPLC analysis revealed that transepithelial apical glucose accumulation (B to A) was significantly less with 5mM D‐glucose (0.01 ± 0.02 μmoles per hour cm‐2) compared to 5mM L‐glucose (0.16 ± 0.05 μmoles per hour cm‐2, p<0.05, n = 4). To try and account for this discrepancy we used proton NMR and identified metabolites formed from D‐glucose such as lactate, alanine, acetate and scyllo‐inositol (a phospholipid precursor) in the apical chamber. These data indicate that transepithelial transport and apical accumulation of glucose in the lung epithelium is reduced by intracellular metabolism and the production and release of glucose metabolites. Funded by Wellcome.
Development of a simulation for teaching fluid therapy
Authors: Rawson Richard E. , Dispensa Marilyn E. , Goldstein Richard E. , Nicholson Kimberly W. , Vidal Noni Korf
Published: June 2021

The course “Management of Fluid and Electrolyte Disorders” is taught using lectures and paper‐based case scenarios. While the latter provide an excellent basis for application of basic‐science concepts, they lack key components of genuine clinical cases that, by nature, are diverse, change over time, and respond in unique ways to therapeutic intervention. We developed a dynamic mathematical model using STELLA® software that simulates normal and abnormal fluid and electrolyte balance in the dog. The simulation outputs real‐time clinical data allowing students to develop a therapeutic plan based on their clinical assessment. Students administer fluids and supplements and the model responds in real‐time, requiring regular reassessment and intervention. Level of success is determined by clinical outcome including improvement, deterioration, or death. Underlying variables can be exploited to illustrate and apply concepts such as compartment volumes, endocrine responses, and dynamic mass balance. We expect that the simulation will allow students to improve clinical competency, apply fundamental principles in disparate contexts, and practice the “art” of fluid therapy without use of live animals.
The development of applied cellular and molecular medical experiences as a supplement to the medical curriculum
Authors: Henry Matthew K , Mortensen Luke
Published: June 2021

The first and second year curriculum at many medical institutions offers little opportunity to cover in depth basic scientific principles. Yet, there is an increase in the requirement for medical students to participate in applied experiences, such as human simulation and standardized performance assessment laboratory exercises. We are interested in using a similar applied approach to reinforce the importance of basic scientific principles. Here, we present the developmental structure and the evaluation of an elective course offering entitled “Advanced Physiology & Pharmacology: From Molecules to Medicine.” This course offers medical students an opportunity to explore the basic science behind medicine through traditional didactic sessions, literature review, and active participation in laboratory research. Following two course iterations, we were interested in determining if medical students find value in this supplemental course work. Each student was required to evaluate their perceived increased: a) knowledge of physiology/pharmacology (4.30±0.17), b) ability to understand primary literature (4.39±0.16), and c) understanding of experimental and diagnostic techniques (4.23±0.17) on a Likert scale (1–5). Based on student evaluations of the course, medical students appear to be interested in expanding their exposure to basic science and perceive added value to their medical educational.
Student Interactions During Collaborative Group Testing
Authors: Giuliodori Mauricio Javier , Lujan Heidi , DiCarlo Stephen Edward
Published: June 2021

We used collaborative group testing in a veterinary physiology course (65 students) to answer the following research questions: 1.‐ When students change their initial answer following the discussion, which students change; students with initial incorrect responses or students with initial correct responses?, and 2.‐ When is the discussion most effective; when students initially agree or disagree?. For the collaborative testing format, students answered questions in the traditional format as individuals. Immediately after completing the exam as individuals, students answered the same questions in groups of two, and finally, the same questions were discussed by the instructor and the students. We found that students with initial incorrect responses changed their answers more often than students with initial correct responses (OR: 7.58, P=0.001). Furthermore, student feedback is more beneficial when group members have different initial responses (P<0.05). Finally, when group members have different initial responses more students change to the correct answer than to the incorrect answer (77% vs. 23%, P<0.001). In conclusion, student discussion during collaborative group testing significantly enhanced student performance.
Integrating Teaching and Research at the Post‐doctoral level: The Fellowships in Research and Science Teaching (FIRST) Program
Authors: Holzman Jennifer L. , Brommer Chad L. , Eisen Arri , Haynes J. K. , Eaton Douglas C.
Published: June 2021

The Fellowships in Research and Science Teaching (FIRST) program presents post‐doctoral fellows with a unique experience that combines research and teaching in preparation for positions in academia. FIRST fellows participate in a traditional, mentored research setting at a consortium institution (Emory, Morehouse School of Medicine, and the Atlanta University Center), and as AUC teachers – at Spelman or Morehouse Colleges, or Clark Atlanta University. The complementation of these two critical aspects of academic life allows fellows to be trained in teaching pedagogies and techniques, and to gain hands‐on mentored experiences in the classroom, while balancing the demands of a research program. By receiving comprehensive feedback about their teaching, fellows can improve their teaching skills during their time in the program, setting them up for successful teaching jobs in the future. Fellows also participate in workshops and seminars, as well as continue to publish their research and present their work at national meetings. Of the 52 fellows that have completed the program, 65% have obtained jobs at a diversity of academic institutions, and continue to be active and innovative in both their research and teaching endeavors. Furthermore, FIRST fellows have exhibited success obtaining extramural funding (80%) at four times the rate of other NIH grant applicants.

Funded by: NIH, NIGMH K12‐GM00680‐06
The use of web‐based video files in lieu of face to face lectures in a medical physiology course ‐ one year's experience
Authors: Nazian Stanley J , Nicolosi Gregory R
Published: June 2021

In response to declining attendance at live lectures and to increase interactive learning, during the 2006/07 academic year lectures in the GI, Endocrinology and Reproduction blocks of the first year Medical Physiology course were delivered exclusively on line. Narrated presentations were prepared by the faculty that had presented these lectures the previous year and subsequently transformed into wmv files and posted on a secure intranet. Students viewed these files at their convenience. For each area, students were expected to meet with one of the participating faculty in groups of approximately 30 students for an intense, highly interactive session covering the material. To facilitate assessment, tests administered in these areas contained questions that were identical to those given the previous year. Performance on the two sets of questions was compared. Questions from the Pulmonary block served as an internal control. Students achieved essentially the same scores in both years: Pulmonary: 79.2 ± 2.6% vs. 79.3 ± 2.8%; GI: 77.5 ± 3.8% vs. 77.9 ± 4.4%; Endo/Repro: 81.7 ± 2.6% vs. 82.5 ± 2.4%. Medical students found the group conferences to be useful: 3.5 (5 point Likert Scale) but the response was mixed to the idea of expanding the system to the entire course: 2.8. Our experience suggests that delivering web‐based “lectures” has no adverse impact on student performance on examinations.
The TRIPSE (Tri‐Partite Problem‐Solving Exercise) in a Large Class Setting
Authors: Nastos Stash , Rangachari P K
Published: June 2021

The TRIPSE is a process‐oriented problem solving exercise that mimics the scientific process: 1) Given limited information, students frame a set of possible explanations. 2) They then select their best explanation and either design experimental tests or propose avenues for further explanation. 3) Given additional information, they re‐assess their original answers. This exercise has generally been used in courses with classes of 15 to 25 students. We report our experiences using the exercise in an introductory biology class of 204 freshmen. Prior to the actual exercise, students were given a practice run followed by a feedback session. We (SN/PKR) graded all TRIPSEs independently. In an exit survey, students rated different evaluation tools used in this course (journals, critiques, abstracts, TRIPSEs, posters, etc) for their learning value in comparison to standard MCQs. The TRIPSE received the highest ratings suggesting that they valued it greatly. We later had a 3rd assessor, who was not involved in the course, grade the answers to gauge inter‐rater reliability. This exercise, which students find valuable, could be readily adapted to large classes.
First principles for promoting meaningful learning
Authors: Modell Harold I.
Published: June 2021

When meaningful learning or learning with understanding takes place, students are able to apply information that is being acquired to solve novel problems. Most teachers agree that their goal is to have students learn their discipline(s) in ways that will be beneficial to their students’ lives. To reach this goal, teachers must establish a learning environment in which students will engage in meaningful learning. Such an environment has certain characteristics related to student behavior, the learning process, and the role of the instructor in the process. The design of a successful environment in this regard is derived from a set of first principles of classroom practice. These represent the assumptions and definitions on which all activities in the classroom are based. The following principles (presented in no particular order) are what I follow to promote a meaningful learning environment: 1) Students must take responsibility for their own learning. 2) Our job is to help the learner to learn. 3) Students come to us with prior knowledge. 4) Life is cumulative! 5) We must help students make their current mental model visible. 6) We must create safe learning envrionments.
Addressing the shortage of qualified staff in medical school anatomy laboratories and a proposed solution
Authors: Homer Richard W , Homer Heather , Mortensen Nathan T , Calderon Armando M , Dowdle Blake , Morton David A
Published: June 2021

Due to a limited number of instructors, the University of Utah has found it difficult to supervise medical students during cadaver dissections. To address this problem, qualified graduate and undergraduate students were hired. During the fall of 2006 a “teaching assistant (TA) dissection course” was implemented enabling new TA's to review the anatomy and dissection sequence prior to teaching the students. The lessons learned during the fall of 2006 more efficiently prepared the TA dissection course during fall 2007. The important task of establishing a weekly mandatory meeting to cover the dissection sequence and identification of structures was continued. In addition, added responsibility was given to the TA coordinator in organizing and directing TA dissections. Under the direction of the anatomy course director, TA's practiced drawing and describing nervous pathways along with blood supply and lymphatic drainage on a dry erase board. TA's would perform the dissection while others reviewed atlases and texts with each group sharing pertinent anatomy and clinical correlations. At the conclusion of the course a questionnaire was administered to each TA regarding their preparedness and confidence with laboratory content as a result of this class. Each participant indicated a positive experience.
Dissection in the UNAM_Mexico City
Authors: Herrera Patricia M
Published: June 2021

Anatomy dissection is being questioned all over the World. In the UNAM in Mexico City we still have the possibility of obtaining cadavers and we endorse the practice of dissection. In the past the dissection of regions was separated from the actual themes that were covered in class. We have proposed for the past 5 years to dissect exactly the same region that has been covered in the lectures. We divide the 40 student group into 4 groups each responsible for the dissection of a corpse. They rotate the different roles and have to finish finding the elements presented to them in a check list. They are examined precisely in the identification of those elements that represent the core knowledge of the Anatomy Course.

We compare the results obtained by those groups with the grades of the groups that dissect regions unrelated to the class topics.

The results show that there is a statistically significant difference at p= 0.5

We conclude that dissecting is important to the learning of anatomy.
Prosection anatomy – 167 students and 4 cadavers
Authors: Daly Frank J
Published: June 2021

Gross anatomy for physical therapy (DPT), occupational therapy (MOT), physician assistant (MPA) and athletic training students (ATC) occurs at the same time as medical gross anatomy course at the University of New England. Medical students perform dissections on 30 cadavers (4 students/body) and run in a traditional course from August to November. To accommodate the large numbers of students (125 medical, 30 DPT, 35MOT, 50 MPA and 50 ATC) without a corresponding large number of cadavers and costly lab space, modification of a traditional dissection course must be made. These students work from cadavers that are previously dissected, typically 3–6 months prior to each course. The four cadavers are only partially dissected to try to preserve their integrity for use in later courses. Cadavers are dissected to show specific structures to correspond with an in‐house lab manual, usually broken down by compartment or region. Students are walked through the prosections during scheduled lab time by faculty, lab instructors or clinical assistants. Historically, students have performed well on license exams prior to their start as health care professionals. This has worked for the University of New England as both a cost savings and space saving technique.
A Gross Anatomical and Histopathological Investigation of Cause of Death in a Cadaver in an Undergraduate Anatomy Lab
Authors: Green Erin Kelly , Jordan Clayton W. , Oldebeken Scott R. , Paudyal Sharad C. , Roehrs Jennifer E. , Potterfield April
Published: June 2021

An undergraduate gross anatomy dissection was performed at Westminster College as part of a human anatomy class, with one objective being to determine the cause of death in the cadaver. The cadaver was procured from Washington University. The dissection proceeded according to the details outlined in Grant's Dissector. During dissection several significant anatomical abnormalities were noted. A polytetrafluoroethylene (PTFE) vein graft was present in the left forearm joining the radial artery and cephalic vein, indicating dialysis. The kidneys were irregularly shaped and had various‐sized cysts (0.1–5.0 cm) and dense, black nodules on their surfaces, as well as internal abnormalities. These factors combined with other abnormalities (left ventricular hypertrophy [2.0 cm thick myocardium compared to a normal range of 0.6–1.1 cm], calcium deposits, and plaque in the abdominal aorta and renal arteries), support our preliminary findings of renal failure as the cause of death. Histopathological analysis of several samples will be conducted to support these findings.
An immunohistochemical study of human costotransverse joints
Authors: Smith Michael P , Sawyer Barbara , Brismee JM , Sizer Phil , Dedrick Gregory
Published: June 2021

The costotransverse joint (CTJ) is the articulation between the posterior tubercle of the ribs with the first through 10th costal facet of the thoracic transverse processes. Anatomical description of the CTJ articulation verifies an extensive network of ligaments that are characteristic to other synovial joints. A few authors suggest the CTJ is a pain generating structure and a synovial joint. However, there is no histological evidence to suggest that the CTJ is a synovial joint or a pain generating structure. The purpose of this study was to determine if there is evidence of pain generating fibers (somatic and sympathetic) within the CTJ. In the current study, CTJ tissue from levels one through ten was obtained from two fresh cadavers and fixed in 4% paraformaldehyde. The fixed tissue was processed and embedded in paraffin, cut and mounted on slides. Immunochemical studies demonstrated immunostained neural elements in the CTJ using antisera against neuropeptide Y, substance P, and calcitonin gene‐related peptide. This study provides preliminary evidence for the hypothesis that the CTJ possesses immunoreactive features characteristic of a pain generating structure.
Mechanical loading prevents decreased bone formation responses of osteoblasts by a bone morphogenic protein‐dependent mechanism
Authors: Patel Mamta , Talish Roger , Rubin Clinton , Jo Hanjoong
Published: June 2021

Bone loss due to osteoporosis, disuse, and microgravity is a significant health problem. A low magnitude and high frequency (LMHF) mechanical loading has been shown to prevent bone loss in animals and humans. However, the underlying mechanisms and the target cells through which LMHF loading alleviates bone loss are not known. We hypothesized that direct application of LMHF loading to osteoblasts alters cell response, preventing decreased bone formation induced by disuse. To test our hypothesis, pre‐osteoblast 2T3 cells were exposed to the Random Positioning Machine (RPM) and intervened with LMHF loading (0.3g, 30Hz, 10 min). The RPM decreased bone formation markers as determined by alkaline phosphatase (ALP) activity and mineralization even in the presence of BMP4 (20ng/ml). However, LMHF loading prevented the RPM‐induced decrease in both markers. Mineralization induced by LMHF loading was enhanced by treatment with BMP4 and blocked by the BMP antagonist noggin, suggesting a role for BMPs. LMHF loading also rescued the RPM‐induced decrease in ALP, runx2, osteomodulin, parathyroid hormone receptor 1, and osteoglycin. These findings show that osteoblasts respond directly to LMHF loading, providing mechanistic insight into how LMHF loading prevented bone loss in vivo. The genes identified here provide potential targets for pharmaceutical treatments that may be used with LMHF loading to better treat bone loss.
Down‐regulation of MLH1 in a relapsed leukemia patient is associated with a three‐nucleotide deletion in the MLH1 3′ untranslated region
Authors: Gu Liya , Mao Guogen
Published: June 2021

Defects in DNA mismatch repair (MMR) are the molecular basis of certain cancers, including hematological malignancies. These defects are often caused by mutations in the coding regions or promoter hypermethylation of of MMR genes. However, in many cases, despite that a hypermutable phenotype is detected, no mutations/hypermethylations of MMR genes can be identified. We report here a novel mechanism leading to MMR deficiency: mutations in the 3′ untranslated region (3′‐UTR) of the MLH1 gene. A relapsed leukemia patient displayed microsatellite instability, but no genetic and epigenetic alterations in key MMR genes were identifiable. Instead, a 3‐nucleotide (ttc) deletion in the 3′ UTR of MLH1 was identified in the patient's blood samples. The 3′ UTR mutation was therefore examined for its role in regulating the MMR function. We show that the mutant MLH1 3′ UTR significantly reduced expressions of both a firefly luciferase reporter gene and an ectopic MLH1 gene in model cell lines. Consistent with these observations, a significant reduction in the steady‐state level of MLH1 mRNA was found in white blood cells of the patient. These findings suggest that the mutant MLH1 3′‐UTR can cause a severely reduced/defective MMR activity conferring leukemia relapse, likely by down‐regulating MLH1 expression at the mRNA level. Although the exact molecular mechanism by which the mutant 3′‐UTR down‐regulates the MLH1 mRNA is not known, our findings provide a novel marker for cancers with MMR defects.
Epigenetic link between DNA methylation and histone modifications
Authors: Cheng Xiaodong , Jia Da , Collins Robert E , Li Keqin , Horton John R , Zhang Xing
Published: June 2021

Genetic imprinting, found in flowering plants and placental mammals, uses DNA methylation to yield parent‐of‐origin‐dependent gene expression. DNA methyltransferase 3a (Dnmt3a) and its regulatory factor, DNA methyltransferase 3‐like protein (Dnmt3L), are both required for de novo DNA methylation of imprinted genes in mammalian germ cells. Dnmt3L interacts specifically with unmethylated lysine 4 of histone H3 through its N‐terminal Cys‐rich domain that has strong structural similarities to the PHD‐like domain of BHC80, a component of LSD1‐mediated histone lysine demethylase complex. The C‐terminal domain of Dnmt3L is shown by crystallography to interact with the catalytic domain of Dnmt3a, demonstrating that Dnmt3L has dual functions of binding of unmethylated histone tail and activating DNA methyltransferase. The complexed C‐terminal domains of Dnmt3a and 3L showed further dimerization through Dnmt3a‐3a interaction, forming a tetrameric complex with two active sites. Substitution of key non‐catalytic residues in the Dnmt3a‐3L interface or Dnmt3a‐3a interface eliminated enzymatic activity. Molecular modeling of a DNA‐Dnmt3a dimer suggested that the two active sites are separated by approximately one DNA helical turn. Studies were supported partially by NIH grants (GM049245 and GM068680) and Georgia Research Alliance.
ARID domain of H3K4 demethylase RBP2 binds to GC rich DNA
Authors: Tu Shengjiang , Yuan Chunhua , Teng Yu‐Ching , Cheng An‐Ning , Wu Ying‐Ta , Juan Li‐Jung , Tsai Ming‐Daw
Published: June 2021

In an effort to identify human trithorax response elements (TREs) and study transcription regulation by histone H3K4 demethylase RBP2, a trithorax group protein, we studied RBP2 by DNA binding, solution NMR, immunofluorescence, and reporter assays. In this study, PCR‐assisted DNA binding selection revealed that RBP2 DNA binding domain ARID selectively binds to GC rich DNA. Subsequent electrophoretic mobility‐shift assays verified RBP2 ARID binds to BRD2 promoter via a consensus G/C run motif. Solution NMR structure of RBP2 ARID showed it had 6 helices and 2 loops. Further NMR titration on ARID‐DNA complex identified that the binding interface is consisted of loop 1, helix 4, loop 2, and helix 5. Importantly, we found loop 2 contributes significantly to DNA binding affinity. One residue in loop 2 could be used to predict ARID DNA binding specificity. Moreover, in vivo immnofluorescence and reporter assays indicated that ARID is necessary for proper RBP2 H3K4 demethylase activity. In summary, the RBP2 ARID has regulatory roles and its consensus motif identified here is one of the first human TRE motifs. Funds from NIH and Academia Sinica
Structural and biochemical investigations of the Rtt109‐histone chaperone complexes
Authors: Berndsen Christopher , Tsubota Toshiaki , Kaufman Paul , Keck James L , Denu John M
Published: June 2021

Reversible acetylation of lysine 56 of histone H3 (H3‐K56) is critical for DNA replication, repair and transcription in S. cerevisiae. Recent evidence suggests the acetylation of H3‐K56 is catalyzed by Rtt109‐histone chaperone complexes. Rtt109 bears no apparent sequence similarity to known histone acetyltransferases (HATs), thus it likely represents a novel HAT. The association of Rtt109 with either of the histone chaperones Asf1 or Vps75 is critical for increasing the efficiency of H3‐K56 acetylation in vitro and in vivo. While Asf1 has been directly linked to H3‐K56 acetylation in vivo, the functions of the Rtt109‐Vps75 complex are unclear. Using X‐ray crystallography and biochemical analyses, we are characterizing the structure and functions of the Rtt109‐Vps75 complex in vitro. We find that while Rtt109‐Vps75 prefers to acetylate H3‐K56, Rtt109‐Vps75 can also efficiently acetylate K9 and K27 of H3, while Rtt109‐Asf1 greatly prefers H3‐K56 over other sites within H3. Furthermore, the tail of H3 is required for efficient acetylation of H3‐K56 by Rtt109‐Vps75 but not for Rtt109‐Asf1, suggesting that substrate recognition and site specificity of Rtt109 may be modulated by the associated histone chaperone. This research is funded by grants from the National Institutes of Health and a pre‐doctoral fellowship from the American Heart Association.