bluuebunny/crossref_metadata_2025 huggingface dataset:
Abundance of Sea Kraits Correlates with Precipitation
Authors: Lillywhite Harvey B. , Tu Ming-Chung
Published: June 2021

Recent studies have shown that sea kraits (Laticauda spp.) – amphibious sea snakes – dehydrate without a source of fresh water, drink only fresh water or very dilute brackish water, and have a spatial distribution of abundance that correlates with freshwater sites in Taiwan. Here we report six years of longitudinal data on the abundance of sea kraits related to precipitation at these sites in the coastal waters of Orchid Island, Taiwan. The abundance of sea kraits varies from year to year and correlates positively with previous 6‐mo cumulative rainfall, which is an inverse index of drought. Grouped data for snake counts indicate that mean abundance in wet years is nearly 3X greater than in dry years. These data complement previous findings and suggest that freshwater dependence influences the abundance of sea kraits on both spatial and temporal scales. Increasing evidence for freshwater dependence in these and other marine species have important implications for the possible impact of climatic change on sea snake distributions. Funding was provided by the National Geographic Society (CER grant #8058‐06 to HBL), the U.S. National Science Foundation grant IOS‐0926802 to HBL, and Taiwan NSC grant 97‐2621‐B‐003‐004‐MY3 to M‐C Tu.
The Role of Polyamines in Osmotic Stress Tolerance in Gulf Killifish Fundulus grandis
Authors: Guan Ying
Published: June 2021

Polyamines are a family of low molecular weight organic cations produced in part by the coordinated actions of arginase II (Arg II) and ornithine decarboxylase (Odc). Although little is known of their functions in fish, polyamines have been implicated in diverse physiological processes. We describe a possible role of polyamines in hypoosmotic tolerance in the gulf killifish, Fundulus grandis, a species known to tolerate acute fluctuations in wide salinity range. Adult fish were reared in 5 ppt water for at least 1 month, and acutely transferred to 0.1, 0.5, 1, 2, and 5 ppt. Fish were randomly sampled from each salinity at 6 h, 1 d, 3 d, and 7 d post‐transfer. We have shown that relative Arg II and Odc mRNA levels, and Odc protein activity are highly up‐regulated in the gills. Hypoosmotic exposure also increases the concentration of the polyamines, putrescene, spermidine, and spermine. Although gill putrescine levels remain elevated throughout the first 7 days post ‐ transfer to freshwater, the concentrations of spermidine and spermine decrease with time suggesting a catabolic regulation of compounds. We are investigating the role of polyamine catabolism in apoptosis during salinity‐induced gill remodeling.
Using a selective breeding strategy to create 7th generation rats that voluntarily run low and high nightly distances
Authors: Roberts Michael Dominick , Gilpin Leigh , Knouse John , Haynes Anthony , Toedebusch Ryan , Ebone Laura , Moore Craig , Naples Scott , Booth Frank W.
Published: June 2021

The objective of this study was to selectively breed rats that voluntarily run high and low distances as a model of exercise and inactivity addictions, respectively. The selection experiment employed was adapted from Koch and Britton [Physiol Genomics. 2001 Feb 7;5(1):45–52], and this strategy was used for HVR and LVR propagation through generations (Gen)1–7. Data from Gen7 HVR and Gen7 LVR rats indicate that HVR rats run 6‐fold greater distances relative to LVR rats on days 5–6 of running [mean ± standard deviation; HVR (n=97): 6.0 ± 1.0 km, LVR (n=108): 1.0 ± 0.9 km, p < 0.001]. Interestingly, Gen7 HVR animals were significantly heavier than Gen7 LVR animals prior to running [HVR (n=81): 96.5 ± 20.6 g, LVR (n=108): 86.9 ± 12.1 g, p < 0.001] despite the similar ages of both lines. Thus LVR rats being heavier than HVR rats prior to running in Gen7 is not a viable explanation for running differences between lines. It will be of future interest to identify central mechanisms which may help explain running motivation differences that exist between our HVR and LVR lines. These efforts will attempt to further understand the etiology of societal sedentarism and potentially assist in the development of therapies meant to increase physical activity.
Inhibition of calpain or caspase‐3 protects against immobilization‐induced muscle atrophy
Authors: Talbert Erin E , Smuder Ashley J , Min Kisuk , Kwon Oh-Sung , Powers Scott K.
Published: June 2021

Increased proteolysis is crucial to disuse skeletal muscle atrophy. However, the roles that the proteases calpain and caspase‐3 play during inactivity‐induced muscle atrophy remain unclear. We hypothesized that administering either a calpain or caspase‐3 inhibitor would protect the soleus muscle against atrophy and mitochondrial dysfunction following 7 days of hindlimb immobilization (i.e. casting). Immobilization significantly (~13%) decreased the cross‐sectional area of soleus type I fibers. Importantly, independent inhibition of calpain or caspase‐3 prevented this atrophy. Casting decreased soleus mitochondrial coupling by ~52%. Inhibition of either calpain or caspase‐3 attenuated this casting‐induced mitochondrial dysfunction. Interestingly, inhibition of calpain prevented the casting‐induced activation of caspase‐3. Further, caspase‐3 inhibition prevented the immobilization‐induced activation of calpain. In conclusion, these results indicate that calpain and caspase‐3 play important roles during inactivity‐induced soleus muscle atrophy. Further, our data suggest that there is a regulatory cross‐talk signaling mechanism though which calpain and caspase‐3 regulate the activity of each other.

Supported by NIH R01 HL087839 (SKP) and a UF Alumni Fellowship (EET).
Onset and recovery of muscle hemodynamic and contractile properties of males and females following a submaximal isometric contraction
Authors: Crenshaw Albert Guy , Pettersson Linda , Johnson Pete
Published: June 2021

Electrical stimulation determined contractile properties and near infrared spectroscopy determined hemodynamics of the forearm extensor carpi ulnaris muscle of 7 males (M) and 7 females (F) before (pre) and after (immediate – post0, 30 min – post30, and 60 min – post60) a 12‐min isometric contraction at 15% MVC. Contractile data entailed twitch force (Force), contraction time (CT) and ½ relaxation time (½Rx). Hemodynamic data, generated with upper arm venous occlusion (50 mmHg for 1 min), consisted of slopes calculated for total hemoglobin (HbT) and deoxyhemoglobin (Hb) as estimates of blood flow and oxygen consumption. Data are below with * indicating significant differences compared to pre (†–higher than pre for ½Rx and lower for Hbslope).

pre
post0
post30
post60

Force (N)

M
2.89 (1.2)
2.82 (1.1)*
2.64 (1.1)*
2.84 (1.3)*

F
1.39 (0.3)
1.31 (0.3)*
1.28 (0.5)*
1.37 (0.4)*

CT (ms)

M
58.34 (8.3)
55.69 (9.4)*
57.71 (9.5)*
58.33 (8.2)*

F
57.16 (2.7)
54.38 (3.4)
56.27 (2.6)
57.41 (2.8)

½Rx (ms)

M
44.77 (8.8)
41.68 (8.5)*
42.48 (8.0)*
45.36 (8.7)†

F
57.19 (10)
51.75 (9.2)*
53.90 (8.2)*
56.45 (9.3)*

HbTslope (% • s−1)

M
0.31 (0.1)
0.36 (0.1)*
0.30 (0.1)
0.29 (0.1)

F
0.27 (0.1)
0.30 (0.1)*
0.30 (0.1)
0.28 (0.1)

Hbslope, (%• s−1)

M
0.21 (0.1)
0.25 (0.1)*
0.19 (0.1)†
0.21 (0.1)†

F
0.18 (0.1)
0.20 (0.1)*
0.18 (0.1)*
0.17 (0.1)†

Prolonged changes in contractile data combined with a quicker recovery of hemodynamics suggest low frequency fatigue.
Variation of global mRNA expression in biopsies from the same human muscle
Authors: Sundberg Carl Johan B , Lindholm Malene , Solnestam Beata Werne , Lundberg Joakim , Fischer Helene
Published: June 2021

Human skeletal muscle biopsies obtained from the same muscle are commonly assumed to be very similar from a phenotypic and gene expression perspective, despite very few studies showing this. Therefore, our aim was to investigate differences in global gene expression pattern between and within legs of the same individual.

Five sedentary subjects were included. 2 skeletal muscle samples from the vastus lateralis muscle were taken from each leg (i.e. 2 + 2 biopsies). The time between the first and last biopsy for each individual was 10 to 15 minutes. Isolated RNA was subjected to global mRNA sequencing by synthesis (RNAseq).

In general, there was a very high degree of gene expression level similarity (>99 %) between biopsies within and between legs. Within the same leg, there were 7 genes with a significantly different expression for all subjects, e.g. STAT1, PRG4 (proteoglycan 4) and MYBPH (myosin binding protein H). Between legs, there were 18 differentially expressed genes. STAT1 and PRG4 were found here too, along with PDK4 and ABRA (actin‐binding Rho activating protein). Within the same leg, the number of genes with a significant differential expression varied from 119 to 4 genes. The same two individuals also had the largest and smallest variation comparing two legs.

In conclusion, biopsies taken from the same individual have highly similar gene expression patterns. However, some individuals display larger variation.
Blunted hypoxic ventilatory drive in adult humans with a history of premature birth
Authors: Beshish Arij , Bates Melissa L. , Farrell Emily T. , Pegelow David F. , Eldridge Marlowe W.
Published: June 2021

The hypoxic ventilatory drive (HVD) is blunted in rats exposed to 100% O2 during the perinatal period. This has not been examined in humans. We recruited adults(n=5) born prematurely (<1500 g and <32 weeks gestational age) and given supplemental O2 at birth and age‐matched controls (n=4) to test the hypothesis that subjects born prematurely have a diminished HVD. Subjects performed four breathing trials. In the first pair, subjects were exposed to 7 minutes of 21% and 12% O2 with minute ventilation (VE) and arterial blood gases measured in the last 2 minutes. As expected, we found that all control subjects increased VE in hypoxia (n=4, 0.03±0.1 L/min/ΔmmHg PO2) while, surprisingly, 2/3 premature subjects decreased VE in hypoxia (−0.08±0.03 L/min/ΔmmHg PO2). On a separate day, the trial was repeated with end tidal CO2 (ETCO2) in hypoxia matched to the normoxic levels. Both groups increased VE in hypoxia although the response tended to be lower in the premature group (n=5, 0.06±0.04 L/min/ΔmmHg PO2 vs n=4, 0.08±0.02 L/min/ΔmmHg PO2). Our data support the hypothesis that perinatal hyperoxia blunts the HVD in humans. Interestingly, this effect is diminished when the ETCO2 is controlled, indicating a potential role for the central chemoreceptors. As our sample size expands, we intend to stratify our populations to determine if the duration of perinatal hyperoxia is important.
Impaired cardiac norepinephrine transport in the hypertensive rat
Authors: Shanks Julia , Mane Saras , Ryan Rebecca , Paterson David
Published: June 2021

Hypertension is associated with upregulation of sympathetic cardiac responsiveness both pre and post‐synaptically. Enhanced cardiac norepinephrine (NE) release has been reported in prehypertensive and hypertensive spontaneously hypertensive rats (SHR) compared to normotensive controls (WKY). This phenotype might result from defective NET‐1 transport (uptake‐1). A novel fluorescent substrate of the NE reuptake transporter (NET) was used to investigate NE reuptake kinetics in isolated sympathetic stellate and superior cervical ganglia (SCG) neural cells from SHRs and age‐matched WKY rats. Pre‐hypertensive SHR, and those with established hypertension were studied. Decreased rate of NET (~24%) in stellate ganglion neurons from the SHR at both developmental ages (4 week SHR, n=24, WKY, n=21, P<0.05, unpaired t test, 16 week SHR, n=20, WKY, n=19, P<0.05, unpaired t test) was observed when compared to the WKY response. No difference in re‐uptake rate was seen in cells from the SCG, a major sympathetic ganglia whose innervations predominantly supplies the face. Electrical stimulation of stellate neurons exacerbated the re‐uptake difference of NE between SHR and WKY cells. These results suggest that cardiac sympathetic hyper‐responsiveness is in part due to slowed NE reuptake in the SHR, and this defect occurs before the actual onset of hypertension. Furthermore the NET impairment was not observed in another (non‐cardiac) sympathetic ganglia.
Sex hormones induce gender‐related difference in renal expression of a novel prostaglandin transporter, OAT‐PG, influencing basal PGE2 concentration
Authors: Asano Shinji , Hatano Ryo , Onoe Kimitaka , Matsubara Mitsunobu
Published: June 2021

A rat prostaglandin‐specific transporter (rOAT‐PG) transports PGE2 and colocalizes with 15‐hydroxyprostaglandin dehydrogenase (15‐PGDH), a metabolic enzyme for PGs, in proximal tubules, suggesting that rOAT‐PG is involved in the PGE2 clearance to regulate its physiological function in the renal cortex. We found that mRNA and protein expression levels of rOAT‐PG in the renal cortex were much higher in male rats than in female rats whereas there was no gender difference in the expression level of cyclooxygenase‐2, a key enzyme producing PGE2, and 15‐PGDH in the renal cortex. Tissue PGE2 concentration in renal cortex was lower in male rats than in female rats, suggesting that renocortical PGE2 concentration is primarily determined by the expression level of OAT‐PG which is differently regulated between male and female rats. Castration of male rat led to the remarkable reduction of OAT‐PG expression and significant increase of renocortical PGE2 concentration. These alterations were recovered by the testosterone supplementation. These results altogether suggest that OAT‐PG is involved in local PGE2 clearance in the renal cortex.
Role of drug transporters in the systemic disposition of fluoroquinolones
Authors: Mulgaonkar Aditi , Venitz Jurgen , Sweet Douglas H
Published: June 2021

Previous meta‐analysis of clinical literature found 17 fluoroquinolone (FQ) antimicrobials with sufficient pharmacokinetic data to allow categorization based on renal clearance mechanisms: net renal tubular secretion, net reabsorption, or no net transport. Results suggested interactions with basolateral and apical drug transporters (DTs). Recent literature has implicated several DTs from the solute carrier (SLC) family in disposition of FQs. The objective of this study was to determine if the organic anion transporters (SLC22), OAT1 and 3, interact with the identified FQs, and if so, characterize the kinetics of these interactions using stably transfected cell lines. Preliminary inhibition studies on mouse (m)Oat1, mOat3 and human (h)OAT3 identified sparfloxacin (S), lomefloxacin (L) and levofloxacin (LV) as FQs exhibiting marked transporter interaction. Concentration‐dependency experiments (1 μM [H3] estrone sulfate, 1 min uptake) with mOat3 revealed IC50 values of 74.7 ± 33, 558 ± 51 and 1245 ± 45 μM for S, L and LV (respectively) and 250 ± 21 μM for L on hOAT3. The ~2‐fold difference in IC50 for lomefloxacin on hOAT3 vs. mOat3 suggested possible species differences in renal handling of FQs. Also, some FQs with clinically well‐characterized renal elimination in humans exhibit significant interactions with the studied DTs and these DTs may constitute the first step in their renal tubular secretion in‐vivo.
Fructose‐rich Diet From Weaning To Adulthood: Effects On Sympathetic Activity In Rats
Authors: Andrade Romario Pacheco , Araujo Iara Cristina , Lopes Ingrid Jarzinski , Lopes Bruna Kogici , Santos Fernando , Irigoyen Maria Claudia , Fiorino Patricia , Farah Vera
Published: June 2021

Administration of a diet rich in fructose initiates a series of metabolic events resulting in hypertriglyceridemia, hypertension and glucose intolerance. The purpose of this study was to evaluate the cardiovascular and autonomic modulation induced by fructose intake during 8 weeks. Weaned (21 days old) male Wistar rats (50–60g) were divided in 2 groups and they were followed for 8 weeks: Control (CG) and Fructose (FG, 10% in drinking water). Arterial pressure (AP) and heart rate (HR) were directly recorded using a data acquisition system (Windaq, 2 KHz) in conscious and awake rats. Cardiac autonomic modulation was evaluated using spectral analysis of HR variability and after pharmacological blockade (Atenolol, 8mg/Kg and Atropine, 4mg/Kg). FG showed an increase of mean AP (120±3.1 mmHg) when compared to CG (98±1.6 mmHg). No changes in heart rate were observed, although autonomic balance increased in FG (0.12±0.02) compared to CG (0.04±0.01). Sympathetic tonus was increased in FG (86±7.2 bpm) when compared with CG (49±3.6 bpm) with no differences in vagal tonus. However, intrinsic HR was significantly lower in FG (375±7.2 bpm) when compared with CG (405±8.2 bpm). In conclusion, these data suggest that metabolic changes induced by frutose‐rich diet during life spam in rats are accompanied by hypertension associated with sympathetic overactivity.

Supported by: FAPESP(2009/52556‐7) and PIBIC/Mackpesquisa
Evaluation of ENaC and fluid secretion along the mouse CD: Endothelin‐1 down‐regulates Na reabsorption
Authors: Lynch I. Jeanette , Cain Brian D. , Wingo Charles S.
Published: June 2021

Na homeostasis and the renin‐angiotensin‐aldosterone system are important for systemic arterial blood pressure. Aldosterone and endothelin‐1 (ET‐1) have opposing actions on Na reabsorption (JNa) in the kidney. Aldosterone increases JNa in the collecting duct (CD) by stimulating the epithelial Na channel (ENaC). CD ET‐1 production is stimulated by aldosterone and inhibits ENaC thereby increasing Na and water excretion in part via the ETB receptor. To date, direct measurement of the quantitative effect of ET‐1 on transepithelial JNa in the native mouse CD has not been determined. These studies were performed to examine benzamil‐sensitive (1 μM) JNa in the CD and to test whether ET‐1 (20 nM) inhibits JNa. JNa was measured as 22Na flux in isolated in vitro microperfused CD segments. We observed: 1) the CD exhibits JNa in male and female mice that is regulated, in part, by a benzamil‐sensitive pathway (presumably ENaC) and exhibits fluid secretion that is greater in females than males; and 2) ENaC‐mediated JNa is greater in the CCD than in the OMCD; however, benzamil‐insensitive JNa is present in the CCD and not in the OMCD; and 3) in the presence of ET‐1, ENaC‐mediated JNa and fluid secretion are both inhibited. We conclude: 1) Na reabsorption is mediated by ENaC in the CCD and OMCD and also by an ENaC‐independent mechanism in the CCD; and 2) ET‐1 inhibits ENaC‐mediated Na reabsorption in the CCD. Supported by NIH R01DK82680.
Antioxidant Therapy Targeted to the Folate Receptor Ameliorates Intestinal Inflammation and Injury in a Model of Colitis
Authors: Knight Sarah , Kundu Kousik , Weiss Daiana , Murthy Niren , Taylor W. Robert
Published: June 2021

Intestinal colitis is associated with upregulation of oxidative metabolites that give rise to inflammatory cell infiltration and cellular injury. We designed a novel antioxidant conjugate to selectively target cells at risk from oxidative damage via the folate receptor. We conjugated the antioxidant tempol with folate by an amide bond able to withstand extremes in pH. We used C57bl/6J mice treated daily by gavage with 0.5μM tempol, folate or tempolfolate conjugate for 3 days prior to and during 4 days of 30% dextran sodium sulphate (DSS) administration in drinking water. Body weight was unchanged however colon length was significantly reduced from 7.8cm in controls to 6.9cm in DSS treated mice (P<0.05). Tempol treatment did not prevent a reduction in colon length (P<0.05), however colon length in the tempol‐folate group was not different from control. We observed an increase in colon myeloperoxidase (MPO) activity from control levels of 0.19U/min to 0.36U/min in DSS treated mice. Mice treated with tempol had MPO activity higher than control levels at 0.25U/min compared with 0.14U/min in the tempol‐folate treated group. These data indicate that tempol‐folate reduced the inflammatory response to DSS treatment. Occult blood was also reduced by tempol‐folate compared with tempol. In conclusion, we have developed a novel therapeutic strategy to target oxidative damage and prevent injury in a mouse model of colitis.
Gli1 is activated via the non‐classical TGFβ/smad 3 pathway rather than the sonic hedgehog pathway in a murine model of hepatic encephalopathy
Authors: McMillin Matthew , Galindo Cheryl , Frampton Gabriel , Pae Hae Yong , DeMorrow Sharon
Published: June 2021

Hepatic encephalopathy (HE) is a serious neurological complication of liver failure. Our understanding of the events leading to HE is quite limited. The sonic hedgehog (Shh) pathway is upregulated during liver damage and Shh ligands have been found in the blood stream. Our aims were to evaluate the molecular changes leading to HE. Injection of azoxymethane (AOM) into mice caused liver damage and cognitive decline that culminated in coma. There was an increased release of Shh into the serum and an increase in Gli1 expression, the downstream effector of Shh signaling, in the neurons of the frontal cortex in the brain. Inhibition of Shh signaling inhibited the release of Shh ligands in the serum, but failed to prevent the activation of Gli1 or the neurological decline that occurred after AOM treatment. TGFβ could also be detected in the serum of AOM‐treated mice and resulted in the activation and nuclear translocation of SMAD3 in neurons of the frontal cortex. Pretreatment of mice with a neutralizing TGFβ antibody, prior to AOM injection prevented the activation of SMAD3, the increase of Gli1 expression and delayed the cognitive decline of the mice. These data suggest that during acute liver failure, TGFβ is released into the serum and may cause SMAD‐3 activation and the subsequent increase in Gli1 expression in the affected brain regions and that this sequence of events may be important in the neurological progression of HE to coma.
Vascular PI3K‐Akt Signaling Contributes to Peripheral NMDAR‐mediated Pressor Response in Conscious Rats
Authors: McGee Marie , Abdel-Rahman Abdel
Published: June 2021

Evidence has implicated the PI3K‐Akt‐NOS signaling pathway in neuronal responses mediated via NMDAR activation. The involvement of this pathway in the peripheral NMDAR‐mediated pressor response is unknown. We tested the hypothesis that vascular PI3K‐Akt‐NOS signaling contributes to NO and ROS generation and the subsequent pressor response mediated by peripheral NMDAR activation. Hemodynamic studies were conducted in conscious male Sprague Dawley rats that were pretreated with the selective inhibitor of: (i) eNOS, N5‐(1‐iminoethyl)‐l‐ornithine (L‐NIO), (ii) nNOS, Nw‐propyl‐l‐arginine (NPLA) or (iii) the upstream PI3K‐Akt inhibitor, Wortmannin. L‐NIO produced no change while NPLA pretreatment significantly attenuated the NMDA‐mediated pressor response. These findings are the first to implicate nNOS‐derived NO in the peripheral NMDAR signaling. Pretreatment with the PI3K inhibitor, Wortmannin significantly suppressed the NMDA mediated pressor response and the associated increases in vascular nitrate and reactive oxygen species levels. Collectively, these novel findings suggest that activation of the PI3k‐Akt‐nNOS signaling pathway serves as a molecular mechanism for the peripheral NMDAR‐mediated pressor response in conscious rats [NIH grant AA07839].
Vasoactive effect of SWCNT exposure to oral mucosal arterioles
Authors: Frame Mary D , Sitharaman Balaji , Chowdhury Sayan M , Avti Pramod
Published: June 2021

Our purpose was to determine whether lipid coated single‐walled carbon nanotubes (SWCNTs) were vasoactive in oral mucosal arterioles; this would mimic the oral component to inhalation exposure. Observations were made of terminal arterioles in the hamster cheek pouch using intravital microscopy (pentobarbital 70mg/kg, N=12). SWCNTs were suspended in 0.1% DSPE‐PEG (1,2‐distearoyl‐sn‐glycero‐3‐phosphoethanolamine‐N‐ [amino(polyethylene glycol)] in physiological saline. The SWCNT‐PEG‐DSPE solution was applied to the abluminal surface of arterioles (micropipette) at 0, 10, 50 and 100 ug/ml SWCNT for 60 seconds. [Previously, 7–10 ug/ml SWCNT i.v. was not toxic over 28 days exposure.] Diluent alone was not significantly vasoactive. A sustained dilation peaked at 50 ug/ml, and decreased at 100 ug/ml. Dilation was significantly greater in the smaller terminal arterioles than in the larger upstream arcading arterioles which feed the terminal arterioles. Complete recovery from dilation was observed with removal of the SWCNT. Finally, comparing vasoactive responses before vs. after SWCNT exposure, there was a diminished dilation to both bradykinin and adenosine, while constriction to phenylephrine was unaffected. SWCNT exposure induces a dose dependent dilation with residual compromised dilatory capability. Thanks to NIH HL55492; AHA 0655908T; NIH 1DP2OD007394‐01; DoD BCRP (W81XWH‐10‐1‐0521).
Brassinosteroid enhances C57BL/6J mice treadmill endurance
Authors: Esposito Debora , Tuazon Marc , Henderson Gregory C , Komarnytsky Slavko , Raskin Ilya
Published: June 2021

The 28‐homobrassinolide (HB) has been show to have multiple anabolic effects on protein synthesis and lean body mass in healthy rats. Using 7 wk old male C57BL/6J mice, we examined the effects of 28 d of HB treatment on grip strength, exercise performance, and HB‐gene interactions in mouse hindlimb muscles. Animals received 60 mg/kg of HB in corn oil (vehicle) via oral gavage daily and underwent bi‐weekly treadmill exercise on d 1 (baseline), 14 and 28 of the experiment. Significant increases in treadmill performance and enhanced grip strength were observed with HB. Components of PI3K/Akt, AMPK, and myogenesis signaling pathways were significantly upregulated by HB treatment in skeletal muscle tissues. These findings indicate that brassinosteroid alone induces an integrated response that includes structural and metabolic changes in skeletal muscles resulting in greater endurance capacity. This study was supported in part by the 5P50AT002776‐05 grant from NCCAM.
Intravenous delivery of AAV9 encoding urocortin 2 increases cardiac function in normal mice
Authors: Lai N. Chin , Hammond H. Kirk , Tang Tong , Guo Tracy , Tang Rouying , Parikh Jay , Gao Meihua
Published: June 2021

Gene transfer for the treatment of cardiovascular diseases is conceptually attractive, but difficulty in obtaining high yield expression in the heart in a manner that can be easily and safely applied has been an impediment to progress. In this study, we tested the efficacy of an adeno‐associated virus 9 (AAV9) vector encoding urocortin 2 (Ucn2), which is a vasoactive peptide in the corticotropin‐releasing factor family that has beneficial effects in animals and patients with heart failure. AAV9.Ucn2 (5×1011 genome copies) was delivered by intravenous injection. Four weeks after gene transfer, AAV9 vector DNA (qPCR) was increased by 115,854‐fold and Ucn2 mRNA by 10,523‐fold in liver vs control. Ucn2 DNA and mRNA were increased 15‐ and 34‐fold, respectively, in the left ventricle (LV). Also seen were increases in: plasma Ucn2, LV +dP/dt, LV −dP/dt, and LV developed pressure (Table).

Conclusion

Intravenous delivery of AAV9.Ucn2 provides sustained increases in serum Ucn2 and LV contractile function, indicating a potential role for paracrine‐based gene transfer for the treatment of heart failure.

Control
AAV9.Ucn2
Fold Δ*

Liver AAV9 DNA (copies/ng)
0.26±0.05
3×105±3906
115,854

Liver Ucn2 mRNA (relative copies)
166±17
17×105±15×104
10,523

LV AAV9 DNA (copies/ng)
18±6
301±45
15

LV Ucn2 mRNA (relative copies)
34±9
958±299
34

Plasma Ucn2 (ng/ml)
2.6±0.1
6.0 ±0.3
2.3

LV +dP/dt (mmHg/s)
4,103±335
6,115±334
1.5

LV −dP/dt (mmHg/s)
3,744±335
5,631±365
1.5

LV Pressure (mmHg)
70±3
88±3
1.3

Heart Rate (beats/min)
439±9
420±14
0.9

Control (n=14); AAV9.Ucn2 (n=9); except DNA: (n=9, ea group) & plasma Unc2 (n=4 ea group);

p<0.05 to <0.001, except for heart rate
Secretin Regulates the Expression of the Renin‐Angiotensin System in Rat Cardiac Fibroblasts
Authors: Afroze Syeda , Nizamutdinov Damir , Feng Hao , Gerilechaogetu Fnu , Jensen Kendal , Munshi MK , Golden Honey , Dostal David , Alpini Gianfranco , Glaser Shannon
Published: June 2021

Secretin (SCT) is a peptide hormone produced mainly by the S cells of the duodenum that exerts its biological effects through the G‐protein coupled secretin receptor (SCTR). Studies have indicated that SCT stimulates cardiac adenylyl cyclase activity and exerts a positive inotropic action in the heart. The expression of SCT/SCTR or the effect of SCT on cardiac fibroblasts has not been fully explored. The AIM of our study was to determine the expression of SCT and SCTR in cardiac fibroblasts and to explore the effect of SCT on cardiac fibroblast expression of the renin‐angiotensin system (RAS).

Results

Rat cardiac fibroblasts express SCT and SCTR by qPCR, IF and FACS. SCT (0.43 ± 0.21 ng/ml) is secreted into supernatants collected from fibroblasts after 6 h. SCT stimulates an increase in intracellular cAMP (but not IP3) and induces an increase in PCNA expression (an indicator of increased cell proliferation) at 6 h. SCT decreases gene expression of the profibrogenic RAS components, AT1a, AT1b and ACE and upregulates expression of AT2.

Conclusions

The findings suggest that SCT may have antifibrotic properties, which are protective during cardiac dysfunction.
In vivo screening of chromatin remodelers in zebrafish reveals proteins governing cardiac growth
Authors: Monte Emma , Mouillesseaux Kevin , Kimball Todd , Vondriska Thomas M , Chen Jau-Nian , Franklin Sarah
Published: June 2021

Cardiac hypertrophy is a common precursor to heart failure. Hypertrophic cardiomyocytes undergo a global change in gene expression (conserved between humans and animal models of disease) that recapitulates aspects of early development. To investigate the role of chromatin reorganization in this phenotype, mass spectrometry was used to quantify changes in the chromatin proteome during heart disease in mice. Candidate chromatin remodelers, including nucleolin (Ncl), which exhibited a four‐fold increase in the hypertrophied mouse heart, were further tested in zebrafish.

In situ hybridization for Ncl demonstrated expression in the developing heart. Morpholinos were used to knockdown (KD) Ncl in zebrafish embryos, resulting in a global increase in histone H3K9 trimethylation, indicative of a more heterochromatic genome. In situ hybridization revealed that the normal restriction of BMP4 to the atrial/ventricular boundary was lost in KD embryos, suggesting delayed ventricular differentiation. Microscopy of 48‐hour embryos showed defects in chamber development in 21% of fish, including delayed (15%) or reversed (5%) looping. Impaired contractility led to blood pooling and edema in a small (3%) population of fish. We conclude that alterations in Ncl expression and activity contribute to chromatin remodeling in the setting of mammalian heart hypertrophy and zebrafish development. Supported by NIH/NHLBI.
Novel Rapid‐Multiple‐Short‐Cycle Preconditioning Stimuli Induces Robust Cardioprotective Signaling Mechanisms and Protects the Heart against in vivo Ischemia Reperfusion Injury: An Effective Approach with Clinical Justification
Authors: Zweier Jay L. , Yang Fuchun , Nishijima Yoshinori , Chen Chun-An , Yang Changjun , Varadharaj Saradhadevi , Talukder M.A. Hassan
Published: June 2021

Despite the large body of basic research, the well‐proved phenomenon of ischemic preconditioning (IPC) still lacks its clinical benefits. In clinics, balloon inflation/deflation cycles during coronary interventions are restricted to <1‐min in duration; and multiple balloon inflations/deflations are required for complex lesions. Recently, we have shown that rapid brief 60‐sec cycle of ischemia/reperfusion (I/R) induces small burst of reactive oxygen species (ROS) formation that could trigger IPC like protection. Therefore, to investigate whether a novel rapid‐multiple‐short‐cycle preconditioning (RPC) stimuli induces in vivo cardioprotection, rats were subjected to in vivo regional I/R (30‐min I and 24‐hr R) preceded by RPC (6 cycles of 60‐sec I and 60‐ sec R) or classical IPC (C‐IPC; 3 cycles of 5‐min I and 5‐min R). Cardiac function, infarction, ROS generation, survival kinase and nitric oxide synthase (NOS) pathways were characterized. RPC conferred robust cardioprotection compared to C‐IPC. This was manifested by reduced myocardial infarction, improved postischemic cardiac function, reduced ROS generation, effective activation of Akt and ERK1/2, as well as enhanced NOS expression and activation. Thus, this novel IPC stimuli RPC is highly effective in preserving myocardial viability and function in the setting of acute myocardial I/R. (Funding: NIH grants HL63744, HL65608, and HL38324 to JLZ)
In cardiomyocytes LPS‐induced activation of IκBα and COX‐2 expression are mediated in part by the hexosamine biosynthesis pathway
Authors: Lu Xiaoyin Tina , Zou Luyun , Chatham John C
Published: June 2021

We have previously reported that hyperglycemia (HG) augmented the induction iNOS by LPS and this was due at least in part by increased flux through the hexosamine biosynthesis pathway (HBP). COX2 is induced by a number of stresses such as ischemia, pro‐inflammation cytokines, and mitogenic responses and this is regulated in part by NF‐κB signaling. Therefore, the goal of this study was to determine if in neonatal rat ventricular cardiomyocytes (NRVM) COX2 expression was also regulated by the HBP. LPS treatment for 6hrs (2μg/ml) significantly increased both P‐IκBα and COX2 expression in NRVM and whereas HG augmented the LPS‐induced increase in P‐IκBα its effect on COX2 was less pronounced. Interestingly however, inhibition of GFAT with either DON (20 μM) or Azserine (40 μM) substantially decreased the LPS‐induced increased COX‐2 and P‐IκBα both under normal and HG conditions. These results suggest that activation of the HBP contributes to the pro‐inflammatory effects of LPS and that this is augmented at least in part under HG conditions.
Elevated reactive oxygen species and increased mononuclear NADPH oxidase expression in type 2 diabetes patients
Authors: Deo Shekhar H , Zimmerman Matthew C , Fadel Paul J
Published: June 2021

Type 2 diabetes (T2D) patients exhibit increased oxidative stress. However, the sources and mechanisms contributing to the elevation in oxidative stress remain unclear. Herein, we sought to examine the potential contribution of the angiotensin II‐NADPH oxidase pathway in mediating the increase in oxidative stress in T2D. In 5 T2D and 5 age‐matched controls, protein expression of NADPH oxidase subunits (gp91phox, p22phox, p47phox and p67phox) and angiotensin II type 1 receptor (AT1R) were assessed at rest from peripheral blood mononuclear cells using western blot. In addition, electron paramagnetic resonance (EPR) spectroscopy was used to measure blood total reactive oxygen species (ROS) and superoxide. T2D showed elevated resting total ROS (1.3±0.2×107 T2D vs. 0.6±0.1×107 controls, EPR a.u.; P<0.05) and superoxide (0.3±0.1×107 T2D vs. 0.1±0.2×107 controls, EPR a.u.; P<0.05) as well as protein expression of all NADPH oxidase subunits (P<0.05 vs. controls). AT1R expression was not different between groups. However, for all subjects, AT1R expression was significantly related to all NADPH oxidase subunits (e.g., gp91phox; R2 = 0.53; P<0.05), which in turn was related with total ROS (gp91phox; R2 = 0.59; P<0.05). These preliminary findings suggest that T2D exhibit greater ROS and superoxide that contributes to resting oxidative stress potentially via an up‐regulation of the AT1R‐NADPH oxidase pathway.
Real‐time continuous monitoring of myocardial blood flow response to dynamic exercise with laser Doppler flowmetry in conscious rats and mice
Authors: Tsuchimochi Hirotsugu , Sonobe Takashi , Shirai Mikiyasu
Published: June 2021

During exercise, coronary blood flow increases to match the augmented myocardial oxygen demand due to the increase in cardiac workload. The mechanisms involved in how the coronary circulation achieves an adequate distribution of blood flow to all working myocardial cells remains largely unknown. The use of genetically‐modified animals will be needed to clarify its mechanisms. Here, we have developed a versatile technique to measure regional myocardial blood flow (MBF) in conscious small laboratory animals with laser Doppler flowmetry (LDF). Each animal was anesthetized with isoflurane and a pair of LDF probes (JF1, Omegawave Inc., Japan) was placed on the left ventricular free‐wall and the tips of the probe were fixed with cyanoacrylate glue. We succeeded in measuring regional MBF which occurred predominantly during diastole in both conscious rats and mice. MBF, as well as heart rate, increased rapidly at the onset of treadmill exercise, and the magnitudes of the responses to exercise were dependent on the treadmill speed. Moreover, beta‐adrenergic blockade with atenolol (4 mg/kg i.p.) significantly attenuated the MBF response to treadmill exercise. In conclusion, LDF with the use of flexible probe will become a powerful tool for investigating the regional MBF responses to various stimulus in the conscious state as well as under anesthesia of experimental animals.
ZINC, IGF‐1, AND FOOD INTERVENTION IN MALNOURISHED PREGNANT WOMEN ON CHILDREN BODY HEIGHT AFTER 6 YEARS IN INDONESIA
Authors: Daud Nurpudji Astuti
Published: June 2021

This study is to investigate effects of zinc and food supplementation on malnourished pregnant women to child body height after 6 years old in Takalar district, Indonesia. Cohort retrospective as a design study (subjects was from double blind study 2003). Samples were children 6 years old, divided into three groups: (1) had zinc, food supplementation history (54 subject), (2) group had food supplementation (20 subject), (3) without zinc and food supplementation (50 subject). Child Growth status is calculated by Marshall and Schwan formula.

The study showed, average of weight gain during 6 years period for boys in group one, two and three were 0.181, 0.181, and 0.177 respectively. For girls were 0.177, 0.169, and 0.168. Children born from mother with zinc and food supplementation had high zinc and IGF‐1 level compared without zinc or food supplement history. The average of height gain (cm/month) for boys in group 1, 2 and 3 were 0,786, 0,786 and 0,773 respectively. For girls were 0.791, 0.739 and 0,672. There were no significant differences of food intake for child growth (weight and height gain status). Rate of weight and height growth of children in group 1 and 2 were better and higher than children in group 3.

Nutrition intervention for malnourished pregnant women continuously (zinc and food supplementation) to prevent growth retardation in children is needed.
Mechanisms related to the thyroid hormone (TH)‐induced vasorelaxation: contribution of reactive oxygen species (ROS) and purinergic signaling
Authors: Basso Cristina Ribas Fürstenau , Barreto-Chaves Maria Luiza Morais
Published: June 2021

This study aims to determine the contribution of ROS and purinergic signaling to the TH‐induced vasorelaxation. In vitro (T3 added to VSMC primary culture) and in vivo (experimental hyperthyroidism developed in rats) protocols were performed to evaluate Nox1 and 4, and P2Y1,2, and 6 expression by Western blot. Preliminary results show that T3 (10‐7 M for 24 hours) diminished both Nox1 and Nox4 expression in VSMC (n=3). The experimental hyperthyroidism was confirmed by classical cardiac hypertrophy induced by T3 treatment (7 ìg/100 g body weight/day, 14 days ) since the ratio body weight (g)/ tibia's length (mm) was significantly higher in hyperthyroid rats (33.17 ± 3.34, n=15 vs 27.49 ± 2.24, n=14 in control group, p<0.05). Besides, total T4 levels measured in blood serum were significantly diminished in hyperthyroid animals (4.88 ± 0.599 in control group vs non‐detectable levels in T3‐treated rats), confirming that the feedback loop in TH production and secretion was satisfactorily working. Analysis of protein expression showed a diminution in P2Y6 expression and a modulation in Nox4 expression in hyperthyroid rats. Although additional experiments need to be performed, the present data point out the participation of both systems (ROS production and purinergic signalling) in the TH‐induced vasorelaxation. Source of research support: FAPESP and CNPq.
Racial Differences in Blood Pressure after Exercise in Young Individuals
Authors: Yan Huimin , Ranadive Sushant M , Sun Peng , Lane Abbi D , Kappus Rebecca M , Fahs Chris A , Rossow Lindy , Fernhall Bo
Published: June 2021

Hypertension prevalence is higher in Caucasians (CA) than in Chinese (CH), and high blood pressure (BP) is associated with greater cardiovascular disease risk. However, little is known about the response of BP to an acute bout of exercise.

18 college age young CH (height: 170 ± 2 cm, weight: 63 ± 3 kg, BMI: 21.8 ± 0.6 kg/m2) and 19 CA (height: 171 ± 3 cm, weight: 73 ± 4 kg, BMI: 24.7 ± 0.8 kg/m2) underwent measures of BP and central (carotid‐femoral) and leg (carotid‐femoral) pulse‐wave velocity (PWV) before, 15 min, and 30 min after a graded maximal aerobic exercise test (EX) on a stationary bike by applanation tonometry.

Diastolic BP was increased in CA but decreased in CH following exercise (CH: from 64 ± 2 to 61 ± 3 to 62 ± 2 mmHg; CA from 65 ± 2 to 67 ± 2 to 67 ± 2 mmHg, race by time interaction, p<0.05). Systolic BP changed with time (CH: from 112 ± 3 to 115 ± 4 to 111 ± 3 mmHg; CA from 118 ± 3 to 121 ± 3 to 116 ± 3 mmHg, p<0.05), but no racial differences. A significant time by race interaction was found in leg PWV (CH: from 8.5 ± 0.4 to 6.9 ± 0.4 to 7.3 ± 0.4 m/s; CA: from 8.6 ± 0.4 to 8.3 ± 0.3 to 8.0 ± 0.3m/s, p< 0.05). Central PWV was not statistically different between CA and CH. Statistically adjusting for BMI did not change these findings.

This is the first study to note racial differences in the BP and peripheral arterial response to exercise, suggesting increased BP in young CA and differences in peripheral arterial stiffening following exercise compared to CH.
Acute aerobic exercise increases AdipoR1 and RAGE proteins and decreases HSP60 protein in skeletal muscle of physically inactive older adults
Authors: Markofski Melissa M , Timmerman Kyle L , Reidy Paul T , Dickinson Jared M , Walker Dillon K , Timmerman Jennifer Z , Rasmussen Blake B , Volpi Elena
Published: June 2021

Aerobic exercise training reduces inflammation and improves insulin sensitivity. Less known are the acute effects of aerobic exercise on skeletal muscle in older, physically inactive adults. We present preliminary data from a study designed to determine the effects of acute aerobic exercise on skeletal muscle markers of inflammation in non‐obese (n=9; BMI: 26.4±1.9 kg·m−2) inactive (3822±1502 steps/day; VO2peak: 22.7±3.5 mL·kg−1·min−1) older adults (72.9±3.9 yrs). After collection of a basal, resting muscle biopsy from the vastus lateralis, subjects walked on a treadmill for 45 min at 70% heart rate reserve, and muscle biopsies were collected 1 and 3 hrs post‐exercise. Protein expression was determined by Western blot. 1 hr post‐exercise, AdipoR1 and receptor for advanced glycation end product (RAGE) were 40.1% and 24.7% higher (p<0.05) than basal. When compared to basal, 3 hrs post‐exercise AdipoR1 was 42.6% higher, heat shock protein 60 (HSP60) was 41.4% lower, and there was a trend for elevation of toll‐like receptor 4 (32.3%). In conclusion, an acute bout of aerobic exercise in physically inactive older adults affects receptors and a ligand (e.g. HSP60) associated with inflammation. These acute effects on inflammation may be necessary for aerobic exercise to initiate muscle adaptation to increased activity in previously sedentary older adults. Future studies are warranted.

NIH R01 AG030070, P30 AG024832
Cervical interneuron bursting during hypoxia in anesthetized rats
Authors: Sandhu Milap S , Gonzalez-Rothi Elisa J , Lee Kun-Ze , Maling Nicholas , Lane Michael A , Reier Paul J , Baekey David M , Sanchez Justin C , Fuller David D
Published: June 2021

Cervical spinal laminae VII‐X contain interneurons that are synaptically coupled to phrenic motoneurons (PMNs). Neurophysiological studies suggest these cells are not necessary for transmission of inspiratory drive, but their role in modulating PMN output during respiratory stimulation or plasticity is unknown. In ongoing studies, we are exploring the role of cervical interneurons in hypoxia‐induced phrenic plasticity. Since spinal serotonin (5‐HT) can induce phrenic plasticity, we first examined whether pre‐phrenic cervical interneurons are innervated by 5‐HT. 5‐HT immunoreactivity was robust throughout cervical laminae VII‐X and in the immediate vicinity of pre‐phrenic interneurons identified with a transynaptic retrograde tracer (pseudorabies virus, delivered to the diaphragm). Cervical interneuron activity was recorded in anesthetized, paralyzed, vagotomized and ventilated adult rats. Initial results indicate that tonic and phasic bursting interneurons are distributed throughout the C3‐C5 intermediate gray matter. During respiratory stimulation with hypoxia a subset of the cervical interneurons showed an increase in burst frequency, and in some cases switched from tonic to phasic bursting. Cervical interneurons are anatomically positioned to modulate phrenic motor output, but their potential role in initiating and/or maintaining phrenic plasticity remains to be determined.
Effects of edible mushrooms on plasma levels of anti‐bovine collagen II IgG antibodies in DBA1 mice with collagen‐induced arthritis
Authors: Kuvibidila Solo
Published: June 2021

Collagen‐induced arthritis (CIA) in mice is commonly used to study rheumatoid arthritis (RA) biology. Plasma levels of anti‐collagen II antibodies (antiCIIA) are usually elevated in CIA and RA patients. While blockade of anti‐CIIA prevents CIA, antiCIIA injection promotes it. We previously reported that 5% dry shiitake (SM) and white button (WBM) mushrooms reduced CIA incidence and severity in DBA1 mice. The current study was conducted to assess the role of antiCII IgG in reduced CIA severity in mice fed mushroom‐fortified diets. Plasma antiCII IgG and cytokines were measured by ELISA in mice fed either the AIN76 diet or the same diet fortified with 5% WBM, SM, and portabella (PM) (n = 6–8) for 6 weeks, and immunized with 100 μg bovine CII, followed by 50 μg LPS on d 21, and euthanized on d 31 post‐CII. In mice with CIA index ≥ 3, mushrooms slightly reduced plasma antiCII IgG (14% SM [p <0.05], 10% WBM, 6% PM) compared to those fed the AIN76 diet. Within the same dietary treatment group, plasma antiCII IgG levels were not different between immunized mice without and those with swollen paws. While WBM and SM decreased sICAM1 and TNF‐α by 15–53% and increased IL‐6 levels by 1.6–3fold (p <0.05), PM only had minor effects. CIA index was positively associated with antiCII IgG and IL‐6 (p<0.01) but not TNF‐α levels. Data suggest that mushrooms very likely modulate CIA through IL‐6 but not antiCII IgG; but an effect on IgG isotypes cannot be ruled out. Supported by the Mushroom Council/USDA Grant 580790706 and funds from Oklahoma State University