bluuebunny/crossref_metadata_2025 huggingface dataset:
Breast cancer cell homing to bone in vivo is modulated by zoledronic acid (676.6)
Authors: Reeves Kimberley , Albon Jacob , Eaton Colby , Brown Nicola
Published: June 2021

Cancer metastasis to the skeleton causes mortality in breast cancer, however the mechanisms involved in bone metastases development remain unclear. A metastatic initiating cell (MIC) phenotype CD44+/CD24‐/ALDHhigh has been identified in breast cancer. Zoledronic acid (zol), a bone anti‐resorptive bisphosphonate is currently the only agent with proven benefit in breast cancer. This study investigates whether zol modulates MIC homing in the early stages of breast cancer bone metastasis using the Dorsal Skinfold Chamber (DSC) metatarsal model in vivo.

Following DSC metatarsal implantation, mice were treated weekly either before or after intra‐cardiac MDA‐MB‐231 tumour cell injection (1x105) for 4 weeks (Zol: 25µg/kg/injection=100μg/kg or PBS control). MDA‐MB‐231 cells were sorted into ALDHhigh and ALDHlow subpopulations using the ALDEFLUORkit and then injected into groups of mice (n=6), treated with zol before tumour cell injection. Chamber tissue and metatarsal recordings were made every 48hr for up to 4 weeks to identify tumour cell homing. At the end of the study, tissue was harvested and processed for µCT, multi‐photon analysis, histology and immunohistochemistry.

Zol treatment reduces MDA‐MB‐231 cells homing to bone by 76% (before) and 62% (after) compared to control animals. ALDHhigh cells home to bone in significantly (p<0.05) higher numbers than ALDHlow cells (Day 25: 10.7±1.3 cells vs. 1.1±0.6 cells). Week zol administration (before tumour cell injection) does not reduce MDA‐MB‐231 cells bone‐homing.

Weekly treatment with zol reduces breast tumour cell homing to bone, whereas ALDHhigh cell homing appears resistant to zol treatment in this model.
SGLT inhibitors attenuate NO‐dependent vascular relaxation in mouse pulmonary artery (677.3)
Authors: Han Ying , Cho Young‐Eun , Ayon Ramon , Dai Anzhi , Yuan Jason X.‐J. , Makino Ayako
Published: June 2021

Sodium‐glucose cotransporter 2 (SGLT2) inhibitors are a new class of oral drugs for the treatment of type 2 diabetes and they reduce plasma blood glucose levels by inhibiting renal glucose reabsorption. However, there are no data focusing on microvascular or cardiovascular outcomes. The present study is designed to determine whether SGLT inhibitors, phlorizin (nonspecific SGLT1/SGLT2 inhibitor) and canagliflozin (SGLT2 specific inhibitor), regulate vascular relaxation in mouse coronary and pulmonary arteries. Isometric tension experiments were performed to evaluate vascular function. Phlorizin significantly decreased sodium nitroprusside (SNP, NO donor)‐dependent vascular relaxation in pulmonary arteries; however phlorizin did not alter SNP‐dependent relaxation in coronary arteries. Contrary to our expectation, phlorizin had no effect on either 8‐Bromo‐cGMP (a cell‐permeable cGMP analogue) or papaverine (phosphodiesterase inhibitor)‐induced relaxation in mouse pulmonary arteries. Canagliflozin inhibited SNP‐dependent vascular relaxation in a dose dependent manner. RT‐PCR data showed that SGLT2 mRNA expressed in small intestine, HEK293, pulmonary endothelial cells and pulmonary smooth muscle cells, while SGLT1 mRNA expressed only in small intestine. These results indicate that SGLT2 regulates NO‐dependent relaxation in mouse pulmonary arteries.

Grant Funding Source: Supported by the NIH/NHLBI grant HL115578
Vasomotion in vivo ő the role of the endoplasmic reticulum (677.9)
Authors: Aalkjaer Christian , Nyvad Jakob , Straarup Marthe , Matchkov Vladimir
Published: June 2021

Low frequency high amplitude oscillation with antiphase oscillations of Ca2+ in endothelial and smooth muscle cells are seen in rat mesenteric small arteries in vitro after inhibition of the endoplasmic reticulum (ER) Ca2+ APTase. We here investigated the role of ER for vasomotion in vivo. Rats were anesthetized with Hypnorm/Dormicum (HD), Ketamin/Xylazin (KX) or Isofluran (ISF) and the intestine exteriorized through a laparotomy. A small mesenteric artery segment was positioned in a 100 μl reservoir and the diameter monitored. Constriction to norepinephrine (NE) was substantially reduced with HD compared to KX and ISF, and further experiments were done with KX. At intermediate NE concentrations vasomotion occurred. With cyclopiazonic acid (CPA) present frequency was reduced and amplitude increased. Inhibition of small and intermediate conductance K+ channels had no effect on vasomotion without CPA, but reduced the amplitude and increased the frequency in the presence of CPA. We conclude that HD is not a suitable anesthetic for studying adrenergic mechanisms in rats, and that small and intermediate conductance K+ channels are important for vasomotion in vivo, but only when the endoplasmic reticulum is inhibited.
Divergence in depressive symptom severity and vascular dysfunction with gender in rats with unpredictable chronic mild stress (680.20)
Authors: Stanley Shyla , Brooks Steven , Butcher Joshua , D'Audiffret Alexandre , Skaff Paulina , Chantler Paul , Frisbee Jefferson
Published: June 2021

Chronic stress leads to many negative health outcomes proportional to the severity of developed depressive symptoms. This includes vascular reactivity, as endothelial function is compromised by 8 weeks of unpredictable chronic mild stress (UCMS). To extend this, we imposed 8 weeks of UCMS on healthy rats of both genders from 9 weeks of age. In males, plasma cortisol and depressive symptom severity (coat status, anhedonia, delayed grooming) were elevated. Endothelial‐dependent dilation to methacholine/acetylcholine was impaired in conduit arteries/skeletal muscle arterioles, indicating a near complete loss of NO bioavailability and increased production of TxA2 with elevated ROS. Endothelium‐independent dilation was intact. However, in females, depressive symptoms and plasma cortisol increases were more severe, although impairments to vascular reactivity were attenuated. The effects of elevated ROS and constrictor prostanoids were also blunted. These results suggest that, in comparison to males, female rats are more susceptible to chronic stress in terms of the severity of depressive behaviors, but that the subsequent development of vasculopathy is blunted.

Grant Funding Source: Supported by National Institutes of Health and American Heart Association
Evaluation of the cardiovascular changes in juvenile and adult female rats submitted to chronic intermittent hypoxia (680.3)
Authors: Souza George , Bonagamba Leni , Amorim Mateus , MORAES DAVI , Machado Benedito
Published: June 2021

Chronic intermittent hypoxia (CIH) induces hypertension in juvenile and adult male rats due to an increase in sympathetic activity. There is experimental evidence that changes in respiratory network are driving the sympathetic overactivity in this model. Female rats are more resistant to the development of hypertension in several experimental models. In this study, we evaluated the cardiovascular changes in juvenile and adult female rats in response to CIH. Juvenile female rats were exposed to CIH for a 10 days protocol and adult female rats to a 35 days protocol and at the end of the protocols direct measurements of arterial pressure were performed in control and CIH rats. Juvenile female rats exposed to CIH (n=30) presented significant increase in arterial pressure compared to their respective control [n=34 (94±1 vs 86±1 mmHg, p<0.05)], as well adult female rats exposed to CIH (n=27) when compared to their respective control [n=26 (111±1 vs 104±1 mmHg, p<0.05)]. We concluded that juvenile and adult female rats submitted to CIH develop high arterial pressure similarly to male rats after CIH, suggesting that sexual characteristics and hormones are not playing a protective role in this experimental model of hypertension.

Grant Funding Source: Supported by CAPES, CNPQ and FAPESP
Effects of gender on mitochondria‐derived middle cerebral artery function in rats (680.7)
Authors: Rutkai Ibolya , Dutta Somhrita , Walter Korey , Katakam Prasad , Busija David
Published: June 2021

Significant gender differences in the composition and functionality of cerebral arteries have been shown, but effects of mitochondrial‐derived responses have not been examined. Our study tested whether mitochondrial mechanisms promoting changes in cerebral vascular tone are different in male and female rats. We examined the effects of a mitoKATP opener, diazoxide (DZ), bradykinin (BK), and sodium nitroprusside (SNP) on isolated, pressurized MCAs from 9 week old male and female Sprague‐Dawley rats. We measured levels of mitochondrial proteins of the voltage dependent anion channel (VDAC), Complex V, the fission protein DRP‐1, and endothelial and neuronal nitric oxide synthase (NOS). Levels of Complex V and VDAC were similar in male and female rats, but the protein expression of phosphorylated DRP‐1, total and phosphorylated eNOS and nNOS in MCAs were significantly higher in females than in males. However, DZ induced vasodilation was greater in male MCAs (21.1±4.5 μm to 50μM, 44.8±4.7 μm to 100 μM) compared with female MCAs (13.3±2 μm to 50 μM, 35±3.2 μm to 100 μM) whereas vasodilator responses to BK and SNP were similar for both sexes. Treatment with L‐NAME and the denudation of endothelium abolished the DZ induced vasodilation to all of the DZ concentrations in female MCAs. Our data suggest that mechanisms underlying gender differences in mitochondrial derived vasoreactivity are associated with NO production.

Grant Funding Source: NIH grants HL‐077731, HL‐030260, HL‐065380, and HL‐093554.
Evaluation of the respiratory and cardiovascular activities in juvenile rats submitted to sino‐aortic denervation (681.9)
Authors: Amorim Mateus , Bonagamba Leni , Souza George , Moraes Davi , Machado Benedito
Published: June 2021

Rats submitted to sino‐aortic denervation (SAD) present normal basal sympathetic activity and mean arterial pressure (MAP) few days after the surgical procedure. In the present study, we evaluated the cardiovascular and respiratory changes 3 and 10 days after SAD. Male juvenile Wistar rats were submitted to SAD or sham surgery and four groups were evaluated 3 (SAD and Sham) and 10 days (SAD and Sham) after the surgery. Cardiovascular and respiratory parameters were recorded in awake rats and the arterial baroreflex was tested to verify the efficacy of SAD. It was observed that MAP in all experimental groups was similar. The variability of MAP was significantly higher in SAD than in Sham rats at 3 (4.63±0.27 vs 2.86±0.16 mmHg) and 10 days (5.68±0.56 vs 2.68±0.18 mmHg). No major changes in the respiratory parameters were observed in SAD at 3 days after the surgery, but the respiratory frequency (114±4 vs 136±5 cpm) and minute volume (1457±67 vs 1998±165 mL/Kg/min) were significantly reduced 10 days after SAD. The durations of expiration (0,34±0,01 vs 0,27±0,01 sec), inspiration (0,22±0,01 vs 0,19±0,01 sec) and respiratory cycle (0,56±0,02 vs 0,46±0,02 sec) were significantly increased 10 days after SAD. These data suggest that 10 days after SAD changes in the respiratory pattern may contribute to the modulation of the sympathetic activity and prevent major changes in MAP in SAD rats.

Grant Funding Source: Supported by FAPESP, CAPES and CNPq
Perivascular inflammation drives sympathetic hyperinnervation and hypertension in obesity (682.1)
Authors: Haddock Rebecca , Matthaei Klaus , Drummond Grant , Hill Caryl
Published: June 2021

Hypertension in obesity occurs as a result of abnormal increases in sympathetic nerve density over arteries controlling blood pressure; the causative factors underlying these changes are unknown. Here we examine the role of inflammatory cell infiltration to changes in perivascular nerve innervation over small mesenteric and renal arteries from C57BL/6 and Rag1‐/‐ mice fed a normal or high fat diet. Pressure myography, electrophysiology and videomicroscopy was used to measure intraluminal pressure, membrane potential, vessel diameter. Nerves were activated by electrical field stimulation. Immunohistochemistry was performed using antibodies against nerve growth factor (NGF), CD3+ (T cells) and F4/80 (macrophages). Sympathetic innervation was examined using anti‐synaptophysin (synaptic vesicles). Images were collected using confocal microscopy and analysed using Image J. Obese C57BL/6 mice were hypertensive. Nerve‐mediated contractions and excitatory junction potentials (EJP) were augmented (P<0.05) and the perivascular sympathetic nerve plexus was denser (P<0.05). Numbers of perivascular NGF‐producing immune cells (T cells and macrophages) were increased (P<0.05). Infiltration of these cells was shown to precede expansion of the nerve plexus (P<0.05). Rag1‐/‐ mice fed a HFD also became obese. However, numbers of perivascular NGF‐producing immune cells were not increased, arteries were not hyperinnervated and mice remained normotensive. Neurogenic hypertension in obesity occurs as a consequence of the infiltration of NGF positive immune cells to the vascular adventitia and expansion of the perivascular sympathetic nerve plexus.

Grant Funding Source: Supported by the NHMRC to REH (466009)
Humor similar to meditation enhances EEG power spectral density of gamma wave band activity (31‐40Hz) and synchrony (684.5)
Authors: Berk Lee , Alphonso Clarice , Thakker Neha , Nelson Bruce
Published: June 2021

Humor associated mirthful laughter (HAML) similar to meditation, both positive brain states, have complex mental processes that can modulate emotions, cognition, memory, sensory perception, consciousness, hormones, immune, and CNS activity. Meditation is used as a non‐pharmacological lifestyle intervention to provide health benefits, from promoting wellness to alleviating symptoms of a variety of medical conditions. Experienced mindfulness and long‐term meditators strive to achieve a brain state of increasing EEG gamma (γ) band activity (31‐40Hz), that is associated with these beneficial changes. We are not aware of any EEG study utilizing the positive brain state of HAML to identify similarities in Power Spectral Density µV2 (PSD) correlates across frequency bins1‐40Hz, but particularly 31‐40Hz γ band. Purpose: This study assessed EEG PSD in 31 university students during HAML and stress experiences for similarities in PSD across bins1‐40Hz. Methods: EEG activity was recorded from 9 cerebral cortical scalp regions F3, Fz, F4, C3, Cz, C4, P3, POz, and P4 with the EEG B‐Alert 10X System™. To reduce bias effects from a single video intervention a cross‐over design was used where each subject was randomly administered two 10 minute videos: Humor and Stress. Results: HAML increased all 9 channels in synchrony for β Band Activity (13‐30Hz) (βBA) (p<0.01) but most significantly for γBA (31‐40HZ), p<0.001. Total frontal region γBA and βBA were different from theta and αBA for HAML p< .05. Conclusions: HAML has correlates in EEG PSD change for specific bandwidths but most significantly and greatest in PSD & synchrony of γBA. HAML, associated with psychophysiological health benefits and happiness, appears to be similar to meditation in increasing PSD γBA & synchrony. We suggest HAML may be another non‐pharmacological lifestyle intervention to provide health, wellness & adjunctive therapeutic benefits. Further EEG research is being conducted to elaborate and expound on these findings.
Evidence for blood‐brain barrier Na‐K‐Cl cotransport, Na/H exchange and Na‐HCO3 cotransport involvement in hyperglycemia exacerbation of cerebral edema formation in ischemic stroke (685.3)
Authors: Chechneva Olga , Yuen Natalie , Tsai Yi‐Chen , Chen Yi‐Je , Anderson Steven , O'Donnell Martha
Published: June 2021

Hyperglycemia (HG) exacerbates cerebral edema formation in ischemic stroke and worsens clinical outcome through poorly understood mechanisms. Our previous studies provided evidence that the blood‐brain barrier (BBB) Na transporters Na‐K‐Cl cotransport (NKCC), Na/H exchange (NHE) and Na‐HCO3 cotransport (NBC) are stimulated by ischemic factors, causing increased secretion of Na and water into the brain and consequent cerebral edema formation. In the present study we examined the effects of HG on cerebral microvascular endothelial cell (CMEC) NKCC, NHE and NBC expression and activity, as well the effects of ischemic factors on the Na transporter activities following HG exposures using methods that included Western blot, qRT‐PCR, radioisotopic flux assays and microspectrofluorometry. We found that HG (30 vs 5 mM glucose; 6, 24, 48 hr and 7 d) increased abundance of bovine and human CMEC NKCC1, NHE1 and NBCn1. These exposures to HG also significantly increased CMEC NKCC, NHE and NBC activities. mRNA levels of NHE1 but not NKCC1 or NBCn1 were significantly increased by 6 and 24 hr of HG. Osmotic controls (with mannitol or NaCl) had no effect on abundance, activity or mRNA levels. HG also caused a greater increase in ischemic factor stimulation of these Na transporter activities. Finally, MRI studies showed that bumetanide and HOE‐642 inhibition of NKCC and NHE reduced HG‐worsened edema in rats with cerebral ischemia.

Grant Funding Source: Supported by NIH and ADA
Carbon monoxide excitatory effect in vasopressin neurons of heart failure rats involves nitric oxide/GABA signaling (686.24)
Authors: Reis Wagner , Biancardi Vinicia , Zhou Yiqiang , Stern Javier
Published: June 2021

We showed that the gas neurotransmitter carbon monoxide (CO) regulates vasopressin (VP) neuronal activity. Neurohumoral activation in the hypothalamic paraventricular nucleus (PVN), characterized by elevated VP secretion, occurs during congestive heart failure (HF). Moreover, blunted nitric oxide (NO)/GABA inhibitory signaling pathways within PVN are major contributing factors. In this study we determined whether PVN expression of heme‐oxygenase‐1 (HO‐1), a CO‐synthesizing enzyme, was changed in HF; whether CO effects on the firing rate (FR) of VP neurons was altered, and whether CO effects were mediated by NO/GABA function. Immunofluorescence and western blots studies showed an increased PVN HO‐1 protein levels during HF (P<0.05). Additionally, patch‐clamp recordings obtained from identified VP neurons in transgenic eGFP‐VP rats, showed that tin‐protoporphyrin IX (20µM, a HO inhibitor) diminished FR in 75% VP responsive cells in HF rats, compared with no effects in 62% VP cells in sham rats. Besides, tricarbonyldichlororuthenium dimer (30µM, a CO donor) increased FR in 71% and 75% VP responsive neurons in HF and sham rats, respectively. Interesting, the majority of VP neurons in HF rats were non responsive to the CO donor after previous application of a NO blocker (62%) or a GABA blocker (80%). In addition, the excitatory CO effect was accompanied by a reduction of GABA currents in 93% of VP neurons in HF rats. Taken together, our results showed overexpression of PVN HO‐1 and excitatory CO responses on VP neurons in HF rats, likely involving changes in NO/GABA‐mediated signaling.

Grant Funding Source: AHA 12POST12060540
Hydrogen sulfide induces miR194 in diabetic nephropathy and mitigates renal fibrosis (689.10)
Authors: Kundu Sourav , Narayanan Nithya , Pushpakumar Sathnur , Sen Utpal
Published: June 2021

Introduction: Recent evidences suggest that hydrogen sulfide (H2S) has a therapeutic potential for several clinical complications including renovascular fibrosis. Similarly, micro RNAs (miRs) has also gained substantial biomedical research interest with relevance to renal fibrosis. In this regard miRs control reactive oxygen species (ROS) and high level of ROS persuades poly‐ADP‐ribose‐polymerase‐1 (PARP1) which induces extracellular matrix (ECM) associated with vascular fibrosis. Among miRs, miR194 is prevalent in the mammalian kidney; however, its role in diabetic nephropathy and whether H2S has any potential implication on miRs regulation associated with diabetic renal fibrosis is unknown. Hypothesis: The objective of this study was to determine whether miR194 has potential roles in diabetic nephropathy which leads to PARP 1 activation by ROS induction causing renal fibrosis. Materials and methods: C57BL/6J (Wild type, WT) and diabetic (Akita, C57BL/6J‐Ins2Akita) mice were used in this study. Animals were treated without or with 0.05 g/L of NaHS in drinking water for 4 weeks. In vitro studies were performed using mouse glomerular endothelial cells (MGECs). Results: Expression of miR‐194 was significantly decreased in Akita animals compared to control as revealed by real time PCR. Increased expression of miR194 has been observed following H2S treatment. In diabetic animals increased ROS and PARP1 levels were measured. Levels of matricellular proteins as well as genes, responsible for collagen re‐alignment, showed higher expression in diabetes and significantly reduced following H2S treatment. We conclude that decrease in miR194 in diabetes contributes to increased expression of ECM components in renal fibrosis and treatment with H2S ameliorates these effects.

Grant Funding Source: HL‐104103
A dual COX‐sEH inhibitor improved glycemic status and reduced kidney injury in Zucker diabetic fatty rat (689.4)
Authors: Khan Abdul , Hwang Sung , Hammock Bruce , Imig John
Published: June 2021

Cyclooxygenase (COX) and soluble epoxide hydrolase (sEH) inhibitors have therapeutic potentials. We synthesized a novel dual COX‐sEH inhibitor, PTUPB and investigated its efficacy in type 2 diabetic (T2D) Zucker Diabetic Fatty (ZDF) rats. Male ZDF rats were treated with vehicle or PTUPB (3 mg/d, i.p.) for 56 days. On day 56, ZDF rats were diabetic (fasting blood glucose, 287±45 mg/dL) compared to Zucker Diabetic Lean rats (ZDL, 99±6 mg/dL), and PTUPB treatment improved glycemic status in ZDF rats (146±6 mg/dL). ZDF rats also developed hypertension and endothelial dysfunction that were reduced by PTUPB. The ZDF rats demonstrated kidney injury compared to ZDL with elevated albuminurea (4±2 vs 44±4 mg/d), nephriurea (16±4 vs 496±127 µg/d), marked renal fibrosis, cast formation and glomerular injury compared to ZDL rats, and PTUPB treatment reduced this injury by 40‐50%. The ZDF rats also demonstrated increased inflammation and oxidative stress with elevated levels of urinary monocyte chemoattractant protein‐1 excretion (319±75 vs 862±300 ng/d), renal macrophage infiltration (37±4 vs 53±2/mm2) and kidney malondialdehyde/protein ratio (5±1 vs 10±1 µmol/mg) that were decreased with PTUPB treatment by 20‐50%. The data obtained demonstrate the ability of a novel dual COX‐sEH inhibitor to improve glycemic status and reduce kidney injury in T2D.
The role of complement C3 in focal inflammation and development of kidney cysts induced by Pkd1 inactivation (690.3)
Authors: Wu Yong , Xu Jen , Li Suyang , Hsieh Tsung‐Hsiu , Lu Tzongshi , Kong Tianqing
Published: June 2021

Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common life‐threatening genetic diseases and is characterized by early formation and gradual enlargement of multiple kidney cysts, which eventually results in end‐stage renal disease (ESRD). Mutations in PKD1 (polycystin‐1, PC1) or PKD2 (polycystin‐2, PC2) account for 85% and 15% of this disease respectively. PC1 is a large integral membrane protein with multiple functions. However, the mechanism for development of multiple kidney cysts induced by PKD1 mutation is not fully understood. In order to find important proteins in renal cystic fluid from ADPKD mice induced by Pkd1 inactivation, we did mass spectral analysis. Our results showed that complement C3 and its cleaved products are the most abundant protein in the cystic fluid, which was further confirmed by Western blots in kidney tissue from ADPKD mice and ADPKD patients. Furthermore, we found C3 and its cleaved fragments are significant increased around kidney cysts epithelium line by immunostaining, and its distribution is consistant with focal fibrosis. As reported previously, the expansion of kidney cysts is a slow inflammation and fibrosis process. Our finding indicates that C3 and its cleaved products may play critical role for this process. Inhibition of focal C3 and its cleavage could be a unique regimen for ADPKD treatment.
Novel epoxyeicosatrienoic acid analog ameliorates renal injury in a rat model of radiation nephropathy (690.7)
Authors: Wahl Geneva , Khan Abdul , Fish Brian , Cohen Eric , Falck John , Imig John
Published: June 2021

Arachidonic acid metabolites epoxyeicosatrienoic acids (EETs) protect the kidney in different pathologies. This study investigated the ability of a novel EET analog, EET‐A to mitigate experimental radiation nephropathy caused by total body irradiation (TBI, 11 Gy). Four groups of rats, normal male WAG/RijCmcr rats, TBI rats treated with vehicle (TBI‐V), EET‐A (10mg/kg/d p.o.) treated TBI rats (TBI‐EET‐A), and Captopril (30mg/kg/d p.o.) treated TBI rats (TBI‐Cap) were studied. Kidney damage was assessed from blood urea nitrogen (BUN), albuminuria, nephrinuria, and histopathological changes 12 weeks post TBI. The TBI‐V group had a 3‐fold increase in BUN level compared to control (158±14 vs 50±3 mg/dL). BUN level was lower in TBI‐EET‐A (85±6 mg/dL) and TBI‐Cap (52±2 mg/dL) compared to TBI‐V group. Albumin/creatinine ratio increased 94‐fold in TBI‐V group compared to control (6451±800 vs 20±5 μg/mg), and was reduced in TBI‐EET‐A (1652±382 μg/mg) and TBI‐Cap (103±28 μg/mg) groups. Nephrinuria increased 30‐fold in the TBI‐V compared to control group (15390±1897 vs 48±9 μg/d), and EET‐A (7071±2122) and captopril (222±107) decreased it. Moreover, renal tubular and glomerular injury was reduced in TBI‐EET‐A and TBI‐Cap groups compared to TBI‐V group. These data demonstrate that a novel EET‐analog, EET‐A reduces kidney injury in experimental radiation nephropathy caused by TBI.
Circadian variations in renal hemodynamics in healthy volunteers ő preliminary findings (692.8)
Authors: Eckerbom Per , Cox Eleanor , Weis Jan , Bjerner Tomas , Hansell Peter , Subasic Irfan , Palm Fredrik , Francis Susan , Liss Per
Published: June 2021

Renal blood flow is tightly controlled by at least two different intrinsic mechanisms, which should guarantee a stable blood flow. However, it is currently unknown if renal hemodynamics is influenced by circadian rhythm like many other tissues. Therefore, we measured total and cortical renal blood flow using completely non‐invasive magnetic resonance technique in six healthy male volunteers every fourth hour for a total of 24 hours (six measurements in total). Total renal blood flow was measured using standard phase contrast technique in renal arteries, whereas cortical renal blood flow was measured using the newly developed arterial spin‐labeling technique.

Total renal blood flow averaged 13.3±1.4 and 13.0±1.0 ml/min for right and left kidney, respectively, with a circadian variation of 18±5 and 14±3%. Cortical renal blood flow in the right kidney averaged 323±17 ml/min/100g with a variation of 10±2% between the different measurements. None of these variations reached statistical significance.

In conclusion, neither total nor cortical renal blood flows display circadian variations. Thus, circadian variations in renal hemodynamics seem less likely to affect the results from studies investigating renal blood flow and related parameters.
The effect of inosine and allopurinol on xanthine oxidoreductase gene expression and mitochondrial function in liver and kidney tissues of broiler chickens (693.16)
Authors: Settle Tabatha , Radke William , Falkenstein Elizabeth , Klandorf Hillar
Published: June 2021

Uric acid is generated in the xanthine/hypoxanthine reactions catalyzed by xanthine oxiodreductase (XOR) as the end‐product of purine degradation. The purpose of the first study was to determine the effects of allopurinol and inosine on XOR activity and gene expression in liver and kidney tissue. In the first study, Cobb x Cobb (n=15; 5weeks old) were separated into three treatments (n=5); control (CON), INO (inosine at .6M/kg feed), or INOAL (inosine and allopurinol 50mg/kgBW). Liver uric acid concentrations were reduced (p< 0.05). There was a significant (p 蠄 0.05) increase in XOR gene expression in the liver tissue of INOAL birds compared to CON and INO groups. There was no difference in XOR expression between treatments in kidney tissue. The purpose of the second study was to assess mitochondrial function. Cobb x Cobb broilers (n=12; 4 weeks old) were separated into two treatments (n=6); control (CON) and AL (allopurinol 35mg/kg BW). Mitochondria were freshly isolated from liver tissue and assessed for State III and State IV respiration using polarography. There was a reduction in State III respiration (p=0.01) and State IV respiration (p=0.007) in allopurinol‐treated birds compared to the control. Lowered uric acid in the liver can increase oxidative stress within this tissue subsequently resulting in mitochondrial dysfunction and an upregulation of XOR gene expression.
High glucose treatment enhances autotaxin and VEGF‐C expression in PC‐3 human prostate cancer cell (693.20)
Authors: Lin Chu‐Cheng , Lee Hsinyu
Published: June 2021

Clinical evidences suggest that lymphangiogenesis and lymphatic metastasis are important processes during the progression of prostate cancer. Vascular endothelial growth factor (VEGF)‐C was shown to be a key regulator in these processes. Our previous studies demonstrated that lysophosphatidic acid (LPA), a low‐molecular‐weight lipid growth factor, enhances VEGF‐C expression in three different human prostate cancer cell lines. In PC‐3 human prostate cancer cells, the enhancement effects of LPA were mediated through LPA1/3‐, PLC‐, PKC‐, Nox‐, ROS‐, and LEDGF‐dependent pathways. Furthermore, autotaxin (ATX), an enzyme responsible for LPA synthesis, also participates in regulating VEGF‐C expression. Herein, we further demonstrated that the mRNA levels of ATX and VEGF‐C were increased in PC‐3 cell by 0.02 M of high glucose treatments within 2 hours. These processes can be suppressed by pretreatment with ROS scavenger, N‐acetylcysteine. These results suggested that the unregulated glucose metabolic conditions might lead to LPA synthesis and therefore the subsequent pathological conditions of prostate cancer. These novel findings could potentially provide new strategies for prostate cancer treatments.
PMN recruitment to small airway epithelial cells in response to cigarette smoke extract (694.8)
Authors: Marentette John , Rastogi Prerna , McHowat Jane
Published: June 2021

Cigarette smoke causes airway inflammation and recruitment of inflammatory cells, requiring migration across the endothelial and epithelial cell barriers. We propose that increased platelet‐activating factor (PAF) production in the airways of smokers enhances inflammatory cell recruitment. Human small airway epithelial cells (HSAEC) incubated with cigarette smoke extract (CSE) demonstrated increased PAF accumulation that was inhibited when cells were pretreated with (S)‐bromoenol lactone ((S)‐BEL, inhibits calcium‐independent phospholipase A2β (iPLA2β). HSAEC incubated with CSE also showed a significant inhibition of PAF acetyl hydrolase (PAF‐AH, degrades PAF). This suggests that increased PAF accumulation is a combination of increased synthesis (iPLA2β activation) and decreased degradation (PAF‐AH inhibition). 舲舲Polymorphonuclear leukocyte (PMN) adherence to HSAEC was increased with CSE and inhibited when HSAEC were pretreated with (S)‐BEL or PMN were pretreated with Ginkgolide B to block the PAF receptor. CSE also increased PMN transmigration across HSAEC in a basolateral to apical direction.舲舲 Taken together, our data demonstrate that PMN accumulation in the airways of smokers depends on increased PAF accumulation in HSAEC and suggest that Ginkgo biloba is a potential therapy to manage airway inflammation.
Effect of orthostasis on endothelial function (695.7)
Authors: Anyaehie Ugochukwu , Goswami Nandu , Hinghofer‐Szalkay Helmut
Published: June 2021

Vascular endothelium has multiple functions and may play a role in the pathophysiology of orthostatic intolerance. We investigated the effect of orthostasis on endothelial function using EndoPAT, a non‐invasive and user‐independent method, and across gender. As sex steroid hormones are known to affect endothelial function, this study examined the potential effect of these hormones on the endothelial response to orthostasis by including females at different phases of the menstrual cycle (follicular and luteal) and females taking an oral contraceptive. A total of 31 subjects took part in this study (11 males, 11 females having normal menstrual cycles and 9 females taking oral contraceptive). Each subject made two visits for testing; in the case of females having normal menstrual cycles the first session was conducted either 1‐7 (follicular) or 14‐21 days (luteal) after the start of menstruation, and the second session two weeks later, i.e., during the other phase, respectively. Endothelial function was assessed at baseline and following a 20‐min orthostatic challenge (active standing). The EndoPAT index increased from 1.71 ± 0.09 (mean ± SEM) at baseline to 2.07 ± 0.09 following orthostasis in females (p<0.001). In males, the index increased from 1.60 ± 0.08 to 1.94 ± 0.13 following orthostasis (p<0.001). There were no significant differences, however, in the endothelial response to orthostasis between females and males, menstrual cycle phases and the usage of oral contraceptive. Results suggest that orthostasis can induce increase in vasodilatatory endothelial response in both females and males. Further studies are needed to determine the potential role of this post orthostasis endothelial response in the pathophysiology of orthostatic intolerance.
Reciprocal relationship between reactive oxygen species and cyclooxygenase‐2 after lead exposure on vascular smooth muscle cells (700.6)
Authors: Simões Maylla , Aguado Andrea , Zhenyukh Olha , Alonso Maria , Briones Ana , Fiorim Jonaína , Azevedo Bruna , Vassallo Dalton , Salaices Mercedes
Published: June 2021

Studies reported that chronic lead (Pb) administration affects cardiovascular parameters. We evaluate the effects of chronic exposure to lead in vascular smooth muscle cells (VCMS) on oxidative stress and cyclooxygenase‐2 (COX‐2) pathways. Aortas were isolated from male Sprague‐Dawley rats and processed to obtain primary cultures of VCMS. The cells were stimulated with Pb (200 ng/ml) for 48 h. Pb increased the gene expression of COX‐2 and subunits of NOX‐1 and NOX‐4 of NADPH Oxidase and the protein expression of Mn‐SOD and COX‐2. Celecoxib (10μmol/L) reversed the upregulation of NOX‐1 and NOX‐4 whereas tempol (10μmol/L) reversed the upregulation of COX‐2. NADPH oxidase activity and superoxide anion production was also enhanced by Pb and normalized by Celecoxib and Rofecoxib (10μmol/L), ML171 (10μmol/L) and/or Tempol, Mito‐TEMPO (5μmol/L). Pb also induces the activation of ERK1/2 and p38MAPK signaling pathways that were involved in the augmented expression of COX‐2, NOX‐1 and NOX‐4. Pb exposure did not induce proliferation or migration of VCMS. Our results suggest a relationship between lead, COX‐2 and oxidative stress, probably involving mechanisms with MAPK signaling pathways. Since this may be involved in hypertension development, lead exposure should be considered an environmental risk factor for cardiovascular disease.

Grant Funding Source: MINECO (SAF 2012‐36400) ISCIII (RD/0042/0024), FMM, CAPES, PRONEX‐FAPES/CNPq
Effect of oral administration of Moringa aloevera leaf extract on neutrophil phagocytic activity in petrol exposed Wister rats (701.8)
Authors: Azeez Oyebisi , Anigbogu Chikodi , Adedeji Adebayo , Saka Waidi , Akhigbe Roland
Published: June 2021

Reduced immunity has been implicated in rats following inhalation of petrol. Angiotensin II AT1 receptor blockade reduces inflammation in hypertension. The aim of this work is to assess the phagocytic effect of neutrophil in rats exposed to petrol and moringa alovera leave concurrently; petrol and candesartan and petrol and captopril.

Methodology: 25 adult male rats were grouped to 4 with 4rats per group. All groups were exposed to petrol for 5 minutes everyday for 2 weeks except control group. Group 1 (control) Group 2, 3 and 4 had moringa aloevera plus feed, candesartan pre‐treatment, captopril pretreatment respectively while group 5 had petrol exposure only. At the end of the exposure, rats were anaesthesized with chloroform and blood collected into EDTA bottles from the heart. In vitro phagocytosis assay was conducted using 0.1ml of autologus chemotactically inactive serum. Phagocytosis ability (PHC) was considered as the percentage of phagocyting neutrophils by microscopic field at 405nm per charge.

Results: There was significant increase in the phagocytotic activity of the moringa aloevera (+7.2%) group compared with control (2.15%). A reduction that was not significant was seen in the captopril group.

Conclusion: Moringa alovera ameliorates the inflammation induced by exposure to inhalation of petrol by increasing phagocytotic activity of Neutrophils. The effect in the captopril [(Angiotensin converting enzyme inhibitor (ACE inhibitor)] group was higher compared with the Candesartan an AT1 receptor blocker. The moringa aloevera effect seem to have been carried out through the ACE inhibitor.
Dipeptidyl peptidase IV inhibition ameliorates cardiorenal function in experimental heart failure (701.9)
Authors: Arruda‐Junior Daniel , Socas Leonardo , Dariolli Rafael , Antonio Ednei , Tucci Paulo , dos Santos Leonardo , Girardi Adriana
Published: June 2021

We have recently reported that long‐term dypeptidyl peptidase IV (DPPIV) inhibition mitigates the development and/or progression of HF in rats subjected to myocardial injury. The present study aimed to investigate whether the DPPIV inhibitor vildagliptin is capable of improving cardiorenal function in rats with established heart failure. Experimental HF was induced in male Wistar rats via left ventricular (LV) myocardial injury using radiofrequency catheter ablation. Sham‐operated rats underwent left thoracotomy and were mock‐ablated. Echocardiographic analysis were performed six (Phase I) and ten (Phase II) weeks after surgery. After the Phase I analysis, HF rats were randomly divided into four groups and treated with vildagliptin (VIL) at daily doses of 20, 80 or 120 mg/kg of body weight or vehicle. Compared to Phase I, Phase II LV ejection fraction reduced in HF groups treated with water or VIL20, remained unaltered in HF rats treated with VIL80 and increased in rats treated with the higher dose of vildagliptin, VIL120. All vitagliptin‐treated rats exhibited lower lung water content and heart weight to body weight ratio as compared to HF rats treated with water. Treatment with VIL80 and VIL120 remarkably reduced BNP circulating levels as compared to vehicle treated HF rats (167±40 and 152±42 vs. 372±75 pg/mL, respectively). HF rats displayed sodium retention and lower glomerular filtration rate compared to Sham. Treatment with VIL120 was capable of restoring renal function in HF rats. Taken together, these results suggest that DPPIV inhibition improves cardiorenal function in rats with established HF. Thus, DPPIV inhibitors may be effective not only for the prevention but also for the treatment of HF in rats.

Grant Funding Source: Supported by FAPESP.
Differential regulation of myofibrillar and mitochondrial protein synthesis following acute endurance exercise (702.3)
Authors: Philp Andrew , Schenk Simon , Breen Leigh , Hamilton Lee , Scott Fraser , Laverone Erin , Phillips Stu , Baar Keith
Published: June 2021

Endurance exercise leads to rapid transcriptional‐ and translational‐mediated remodeling in skeletal muscle. The purpose of this study was to investigate the role of the mechanistic target of rapamycin (mTORC1) in the molecular regulation of myofibrillar and mitochondrial protein synthesis in the first 6h after endurance exercise. Female C57BL/6 mice performed one‐hour of treadmill running after administration of rapamycin or vehicle. A flooding dose of L‐[ring‐13C6]phenylalanine was administered via IP injection and blood and gastrocnemius muscle collected 0.5, 3 and 6h post exercise and in a separate, non‐exercising control group (n=4/time). Skeletal muscle was analyzed for phenylalanine incorporation into the myofibrillar and mitochondrial pools via GC‐C‐IRMS. Myofibrillar fractional synthesis rate (FSR) increased rapidly in the vehicle group post‐exercise and remained elevated at all post‐exercise times. Rapamycin had no effect on basal FSR, but attenuated the rise in myofibrillar protein FSR at 0.5 and 3h post exercise. Interestingly, rapamycin did not affect mitochondrial protein FSR, which was increased at 3h and 6h post exercise in both groups. Taken together our data suggest that the initial increase (0‐3h) in myofibrillar FSR after endurance exercise is mTORC1 mediated, whereas at latter time points (3‐6h) myofibrillar and mitochondrial FSR are both stimulated, independent of mTORC1.
Use of microarray for the evaluation of long‐haul shipping stress in beef calves (703.2)
Authors: Onaga Jonathan , Sy Tracy , Richards Tim , Tiirikainen Maarit , Turano Helen , Lee C. , Stokes Ashley
Published: June 2021

Objectives of this study were to evaluate the impact of long‐haul shipping on gene expression of leukocytes from weaned calves using a bovine‐specific microarray before and after transport. Two shipments of weaned, Angus and Charolaise, female calves (n=48) were transported to Washington or California. Each shipment, the calves were divided and placed into two separate, bi‐level, four‐compartment, custom cowtainers. Blood was collected prior to shipping (P), upon arrival to the receiving yard (A), and 6‐days post‐arrival (PA). Leukocytes were isolated, total RNA extracted, quantified, and analyzed for quality. An Affymetrix 26,773 WT bovine‐specific array was utilized using manufacturer’s protocols. Using a conservative fold‐change of 2 and a p‐value of less than or equal to 0.05 as significant, only 2 genes were significantly different across the study. FOS was decreased 3.05‐fold P to A, but then increased back 2.92‐fold A to PA. P to A, haptoglobin decreased 2.27‐fold with a p‐value of 0.047. Considering the sensitivity and specificity of the microarray, these changes are very minor physiologically and reflect minimal impact of shipping on these study animals. The findings of this study support our previous work that the currently utilized shipping protocol to transport calves from Hawaii to the Mainland US does not cause large changes in gene expression due to shipping stress.

Grant Funding Source: Supported by National Cattlemen's Beef Association
Impact of stress on mast cell and vitamin D receptor expression in an animal model of endometriosis (703.4)
Authors: Seguinot Inevy , Rivera Yadmarie , Cruz Suheil , Cruz Myrella , Hernandez Siomara , Hernandez Anixa , Isidro Angel , Flores Idhaliz , Appleyard Caroline
Published: June 2021

Endometriosis is characterized by peritoneal inflammation, infertility and chronic pelvic pain that substantially affect quality of life. Since stress can affect the immune system and recruitment of inflammatory cells it may contribute to this condition. Vitamin D receptor (VDR) is associated with anti‐proliferative and anti‐inflammatory properties and is expressed in the cycling endometrium. AIM: Investigate the impact of stress on mast cells and VDR in an animal model of endometriosis. METHODS: Endometriosis was induced in female Sprague‐Dawley rats by suturing uterine horn tissue next to the intestinal mesentery. Sham rats had sutures only. Rats were exposed to either an uncontrollable or controllable stress protocol for 10 days (endo‐stress) or no stress (endo‐no stress). At sacrifice (day 60) all rats were examined for cysts. Colon and uterine tissue were analyzed for damage and myeloperoxidase (MPO) levels. Mast cell infiltration was measured by staining with toluidine blue or immunofluorescence, and VDR expression by immunohistochemistry. RESULTS: Endo‐stress rats developed more and bigger cysts than endo‐no stress rats. Uncontrollable stress resulted in higher colonic damage (p<0.05) and uterine MPO compared to no stress, while controllable stress showed no difference. Uncontrollable stress increased mast cell infiltration in colonic tissue compared to controllable stress (p<0.05), endo‐no stress (p<0.05) and sham‐no stress (p<0.01). There was increased mast cell infiltration in cysts. Endo‐no stress group had higher expression of VDR in uterus stroma nuclei and glands (p<0.05 vs controllable stress; p<0.05 vs sham, in glands only). CONCLUSION: Stress exacerbates development of cysts in an animal model of endometriosis through mechanisms involving mast cell recruitment, release of cell mediators and VDR down‐regulation.

Grant Funding Source: Supported by R15AT006373 & R25GM082406
Spinal μ‐opioid receptor sensitive locomotor muscle afferents decrease responsiveness of motor cortical cells and diminish cortical voluntary activation (705.1)
Authors: Sidhu Simranjit , Weavil Joshua , Mangum Tyler , Morgan David , Jessop Jacob , Richardson Russell , Amann Markus
Published: June 2021

We investigated the influence of group III/IV locomotor muscle afferents on the development of supraspinal fatigue and changes in motor cortical cell responsiveness by comparison of cortical versus spinal excitability during sustained cycling exercise. Six males performed constant‐load cycling exercise (80% peak power‐output) to exhaustion under control conditions (CTRL) and with lumbar intrathecal fentanyl (FENT) impairing feedback from µ‐opioid receptor‐sensitive lower limb muscle afferents. Pre‐to‐post exercise changes in voluntary activation (VA) and potentiated quadriceps twitch force (∆Qtw) were assessed via transcranial magnetic (TMS) and electrical femoral nerve stimulation (FNS). During cycling, vastus lateralis responses evoked by TMS (MEP) and electrical cervicomedullary stimulation (CMEP) were normalized to FNS‐evoked M‐waves. Although maximum voluntary quadriceps torque was similarly reduced following both trials (~14%; P<0.05), ∆Qtw was greater in FENT vs. CTRL (51 ± 5% vs. 62 ± 7%; P<0.05). Furthermore, VA was reduced by 6% in CTRL (P<0.05) but remained unchanged in FENT. MEP increased throughout the exercise in FENT (14 ± 5%; P<0.05) but remained unchanged in CTRL. CMEP did not change in either session. Our findings suggest that group III/IV muscle afferents compromise motor cortical cell responsiveness during locomotor exercise and facilitate supraspinal fatigue.

Grant Funding Source: HL_103786; HL‐116579
Characterizing rapid onset vasodilation to single muscle contractions in the leg in humans (705.10)
Authors: Credeur Daniel , Holwerda Seth , King Phillip , Crutcher Kiera , Padilla Jaume , Fadel Paul
Published: June 2021

Rapid onset vasodilation (ROV) following single muscle contractions has been examined in the forearm of humans, but has not been characterized in the leg. Given known vascular differences between arms and legs as well as the greater susceptibility of legs to atherosclerosis, we sought to characterize ROV in the leg. To do this, 9 healthy men performed random ordered single muscle contractions at 5, 10, 20, 40, and 60% of their maximum voluntary contraction (MVC) in the leg (knee extension) and forearm (handgrip). Continuous blood flow (BF) measures were made via duplex Doppler‐ultrasound from the brachial and femoral artery during 15 sec prior to and 45 sec after release of each contraction. Three trials were performed at each intensity. Limb BF was normalized to lean mass of forearm and thigh region through DEXA. Single muscle contractions produced an intensity dependent increase in BF, peaking ~4 sec post contraction in both limbs. Initial BF responses (1st heart beat) were significantly greater in the leg, most notable at lower MVC intensities (e.g. 10%‐MVC initial response; Arm=+19±5% vs. Leg=+75±15%, P=0.01). However, peak changes in BF were similar between limbs for each contraction intensity (e.g. 60%‐MVC peak; Arm=+207±33% vs. Leg=+208±33%, P>0.05). These preliminary findings indicate that peak BF responses to single muscle contractions are similar between limbs establishing the presence of ROV in the leg.

Grant Funding Source: NIH grant RO1 HL093167 (PJF)