bluuebunny/crossref_metadata_2025 huggingface dataset:
Mongolian low birth weight young adults have higher risk of hypertension compared with Han Chinese and Japanese counterparts
Authors: Bao Sarina , Sato Haruka , Sasaki Konosuke , Kanno Emi , Maruyama Ryoko
Published: June 2021

Low birth weight (LBW) was confirmed as a risk of high blood pressure (BP) in later stages of life (Barker DJ et al, 1989). Low‐grade inflammation and deterioration of autonomic regulation play an important role in hypertension. However, the associations with birth weight are poorly understood. We examined these relationships among Mongolian, Han Chinese and Japanese young adults, and investigated whether ethnicity affected these relationships. We measured BP and heart rate variability at rest and during postural change from a supine to a sitting position in 16 Mongolian, 4 Han Chinese and 21 Japanese healthy volunteers aged 18–34 years. Blood cell counts, and total cholesterol, high‐density lipoprotein cholesterol (HDL‐C), low‐density lipoprotein cholesterol, triglyceride, and high sensitivity C‐reactive protein levels were measured. Mongolians had lower levels of HDL‐C than did the Japanese (p < 0.01) and the Han Chinese. In Mongolians, the platelet count was higher in the LBW group than in the normal birth weight (NBW) group (p < 0.05). Following postural change, systolic blood pressure and heart rate showed no significant increases in the Mongolian and Japanese LBW groups, whereas the NBW groups had normal responses (p < 0.05). The Mongolian LBW group displayed a slight decrease in sympathetic nerve activity from a supine to a sitting position, although it increased in the Japanese LBW group (p < 0.05). These relationships were not able to examine in Han Chinese because of the small sample size. We suggested that Mongolian low birth weight young adults have higher risk of hypertension compared with Han Chinese and Japanese counterparts.
Prolonged Walking Attenuates Hypersympathetic State In Swine With Bilateral Femoral Artery Occlusion
Authors: Yang Hsiaotung , Bowles Douglas , Laughlin M , Terjung Ronald
Published: June 2021

An increased sympathoexcitation state (SES) has been found in patients with peripheral

arterial disease (PAD). Symptoms include an exaggerated response of blood pressure (BP) during increasing physical

activity. This high SES is detrimental to PAD patients. To determine whether physical training could alleviate SES, 11 Yucatan male pigs received bilateral femoral artery occlusion (BiFemOccl), and were either limited to cage activity (Sed, n=5) or walked

about 30 min/d (3 mph) on treadmill for 12 wks (Tr, n=6). Arterial baroreflex function was assessed by intravenously infusing adenosine to trig sympathetic drive (1.5 mg /kg) or phenyleprine (PE, 0.1 mg/kg) for parasympathetic drive. BP and heart rate (HR) were recorded through a catheter in the thoracic aorta while animals stood on treadmill. Net change of HR was calculated to change of BP in each animal. Even though resting BP (125 ±5 mm Hg) and HR (110±5 bpm) showed no differences between Sed and Tr group at either prior or post training, the slopes of HR response to reduce BP by adenosine were ‐1.08±0.22 and ‐0.57±0.03 for Sed and Tr group respectively (p<0.05). The HR responses to increased BP by PE were similar between Sed and Tr groups (Slope ‐0.71 ±0.04). The results indicate that swine with BiFemOccl shows exaggerated HR response to baroreceptor unloading and the high SES can be tempered by exercise training. Thus, a chronic exercise program could provide benefit to patients with PDA by reducing the risk of cardiovascular accidents due to the high SES.

Supported by NIH grant P01 HL 52490.
Effect of Carotid Body Chemoreceptor Inhibition on Cardiac Baroreflex Sensitivity in Resting Humans
Authors: Chantigian Daniel , Johnson Maja , Limberg Jacqueline , Joyner Michael , Johnson Blair
Published: June 2021

Activation or inhibition of the carotid body chemoreceptors (CBCs) may impact cardiac baroreflex sensitivity (cBRS). The purpose of this study was to test the hypothesis that inhibition of the CBCs will elicit a change in cBRS when compared to normal resting conditions. Young, healthy subjects (n=16, 31±2, 6 women) were given IV saline and a range of IV dopamine (DA) (1‐4 µg/kg/min) for 15 minutes prior to and throughout five hypoxic ventilatory response (HVR) tests. Subjects were initially infused with IV saline. After each HVR, the DA infusion rate was increased by 1 µg/kg/min. A 3‐lead EKG and beat‐by‐beat blood pressure (NexFin) were used to determine cBRS. Data were analyzed from the 5 minutes prior to the HVR associated with the DA dose that elicited the greatest attenuation of the HVR when compared to saline. Heart rate was not significantly different during saline vs. DA (62±3 vs. 65±3, p=0.25). Diastolic blood pressure (77±2 vs. 72±1 mmHg), mean arterial pressure (98±2 vs. 92±2 mmHg), and cBRS “up” sequence (38±7 vs. 21±3 ms/mmHg) were significantly lower during saline vs. DA (p<0.05 for all). cBRS values from the RRI spectrum were not statistically distinguishable between saline and DA (26±4 vs. 21.1±3, p=0.06). In summary, inhibiting the CBCs via DA infusion in young, healthy subjects results in a fall in blood pressure and a reduction in cBRS in response to a rise in blood pressure. These data suggest the CBCs contribute to cBRS and play an important role in modulating parasympathetic control of blood pressure. Funding: NIH R01DK090541
Methodological Considerations for Assessing Measures of Spontaneous Cardiac Baroreflex Sensitivity in Humans
Authors: Kent Ivan , Holwerda Seth , Fadel Paul
Published: June 2021

The arterial baroreflex is a negative feedback control system that plays an important role in short‐term maintenance of arterial blood pressure (BP). Analyses assessing spontaneous beat‐to‐beat fluctuations in BP and heart rate (HR) are increasingly being used to estimate cardiac baroreflex sensitivity. Published results have commonly used 10 min recording durations for these analyses; however, some research protocols require shorter recording durations. Since the validity of using shorter compared to longer recording durations has not been established, we tested the hypothesis that reduced baseline durations diminish the reliability of spontaneous cardiac baroreflex sensitivity measures. Beat‐to‐beat HR and BP were measured continuously during supine rest in 10 healthy individuals for 10 min, and spontaneous cardiac baroreflex sensitivity was estimated using the sequence technique and low frequency transfer function analysis (0.04–0.15 Hz). The 10 min baseline for each subject was divided into two 5‐min segments and two 3‐min segments, and each analysis was repeated. Statistical analysis revealed similar spontaneous baroreflex sensitivity among the various baseline durations for both analyses (P>0.05). For example, spontaneous cardiac baroreflex sensitivity for the sequence technique was 18±3 ms/mmHg for 10 min, 18±2 ms/mmHg for 5 min and 18±3 ms/mmHg for 3 min durations (P>0.05). However, the reliability over the various recording durations as assessed by intra‐class correlation was acceptable only for the sequence technique. These preliminary findings suggest that shorter baseline durations appear adequate to assess spontaneous cardiac baroreflex sensitivity when using the sequence technique.
Spectral Analysis of Barosensitive Neurons in the Rat Rostral Ventrolateral Medulla
Authors: Huereca D , Barman S , Mueller P
Published: June 2021

Regulation of blood pressure occurs primarily via the activity of bulbospinal rostral ventrolateral medulla (RVLM) neurons that drive sympathetic nerve activity (SNA). Central and peripheral stimuli influence the activity of RVLM neurons and are reflected in their cardiac and respiratory modulation. Dysregulation of RVLM neurons can lead to increased SNA and the development and maintenance of cardiovascular disease. In this study we used spectral (frequency‐domain) analysis to identify cardiac‐ and/or respiratory‐related (CR, RR) activity in barosensitive RVLM neurons (n=22) and splanchnic SNA of 16 Inactin‐anesthetized rats in which we also recorded arterial pressure (AP) and an index of respiration. SNA had RR activity (0.40±0.05; mean ± SE) and/or CR activity (0.63±0.05) as evidenced by a coherence value > 0.1 at the frequency of respiration and the heart rate, respectively, in the corresponding coherence functions. RVLM unit activity was also shown to be RR (0.40±0.14; n=4) or CR (0.41±0.07; n=4). Moreover, RVLM unit activity cohered to SNA at the frequency of the heart rate (0.39±0.08; n=4) or respiration (0.21; n=1). AP‐triggered analysis (time‐domain analysis) revealed CR activity in 4 other RVLM neurons. These data demonstrate our ability to study the relationships among rat RVLM unit activity, SNA, AP, and respiration, which will allow us to explore whether differences in the strengths of these relationships contribute to changes in SNA and AP under different physiological conditions.

HL096787
Changes in Directly Recorded Sympathetic Nerve Activity and Noradrenaline Spillover to the Heart and Kidney during Heart Failure
Authors: Ramchandra Rohit , Hood Sally , Xing Tao , Lambert Gavin , May Clive
Published: June 2021

Heart failure (HF) is associated with increased cardiac and renal noradrenaline (NA) spillover, both of which are independent predictors of mortality. We hypothesized that in HF changes in the mechanisms controlling NA release and uptake would lead to differences between directly recorded sympathetic nerve activity (SNA) and NA spillover to the heart and kidney. Experiments were conducted in conscious sheep implanted with cardiac SNA (CSNA) and renal SNA (RSNA) recording electrodes. Together with recordings of SNA and arterial pressure, the spillover of NA from both the heart and the kidney was determined. HF was associated with a two‐fold increase in CSNA (42±8 vs. 95±11 bursts/100hb, p<0.05) but a five‐fold increase in cardiac NA spillover (3.1±0.8 vs. 18.1±4.1 pmol/min, p<0.05). Infusion of sodium nitroprusside in the normal animals significantly increased CSNA to levels similar to those in HF, but the level of cardiac NA spillover (7.1±1.6) reached only 50% of that in HF. Further increases in CSNA in the HF animals were not associated with any further increases in cardiac NA spillover. In HF, RSNA was unchanged (95±8 bursts/100hb), but interestingly spillover of NA from the kidney was significantly increased (10.1±2.4 vs. 52.5±10.2 pmol/min, p<0.05). The mismatch between CSNA and cardiac NA spillover in HF suggests that other mechanisms such as the NA transporter may play an important role in mediating the increased cardiac NA spillover. Further experiments are required to determine what causes the discordance between the baseline levels of RSNA and renal NA spillover in HF.
Insulin Resistance Impairs Arterial Stiffness and Peripheral Vascular Resistance in Metabolic Syndrome Patients
Authors: Rodrigues Sara , Cepeda Felipe , Dutra‐Marques Akothirene , Carvalho Jefferson , Toschi‐Dias Edgar , Maki‐Nunes Cristiane , Bortolotto Luiz , Costa‐Hong Valeria , Rondon Maria , Alves Maria , Negrao Carlos , Tombetta Ivani
Published: June 2021

Besides autonomic alterations, metabolic syndrome (MetS) causes changes in the vascular system directly related to cardiovascular events and death. Since insulin resistance is strongly associated with sympathetic hiperactivation, we tested the hypothesis that the presence of impaired fasting glucose (IFG) is the main cause of structural and functional worsening on large and small vessels. Never treated, newly diagnosed MetS (ATP‐III) patients were divided into: fasting glucose >100mg/dL (MetS+IFG, n=28, 49.1±1.3 yrs) and fasting glucose <100mg/dL (MetS‐IFG, n=22, 46.1±1.4 yrs). A healthy control group was also involved (C, n=17, 49.6±1.4 yrs). We measured the arterial stiffness (PWV, pulse wave velocity), muscle sympathetic nerve activity (MSNA, microneurography), forearm blood flow (plethysmography), mean blood pressure (MBP, oscillometric), and peripheral vascular resistance (VR=MBP/forearm blood flow). MetS+IFG had similar PWV vs. MetS‐IFG, but had higher PWV than C (9.6±0.3, 9.1±0.2, and 8.7±0.2, P=0.02). Similarly, MetS+IFG had no different VR vs. MetS‐IFG, albeit had higher VR vs. C group (72.1±3.9, 57.5±5.4 and 47.4±9.2, P=0.01). Further analysis showed that PWV was correlated with MSNA (R=0.47; P<0.01). In conclusion, sympathetic hyperactivation may be, at least partially, the base mechanism that increase arterial stiffness. Impaired fasting glucose appears to be the main factor to aggravate the cardiovascular risk in metabolic syndrome patients.
Lesion of the Area Postrema Reduces Cardiac Sympathoexcitation and Improves Cardiac Function in Heart Failure
Authors: Abukar Yonis , Thomas Colleen , Ramchandra Rohit , Yao Song , May Clive
Published: June 2021

Background and Aims

Heart Failure (HF) is associated with increased cardiac sympathetic nerve activity (CSNA), but the mechanisms causing this are unclear. The area postrema (AP), a brainstem circumventricular organ, plays an important role in controlling SNA. We hypothesized that the elevated CSNA in HF is mediated in part by the AP and that lesioning this region would a) reduce CSNA in sheep with HF and b) improve cardiac function in rats with HF.

Methods

In sheep with pacing‐induced HF, lesion of the AP or sham lesion (n=6/group) was performed when ejection fraction (EF) had fallen to ~50%. In conscious sheep, blood pressure, heart rate and CSNA were recorded when EF was <40%. In rats, AP lesion or sham lesion (n=18/group) was performed before HF was induced by myocardial infarction (MI) and 11 weeks later left ventricular end‐diastolic pressure (LVEDP) was measured in anaesthetised rats.

Results

In sheep with HF there was a significant decrease in EF (from 74 ± 2 % to 38 ± 1 %, P <0.001) and lesion of the AP significantly reduced the elevated CSNA burst incidence (45 ± 10 vs. 89 ± 3 bursts/100 heartbeats, P<0.01). In rats with HF, LVEDP was significantly lower in the AP lesion group compared with the sham lesion group (11 ± 2 vs 17 ± 2 mmHg, respectively, P<0.05).

Conclusions

Lesion of the AP significantly reduced the elevated CSNA in sheep with HF and resulted in a lower LVEDP in rats with HF. These findings suggest that the AP plays a critical role in setting the high levels of CSNA in HF and indicate that lesion of the AP reduces the decline in cardiac function post‐myocardial infarction, possibly due to the reduction in CSNA.

Support: Melbourne University
Ablation of Sympathetic Neurons Projecting to Mesenteric Blood Vessels Reduces Splanchnic Nerve Evoked Systemic Blood Pressure Increase: Widespread Projection of Prevertebral Neurons to the Mesenteric Circulation
Authors: Hammond Bradley , Kreulen David
Published: June 2021

Celiac ganglionectomy attenuates the increase in arterial pressure (AP) seen in the DOCA‐salt model of hypertension; however, it is unknown which population of splanchnic sympathetic neurons is responsible for the attenuation of DOCA‐salt hypertension. Therefore, we ablated sympathetic post ganglionic neurons specifically innervating mesenteric blood vessels by applying an antibody to dopamine beta hydroxylase conjugated to the neurotoxin saporin (DβH‐Sap) on two, 7 mm segments of mesenteric arteries and veins separated from paravascular nerve trunks. Control animals received IgG‐saporin instead of DβH‐Sap. 3wks after application of saporin conjugates, rats were anesthetized, fitted with a femoral catheter, and the greater splanchnic nerve was dissected. The number of neurons in the celiac ganglia and splanchnic ganglion was considerably less in DβH‐Sap treated animals compared to IgG‐saporin treated control animals. Also, there was nearly complete elimination of TH‐positive perivascular nerve fibers. Stimulation of the greater splanchnic nerve (3Hz, 5v, 1ms for 20s) increased AP in IgG‐sap treated controls (6.2 ± 0.6 mmHg, N=4), while DβH‐Sap treated animals saw a decrease in AP (‐1.5 ± 1.5 mmHg, N=4). Ablation of vascular‐projecting neurons in prevertebral ganglia eliminates the systemic pressor response to splanchnic nerve stimulation. This study also shows that nearly all the vascular‐projecting neurons converge on two, 7 mm segments of mesenteric blood vessels, which suggests that individual prevertebral neurons project widely to the mesenteric circulation.
Effect of Carotid Body Resection on Baroreflex Control of Blood Pressure During Hypoglycemia
Authors: Limberg Jacqueline , Taylor Jennifer , Dube Simmi , Basu Ananda , Basu Rita , Curry Timothy , Joyner Michael , Wehrwein Erica
Published: June 2021

Background

Hypoglycemia results in a reduction in spontaneous cardiac baroreflex sensitivity (scBRS) and a shift in the baroreflex working range to higher heart rates; this effect may be mediated by the carotid chemoreceptors. We hypothesized changes in baroreflex control of heart rate with hypoglycemia would be blunted in carotid body resected patients when compared with healthy controls.

Methods

Five patients (3M/2F) with bilateral carotid body resection for glomus tumors and ten healthy controls (7M/3F) completed a 180‐min hyperinsulinemic, hypoglycemic (~3.3 µmol/mL) clamp. Changes in heart rate, blood pressure, and scBRS were assessed.

Results

scBRS was reduced from baseline during hypoglycemia (p<0.01) and was lower in carotid body resected patients when compared with controls (p=0.01). Hypoglycemia‐mediated reductions in systolic and mean blood pressure were significantly greater in patients when compared with controls (p<0.05). Despite lower blood pressures, increases in heart rate with hypoglycemia were blunted in patients (p<0.01).

Conclusion

Reductions in scBRS with hypoglycemia cannot be exclusively attributed to the carotid chemoreceptors; however, the chemoreceptors play a role in resetting the baroreflex working range to higher heart rates during hypoglycemia such that a rise in heart rate is not observed in carotid body resected patients. These data support a contribution of the carotid chemoreceptors to baroreflex control of blood pressure during hypoglycemia and highlight potential widespread effects of carotid body resection in humans.

Funding

DK090541, HL120570
Exogenous L‐Arginine Restores Spontaneous Cardiac Baroreflex Sensitivity in Never‐Treated Hypertensive Men
Authors: Vianna Lauro , Fernandes Igor , Barbosa Thales , Scharf Lucas , Rocha Natália , Secher Niels , Nóbrega Antonio
Published: June 2021

Depressed cardiac baroreflex sensitivity (cBRS) is associated with an increased risk of mortality in patients with hypertension. A candidate mechanism for the centrally mediated impairment in cBRS is reduced synthesis of the neuromodulator nitric oxide. Therefore, we investigated the effect of intravenous L‐arginine, a precursor of nitric oxide, on the cBRS in human hypertension. Beat‐to‐beat blood pressure (Finometer) and heart rate (ECG) were measured in both normotensive (n=8, 44±4 yrs., Norm) and never‐treated hypertensive men (n=8, 47±2 yrs., Hyper) before and during intravenous infusion of L‐arginine (1 g/min; total, 30 g) or saline. Linear regression analysis between RR interval and systolic blood pressure was used to determine the gain (i.e. sensitivity) of the arterial baroreflex control of the heart. Compared to Norm group, patients with hypertension had lower spontaneous cBRS (Norm: 13±1 vs. Hyper: 7.9±1 ms.mmHg‐1; P<0.01). On administration of L‐arginine, cBRS increased in both groups (Norm: 17±3 vs. Hyper: 15±3 ms.mmHg‐1; P<0.01 vs. before infusion) and returned to near resting values after infusion. Increases in cBRS during infusion of L‐arginine were greater in Hyper (P<0.05) and differences between groups disappeared. Importantly, during saline infusion, the spontaneous cBRS remained unchanged in both groups. In conclusion, these data demonstrate that short‐term L‐arginine infusion restores the arterial baroreflex control of heart rate in never‐treated hypertensive men, probably via increased nitric oxide synthesis.
TAK1 Signaling Downstream of PAR‐1 in Endothelial Cells Restores Lung Vascular Barrier Integrity
Authors: Soni Dheeraj , DebRoy Auditi , Wang DongMei , Vogel Stephen , Tiruppathi Chinnaswamy
Published: June 2021

The proinflammatory mediator thrombin increases vascular permeability by disassembling vascular endothelial adherens junctions (AJs). However, the intrinsic signaling mechanisms that mediate the reassembly of endothelial AJs are poorly understood. Here we show that transforming growth factor‐β‐activated kinase (TAK1) activation downstream of protease‐activated receptor‐1 (PAR‐1) signals the reassembly of endothelial AJs. PAR‐1 agonist, thrombin induced a time‐dependent phosphorylation of TAK1 in lung endothelial cells (ECs). Silencing of stromal interaction molecule 1 (STIM1) suppressed thrombin‐induced TAK1 phosphorylation in ECs, indicating that store‐operated calcium entry (SOCE) is essential for PAR‐1‐mediated TAK1 activation. Inhibition of TAK1 augmented thrombin‐induced permeability increase both in vitro and in intact lung microvessels. Consistent with these findings, inhibition of TAK1 impaired reassembly VE‐cadherin at AJs after thrombin treatment. We also observed that thrombin‐induced SOCE was markedly increased in TAK1 deficient ECs. Since global as well as EC‐restricted TAK1 knockout mice are embryonically lethal, to study the in vivo role of EC‐expressed TAK1, we generated tamoxifen‐inducible EC‐restricted TAK1 knockout (TAK1iΔEC) mice. Surprisingly, we observed markedly reduced expression of VE‐cadherin and its associated protein β‐catenin in lungs of TAK1iΔEC mice compared with their wild type littermates. We also observed enhanced lung vascular permeability as assessed by Evens blue dye‐conjugated albumin uptake in TAK1iΔEC mice. These findings support the notion that TAK1 signaling in endothelial cells is required to maintain lung vascular barrier integrity.
Pseudomonas aeruginosa Exoenzymes U and Y Induce a Transmissible Endothelial Tauopathy
Authors: Morrow Kyle , Ochoa Cristhiaan , Balczon Ron , Cauthen Laura , Alexeyev Mikhail , Schmalzer Katherine , Frank Dara , Stevens Troy
Published: June 2021

Here, we tested the hypothesis that Pseudomonas aeruginosa type 3 secretion system effectors ExoY and ExoU induce endothelial tau oligomerization and release, causing transmissible cytotoxicity characteristic of an infectious tauopathy. Both the bacterial delivery of ExoY and ExoU, and the conditional expression of an activity‐attenuated ExoU, induced time‐dependent pulmonary microvascular endothelial cell (PMVEC) gap formation that was paralleled by loss of intracellular tau and concomitant appearance of extracellular tau oligomers. Transfer of tau oligomers in filtered supernatant onto naïve endothelial cells resulted in the intracellular accumulation of oligomeric tau inclusions accompanied by cytotoxicity, as determined by inter‐endothelial gap formation and decreased endothelial network stability in Matrigel. Tau oligomer monoclonal antibodies captured monomeric tau from filtered supernatant, but did not retrieve higher molecular mass tau oligomers, and did not rescue the cytotoxic effects of tau. Since cytotoxic effects were not rescued, we combined an electrophoresis and electroelution approach to recover the high molecular mass tau oligomers, treat PMVECs with these oligomers, and recapitulate cytotoxicity. Thus, we provide the first evidence for a pathophysiological stimulus that induces endothelial tau oligomerization, release and transmissible cytotoxicity, characteristic of an endothelial tauopathy.
Quantitative Functional Cardiac Magnetic Resonance Imaging in Mice with Pulmonary Arterial Hypertension
Authors: Breen Ellen , Scadeng Miriam , Murray Fiona , Bigby Timothy
Published: June 2021

Pulmonary arterial hypertension (PAH) is a condition characterized by elevated mean pulmonary artery pressure, pulmonary vascular remodeling and secondary right heart failure. Mouse models are instrumental in understanding the pathophysiology of PAH and require precise methods to non‐invasively and longitudinally evaluate in vivo cardiac morphology and function. In this study we used magnetic resonance imaging (MRI) to measure in vivo cardiac dimensions in mice with PAH. PAH was induced in C57BL/6 mice using 10% hypoxia and weekly, subcutaneous injections of 20 mg/kg of SU5416 for three weeks. Short axis images at 1 mm intervals from the base to the apex of the heart were collected by proton MRI with a fast low angle SHot (FLASH)‐cine sequence, TE: 2.11 ms and TR: 8 ms. Thirteen images were collected throughout each cardiac cycle. Measurements are presented of right ventricle (RV) lateral wall thickness and ejection fraction. RV wall thickness at the uppermost region of the ventricle was greater in PAH mice than control at end‐diastole (control, 0.30 + 0.05 mm, PAH; 0.48 + 0.06 mm, p< 0.01, n=6) but not different at end‐systole (control, 0.59 + 0.11 mm; PAH, 0.70 + 0.11 mm, n=6). RV ejection fraction was decreased in PAH mice (control, 71.9% + 2.5, PAH 57.6% + 5.4, p=0.04, n=6). These data demonstrate that MR imaging can detect right ventricle remodeling and cardiac dysfunction under in vivo conditions.

Funded by Department of Veteran Affairs
Lithium Potentiates Epithelial–Mesenchymal Transition in the Kidney through Induction of Matrix Metallopeptidase‐9
Authors: Ward John , Swaim Grace , Gooch Jennifer , Blount Mitsi
Published: June 2021

Lithium is a common therapeutic agent that is used to effectively treat patients with various mood disorders. Unfortunately there are many renal side effects of lithium therapy, including tubular atrophy and chronic interstitial fibrosis, which are not fully understood. To identify the intracellular signaling cascades that cause the proliferative action of lithium in the inner medullary collecting duct (IMCD), mIMCD3 cells were incubated with increasing concentrations of lithium chloride for 48 h. We found that lithium inhibited glycogen synthase kinase 3β (GSK‐3β) activity through serine‐9 phosphorylation in a dose responsive manner. Lithium inhibition of GSK‐3β induced ERK phosphorylation a concentration‐dependent manner. Both GSK‐3β and ERK are known to be involved in a variety of biological processes and are suggested to be key in the proliferative and antiapoptotic actions of lithium in the renal collecting duct. A reported downstream target of these kinases is matrix metallopeptidase 9 (MMP‐9), an enzyme that induces tubular cell epithelial‐mesenchymal transition (EMT) and therefore a major contributor to renal fibrosis. We found that protein abundance of MMP9 was elevated with increasing lithium concentrations in mIMCD3 cells. These results suggest that lithium inactivation of GSK‐3β potentiates EMT through ERK induction of MMP‐9 in the collecting duct.
The Homeodomain Protein Cux1 is Required for Cyst Development in an ADPKD Mouse Model
Authors: Vanden Heuvel Greg , Wright Joshua , Andres Erica , Woo Anna , Baybutt Richard , Paul Binu
Published: June 2021

Cux1 is a homeobox gene involved in cell cycle regulation and kidney development. Cux1 is upregulated in the kidneys of both mice and humans with autosomal dominant polycystic kidney disease (ADPKD). However, Cux1 transgenic mice do not develop cystic kidneys, indicating that upregulation of Cux1 alone is insufficient to develop PKD. Mice carrying a deletion of the C‐terminus of Cux1 (Cux1tm2Ejn‐/‐) express a truncated protein that does not localize to the nucleus, resulting in functional loss of Cux1 transcriptional activity. To test the hypothesis that Cux1 is required to develop PKD, we generated collecting duct specific Pkd1 mutant mice (Pkd1CD) that were also homozygous for the Cux1 mutation. When kidneys isolated from newborn Pkd1CD/Cux1tm2Ejn ‐/‐ mice were examined, we found a marked reduction in cystic dilations compared to the kidneys isolated from newborn Pkd1CD mice. We confirmed the Cre activity by crossing the Pkd1CD and Pkd1CD/ Cux1tm2Ejn ‐/‐ mice to the membrane‐targeted Tomato/Green Fluorescent Protein (mT/mG) double fluorescent reporter mouse strain (Gt(ROSA)26Sortm4(ACTB‐tdTomato,‐EGFP)Luo/J), which carries a loxP‐flanked Tomato cassette. Immunofluorescence of kidney sections indicated that collecting duct derived cysts were positive for the excision event in the Pkd1CD mice. In contrast, although collecting ducts were positive for the excision event in Pkd1CD/ Cux1tm2Ejn ‐/‐ mice, none of the positive tubules were cystic. Taken together, our results suggest that Cux1 gene dosage influences the progression of cystic disease in an ADPKD mouse model.
Clomiphene Citrate Induced Kidney Injury in Female Albino Rat
Authors: Lamfon Hawazen , Lamfon Nahid
Published: June 2021

Clomiphene citrate (CC), a selective oestrogen‐receptor modulator, is used in treatment of polycystic ovary syndrome. On the other hand, it showed many adverse effects. The present work investigates the effects of CC on kidney of female albino rats. Many biochemical and histological changes were induced in these animals when treated with CC at doses of 10, 50 and 100 mg/kg body weight. Treating animals with doses of 50 and 100 mg/kg body weight caused elevations in serum creatinine and urea. The histological changes in the kidney of treated animals included intertubular leucocytic infiltrations, congestion of blood vessels and degeneration of renal tubules. Moreover, atrophy of glomeruli was recorded. In conclusion, biochemical and histopathological alterations indicate that clomiphene citrate causes renal damage in albino rats. The present work is supported by Umm Al‐Qura University and King Abullaziz medical City / National Guard Health Affair/ Riyadh
Nanoparticle Interactions With Renal Epithelial Cells in vivo
Authors: Nair Anil , Keliher Edmund , Core Amanda , Brown Dennis , Weissleder Ralph
Published: June 2021

Nanotechnology approaches are actively being pursued for drug delivery, novel diagnostics, implantable devices, and consumer products. While considerable research has been performed on the effects of these materials on targeted tumor or phagocytic cells, little is known about their effects on renal cells. This becomes critical for supersmall (< 10 nm) nanoparticles (NP), designed to be renally excreted. To test NPs effect on kidney function, we injected mice with either 5 nm dextran NPs (DNP) that are similar in composition to FDA approved materials or poly(amido amine) dendrimer NPs (PNP) of comparable size. These fluorophore tagged NPs were filtered and internalized by renal cells in a dose and time dependent way. The biological effects were quantitated by immunofluorescence, measuring kidney injury markers and performing functional tests. DNP administration resulted in dose dependent increases in urinary output, while cellular albumin endocytosis was increased. The expression of megalin was also increased but AQP1 expression was unaffected. The effects after PNP administration were similar but additionally resulted in increased clathrin expression, increased endocytosis and decreased AQP1 expression. We conclude that there are no major detrimental effects of DNP on overall kidney function but changes in protein expression do occur. These studies provide a framework for the testing of additional NP preparations as they become available.
Seven Encapsulated and Nonencapsulated (iso)microRNAs in Human Samples Predict Coronary Atherosclerosis and Cardiac Events
Authors: James Amanda Marie , Thomas Sheena , Rooney Kimberly , Quyyumi Arshed , Wu Hao , Johnson Zachary , Searles Charles
Published: June 2021

Coronary Artery Disease (CAD) is a chronic inflammatory process that modifies microRNA (miR) abundance, intracellularly, in vascular cells, and extracellularly, in plasma. Extracellular miRs are encapsulated in microvesicles (MV) or protein‐bound, and variations in miR profiles are promising as novel biomarkers of CAD severity. We hypothesized that CAD‐associated variations in the miR profile are related to changes in the abundance of miR isoforms (isomiRs). IsomiR sequences differ by 1‐3 nucleotides from the consensus miR sequences. Using RNA‐deep sequencing, the change in abundance/frequency of isomiRs was assessed in human blood samples from cohorts with varying CAD severity. This analysis highly expressed 18 miRs whose dominant form was an isomiR of a consensus miR. Using custom‐designed RT/PCR primers for the 18 isomiRs, unfractionated and fractionated plasma from patients with/without significant CAD were assessed for isomiR concentrations. Seven isomiRs (miR‐10b, 30d, 93, 143, 181a, 182, and 744) were detectable and distinctly different in abundance between cohorts. Subgroupings of these 7 isomiRs (based on abundance/frequency within each plasma fraction) were examined to develop an isomiR biomarker panel. Statistical comparison and correlation analysis of this subgrouping had an accuracy of 69‐100% in predicting relevant clinical outcomes such as CAD severity, MV counts (inflammation marker), or myocardial infarction. These data support the development and clinical use of an isomiR biomarker panel to assess CAD risk.
Genetic fine‐mapping and gene identification of adiposity traits in outbred rats
Authors: Solberg Woods Leah , He Hong , Holl Katie , Littrell John , Prokop Jeremy , Zagloul Maie , Keele Greg , Baur Brittany , Fox Joseph , Robinson Melanie , Levy Shawn , Valdar William
Published: June 2021

Obesity and overweight are major risk factors for multiple diseases including cardiovascular disease and type 2 diabetes. Genetic factors are known to contribute significantly to obesity and related traits with heritability ranging from 50‐80%. Human genome‐wide association studies have identified over 50 genes for traits related to adiposity. These genes, however, explain less than 10% of the heritable variance. Heterogeneous stock (HS) rats are outbred from eight inbred strains and allow genetic fine‐mapping to only a few Megabases. The goal of the current study was to use HS rats to fine‐map adiposity traits genome‐wide and identify underlying candidate genes. We measured body weight, body mass index and visceral adiposity in 1090 male HS rats and genotyped all rats using the Affymetrix 10K SNP array. Linkage disequilibrium mapping by mixed model regression was used to identify significant loci. We identified one or more significant loci for all traits, with an average confidence interval of only 2.2 Mb. Using expression data and founder sequence, we identified potential candidate genes within several loci, including those that have previously been identified in human genome‐wide association studies. Protein modeling was used to support a causal role for some of the genes. These results demonstrate the power of HS rats for fine‐mapping metabolic traits and rapidly identifying candidate genes.

Supported by NIH R01 DK‐088975 and the Individualized Medicine Institute at MCW.
Claudin 3 CRISPR Knockout Mice are Healthy and Fertile
Authors: Kerr Jenessa , El Andalousi Jasmine , Yamanaka Yojiro , Ryan Aimee , Gupta Indra
Published: June 2021

Claudins regulate the passage of ions and solutes between neighboring cells through tight junctions. The nephron is the functional unit of the kidney that filters the blood and removes excess water, salts and wastes. The filtering properties of the nephron are determined by the unique combination of claudins expressed within each segment. Claudins are expressed in the nephric duct in advance of kidney function. Nephric duct derivatives undergo branching morphogenesis and ultimately each branch tip induces a mesenchymal‐to‐epithelial transition event required for nephron formation. In the chick embryo, a reagent that specifically removes Claudin 3 and 4 from tight junctions blocked nephric duct formation. This led us to hypothesize that claudins are required for kidney development in the mouse. Claudin‐4 knockout mice do not have an embryonic phenotype, therefore, Claudin‐3 was prioritized for investigation. Claudin‐3 compound heterozygote mice were created using the CRISPR/Cas9 system of genome editing. Male and female founder mice were generated with distinct frameshift mutations in both Claudin‐3 alleles. Neither of these founders showed an obvious phenotype. Pups derived from mating these founders appeared grossly normal. Histological analysis of the kidneys did not reveal a nephron phenotype. In conclusion, Claudin‐3 null mice generated by CRISPR/Cas9 are healthy and fertile. We hypothesize that the lack of Claudin‐3 is compensated by other claudins. This will be tested by creating double knockouts using the CRISPR/Cas9 system, which would be predicted to create a more severe phenotype and allow further understanding of the important role of claudins during kidney development.
A Potential Role of Histone Methyltransferase, Dot1l, in Renal Regulation of Acid‐Base Balance
Authors: Nakhoul Nazih , El‐Dahr Samir , Brown Karen , Islam M Toriqul , Khan Altaf , Hering‐Smith Kathleen , Hamm L.Lee , Abdulnour‐Nakhoul Solange
Published: June 2021

The histone methyltransferase Dot1l is a key epigenetic factor that plays a critical role in embryogenesis, angiogenesis, expression of the collecting duct epithelial Na channel (ENaC)and possibly hypertension. Deletion of Dot1l from renal cells expressing AQP2 resulted in a phenotype whereby kidneys had fewer principal cells and more intercalated cells (IC). It was reported that Dot1l, forming a complex with Af9 (a DNA binding protein) mediated transcriptional repression of ENaC‐α. Because of this transcriptional effect we hypothesized that targeted deletion of Dot1l from the collecting duct may affect expression of other transporters in the collecting duct. To test this hypothesis, we crossed Hoxb7‐Cre to Dot1F/F mice to generate collecting duct Dot1l knockout (Dot1l‐/‐) mice. We then placed control (WT) and renal Dot1l‐/‐ mice on acid diet for 3 days and examined the expression and distribution of Rhbg and Rhcg by immunohistochemistry and Western analysis. Rhbg and Rhcg are NH3/NH4+ transporters located respectively at the basolateral and apical membranes of IC. In response to acid diet Dot1l‐/‐ mice showed an increase in net NH4+ excretion. Rhcg and Rhbg were abundantly expressed but there was no change in Rhcg among the different groups. Rhbg expression was similar in WT & Dot1l‐/‐ under normal diet but was significantly increased in Dot1l‐/‐ mice on acid diet. These data suggest that Dot1l pathways may play a role in affecting renal acid‐base transport. This may be due, at least in part, to the increased expression of Rhbg transporters in DOT1l‐/‐ but a direct effect on transporter function is also possible.

Supported by VA Merit Grant
The SGK1/NEDD4‐2 pathway is crucial in regulating renal potassium secretion
Authors: Staub Olivier , Al‐Qusairi Lama , Basquin Denis , Stifanelli Matteo , Loffing Johannes
Published: June 2021

Dietary K+ load results in hyperkalemia, with consequent aldosterone release in order to stimulate K+ secretion in the distal nephron. The molecular mechanisms of this regulation are still not fully elucidated. Here, we aim to identify the role of the aldosterone induced SGK1 kinase and its target, the ubiquitin‐protein ligase NEDD4‐2 in the regulation of short and long term K+ secretion. To avoid the compensatory mechanisms which may mask the role of SGK1 and Nedd4‐2 during kidney development, we employed the previously described inducible nephron specific SGK1‐KO and Nedd4‐2‐KO mouse models in which a deletion of the target gene in renal tubules was observed. Our results indicate that short‐term K+ regulation (30 min to 2h) is not altered in SGK1 KO mice. However, these animals exhibit 35% decrease in urinary K+ excretion after 2 days of exposure to high K+ diet (5%) leading to hyperkalemia. Molecular analysis of WT and SGK1 KO mice after 2 days of HK diet revealed that the cleavage and the membrane localization of ENaC alpha and gamma subunits are decreased in the KO mice. On the same line, Nedd4‐2 phosphorylation was also decreased suggesting that NEDD4‐2 mediated internalization of ENaC is more active in mutant mice. Interestingly, when Nedd4‐2 KO mice were challenged by low K+ diet (<0.1%), they showed an abnormal ability to handle K+ deprivation. More specifically, they exhibit hypokalemia and urinary K+ loss likely mediated by increased ENaC activity as demonstrated by the increased membrane localization of alpha and gamma ENaC subunits. In conclusion, our data suggest that SGK1 and its target NEDD4‐2 control K+ homeostasis through the regulation of ENaC membrane localization
Re‐investigation into the Ligand Selectivity of OAT2
Authors: Hotchkiss Adam , Li Mansong , Ingraham Leslie , Pelis Ryan
Published: June 2021

There are discrepancies in the literature regarding ligand selectivity of organic anion transporter 2 (OAT2). The purpose of this study was to re‐examine the interaction of ligands with both transcript variants of OAT2 since it is not known whether they are functionally distinct. The two splice variants of human OAT2 (OAT2‐TV1 and OAT2‐TV2) were stably transfected into three cell lines (CHO, HEK and MDCK), and the interaction of previously identified substrates and inhibitors was assessed. Expression of both transcript variants was confirmed at the mRNA level, but only OAT2‐TV1 protein was detected at the cell surface. Uptake of penciclovir was ~1.5‐20 fold higher in OAT2‐TV1 cells compared to control, but para‐aminohippurate (PAH), estrone sulfate (ES), succinate and glutarate transport was not observed. Eight Krebs cycle intermediates (1mM) failed to cis‐inhibit penciclovir uptake. Conversely, cis‐inhibition by a structurally diverse set of xenobiotic drugs, including anions, cations and neutral compounds suggests that OAT2‐TV1 is broadly selective. Treatment of cells with sulfo‐NHS‐SS biotin inhibited penciclovir transport. Co‐treatment of cells with furosemide along with sulfo‐NHS‐SS biotin ameliorated transport activity, suggesting that lysine residue(s) line the ligand binding surface. These results show that OAT2‐TV2 is non‐functional in mammalian cells, likely due to a trafficking defect. Additionally, lack of mediated transport or cis‐inhibition by several previously reported compounds highlights the necessity to re‐examine ligand selectivity of OAT2‐TV1 to better understand its importance in pharmacokinetics. Supported by CIHR and NSHRF.
Angiotensin II‐induced Hypertension Impairs Hematopoietic Stem Cell Homing and Engraftment
Authors: Kim Seungbum , Cogle Christopher , Raizada Mohan
Published: June 2021

Immunosuppressive drugs such as cyclosporine commonly used in hematopoietic stem cell transplantation (HSCT), increase patients' blood pressure. Despite this hypertensive side effect, little is known about the influence of hypertension (HTN) on HSC and bone marrow (BM) activity. The objective of this study was to investigate if HTN exerts deleterious influences on HSC biology and its engraftment compromising efficient transplantation. Ang II induced HTN resulted in increased proliferation of HSCs as evidenced by 83% increase in Sca‐1+, c‐Kit+, Lin‐ (SKL) hematopoietic stem/progenitor cells (HSPC) and 207% increase in CD150+, CD48‐ SKL long‐term HSC (LT‐HSC) in mice. In contrast, hypertensive mice that were lethally irradiated and reconstituted with HSC had 65% diminished engraftment efficacy. The retrospective analysis of HSC transplanted patients showed that graft failure patients tended to have HTN when compared to the well‐recovered group (50% vs. 25%). Time‐lapse in vivo imaging of mouse tibia demonstrated that HSC in HTN mice failed to engraft to the BM osteoblastic HSC niche due to untimely proliferation and differentiation from Ang II infusion. In addition, HSCs pretreated with Ang II for 18 hours in vitro showed diminished BM reconstitution ability showing that Ang II directly modulates the HSC engraftment potential. These observations demonstrate that 1) chronic Ang II‐induced HTN regulates HSC proliferation, 2) although the hypertrophic effect of Ang II increases the HSC numbers, it impairs the homing ability and reconstitution potential of donor HSCs. They suggest that HTN and BP control should be important consideration in clinical application of HSCT.
Cellular Reprogramming of Epithelial Cells Leading to Conditional Immortalization is Accompanied by Changes in Multiple Pathways
Authors: Upadhyay Geeta , Glazer Robert , Petricoin Emanuel , Jablonski Sandra , Krawczyk Ewa , Graham Garrett , Khurana Anikita , Ligaba Segni
Published: June 2021

Co‐culture of human primary epithelial cells with irradiated 3T3 fibroblast feeder cells (J2 cells) and the Rho kinase inhibitor Y‐27632 (Y) allows for the unrestricted growth of cells of epithelial origin by the process termed conditional reprogramming. To better understand the nature of the signaling processes associated with conditionally reprogrammed cells, the effect of the two critical components of the co‐culture conditions, J2 cells and Y, on the growth of human foreskin keratinocytes (HFKs) was evaluated by gene expression profiling, reverse‐phase protein arrays and siRNA screening. J2 cells and Y acted cooperatively to down‐regulate differentiation, and upregulate proliferation and cell adhesion, including increased pT308Akt and pERK, and reduced TGF‐β pathway signaling. These findings establish a mechanistic basis for the unlimited growth potential of human epithelial cells that will be invaluable to assess the effect of genetic changes in pathologic tissues and their response to therapeutic agents.
Chemoprotective natural compounds targeting DNA damaged stem cells‐ the cells of origin of colon cancer
Authors: Kim Eunjoo , Davidson Laurie A Davidson , Patil Bhimanagouda , Jayaprakasha Guddadarangavvanahally , Callaway Evelyn , Turner Nancy , Chapkin Robert
Published: June 2021

The majority of colon tumors are triggered by aberrant Wnt signaling in intestinal stem cells, which is an extremely efficient route towards initiating intestinal cancer. Curcumin, a component of turmeric, and n‐3 polyunsaturated fatty acids (PUFAs), mainly found in fish oil, exert chemoprotective and anti‐inflammatory properties in part by modulating apoptotic, Wnt, and/or NFkappaB related pathways. However, the chemoprotective effects of diet on colonic adult stem cells‐the cells of origin of colon cancer are unknown. Therefore, we determined whether the combination of fish oil (FO) and curcumin (C) reduce colon cancer risk by modulating targeted apoptosis, DNA damage repair and proliferation in colonic Lgr5+ stem cells at the initiation stage of cancer compared to corn oil (CO, control) by utilizing Lgr5‐EGFP‐IRES‐creERT2 knock‐in mice fed diets containing CO±C or FO±C for 3 weeks followed by carcinogen (azoxymethane) injection. Interestingly, FO+C combination synergistically increased apoptosis (TUNEL) by 5‐fold, and promoted targeted DNA repair (MGMT+) by 9‐fold in DNA damaged (gamma H2AX+) stem cells compared to control. In general, stem cells were more responsive to environmental (carcinogen & diet) cues compared to non‐stem cells. These novel results demonstrate the unique ability of colonic stem cells to respond to external cues.