bluuebunny/crossref_metadata_2025 huggingface dataset:
Design and Synthesis of a Dual‐Ligand Inhibitor for Acetyl‐CoA Carboxylase
Authors: Silvers Molly A , Taylor Carol M , Waldrop Grover L
Published: June 2021

An urgent demand for the development of new antibiotics has been caused by the increase in drug‐resistant pathogenic bacteria. Targeting the enzymes involved in fatty acid synthesis offer a solution for developing new inhibitors for drug discovery. A novel approach is to focus on the multifunctional enzyme acetyl‐CoA carboxylase, which catalyzes the first committed step in fatty acid synthesis. The bacterial form of the enzyme consists of three proteins: biotin carboxylase, biotin carboxyl carrier protein, and carboxyltransferase. By joining known inhibitors for biotin carboxylase and carboxyltransferase together with a linker via chemical synthesis, we will generate a dual‐ligand inhibitor with the potential to halt fatty acid synthesis in pathogenic bacteria. The expectation for this dual‐ligand inhibitor is increased potency relative to the component inhibitors. Moreover, this work may lead to an antibacterial agent that minimizes or defers the development of resistance by pathogenic bacteria. This work is funded in part by National Science Foundation (MCB‐0841134).
An interactive and probabilistic strategy for visualizing linear biological sequence motifs
Authors: Schwartz Daniel , O'shea Joseph P. , Quader Saad A. , Mingir Ahmet C. , Church George M. , Chou Michael F.
Published: June 2021

Short functional patterns of nucleotides or amino acids, also known as motifs, are found ubiquitously on DNA, RNA, and protein sequences. These motifs often serve as molecular interaction beacons, thus providing a degree of specificity between the macromolecular interactions found in the seemingly stochastic cellular environment. To date, the most common means of visualizing biological sequence motifs has been through sequence logos (Figure 1, top). While sequence logos have been an effective means of motif visualization for a subset of biological patterns, several limitations, including the assumption of uniform background probabilities and the inability to visualize underrepresented sequences, have limited their effectiveness – particularly in the field of post‐translational modifications. Here we present the pLogo, or probability logo, a interactive and statistically based representation of sequence motifs (Figure 1, bottom). pLogos share a number of unique features including i) residue/nucleotide heights which approximate their statistical significance, ii) underrepresented residues/nucleotides which are depicted below the x‐axis, and iii) conditional probabilities which can be easily manipulated and visualized by users. A pLogo web server will be created at http://plogo.uconn.edu. This work was funded in part by grants from the UConn Research Foundation and Genomes to Life from the US DoE.
Calcium Promotes Oxidative Stress and Metabolic Dysfunction in Lipotoxicity
Authors: Egnatchik Robert A. , Leamy Alexandra K. , Jacobson David A , Young Jamey D.
Published: June 2021

Elevated saturated fatty acids (SFA), but not monounsaturated fatty acids, induce apoptosis in a variety of cell types. In H4IIEC3 cells, lipoapoptosis is characterized by decreased ER calcium, increased TCA cycle flux, and the accumulation of reactive oxygen species (ROS). Our hypothesis is SFA‐initiated ER stress results in a net flux of calcium to the mitochondria, which increases oxidative metabolism, leading to ROS‐dependent apoptosis. Our objective is to define the mechanism of deregulated mitochondrial metabolism in promoting lipotoxicity defined by ROS accumulation and O2 uptake as proxy of mitochondrial activity. Scavenging SFA‐induced ROS with antioxidants (vitamin E, n‐acetyl cysteine) reduced apoptosis but not increased mitochondrial activity. Combining SFA and phenformin to inhibit Complex I activity eliminated reduced the lipotoxic phenotype. Additionally, cells were treated with the intracellular calcium chelator BAPTA to examine the role of depleted ER calcium in apoptosis. BAPTA treatments attenuated SFA‐induced ROS accumulation, oxygen consumption, caspase activation, and cell death. Our results suggest that SFA‐induced lipoapoptosis in hepatic cells is dependent on calcium‐stimulated mitochondrial activation and ROS accumulation. This work is funded by NSF Career Award 0955251 (Young).
Characterization of thyroid hormone receptor export pathways
Authors: Nelson Hallie Nicole , Allison Lizabeth A
Published: June 2021

Thyroid hormone receptor alpha (TRα) and beta (TRβ) are critical in many aspects of metabolic control and development. TRα can use a CRM1‐dependent or a CRM1‐independent pathway for nuclear export. To determine if exportin 5 mediates TR's export by the latter pathway, Myc‐tagged exportin 5 was over‐expressed in HeLa cells. The distribution and transcriptional activity of GFP‐TRα or TRβ was analyzed by fluorescence microscopy and CAT‐ELISA, respectively. TR is primarily nuclear at steady‐state, but TR had a more cytoplasmic distribution when exportin 5 was overexpressed. In addition, CAT reporter gene expression under control of a thyroid hormone response element was markedly decreased when exportin 5 was over‐expressed, indicating less TR was present in the nucleus. TR has multiple nuclear export sequences (NESs) in the ligand‐binding domain. We tested the helix 12 NES (NES‐H12) and the NES located in the 3rd and 6th helices (NESH3/ H6) on their direct interactions with exportin 5. TR with a NES‐H12‐disrupting mutation had a more cytoplasmic distribution when exportin 5 was over‐expressed, suggesting that exportin 5 utilizes NES‐H3/H6 during export. By determining the nuclear export mechanism of TR by exportin 5 and other exportins, further knowledge into regulation of thyroid hormone‐responsive gene expression will be gained. This work was funded in part by NSF MCB 10144844 and NIH 2R15DKO58028–03 to L.A.A.
Diabetes complication‐related healthcare utilization among low‐income older Medicare beneficiaries in need of food assistance
Authors: Sattler Elisabeth Lilian Pia , Lee Jung Sun , Bhargava Vibha
Published: June 2021

Little is known about diabetes management in low‐income older adults who are at high risk of diabetes and poor access to food assistance and healthcare. This study described factors associated with diabetes complication‐related healthcare utilization in low‐income older adults in need of meals services using data from the GA Advanced POMP and Medicare claims. Participants were Medicare beneficiaries with a diagnosis of diabetes who requested meals services in GA in 2008 (n=246, mean age 74.1 ±7.8 years, 66.7% white, 76.4% female). Healthcare utilization was measured based on previously identified diabetes complications for 4 different types of medical services. About 81% used services for 1+ complication (range 0–18; median 3.0). Cardiovascular (62.6%), neuropathic (35.8%), and nephropathic (30.5%) complications were most prevalent. Age <75 years, being male, poor self‐reported health, and requesting home delivered meals were associated with a higher level of diabetes complication‐related healthcare utilization. In multivariate analyses, only age <75 years remained a significant factor (OR 1.9; 95% CI 1.2, 3.1). As participants <75 years were more likely food insecure and receiving subsidies for healthcare and medications, more research is warranted to better understand their healthcare needs, access, utilization, and outcomes related to diabetes management. This study was funded by UGA and GA DAS.
Maternal iron status and depressive symptoms in rural Bangladesh
Authors: English Laural Kelly , Murray‐Kolb Laura E , Chang Stephanie , Hamadani Jena , El Arifeen Shams , Baqui Abdullah , Black Robert E
Published: June 2021

Iron deficiency and maternal depression are prevalent in developing countries like Bangladesh. Our aim was two‐fold: 1) determine the relationship between iron status and maternal depressive symptoms; 2) assess the Center for Epidemiological Studies Depression Scale (CESD) in women of rural Bangladesh. Mother‐infant dyads taken from a larger RCT (Chang et al., 2010) to improve diarrhea and anemia in rural Bangladesh were analyzed. Depressive symptoms were collected at baseline (n=113), 3 mo (n=319), and 6 mo (n=328). Maternal ferritin and transferrin receptor were collected between baseline and 3 mo (n=307) and at 6 mo (n=108). Factor analysis of CESD structure resulted in a 3‐factor solution: interpersonal difficulty/somatic inhibition, depressed affect, and well‐being. We found a 70.3% prevalence of high depressive symptoms (CESD ≥ 16) but low prevalence (11.2%) of iron depletion (ferritin < 20 ng/mL). Nonetheless, iron depleted (ID) women had higher depressive symptoms than iron sufficient (IS) women at every time point (ANOVA, p<0.04). Multivariate regression adjusted for covariates (child age & sex, WAZ, assets), revealed that body iron at 6 mo (assessed with Cook's equation) was related to the “depressed affect” factor scores at 6 mo (p<0.048). This relationship between iron depletion and CESD symptomatology warrants validation of CESD factor structure and cutoff point in Bangladesh.

Grant Funding Source: USAID to the Johns Hopkins Global Research Activity Cooperative Agreement
Maternal vitamin D status and inflammation across adolescent pregnancy
Authors: Akoh Christine , Pressman Eva , Cooper Elizabeth , Queenan Ruth Anne , O'Brien Kimberly
Published: June 2021

Vitamin D is known to play a role in both innate and adaptive immunity, but the role of vitamin D in the inflammatory process during pregnancy is unclear. The purpose of this study was to examine associations between vitamin D status and inflammation across pregnancy using extant biospecimens obtained from a cohort of 168 pregnant adolescents (≤18 y). In this longitudinal study, serum was collected from adolescents at mid‐gestation (MG~26 weeks), delivery, and from cord blood; 66.1% of teens were African American, 33.9% Caucasian, and 24.1% Hispanic. Half of the teens and 60% of their newborns were 25(OH)D insufficient (≤20 ng/mL). Analyses are on‐going to measure inflammatory cytokines (IL‐6, IL‐10, TNF‐α, and CRP) in archived maternal and cord serum. To date maternal 25(OH)D at MG was positively correlated with TNF‐α in cord blood (p=0.0284, n=89). Maternal 1,25(OH)2D was positively associated with both TNF‐α (p=0.0364, n=96) and CRP at MG (p=0.0305, n=86) but was significantly negatively correlated with TNF‐α at delivery (p=0.0004, n=92). These results suggest that vitamin D may impact inflammatory activity in pregnant adolescents and their infants at birth.

Grant Funding Source: USDA AFRI 2012–67017‐30216
Dietary patterns and weight change in older adults ≥ 75 years old: What's the relationship?
Authors: Hsiao Pao Ying , Coffman Donna L. , Wood G. Craig , Mitchell Diane C. , Still Christopher D. , Jensen Gordon L. , Hartman Terryl J.
Published: June 2021

Weight change is a common health concern among the elderly, yet little is known about the relationship between weight change and dietary patterns (DP) in older adults, especially the very old (≥ 75 y). This investigation examined the association between DP and 5‐ year weight change in community‐dwelling older adults (n=270; mean ± SD age: 78.6±3.9 y) of the Geisinger Rural Aging Study, a longitudinal cohort study. Using baseline 24‐hour dietary recall data, cluster analysis was used to derive ‘Sweets and dairy’ (SD), ‘Western’ (W) and ‘Health‐conscious’ (HC) DP. Weight change data was extracted from electronic medical records and was characterized using three methods: 1) overall 5‐y weight change, 2) 10 pounds gained or lost (y/n) and 3) 10% gained or lost (y/n). Kaplan‐Meier plots and Cox proportional hazards regression were used to examine the association between DP and weight change. Mean overall weight change was not significantly different across DP: −5.56 (all), −7.2, −4.8 and −3.3 pounds for the SD, the W, and the HC DP, respectively (p=0.17). In gender‐stratified analyses, compared to the HC DP, females characterized by the SD and W DP, were three and two times more likely to lose 10 pounds (Hazard ratio (95% CI): SD=3.2 (1.5–6.7); W=2.4 (1.1–5.6)), respectively. Findings suggest that a HC DP may be associated with weight stability in females, which provides additional impetus for promoting healthy DP in older women.

Grant Funding Source: USDA #1950–51530‐010–02G
Maternal metabolic changes with dietary intake of blueberry during pregnancy and lactation predispose adult progeny to lower mammary tumor growth rate
Authors: Montales Maria Theresa E , Melnyk Stepan , Prior Ronald L , Simmen Rosalia CM
Published: June 2021

We have shown the lower growth rates of tumors that developed from Wnt1‐transgenic (Tg) offspring of dams consuming whole blueberry powder (3% BB) during pregnancy and lactation, compared to those of control (Casein) dams. Dietary exposure at post‐weaning through lifetime did not mimic the effects of early BB exposure. The basis for maternal BB effects on mammary tumor growth was examined in dams exposed to BB or control diet as young adults (beginning at age 8 weeks) and throughout pregnancy and lactation. Dams at weaning were evaluated for growth and metabolic parameters. BB‐fed dams had lower abdominal and retroperitoneal fat pad weights than control‐fed dams, although their body weights did not differ. BB dams also displayed increased insulin sensitivity (lower blood glucose, serum insulin, HOMA) and higher serum glutathione and methionine levels. Isolated epithelial cells from pre‐neoplastic mammary tissues of Tg offspring exposed to maternal control and BB diets were analyzed for percentage of basal (CD29highCD24+) and luminal progenitor (CD29lowCD24+) populations. Maternal BB exposure (relative to control) resulted in lower percentage of the luminal subpopulation (6.4% vs 21%), which comprise the tumor bulk in Tg tissues. Results implicate the influence of the favorable metabolic and anti‐oxidative maternal environment with BB diet on developing mammary glands for the improved mammary tumor outcome.

Grant Funding Source: USDA‐CRIS 6251–51000‐005–004S, Arkansas Biosciences Institute
Fiber decreases the antioxidant capacity of phenolic acids in an alkaline milieu in vitro
Authors: Palafox‐Carlos Hugo , Kamil Alison , Blumberg Jeffrey , Chen C‐Y. Oliver
Published: June 2021

Phenolic acids and dietary fiber contribute to the health benefits from consuming plant foods. However, fiber may interact with phenolic acids via an affinity to their functional antioxidant moieties. We tested whether fiber (soluble pectin or insoluble cellulose) under different pH in vitro affects the Ferric Reducing Antioxidant Power (FRAP) and Total Phenol (TP) content of 3 phenolic acids: gallic (GA), protocatechuic (PA), and p‐coumaric (CA) acids with 3, 2, and 1 hydroxyl moieties, respectively. Phenolic acids at 30 μmol/L were incubated with 1, 2, or 3% (w/v) fiber at pH 1.5, 3, 5, 7 or 9 for 2 h. After centrifugation, supernatants were collected for the determination of FRAP and TP. The effect of fiber on FRAP and TP of the test compounds was dependent on fiber type and concentration, pH, and phenolic acid. Overall, pectin ≥2% and pH 9 had the greatest impact on study outcomes; e.g., at pH 9, 3% pectin decreased the FRAP of GA, PA, and CA by ≥65% and TP by ≥62%. At pH 1.5, 1% cellulose did not significantly affect the TP of GA and PA with reductions of ≤4%. Thus, fiber in an alkaline milieu decreased the antioxidant capacity of phenolic acids in vitro. This relationship may be relevant to the bioactions of these phytochemicals in the ileum in vivo. Further studies using infrared spectrometry and NMR are warranted to elucidate the chemical/physical interactions between dietary fiber and phenolic acids. Supported by the USDA.
In vitro and in vivo anti‐inflammatory effect of 4′‐hydroxylnobiletin, a major colonic metabolite of nobiletin
Authors: Wu Xian , Song Mingyue , Zheng Jinkai , Guo Shanshan , Xiao Hang
Published: June 2021

Nobiletin, a citrus flavonoid has been associated with various biological activities including anti‐inflammation. We have found that 4′‐hydroxylnobiletin (4HN) was a major colonic metabolite of nobiletin in mice after oral administration, and the colonic levels of 4HN were much higher than nobiletin. Herein, we studied anti‐inflammatory effects of 4HN in lipopolysaccharide (LPS)‐stimulated RAW 264.7 macrophages and 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA)‐induced skin inflammation in mice. Treatments of 4HN dose‐dependently inhibited LPS‐induced NO production in macrophages. These inhibitory effects were much stronger than those produced by nobiletin. Western blot analysis, ELISA and real‐time qRT‐PCR showed that 4HN significantly reduced the expression levels of iNOS, COX‐2, IL‐1 and IL‐6, and inhibited activation of NF‐κB, Akt and JNK in LPS‐stimulated macrophages. Furthermore, our results demonstrated that topical application of 4HN decreased TPA‐induced mouse ear edema by more than 80%. This inhibitory effect was associated with inhibition on TPA‐induced expression of iNOS, COX‐2, IL‐1, IL‐6, and TNF‐α in the mice. Overall, our results demonstrated that 4HN had strong anti‐inflammatory effects both in vitro and in vivo, which provided a solid scientific basis for using nobiletin to inhibit inflammation related diseases in the colon such as colitis and colon cancer.

Grant Funding Source: AICR, NIH, NAS, USDA
Mode of administration affected the capability of soybean‐derived peptide lunasin to prevent metastasis of human colon cancer cells in a mouse model
Authors: Dia Vermont P. , de Mejia Elvira Gonzalez
Published: June 2021

The objective was to evaluate soybean‐derived peptide lunasin to inhibit KM12L4 human colon cancer cell metastasis in mouse model of colorectal cancer. Mouse was injected with 1 × 106 KM12L4 colon cancer cells in the spleen and randomization was made after 4 days. Mice that received 4 mg lunasin/kg body weight per day (i.p.) had a reduced number of liver metastatic tumors compared to group that received PBS (P = 0.047). Liver‐tumor tissue showed lunasin (4 mg/kg bw, per day, i.p.) reduced the expression of histone H3K18 (P = 0.002) and H4K8 (P < 0.0001) with reduction in the expression of p300 enzyme (P = 0.039). Orally administered lunasin for 28 days reduced the number of liver metastatic nodules from 18 (PBS, n = 6) to 8 (8 mg/kg bw, n = 5, P = 0.293) to 1 (20 mg/kg bw, n = 3, P = 0.247); however, these values were not statistically significant. Gavage lunasin (8 mg/kg bw) did not significantly affect expression of proliferating cell nuclear antigen, pro‐apoptotic Bax, anti‐apoptotic Bcl‐2 and H3K18 and significantly increased the expression of H4K8 in liver‐tumor tissue. No liver‐tumor tissue was present for analysis after gavage lunasin (20 mg/kg bw). In conclusion, intraperitoneally‐administered lunasin (4 mg/kg bw) was effective in preventing colon cancer metastasis but not after oral administration of the bioactive peptide lunasin up to 20 mg/kg bw.

Grant Funding Source: USDA
Identifications of Novel SNPs in Portuguese Essential Hypertensive Patients
Authors: Pinho Maria Joao , Vaz‐da‐Silva Manuel , Soares‐da‐Silva Patricio
Published: June 2021

Essential hypertension (EH) is an etiological risk factor that poses a serious threat to human health. The aim of this study therefore was to investigate the correlation between the SNPs in hypertension related genes and Portuguese essential hypertensive subjects. Genomic DNA was isolated from PBMCs from a total of 79 hypertensive subjects (divided into: group 1, medicated HTA I and II; group 2, medicated HTA III and group 3, non‐hypertensive with cardiac disease) and 28 non‐hypertensive subjects. SNPs of interest were investigated using a PCR and Sanger sequencing based approach. The results of the analysis of the allele frequencies of MTHFR rs1801133 (C/T) and ATGR1 rs5185 (G/T), previously identified as hypertension‐associated SNPs, revealed that there were significant differences between goup1 and control. Six unidentified SNPs were found in SIK1. Among these novel variants the frequencies of allele C/T at chromosome position 44848234 lower in the hypertensive subjects (3.6% in group 1; 0% in group 2; vs 35.7% in control group), suggesting a protective role for this SNP. The results of this study partially confirm previous SNP profiling studies identifying SNPs that are associated to EH. However, we report several novel SNPs in hypertensive subjects these SNPs require further validation and functional analysis to evaluate whether they play a role in the pathogenesis of EH. Supported by FCT (PIC/IC/83204/2007)
Manganese accumulations in gill mitochondria of Crassostrea viginica
Authors: Nuhar Ahmed , Boisette Beatrix , Carroll Margaret A. , Catapane Edward J.
Published: June 2021

Manganese (Mn) is a neurotoxin causing Manganism in people exposed to high levels in the environment. Mn targets dopamine (DA) neurons in basal ganglia. Oxidative stress is implicated as factors of Mn toxicity and DA dysfunction. Mitochondria play a role as cause and target of oxidative stress damage. The mechanisms of damage is attributed to Mn's capacity to produce toxic free radicals and induce mitochondrial dysfunction. Mn is reported to accumulate in mitochondria and represent the primary pool of Mn in cells. Controversy exists to the extent of Mn accumulation in mitochondria. Some report Mn accumulates in nuclei and cytoplasm, not mitochondria. Our lab uses the oyster Crassostrea virginica as a test animal to study Mn neurotoxicty. We found Mn disrupts the DA system as well as mitochondrial respiration. To study where Mn accumulates within cells we used differential centrifugation and atomic absorption spectrometry. Gills were homogenized and centrifuged to isolate nuclear, mitochondrial and post‐mitochondrial fractions. Fractions were analyzed for Mn. To determine if isolated mitochondria accumulate Mn we prepared treated mitochondrial suspensions with up to 300 mM Mn. Results show a dose dependent accumulation of Mn in mitochondria of up to 5000%. Two day treatments with 500 and 1000 μM Mn increased Mn (μg/gdw) in gill from a baseline of 5.8 to 41.6 and 133.8, respectively, and centrifugation revealed Mn accumulations were primarily in nuclear and mitochondrial fractions. The study shows mitochondria accumulate Mn. In vivo treatments reveal accumulations with both the nuclear and mitochondrial fractions. The work was supported by 0516041171 of NYSDOE and 0622197 of DUE Program of NSF.
Involvement of p38 MAP kinase, but not ERK‐1/2, in histamine‐induced endothelial actin reorganization and barrier disruption
Authors: Lawrence Curtis , Madonia Eyong , Olubadewo Joseph O , Breslin Jerome W
Published: June 2021

We tested the hypothesis that histamine disrupts the endothelial barrier by activation of the p38 MAP kinase and ERK‐1/2. Transendothelial electrical resistance (TER) of human umbilical vein endothelial cells (HUVEC) grown on small gold electrodes served as an index of barrier function, before and after addition of 10 μM histamine. The role of p38 MAP kinase was tested with 6 μM SB203580 and ERK‐1/2 with 10 μM PD98059 or 1 μM U0126. Specific, dual phosphorylation of p38 MAP kinase or ERK‐1/2 on their activation sites was detected by Western blotting. Dynamics of GFP‐actin and VE‐cadherin‐GFP expressed in HUVEC were also evaluated. The results show that histamine increased phosphorylation of both p38 MAP kinase and ERK‐1/2, which was blocked by pretreatment with SB203580, or PD98059/U0126, respectively. Histamine‐induced decreases in TER were inhibited by SB203580, but not affected by PD98059 or U0126 pretreatment. Histamine did not change VE‐cadherin‐GFP organization, but did briefly stop GFP‐actin‐rich edge protrusions. However, these protrusions were not decreased by histamine when the cells were pretreated with SB203580. The data suggest that p38 MAP kinase, but not ERK‐1/2, mediates histamine‐induced endothelial barrier disruption by reducing endothelial cell spreading motions. Supported by NIH grants R01HL098215 and P20GM103424, NSF grant HRD0928797, and the Louisiana Board of Regents Support Fund.
The Kelch‐Like 1 Actin Binding Protein Modulates Neuronal Voltage Gated Calcium Channels
Authors: Piedras‐Renteria Erika Sofia , Perissinotti Paula Patricia , Ethington Elizabeth Gibbs , He Yungui , Martin Jody , Cribbs Leanne , Koob Michael D
Published: June 2021

Kelch‐like 1 (KLHL1) is a neuronal actin‐binding protein first identified in humans as part of the genetic locus of the neurological movement disorder Spinocerebellar Ataxia type 8. KLHL1 is expressed in the cytosol, axons and dendrites. KLHL1 KO causes loss of post‐synaptic structures, loss of motor coordination and gait abnormalities. KLHL1 is an ion channel modulator and upregulates HVA and LVA calcium currents in vitro by increased recycling endosome activity and by alteration of the kinetics of τdeactivation. We analyzed the endogenous HVA and LVA calcium currents in neurons from WT and KLHL1 KO mice, and compared these data with the acute down‐regulation of KLHL1 using a knock‐down shRNA (KD) strategy. KLHL1 KD caused decreased HVA and LVA calcium current expression. In contrast, the KO had no differences in total LVA current density, yet changes in current kinetics suggest there is a down‐regulation of α1H and compensatory expression of α1I. HVA current density was also decreased in KO (40%) at 4–6 DIV but not at 7–10 DIV, suggesting this deficit was also compensated for after 7 DIV. Our data confirm that KLHL1 modulates LVA and HVA channel function in vivo; the compensatory modulation of currents in the KO may contribute to the mild motor phenotype seen in these mice.

This material is based upon work supported by the National Science Foundation under Grant No. 1022075
Hypersensitivity of bronchopulmonary C‐fibers induced by an increase in airway temperature in ovalbumin (Ova)‐sensitized Brown Norway rats.
Authors: Lin Yu‐Jung , Lee Lu‐Yuan
Published: June 2021

Our recent study has reported that hyperventilation of humid warm air (WA) triggered cough and reflex bronchoconstriction in patients with mild asthma (AJRCCM 2012). We suggested that sensitization of the bronchopulmonary C‐fibers by increasing airway temperature is involved, but direct evidence is lacking. This study was carried out to determine whether hyperventilation of WA enhanced the pulmonary C‐fiber sensitivity in an animal model of asthma (Ova‐sensitized Brown‐Norway rats). Isocapnic hyperventilation of WA for 3 minutes rapidly raised airway temperature to a peak of ~42°C which significantly elevated the base‐line fiber activity (FA) of pulmonary C‐fibers in Ova rats immediately after the termination of WA challenge, but not in the control rats. Furthermore, the pulmonary C‐fiber response to right atrial injection of capsaicin was significantly higher in Ova rats than control rats before the WA challenge; this difference in the FA response to capsaicin was further amplified after WA in Ova rats (Δ=4.98±1.78 imp/s before, and 10.34±2.21 imp/s after WA; P<0.05, n=9). A similar pattern of the WA‐induced potentiation of the FA response to phenylbiguanide was also founded in Ova rats. These results suggest that Ova‐sensitization increases the excitability of pulmonary C‐fibers, and the hypersensitivity is further enhanced by an increase in the airway temperature. (Supported by NIH and DoD grants)
Hypersensitivity of pulmonary C fiber induced by arterial hypotension in anesthetized rat
Authors: Lin Ruei‐Lung , Lin Yu‐Jung , Lee Lu‐Yuan
Published: June 2021

This study was carried out to investigate whether arterial hypotension altered the sensitivity of pulmonary C‐fibers. Rats were anesthetized, ventilated, and their femoral arteries connected to a height‐adjustable reservoir for lowering the arterial blood pressure (ABP). Arterial hypotension (decreasing ABP to ~50 % of its control baseline) generated by lowering the level of blood reservoir induced a striking elevation of the base‐line activity of pulmonary C fibers and their responses to intravenous (iv) injection of capsaicin (0.25–1.00 μg/kg). The hypersensitivity developed shortly after the onset of hypotension (control, 2.18 ± 0.56 imp/sec; hypotension, 10.82 ± 1.72 imp/sec; n = 11, P < 0.05), sustained for the entire 20‐min duration of hypotension, and recovered slowly after ABP was returned to control. Furthermore, pulmonary C‐fiber sensitivity to lung hyperinflation (30 cmH2O), iv injections of phynylbiguanide (1.0–8.0 μg/kg), lactic acid (5–10 mg/kg) and adenosine (0.1–0.3 mg/kg) were all markedly potentiated by hypotension in a similar pattern. Infusion of sodium bicarbonate (70 μM/kg/min for 30 min) and ventilation with 100% O2 prevented systemic ischemic acidosis and tissue hypoxia, respectively, but they failed to attenuate the hypotension‐induced hypersensitivity. The mechanism(s) underlying the C‐fiber hypersensitivity remains to be determined. (Supported by NIH and DoD grants)
Development of an in vitro analog concomitantly expressing the cellular correlates of sensitization and suppression of feeding in Aplysia californica
Authors: Weisz Harris Alexander , Wainwright Marcy , Mozzachiodi Riccardo
Published: June 2021

When presented with aversive stimuli, the marine mollusk Aplysia californica shows increased defensive responses (sensitization) and suppression of feeding. The cellular correlates of sensitization and suppression of feeding include increased excitability of the tail sensory neurons (TSNs, which mediate defensive responses) and decreased excitability in neuron B51 (a cell critical to the Aplysia feeding circuit), respectively. The goal of this project was to develop a reduced preparation of the isolated Aplysia nervous system (buccal, pleural‐pedal, and cerebral ganglia) in which the effects of aversive stimuli within the defense and feeding neural circuits could be mimicked in vitro by electrical stimulation of afferent nerves. We successfully observed significant concomitant increase in excitability of the TSNs (measured by spikes evoked) and decrease in excitability of B51 (increase in the burst threshold) 15 min post stimulation. Currently, we are examining whether this in vitro analog expresses long term (24 h) correlates. This in vitro analog will facilitate further analyses of cellular and molecular modifications resulting from aversive stimulation, as it would allow manipulations not feasible in vivo. Supported by NSF Grant IOS 1120304 to RM.
Biochemical and mood changes dissociate during exposure to intense military training
Authors: Biedenkapp Joseph , Grohmann Kyle , McClung James , Farina Emily , Murphy Nancy , Cummings Christopher , Lieberman Harris
Published: June 2021

Exposure to intense stress is known to alter affective and cognitive behavior more profoundly in some individuals than others. Recently, we had the opportunity to examine the biochemical, cognitive and mood changes (Profile of Moods) associated with intense military training known as SERE school (Survive, Evade, Resist, Escape). SERE is a unique environment to examine Soldiers’ responses to stress in a realistic, standardized and carefully monitored military context in which students apply newly learned resistance skills to a variety of captivity scenarios. From baseline to the end of SERE (~2wks), serum testosterone and prolactin were both substantially depressed from 468ng/ml to 156ng/ml and 13.8ng/ml to 5.46ng/ml, respectively (p <.001). In contrast, epinephrine and norepinepherine remained elevated from 41pg/ml to 67pg/ml and 468pg/ml to 754.8pg/ml, respectively (p<.001). Interestingly, changes in mood states were not reflective of these prolonged endocrine alterations, as depression, tension, anger, and confusion returned to approximately baseline levels at recovery. This suggests that changes in these hormonal levels are not reliable biomarkers of mood in this elite population. This research was supported by the Office of Naval Research and US Army Medical Research and Material Command.
Role of Salicylic acid in Chromatin Modification and Remodeling
Authors: Kumar Dhirendra , Haq MD Imdadul , Hotz Tazley A , Hossain Mir A , Zhao Bingqing , Donald David
Published: June 2021

SABP2 is a 29 kDa tobacco protein which catalyzes the conversion of methyl salicylate into salicylic acid (PNAS 2005). RNAi silencing of SABP2 makes the plants more susceptible to pathogens and they fail to exhibit robust systemic acquired resistance (SAR) response (PNAS, 2005). Pathogen induced MeSA being lipid mobile quickly travels throughout the plant and also to neighboring plants. In uninfected tissues, MeSA is converted back to SA by SABP2 to induce SAR (Science, 2007). Using yeast two‐hybrid screening a number of putative SABP2 interactors were identified. One of the putative SABP2 interactor shows similarity to Histone deacetylases (HDAC). HDAC catalyze deacetylation of acetylated proteins. Acetylation and deacetylation have been implicated in chromatin modifications causing epigenetic changes in gene expression. Acetylation of histones is associated with increased transcriptional activity while deacetylation results in repression. HDAC are regulated by salicylic acid suggesting a direct role in biotic stress. HDAC suppress expression of several genes associated with SA biosynthesis in Arabidopsis. Pathogens target the expression of HDAC to suppress expression of host defense genes. Understanding these biochemical pathways would help in developing crop plants with better disease resisting capabilities. This research is supported by a grant from National Science Foundation.
Biogenesis of the H‐cluster of the [FeFe]‐hydrogenase
Authors: Broderick Joan B. , Shepard Eric M. , Duffus Benjamin R. , Ghose Shourjo , Joshi Neelambari , Boyd Eric S. , Peters John W.
Published: June 2021

The organometallic H‐cluster at the active site of [FeFe]‐hydrogenase consists of a [4Fe‐4S] cluster bridged to a 2Fe subcluster that is further coordinated by cyanide, carbon monoxide, and a nonprotein dithiolate ligand. Biosynthesis of this cluster requires three accessory proteins, two of which (HydE and HydG) are radical S‐adenosylmethionine (radical SAM) enzymes. The third, HydF, is a GTPase that binds iron‐sulfur clusters. EPR and FTIR spectroscopic studies have revealed that HydE and HydG serve to synthesize a CO and CN− ligated precursor on HydF that is ultimately transferred to the hydrogenase to effect activation. HydG has been shown to synthesize CO and CN− ligands from tyrosine using radical SAM chemistry. HydE is thought to synthesize the dithiolate ligand of the H‐cluster. Together, these results point to a mechanism for H‐cluster assembly in which radical SAM chemistry is utilized to modify standard iron‐sulfur clusters produced by the ISC machinery to generate the unique organometallic H‐cluster. This work has been supported by the Chemical Sciences, Geosciences, and Biosciences Division, Office of Basic Energy Science, Office of Science, U.S. Department of Energy.
The structure, regulation and activity of non‐canonical inteins
Authors: Mills Kenneth V. , Reitter Julie N. , Connor Katherine R. , Nicastri Michael C. , Williams Jennie E. , Colelli Kathryn M. , Marieni Michelle D.
Published: June 2021

Protein splicing is the post‐translational excision of an intervening polypeptide (intein) from its flanking domains (the exteins), concomitant with extein ligation. The third step of splicing is Asn cyclization coupled to cleavage of the peptide bond linking the intein and C‐extein. Both the Methanoculleus marisnigri (Mma) and the Pyrococcus abyssi (Pab) PolII inteins promote efficient splicing with C‐terminal Gln in place of Asn. The activity of both inteins is regulated by disulfide bond formation, which might suggest a regulatory role for splicing. The Pab PolII intein splices only at elevated temperatures, allowing for isolation of a stable precursor and for study of each step of splicing in vitro. An NMR solution structure of the Pab PolII intein reveals a particularly rigid structure, a disordered loop absent in a highly similar P. horikoshii intein, and a β‐hairpin specific to inteins from archaebacteria.

This material is based upon work supported by the National Science Foundation under grant MCB‐0950245 and the Camille and Henry Dreyfus Foundation.
Experimental and computational evidence that ribonuclease A alters the transition state for RNA 2′‐O‐transphosphorylation
Authors: Harris Michael E , Gu Hong , Wong Kin‐Yu , Radak Brian , Dissanayake Thakshila , Kellerman Daniel , Zhang Shuming , Dai Qing , Miyagi Masaru , Anderson Vernon E , York Darrin , Piccirilli Joseph
Published: June 2021

Ribonuclease A (RNase A) is a widely used model system illustrating basic principles of enzyme catalysis. Yet, ambiguity regarding the transition state it stabilizes precludes a complete understanding of its mechanism. Here, we present kinetic isotope effect analyses that provide direct evidence for a late, anionic transition state for RNase A catalyzed RNA 2′‐O‐transphosphorylation similar to catalysis by hydroxide in solution, but with dramatically less charge accumulation on the 5′O. Molecular simulations indicate stabilization of negative charge on the NPOs by H‐bonding with a protonated His12 and Lys41. Quantum chemical calculations consistent with the experimental KIEs provide a unified mechanistic interpretation whereby P‐O5′ bond cleavage is concerted with proton transfer from a protonated His119. Thus, theory and experiment provide strong support that RNase A exploits both direct electrostatic stabilization of the anionic transition state and general acid catalysis, resulting in an altered transition state relative to that observed in solution. This work was supported by grants NIH GM096000 to M.E.H., NSF MCB0717850 to V.E.A., NIH AI081987 to J.A.P. and NIH GM064288 to D.M.Y. D.L.K was supported by NIH training grant T32‐GM008056.
2010 Dietary Guidelines for Americans and decreased inflammation
Authors: El‐Abbadi Naglaa H. , Jacques Paul F. , Lin Honghuang , Sakakeeny Lydia , Troy Lisa M.
Published: June 2021

The Dietary Guidelines for Americans (DGA) aim to reduce chronic diseases. Inflammation is an established risk factor for Type 2 diabetes, hypertension, and CVD. We examined cross‐sectional associations between adherence to the 2010 DGA and inflammation among 2,200 Framingham Offspring study participants (mean age 61.3 years, 46% male). Adherence to the 2010 DGA was assessed by the 2010 Dietary Guidelines for Americans Index (DGAI_2010), possible range 0 worst to 100 maximum adherence (mean 59.1, range 24.3–86.3). Eleven inflammatory biomarkers were ranked and compiled into an inflammation score (IS): CRP, ICAM, CD40P, IL‐6, MCP1, MPO, P‐Selectin, TNFR2, OPG, and LPL‐A2 mass and activity, possible range 0 lowest to 11 highest level of inflammation (mean 4.9, range 1.4–8.9). Using multivariable linear regression, we observed an inverse association between continuous DGAI_2010 and IS adjusted for age, sex, BMI, smoking and NSAIDS use (p<0.001). Analysis of adjusted means IS by quintiles of the DGAI_2010 (quintile means 43.2, 53.5, 59.7, 65.6 and 73.3) showed lower IS in the highest two DGAI_2010 categories (IS means 4.7 and 4.8) vs lowest (IS mean 5.1) with Tukey adjustment (p<0.001). Greater adherence to the 2010 DGA may decrease chronic disease prevalence due to reduced risk of inflammation. Supported by grants from NIH (N01‐HC 25195, RO1 HL64753; R01 HL076784; R01 AG028321) and USDA (agreement 58–1950‐0–014).
Profiling heat loss in an extreme polar mammal, the Weddell seal
Authors: Hindle Allyson , Horning Markus , Mellish Jo‐Ann
Published: June 2021

A major weakness in predicting habitat change impacts in polar regions is that little is actually known about energetic costs of basic thermoregulation in high‐latitude seals and other apex predators. Characteristics of body size and condition (surface area: volume ratios, insulation) are likely to affect heat loss and manifest as altered thresholds for thermal homeostasis. We monitored the thermal interface between animal‐environment directly via skin surface heat flux sensors (HFS) and indirectly with infrared thermography (IRT) in Weddell seals (n=42) over 2–7d. Utilizing 4 life history categories—pups, juveniles, non‐reproductive adult females (skip breeders) and post‐weaning females—we tested for effect of body size and known insulation patterns (blubber depth via imaging ultrasound). IRT surface temperature maxima and range suggested 4 sites of interest for HFS placement and continued monitoring: axilla (variable hot), head (hot), flank (variable cold) and neck (cold). IRT‐generated surface temperature distribution histograms further permitted the partitioning of body surface area such that an integrated thermal profile could be modeled from HFS data collected at these 4 sites. With this tool we can for the first time examine thermal homeostasis in free‐ranging seals over a range of potentially changing environments. Funded by NSF Polar Programs.
Thermoregulation in polar seals: Profiling surface heat in Weddell seals
Authors: Mellish Jo‐Ann , Hindle Allyson , Horning Markus
Published: June 2021

Our understanding of thermoregulation in marine mammals is ambiguous at best, due to the conflicting needs of homeostasis in air and water. Infrared imaging (IRT) has become increasingly applied to wildlife research given the non‐invasive nature of the technology, which is particularly desirable for work with species that are at risk and/or difficult to handle. Our prior work identified species‐specific patterns of heat loss by season and body condition under controlled conditions. As part of a larger study, we are assessing the use of IRT under extreme Antarctic field conditions to determine baseline and variation in thermal profiles in animals of known condition and physiological state. Weddell seals show consistent hot spots around the poorly insulated head and well‐insulated but active axilla. The trunk and neck are typically cool, whereas flippers are variable. Best fit LME showed ambient temperature, relative humidity and blubber depth to be the primary predictors of IRT signatures. Animal, body site and wind speed were not significant. Profiles were similar between wet and dry seals, however, dry animals can have surface temperatures that are only marginally above ambient, greatly minimizing heat loss. Funded by NSF.
Matrix metalloproteinase‐2 is not active in the diaphragm during mechanical ventilation
Authors: Sollanek Kurt J , Smuder Ashley J , Hudson Matthew B , Nelson W Bradley , Carrigan Christopher T , Urso Maria L , Powers Scott K
Published: June 2021

Prolonged mechanical ventilation (MV) results in diaphragmatic atrophy and contractile dysfunction, which occurs, in part, due to the activation of several proteolytic systems. New evidence shows matrix metalloproteinase‐2 (MMP‐2) can contribute to muscle fiber atrophy and contractile dysfunction in animal models of muscle disuse. Therefore, this experiment determined if prolonged MV alters mRNA expression and protein abundance of MMP‐2 and its regulators, TIMP‐2 and MT1‐MMP. Our results reveal that prolonged MV (12 hours) did not alter MMP‐2 mRNA expression or protein abundance in the diaphragm. Furthermore, prolonged MV did not promote an increase in mRNA expression of TIMP‐2 in the diaphragm. However, prolonged MV did result in a small increase in MT1‐MMP mRNA expression, but this increase may not be physiologically significant (<2‐fold). Finally, MV did not result in a significant change in diaphragmatic protein abundance for TIMP‐2 or MT1‐MMP. Collectively, our results demonstrate that MV does not activate MMP‐2 or its regulators. Supported by NIH RO1 HL087839 (SKP) and MRMC (MLU).
Gene array analysis reveals consensus heat stress response in rat liver, lung, heart, and kidney
Authors: Stallings Jonathan Daniel , Rakeesh Vineet , Helwig Bryan G. , Baer Christine E. , Lewis John A. , Leon Lisa R. , Reifman Jaques , Jackson David A.
Published: June 2021

Heat stroke (HS) is characterized by severe hyperthermia and a systemic inflammatory response syndrome that often results in multi‐organ damage (MOD). To identify molecular mechanisms mediating MOD, we examined global gene expression in rat liver, heart, lung, and kidney at a core temperature of 41.8°C (Tc,Max), 24h, and 48h. The largest gene response was in the liver, which correlated with a computational model of heat distribution at Tc,Max. In all organs, HS induced a consensus response of 66 transcripts, which were functionally enriched with annotated heat shock, anti‐apoptosis, and inflammation genes. We identified Zfand2a, Slc5a3, and SNORD14, novel genes not previously associated with the in vivo HS response. Each organ also demonstrated a unique HS response, such as HS‐induced Crayb (HSP20), HMGB1‐like protein, and IL10 in liver, all known inflammatory mediators. Consensus and organ specific markers may be exploited to assess HS‐induced MOD and subsequent recovery. Research supported by MRMC. Research was conducted in compliance with the Animal Welfare Act, and all other Federal requirements. The views expressed are those of the authors and do not constitute endorsement by the US Army.
Behavioral economics and social marketing campaign increases selection and consumption of school lunch garden bar Items among elementary school students (1019.19)
Authors: Andre Joshua , Wolff Cindy , Goto Keiko , Bianco Stephanie , Frigaard Martin , Hansen Garth , Riley Rachel , St. Cin Sheila
Published: June 2021

The current study assessed the effects of behavioral economics (BE) and social marketing (SM) on the selection and consumption of garden bar fruits and vegetables in elementary school students. A quasi‐experimental study evaluated the immediate and lasting effects of BE and SM on student selection and consumption of garden bar items. The BE component provided verbal prompts to take fruits and vegetables and increased availability of garden bar favorites. SM posters acted as reminders and guided selection of garden bar items. Two intervention levels were implemented: BE + SM (n=168) and SM only (n=275), and these were compared to a control (n=205). Selection and consumption were recorded using digital images of student lunch trays. Observations occurred at pre intervention, post, and follow‐up. Mixed ANOVA was used for statistical analysis. Pre‐to‐post changes in fruit selection and consumption, as well as vegetable selection, were significantly higher among BE + SM students compared to control students. Those pre‐to‐post changes remained at follow‐up. Although vegetable selection significantly increased pre to post among SM students, the change did not remain at follow‐up. A positive, but not significant, pre‐to‐follow‐up change in vegetable consumption was observed among the intervention groups. Together BE and SM appear to foster lasting effects on student selection and consumption of fruits and vegetables.

Grant Funding Source: United States Department of Agriculture