bluuebunny/crossref_metadata_2025 huggingface dataset:
Predicting plasma profiles following oral dosing for drug liver transporter substrates using physiologically based pharmacokinetic modeling
Authors: Gosset James Richard , Jones Hannah M , Lai Yurong , Barton Hugh A
Published: June 2021

The aim of this work was to assess the prediction of oral plasma profiles utilizing a previously developed physiologically based pharmacokinetic (PBPK) approach using in vitro sandwich cultured hepatocytes (SCHH) to predict intravenous plasma profiles for seven liver transporter substrates (Jones et al., 2012).

Utilizing physicochemical, in vitro passive permeability and solubility inputs, descriptors of oral absorption were predicted using a physiologically based gastrointestinal tract model. These inputs were then incorporated into the published transporter PBPK model with a single compartment for oral absorption.

The validity of these absorption predictions were shown to be satisfactory when assessed by combining them with the transporter parameters obtained from fitting the intravenous plasma data. Prospective predictions based on SCHH utilizing scaling factors developed in the literature and predicted oral absorption parameters were performed as a test of both aspects in predicting oral plasma time course. The average fold errors for the seven compounds increased somewhat to 2.5, 3.3, and 2.4, respectively.

An early prediction method has been developed using physicochemical and in vitro inputs to estimate plasma profiles and pharmacokinetic parameters for intravenous and oral dosing drugs that are substrates for liver transporters.
Regulation of NADPH‐cytochrome P450 oxidoreductase by glucocorticoids in rat liver
Authors: Vonk Alex , Riddick David S
Published: June 2021

NADPH‐Cytochrome P450 oxidoreductase (POR) is the obligate electron donor for all microsomal P450s, key catalysts of endobiotic and xenobiotic metabolism. Adrenalectomized rats show decreased hepatic POR activity that is rescued by cortisone. Dexamethasone (DEX), a synthetic glucocorticoid known to activate both the glucocorticoid receptor (GR) and the pregnane X receptor (PXR), increases hepatic levels of POR mRNA in rats and POR protein in wild‐type and Gr‐null mice. To study the roles of GR and PXR in POR regulation, we treated male rats with the GR‐selective agonist triamcinolone acetonide (TA), the PXR‐selective agonist pregnenolone‐16α‐carbonitrile (PCN), and several doses of DEX. At 6 h after dosing, PCN caused a 3‐fold induction of POR mRNA and a trend for increased POR mRNA levels was seen in response to TA. Neither PCN nor TA alone affected POR protein levels or cytochrome c reduction activity. A low dose of DEX (1 mg/kg), shown to activate GR but not PXR, caused a 3‐fold induction of POR mRNA at 6 h, whereas a high dose of DEX (50 mg/kg), shown to activate both GR and PXR, increased POR mRNA levels by nearly 5‐fold at 6 h and 4‐fold at 24 h. POR protein levels and cytochrome c reduction activity were increased at 24 h after the high dose of DEX. Both GR and PXR may contribute to the rat hepatic POR mRNA induction, but activation of either receptor alone seems insufficient to elevate POR protein and activity. [Support: CIHR]
Lisinopril augments the combined effect of nitric oxide and potassium channels on endothelium‐dependent relaxation in spontaneously hypertensive rats
Authors: Albarwani Sulayma , Al‐Husseini Isehaq , Al‐Siyabi Sultan , Al‐Ismail Azza , Tanira Musbah O
Published: June 2021

Endothelium‐dependent relaxation is reported to be impaired in hypertension. However, there is equivocal evidence on the exact nature of the underlying dysfunction and whether it could be affected by antihypertensive drugs specifically angiotensin‐converting enzyme inhibitors. This study was conducted to evaluate the contribution of NO and potassium channels to endothelial function after lisinopril treatment in spontaneously hypertensive rats. Male animals (10–12 weeks old) were treated with lisinopril (20mg/Kg/day) for 10 weeks. ACh (0.1 nM–10μM)‐induced relaxation of third order mesenteric arteries was measured on a wire myograph with isometric recording in the presence and absence of different blockers. In the presence of inhibitor of nitric oxide synthase (LNMMA 100uM), the maximum relaxation of arteries from untreated (UT) and treated (T) rats to ACh was reduced by 19±3.5% and 5.0±01.6% respectively, while in the presence of SK and IK channel blockers (apamin 100nM and TRAM‐34 10μM) it was reduced by 35±3.5% and 34%±6.25%; respectively (n=10–22). With combined blockade, ACh‐induced relaxation was inhibited by 29±6.2% UT and 65±3.2 % T. Our results suggest that chronic treatment with lisinopril, although it reduced the relative contribution of nitric oxide alone it enhanced the combined effect of nitric oxide and potassium channels towards more endothelium‐dependent relaxation.
Ionizing radiation exposure alters coronary and cardiac function
Authors: Kuo Maggie , Abraham Theodore P , Shoukas Artin A , Berkowitz Dan E
Published: June 2021

Radiation‐associated coronary artery disease is a significant side effect of cancer radiotherapy. Several studies have shown elevated reactive oxygen species (ROS) levels in the vasculature and impaired endothelial function after ionizing radiation (IR) exposure. Increased ROS levels and resulting oxidative stress and endothelial dysfunction are precursors to coronary artery and other cardiovascular diseases. The purpose of this study is to describe the effects of IR exposure on coronary artery and myocardial function. Rats were exposed to 5Gy gamma radiation. Coronary function was evaluated through drug‐induced vasodilatation response in vivo using high‐resolution transthoracic echocardiography. Adenosine triphosphate (ATP), adenosine, and dobutamine were administered intravenously to characterize endothelial‐mediated and smooth muscle‐mediated vasorelaxation. Dobutamine was also used to evaluate cardiac function. 5Gy rats exhibited increased resting coronary tone along with attenuated ATP, adenosine, and dobutamine‐induced vasodilatation. 5Gy rats also had smaller end‐diastolic and end‐systolic diameters but higher contractility. Data support that IR exposure results in coronary vascular dysfunction with myocardial compensation.
Effect of glutamine supplementation and resistive training in signaling pathways of protein synthesis and degradation in rat skeletal muscle
Authors: Pithon‐Curi Tania C , Rodrigues Carlos F , de Sousa Luis G O , Vasconcelos Diogo A , Vinolo Marco A , Rodrigues Alice C , Nachbar Renato T , Guimarães‐Ferreira Lucas , Gerlinger‐Romero Frederico , Sellitti Donald F , Hirabara Sandro M , Curi Rui
Published: June 2021

The effect of physical training and glutamine supplementation on signaling pathways involved in protein synthesis (Akt, 4EBP‐1 and S6) and degradation (FOXO, MuRF‐1 and MAFbx) of rat skeletal muscle was determined by real time PCR and western blotting in the following groups: 1) control (C); 2) trained (T); 3) control supplemented with glutamine (G); and 4) trained supplemented with glutamine (GT). Exercise protocol was conducted once every 3 days, during 5 weeks. Each training session consisted of six climbs in a vertical ladder (1.1 m; 80° from horizontal). EDL muscle mass was not changed, but a significant increase in the cross‐sectional area of the fibers was observed. Expression of FOXO‐1, Akt, atrogin‐1, Murf‐1, 4E‐BP1, and S6 was not altered. In contrast, phosphorylated S6 and 4EBP‐1 expression was significantly increased in the G and GT groups, and 26S proteasome activity decreased in the T group as compared to C group. Our results suggest that glutamine supplementation led to an increase of the phosphorylation of state 4EBP‐1 and S6. The 5 weeks of a physical training protocol described in this study in rats is a useful model to investigated the molecular mechanisms underlying the increased muscle mass due to long‐duration exercise. This research was supported by FAPESP, CNPq, CAPES and Guggenheim Foundation.
Quantitative assessment of cutaneous C‐fiber function in human skin via direct sudomotor activation
Authors: Kunz Justin D , Gillespie Michael A , Mack Gary W
Published: June 2021

Small sympathetic cholinergic C‐fiber function was examined using intradermal electrical stimulation to activate sudomotor nerves in the forearm skin of healthy humans. We measured local sweat rate (SR) with a 0.7 cm2 capsule containing a humidity sensor and a thermocouple and flushed with dry gas. Two small stainless steel needles were place into the skin (7 mm apart, 2 mm deep) and stimulated at a constant current intensity of 2.5 mA for 30 s and 120 s at frequencies of 0.2, 1, 2, 4, 8, 16, 32, and 64 Hz. The stimulus‐response characteristics were evaluated using a non‐linear four‐parameter logistic curve fitting equation. Peak local SR occurred between 32 and 64 Hz and was similar for 30 and 120 s stimulus duration averaging 0.46 ± 0.2 and 0.49 ± 0.2 mg • min−1 • cm−2, respectively. The EC50 averaged 8.3 ± 0.3 Hz during the 30 sec stimulus but decreased to 3.6 ± 0.05 Hz (p<0.05) during the 120 s stimulus. The area under the SR‐time curve following atropine sulfate (2 μg/ml) was lower than control (p<0.05) and not different from zero. These data support the hypothesis that the SR response to intradermal electrical stimulation reflects, exclusively, the effect of direct depolarization of sympathetic cholinergic C‐fibers and release of acetylcholine. As such, this model can be used to provide a quantifiable assessment of small C‐fiber function in human skin.
Improving mucosal function mitigates radiation‐induced gastrointestinal toxicity
Authors: Yin Liangjie , Vijaygopal Pooja , Menon Rajeesh , Zhang Amy , Zhang Mei , Huang Luqiang , Chen Chun , Zhang Lurong , Okunieff Paul , Vidyasagar Sadasivan
Published: June 2021

Our objective was to determine the optimal amino acid (aa) formula to mitigate the effect of radiation on the electrolyte and nutrient absorption and mucosal barrier functions of the gastrointestinal (GI) tract.

Studies were conducted on small intestinal mucosa of irradiated NIH Swiss mice (0, 1, 3, 5 or 7 Gy). The mucosal barrier mechanism was studied for dilution potential, conductance, and plasma endotoxin and IL‐1β; electrolyte, glucose, and aa absorption were studied with Ussing chamber flux studies and transepithelial electrical measurements. Amino acids that improved GI functions were used to formulate a drink that was tested in survival studies.

Radiation (0 vs 7 Gy) decreased anion selectivity (0.48±0.05 vs 0.91±0.07) and increased paracellular conductance (30.5±4.1 vs 54.0±10.1). Plasma endotoxin levels were elevated at 7 Gy (20 vs 120). Radiation (0 vs 7 Gy) decreased Na absorption (9.8±0.8 vs − 0.6±0.3) and increased electrogenic Cl secretion (10.1±1.5 vs 0.1±0.01). Glucose absorption exhibited a dose‐dependent decrease and was abolished in 7‐Gy mice (140.7±32.2 vs 3.2±0.8). Lysine, aspartic acid, glycine, isoleucine, threonine, tyrosine, valine, serine, and tryptophan improved GI functions. Survival studies using the aa mixture showed increased survival for lethally irradiated mice.

The aa mixture improved GI mucosal function and mitigated radiotoxicity.

Support provided by Enterade, LLC.
The Effect of Caffeine on Protein Synthesis Rates in Rat Gastrocnemius Muscle Tissue
Authors: Bui Steve , Shimkus Kevin L , Lee Chang Woock , Lee Teak V , Chen Vincent CW , Fluckey James D , Riechman Steve E
Published: June 2021

Background

Few studies have observed caffeine's effects on resistance exercise (RE). Some studies have shown caffeine may affect several key protein synthesis pathways. Since RE is commonly performed to increase muscle protein synthesis, it would be important to determine how caffeine affects protein fractional synthesis rates (FSR).

Methods

Thirty‐six male Sprague‐Dawley rats were assigned into six groups: no exercise no caffeine (NENC), no exercise low caffeine (NELC), no exercise high caffeine (NEHC), exercise no caffeine (ENC), exercise low caffeine (ELC), and exercise high caffeine (EHC). Animals in the exercise groups performed four sessions of jump squats with weighted vests. On each exercise day, caffeine groups were given three caffeine injections (LC: 2mg/kg; HC: 6mg/kg). All animals were given deuterium oxide as a protein tracer on the last RE session, and the gastrocnemius muscle was collected 24 hours later and FSR was analyzed.

Results

FSR was significantly different between exercise and no exercise groups (E: 10.33±0.49; NE: 8.54±0.33, p<0.05); however, there was no effect of caffeine intake on FSR (NENC: 8.36±0.82, NELC: 8.65±0.75; NEHC: 8.56±0.82/ ENC: 10.75±0.75; ELC: 9.99±0.75; EHC: 10.25±0.75, p>;0.05).

Conclusion

Our data suggests that habitual caffeine intake plays little to no role in post‐RE protein FSR. This observation could be due to an increased tolerance level for caffeine.
Interactions of mechanically induced coughing and sneezing
Authors: Simera Michal , Poliacek Ivan , Dobrolubov Boris , Veternik Marcel , Jakus Jan
Published: June 2021

Mutual interactions of cough and sneeze were studied on 11 spontaneously breathing pentobarbitone anesthetized cats (3.5±0.2 kg). Reflexes were induced mechanically in the nose and tracheobronchial airways, respectively. The amplitude of styloglossus muscle electromyogram (EMG) moving average during the expulsive phase of sneeze was about 16‐fold higher than that during cough (p<0.01; 9 cats). Higher inspiratory efforts occurred during coughing (p<0.01) vs. those in sneeze. Mechanical nasal stimulation (when no sneeze was evoked) had no significant effect on cough (7 cats). In the combined trials (both cough and sneeze were executed; 7 cats), the responses were classified as either sneeze or cough (no hybrid responses occurred) and they were potentiated comparing to control reflexes. Higher expiratory maxima (esophageal pressure and the amplitudes of abdominal muscles EMG) compared to control coughs and sneezes were found (p<0.05). In sneeze during combined trials also inspiratory efforts and the number of sneezes were increased (p<0.05). We conclude that mechanosensitive nasal afferents have limited effect on the tracheobronchial cough response. However, co‐expression of coughing and sneezing results in a variety of enhanced cough and sneeze responses supposedly representing improved airway defense.

“This work was supported by the Slovak Research and Development Agency under the contract No. APVV‐0189–11“ The project is co‐financed from EU sources“ – ERDF – European Regional Developmental Fund, and by VEGA 1/0126/12
Heart Failure with Preserved Ejection Fraction Induces Molecular, Mitochondrial, Histological, and Functional Alterations in Rat Diaphragm Muscle
Authors: Bowen T Scott , Rolim Natale , Fischer Tina , Bækkerud Fredrik , Medeiros Alessandra , Werner Sarah , Bronstad Eivind , Rognmo Oivind , Mangner Norman , Linke Axel , Schuler Gerhard , Silva Gustavo , Wisloff Ulrik , Adams Volker
Published: June 2021

Peripheral muscle dysfunction is a key mechanism contributing to exercise intolerance in heart failure patients with preserved ejection fraction (HFpEF), however the underlying mechanisms remain unknown. We used an animal model to better understand potential molecular, mitochondrial, histological, and functional alterations induced by HFpEF in the diaphragm.

Methods

Female Dahl salt‐sensitive rats were fed a low‐ or high‐salt diet of 0.3% or 8% NaCL for 28 weeks. Heart function and diaphragm alterations were subsequently assessed.

Results

Compared to low‐salt rats (CON; n=10), high‐salt rats (n=11) developed HFpEF, as demonstrated by impaired diastolic function, preserved left ventricular EF, left ventricular hypertrophy, elevated systolic blood pressure, increased pulmonary congestion, and elevated plasma natriuretic peptides (P<0.05). HFpEF rat diaphragm demonstrated (P<0.05): 1) a fiber type distribution shift from fast‐to‐slow twitch; 2) a decreased pro‐oxidative but increased anti‐oxidant capacity; 3) reduced proteasome activation; 4) impaired in situ mitochondrial respiration at complex I: 5) in vitro muscle weakness and increased fatigability in fiber bundles.

Conclusion

These data suggest a skeletal muscle myopathy exists in HFpEF and identifies novel mechanisms of exercise intolerance in HFpEF.
Hyperglycaemia Elevates Epithelial Lactate Secretion and Alters pH of Airway Surface Liquid
Authors: Garnett James , Baker Emma , Baines Deborah
Published: June 2021

Glucose in airway surface liquid (ASL) is normally ~0.4 mM, but rises (up to 4mM) when blood glucose is elevated (hyperglycaemia) and in respiratory disease, making the lung more susceptible to respiratory infection. We investigated whether hyperglycaemia also modified ASL lactate concentrations and the pH, of bronchoalveolar lavage (BAL) from wild type (WT) and glucokinase deficient GK+/‐ mice, and the ASL of primary human bronchial epithelial cells (HBE) and cell lines H441 & Calu3, cultured at air‐liquid‐interface.

Blood glucose was elevated in GK+/‐ compared to WT mice (WT= 9.4±0.2 mM, GK+/‐= 15.1±0.6 mM; p<0.001, n=12). BAL (but not blood) lactate was elevated in GK+/‐ mice (0.052±0.002 to 0.09±0.008 mM; p<0.05, n=6) and there was a correlation between blood glucose and BAL lactate. There was no difference in BAL inflammatory cell counts between GK+/‐ and WT mice (n=6), indicating another source of lactate in GK+/‐ lung.

In vitro, elevation of basolateral glucose from 5 to 15 mM increased ASL lactate concentrations (Calu‐3= 2.2±0.7 to 7.1±1.2 mM; H441= 3.7±0.3 to 8.3±1.0 mM; HBE= 0.51±0.11 to 2.2±0.12 mM; P<0.05, n=7‐13) after 6 hours, and also produced an alkalinisation of Calu‐3 and HBE monolayer ASL, but acidified Cystic Fibrosis (CF) HBE ASL. This acidification could be mimicked in normal HBE monolayers by removing HCO3‐ from the bathing solution. Inhibition of MCT1/2 with AR‐C155858 (100 nM) partially overcame the effects of hyperglycaemia on ASL lactate and pH.

These data indicate that hyperglycaemia increases lactate in ASL via secretion through epithelial MCT1/2 transporters. Also, the airway epithelium compensates for MCT driven H+ secretion by secreting HCO3‐, which is defective in CF epithelia.

Funded by MRC‐MICA MR/K012770/1.
The Lung Macroautophagy Substrate Proteome: A SYSTEMS BIOLOGY Analysis
Authors: Haspel Jeffrey , Chettimada Sukrutha , Lane Bill , Choi Augustine
Published: June 2021

Rationale

Macroautophagy is a lysosome‐based catabolic process that is implicated in the pathogenesis of multiple diseases. Although macroautophagy is ubiquitous, its physiologic role may be specialized in various tissue types and we know little of how this process contributes specifically to homeostasis in the lung. Since standard markers of macroautophagy cannot convey information about tissue‐specific patterns of catabolism we used a proteomic approach to compare autophagosome substrate turnover in mouse lung to that of mouse liver.

Methods:

We measured macroautophagic activity in the lungs and liver of GFP‐LC3 expressing transgenic mice using a leupeptin based protocol we developed previously. GFP‐LC3+ vesicles were were analyzed via 1D‐SDS PAGE followed by label‐free LC‐MS‐MS.

Results:

The macroautophagy substrate proteome of the lung was highly divergent than that of livers. Specific to lung macroautophagy, tight junction components were part of the substrate proteome in this organ. Lung macroautophagy had a higher predilection than liver for secreted proteins, possibly reflecting increased input from retrograde endocytic traffic to amphisomes. There was also a greater contingent of leukocyte‐associated proteins among lung macroautophagy substrates.

Conclusions

A proteomics‐based approach successfully identified organ specific variations in macroautophagy. We believe this approach will prove useful for understanding how disease states reprogram macroautophagy, and conversely how macroautophagy regulation brought about by interventions are likely to affect cellular physiology in different cell types.
Neuronal Na+ Channels as a Novel Cardiac Antiarrhythmic Target
Authors: Radwanski Przemyslaw , Brunello Lucia , Knollmann Björn , Gyorke Sandor
Published: June 2021

Background

Certain arrhythmias caused by abnormal impulse formation including catecholaminergic polymorphic ventricular tachycardia (CPVT) are associated with diastolic Ca2+ release (DCR) from the sarcoplasmic reticulum. Surprisingly, CPVT patients often respond to treatment with Na+ channel blockers. However, the relationship between Na+ influx and disturbances in Ca2+ handling immediately preceding arrhythmias in CPVT remains elusive.

Methods

Confocal microscopy and patch‐clamp recordings ofventricular myocytes isolated from CPVT mouse model with genetic ablation of cardiac calsequestrinwere used to assess Ca2+ handling and Na+ current density during various pharmacological interventions, while surface electrocardiograms were performed during catecholamine challenge.

Results

A subpopulation of Na+ channels (neuronal‐ and not cardiac‐type Na+ channels) colocalize with RyR2. Disruption of the cross‐talk between these by neuronal Na+ channel blockade abolished persistent Na+ current and reduced DCR in cells (Fig a).

image

Such reduction of DCR on cellular level translated in decreased arrhythmias in vivo (Fig b). On the other hand, augmentation of the neuronal Na+ channel activity with β‐Pompilidotoxin (β‐PMTX) increased DCR and caused frequent arrhythmias.

Conclusion

These data suggest that neuronal Na+ channels can potentially serve as an antiarrhythmic target. As current antiarrhythmic drugs are all cardiac‐type Na+ channel blockers, this is a major conceptual shift in cellular arrhythmogenesis.
Clonal Growth Capacity and Phenotype of Ex Vivo Expanded Oral Mucosal Tissue
Authors: Islam Rakibul , Utheim Tor , Eidet Jon , Lippestad Marit , Messelt Edward , Griffith May , Dartt Darlene
Published: June 2021

Purpose

Cultured oral mucosal epithelial cells can treat stem cell deficiency of the cornea. The present study aims to determine the best site for harvesting oral mucosal cells and the effects of four media on these cultured cells.

Methods

Pieces of oral epithelium were used from four locations: buccal mucosa (BM), hard palate (HP), lower lip (LL), and transition zone of the lower lip (TZ) of Sprague‐Dawley rats. Explants were grown in four different culture media for six days. The media were 1) RPMI 1640; 2) EpiLife; 3) oral keratinocyte medium (OKM); and 4) DMEM and Ham 12 (1:1 mixture). Morphology, phenotype, and colony forming efficiency were analyzed.

Results

DMEM and RPMI yielded higher explant outgrowth than OKM and EpiLife. Using DMEM and RPMI, fold growth was superior from LL biopsies (36.43 ± 7.42 and 32.34 ± 12.22, respectively) compared to biopsies from HP (7.46 ± 0.69 and 10.14 ± 1.14, respectively), BM (11.15 ± 2.94 and 22.40 ± 8.45, respectively) and TZ (19 ± 3 and 15 ± 4.02, respectively) (p<0.05; N=4‐11). The percentage of PCNA+ cells was lower near the explant than at the leading edge whereas the percentage of NGF P75+ was not significantly different in any of the locations. In DMEM, cells from TZ showed higher colony forming efficiency (0.059% ± 0.004%) than cells from BM, HP and LP (p<0.05; N=5).

Conclusions

The transition zone of the lower lip, which showed the highest colony forming ability, may be most effective in treating limbal stem cell deficiency.
B Vitamin Damage and its Repair or Pre‐Emption
Authors: Hanson Andrew
Published: June 2021

Two increasingly recognized facts about metabolism are that (i) metabolites are damaged by spontaneous and enzymatic side‐reactions that form potentially toxic products, and (ii) cells have conserved enzymes that repair or pre‐empt such damage. B vitamins and the corresponding cofactors are highly damage‐prone, yet must be kept intact because they are essential. It is therefore no surprise that B vitamin/cofactor metabolism provides iconic examples of metabolite damage and its repair or pre‐emption, such as the following.

Repair of NADH and NADPH. These cofactors undergo enzymatic and chemical reactions that produce R and S forms of NAD(P)H hydrates [NAD(P)HX], which can inhibit dehydrogenases. The hydrates are repaired by the tandem action of an ADP‐dependent dehydratase that converts (S)‐NAD(P)HX to NAD(P)H and an epimerase that interconverts the R and S forms. The dehydratase and epimerase are triple‐targeted to plastids, mitochondria, and cytosol, enabling them to reconvert NAD(P)HX to NAD(P)H throughout the plant cell.

Pre‐empting damage in riboflavin biosynthesis. The first two intermediates of riboflavin biosynthesis in plants and bacteria are reactive potential damage agents, and may be over­produced because the biosynthetic enzymes lack feedback controls. Excesses of these intermediates are disposed of by COG3236, a protein that is fused to different riboflavin pathway enzymes in plants and bacteria. COG3236 cleaves the N‐glycosidic bond of riboflavin intermediates, yielding fairly harmless products and thus pre‐empting damage to the riboflavin pathway.
Electrical Disturbances in the Brain and Heart in Long QT Syndrome: A Dangerous Synergy
Authors: Auerbach David , McNitt Scott , Dirksen Robert , Moss Arthur
Published: June 2021

Patients with genetic ion channel diseases, such as Long QT Syndrome (LQTS), experience electrical disturbances in the brain (seizures) and heart (arrhythmias) that can lead to sudden death. Mutated genes in LQTS1‐3 are expressed in the heart and brain. The objective was to determine the co‐prevalence and severity of seizures and arrhythmias in LQTS. We hypothesized: 1) LQTS mutations and QT prolongation are associated with seizures; 2) LQTS subjects with seizures have an increased risk of cardiac arrhythmias.

Prevalence of seizures and arrhythmias were assessed using the Rochester LQTS Registry. LQTS analyses were limited to LQTS1‐3 subjects (LQTS+,n=965) and those with no known LQTS mutation (LQTS‐,n=936). Patients were assigned to the seizure group if they answered yes to seizures/epilepsy or were prescribed an anti‐seizure medication.

LQTS+, particularly LQTS2, patients were at a 3‐4x higher total risk of seizures, compared to LQTS‐ subjects (p<0.0001), and at each 10 year mark over 40 years of follow‐up (p<0.01). There was a positive correlation of QTc to the percentage of subjects in the seizure group (R2=0.998). The percentage of LQTS genotype/phenotype positive subjects in the seizure group was 4x greater than LQTS genotype/phenotype negatives (p<0.0001). Those in the seizure group had longer QTc and a 4‐9x higher risk of cardiac arrhythmias, compared to non‐seizure group, with a higher probability of arrhythmias over time in LQTS1 & LQTS2 (p<0.001). A complex network of factors coalesces to promote neuro‐cardiac electrical disturbances.
Selective Delivery of Copper Enhances Cytotoxicity of Millimolar Concentrations of Ascorbate towards Cancer Cells
Authors: Michels Alexander , Yu David , Frei Balz
Published: June 2021

Millimolar concentrations of ascorbic acid (millimolar ascorbate) in cell culture media and biological fluids may cause the generation of hydrogen peroxide (H2O2)via ascorbate auto‐oxidation and reduction of molecular oxygen. Although many cancer cell lines appear to be particularly susceptible to H2O2 exposure, the cytotoxicity of millimolar ascorbate may also be observed in normal cell lines. Delivery of redox‐active copper specifically to cancer cells, but not normal cells, may increase the cancer cells' susceptibility to killing by H2O2 due to increased intracellular production of hydroxyl radicals and oxidative DNA damage. Consistent with this hypothesis, we observed that pre‐treatment of colon cancer cells (Hct 116 or HT29) with the clinically‐used copper compound, copper(II)‐diacetyl‐bis‐methylthiosemicarbazone (CuATSM), increased the cytotoxic effects of millimolar ascorbate as measured by MTT and luciferase‐based methods. Since CuATSM does not increase the apparent rate of ascorbate oxidation in cell culture media, extracellular H2O2 production is not increased. Rather, the cytotoxic effects of CuATSM treatment are due to the intracellular accumulation of copper ions. The addition of millimolar ascorbate or H2O2 results in an acceleration of the copper‐mediated oxidative damage and cytotoxicity. The reduction of CuATSM and subsequent accumulation of copper is thought to be specific to cancer cells in culture and in vivo. Overall, our results suggest that CuATSM may be used in vivo to deliver redox‐active copper to cancer cells, thereby increasing the efficacy of intravenous administration of ascorbic acid as a cancer therapy.
The in vivo effect of oil palm phenolics (OPP) in atherogenic diet induced rat model of Alzheimer's Disease (AD)
Authors: Wu Yan , Srirajavatsavai Vindhyaja , Monplaisir Kenechukwu , Sambanthamurthi Ravigadevi , Gupta Smiti
Published: June 2021

OPP, a water soluble fraction from oil palm, rich in phenolics has been found to possess significant antioxidant activities. Since AD and the associated amyloid plaque buildup has an underlying component of oxidative stress, we hypothesized that OPP may reduce amyloid beta burden in vivo. The aim of this study was to investigate the change in behavior and pathology upon dietary supplementation of OPP in an atherogenic high cholesterol diet induced rat model of Alzheimer's disease. For this, 24 male Brown Norway rats were randomly assigned to 3 groups: the cholesterol group (2% cholesterol), cholesterol + OPP (5% OPP) and cholesterol + curcumin (2% curcumin), and maintained on their respective diets for 6 months. Curcumin has shown to have therapeutic benefits in AD and was used as a positive control. Morris water maze test performed once a month (3 trials/day for 5 days) to assess hippocampal dependent spatial learning showed that OPP supplemented rats exhibited improved learning abilities as compared to the cholesterol group (p<0.05). Hippocampal ß Amyloid (Aß) plaque burden, as observed by Congo red staining, was reduced in both OPP and curcumin supplemented animals. This was validated by ELISA assay. Further, mRNA expression levels of APP and BACE1, involved in Aß formation in AD were significantly decreased in the OPP and curcumin supplemented rats. The data indicates that dietary OPP have potential in improving an age‐dependent decline in spatial cognition and Aß deposition in hippocampus.
Race‐Ethnic Patterns in Iron Intake and Status in US Women: Findings from NHANES 2003‐06
Authors: Ahluwalia Namanjeet , Sternberg Maya , Raghavan Ramkripa , Looker Anne , Pfeiffer Christine
Published: June 2021

Race‐ethnic differences in iron status of US women have been reported previously. We examined the association of race‐ethnicity with iron intake from diet and supplements and with iron status in a nationally representative sample of non‐pregnant, non‐lactating 20‐49 y old women who participated in NHANES 2003‐06 (n~1850). Iron intake from supplements (SupplFe) over the last 30 d was recorded at an in‐home interview. Dietary iron (DietFe) intake was estimated via a 24‐h recall obtained using USDA's multiple pass method. Blood was analyzed for serum ferritin (FER), transferrin saturation (TS), and soluble transferrin receptor (sTfR). Body iron (mg/kg) was estimated as the molar ratio of sTfR to FER. Significant correlations were noted between total iron intake (diet and supplements) and some iron status indicators: TS (rho=0.16, p<0.0001) and FER (rho=0.08, p<0.02); DietFe intake was not associated with iron status. DietFe intake was related to race‐ethnicity overall (p=0.01) and in iron‐supplement users (p=0.02), with non‐Hispanic blacks (NHB) having lower DietFe intake compared to non‐Hispanic whites (NHW) and Mexican Americans. Interestingly, SupplFe intake was not related to race‐ethnicity. Iron status indicators (TS, sTfR, and body iron) were associated with race‐ethnicity overall and in iron‐supplement users and nonusers (except for body iron) (p<0.01), with NHB having lower iron status versus NHW. In conclusion, race‐ethnicity was related to iron intake and status and patterns varied with iron supplement use. NHB race‐ethnicity tended to have lower iron intake and status, which deserves further attention.
Relationship between Health Status, Quality of Life, Sleep, and Academic Success in Undergraduates
Authors: Shaffer Kendal , Kuhlman Amanda , Leone Ryan , Tucker Robin , Morgan Amy , Ludy Mary‐Jon
Published: June 2021

Academic achievement is related to career success; thus, peak college performance is critical. Information on relationships between grade point average (GPA), health‐related variables, quality of life (QOL), and sleep is conflicting. Limited research exists on the role QOL plays in GPA and primarily focuses on grade school students. This study's purpose was to determine associations between these variables at a collegiate level. Students (n=39F, 14M; 90.6% Caucasian; 77.4% freshmen, 22.6% sophomores) were recruited through campus advertising. Participants underwent anthropometric testing including body mass index (BMI 23.7±4.5 kg/m2) and percent body fat (%BF) by air‐displacement plethysmography (27.7±7.8%F, 16.8±8.7%M). Alcohol use, sleep (7.1±1.1 hr/day), and QOL were assessed via validated questionnaires. GPA was provided by the registrar (3.6±0.5). Results indicate moderate negative correlations between GPA and BMI, alcohol use, and %BF (r蠅‐0.297, p<0.05 for all), with a strong positive correlation between GPA and sleep quality (r=0.505, p<0.001). QOL was strongly negatively associated with sleep quality (r=‐0.434, p<0.001). Health‐related risks including high BMI, high %BF, high alcohol use, poor QOL, and poor sleep were demonstrated in nearly a third of students. Interventions aimed at altering health‐related behaviors may be helpful in improving academic performance.
Effects of Pre‐conceptual Micronutrient Supplementation on Maternal Depression in Vietnamese Women
Authors: Ramakrishnan Usha , DiGirolamo Ann , Nguyen Phuong , Gonzalez‐Casanova Ines , Nguyen Hieu , Pham Hoa , Truong Truong , Nguyen Son , Martorell Reynaldo
Published: June 2021

Micronutrient malnutrition has been associated with maternal depressive symptoms (MDS), but little is known about the effects of pre‐conceptual micronutrient supplementation. We examined the effects of pre‐conceptual micronutrient supplementation on MDS during pregnancy and postpartum using data from a double‐blind controlled trial (PRECONCEPT) in which 5011 Vietnamese women were randomized to receive weekly supplements containing either a) multiple micronutrients MM b) iron and folic acid IFA or c) folic acid FA until conception, followed by daily IFA supplements through delivery. Maternal depression was assessed using CES‐D at baseline and 12 m post‐partum and the EPDS during pregnancy and 3 m postpartum. The proportion of women ever experiencing elevated MDS during pregnancy (n=1616), postpartum (n= 1465) was 17.8% and 5.4%, respectively. No significant differences were seen by treatment. Among women at risk for depression at baseline (CES > 10; n=157), 29.1 and 30.1% experiencing elevated MDS during pregnancy in the MM and IFA groups, respectively, compared to 48.1% for FA only (p=0.07 for interaction). These relationships were attenuated (RR = 0.45, 95% CI: 0.2, 1.02 for MM vs FA and RR=0.47, 95% CI: 0.21, 1.06 for IFA vs FA) after adjusting for differences in age at marriage and serum ferritin between treatment groups. These findings suggest that women at risk for depression before they conceive may benefit from pre‐conceptual micronutrient supplements containing iron. Funded by Mathile Institute, Micronutrient Initiative, and Nestle Foundation.
Multiple Micronutrients And Early Learning Interventions Promote Infant Micronutrient Status And Development
Authors: Fernandeez‐Rao Sylvia , Hurley Kristen , Nair Madhavan , Balakrishna N , Tilton Nicholas , Radhakrishna V , Reinhart Greg , Harding Kimberly , Black Maureen
Published: June 2021

Background

Malnutrition and lack of early learning (EL) opportunities are major causes for children not reaching their developmental potential.

Objective

To evaluate the impact of multiple micronutrient powder (MNP) and EL intervention on infants' micronutrient status and development.

Methods

497 infants (6‐12 mos) were enrolled from 26 villages in rural India and randomized using a 2X2 design to receive MNP (iron, zinc, vitamins A,B2,B12, C and folic acid ) vs. placebo (B2) and EL vs. control. The EL intervention was based on the UNICEF‐developed Care for Child Development and delivered through biweekly home visits. Baseline, post‐intervention (6 mos) and follow‐up (12 mos) evaluations included 2ml venous blood and Mullens Scales of Early Learning. Data were analyzed using 2‐way ANOVA, adjusting for baseline, with MNP X EL interactions to assess intervention synergy.

Results

No baseline differences (66% anemic (Hb<11 g/dL), 31% inadequate iron stores (ferritin>12 µg/L), 20% stunted (HAZ<‐2), 19% underweight (WAZ<‐2). At follow‐up, the prevalence of micronutrient (MN) deficiencies was significantly lower in the MNP group. Significant interactions in motor and language performance showed that children who received either or both interventions had better scores than children who received neither.

Table 1. Percent MN Deficiencies at Follow‐Up * p < .05

MNP
Placebo

Infant
%
%

Hemoglobin (< 11g/dL) *
50.2
74.2

Ferritin (<12 ug/L) *
32.6
81.6

Transferrin receptor (> 2mg/L) *
44.1
81.7

Vitamin B12 (<200p/mL) *
2.6
9.1

Zinc (<65 ug/dL)
2.1
1.8

Table 2. MNP x EL Interaction at Post Intervention: Mean (SE) * p<.05

MNP
Placebo

EL
Control
EL
Control

Gross Motor
50.2 (1.0)
51.8 (1.0)
51.7 (1.0)
48.7 (1.0) *

Language
38.9 (0.6)
39.3 (0.6)
40.3 (0.6)
48.1 (0.6) *

Conclusion

Home MNP and EL can improve infant MN status and development.
The Ability of Serine Proteases to Induce an Increase in Barrier Function is Dependent on the Tight Junction Protein Occludin
Authors: Ronaghan Natalie , Shang Judie , Iablokov Vadim , MacNaughton Wallace
Published: June 2021

Inflammatory mediators disrupt the tight junction and may contribute to the pathogenesis of IBD through modulation of barrier function. We have found that serine proteases such as trypsin and matriptase induce an increase in barrier function in intestinal epithelial cell lines. However we do not know if they can reverse barrier disruption induced by inflammation, or how they modulate components of the tight junction (TJ). The SCBN canine epithelial cell line was mounted in Ussing chambers and barrier function assessed by 4 kDa FITC‐dextran flux and transepithelial electrical resistance (TER) measurement. Apical treatment with trypsin (45 BAU/mL) or matriptase (0.5 BAU/mL) induces a rapid and sustained increase in TER and decrease in FITC‐dextran flux. Treatment of cells with IFNγ and TNFα for 48 hours induced a dose‐dependent increase in permeability and redistribution of claudin‐4, tricellulin, and occludin from the TJ, as assessed by confocal immunofluorescence. The higher the doses of cytokines, the less able trypsin or matriptase could induce an increase in TER or decrease in FITC flux. Knockdown of occludin using siRNA resulted in a 39.8% reduction in baseline TER but no change in FITC‐dextran flux. Occludin knockdown induced a 74.1% and 90.6% reduction in the ability of trypsin and matriptase, respectively, to induce an increase in TER. Phosphorylation status of myosin light chain was assessed by Phos‐tag acrylamide Western blotting and no changes were observed with serine protease treatment. Thus, during inflammatory conditions, cytokines alter tight junctional structure and occludin localization; causing serine proteases to lose their ability to enhance epithelial barrier function.
From Lipidomics to Cellular Functions: Lipids as Modulators of Protein Activity
Authors: Brügger Britta
Published: June 2021

Given the fact that membranes are composed of hundreds of different lipid species, it is believed that beyond their well‐established functions as structural membrane components and solvents for membrane protein, individual lipid species fulfill specific roles in cellular membranes. Lipidomics analysis of COPI‐coated vesicles and their donor Golgi membranes revealed a role of COPI vesicles in intracellular lipid sorting. While the majority of sphingomyelin species together with cholesterol were segregated from COPI vesicles, one single sphingomyelin species, namely N‐stearoyl‐sphingomyelin, was significantly enriched in the vesicular membrane. In order to elucidate the underlying molecular mechanism of sorting of this lipid species into COPI vesicles, we have established novel in vivo and in vitro chemical biology lipid tools. Capitalizing on these tools, we discovered a direct and highly specific interaction of N‐stearoyl‐sphingomyelin with the transmembrane domain of p24, a protein involved in vesicular trafficking. p24 is a type I transmembrane protein that continuously cycles within the early secretory pathway and is suggested to act in its dimeric form as coat and cargo receptor in COPI vesicle biogenesis. Our data suggest that binding of sphingomyelin to p24 facilitates formation of p24 homodimers, and thereby triggers initiation of COPI vesicle budding

This specific protein‐lipid interaction within the hydrophobic phase of the membrane is defined by a signature within the TMD of p24 that is responsible for lipid selection. Bioinformatic analyses using a motif‐probability algorithm predict that this signature represents a conserved sphingolipid‐binding motif in mammalian membrane proteins, suggesting a widespread function of sphingolipids in modulating membrane protein activities.
Efficient Conversion of Human Fibroblasts to Functional Neurons by Sequential Inactivation of PTB/nPTB‐regulated RNA Programs
Authors: Fu Xiang‐Dong , Xue Yuanchao , Qian Hao , Hu Jing
Published: June 2021

Cellular reprogramming can be accomplished by using a set of cell lineage‐specific transcription factors. Our recent work reveals a key RNA program regulated by the RNA binding protein PTB, which is not only necessary but also sufficient to trans‐differentiate diverse cell types into neuronal‐like cells or even functional neurons (Xue et al., Cell 152:82‐96, 2013). In this RNA program, PTB down‐regulation allows the neuronal‐specific microRNA miR124 to more efficiently target its substrates, including multiple components of the REST complex responsible for repressing a large array of neuronal‐specific transcription factors in non‐neuronal cells. Down‐regulated REST also further depresses miR124, thus creating a sustainable signal for reprogrammed cells to maintain the neuronal identity.

One application of this new neuronal induction protocol is to convert patient fibroblasts into neurons for mechanistic studies of neurodegenerative diseases. Interesting, unlikely mouse cells, human fibroblasts can only be converted to neuronal‐like cells with complex neuronal morphology but without functional neuronal properties, suggesting the existence of another gatekeeper(s) that uniquely operates for neuronal maturation in human cells. We now demonstrate that this gatekeeper consists of the PTB paralog called nPTB and a specific transcription factor Brn2, which together forms another feedback loop critical for neuronal maturation. Either overexpression of Brn2 or down‐regulation of nPTB in PTB down‐regulated cells induces mature neuronal marks and a full spectrum of neuronal activities. These data demonstrate a sequential control mechanism for neuronal induction and maturation in human cells.
Knockout of Histidine Decarboxylase (HDC) Reduces Hepatic Fibrosis in Bile Duct Ligated (BDL) Mice
Authors: Francis Heather , Hargrove Laura
Published: June 2021

Background

Histamine (HA) is formed by the conversion of histidine by histidine decarboxylase (HDC). Cholangiocytes (i) express HDC; (ii) secrete HA; and (iii) proliferate after HA treatment. In cholestatic rodents, there is enhanced biliary hyperplasia and fibrosis.

Aim

to demonstrate that knockdown of HDC inhibits hepatic fibrosis.

Methods

HDC‐/‐ mice along with matching wild‐type (WT) were subjected to sham or bile duct ligation (BDL). Immunohistochemistry was performed for evaluating (i) bile duct mass by CK‐19 and (iii) collagen content by Masson's Trichrome. We measured: (i) serum histamine levels; and (ii) lobular damage, necrosis, inflammation and fibrosis (Sirius Red staining) in liver sections. In cholangiocytes, we studied the expression of a‐SMA, collagen 1, fibronectin and matrix metalloproteinases (MMPs) by qPCR. HA levels were assessed by EIA in serum and cholangiocyte supernatants. In vitro, cholangiocyte cultures were stably transfected with shRNA‐HDC to knockdown HDC expression. After transfection we evaluated: a‐SMA, collagen 1, fibronectin and MMPs by qPCR.

Results

In BDL HDC‐/‐ mice, there was decreased collagen content, necrosis, inflammation and fibrotic reaction compared to BDL WT mice.

image

HA levels were decreased in BDL HDC‐/‐. In BDL HDC‐/‐ cholangiocytes, there was a reduction of a‐SMA, collagen 1, fibronectin and MMP expression compared to BDL WT. In vitro, knockdown of HDC decreased a‐SMA, collagen 1, fibronectin and MMP expression.

Conclusion

Loss of HDC decreases BDL‐induced hepatic fibrosis. Our data suggests that HDC is a key regulator of the fibrotic reaction seen during cholangiopathies and may be a potential target for treatment.
Non‐glycolytic Function of Aldolase A Promotes Lung Cancer Metastasis through Down‐regulation of PLD2 Enzyme to Activate PLD1
Authors: Chang Yu‐Chan , Jan Yi‐Hua , Su Chia‐Yi , Hsiao Michael
Published: June 2021

In cancer, glycolysis rate is increased regardless of the oxygen availability, a phenomenon known as the Warburg effect. Several enzymes in glycolytic pathway have been investigated as potential therapeutic targets for cancer. However, the impact of targeting each glycolytic enzyme for cancer treatment remains unclear. Here, we perform systemic RNAi and pharmacological screening targeting every glycolysis pathway enzyme in lung cancer to found aldolase A (ALDOA) knockdown most significantly inhibits cell invasion/migration. However, this effect was not due to aldolase enzyme activity inhibition. Moreover, ALDOA knockdown does not affect glucose consumption, lactate production or intracellular ATP concentration, indicating a non‐glycolytic cause for lung cancer invasion/migration inhibition. We further perform immunoprecipitation and mass spectrometry using ALDOA as a bait in invasive lung cancer cells and found phospholipase D (PLD) and several ALDOA‐interacting proteins that may affect cancer invasion and metastasis. We further identify ALDOA interacts with Phospholipase D2 (PLD2) through PH domain and inhibits its enzyme activity. Instead, PLD1 upregulated to compensate the PLD enzyme activity and phosphatidic acid (PA) production to activate beta‐catenin/TCF‐4 signaling axis that are essential for ALDOA induced lung cancer cell invasion/metastasis. Taken together, our results uncover a novel non‐glycolytic function of ALDOA to promote lung cancer metastasis via PLD2 interaction and PLD1 activation that provides new mechanistic insights into lung carcinogenesis, with implication of novel strategy for lung cancer treatment.