bluuebunny/crossref_metadata_2025 huggingface dataset:
Snail Regulates Type I Collagen Expression in Cardiac Fibroblast and Myofibroblast
Authors: Katwa Laxmansa C , Salameh Muhammad M , Ferrell Paul D
Published: June 2021

Snail is a transcription factor that binds to the E‐box sequence to regulate the promoter activity of target genes and is involved in cell signaling, wound healing, cell migration, apoptosis, and EMT. We evaluated the gene expression in Sprague‐Dawely rat myocardial infarction (MI) models. Snail expression was found up regulated in MI heart tissue relative to non‐MI. We then investigated the regulatory role of Snail in angiotensin II (AngII) induced collagen expression. Primary cultures of cardiac fibroblast and myofibroblasts (isolated from site of MI) treated with 10−7 M AngII, a strong stimulator of collagen I expression, resulted in significant increase in Snail expression, confirmed by RT‐PCR. Fibroblast and myofibroblast treated with losartan, an AngII type 1 receptor (AT1) blocker, resulted in reduced Snail expression. Snail siRNA transfection confirmed its regulatory role in AngII induced collagen I expression, which resulted in significant down regulation of collagen I expression (*p= <0.001). These data demonstrate AngII induced collagen expression is regulated by Snail, which is under the control of AT1 receptor mediated signaling pathways in cardiac fibroblast and myofibroblasts.
Shank2 regulates NaPiIIa abundance and endocytosis in OK cells
Authors: Dobrinskikh Evgenia , Cranston DeeAnn , Rachelson Joanna , Moldovan Radu , Lei Tim , Doctor R. Brian
Published: June 2021

Sodium phosphate co‐transporter IIa (NaPiIIa) is concentrated in the apical microvilli of renal proximal tubule (PT) cells under low serum phosphate (Pi) conditions but is internalized to and degraded within lysosomes under high Pi conditions. In vivo rat studies indicate that Shank2, a PDZ domain protein that binds the C‐terminal tail of NaPiIIa, undergoes a parallel pattern of internalization. The present study utilizes OK cells to determine the functional significance of Shank2 on NaPiIIa. Under low Pi (0.1 mM) conditions, KD of Shank2 expression resulted in a parallel decrease in NaPiIIa abundance. 3 hours after shifting to high Pi (2.0 mM) media, western blotting showed both NaPiIIa and Shank2 undergo degradation. After 1 hour in high Pi media, confocal imaging of fixed cells shows that both native and expressed NaPiIIa and Shank2 are entered the sub‐apical domain. Live‐cell imaging shows that GFP‐NaPiIIa and mRFP‐Shank2 traverse through the sub‐apical domain together as a dual‐labeled endosomes. Fluorescence cross‐correlation spectroscopy (FCCS) provided quantitative evidence at the molecular level of NaPiIIa and Shank2 co‐migration through the sub‐apical domain. Together these findings indicate that, under low Pi conditions, Shank2 contributes to the retention of NaPiIIa within PT cells and that, under high Pi conditions, Shank2 is positioned to impact the endocytosis and trafficking of NaPiIIa.
The cutaneous vascular response to heat stress does not explain sex‐related differences in sudomotor activity
Authors: Gagnon Daniel , Kenny Glen P
Published: June 2021

Changes in skin blood flow can independently modulate changes in sweat production. These findings could explain the lower whole‐body sudomotor activity recently observed in females during exercise. We therefore examined potential sex‐differences in local cutaneous and whole‐limb vascular conductance during exercise in the heat (40°C). Eight males and eight females (follicular phase) performed three successive 30 min exercise bouts at fixed rates of metabolic heat production equal to 200, 250, and 300 W·m−2. Local sudomotor activity (ventilated capsule) and cutaneous vascular conductance (CVC, laser‐doppler) were measured on the back, chest, and forearm, while whole‐limb vascular conductance was measured at the forearm (FVC, venous occlusion plethysmography). Despite a similar requirement for heat loss during each exercise bout, increases in local sudomotor activity were significantly lower in females at each measurement site (p≤0.001). However, these differences were paralleled by similar CVC responses on the back (43 ± 2 vs. 47 ± 5%, p=0.195), chest (52 ± 5 vs. 48 ± 4%, p=0.705), and forearm (51 ± 3 vs. 60 ± 6%, p=0.212). Furthermore, FVC did not differ between sex throughout exercise (0.09 ± 0.01 vs. 0.10 ± 0.01 L·100 mL−1·min−1·mmHg−2, p=0.278). These results suggest that the lower sudomotor activity observed in females during exercise is not due to a different cutaneous vascular response. Supported by NSERC grant RGPIN‐298159‐2009.
Dendritic release of VP mediates crosstalk between neuroendocrine and presympathetic PVN neurons: Role in osmotically‐driven homeostatic responses
Authors: Son Sook Jin , Filosa Jessica A. , Zheng Hong , Patel Kaushik P. , Stern Javier E.
Published: June 2021

Dendritic release of vasopressin (VP) in the SON and PVN nuclei plays an important role in the autoregulation of VP neuronal activity. Here, we tested the hypothesis that dendritically‐released VP acts in a diffusible manner to influence the activity of neighboring presympathetic neurons. Patch‐clamp recordings in brain slices showed that focally applied VP depolarized and increased presympathetic PVN‐RVLM neurons (~450%, P< 0.01), an effect that persisted in the presence of GABA/glutamate receptor blockers. Activation of a single, identified eGFP‐VP neuron using photolysis of caged NMDA induced a Ca2+‐dependent, V1a‐mediated increased firing discharge in neighboring PVN‐RVLM neurons. Blockade of V1a receptors per se reduced PVN‐RVLM firing discharge (~75%, P< 0.02). Increasing (high K+, aminopeptidase blockade) and decreasing (κ opiod agonist) endogenous VP ambient levels, respectively, further increased/decreased PVN‐RVLM firing activity. Finally, a central osmotic stimulation (NaCl 0.3–2.1 omosl/l ICA) induced a prompt increase in RSNA, and effect that was blunted (~50%) when V1a receptors were locally blocked within the PVN (P<0.001). Collectively, our results demonstrate that dendritic release of VP mediates a functional crosstalk between neurosecretory and presympathetic PVN neurons, playing an important role in the generation of osmotically‐driven homeostatic responses. Supported by NIH HL‐090948 to JES and NIH HL‐62222 to KKP.
Direct recording of renal sympathetic nerve activity in unrestrained, conscious mice
Authors: Hamza Shereen M. , Hall John E.
Published: June 2021

Renal sympathetic nerve activity (RSNA) has been measured in anesthetized mice which exhibit poor cardiovascular stability and low blood pressure which confounds proper interpretation of experimental results. We therefore developed a surgical and experimental protocol for measuring RSNA in conscious, unrestrained mice. Male C57Bl6J mice were chronically instrumented with blood pressure telemeters, a jugular venous catheter and a bipolar electrode for recording RSNA. Mice were placed in a home cage and left to recover for 48–72 hrs. Survival rate was 100%; all mice exhibited normal behavior with no sign of distress 24 hrs following surgery. RSNA was successfully recorded in 80% of the mice at 48 and 72 hrs post‐surgery; viable RSNA was reduced to 70 and 50% at 4 and 5 days post‐surgery, respectively. Mean arterial pressure (MAP, 116 ± 2 mmHg, n=10) was consistent with values previously reported for conscious mice. RSNA increased with the normal physical activities of eating and grooming and was validated by pharmacological manipulation of MAP; reduction in MAP to 62 ± 3 mmHg with nitroprusside increased RSNA by 159 ± 41% above baseline (n=5, p<0.05) whereas an increase in MAP to 137 ± 6 mmHg with phenylephrine reduced RSNA to −82 ± 4% compared to baseline (n=5, p<0.05). Thus, we demonstrate an accessible and effective method for direct assessment of RSNA in conscious, unrestrained mice. NIH HL51971, CIHR.
Aging and the effect of autonomic blockade on central and peripheral pulse wave velocity
Authors: Harvey Ronee , Casey Darren P. , Hart Emma C. , Charkoudian Nisha , Curry Timothy B. , Joyner Michael J. , Barnes Jill N.
Published: June 2021

Pulse wave velocity (PWV) and sympathetic nerve activity increase with age. However, it is unknown if the age‐related increase in sympathetic activity influences PWV. We determined the effect of autonomic blockade on peripheral and central arterial stiffness in young (23 ± 4 yr) and older, postmenopausal (57 ± 3 yr) women before and during administration of trimethaphan camsylate. Arterial tonometry was used to measure carotid‐radial (peripheral) and carotid‐femoral (central) PWV. The groups did not differ in height, weight, BMI, or mean arterial pressure (MAP). At baseline, peripheral PWV did not differ between the groups. However, central PWV was significantly greater in older women compared with young women (8.2 vs. 6.1 m/s, respectively; p<0.05). Trimethaphan infusion caused a substantial drop in MAP; therefore, we corrected PWV for the change in MAP. Peripheral PWV was greater in older women compared with young women (0.12 vs. 0.08 m/s·mmHg−1; p<0.05). In addition, central PWV remained higher in older women during trimethaphan infusion (0.11 vs. 0.08 m/s·mmHg−1 in young; p<0.05). In conclusion, central PWV was greater in older women before and during autonomic blockade, whereas peripheral PWV was only higher in the older women after autonomic blockade. This suggests that peripheral PWV is influenced by sympathetic activity to a greater extent in young vs. older women.

Funded by NIH
Augmented supraorbital skin sympathetic nerve activity responses to symptom trigger events in rosacea patients
Authors: Wilson Thad E. , Toma Kumika , Metzler-Wilson Kristen , Sammons Dawn
Published: June 2021

To investigate if rosacea alters skin sympathetic nerve activity (SSNA) and end‐organ responses, 12 patients and age‐ and sex‐matched controls underwent 2 symptom triggering stressors (mental and physical stress). Subjects performed 2‐min mental arithmetic and 2‐min isometric handgrip (35% of maximal voluntary contraction). SSNA (supraorbital microneurography), forehead skin blood flow (laser‐Doppler flowmetry), and forehead sweat rate (capacitance hygrometry) were measured. Heart rate and mean arterial pressure (MAP) were recorded via finger photoplethysmography. Cutaneous vascular conductance (CVC) was calculated by flux/MAP, and SSNA was normalized to baseline. Heart rate and MAP changes from baseline were not different between patients and controls during either stressor. Rosacea patients showed higher baseline sweat rate than controls (0.20±0.02 vs. 0.10±0.02 mg/cm2/min, p>0.05), and CVC trended higher compared to controls. SSNA during mental stress was higher in patients compared to controls (39±12% vs. 9±3%, p>0.05); similar augmented SSNA responses were observed during handgrip (rosacea = 31±13% and controls = 2±1%, p>0.05). These data identify trigger induced hyper‐responsiveness of sympathetic outflow in the face of rosacea patients and thus may be part of the etiology of the disease.

Funded by a grant from the National Rosacea Society.
Dissection of the Renal Failure‐3 QTL
Authors: Yeo Nan Cher , O'meara Caitlin , Lazar Jozef , Jacob Howard J.
Published: June 2021

Rf‐3, a quantitative trait locus (QTL) on the rat chromosome 3, affects renal disease susceptibility in the FHH (Fawn Hooded Hypertensive) rat. Synergistic interaction between Rf‐3 and other loci such as Rf‐1 had shown to be important in the disease development. Therefore, we used a triple congenic strain, in which Rf‐1, Rf‐3, and Rf‐4 were introgressed onto the genetic background of the renal disease resistant strain ACI (August Copenhagen Irish), to elucidate the genetic control of Rf‐3. The aim of this study was to reduce the Rf‐3 congenic region and identify the disease‐causing gene(s). By developing set of congenic strains we reduced the Rf‐3 region from 110 Mbp to 3.2 Mbp. The congenic strain carrying the reduced Rf‐3 region showed 2 and 3.5‐fold increase in proteinuria and albuminuria, respectively, compared to the control group. Kidney histology also revealed significantly increased protein casting and glomerular sclerosis in the minimal congenic strain. Sequence analysis of the congenic region revealed 2995 sequence variants between the FHH and ACI. We prioritized the variants based on sequence conservation across species and their location. Our analysis found 42 variants in 13 different genes. Two of them, Defb28 and Bcl2l1, are present in the syntenic mouse and human renal function QTL, which makes them potential candidates for future study. Research support: NHLBI‐5R01HL069321 to H.J.J.
Hydrochlorothiazide exacerbates metabolic syndrome in the SHROB rat model
Authors: Ernsberger Paul , Agarwal Akshita , Lute Beth , Koletsky Richard J
Published: June 2021

Metabolic syndrome consists of hypertension, insulin resistance, dyslipidemia and fatty liver. Thiazides are the leading treatment for hypertension, but some clinical studies have suggested adverse metabolic effects. We sought for the first time to characterize the metabolic effects of thiazides in an animal model of metabolic syndrome. SHROB rats are obese, hypertensive and express every component of human metabolic syndrome. Adult female SHROB rats (N=6 per group) were fed a purified diet (AIN 93) replete with electrolytes and treated in their drinking water for 5 weeks with either vehicle (250 ppm saccharine and 100 ppm sodium benzoate) or hydrochlorothiazide (achieved dose of 44±2 mg/kg/d). Systolic blood pressure fell progressively from 184±5 to 142±2 mmHg with thiazide treatment while remaining unchanged in controls. Heart rate was not affected. Body weight, food intake, and water intake did not differ between groups, nor did fasting glucose or insulin. Oral glucose tolerance testing (6 g/kg by gavage after an 18h fast) showed increased plasma glucose levels in the thiazide group (at 60 min post load, 394±48 vs 214±51 mg/dL in controls, P<0.05 by t‐test). Insulin levels at the same time were half again higher in SHROB treated with thiazide (90±20 vs 61±14 ng/mL in controls, NS, but two‐way ANOVA shows a difference, P<0.05 by F‐test). Relative liver mass was increased in the thiazide group relative to controls (43±3 vs 35±3 g/kg body weight, P<0.05 by t‐test). Thus, thiazide was an effective antihypertensive therapy in the SHROB model of metabolic syndrome but worsened glucose tolerance and insulin resistance and enlarged the already fatty liver. The secretory response of the pancreas to glucose was apparently not impaired or even increased, suggesting that insulin signaling may be affected by thiazides.

Research support for this study was given by Ophthalmology Education Worldwide
Stress induced increase in protein O‐linked‐N‐acetylglucosamine (O‐GlcNAc) levels is CaMKII dependent
Authors: Zou Luyun , Zou Chenhang , Chatham John C.
Published: June 2021

Acute cellular stress increases O‐linked‐N‐acetylglucosamine (O‐GlcNAc) modification of proteins; however, the mechanisms by which this occurs is not known. We have previously shown that in cardiomyocytes glucose deprivation (GD) is a potent stimulus for increasing cellular O‐GlcNAc levels and that this response was Ca2+ dependent. To determine whether Ca2+ contributed to increased O‐GlcNAcylation in response to other stress stimuli and in other cell types, C2C12 and HEK293 cells were subjected to either GD or heat shock in the presence or absence of KN93 a CaMKII inhibitor. Similar to cardiomyocytes both C2C12 and HEK293 cells exhibited a marked increased in O‐GlcNAc levels following 24hrs GD and in both cases this was significantly attenuated by treatment with KN93. Heat shock in HEK293 cells (42C for 1 hour) and C2C12 cells (45C for 1 hour) exhibited a clear time dependent increase in O‐GlcNAc following recovery at 37C, reaching a maximum at 30min and 3hrs in HEK293 and C2C12 cells respectively. Pretreatment with KN93 significantly attenuated the heat shock induced increase in O‐GlcNAc levels in both C2C12 and HEK293 cells. These data suggest that both GD and heat shock induced increase in O‐GlcNAcylation are mediated at least in part by Ca2+ induced activation of CaMKII; thereby providing new insights into the mechanisms underlying regulation of O‐GlcNAc cycling in cells subjected to acute stress.
Role of the TRPC4 proline rich region in regulation of the endothelial store‐operated calcium entry
Authors: Wang Xiaogan , Chen Hairu , Wu Songwei , Cioffi Donna L.
Published: June 2021

Endothelial cells line blood vessels and form a semi‐permeable barrier between blood and underlying tissues. Calcium entry through store‐operated calcium (SOC) channels mediates endothelial gap formation. The endothelial ISOC channel is a calcium‐selective SOC channel, and transient receptor potential canonical (TRPC) proteins are subunit candidates. While it is known that phosphorylation negatively regulates TRPC3 channel activity, the role of TRPC phosphorylation in regulation of the endothelial ISOC channel is unknown. Therefore we performed the Phos‐tag™‐based detection of phosphorylation with rat pulmonary artery endothelial cells (PAEC). We found that among TRPC1, TRPC3 and TRPC4, TRPC4 is the major phosphorylation target in the endothelial ISOC channel. TRPC4 carries a conserved serine in the proline rich region (PRR) as a putative phosphorylation site close to the channel pore. Transfection of the TRPC4 PRR peptide into PAECs suppressed the SOC entry and ISOC current. Collectively these data suggest that TRPC4 PRR regulates the endothelial SOC entry possibly through phosphorylation at the serine residue. Supported by 5R00HL089361.
Role of membrane permeability and extracellular diffusion in nitric oxide transport to the red blood cells
Authors: Deonikar Prabhakar , Kavdia Mahendra
Published: June 2021

Nitric oxide (NO) availability for vasodilation in vasculature is dependent on its consumption by red blood cell (RBC) hemoglobin. Extracellular diffusion in the plasma, cell free layer near the vessel wall and the RBC membrane permeability (Pm) reportedly lower NO consumption by RBCs. To understand the contribution of RBC membrane permeability and extracellular diffusion from the plasma layer surrounding the RBC towards NO uptake by RBCs, we developed a mathematical model simulating NO transport to a single RBC. Simulations were performed for Pm values ranging from 0.0415–40 cm/s and hematocrit (Hct) ranging from 1–45% and NO concentrations at the RBC membrane (CNOm inside and CNOm outside) were estimated. NO concentration gradients in the plasma layer increased with Pm. CNOm inside increased while CNOm outside decreased with Pm. At Pm =40 cm/s, CNOm inside and CNOm outside were similar to each other and both increased with Hct, indicating that extracellular diffusional resistance provides a significant transport barrier for NO at higher RBC membrane permeability. At Pm = 0.0415 cm/s, estimated CNOm outside were much higher than CNOm inside and both were independent of Hct for Hct ≥ 15%, suggesting that at lower RBC membrane permeability, for Hct ≥ 15%, RBC permeability is a major barrier for resistance for NO transport. Insights from this study may prove useful in understanding interaction of NO with RBC. Supported by AHA 0530050N & R15 HL087287 grants.
Acupuncture stimulates eNOS translocation and NO production along meridians
Authors: Duran Ricardo G , Korayem Adam H , Duran Walter N , Kim David D
Published: June 2021

The meridian theory is the central theory of many complementary and alternative medicines and it is the backbone of acupuncture and Oriental medicine. However, the biological mechanisms of the meridian system remain elusive. We hypothesized that acupuncture stimulates endothelial nitric oxide synthase (eNOS) translocation and NO production via the meridian communication system. We applied different time sequence electroacupuncture (DTS‐EA) on the Stomach 36 point (ST‐36) and measured periarteriolar nitric oxide concentration ([NO]) in the hamster cheek pouch. We found that 1) DTS‐EA increased [NO] in hypertensive hamsters, but not in healthy control animals; 2) the second EA treatment caused faster rise in [NO] compared to the first EA treatment; 3) importantly, DTS‐EA increased total eNOS and phosphorylated eNOS in cheek pouch and stomach (both in the stomach meridian), but not in liver (a non‐stomach meridian organ); 4) DTS‐EA enhanced the preferential translocation of eNOS to the Golgi‐enriched fraction, a location consistent with the onset of vasodilation. Our data confirm that activation of eNOS is an important signaling mechanism stimulated by acupuncture, and demonstrate that acupuncture signals are transmitted via specific meridians. (Supported by NIH grants 5R21 AT004609 and 5R01 HL070634).
Benfotiamine: a novel cardioprotective agent that blunts hyperglycemia‐induced cardiac dysfunction
Authors: Mapanga Rudo Fiona , Essop M Faadiel
Published: June 2021

Increased blood glucose levels during and after an ischemic insult amplify the risk of future cardiovascular disease events. We previously found that hyperglycemia triggers oxidative stress that enhances flux through the hexosamine biosynthetic pathway (HBP), leading to increased myocardial apoptosis. Since benfotiamine (BFT) decreases HBP flux, we hypothesized that it attenuates cardiac cell death and enhances contractile function in response to ischemia‐reperfusion under hyperglycemic conditions. We perfused isolated rat hearts with Krebs Henseleit buffer ± 11 mM (control) and 33 mM glucose (hyperglycemia), respectively (Langendorff system), for 90 min, followed by 30 min global ischemia and 60 min reperfusion. 100 μM BFT was administered immediately after ischemia for 20 min of reperfusion. Separate experiments were performed to assess infarct size with BFT treatment. This study found decreased functional recovery of heart function under hyperglycemic conditions vs. matched controls (p<0.05) following ischemia‐reperfusion. BFT treatment significantly improved the heart's functional recovery and reduced infarct sizes by 21 ± 3.2% under hyperglycemic conditions (p<0.05 vs. controls). Our study demonstrates that benfotiamine is a promising cardioprotective agent that may ultimately benefit pre‐and full‐blown diabetic patients suffering from cardiovascular disease complications.
Mechanisms of Skeletal Muscle Contraction‐Induced Rapid Vasodilation in Humans: Role of Vascular Hyperpolarization
Authors: Kirby Brett S , Crecelius Anne R , Richards Jennifer C , Garcia Leora J , Luckasen Gary J , Larson Dennis G , Dinenno Frank A
Published: June 2021

A monophasic increase in skeletal muscle blood flow is observed following a brief single forearm contraction in humans. Evidence in dogs indicates an obligatory role of vasodilation via smooth muscle hyperpolarization, and recent in situ data suggest that activation of Na+/K+ATPase and KIR channels during muscle contraction contributes to this vasodilation. We tested the hypothesis that inhibition of Na+/K+ATPase and KIR channels via intra‐arterial infusion of ouabain and barium chloride (BaCl2), respectively, and thus vascular hyperpolarization decreases contraction‐induced rapid vasodilation in the human forearm. We measured forearm blood flow (Doppler ultrasound) and calculated vascular conductance 10s prior to, and for 30s after a single 1‐s dynamic forearm contraction at 10, 20, and 40% maximum voluntary contraction (MVC) in 12 young adults. Inhibition of hyperpolarization reduced total forearm vasodilation by ~74, 62, and 55% (10, 20, and 40% MVC, respectively). Additional blockade of nitric oxide synthase and cyclooxygenase further attenuated the response to a total reduction of ~80% for all intensities. The present study demonstrates a significant contribution of vascular hyperpolarization to contraction‐induced rapid vasodilation, and that collective blockade of nitric oxide, prostaglandins, and vascular hyperpolarization nearly abolishes this phenomenon in humans.

HL102720
Curcumin suppresses NOD1‐mediated chemotactic genes expression in 3T3‐L1 adipocytes
Authors: Hu Pan , Zhao Ling
Published: June 2021

We have reported that activation of Nucleotide‐Oligomerization Domain containing protein 1 (NOD1) induced mRNA expression and protein secretion of monocyte chemotactic protein‐1 (MCP‐1) in adipocytes. Here we study 1) whether NOD1 activation induces up‐regulation of other chemotactic genes [MCP‐2, MCP‐3, and serum amyloid A3 (SAA3)]; 2) the roles of other cell components of adipose tissue, pre‐adipocytes and macrophages in NOD1‐ mediated effects; 3) whether curcumin, a known anti‐obesity and anti‐inflammatory phytochemical, inhibits NOD1‐mediated chemotactic genes expression. Our results show that in addition to MCP‐1, NOD1 activation also time‐ and dose‐dependently induces MCP‐2, MCP‐3 and SAA3 mRNA expression in 3T3‐L1 adipocytes and this induction is mediated by enhanced promoter activity as well as mRNA stability. No significant changes in mRNA expression of the chemotactic genes are found in pre‐adipocytes and macrophages upon NOD1 activation. Moreover, curcumin suppresses NOD1‐mediated MCP‐1 up‐regulation in adipocytes, partly due to its suppression on MCP‐1 promoter activity induced by NOD1 activation. Our studies suggest a novel role of NOD1 activation in adipose inflammation in obesity and support the anti‐inflammatory effects of curcumin in obesity prevention and treatment. The work is supported by The University of Tennessee faculty start‐up funds to LZ.
Full‐length HGF mRNA persists after genetic recombination in mice
Authors: Nejak-Bowen Kari N , Orr Anne V , Bowen William C , Michalopoulos George K
Published: June 2021

Hepatocyte growth factor (HGF) has been shown to be indispensible for liver regeneration because it serves as the main mitogenic stimulus driving hepatocytes to proliferate. We hypothesize that ablating HGF in adult mice would have a deleterious effect on liver homeostasis. HGFflox/flox mice were bred to mice transgenic for either Mx‐cre or Tam‐cre and then treated with a regimen of either p(I):p(C) or tamoxifen, respectively. Analysis of liver genomic DNA from animals 10 days after treatment showed that a majority (70–80%) of the DNA underwent cre‐induced genetic recombination. Intriguingly, however, analysis by RT‐PCR showed the presence of both the unrecombined and recombined form of HGF mRNA after treatment. HGF protein levels were also unchanged between control and treated (KO) mice. Separation of liver cell populations into hepatocytes and non‐parenchymal cells showed equal recombination of HGF genomic DNA in both cell types. The presence of unrecombined HGF mRNA persisted in the liver even after 2/3 partial hepatectomy (PH). HGF protein expression in control and KO livers after PH was equivalent, correlating with the persistence of full‐length HGF mRNA. Additionally, ablation of HGF genomic DNA had no impact on proliferation during regeneration. Thus, HGF mRNA may be persisting due to long‐term mRNA stability or de novo production of full‐length HGF from unrecombined stellate cells.
The use of gaze tracking to quantify learning
Authors: Zumwalt Ann C , Wice Mitchell , Bergethon Peter
Published: June 2021

The long‐term goal of this ongoing study is to quantify how individuals of varying levels of expertise interact with visual data, to ultimately establish an objective assessment of expertise and learning. The current study investigates how gaze patterns shift as an individual becomes more familiar with an image. Specifically, this study tests the hypothesis that as an individual sees an image repeatedly they will examine the image for less time, and will fixate on fewer locations on the image for relatively longer durations of time. Subjects (n=13) were shown a random series of 100 images in which four images were repeated ten times and the rest of the images were seen once. Subjects were instructed to freely examine each image and advance to the next at any time, and their gaze patterns were recorded as they viewed the images. Outcome variables were the duration of time the subject viewed an image, number of gaze fixations, and the durations of those fixations. When the data were pooled the variation in each variable was too extensive to reveal clear trends from the first through tenth viewing, indicating the need for a larger sample size. However, the study did reveal that some individuals behaved as predicted, whereas others did the opposite: with each repeated view of an image, they investigated it for more time. While the sample size was inadequate to confidently quantify these possible trends, follow‐up studies are ongoing.

Grant Funding Source: n/a
Long non‐coding RNAs in epigenetic regulation
Authors: Shiekhattar Ramin
Published: June 2021

Spatial and temporal regulation of gene expression is achieved through the instruction provided by the distal transcriptional regulatory elements known as enhancers. How enhancers transmit such information to their targets has been a subject of intense investigation. Recent advances in high throughput analysis of the mammalian transcriptome have revealed a surprising result indicating that a large number of enhancers are transcribed to non‐coding RNAs. While long non‐coding RNAs were initially shown to confer epigenetic transcriptional repression, recent studies have uncovered a role for a class of such transcripts in gene‐specific activation often from distal genomic regions. We will discuss our recent findings on the role of long non‐coding RNAs in transcriptional regulation with an emphasis on new developments on the functional links between long non‐coding RNAs and enhancers.
Feeding Soy Protein Isolate Prevents Impairment of Bone Acquisition by Western Diets as a Result of Insulin Signaling in Bone
Authors: Zhang Jian , Lazarenko Oxana P. , Badger Thomas M. , Ronis Martin J.J , Chen Jin-Ran
Published: June 2021

Excessive consumption of high fat/high cholesterol “Western” diets during postnatal life results in increased energy intake, development of obesity and systemic insulin resistance. However, how this diet impairs bone development and remodeling is not well understood, and no effective dietary interventional strategy has emerged. In the current study, male rats pair‐fed a Western diet (WD) for 7 wks start from post natal day 20 displayed obesity with significantly increased gonadal and abdominal fat mass and body weight compared to control (p<0.05). WD rats had increased in bone marrow adiposity, insulin resistance and impaired glucose tolerance. Supplementation of soy protein isolates (SPI) ameliorated the effects of WD. Inhibition of WD‐induced impairment of bone acquisition by SPI was associated with increased undercarboxylated osteocalcin secretion and increased IRS1/Akt insulin signaling in osteoblasts. SPI normalized non‐esterified free fatty acid (NEFA) levels in WD rats. These results suggest that the effects of SPI diet on metabolism and body composition in WD rats are due to increased of secretion of active osteocalcin and stimulated insulin signaling in bone. Supported in part by ARS CRIS #6251‐51000‐005‐03S.

Grant Funding Source: USDA / ARS CRIS
Dissemination of an Effective Weight Management Program for Mexican American Children in Schools
Authors: Palcic Jennette , Johnston1 Craig A , Breslin Whitney , El-Mubasher Abeer , Foreyt John P
Published: June 2021

Baseline

The rates of child obesity are epidemic in the United States, and Mexican American children are at particular risk. We have found an intensive, multi‐component, school‐based, weight management intervention to be efficacious at reducing standardized body mass index (zBMI) in overweight children. Our next step was to examine the ability of PE teachers to implement this program (dissemination condition (DS)).

Objective

The current study sought to examine the efficacy of our intervention led by a PE teacher.

Methods

Students were randomized to either a research staff led intervention (SLI) or the DS condition. The SLI received classes 3 days a week. Students in the DS condition received the same curriculum but led by a PE teacher. Repeated measures ANOVA was used to evaluate weight changes using zBMI from baseline to 3, 6, and 12 months.

Results

Students in both conditions significantly decreased their zBMI at 3, 6, and 12 months (F=50.7, p<.0001). Independently, both conditions demonstrated a significant decrease in zBMI at 3, 6, and 12 months (ILI: F=46.0, p<.001 and DS: F=12.8 p<.001). The SLI condition significantly out performed the DS condition (F=5.34, p<.01).

Conclusions

Overall, the results suggest that our intensive school‐based weight management program can be disseminated. However, there does still appear to be an advantage to the intervention when implemented by health professionals.
Food Aid Programs: Sustaining Impacts after Program Exit
Authors: Rogers Beatrice Lorge , Coates Jennifer , Houk Katie , Kegode Elizabeth
Published: June 2021

One of the persistent challenges to food assistance programs is to ensure that the positive impacts achieved by the programs are sustained after the programs end. The present study reviews experiences of three countries with PL480 Title II programs in the two years after the food was withdrawn: Bolivia, Kenya, and Honduras. Qualitative data were collected at exit and one year later. Key factors predicting sustainable impacts include visible positive change in the beneficiary communities; an assured source of resources from profits, user fees, or government budgets (depending on government resources and capacity). Both visible benefit and assured resources were necessary for sustainability. Programs were more sustainable if the participants were trained in both technical and managerial aspects of their activities. Gradual exit produced more sustainable activities and impacts than more abrupt departure. The use of food as an incentive for participation risks creating unsustainable expectations among beneficiaries. The study will conclude with a third round of qualitative and quantitative data collection in the summer of 2011. Funding was from USAID Food for Peace through the FANTA2 Program.
Regulation of antioxidant responses in prostate cancer: Nrf2‐ dependent and independent effects of lycopene
Authors: Gong Xiaoming , Marisiddiah Raju , Wiener Doris , Rubin Lewis P
Published: June 2021

Control of intracellular reactive oxygen species (ROS) concentrations is critical for cancer cell survival. Evidence from epidemiological, experimental studies and clinical trials suggests that lycopene may modify the risk or delay progression of prostate cancer. However, the molecular mechanisms involved are still unclear. The aim of this study is to determine the effects of lycopene on endogenous ROS and antioxidant responses in normal prostatic epithelial cells and in prostate cancer (PC) cells. Here, we show that levels of ROS generation were much higher in PC cells than in normal prostate cells (PrEC). Lycopene reduces ROS levels in a dose‐dependent manner in androgen‐dependent LNCaP but not in androgen‐independent DU145 and PC3. Intracellular levels of reduced GSH were highest in DU145 and associated with abundant levels of nuclear (NF‐E2)‐related factor 2 (Nrf2). Lycopene treatment did not significantly affect cell GSH or nuclear Nrf2 in either normal or PC cells. Using a cell‐based ARE‐reporter assay, we found that lycopene treatment increased ARE transcriptional activity in LNCaP but decreased ARE activity in DU145 cells. These findings indicate that lycopene may impair the PC cell survival machinery through regulating intracellular ROS and antioxidant response. Certain effects appear to be independent of Nrf2 signaling. (Supported by NIH, Muma family Endowment, USF)
Saturated fatty acids activate TLR‐mediated proinflammatory signaling pathways
Authors: Hwang Daniel , Huang Shurong , Rutkowsky Jennifer M. , Snodgrass Ryan G. , Ono-Moore Kikumi D. , Schneider Dina A. , Newman John W. , Adams Sean H.
Published: June 2021

Toll‐like receptor 4 (TLR4) and TLR2 were shown to be activated by saturated fatty acids (SFAs) but inhibited by docosahexaenoic acid (DHA). However, one report (ATVB 11:1944, 2009) suggested that SFA‐induced TLR activation in cell culture systems is due to contaminants in BSA used for conjugating fatty acids. This report casted doubt about proinflammatory effects of SFAs. Our studies herein demonstrate that sodium palmitate (C16:0) or laurate (C12:0) without BSA conjugation induced phosphorylation of IκBα, JNK, ERK, and NFκB p65 and TLR target gene expression in THP1 monocytes or RAW264.7 macrophages, respectively when cultured in low FBS (0.25%) medium. C12:0 induced NFκB activation through TLR2 dimerized with TLR1 or TLR6, and through TLR4. Since BSA was not used in these experiments, contaminants in BSA have no relevance. Unlike suspension cells (THP‐1), BSA conjugation is required for C16:0 to induce TLR target gene expression in adherent cells (RAW264.7). BSA‐conjugated C16:0 transactivated TLR2 dimerized with TLR1 or TLR6, and through TLR4 as seen with C12:0. These results and additional studies with LPS sequester polymixin B and MyD88−/− macrophages indicated that SFA‐induced activation of TLR2 or TLR4 is a fatty acid‐specific effect, but not due to contaminants in BSA or fatty acid preparations. (USDA‐ARS‐WHNRC Program Funds and NIHDK 064007)
The Mammalian O‐Mannosylation Pathway: Protein Substrates and Functional Glycans
Authors: Wells Lance
Published: June 2021

O‐linked mannose modification imparts essential functions to targeted glycoproteins. The protein substrates, enzymes, and glycan products of the O‐mannosylation pathway underlie a variety of cellular processes and their perturbation contribute to diverse pathophysiological conditions including viral‐host interactions, tumor metastasis, axon guidance in the brain, muscle integrity, and cardiac function. Despite the broad physiologic and pathophysiologic impact of the O‐mannosylation pathway, very little is known regarding the key steps in the production of functional O‐mannose glycans. This talk focuses on identification strategies for the protein substrates for O‐mannosylation in mammalian brain and muscle, efforts to delineate the unknown enzymes in the pathway, and the assignment of the functional Omannose initiated glycan(s) structures. Towards these goals, several key tools including mass spectrometry‐based glycoanalytics, recombinant enzymes and protein expression systems, chemoenzymatic tagging methods, complex glycopeptide synthesis, and anti‐O‐glycan antibodies will be discussed. These research efforts will be highlighted that focus on providing new understanding regarding the mechanisms by which defects in the O‐mannosylation biosynthetic pathway contribute to multiple disease processes.
Diagnosis and monitoring of mucopolysaccharidoses using disease‐specific non‐reducing end carbohydrate biomarkers
Authors: Esko Jeffrey D. , Lawrence Roger , Lamanna William C. , AL-Mafraji Kanar , Boons Geert-Jan , Dierks Thomas , Brown Jillian R. , Crawford Brett E.
Published: June 2021

A significant need exists for improved biomarkers for differential diagnosis, prognosis and monitoring of therapeutic interventions for mucopolysaccharidoses (MPS), inherited metabolic disorders that involve lysosomal storage of glycosaminoglycans. Here, we report a simple reliable method based on the detection of abundant non‐reducing ends of the glycosaminoglycans that accumulate in cells, blood, and urine of MPS patients. In this method, glycosaminoglycans were enzymatically depolymerized releasing unique mono‐, di‐, or trisaccharides from the non‐reducing ends of the chains. The composition of the released mono and oligosaccharides depends on the nature of the lysosomal enzyme deficiency, and therefore they serve as diagnostic biomarkers. This strategy has been used to validate the cause of secondary dermatan sulfate storage in Sanfilippo disease by matching the non‐reducing end of this glycosaminoglycan with deficient iduronate‐2‐sulfatase activity. Furthermore, non‐reducing end analysis has been used to identify a novel MPS disorder. These findings underline the potential for this technology to aid in diagnosis, therapy and identification of new pathological mechanisms in lysosomal storage diseases as well as the discovery of novel MPS disorders.
Moral and sexual education as a factor in the prevention of sexually transmitted infections (STIs) and HIV infection among inmates of SOS «Astanaˈs Village
Authors: Dalenov Erbolat , Kalashnikova Anastasia , Ilderbayev Oralbek
Published: June 2021

In Astana city the sociological knowledge monitoring has been held to determine the level of awareness and evaluation of risk behavior factors before and after the preventive activities among young people. To this end, the survey was conducted among young people: the main group included 42 respondents‐students SOS “Childrenˈs Village Astana” (14 –18 years), the control group‐52 pupils (17 years). After questioning to determine the baseline level of knowledge, special seminar on the prevention of STIs/HIV was held in the control group, but in the main group for 7 years the subject of moral and sexual education is taught. As a result of study, before preventive activities: the level of awareness in the main group was lower than in the control group, 100% and 62.8%, respectively. The risk factors are low in both groups, as noted above, due to the fact that the majority of respondents did not have sexual relations. Analysis of risk behavior showed that this group does not have reported growing numbers of young people entered into sexual relationships, and among students the number of respondents who entered into sexual relationships has increased‐up to 14 teenagers in connection with the increased number of people with sexual debut at the age of “16–18 years”.
A Survey of Memorial Ceremonies in United States Anatomy Programs
Authors: Jones Trahern W , Nwojo Macaulay , Pawlina Wojciech , Lachman Nirusha
Published: June 2021

Anatomy programs that incorporate dissection of donated human tissue often hold ceremonies of gratitude and reflection for body donors. The content of these ceremonies often include learners’ reflections on mortality, respect, altruism, and personal growth told through various humanities modalities. The task of planning is usually faculty‐ and student‐led with participation from other health care students. Objective information on current memorial ceremonies for body donors in anatomy programs in the United States appears to be lacking. The number of programs in the United States that currently plan these ceremonies and information on trends in programs undertaking such remain unknown. Anatomy program directors throughout the United States were issued a brief, voluntary, thirteen‐item questionnaire on memorial ceremonies held at each institution. Qualitative data regarding the number of anatomy programs which hold their own memorial ceremonies, the content of such ceremonies, how they are planned, level of confidentiality maintained, and the types of participants who attend these events will be presented. This information may serve as a guide to institutions considering initiating a ceremony of their own.

Grant Funding Source : Department of Anatomy
The relative efficiency of homology‐directed repair has distinct effects on proper anaphase chromosome separation
Authors: Laulier Corentin , Cheng Anita , Stark Jeremy Michael
Published: June 2021

Since several tumors show deficiency in homologous recombination (HR), it is important to understand how HR deficiency can lead to cancer. Notably, cells deficient in HR show increased chromosomal instability (CIN), which is a hallmark of cancer. To examine the role of HR in limiting CIN, we considered that HR defects could cause at least two types of anaphase abnormalities: DNA linkages and loss of spindle bipolarity. DNA linkages can be visualized using DAPI, and/or by immunostaining of the helicase PICH, which can form ultrafine bridges (UFBs). We examined HEK293 cells with inducible expression of dominant‐negative alleles of the strand exchange factor RAD51: RAD51K133R and RAD51K133A, which cause a mild and severe HR defect, respectively. RAD51K133R causes a rapid increase in UFBs, but does not cause loss of spindle polarity. Similarly, BRCA1 or BRCA2 mutant cells displayed increased UFBs frequencies. In contrast, RAD51K133A expression causes UFBs, DAPI bridges, multipolar anaphases, and cell death. Based on these findings, we propose that mild HR defects allow proliferation of cells that are prone to CIN due to defective chromosome separation during anaphase, thus promoting tumor development. In contrast, severe HR defects lead to multipolar anaphases that result in cell death. We are currently examining anaphase abnormalities in cancer cells to develop biomarkers for HR deficiency.
The Desmosomal Protein, PERP, is Required for the Pro‐ Inflammatory Response to Salmonella Typhimurium Infection
Authors: Hallstrom Kelly , Srikanth C. V. , Agbor Terence , Demma Zachary , McCormick Beth A
Published: June 2021

Salmonella enterica serovar Typhimurium (S. Typhimurium) is one of several Salmonella strains responsible for over 40,000 cases of Salmonella‐induced gastroenteritis in the United States yearly. Salmonella invade the apical surface of intestinal epithelial cells and trigger inflammation through the activity of type three secreted effectors. One such effector, SipA, activates signal transduction cascades that ultimately lead to the recruitment of neutrophils to the site of invasion. Previous studies showed that SipA acts extracellularly to induce neutrophil migration. Recent studies from our lab demonstrated that SipA requires processing by CASPASE‐3 to be fully functional, and that SipA induces release of activated CASPASE‐3 at the mucosal surface. In order to understand how SipA triggers these events without being translocated into the cell, we sought to identify an interacting host protein.

We have determined that the desmosomal protein, PERP, is a binding partner of SipA. We have further observed that PERP accumulates at the apical surface during Salmonella infection in a SipA‐dependent manner, and that loss of PERP via siRNA knockdown results in decreased neutrophil migration during Salmonella infection. Additionally, we see reduced levels of activated CASPASE‐3 when PERP is knocked down. These results suggest PERP plays a crucial role in the pro‐inflammatory response to Salmonella infection.

Funding Sources: NIH, NIDDK, Crohn's and Colitis Foundation of America
Kruppel‐like transcription factor KLF8 regulates of early adipocyte differentiation
Authors: Kim Hyo Jung , Choi Hyeonjin , Lee Yoo Jeong , Lee Hye-min , Yu Jung Hwan , Kim Jae-woo
Published: June 2021

KLF8 is a zinc‐finger transcription factor known to play an essential role in cell cycle and differentiation. However, its physiological roles and functions in 3T3‐L1 cells remain relatively unclear. In the present study, we showed that KLF8 acts as a key regulator controlling adipocyte differentiation. In 3T3‐L1 preadipocytes, KLF8 expression was induced at an early stage of differentiation which was followed by expression of PPAR¥ã2 and C/EBP¥á. The mRNA level of KLF8 dramatically increased at 36 h and 48 h during differentiation and the protein level also gradually increased after 36 h. This induction and differentiation was attenuated significantly by the presence of small interfering RNA against KLF8, whereas overexpression of KLF8 induced 3T3‐L1 differentiation. Furthermore, overexpression of KLF8 induced PPAR¥ã2 and C/EBP¥á promoter activity and the direct binding site in C/EBP¥á promoter exists between (−205) and (−89) region. Taken together, this study provides KLF8 as a key component of the transcription factor network that controls the terminal differentiation during adipogenesis.

This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MEST) (No. 2011‐0030711 and 2011‐0015665) and by Korean Research WCU grant (R31‐2008‐000‐10086‐0) from the Korean Ministry of Education, Science, and Technology.