bluuebunny/crossref_metadata_2025 huggingface dataset:
Resveratrol optimizes the pharmacotherapy of captopril in renal hypertensive rats
Authors: Garcia Arthur F. E. , Restini Carolina B.A.
Published: June 2021

We evaluate the probable cardiac benefit by the antioxidant Resveratrol (RESV) treatment and Captopril (CPT). It was measured Cardiac and Ventricular Hypertrophy Indexes (CHI; VHI) and Systolic Blood Pressure (SBP). Male Wistar rats (180g) had undergone surgery for implantation of a silver clip in the left renal artery. Control rats were undergone fictitious laparotomy (2K). Six weeks after surgery, 2K‐1C (SBP >; 160 mmHg) and 2K rats were submitted to different treatments by gavage, 3 times/week: RESV (20mg/kg); vehicle; CPT (6 or 12 mg/kg) and the association of the CPT doses + RESV (20mg/kg). 3 weeks after, SBP, CHI and VHI were measured. CHI was reduced in RESV 2K (3.0 ± 0.05, n=9) when compared to RESV 2K‐1C (3.4±0.1, n=18) and 2K‐1C (3.4 ±0.2, n=9) or 2K (3.2 ± 0.3, n=6) untreated. CPT equalized to the other groups CHI values to the RESV 2K.VHI also was reduced in RESV 2K (2.2 ± 0.05, n = 9) compared to RESV 2K‐1C (2.7 ± 0.1, n = 18) and 2K‐1C (2.9 ± 0.2, n = 9) or 2K (2.7 ± 0.2, n = 6) untreated. CPT presented the same effects on VHI and CHI values. Untreated 2K‐1C, CHI and VHI, was increased compared to the 2K‐1C RESV or RESV + CPT (VHI: 2.2 ± 0.1; CHI: 2.9 ± 0.1, n = 4) treated. RESV effectively reduced the SBP in 2K‐1C (99.0 ± 4.1, n = 10) when compared to RESV 2K (100.0 ± 10.4, n=8) and 2K (105.0 ± 5.0, n=8) or 2K‐1C (170.0 ± 10.8 n = 4) untreated. In the 2K‐1C, SBP reduction by CPT (140.0 ± 4.5, n=4) was less effective than by RESV or CPT + RESV (154.0 ± 13.7, n=5). RESV has potential to reduce SBP on 2K‐1C. CPT or RESV had effectively reduced the CHI and VHI in 2K‐1C and 2K rats, suggesting both treatments are acting in the same via to attenuate the hypertrophy.

Support: PIBIC‐CNPq (02–2012). Ethics Committee approval: 007/2010.
An important role of renal angiotensin‐converting enzyme in the development of salt‐sensitivity during renal parenchyma inflammation
Authors: Giani Jorge Fernando , Janjoulia Tea , Fuchs Sebastien , Peti‐Peterdi Janos , Bernstein Kenneth E. , Gonzalez‐Villalobos Romer A
Published: June 2021

Studies suggest that intrarenal renin‐angiotensin system (RAS) activation plays an important role in the establishment of salt sensitivity associated with renal inflammation. To determine this, we studied the responses of mice lacking renal ACE (ACE 10/10) to a high‐salt (HS) diet after a transient exposure to L‐NAME. In this model, known as Post L‐NAME hypertension, the appearance of salt sensitivity is associated with renal inflammation and RAS activation. Wild‐type (WT) and ACE 10/10 mice (n=12) were subjected to a 4‐wk L‐NAME treatment (50 mg.Kg‐1.day‐1) followed by a 1‐wk washout and a 3‐wk HS diet (4% NaCl). In WT mice, L‐NAME treatment increased systolic blood pressure (Δ: 22±2 mmHg; p<0.05), that returned to baseline during washout and increased again in response to HS diet (Δ: 21±3 mmHg; p<0.05). In contrast, ACE 10/10 mice were resistant to L‐NAME treatment (Δ: 5±2 mmHg; p=NS) and the HS salt diet (Δ: −3±1 mmHg; p=NS). Further, when compared to ACE 10/10 mice, WT mice displayed higher renal TNF‐α content (54±6 vs. 36±4 pg/mg; p<0.05; ELISA), macrophage infiltration (118±10 vs. 86±8 F4/80 positive cells/field, immunohistochemistry) and renal angiotensinogen (2.82±0.16 vs. 1.30±0.14 fold over basal; p<0.01; Western blot). These results demonstrate that renal ACE plays a fundamental role in the development of hypertension and the salt sensitivity observed during renal inflammation in the Post L‐NAME model
Regulation of AT1‐Receptors by ARAP1 is involved in vasodilatation during sepsis‐induced hypotension
Authors: Mederle Katharina , Doblinger Elisabeth , Kattler Veronika , Höcherl Klaus , Schweda Frank , Castrop Hayo
Published: June 2021

Hypotension in septic patients results from hypovolemia, vasodilatation and hyporeactivity to vasoconstrictors. Altered expression of angiotensin receptors might contribute to reduced vascular tone during sepsis. The AT1 receptor‐associated protein 1 (ARAP1) is expressed in vascular smooth muscle cells and increases the surface expression of the AT1‐receptor. Here we assessed the role of ARAP1 in sepsis‐induced hypotension.

Mean arterial blood pressure (MAP) (102±2 vs.103±2 mmHg; n=6; p=.66) and glomerular filtration rate (296±20 vs. 315±30 μl/min; n=10; p=.61) were similar in ARAP1−/− and WT mice. Plasma renin concentration was increased in ARAP1−/− mice compared to WT mice (66±6 vs. 41±4 ng AngI/ml/h; n=23; p=.001). Dose‐dependent changes in vascular resistance were right‐shifted in isolated perfused kidneys from Arap1−/− mice. LPS‐treatment (3mg/kg) induced a marked downregulation of ARAP1 expression in different organs. Following LPS, MAP (84±2 vs. 90±1 mmHg; n=10; p=.03) and heart rate (446±13 vs. 492±10 bpm; n=10; p=.01) decreased to lower levels in ARAP1−/− mice compared to WT mice.

Under baseline conditions, ARAP1−/− mice seem to normalize blood pressure by compensatory activation of the RAS. However, sepsis‐induced hypotension is enhanced in ARAP1−/− mice. Downregulation of ARAP1 expression during sepsis may contribute to hypotension by causing reduced vascular sensitivity to Ang II.
HIF‐1alpha and AMPK interactions regulate cellular hypoxia adaptation in the subtotal nephrectomy model of CKD
Authors: Singh Prabhleen , Satriano Joseph , Li Hui , Hallows Kenneth
Published: June 2021

HIF‐1α activation in subtotal nephrectomy (STN) ameliorates hypoxia and improves GFR. The AMPK pathway facilitates adaptation to hypoxia, and there is significant overlap between the actions and targets of HIF‐1α and AMPK. Here we investigate the effects of HIF‐1α activation on cellular physiology and the interaction between HIF‐1α and AMPK in early STN kidney.

Kidneys harvested from control, 1‐week STN, and STN treated with DMOG (HIF‐1α inducer) were examined by Western blotting. The expression of molecular markers for hypertophy (p70S6k) and apoptosis (cleaved caspase‐3) were elevated in STN, while a marker for autophagy (LC3II/LC3I) was reduced. HIF‐1α induction completely reversed these processes. The activated AMPK (pAMPK) to total AMPK ratio was low in STN and increased by HIF‐1α activation. All results were significant at p<0.05.

Interactions between HIF‐1α and AMPK in HK2 and MDCK with AMPK knockdown (KD) cell lines were investigated. Consistent with the in vivo data, DMOG treatment increased pAMPK levels, increased LC3II/LC3I and lowered p70S6k in HK2 cells. These effects were reversed by HIF‐1α inhibition by YC‐1, with a dramatic increase in cleaved caspase3. AMPK activation by AICAR blocked the increase in cleaved caspase3 induced by YC‐1, and this was reversed in the AMPK KD cells. Inhibition of LC3II/LC3I by YC‐1 was AMPK‐independent. Finally, both AMPK and HIF‐1α activation inhibited p70S6K.

We propose that HIF‐1α activation in the STN kidney facilitates hypoxic adaptation and cell survival by inhibiting hypertrophy and apoptosis and activating autophagy and the AMPK pathway. These novel findings suggest that HIF‐1α and AMPK closely interact with each other to mount a coordinated cellular response to hypoxic stress.
Dopamine inhibits basolateral potassium channels in the murine distal nephron
Authors: Zaika Oleg L. , Mamenko Mykola , Pochynyuk Oleh
Published: June 2021

It is recognized that dopamine promotes natriuresis by inhibiting NHE3 and Na+‐K+‐ATPase in the proximal tubule. In contrast, less is known about putative dopamine actions on the water‐electrolyte transport in the distal nephron. Basolateral side of the epithelial cells in the distal nephron is exposed to influence of dopamine secreted from sympathetic nerve endings. Furthermore, dopamine receptors are abundantly expressed at the basolateral membrane of the connecting tubule and the collecting duct cells. Basolateral K+ channels in the distal nephron are critical for potassium recycling and controlling basolateral membrane potential to promote electrogenic Na+ reabsorption. Using patch clamp electrophysiology in freshly isolated distal nephrons from mice, we detected ~20 pS and 40 pS basolateral K+ channels most likely representing Kir4.1, Kir4.1/5.1, respectively. Dopamine reversibly inhibited both channels when applied in a low micromolar range. Importantly, dopamine also decreased the amplitude of unitary currents approximately by 30% indicating depolarization of basolateral membrane and reduction of transepithelial voltage. We propose that consistently with its actions in the proximal tubule, dopamine reduces Na+ transport in the distal nephron, in part, by inhibiting basolateral K+ conductance and decreasing driving force for sodium reabsorption.
Mechanosensing with restart of flow drives KATP channel induced NOX2 activation in a model of Lung Ischemia Reperfusion
Authors: Chatterjee Shampa , Wang Hui , Hong Nankang , Yu Kevin , Debolt Kristine , Feinstein Sheldon I , Christie Jason D , Cantu Edward , Fisher Aron B
Published: June 2021

From our earlier studies in lung ischemia, we concluded that endothelial mechanotransduction with stop of flow in the lung activates the generation of reactive oxygen species (ROS) as a result of KATP closure that leads to NOX2 activation. As the next phase, we evaluated the role of mechanosignaling with reperfusion (I/R). We now propose that the increase in shear with reperfusion triggers a mechanosignaling cascade similar to that associated with ischemia. Isolated perfused mouse lungs and pulmonary endothelial cells showed a 2 fold increase in ROS production (as monitored by ROS sensitive dyes H2DCFDA, H2DFFDA and DHE) with ischemia that is further enhanced by 3–3.5 fold upon reperfusion. This was not observed in lungs from KATP channel null and NOX2 mice. Lungs subjected to ischemia or I/R show ~1.8 fold increase in lung damage (as assessed by 8‐isoprostanes, TBARS, permeability) with ischemia and a ~2.5 fold increase during the reperfusion period. Understanding the initiation of reperfusion signaling has potential application in lung transplantation which requires reperfusion of a previously ischemic lung; thus this model is relevant to design strategies that can reduce NOX2 activation and resultant injury.
EFAVIRENZ, ATAZANAVIR AND RITONAVIR IMPAIR SARCO(ENDO)PLASMIC RETICULUM Ca2+ CYCLING
Authors: Bidasee Keshore , Shao Chun Hong , McMillan JoEllyn , Cibrowski Pawel , Gendelman Howard , Tian Chengju
Published: June 2021

Efavirenz (EFV), atazanavir (ATV) and ritonavir (RTV) are key components of combination anti‐retroviral therapy used to reduce viral replication in therapy naive HIV‐infected individuals. However, their beneficial effects are being compromised by clinically important adverse side‐effects including cognitive decline, lipoatrophy, cardiac dysfunction and skeletal muscle weakness. These undesired effects have been attributed to mechanisms including induction of the unfolded protein response (UPR), autophagy, and mitochondrial dysfunction. What is less clear is whether these mechanisms are initiating causes or secondary consequences stemming from the actions of these drugs at upstream cellular targets. This study assesses whether EFV, ATV and RTV alter sarco(endo)plasmic reticulum (SR/ER) Ca2+ content, which if compromised will induce these untoward effects. At pharmacologically‐relevant concentrations, EFV (0.24 μM), ATV (1.5 μM) and RTV (0.8 μM) bind to and modulate ryanodine receptors, the channels through which Ca2+ is released from the SR/ER, with varying potencies. These drugs also dose‐dependently activated and/or inhibited SERCA1 and SERCA2, ATPases that translocate Ca2+ from the cytoplasm to the lumen of the SR/ER. Exposure of quiescent C2C12 myotubes and rat ventricular myocytes to EFV, RTV, and ATV elevated cytoplasmic Ca2+ and decreased the SR/ER Ca2+ content by >;50%. Overnight exposure of C2C12 cells with EFV and RTV also triggered the unfolded protein response. These data provide new mechanistic insights into the tissue/organ‐selective adverse effects of three first‐choice antiretrovirals. Supported in part by NIH HL085061
Acute Administration of PPARγ Agonist Rosiglitazone in Isolated Hearts Differentially Aggravates Cardiac Ischemia Reperfusion Injury in a Consomic Rat Model
Authors: Cheng Qunli , Elorbany Reem , Stowe David , Camara Amadou , Weihrauch Dorothee , Riess Matthias
Published: June 2021

The effect of peroxisome proliferator‐activated receptor‐gamma (PPARγ) agonist rosiglitazone (RGZ), a thiazolidinedione (TZL), on cardiac ischemia/reperfusion (IR) injury is controversial. We tested effects of RGZ on post‐IR function and infarction in a consomic rat model that exhibits genetically determined cardioprotection. 15 min after 2 × 5‐min boluses of 50 μM RGZ or vehicle (Con), isolated hearts from Brown Norway (BN), Dahl Salt Sensitive (SS), and consomic (SS‐6BN) rats underwent 30 min global I and 120 min R. Systolic and developed left‐ventricular pressure (LVP), contractility, and coronary flow at 120 min R were decreased while diastolic LVP and infarct size increased in all Con groups, more so in SS than in BN or SS‐6BN hearts. RGZ enhanced IR injury more in SS‐6BN than in SS or BN hearts. Concurrent fluorescent redox state measurements suggested incomplete washout of RGZ and mitochondrial uncoupling before IR. These data confirm prior reports of deleterious effects of TZL on cardiac IR injury that appear to vary by species, dose, timing and route of administration, model used and PPARγ‐independent actions. These results also suggest genetic predisposition plays a role in the adverse response to TZLs. If translated clinically, PPARγ agonists may adversely enhance cardiac IR injury in genetically susceptible patients. Research supported by Department of Veterans Affairs (MR: CARA‐026–10F) and NIH.
Deficiency in caveolin‐1 or type 2 diabetes augments vasoconstrictor responses to sphingosine‐1‐phosphate
Authors: Lamping Kathryn G , Nelson Peter M , Nuno Daniel W
Published: June 2021

Sphingolipids are not only integral membrane components but also vasoactive signaling molecules. We tested the hypothesis that vascular responses to sphingosine‐1‐phosphate (S1P) are augmented in diabetics through abnormal rho kinase and caveolar regulation. We measured responses of isolated pressurized resistance arteries from gracilis muscle of wild type (WT, diameter 101±23μm), caveolin‐1 deficient which lack caveolae (Cav−/−, diameter 130±30μm) and a model of type 2 diabetes in mice [leptin receptor deficient (db/db), diameter 123±11μm] to S1P (1nM‐1μM) and KCl (25–100 mM) in the absence and presence of a rho kinase inhibitor, H1152 (1 μM). S1P produced concentration‐dependent vasoconstriction that was increased 60–70% (P<0.05) in arteries from Cav−/− and db/db mice. H1152 had minimal effect on constriction of arteries from WT mice but inhibited responses in arteries from Cav−/− and db/db mice to the levels of WT. Vasoconstriction to KCl was similar in all groups. We conclude that in type 2 diabetics, responses of resistance arteries in skeletal muscle to S1P are augmented. The increased constriction may be due, in part, to abnormal caveolar localization and/or rho kinase activation. Support: Dept of Veterans Affairs
Functional and structural alterations in urinary bladder of an obese Zucker rat model
Authors: Jiuan‐Miaw Liao
Published: June 2021

Diabetic bladder dysfunction (DBD) is a most common complication of diabetes mellitus (DM), and is seen in 40–80% of diabetic patients. It is characterized by impaired bladder sensation, increased bladder capacity, decreased bladder contractility and increased residual urine. The goals of these studies were to test the hypothesis that the obese Zucker rat is an appropriate model for type 2 diabetic bladder investigations and to define the alterations in bladder structure and functional responses in this setting. Histological examination of bladder cross sections was shown that total bladder wall thickness and threshold pressure of voiding were significantly increased in obese rats, while voiding time, maximal intravesical pressure and intercontraction intervals significantly declined as compared to age‐matched lean rats, respectively. Contractile responses of obese DM strips to Aceltylchine (Ach) and KCl were significantly less than those of bladder strips from lean animals. In addition to generating less force, it can also be seen that strips from the obese DM strips showed slower rate of force development than strips from lean animals. Two major myosin isofroms, SM1 and SM2 myosin heavy chains (MHCs) were separated, estimated and the ratios averaged shown that the SM2‐to‐SM1 isoform ratios were declined in obese bladders. After 12 % SDS‐PAGE, transfered and immunoreacted with antibody against tropomyosin (Tm) and α‐actin (as an internal control), the Tm/α‐actin ratios increased in obese bladders as compared to age‐matched lean bladders. All the data indicated that the obese Zucker rat is a relevant model for type 2 diabetic bladder investigations than the streptozotocin‐induced animal model.
Quercetin Flavonoid Has In Vivo Antioxidant Properties By Increasing Hepatic Expression Of Gpx And Hepatic Gpx Enzyme Activity In Old Rats
Authors: Nassef Noha , Louka Manal , Habib Haidy
Published: June 2021

Background

Quercetin, one of the major flavonoids present in the human diet, has been associated with a wide range of biological properties, including antioxidant, anti‐inflammatory and anticarcinogenic actions. The antioxidant action is considered as the most likely mechanism for the beneficial effects of quercetin. However, little is known on its activity in vivo. Therefore, the present study aimed to investigate the in vivo effects of quercetin on mRNA and activity levels of hepatic GSTa2 in aged rats. This study was carried out on 28 male Wistar albino rats of 24‐months age. Rats were allocated into 2 groups: 1‐Quercetin treated group (14 rats): Received Quercetin (50 mg/kg by gavage for 7 days). 2‐Control group (14 rats): Received 1 ml/kg distilled water by gavage for 7 days.

Results

Quercetin treatment resulted in significant increase in relative hepatic mRNA levels for glutathione peroxidase (GPx) and liver GPx enzyme activity (1.67 ± 0.78 versus 1 ± 0.05, p< 0.01 and 375.92±27.84 versus 242.78 ±19.79 mU/mg protein, p<0.01 respectively). On the other hand, it significantly decreased relative hepatic mRNA levels for Gsta2, hepatic GST enzyme activity and hepatic glutathione concentrations (0.45 ± 0.16 versus 1 ± 0.04, p< 0.01, 127.78±12.9 versus 181.85±17.23 mU/mg protein, p<0.01 and 19.55 ±3.96 versus 23.78 ± 3.28, p<0.01 respectively). No significant difference was found in BMI or in plasma concentration of ox‐LDL between the two groups of aged rats.

Conclusion

Our results indicate that quercetin flavonoid has in vivo antioxidant properties by increasing hepatic expression of GPx and hepatic GPx enzyme activity in old rats.
Overexpression of CuZnSOD in median preoptic nucleus improves cardiac function after myocardial infarction in the rat
Authors: Collister John , Hartnett Cristina , Nahey David , Stauthammer Christopher , Kruger Maxie , OSullivan Gerry , Parker Josh , Tian Jun , Zimmerman Matthew
Published: June 2021

It has been shown that central overexpression of the superoxide scavenging enzyme, Cu/Zn superoxide dismutase (SOD), in the subfornical organ improves cardiac function in a mouse model of heart failure (HF). A downstream hypothalamic site, the median preoptic nucleus (MnPO), may act as a relay center for reactive oxygen species to act as mediators in the pathophysiology of HF. To test the hypothesis that elevated superoxide in the MnPO is related to the pathophysiology of HF and decreased cardiac function, we injected adenovirus encoding CuZnSOD (AdSOD, n=2) or control empty vector (AdE, n=3) into the MnPO of normal rats. Subsequently, rats were subjected to coronary artery ligation to create a myocardial infarct (MI) of the left ventricle. Cardiac function was monitored at 2 week intervals via echocardiography. Upon completion, rat brains were examined for SOD expression in MnPO via immunofluorescence and histopathological analyses of cardiac infarct size were conducted. Baseline (EF) ejection fractions (%) of AdSOD and AdE rats were 62±4 and 67±2, respectively. Two weeks after MI, EF was significantly decreased in both groups of rats (AdSOD: 38±3, AdE: 48±4). In contrast by 6 weeks post MI, EF had improved to 52±1 in AdSOD rats yet was only 41±6 in AdE rats. In conclusion, despite decreases in EF early after MI, overexpression of SOD in the MnPO was related to an improvement in left ventricular function 6 weeks post MI.
Altered activation of a different type of nitric oxide synthase over the course of repeated hepatic ischemia/reperfusion in rats
Authors: Miyake Takashi , Yokoyama Yukihiro , Kokuryo Toshio , Mizutani Tetsushi , Imamura Atsushi , Nagino Masato
Published: June 2021

Objective

The aim of this study was to determine the intrahepatic kinetics of nitric oxide synthase during ischemia/reperfusion (I/R).

Methods

Intrahepatic nitric oxide (NO) levels were measured using a selective NO sensor. Changes in hepatic microcirculation were investigated using intravital microscopy. Hepatic expression of endothelial nitric oxide synthase (eNOS), phospho‐eNOS, and inducible NOS (iNOS) were evaluated by western blot, reverse transcription polymerase chain reaction (RT‐PCR), and immunohistochemical analysis.

Results

Intrahepatic NO levels gradually increased and then reached a plateau approximately 15 min after starting ischemia. The sinusoids after reperfusion were dilated compared with the sinusoids before ischemia. The expression of phospho‐eNOS during I/R markedly increased during the first ischemia, and then the levels attenuated during the subsequent repeating I/R cycles; however, the expression of iNOS gradually increased, as observed by western blot, RT‐PCR, and immunohistochemical analysis.

Conclusion

Our results indicate that eNOS, through activation of the phospho‐eNOS system, promptly responds to ischemic stress in the super‐acute phase of ischemia. Subsequently, repeating I/R stress induces iNOS gene expression, although it may take at least an hour to upregulate the iNOS protein.
Interactions between L‐arginine supplementation and physical training in the mitochondrial biomarkers and nitric oxide bioavailability in rat skeletal muscle
Authors: Zanesco Angelina , da Silva Carmem V , Delbin Maria A , La Guardia Paolo G , Davel Ana P , Priviero Fernanda B
Published: June 2021

Nitric oxide (NO) and physical exercise regulate mitochondrial function with improvement of resistance to fatigue and muscular wasting. We studied the relationship between physical performance and the protein expressions of mitochondrial function markers (mtTFA and ATP synthase subunit c) in gastrocnemius muscle from exercised rats supplemented with L‐arginine during 8 weeks. We also analyzed the nNOS/NO pathway by measuring the content of tissue nitrite/nitrate as well as proteins expression of Cu/Zn superoxide dismutase (Cu/Zn‐SOD), neuronal nitric oxide synthase (nNOS) and mitochondrial manganese superoxide dismutase (Mn‐SOD) in the cytoplasm and mitochondrial fractions from skeletal muscle. Wistar rats were divided into: control sedentary (SD), sedentary supplemented with L‐arginine (SDLA), trained (TR) and trained supplemented with L‐arginine (TRLA). L‐arginine supplementation‐plus physical training promotes an up‐regulation of the protein expressions of ATP synthase subunit c, nNOS, Cu/Zn‐SOD, Mn‐SOD and mtTFA in gastrocnemius muscle that was associated with improvement in fatigue resistance and reduction in body weight. Our data shows that run physical training in combination with L‐arginine supplementation initiate a sequence of biochemical events in isolated gastrocnemius muscle that culminate in improvement of fatigue resistance as well as in lowering body weight gain.
Activation of PDK1 mediates VSMC migration and may contribute to vascular remodeling following injury
Authors: Weber David S , Sullivan Lydia M , Bennett Justin R , McCarthy Cullen
Published: June 2021

We hypothesized that increased 3‐phosphoinositide‐dependent kinase 1 (PDK1) activation contributes to arterial wall thickening following vascular injury. Vascular smooth muscle cells (VSMCs) were pretreated with BX‐912, a small molecule inhibitor of PDK1, prior to stimulation with PDGF. Western analysis confirmed inhibition of PDK1 activity in the presence of BX‐912 by decreased phosphorylation of PDK1 (ser241) by ~60% and Akt (thr308) by ~ 70%. Conversely, Akt phosphorylation at ser473 was only mildly affected. To examine VSMC migration, both scratch wound assays and modified Boyden chamber studies were completed. In both instances, wound‐ and PDGF‐induced chemotaxis was attenuated following BX‐912 treatment. Following wire injury of the carotid artery in C57BL6/J mice, in vivo PDK1 phosphorylation (ser 241) was assessed by immunoblotting in both injured and contralateral control carotids at 3, 7, and 14 days post injury. PDK1 phosphorylation was markedly increased at day 3 (~5 fold vs. non‐injured), remained slightly elevated at day 7 (~2 fold) and returned close to baseline by later timepoints following injury. Our findings indicate that BX‐912 is effective in attenuating PDK1 activity and chemotaxis in VSMCs and that increased PDK1 activity occurs following vascular injury. Thus, PDK1 inhibition, via BX‐912 treatment, could potentially be utilized to attenuate wall remodeling in response to injury. Support: NIH HL084159
Role of vascular hyperpolarization, nitric oxide, and prostaglandins in reactive hyperemia in healthy humans
Authors: Crecelius Anne R , Richards Jennifer C , Luckasen Gary J , Larson Dennis G , Dinenno Frank A
Published: June 2021

Reactive hyperemia (RH) in response to forearm ischemia can be an important cardiovascular risk marker yet the underlying vasomotor signaling mechanisms have been difficult to elucidate. We tested the hypothesis that vascular hyperpolarization via activation of KIR channels and Na/K‐ATPase (Na/K) contributes to peak and total (150 sec area under curve; AUC) RH in young healthy humans. Subjects (3 groups of n=8) were instrumented with a brachial artery catheter for local inhibition of KIR channels (BaCl2), Na/K (ouabain), nitric oxide synthase (L‐NMMA) and cyclooxygenase (COX; ketorolac). Forearm blood flow (FBF; venous occlusion plethysmography) was measured at rest and immediately following 5 min of forearm ischemia. Compared to control, inhibition of KIR channels reduced peak RH (ΔFBF; −48±8%; P<0.05) whereas no reduction occurred with inhibition of Na/K (+2±6%). Combined KIR and Na/K inhibition further reduced peak RH (−61±6%). Individual inhibition of KIR channels (−61±3%) or Na/K (−44±12%) reduced AUC and combined inhibition had a greater effect (−87±4%). NOS+COX inhibition did not reduce peak RH (+18±8%) or AUC (−10±12%) and did not further reduce the responses observed during combined KIR channel and Na/K inhibition. Our collective results reveal a novel role for vascular hyperpolarization via KIR channels as the primary pathway involved in both the peak and sustained RH in humans. Support: HL102720
Proteomic analysis of aortic and coronary resistance microvessel tissue in type 2 diabetic db/db mice
Authors: Trask Aaron J. , Cismowski Mary J. , Halleck Kathryn , Lucchesi Pamela A.
Published: June 2021

The current study analyzed alterations in protein expression between normal and type 2 diabetic aortas and coronary resistance microvessels (CRMs) as we have previously shown differential remodeling in these vessels. Aortas and CRMs (n=2 pools of 4 vessels each) were isolated from control Db/db and diabetic db/db mice, lysed, and 16 Âμg protein for each sample was separated by 10% SDS‐PAGE. Bands were excised from Sypro‐stained gels were subjected to nano LC‐MS/MS at the Ohio State University Mass Spec & Proteomics Facility. Significant increases in extracellular matrix proteins fibronectin (10.3‐fold) and prolargin (1.8‐fold), and in the cytoskeletal proteins gelsolin (2.3‐fold) were observed in diabetic relative to normal aortas. Diabetic CRMs expressed a 71% decrease in cysteine‐rich protein 2 transcription factor and ~50% lower levels of cytoskeletal proteins Î±‐actinins‐1, 4, and filamin A relative to control CRMs. Gelsolin was only expressed in control CRMs. These data reveal a protein expression profile that may explain a pro‐fibrotic milieu in diabetic macrovessels (e.g. aortas), while increased transcription factor and decreased cytoskeletal protein expression in diabetic CRMs may suggest a distinct VSMC phenotype that may partially explain the adverse inward hypertrophic CRM remodeling associated with a reduction in wall stiffness. Support: NIH T32HL098039 to AJT; NIH R01HL056046, NCH to PAL
Microvascular Smooth Muscle Gene Program is Highly Sensitive to Blood Flow
Authors: Reho John J , Fisher Steven A
Published: June 2021

Microvascular smooth muscle (MVSM) is phenotypically and functionally distinct from that of the large arteries yet the deterministic input signals are not known. We use Myosin Phosphatase as a model to address this question as it is a primary mediator of vasomotor responses. Isoforms of the myosin phosphatase regulatory subunit (Mypt1) are generated by alternative splicing of exon 24 (E24) and determine MVSM responses to NO/cGMP. Mesenteric artery MVSM expresses the E24+ (fast) isoform; large arteries express the E24‐ (slow) isoform. Ligation of the downstream MA2s causes switching of the upstream MA1s to the E24‐ isoform suggesting the hypothesis that the unique (fast) contractile phenotype of MVSM is determined by blood flow. To test this we developed a model of reduced perfusion pressure in which placement of an occluder around the abdominal aorta allows for transient reductions in blood flow within the rat mesenteric arcade. Brief (20 & 60min) and repetitive (60min; 5days) perfusion pressure reduction via the balloon occluder causes a switch in the splicing of E24 in Mypt1 of mesenteric arteries from the E24+ to E24‐ isoform 6 hours after pressure reduction in both brief (20% E24+ vs 80% control; p<0.05) and repetitive (45% E24+; p<0.05) protocols. Given these effects of reduced perfusion pressure we propose that the unique properties of the MVSM are highly dependent upon the pulsatile blood flow generated by the heart
Critical role of Kv channels in cerebrovascular dysfunction associated with ischemic small vessel disease in a mouse genetic model
Authors: Dabertrand Fabrice , Bonev Adrian D , Krøigaard Cristel , Brayden Joseph E , Joutel Anne , Nelson Mark T
Published: June 2021

Dominant mutations in NOTCH3 gene induce the most common heritable cause of stroke and vascular dementia, referred to as Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy (CADASIL). Using a recently developed mouse model of CADASIL (Joutel, JCI, 2010), we examined the consequences of this mutation on the function of intracerebral arterioles. Elevation of intravascular pressure to 20 mm Hg constricted isolated arterioles from control and CADASIL mice to a similar extent. However, above 30 mm Hg, CADASIL arterioles constricted significantly less than control. At 40 mm Hg, CADASIL arterioles were 38% less constricted, and the smooth muscle membrane potential was 10 mV hyperpolarized compared to control. We hypothesize that this difference was due to increased activity of either calcium‐sensitive BK channels or voltage‐dependent potassium (Kv) channels. BK channel blockers had no effect on the diameter of CADASIL arterioles. Kv current density was 30% greater in myocytes from CADASIL arteries than in control, and activation of epidermal growth factor receptor which causes Kv channel endocytosis restored myogenic tone in CADASIL arterioles. These results support the concept that the CADASIL mutation decreases pressure‐induced vasoconstriction through upregulation of smooth muscle Kv channels. Supported by AHA 09POST2290090 (FD), the NIH R37DK053832, PO1HL095488, RO1HL44455, RO1HL58231, the Totman Trust for Medical Research and the Fondation Leducq for the Transatlantic Network of Excellence on the Pathogenesis of Small Vessel Disease of the Brain.
Early changes to circadian rhythm account for elevated blood pressure and sympathetic activity during a high fat diet in rabbits
Authors: Lim Kyungjoon , Burke Sandra L , Barzel Benjamin , Armitage James A , Head Geoffrey A
Published: June 2021

Consumption of a high fat diet (HFD) by rabbits results in rapidly increased blood pressure (BP), heart rate (HR) and renal sympathetic activity (RSNA). We determined how early these changes occurred and if they were related to changes in circadian rhythms. Rabbits were meal‐fed a 13.5% HFD and BP, HR and RSNA were measured daily via implanted telemeters in the home cage for 3 weeks and for 1 week after return to a normal diet. Baseline BP, HR and RSNA over 24h were 71±1mmHg, 205±4b/min and 7±1nu. Circadian rhythms were entrained to the feeding cycle and values increased from the preprandial minimum to the postprandial maximum by 4±1mmHg, 51±6b/min and 1.6±0.6nu. Within 2 days of commencing a HFD, 24h BP, HR and RSNA had increased by 2%, 18% and 22% respectively and the preprandial minimum was elevated by 79–127%. This inhibition lasted for 3 weeks of HFD and accounted for all of the hypertension and 50% of the RSNA activation. Circadian rhythms of BP and HR, but not RSNA, switched to being synchronised with the light‐dark cycle. Resumption of a normal diet for 1 week did not reverse the light entrainment or alter the inhibition of the BP preprandial dip but the HR pattern was progressively restored. Thus, HFD‐induced hypertension, mediated by rapid changes in sympathetic nerve activity, is due to the immediate (within 2 days) and continued loss of preprandial dipping.
Tetrahydrobiopterin attenuates sympathetic vasoconstrictor responsiveness in resting and contracting skeletal muscle
Authors: Jendzjowsky Nicholas Gregory , DeLorey Darren Scott
Published: June 2021

We tested the hypothesis that acute tetrahydrobiopterin (BH4) supplementation would attenuate sympathetic vasoconstrictor responsiveness in resting and contracting skeletal muscle. Sprague‐Dawley rats (n=15) were anesthetised and instrumented for lumbar sympathetic chain stimulation and measurement of arterial blood pressure and femoral artery blood flow. Femoral vascular conductance (FVC) was calculated. The sciatic nerve was stimulated to contract the triceps surae muscle group at 30 and 60% of maximal contractile force (MCF). The percent change in FVC in response to sympathetic stimulation at 2 and 5 Hz was determined at rest and during contraction before and after BH4 (20mg/kg + 10mg/kg/hr IA) supplementation. BH4 blunted (p<0.05) the vascular response to sympathetic stimulation at rest (CONTROL: 2Hz: −28 ± 5%; 5Hz: −45 ± 5%; BH4: 2Hz: −17 ± 4%; 5Hz: −34 ± 7%) and during contraction at 30%MCF (CONTROL: 2Hz: −14 ± 6%; 5Hz: −28 ± 11%; BH4: 2Hz: −6 ± 6%; 5Hz: −16 ± 10%) and 60%MCF (CONTROL: 2Hz: −7 ± 3%; 5Hz: −16 ± 6%; BH4: 2Hz: −3 ± 3%; 5Hz: −11 ± 6%). In conclusion, acute supplementation of the nitric oxide synthase cofactor BH4 reduced the vascular response to sympathetic stimulation in resting and contracting skeletal muscle. These data demonstrate that BH4 supplementation augmented nitric oxide‐mediated inhibition of sympathetic vasoconstriction. NSERC, Canada.
Chromatin Replication and Epigenome Maintenance
Authors: Alabert Constance , Groth Anja
Published: June 2021

In dividing cells, faithful duplication of the genome must be accompanied by reproduction of the chromatin landscape on new DNA. Our research focuses on how human cells replicate chromatin and ensure transmission of genetic and epigenetic information during cell division.

Recently, we have developed an explorative proteomic approach, Nascent Chromatin Capture (NCC), to profile chromatin replication in human cells. NCC relies on biotin‐dUTP labeling of replicating DNA and cross‐linking, followed by affinity‐purification of tagged chromatin and quantitative proteomics. By comparing nascent newly synthesized chromatin with mature post‐replicative chromatin, we provide association dynamics for 3995 proteins. Highly enriched in nascent chromatin, we find the complete human replisome followed by central players in chromatin assembly and cohesion establishment. In contrast, a large set of chromatin proteins including histone H1 show delayed association. By combining NCC enrichment with experimentally derived chromatin probabilities, we also use the data set to identify new replication factors. Together, this provides an extensive resource to understand genome and epigenome maintenance. In addition, this proteomic instrument provides opportunity to address transmission of histone marks and replication stress responses.
RNA‐seq analysis identifies novel gene signatures common to SHP knockout mice and human steatosis, fibrosis, NASH and cirrhosis
Authors: Smalling Rana , Delker Don , Zhang Yuxia , McGuiness Michael , Liu Shuanghu , Hagedorn Curt , Wang Li
Published: June 2021

Chronic hepatitis C virus and non‐alcoholic fatty liver disease are major causes of cirrhosis and hepatocellular carcinoma. It is vital to identify genetic changes fueling liver disease progression in order to better prevent, diagnose and treat chronic liver disease. Nuclear receptor small heterodimer partner (Shp) null mice are an attractive model as they have altered hepatic lipid and bile acid metabolism and develop spontaneous hepatoma. Using next generation RNA sequencing, we identified common genes that were significantly differentially expressed in Shp−/− mice, hepatitis C cirrhosis, and non‐alcoholic steatohepatitis (NASH). Novel gene signatures and their functional significance were validated in human steatosis, fibrosis, NASH, alcohol and hepatitis C cirrhosis as well as in Shp−/− and SHP transgenic mice, liver fibrosis, inflammation and bile acid injury models. The novel genes include peptidoglycan recognition protein 2 (PGLYRP2), monooxygenase, DBH‐like 1 (MOXD1), dual specific phosphatase‐4 (DUSP4), thrombospondin 1 (THBS1), tetraspanin 4 (TSPAN4), SPARC‐related modular calcium binding protein‐2 (SMOC‐2), latent transforming growth factor‐beta binding protein 4 (LTBP4), and family with sequence similarity 198a (FAM198A).

Conclusion

Our study has identified novel genes implicated in chronic human liver diseases using RNA sequencing and Shp−/− mice as our primary model.

This study was supported by the NIH DK080440 (LW) and CA148068 (CHH).
Student Peer Review and Self‐Reflection in the Gross Anatomy Laboratory: Identifying Student Perceived Strengths and Areas for Improvement
Authors: Edmondson Anna , Mackey John Evan , Shelley Robert , Nichols Carol
Published: June 2021

Giving feedback is a skill health professionals need. While uncomfortable, the small group setting in anatomy lab provides an ideal environment for this in regards to professionalism & communication objectives. We examined perceived strengths & weaknesses of allied health students (physician assistant, occupational & physical therapy) in lab. Students provide feedback via evaluative ratings & comments for lab partners & themselves at the course end. Credit is given for completing it in a timely, professional manner & providing comments. Evaluation questions assess communication/collaboration & professionalism domains. Students scored themselves significantly lower than peers on several questions. Narrative comments were thorough, with more emphasis on strengths. For peers, 62% of positive comments were associated with themes involving professional/leadership skills, followed by personal behaviors (26%) & knowledge (12%). Similar results were seen in self‐reflections. Constructively critical comments were more closely distributed between professional/leadership skills (41%), personal behaviors (33%) & knowledge (26%). While similar results were seen in self‐reflections, more students recognized their own knowledge deficiencies. No difference was noted in student comments based on program. While allied health students may find giving peer feedback difficult, it is thorough & insightful.

Grant Funding Source: N/A
Relationship between spatial abilities and three‐dimensional synthesis of structures
Authors: Langlois Jean , Dagenais Yvan , Lemieux Renald , Martin Marcel , Lecourtois Marc , Yetisir Elizabeth , Bergeron Germain , Hamstra Stanley , Wells George A.
Published: June 2021

Objective

The mental image of a three‐dimensional anatomical structure is a prerequisite to the performance in technical skills. The objective was to assess the relationship of spatial abilities to three‐dimensional synthesis of two‐dimensional views of structures using drawings.

Methods

Fourth‐year medical students (n = 49) were recruited in a prospective study. Spatial abilities were measured with a redrawn Vandenberg and Kuse Mental Rotations Test in two (MRTA) and three (MRTC) dimensions and the Surface Development Test (SDT). As part of a drawing course, participants had to build structures of increasing complexity from simple parts using two‐dimensional views of structures and then to draw isometric views of structures. The accuracy of the drawings was assessed as right or wrong. The maximum score was 24 for MRTA and MRTC, 60 for SDT and 25 for the drawings. The results were expressed as means ± standard deviation and the Spearman's correlation coefficient was used to compare the drawing score to MRTA, MRTC and SDT scores.

Results

The drawing score (14.6 ± 3.7) was related to MRTA (13.5 ± 5.2), MRTC (9.7 ± 4.5) and SDT (43.4 ± 10.0) scores; with a correlation of 0.3728 (p = 0.0083), 0.4248 (p = 0.0023) and 0.5420 (p < 0.0001), respectively.

Conclusion

Spatial abilities were related to three‐dimensional synthesis of two‐dimensional views of structures using drawings.

This study was supported by an internal grant from the Department of Surgery, University of Sherbrooke, Sherbrooke, QC, Canada.
Does exam performance correlate with perception of readiness or pretest quizzes?
Authors: Ball Alexander , Belbeck Larry , Volc Nicole , Helli Peter , Wainman Bruce
Published: June 2021

We tested the hypothesis that student's perceived readiness for examinations was not correlated to exam performance as has been previously reported. In a population of 615 undergraduate nursing, midwifery, baccalaureate health sciences (BHSc) and engineering students in a required human anatomy and physiology course perceived readiness for examinations was surveyed prior to the beginning of the midterm exam. Perceived readiness was weakly correlated with exam performance (r=0.21, p=0.05). Performance on fortnightly quizzes administered in the lab was more strongly correlated with exam performance (r=0.34, p<0.005) than perceived readiness. Though perceived readiness had a small significant correlation for most students groups the BHSc students, who have the highest course average of 84%, showed no correlation (r=0.15, n = 211). The quiz performance was most strongly correlated with the midterm exam performance in the midwifery students (r=0.61, p<0.005, n=22) and the nursing students (r=0.46, p<0.005, n= 322). This research confirms that students are only poorly able to judge exam preparedness and quantitative measures on structured tests are the most useful predictor of student success.
Anatomical knowledge retention in third year medical students prior to OB/GYN and surgery rotations
Authors: Jurjus Rosalyn A , Krapf Jill , Ahle Samantha , Brown Kirsten , Butera Gisela , Goldman Ellen , Lee Juliet
Published: June 2021

The literature indicates that surgical specialists have a low opinion of students’ anatomical knowledge in the clinical setting; quantification of this is lacking. Our multi‐disciplinary team aimed to identify baseline anatomical knowledge in third‐year medical students prior to surgical rotations and in advance of developing an integrated curriculum.

During the first week of the Obstetrics and Gynecology and Surgical rotations, 120 consenting students completed a short multiple‐choice examination consisting of clinically‐ based anatomy questions. Examination scores were calculated for each anatomic category based upon 100% expected knowledge after preparing for the national board examination. Preliminary data indicate a significant anatomic knowledge gap between the preclinical and clinical years: a majority of students scored below 50% in vascular, abdominal, and inguinal anatomy, 11% in questions of lymphatic drainage of the pelvis, 45% in embryology, and 32% in neuroanatomy.

This data will be used to create interactive modules, using principles of adult learning and instructional design, aimed at improving clinically‐relevant anatomic knowledge prior to inpatient surgical rotations. With these endeavors, we aim to show the benefits of longitudinal integration of anatomical modules before surgical specialties in our curriculum.

Grant Funding Source: None
An Interprofessional Approach to Improve Gestational Outcomes
Authors: von Hagen Laura Jane , Penava Deborah , Johnson Marjorie , Mottola Michelle
Published: June 2021

Sixty percent of obese women exceed the recommended criteria for gestational weight gain during pregnancy, potentially compromising optimal fetal growth. Pregnancy represents a unique period of time conducive to lifestyle modification during which we can provide patients with health advice and education; provision of information alone, however is insufficient for long‐term behavior changes. “My Clinic” is an Interprofessional clinic for pregnant women with BMI>;35 developed to improve health outcomes through lifestyle changes. Effective low cost behavior tools that promote healthy lifestyles are needed to focus clinical interventions. The purpose of this study is to assist pregnant women referred to “My Clinic” achieve a healthy pregnancy. Participants will be provided with a series of computer or paper‐based modules aimed at improving their physical activity levels and eating habits. Individually tailored modules include the following topics: time management, goal setting, action planning, self‐talk and overcoming barriers. We predict that participants undergoing specialized care and completing the modules will show increased self‐efficacy and action planning for physical activity and healthy eating, preventing excessive gestational weight gain compared to women receiving standard care in this novel study.

Grant Funding Source: Departmental
Cadaver Lab, White Boards and Pedagogy: Uniting Teachers, Students and Anatomy
Authors: Harper Noah , Morton David A , Nielsen Mark T
Published: June 2021

Human anatomy is not a prerequisite for admittance to medical school, thus matriculated students enter with a wide variety of anatomical backgrounds, many having no experience studying anatomy. Most medical gross anatomy courses take a regional approach to cadaveric dissection, which can leave students struggling to connect individual, regional structures to associated functional systems. Time constraints of rigorously scheduled, preclinical curricula leave little opportunity to create training sessions that address gaps in “big picture” knowledge. It is possible to address some of these issues by training instructors to incorporate brief anatomical stories and simple yet powerful illustrations that unite embryological, regional and systemic concepts while simultaneously highlighting elegant anatomical patterns of the body. Additionally, these novel stories and illustrations promote consistency in teaching approaches, especially when applied to impromptu “teachable moments” often encountered in laboratory settings. Illustrations that highlight the development and final form of ventral strap, subvertebral, four‐layer lateral wall, and epaxial musculature can be used to simplify trunk anatomy, including the neck where pharyngeal arch migration adds layers to a basic pattern. Moreover, the narrative can be further applied to connect and simplify patterns of innervation and vascularization.

Grant Funding Source: Department of Neurobiology and Anatomy