bluuebunny/crossref_metadata_2025 huggingface dataset:
Evidence that hypoxia‐induced pulmonary vasoconstriction in humans is primarily a post‐capillary event
Authors: Taylor Bryan Joseph , Snyder Eric , Olson Thomas , O'Malley Kathy , Johnson Bruce
Published: June 2021

Background

Exposure to hypoxia causes pulmonary vasoconstriction (HPV) and may contribute to the pathogenesis of HAPE. The site of HPV, however, is controversial. While the majority of data suggests HPV occurs in the pulmonary arterioles (precapillary), there is evidence that HPV may also occur in postcapillary vessels. Expansion of pulmonary capillary blood volume (Vc) is primarily passive and thus closely linked to cardiac output (Q).

Purpose

To examine relationships between Q and Vc during exposure to hypoxia and fluid loading.

Methods/Results

Healthy adults (n=25, age=31±2 yr) reported to the lab for 2 exposures: 1) 17‐h 12.5% FIO2 in a hypoxia tent, and 2) fluid loading using IV saline (30 ml/kg over ~15 min). Q, Vc and Dm (alveolar‐capillary conductance) were measured using a rebreathe method with small amounts of C2H2, CO and NO gas. Saline infusion caused an increase in Q (32±6%, P<0.01) and Vc (39±13%, P<0.05), and a decrease in Dm (−9±3%, P<0.05). Vc/Q was not different pre‐ vs post‐saline (17±2 vs 18±3, P=0.40). Hypoxic exposure caused an increase in pulmonary arterial pressure (67±11%, P<0.01), Vc (57±20%, P=0.04) and Dm (35±6%, P<0.01) despite no change in Q (0.2±2.8%, P=0.98). Vc/Q was greater post‐ vs. pre‐hypoxia (24±4 vs. 17±2, P=0.06).

Conclusion

The rise in Vc with a constant Q during hypoxic exposure suggests a post capillary site for HPV causing the expansion of the pulmonary capillary bed. HL71478.
Evaluation of Bioimpedance Spectroscopy (BIS) for Determination of Body Fluid Compartment Volumes (BFC) in Rats
Authors: Hu Lufei , Maslanik Thomas , Zerebeckyj Mykolai , Plato Craig F
Published: June 2021

BIS is broadly used in human and large animal research to measure BFC including total body water (TBW), extracellular fluid volume (ECFV) and intracellular fluid volume (ICFV). To date the application of BIS to small research animals has been very limited. We evaluated the sensitivity and reliability of BIS using ImpediVET by evaluating BFC in SD rats (n = 6–8/group) under various conditions including a) differences in BW (267.8±15.1g vs. 372.5±16.0g), b) furosemide (50 mg/kg p.o.; FUR) treatment, c) BFC responses to i.v. volume expansion (20 ml) with differing NaCl concentrations (hypotonic: 0.3%, isotonic: 0.9%, or hypertonic: 2.7%, respectively), and d) comparing BFC values to those obtained using traditional indicator dilution methods for TBW (3H‐water) and ECFV (35S). BIS‐determined BFC consistently agreed with predicted values independent of body size, and detects modest (−10%) FUR‐induced reductions in ECFV (70.8±4.1 ml) compared to weight‐matched control rats (84.0±4.3 ml; p < 0.05). BIS closely detected predicted shifts in ECFV (3.7–23.7%) and ICFV (−3.5–8.4%) in response to i.v. tonicity challenge with differing NaCl concentrations, and showed good agreement (r2 = 0.90) in BFC compared to that determined with 3H and 35S. The current findings indicate BIS evaluation with ImpediVET has as good or better consistency, sensitivity and reliability to evaluate BFC in rats as compared to traditional methods.
Ventilatory Changes After Active Heat Acclimation for Multiday Ultramarathon Races in the Sahara Desert
Authors: Serna Selima D , Clegg Miriam E , White Matthew D
Published: June 2021

To determine the effect of active heat acclimation (HA) on ventilatory responses, 4 endurance‐trained athletes (42.0 ± 5.7 y, mean ± SD) underwent 10 day of activity in 35–40°C and ~15% RH conditions to prepare for multiday ultramarathon races in the Sahara desert. Exercise during HA included 1 h/day of running at ≥50% maximal oxygen consumption (VO2max). Physiological assessments included pre‐ and post‐HA VO2max, heart rate (HR), esophageal (Tes), rectal (Tre), and skin temperatures (4 sites). Pre‐ and post‐HA VO2max values were not significantly different (52.8 ± 4.0 vs 52.0 ± 4.6 mL·kg−1·min−1, p = 0.35). Attainment of HA was confirmed by a significant reduction in exercise HR (p < 0.01) and trends (p=0.12) for reductions in both Thermal Comfort ratings and Borg Ratings of Perceived Exertion over the 10‐d HA period. During incremental exercise, from pre‐ to post‐HA, the ventilatory equivalent for O2 (VE/VO2) increased from 28.6 ± 1.0 to 30.2 ± 1.0 (p = 0.06), whereas the ventilatory equivalent for CO2 (VE/VCO2) showed no change (29.6 ± 0.8 vs 29.8 ± 2.0, p = 0.37). These preliminary data suggest that active HA increases ventilatory responses independently of an exercise training effect.

Supported by NSERC, Canadian Foundation for Innovation.
Changes in central venous pressure during heat stress as a possible predictor of compromised blood pressure control during simulated hemorrhage
Authors: Ganio Matthew Stueck , Brothers R. Matthew , Wingo Jonathan E. , Keller David M. , Crandall Craig G.
Published: June 2021

The extent to which heat stress compromises blood pressure control is quite variable between individuals; some are intolerant to a simulated hemorrhagic challenge while heat stressed, with others being relatively tolerant. The objective of this study was to identify if the magnitude of reduction in central venous pressure (CVP) during heat stress explains the variability in tolerance to a hemorrhagic challenge. Simulated hemorrhage was imposed via lower body negative pressure to pre‐syncope on 20 subjects during passive heating. Tolerance was quantified with a cumulative stress index (CSI), and the 8 subjects with the lowest and the 8 with the highest CSI were categorized as low‐ and high‐tolerant, respectively. The increase in core body temperature was similar between groups (1.45 ± 0.11 versus 1.41 ± 0.09°C, P = 0.44), and by design CSI was greater for the high tolerant group (439 ± 183 vs 86 ± 39 CSI units; P < 0.001). Although heat stress decreased CVP in the low (6.6 ± 2.6 to 0.8 ± 2.1 mmHg) and the high (7.6 ± 2.0 to 2.0 ± 3.0 mmHg) tolerant groups (P < 0.001), the magnitude of this reduction was not different between groups (low: 5.9 ± 1.4; high: 5.6 ± 2.1 mmHg; P=0.53). Contrary to our hypothesis, differences in blood pressure control during simulated hemorrhage are not related to differences in the magnitude of the heat stress‐induced reduction in CVP.

Supported by NIH Grants HL61388 & HL84072 & TREi
Acute volume infusion preserves left ventricular end diastolic volume during combined heat and simulated hemorrhage
Authors: Brothers R. Matthew , Wilson Thad , Dalsgaard Morten , Bundgaard‐Nielsen Morten , Pecini Redi , Secher Niels H , Crandall Craig G.
Published: June 2021

Heat stress compromises blood pressure (BP) control during a simulated hemorrhagic challenge, while acute volume loading attenuates this response. This study tested the hypothesis that the mechanism of preserved BP control during heat stress + volume loading is, in part, through preservation of left ventricular end diastolic volume (LVEDV). LVEDV was measured in nine subjects during normothermia (NT), after an increase in pulmonary artery blood temperature of 1.3 ± 0.2 °C (HS; P < 0.05), and after volume infusion (HS + Inf). In each condition, supine LVEDV was assessed via echocardiography, by the Simpson biplane method, prior to and during simulated hemorrhage, evoked via 30 mmHg lower body negative pressure (LBNP). LVEDV was reduced by HS (NT: 148 ± 26 ml, HS: 126 ± 31 ml; P < 0.05), however, this fall was attenuated by HS + Inf (HS + Inf: 133 ± 25 ml; P > 0.05 relative to NT and HS). During simulated hemorrhage, the reductions in LVEDV were similar between NT (45 ± 20 ml) and HS (52± 20 ml), whereas the reduction in LVEDV after volume infusion (22 ± 22 ml) was attenuated relative to NT and HS (P < 0.05). Importantly, infusion resulted in an LVEDV during LBNP that was significantly elevated (111 ± 26 ml) relative to HS alone (74 ± 25 ml). Improved control of BP following volume infusion during HS occurs in part by attenuating the reduction in LVEDV to a simulated hemorrhagic challenge.

Supported by NIH Grant NIH‐HL61388 & HL84072
Submaximal exercise increases Fos expression in central noradrenergic A1 and A6 nuclei of rats is related to heat storage
Authors: Rodovalho Gisele Vieira , Coimbra Cândido Celso
Published: June 2021

It has been shown that the central noradrenergic groups are involved in thermoregulation. However, the role of these neurons on heat balance during the exercise is unknown. The aim of our study was to evaluate the participation of central noradrenergic system on heat balance during exercise. Adult male Wistar, after familiarized with exercise on a motor‐driven treadmill, were submitted to a session of submaximal exercise until fatigue. During the session core (Tb) and skin tail (Ttail) temperatures were measured and heat storage rate (HSR) was evaluated. Ninety minutes after fatigue point, animals were transcardially perfused, the brainstem removed and A1, A2 and A6 neurons processed immunohistochemically for Fos and tyrosine hydroxylase (TH). Exercise increased the expression of Fos in A1, A2 and A6 groups. However we observed an inverse correlation between Fos expression and heat storage rate only in the A1and A6 groups respectively (.r2=0.92, P <0.01; r2=0.78, P <0.01). Our data indicate that although exercise increases Fos expression in all three noradrenergic brainstem nuclei, only the expression in A1 and A6 are related to heat storage. Financial support: FAPEMIG, CNPq.
Localized tyrosine or tetrahydrobiopterin supplementation augments vasoconstriction in aged human skin
Authors: Lang James A , Holowatz Lacy A , Kenney W Larry
Published: June 2021

Attenuated reflex vasoconstriction in aged skin may be partly mediated by oxidant‐induced reduction in functional substrate and cofactor availability for norepinephrine (NE) biosynthesis; tyrosine and tetrahydrobiopterin (BH4), respectively. We hypothesize that localized supplementation of these in aged skin would augment reflex‐ (whole‐body cooling) and pharmacologically‐ (tyramine‐‐displaces NE from axon terminals) induced VC. Four microdialysis fibers were placed in the forearm skin of 10 young (Y: 20 yr) and 10 older (O: 72 yr) subjects for infusion of 1) Ringer's (control), 2) 0.5mM L‐tyrosine, 3) 5mM BH4, and 4) BH4 + L‐tyrosine. Cutaneous vascular conductance was calculated (CVC = laser Doppler flux/MAP) and normalized to baseline (%ΔCVCbase). VC was attenuated at the control site in O during whole‐body cooling (Y: −39 ± 3, O: −17 ± 3 %ΔCVCbase; P<0.01) and tyramine perfusion (Y: −41 ± 3, O: −21 ± 4 %ΔCVCbase; P<0.01). Similar to BH4 (cold, Y: −37 ± 3, O: −36 ± 3; tyramine, Y: 41 ± 2, O: −36 ± 3 %ΔCVCbase), tyrosine also augmented the VC in O during cooling (Y: −37 ± 4, O: −34 ± 4 %ΔCVCbase) and tyramine (Y: −40 ± 4, O: −45 ± 4 %ΔCVCbase), but BH4 + tyrosine did not further augment VC during cooling (Y: −38 ± 4, O: −31 ± 3 %ΔCVCbase) or tyramine (Y: −36 ± 3, O: −36 ± 5 %ΔCVCbase). Localized tyrosine or BH4 augmented VC in aged skin, suggesting that their reduced bioavailability may impair NE synthesis and contribute to attenuated VC.
Influence of oral vitamin C on vasoconstrictor response to local cooling in human skin
Authors: Yamazaki Fumio , Takahara Kazuo , Sone Ryoko
Published: June 2021

Local administration of vitamin C (VC) at a very high physiological concentration (10mM) using intradermal microdialysis technique inhibits the cutaneous vasoconstrictor response to local skin cooling (LC) through adrenoceptor function in humans. But the effect of orally ingesting VC at physiological concentrations on the LC‐induced vasoconstrictor response remains unknown. The purpose of the present study was to test the hypothesis that oral VC enhances the cutaneous vasoconstrictor response to LC by inhibiting the oxidation of norepinephrine. Skin blood flow (SkBF) was measured by laser Doppler flowmetry at three sites (forearm, calf, palm) in healthy volunteers. The three skin sites were locally cooled from 34°C to 24°C at −1°C/min and maintained at 24°C for 20 min before (Pre) and 1.5 h after (Post) oral VC (2g single does) or no VC supplementation (time control study). Cutaneous vascular conductance (CVC) was calculated as the ratio of SkBF to blood pressure and expressed relative to the baseline value before LC. Oral VC enhanced (P<0.05) the reductions in CVC in the forearm (Pre, −48%; Post, −55%), calf (Pre, −43%; Post, −63%) and palm (Pre, −26%; Post, −54%) during LC. In the time control study, changes in CVC at each site during LC did not differ between the two trials. It was suggested that acute administration of oral VC enhances the vasoconstrictor responses to local cooling in glabrous and nonglabrous skin.
Integrity of paraventricular nucleus is critical for thermoregulation adjustments during exercise
Authors: Guimarães Juliana Bohnen , Wanner Samuel Penna , Coimbra Cândido Celso
Published: June 2021

Exercise induced‐hyperthermia results from the imbalance between heat production and dissipation, being capable of interfering on performance. Anatomical and physiological evidences support that the paraventricular nucleus of hypothalamus (PVN) is involved in central pathways modulating the sympathetic activity of thermoregulatory effectors, including the rat's tail vasodilation that is crucial for heat dissipation during exercise. The aim of this study was to verify the participation of PVN on thermoregulatory adjustments during exercise. Male Wistar rats (250–350 g) were submitted to bilateral electrolytic lesion of PVN (n=4) and with control rats were performed sham surgery (n=6). Core body temperature (Tb) and tail skin temperature (Ts) were continuously measured during exercise (18/min, 5% inclination) until fatigue. Ts was taken as an index of vasodilation and heat dissipation. Time to fatigue was 33.4% lower in PVN‐lesioned rats. However, heat storage rate was higher in PVN‐lesioned rats than control (10.4 ± 1.8 cal/min, PVN‐lesioned rats vs. 4.1 ± 0.2 cal/min, control; p<0.05). We also observed a higher threshold for tail vasodilation in PVN‐lesioned rats (0.6 ± 0.0 ° C PVN‐lesioned vs. 0.3 ± .0.0 °C, control; p<0.05). Our results indicate that integrity of PVN interferes with heat balance during exercise by altering heat dissipation.

Support: CNPq, FAPEMIG, CAPES.
Disinhibition of neurons in paraventricular hypothalamic nucleus (PVH) inhibits cold‐evoked shivering
Authors: Morrison Shaun F.
Published: June 2021

The thermogenesis from skeletal muscle shivering contributes to the maintenance of a homeostatic body temperature during exposure to a cold environment. Activation of neurons in the PVH elicits a potent inhibition of the sympathetic outflow and thermogenesis in brown adipose tissue (Am. J. Physiol., 296:831–843, 2009) and the present study determined whether this effect of PVH neurons to inhibit sympathetically‐mediated thermogenesis extends also to thermogenesis mediated by activation of somatic motoneurons. Cooling‐evoked shivering responses were monitored as the increases in neck and gastrocnemius muscle EMG activities elicited by cooling the trunk skin to ~32¢aC for 1–2 min with a water jacket in isoflurane‐anesthetized rats whose core temperature was lowered to ~36¢aC. Nanoinjection of the GABAA receptor antagonist, bicuculline (60 pmol), into the PVH terminated ongoing skin cooling‐evoked EMG increases and prevented EMG increases to subsequent skin cooling episodes for ~20 minutes. These results indicate that neurons in the PVH can exert a potent inhibitory influence on cooling‐evoked shivering thermogenesis. The physiological stimulus for, and the synaptic integration sites supporting this PVH‐mediated inhibition of thermoregulatory energy consumption remain to be determined. Supported by NIH grant NS40987.
Push‐pull thermoregulatory control by raphé‐projecting preoptic neurons?
Authors: Tanaka Mutsumi , McKinley Michael Joseph , McAllen Robin Michael
Published: June 2021

Cutaneous vasoconstriction is tonically regulated by preoptic neurons (Tanaka et al 2009, Am J. Physiol 296, R1248‐57) via a pathway that relays synaptically in the medullary raphé but not in the dorsomedial hypothalamus (Rathner et al 2008, Am J. Physiol 295, R343‐54). We sought to identify the preoptic neurons responsible for this function in 28 urethane‐anesthetized rats (1–1.5g/kg, i.v.). Extracellular unit recordings were made from 40 preoptic neurons that were antidromically activated by electrical stimulation in the medullary raphé. The neurons’ ongoing activity was measured during episodes where trunk skin and core temperatures were manipulated by passing cold or warm water through a jacket around the animal's trunk. Frontal EEG was recorded at the same time. Seventeen preoptic‐raphé neurons were unresponsive to temperature, and a further four cells gave responses that were secondary to changes in EEG state. Seven preoptic‐raphé neurons behaved as expected for neurons regulating cutaneous vasonstriction: they were tonically active in warm conditions but were inhibited by cooling the trunk skin. Unexpectedly, twelve preoptic‐raphé neurons were activated by cooling the trunk skin. Both ‘warm’ and ‘cold’ preoptic neurons thus project directly to the medullary raphé, and we speculate that both contribute to the control of cutaneous vasoconstriction.

Supported by NHMRC
Neurochemical alterations of sensory nerve in bone of osteoporosis‐induced female rats
Authors: Poonyachoti Sutthasinee , Deachapunya Chatsri , Ueasilamongkol Pimwipa , Chareonpundhu Narattapol , Krishnamra Nateetip
Published: June 2021

Post‐menopausal condition reduces not only bone mass but also skin nerve innervation. Estrogen may play roles in the regulation of nerve integrity in bone. Effects on sensory nerve integrity relevant to osteoporosis by estrogen in six‐month‐old sham and ovariectomized rats (OVX) with or without daily subcutaneously supplementation of 17β‐estradiol (E2, 1, 5 or 15 μg/kg BW) were examined. After 3 months, dual‐energy x‐ray absorptiometry was performed at left femur for bone mass density (BMD) and immunoreactivity (ir) of the neuronal and sensory neurotransmitter marker antibodies, anti‐PGP 9.5, anti‐calcitonin gene related peptides (CGRP) and anti‐substance P (SP) antibody at right femur. OVX rats without E2 supplement showed the lowest BMD at the distal femur, but not the proximal femur. The bone loss in the distal femur was in parallel with decreased PGP‐9.5, CGRP and SP‐ir at the distal femur of the same subjects whereas the up‐regulation of CGRP‐ir was indicated at the proximal femur. Daily E2 injection prevented these changes. The results indicated the presence of bone sensory nerve which was regulated by estrogen. The protection of bone loss induced by estrogen depletion may require the up‐regulation of nerve containing CGRP. This work was supported by Graduate school, Chulalongkorn University.
Simultaneous introduction of a novel high fat diet and wheel running induces anorexia
Authors: Scarpace Erin , Matheny Mike , Cheng Kit‐Yan , Erdos Benedek
Published: June 2021

Wheel running (WR) in rodents influences ingestive behavior, and scheduled feeding induces an anorexia that is associated with increased WR. We examined a new phenomena where the simultaneous introduction of both a novel high‐fat (HF) diet plus WR induces a severe anorexia in both young and aged F344 × BN rats. Rats were conditioned to normal chow (67±4 kcal/day) with and without WR. Chow was replaced with 60% HF and half the sedentary rats were switched to WR and half WR rats switched to sedentary. All groups experienced a hyperphagia except the sedentary introduced to HF plus WR which ate less than 10 kcal/day for three days. When HF and WR was introduced simultaneously, with continued availability of chow, rats ate more food (21±4 kcal/day), but less than WR then HF‐fed control (119±4 kcal/day). Similar results were observed in aged rats even though aged (162±23 m/day) ran considerably less than young (1139±185 m/day), suggesting it is the act of WR rather than the extent of WR. These results demonstrate that the act WR plus the novel introduction of a HF diet induces a severe anorexia in young and old rats despite the ad libitum availability of food. Supported by NIH AG‐26159 and P30 AG028740.
Genotypic variation in DPP4 modulates sensitivity to pharmacologic DPPIV inhibition
Authors: Woodard‐Grice Alencia Vanay , Marney Annis , Kunchakarra Siri , Brown Nancy J
Published: June 2021

Dipeptidyl peptidase IV (DPPIV) is a membrane‐associated serine protease that cleaves dipeptides from the amino terminus of peptides with an X‐Pro (or X‐Ala) motif, including the incretin, glucagon‐like peptide 1 (GLP‐1). Pharmacological inhibition of DPPIV improves glucose homeostasis in Type 2 diabetics by inhibiting the degradation of GLP‐1. We hypothesized that genetic variation in DPP4 modulates sensitivity to pharmacological inhibition of DPPIV. Previously, we reported 7 single nucleotide polymorphisms (SNPs) associated with DPPIV activity and/or antigen levels. We have now genotyped 19 subjects with metabolic syndrome who participated in a prospective, randomized, double‐blind, placebo‐controlled parallel group crossover study. Subjects were randomized to receive the DPPIV inhibitor sitagliptin (100 mg/day) or placebo for 5 days. Sitagliptin decreased DPPIV activity (13.08±1.45 versus 30.28±1.76 nmol/mL/min during placebo, P≤0.001). 3 of the 7 DPP4 SNPs associated with sitagliptin‐induced decreases in DPPIV activity. For example, sitagliptin reduced DPPIV activity to greater extent in individuals heterozygous at rs1861978 [13.57±0.41, 8.44±0.89, and 14.44±1.34 nmol/ml/min for G/G (n=2), G/T (n=9) and T/T (n=8) (P<0.001 for G/T versus others)]. There was no difference in DPPIV activity during placebo treatment [28.67±2.12, 26.41±3.64, and 31.71±3.83 nmol/ml/min in G/G, G/T and T/T]. In addition, the haplotype, rs12469968‐rs873826‐rs6733162, which we have previously reported to associate with ACE inhibitor‐associated angioedema, also associates with sitagliptin‐induced decreases in DPPIV activity (P<0.005). In conclusion, DPP4 genetic variation modifies the response to pharmacological DPPIV inhibition.
Visfatin‐Induced Lipid Raft Redox Signaling Platforms and Hyperpermeability in Glomerular Endothelial Cells
Authors: Boini Krishna M , Zhang Chun , Xia Min , Li Pin‐Lan
Published: June 2021

Adipocytokines have been reported to contribute to the glomerular injury during obesity or diabetes mellitus. However, the mechanisms mediating the actions of various adipocytokines on the kidney remained elusive. The present study was performed to determine whether acid sphingomyelinase (ASM)‐ceramide signaling pathway is involved in local oxidative stress in glomerular endothelial cells (GECs) induced by adipocytokines such as visfatin and adiponectin. Using confocal microscope, visfatin but not adiponectin was found to increase lipid rafts (LRs) clustering on the membrane of GECs in a dose‐dependent manner. Upon visfatin stimulation ASM activity was increased, an aggregation of ceramide or NADPH oxidase subunits, gp91phox and p47phox was observed in LR clusters, which was blocked by prior treatment with LR disruptor filipin, adiponectin, ASM inhibitor amitriptyline or ASM siRNA. Corresponding to aggregation of LR clustering with NADPH subunits in GECs upon stimulation with visfatin, superoxide (O2·−) production was significantly increased (2.7 folds) as measured by ESR. Functionally, visfatin treatment significantly increased the GEC permeability and this action of visfatin was blocked by inhibition of LR redox signaling platform formation. In conclusion, the injurious effect of visfatin, but not adiponection on the glomerular endothelium is associated with the formation of LR redox signaling platforms via LR clustering, which increases the glomerular permeability by disruption of microtubule network in GECs (supported by NIH grants HL‐091464, HL‐75316 and DK54927).
Skeletal muscle protein balance in mTOR heterozygous mice in response to endotoxin and leucine
Authors: Lang Charles H. , Frost Robert A. , Bronson Sarah K. , Lynch Christopher J. , Vary Thomas C.
Published: June 2021

We assessed the role of mTOR in regulating muscle protein synthesis in wild‐type (WT) and mTOR heterozygous (+/−) mice under basal conditions, and in response to endotoxin (LPS) and/or leucine. Lean body mass was decreased in mTOR+/− vs WT mice. Gastrocnemius weight was decreased in mTOR+/− mice due in part to a reduced rate of basal protein synthesis. LPS decreased muscle protein synthesis in WT and mTOR+/− mice to the same extent. Changes in protein synthesis in mTOR+/− mice under basal and LPS‐stimulated conditions were associated with reduced 4E‐BP1 and S6K1 phosphorylation. LPS also decreased PRSA40 phosphorylation and increased phosphorylation of raptor and IRS‐1 (Ser307) to the same extent in WT and mTOR+/− mice. Muscle atrogin‐1 and MuRF1 mRNAs were elevated in mTOR+/− mice under basal conditions implying increased proteolysis, and the LPS‐induced increase in these atrogenes was comparable between groups. Differences in basal muscle protein balance were not due to differences in plasma insulin or IGF‐I, nor to tissue TNFα, IL‐6 or NOS2. LPS impaired leucine's ability to increase muscle 4E‐BP1 and S6K1 phosphorylation in WT mice and this leucine resistance was exacerbated in mTOR+/− mice. Our data support the contention that sepsis‐induced reductions in mTOR activity are central in regulating skeletal muscle mass under basal conditions and especially in response to nutrient stimulation. (GM38032)
Myostatin attenuates hyperplasia and is important for muscle remodeling during compensatory growth
Authors: St‐Louis Mathieu , Solecki Michal , Michel Robin N
Published: June 2021

Myostatin (Mstn) is a TGFβ family growth factor that negatively regulates skeletal muscle growth such that mice lacking this protein display a doubling in muscle mass. To investigate the role for Mstn in muscle fiber remodeling, we compared the effects of functional muscle overload (OV) of the plantaris in Mstn Knockout (Mstn−/−) and C57BL/6 mice. The muscle midbelly area of non‐OV Mstn−/− mice was double that of C57BL/6 counterparts due to a higher number of muscle fibers and fibers displaying greater cross sectional size. OV induced a rapid increase (70%) in plantaris midbelly area of mstn−/− mice due to nascent muscle fibers and without concomitant increases in individual fiber sizes until 6 weeks of OV. C57BL/6 mice displayed a gradual increase (130%) in plantaris midbelly area over 6 weeks in response to OV largely due to muscle fiber hypertrophy. Molecular markers for hyperplasia confirmed this differential response in OV mstn−/− mice. The plantaris of Mstn−/− mice also did not undergo the typical OV‐related fiber type remodeling seen in C57BL/6 mice. These results show that hyperplasia is the major mechanism of muscle growth in OV mstn−/− mice whereas hypertrophy is the prime response in OV C57BL/6 mice. This differential growth pattern may be of importance when considering Myostatin‐related therapies in aging and diseases such as muscular dystrophy. Funded by CIHR, NSERC and CRC.
Role of PPARα in the regulation of glutathione metabolism
Authors: Guelzim Najoua , Huneau Jean‐François , Tomé Daniel , Hermier Dominique
Published: June 2021

Glutathione (GSH) derives from cysteine and plays a key role in redox status. GSH synthesis is determined by the availability of cysteine (from the diet or derived from methionine) and by the activity of rate limiting enzyme glutamyl‐cysteine synthase (GCS). Because PPARα has been shown to regulate the secondary metabolism of certain amino acids, GSH synthesis from cysteine was explored in fasted wild‐type (WT) and PPARα‐null (KO) male mice, fed on a semi synthetic standard diet. The concentration of plasma amino acids and of hepatic thiols was measured. Activity and mRNA levels of GCS were determined in the liver.

Plasma cysteine concentration was identical in both groups, whereas that of methionine was 18% higher in KO mice. Hepatic GSH concentration was 40% higher in KO mice, whereas other thiols (cysteine, homocysteine and cysteinylglycine) were not affected. GCS activity of KO mice was 16% higher for specific activity and 52% higher for total hepatic activity.

In contrast, PPARα deficiency did not affect GCS mRNA level. In conclusion, PPARα deficiency enhanced hepatic GSH storage, probably through an increased synthesis from cysteine. Thus, PPARα may influence the redox status by regulating cysteine metabolism. However, the absence of transcriptional regulation of GCS suggests a more complex control of GSH synthesis. Research support: Valorex & GLN (France).
Advancing Age Enhances AKT Activity and Expression in Skeletal Muscle Without Suppressing Contractile Protein Degradation and Atrogin‐1 Expression
Authors: Merrell Erin , Brown Alicia , DiSanto‐Rose Maria , Reynolds Thomas H.
Published: June 2021

The purpose of this study was to determine if sarcopenia is associated with insulin resistance, impaired AKT signaling, and increased contractile protein degradation rates. Insulin sensitivity was assessed by an insulin‐assisted glucose tolerance test (IAGTT) in middle‐aged (MA, 11–13 months) and aged (AG, 24–26 months) male C57B/6 mice. Contractile protein degradation rates were assessed by measuring 3‐methyl histidine release from isolated soleus (SOL) and extensor digitorum longus (EDL) muscles. During the IAGTT, blood glucose levels were significantly lower in AG mice compared to MA mice, indicating an improvement in insulin sensitivity with advancing age. The increase in insulin sensitivity observed in AG mice was associated with a significant increase in the phosphorylation of AKT in SOL and EDL muscles from AG compared to MA mice. The increase in AKT phosphorylation appears to be due to significant increases in both AKT1 and AKT2 expression. The phosphorylation of FOXO3a was significantly higher in EDL muscles from AG mice compared MA mice, but no age‐related differences in FOXO3a phosphorylation were observed in SOL muscles. The expression of the ubiquitin ligase, Atrogin‐1, was similar in SOL and EDL muscles from AG and MA mice. These results indicate that sarcopenia is not related to a decline in the ability of AKT to suppress Atrogin‐1 expression and contractile protein degradation.
Signal Transducer and Activator of Transcription 5 in Enterocytes Promotes Intestinal Mucosal Wound Healing by Regulation of Epithelial Tight Junction Assembly
Authors: Han Xiaonan , Gilbert Shila
Published: June 2021

Methods

We generated intestinal epithelial‐specific STAT5 knock out mice (STAT5 IEC KO) using cre‐mediated recombination to delete stat5 locus in IEC. STAT5 IEC KO mice were then treated with 3 % dextran sodium sulfate (DSS) either for seven days or for five days followed by 5 days of water. STAT5 IEC KO mice also exposed to NSAIDs chow for two weeks. We significantly knocked down GM‐CSF signaling and STAT5 expression in Caco‐2 and HT‐29 monolayer.

Results

STAT5 IEC KO mice revealed an increased para‐cellular permeability and bacterial translocation in mesenteric lymph nude. STAT5 IEC KO mice were more susceptible to NSAIDs‐induced ileitis and DSS‐induced colitis, precluding mucosal healing after stopping DSS administration compared to littermate control mice (STAT5 IEC WT). Intestinal epithelial cells (IEC) monolayer exhibited a hyperpermeability under either GM‐CSF signaling or STAT5 knocking down; phosphoserine‐MLC was elevated and tight junction (TJ) assembly was disrupted in either IEC monolayer with STAT5 knock down or STAT5‐depleted IEC.

Conclusion

STAT5 signaling in enterocytes is required for maintaining intestinal mucosal barrier function and promotes intestinal mucosal wound healing. Targeting on the activity of STAT5 signaling in enterocytes may be a novel therapeutic approach for treating intestinal barrier dysfunction in the sepsis and inflammatory bowel disease. This work was supported by Crohns and Colitis Foundation of America (XH), CCHMC Trustee Award (XH) and NIH clinical and translational research award KL2 (RR026315, XH).
Interplay between Septin 2 and Aquaporin 5 as a novel mechanism to modulate epithelial barrier function by regulation of microtubular stability.
Authors: Sidhaye Venkataramana , Chau Eric , King Landon Stuart
Published: June 2021

Airflow creates shear forces on the apical surface of airway epithelial cells, the effects of which are largely unknown. In isolated mouse trachea, airflow induced shear stress enhanced epithelial barrier function (evans blue dye‐4% albumin permeability). Primary human airway epithelial cells differentiated in an air‐interface were exposed to shear stress (3 dynes/cm2) using Krebs solution, causing increased barrier function (FITC‐dextran), and reduced the abundance of AQP5, an apical water channel, by sequential activation of TRPV4 and VGCC. With barrier enhancement we observe increased membrane associated septin 2 and destabilization of microtubules. Septin 2, known to mediate epithelial polarity in dividing cells by its interaction with microtubules, has not been previously implicated in altering monolayer barrier function. Our data suggests that AQP5 is a novel microtubule stabilizing protein and that regulation of AQP5 and Septin 2 modulate microtubule organization in response to shear stress, which is critical in regulating barrier function. This important homeostatic mechanism to regulate polarized epithelial responses to known stimuli such as EGF, IFNy and Substance P.
Scaling in the primate masticatory apparatus
Authors: Holmes Megan A , Ruff Christopher B , Deleon Valerie B , Brown Kirsten M , Frohlich Bruno
Published: June 2021

Scaling patterns within the primate masticatory complex have been the focus of a number of comparative and evolutionary studies. The aim of this study was to further elucidate the mechanical factors influencing the scaling of primate mandibular strength and molar area. It is hypothesized that cross‐sectional strength properties of the mandibular corpus will scale isometrically to estimated mandibular bending moments (size) and that both M1 and M2 occlusal area will scale with isometry to both strength properties and bite force estimates. Computed tomography scans of adult Macaca (n=18) and Papio (n=16) were used to collect mandibular bending rigidity (Ix, Iy) and strength (Zx, Zy) properties. Molar area was calculated at the dento‐enamel junction. Genera were chosen for divergent body size, similar diet and close phylogenetic association.

Results indicate that when mandibular strength properties are compared to size indices both macaques and baboons scale with mechanical isometry. In contrast, molar area displays negative allometric trends in relation to strength properties and estimated bite force. This is most likely the result of diverse selective pressures acting on different elements of the masticatory complex. While mandibular cortical bone and cross‐sectional areas are highly responsive to functional demands and adapt to rapid changes in size, molar area is more genetically constrained.
Role of PS Product Inhibition in Accumulation of Phosphatidylserine (PS) in Neuronal Membranes
Authors: Kimura Atsuko Kakio , Kim Hee‐Yong
Published: June 2021

PS is a major membrane component in mammalian tissues. Neuronal tissues are highly enriched with docosahexaenoic acid (DHA, 22:6n‐3) and PS in comparison to non‐neuronal tissues. Such concentration of DHA and PS is known to be important for proper neuronal functions. In mammalian cells, PS synthase (PSS) 1 and 2 catalyze PS synthetic activity, which is inhibited by the product PS through the feedback mechanism. We have previously found that DHA containing PS species (DHA‐PS) highly accumulates in neuronal membranes mainly due to the substrate preference in PS synthesis towards DHA phospholipid species. To understand the biochemical mechanisms underlying enrichment of PS in neuronal membranes, we hypothesized that fatty acyl composition‐dependent product inhibition in PS synthesis also plays a role in PS accumulation. We found that DHA‐PS has weaker inhibitory effect compared to oleic acid (OA, 18:1n‐9) containing PS species (OA‐PS) when tested with mouse liver microsomes. Consistently, DHA‐PS inhibited PSS activity less effectively than OA‐PS when flag‐PSS2 were expressed in mammalian cells, successfully immuno‐purified, and subsequently used as an enzyme source. DHA‐PS as a weaker inhibitor of PSS together with the high specificity towards DHA‐PS synthesis may contribute to the high PS accumulation in neuronal membranes.

Intramural Program of National Institutes of Health
Man‐6‐P‐GlcNAc recognition by domain 5 of the cation‐independent mannose 6‐phosphate receptor
Authors: Castonguay Alicia , Olson Linda , Bohnsack Richard , Kudo Mariko , Gotschall Russell , Canfield William , Dahms Nancy
Published: June 2021

The 300kDa cation‐independent mannose 6‐phosphate receptor (CI‐MPR) targets newly synthesized acid hydrolases displaying mannose 6‐phosphate (Man‐6‐P) modified glycans to lysosomes. The extracellular region of the CI‐MPR is comprised of 15 domains and contains two high and one low affinity Man‐6‐P binding sites (localized to domains 1‐3, 9, and 5 respectively) which bind through four conserved residues (Gln, Glu, Tyr, Arg). Unlike domains 1‐3 and 9, domain 5 exhibits a higher affinity for phosphodiester‐containing lysosomal enzymes than Man‐6‐P. The goal of this study is to determine the residues of domain 5 that are responsible for phosphodiester recognition. Domain 5 containing single amino acid substitutions of residues in the binding pocket were expressed in Pichia pastoris. Surface plasmon resonance analyses revealed that the Y679F mutant exhibited similar affinities to lysosomal enzymes, acid α‐glucosidase (GAA) monoester and GAA diester, as wild‐type domain 5. In contrast, Y679L displayed a 10‐fold decreased binding affinity to GAA diester and no detectable binding to GAA monoester. Furthermore, substitution at position 653 (W653K/T) resulted in no detectable binding to either ligand. Taken together, Y679 and W653 are important for the architecture of the binding pocket of domain 5 and its specificity for diester.
I'm a PC (Pyruvate Carboxylase)…and diabetes was not my idea!
Authors: Andera‐Cato S , Arnold A. , Bach S. , Faught A. , Frisch E. , Her A. , Keller A. , Kennedy E. , Martin T. , McMurray D. , Mitch C. , Orozco C. , Rice C. , Roberts B. , Rodgers A. , Sauer A. , Schulman A. , Suggs A. , Surfus K. , Tucker S. , Wray T. , Vogt G. , St. Maurice M.
Published: June 2021

NIH estimates that 23 million Americans have diabetes, and 6.2 million are undiagnosed. If untreated, diabetes can cause complications, including heart disease and neuropathy. Type two diabetes patients cannot regulate glucose due to insulin resistance or deficiency. Pyruvate carboxylase (PC) plays an important role in insulin release from pancreatic β cells. Abnormal PC activity has been correlated with type two diabetes. PC is a dimer of dimers, each monomer a single chain with four domains: N‐terminal biotin carboxylase (BC), central carboxyltransferase (CT), C‐terminal biotin carboxyl carrier protein (BCCP), and allosteric domains. PC catalyzes the conversion of pyruvate to oxaloacetate (OAA). The process begins when biotin is carboxylated at the BC active site. The BCCP domain transfers the carboxybiotin to an active site in the CT domain. OAA is formed at the CT domain by adding a carboxyl group to pyruvate. Researchers concluded that the BCCP domain swings between active sites on opposite chains, instead of sites on the same chain. The Brown Deer SMART Team (Students Modeling A Research Topic), in collaboration with MSOE, built a model of PC using 3D printing technology illustrating this movement of the BCCP domain. Current research is focused on increasing PC activity through controlling a binding site in the allosteric domain, which may increase insulin production.

Supported by a grant from NIH‐NCRR‐SEPA.
The effects of estrogen, parathyroid hormone, and testosterone on raw cells
Authors: Rose LaKeysha , Anderson Marquissa , Tucci Michelle , Benghuzzi Hamed , Cameron Joseph A.
Published: June 2021

Several clinical and experimental studies suggest that gender affects immune responses. In this regard, there have also been observations of an extraordinarily significant female predominance of inflammatory arthritis. The purpose of our study was to investigate the cellular effects of estrogen (E), testosterone (T), and parathyroid hormone (PTH) on macrophage cells over 24, 48, and 72 hours. Cells treated with T caused enhanced cell proliferation after 24 hours, whereas cells treated with E or PTH caused less cellular proliferation after 24 hours. By 72 hours all treatment groups had less cell number than the untreated control. Cell membranes were not damaged by the increased hormone levels as evidenced by similar MDA values for experimental and control groups. Interestingly, nitric oxide levels were increased in the estrogen treated cells after 72 hours compared to the control cells. Cytological evaluation of the cells revealed increased aggregation of cells and swelling of those treated with T. The increase in nitric oxide levels by E indicates activation of macrophages, which may ultimately lead to the production of inflammatory cytokines. More experiments are necessary to address the effects of E on inflammatory cytokine production by macrophage cell types. Supported in part by NIGMS NIH Grant R 25 GM 50117.
The Ser377Gly of the GATA4 gene is a susceptibility variant for hypertension
Authors: Alrasheed Maha M , Muiya Paul , Gueco Daisy , Andres Editha , Alshahid Maie , Dzimiri Nduna
Published: June 2021

The disease‐predisposing effect of GATA4 to acquiring cardiovascular disease still remains to be elucidated. We initially sequenced the GATA4 coding region to identify variants of interest in a total of 250 individuals, and investigated the impact of 5 SNPs, rs3729855 (C>T), rs3729856 (A>G), rs1062219 (C>T), rs11785481 (C>T) and rs3203358 (C>G) on myocardial infarction (MI) by the real‐time PCR‐based techniques in a total of 3969 Saudi individuals. We first tested the potential confounding impact of congenital heart disease (CHD) on the study by comparing the 113 CHD patients versus 272 individuals harbouring dilated cardiomyopathy DCM. The results pointed to a significant causative association for the rs3729856 (p.S377G) with CHD [Odds ratio(95% Confidence Interval) = [1.76(1.18–2.61); p=0.005]. We further compared the 113 CHD versus the rest of the individuals (3969) and observed a similarly significant association [1.43(1.04–1.97); p=0.026]. Therefore, in order to rule out the influence of CHD on other relationships, we excluded the CHD individuals from further analyses, involving 2926 hypertensive patients (HTN) and 1022 non‐hypertensive individuals. Interestingly, the rs3729856 [1.22(1.06–1.40); p=0.005] similarly conferred risk for HTN. None of the other SNPs was associated with the disease and no confounding effect of coronary artery disease, diabetes of dyslipidaemia was found. These results demonstrate that the GATA4 rs3729856 (A>G) variant independently predisposes individual to acquiring CDH and HTN.
Effect of Advanced Glycation Endproducts on Human Smooth Muscle Cell proliferation by Co‐culture System with Endothelium and Monocytes.
Authors: Nam Mi‐Hyun , Lee Hyun‐Sun , Hong Chung‐Oui , Koo Yun‐Chang , Kim Ji Hoon , Wang Zeng , Yang Sung‐Yong , Lee Yan Houy , Lee Kwang‐Won
Published: June 2021

Atherosclerosis is one of the consequence of long‐term diabetic complications due to the accumulation of lipids and fibrosis in the artery wall. Advanced glycation endproducts (AGEs) may play an important role in the pathogenesis of diabetic complications. Atherosclerosis initiates adhesion of monocytes which induced by AGEs to the endothelium and transmigration across tight‐juction of endothelial cells (EC) into intima. Monocytes differentiated into macrophages which uptook lipoproteins and became foam cells. Smooth muscle cells (SMC) were then migrated from media to surface and secreted various fibrous elements. The aim of this study is to confirm the proliferation of SMC by AGE in a co‐culture system in which monocyte, EC and SMC were employed. THP‐1, human acute monocytic leukemia cells and HUVEC, human umbilical vein endothelial cells were cultured for 48 hours on the 2% gelatin coated upper chamber and HA‐VSMC, human artery vascular smooth muscle cells in the lower chamber. In a group treated with AGEs, SMC were co‐cultured with HUVEC and THP‐1 with 100 μg/mL of AGEs for 48 hours. Proliferation of SMC was measured by MTT assay. We observed proliferation of SMC by AGEs that using our co‐culture system. Moreover, AGEs treated SMC also induced several fibroblast markers on mRNA levels including fibronectin, collagen type 3 by this co‐culture system.
Sigma‐1 receptors regulate hippocampal dendritic spine formation via a free radical‐sensitive mechanism involving Rac1·GTP pathway
Authors: TSAI SHANG‐YI ANNE , HAYASHI TERUO , HARVEY BRANDON K , WANG YUN , WU WELL W , SHEN RONG‐FENG , ZHANG YONGQING , BECKER KEVIN G , HOFFER BARRY J , SU TSUNG‐PING
Published: June 2021

Sigma‐1 receptors (Sig‐1Rs) are endoplasmic reticulum (ER)‐resident proteins known to involve in learning and memory. This study tested if Sig‐1Rs might regulate denritic spine formation in hippocampal neurons and examined potential mechanisms therein. In rat hippocampal primary neurons the knockdown of Sig‐1Rs causes a deficit in the formation of dendritic spines that is unrelated to ER Ca2+ signaling or apoptosis but correlates with the mitochondrial permeability transition and cytochrome c release, followed by caspase‐3 activation, Tiam1 cleavage, and a reduction in Rac1·GTP. Sig‐1R‐knockdown neurons contain higher levels of free radicals when compared to control neurons. Microarray analyses indicate Sig‐1R knockdown highly correlate to oxidative stress response pathways. The application of a hydroxyl free radical scavenger N‐acetyl cysteine (NAC) to the Sig‐1R‐knockdown neurons rescues dendritic spines and mitochondria from deficits caused by the Sig‐1R siRNA. Further, the caspase‐3 resistant TIAM1 construct C1199DN, a stable guanine exchange factor able to constitutively activate Rac1 in the form of Rac1·GTP, and constitutively active Rac1·GTP also rescues the siRNA‐caused dendritic spine deficits, respectively. These results implicate Sig‐1Rs as endogenous key regulators in hippopcampal synaptic plasticity and suggest a free radical‐sensitive ER‐mitochondrion‐Rac1·GTP pathway in the regulations of dendritic spine formation in the hippocampus.
Modes of bacterial cell wall synthesis: combining localization analysis and protein‐protein interactions
Authors: White Courtney L. , Gober James W
Published: June 2021

In rod‐shaped bacteria, a complex of morphogenetic proteins coordinates cell wall biosynthesis during both lateral growth and division. One subset of these proteins (MreBCD) guides lateral synthesis whereas another group (FtsZQW, etc.) coordinates septum formation. We have found that FtsW, an integral membrane protein in Caulobacter crescentus, plays a novel dual role in both processes by interacting with critical components and regulators of peptidoglycan (PG) synthesis. Localization studies show that MreB actin cables collapse to irregular foci along the cell length upon FtsW ablation. This is mirrored by nearly identical changes in nascent PG synthesis, as measured by PG‐precursor labeling. Lateral patterns of PG synthesis, however, were uninterrupted by depletion of the essential cell division protein, FtsZ. Additionally, FtsW interacts with MreB actin and other morphogenetic complex components as inferred by two‐hybrid analysis. Our studies indicate that cell elongation may occur through two distinct modes of PG synthesis: (1) evenly distributed lateral synthesis directed by FtsW, and (2) new synthesis emanating from concentrated foci, directed by FtsZ. We hypothesize that FtsW and FtsZ interact with morphogenetic proteins in distinct functional ways, resulting in two discrete mechanisms of cell wall growth.