bluuebunny/crossref_metadata_2025 huggingface dataset:
Renal ischemia‐reperfusion (I/R) injury induces a rapid activation of local inflammatory markers and causes increased peritubular permeability.
Authors: Tenstad Olav , Bivol Liliana Monica , Iversen The late Bjarne Magnus , Hultström Michael , Reed Rolf Kåre , Wiig Helge
Published: June 2021

Local cytokines were measured in renal hilar lymph and in cortex after 30 min unilateral occlusion of renal artery followed by 120 min reperfusion. The effect of I/R on size selectivity for proteins in glomerular and peritubular capillaries were estimated using HPLC and mass spectrometry. Whereas all measured mediators increased dramatically in renal hilar lymph, plasma and renal cortical tissue samples and approached control levels after 120 min reperfusion, the responses were differentiated. IL‐1beta, MCP‐1 and leptin were markedly increased in plasma before reperfusion, reflecting an extra renal response possibly induced by afferent renal nerve activity from the ischemic kidney. The cortical cytokine content also reached high levels before reperfusion suggesting proteolytic release of local stores. However, only TNF‐alpha showed increased lymph to plasma ratio (L/P>;1) following 30 min reperfusion indicating release of mediators directly into the bloodstream in agreement with abundant cytokine staining of glomerular and peritubular capillaries. The cytokine levels were paralleled by a significant increase in high molecular weight plasma proteins in both lymph and urine indicating temporarily reduced size selectivity of both glomerular and peritubular capillaries. In conclusion, short time renal ischemia is a strong trigger for a local as well as systemic inflammatory reaction.
Short Sleep Duration and Endothelial t‐PA Release in Overweight/Obese Adults
Authors: Bammert Tyler D. , Beckstrom Collin A. , Greiner Jared J. , Stauffer Brian L. , DeSouza Christopher A.
Published: June 2021

Short sleep duration is associated with cardiovascular (CV) morbidity. The specific mechanisms responsible for this increased CV risk are not fully understood. Overweight and obesity (OW/OB) is associated with both sleep disturbances and impaired endothelial function. We determined whether habitual short sleep duration worsens endothelial fibrinolytic function in OW/OB adults. 20 OW/OB adults were studied: 10 normal sleepers (age: 57±2 yr; BMI: 29.6±1.2 kg/m2; sleep/night: 7.9±0.2 hrs) and 10 short sleepers (age: 52±2 yr; BMI: 31.1±1.4 kg/m2; sleep/night: 6.2±0.2 hrs). Short sleep was defined as habitual nightly sleep of less than 7 hours. Endothelial fibrinolytic function was assessed, in vivo, by acute tissue‐type plasminogen activator (t‐PA) release to intrabrachial infusion of bradykinin (BK) and sodium nitroprusside. There were no group differences in basal endothelial t‐PA release between the groups. Importantly, in response to BK the increase in t‐PA release was not significantly different between the OW/OB normal sleepers (from −0.3±0.5 to 37.7±5.3 ng/100 mL tissue/min) and OW/OB short sleepers (from −0.7±0.9 to 36.7±5.0 ng/100 mL tissue/min). Further, total t‐PA release was similar between the groups (236.3±27.8 vs 199.3±36.9 ng/100 mL tissue). Thus, habitual short sleep is not associated with further diminishment in endothelial t‐PA release in OW/OB adults.
Resolving inflammation in stroke through FPR2/ALX
Authors: Gavins Felicity N. E. , Smith Helen K.
Published: June 2021

Members of the formyl peptide receptor family (in particular FPR2/ALX; Fpr2/3 in mouse) act as endogenous anti‐inflammatory/ pro‐resolving agents. We investigated the pharmacological potential of FPR ligands in modulating inflammation in stroke.

Mice underwent bilateral common carotid artery occlusion (5min) and reperfusion (40min or 2h). FPR agonists (Annexin A1 peptide Ac2–26 (AnxA1Ac2–26) and 15‐epi‐lipoxin A4 (15‐epi‐LXA4)) were administered at the start of reperfusion, and resulting cerebral leukocyte‐endothelial (L‐E) interactions were viewed using intravital microscopy.

At 40 min reperfusion, 15‐epi‐LXA4 (0.5μg and 4μg/mouse) reduced inflammation vs. vehicle treatment, although this reduction was not sustained at 2h with 0.5μg/mouse. AnxA1Ac2–26 treatment decreased leukocyte adhesion at 40min and 2h vs. vehicle. Combined treatment of AnxA1Ac2–26 and FPR antagonists Boc2 or WRW4 (FPR2/ALX‐selective) abrogated the effects of AnxA1Ac2–26 at 40min, suggesting an Fpr2/3 mechanism. Abrogation was not sustained at to the same extent at 2h.

To conclude, 15‐epi‐LXA4 and AnxA1Ac2–26 reduced L‐E interactions in stroke, in a dose and time‐dependent manner, and likely via FPR2/ALX. These novel data indicate that that modulation of FPR2/ALX could produce significant protective effects in clinical stroke.

Funded by the BBSRC.
BK channels are not involved in the ryanodine‐induced vasoconstriction of the spiral modiolar artery
Authors: Krishnamoorthy Gayathri , Reimann Katrin , Wangemann Philine
Published: June 2021

We investigated whether ryanodine‐sensitive Ca2+‐sparks activate BK channels to lower smooth muscle global Ca2+ and cause vasodilation in the spiral modiolar artery (SMA). Ca2+‐sparks in the smooth muscle cells of the SMA were abolished by ryanodine at pressures of 40 and 60 cmH2O. At 40 cmH2O, ryanodine, but not tetracaine, abolished Ca2+‐waves, increased global Ca2+ and constricted SMA. Iberiotoxin, a BK channel inhibitor, had no effect on either global Ca2+ or vascular diameter. However, iberiotoxin increased global Ca2+ and caused a vasoconstriction in the presence of apamin, an inhibitor of SK channels. In contrast to effects at 40 cmH2O, ryanodine did not constrict the SMA at 60 cmH2O. The pressure‐dependence of the ryanodine‐induced vasoconstriction was consistent with the pressure‐dependence of the Ca2+‐sensitivity, which was greater at 40 than at 60 cmH2O. Enhancement of the Ca2+‐sensitivity with 1 nM endothelin‐1 enabled ryanodine‐induced vasoconstriction at 60 cmH2O. These results suggest that BK channels are present but not activated by Ca2+‐sparks in the SMA. Instead, regulation of global Ca2+ and vascular diameter by BK channels is modulated by SK channels. Ryanodine‐induced vasoconstriction is regulated by the Ca2+ sensitivity and may be caused by store‐dependent mechanisms.

Supported by NIH‐R01‐DC04280.
Excessive vascular smooth muscle and macrophage proliferation underlies impaired coronary collateral growth in the metabolic syndrome
Authors: Hutcheson Rebecca Lynn , Chaplin Jennifer , Smith Erika , Jadhav Rashmi , Russell James C , Rocic Petra
Published: June 2021

Rationale

Coronary collateral growth (CCG) is impaired in the metabolic syndrome. It is thought that inadequate cell proliferation is a causative factor. Pro‐angiogenic growth factors (GFs) stimulate cell proliferation. Metabolic syndrome patients and animal models exhibit elevated GF levels, but administration of GFs does not promote CCG in the metabolic syndrome.

Objective

To determine whether temporally inappropriate excessive cell proliferation underlies impaired CCG in the metabolic syndrome.

Methods and Results

Normal (SD) and metabolic syndrome (JCR) rats underwent transient, repetitive coronary artery occlusion (RI). We have previously shown that CCG was maximal at day 9 of RI in SD rats but did not occur in JCR rats. Cell proliferation was evaluated by immunohistochemistry (PCNA, Ki‐67) and Western blotting (PCNA). The increase in cell proliferation was transient in SD but greater and sustained in JCR rats. Assessment of cell cycle progression confirmed early and transient cell proliferation in SD vs. sustained proliferation in JCR rats. This was associated with accumulation of proliferating cells in the lumen of small arterioles in JCR rats, which failed to undergo outward expansion.

Conclusions

Excessive cell proliferation in the later stages of collateral remodeling underlies impaired CCG in the metabolic syndrome. R01 HL093052
Blunted Osmopressor Response in Familial Dysautonomia
Authors: Goulding Niamh Elisha , Norcliffe‐Kaufmann Lucy J , Kaufmann Horacio
Published: June 2021

In patients with chronic autonomic failure (AF), drinking water increases mean blood pressure (MBP). This is an “osmopressor” response mediated by osmoreceptors on afferent neurons with their cell bodies in the dorsal root ganglia (DRG). Osmoreceptor activation triggers sympathetic excitation via a spinal reflex. Familial dysautonomia (FD) is a genetic disease that affects the development of primary afferent neurons with cell bodies in the DRG, but leaves efferent sympathetic neurons functionally intact. Our objective was to investigate whether this osmopressor response was present in FD. Beat‐to‐beat BP and RR intervals were recorded for 10 minutes prior and for 30 minutes after ingesting 500ml of water while supine. Thirty minutes after the water, MBP was significantly higher in the patients with AF than in those with FD (Δ15±5mmHg and Δ0±4mmHg respectively, P<0.01). Area under the curve also differed significantly between groups (191±25 and 387±55 mmHg/min, P<0.004). Blunting of the osmopressor response to water in patients with FD suggests that the congenital mutation affects osmosensory neurons.
Effect of Water Deprivation on KCC2 Expression in Hypothalamic Vasopressin Neurons in Rat
Authors: Knapp Blayne , Nedungadi T. Prashant , Cunningham J. Thomas
Published: June 2021

Water deprivation (WD) represents a physiological challenge that results in the sustained release of vasopressin (AVP) that is inhibited by water intake. We and others have previously shown that WD enhances glutamate signaling in AVP magnocellular neurosecretory cells (MNCs). In this study, we tested the hypothesis that WD has profound effects on GABA neurotransmission as well. One mechanism that could alter GABAergic neurotransmission in AVP MNCs is through the chloride extruder K+/Cl− cotransporter (KCC2). Changes in the expression and function of KCC2 could influence the intracellular concentration of Cl− and alter the synaptic inhibitory tone of MNCs located in the supraoptic (SON) and paraventricular nuclei (PVN) of the hypothalamus. Using laser capture microdissection (LCM),7–10 quick immunolabled AVP cells were harvested from the SON and PVN of euhydrated and 48 h WD adult male rats and RT‐PCR was used to test for changes in KCC2 message. We observed a significant elevation in KCC2 mRNA expression in AVP cells of the SON (WD 2.5 ± 0.52; Control 1.0 ± 0.06, p<0.05) but not in the PVN (WD 3.5 ± 1.7; Control 1.0 ± 0.1). Immunofluorescence demonstrated the colocalization of KCC2 and AVP in the SON. Increased expression of KCC2 could be associated with decreased intracellular Cl− in AVP neurons in the SON, thereby serving to maintain or enhance the inhibitory tone of AVP neurons in the SON but not the PVN during WD. HL62569
Leptin increases lumbar sympathetic nerve activity (LSNA) and its baroreflex regulation in female rats: variations with the estrus cycle and pregancy
Authors: Shi Zhigang , Li Baoxin , Brooks Virginia L.
Published: June 2021

Intracerebroventricular (ICV) leptin increases LSNA and LSNA baroreflex gain (BRG) in males; however, leptin's actions in females are unknown. Therefore, we determined if leptin increases LSNA in α‐chloralose anesthetized female SD rats during proestrus (n=6), diestrus (n=6), or late pregnancy (n=4). Baroreflex function was assessed at baseline, and 1 and 2 hr after ICV leptin (3 μg), using a 4 parameter sigmoidal fit of LSNA responses to slow ramp (3–5 min) changes in arterial pressure, induced by iv infusion of nitroprusside and phenylephrine. During proestrus, ICV leptin increased (P<0.05) basal LSNA [to 157±21 (1hr) and 197±18 (2hr) % control], LSNA reflex maximum [from 106±19 to 157±31 (1hr) and 205±48 (2hr) % control], LSNA baroreflex gain [from 2.2± 0.3 to 2.9±0.8 (1hr) and 3.9±0.5 (2hr) %/mmHg], and LSNA baroreflex range [from 106±19 to 157±31 (1hr) and 205±48 (2hr) % control]. In contrast, during diestrus and pregnancy, ICV leptin failed to significantly increase basal LSNA (diestrus, to 117±10 (1hr) and 130±13 (2hr) % control; pregnancy, to 114±16 (1hr) and 101±19 (2hr) % control] and baroreflex control of LSNA. In conclusion, leptin increases LSNA and its baroreflex regulation in females during proestrus, but not during diestrus. Pregnancy is also a leptin‐resistant state. Supported by HL088552 and AHA.
Rapid and slow effects of corticosterone on glutamatergic transmission in NTS
Authors: Fan Yun , Mifflin Steve
Published: June 2021

Rapid, non‐genomic and slower effects of CS on AMPA‐receptor– mediated transmission in NTS neurons were examined in rats using whole‐cell recordings of second‐order NTS neurons receiving arterial chemoreceptor afferent inputs in a brain slice. Measured at least 1 h after 20 min superfusion of the slice with 100 nM CS, CS increased miniature postsynaptic excitatory current (mEPSC) amplitude (control 15.6±1.1pA, n=12, CS 21.2±1.7pA, n=10, p<0.01) but not interval (control 0.77±0.13s, n=12, CS 0.99±0.08s, n=10, p>;0.05). During the 20min superfusion of the slice with 100nM CS, mEPSC amplitude measured during the control period prior to application of CS (15.6±1.1pA, n=5) was not different from that measured during the 4min to 9min following the onset of the CS superfusion (16.4±1.3pA, n=5) or 14min to 19min after the onset of the CS superfusion (16.48±0.83pA, n=5). There were no differences in mEPSC frequency during these time periods. PKC inhibitors chelerythrine choride 1uM or calphostin C 100nM applied 10 min before and during 20 min superfusion with 100 nM CS did not alter delayed CS‐induced increases in mEPSC amplitude. CS does not appear to have a rapid (<20 min) effect on glutamatergic transmission in NTS neurons receiving arterial chemoreceptor afferent inputs; however a 20 min application of CS increased mEPSC amplitude measured at least 1 hr later apparently not mediated by PKC systems. Supported by HL‐088052
Bradykinin B2 receptor contributes to exaggerated muscle mechanoreflex in rats with femoral artery occlusion
Authors: Lu Jian , Xing Jihong , Li Jianhua
Published: June 2021

In peripheral artery disease (PAD), amplified sympathetic nerve activity (SNA) and blood pressure (BP) responses to static exercise are observed. This dysfunction of the exercise pressor reflex is mediated, in part, by muscle mechanoreflex overactivity. Prior studies suggest that bradykinin B2 receptor mediates the reflex SNA and BP responses during the exercise pressor reflex. Thus, we determined if kinin B2 receptor contributes to the augmented mechanoreflex activity in rats with PAD induced by 24 hrs of femoral artery ligation. First, western blot analysis showed that protein expression of B2 displays significant over‐expression in dorsal root ganglion of ligated limbs as compared with control limbs (optical density: 0.94 ± 0.02 in control and 1.87 ± 0.08 after ligation, P < 0.05 vs. control; n = 6 in each group). Second, mechanoreflex was evoked by passive tendon stretch and the reflex renal SNA and BP responses to stretch were examined after arterial injection of HOE‐140 (a B2 receptors blocker) into the hindlimb. HOE‐140 had greater attenuating effects on SNA and BP in ligated rats. i.e. after 5 μg/kg of HOE‐140 renal SNA and BP responses evoked by 0.5 kg of muscle tension were attenuated by 43% and 25% in control vs. 54% and 34% in ligation (P < 0.05 vs. control group; n=11 in each group). In contrast, there is no significant difference in B1 expression in both experimental groups, and arterial injection of R‐715, a B1 receptors blocker, had no significant effects on SNA and BP responses evoked by muscle stretch. Accordingly, results obtained from this study support our hypothesis that heightened kinin B2 expression in the sensory nerves contributes to the exaggerated muscle mechanoreflex in PAD rats. (Supported by NIH P01 HL096570 & AHA EIA 0840130N)
Interactions between carotid body denervation and renal nerve denervation in lowering arterial blood pressure in the adult spontaneously hypertensive rat (SHR)
Authors: McBryde Fiona D , Abdala Ana PL , Hendy Emma B , Engelman Zoar , Fudim Marat , Paton Julian F R
Published: June 2021

We have previously shown that bilateral carotid body denervation (CBD) in SHR reduces arterial pressure (AP). How CBD interacts with the more established technique of renal denervation (RDNX) is unknown. Here we test the hypothesis that CBD will lower AP over and above the fall in pressure achieved with RDNX. In 12 week‐old SHR previously instrumented to record AP telemetrically, denervation surgery (unilateral CBD n=6; bilateral CBD n=7; Sham CBD n=6; RDNX n=5; Sham RDNX n=5) was performed on Day 0. On Day 21 after CBD or Day 14 after RDNX, the renal nerves or carotid bodies were denervated respectively. Chemoreflex elimination was confirmed by i.v. sodium cyanide (120ìg.kg‐1; Sham 15±3mmHg, CBD 1±2mmHg). By Day 21 after bilateral CBD, AP was reduced by −17±2 mmHg from a baseline of 160±5 mmHg (p<0.05); unilateral CBD/Sham rats showed the expected developmental increase in AP of 7±2mmHg/9±3mmHg. Subsequent RDNX reduced AP by a further −8±2mmHg in bilateral CBD rats and by −8±3mmHg in Sham rats (p<0.05). Conversely, in RDNX rats (−7±2mmHg), bilateral CBD produced a further fall in AP of −12±3 mmHg(p<0.05). In summary, unilateral CBD alone seems insufficient to reduce AP in SHR. Bilateral CBD and RDNX appear to have an occlusive interaction. This is important clinically as it suggests further reductions in AP may be obtained in hypertensive patients post renal denervation. Supported by the British Heart Foundation and Cibiem.
Angiotensin II contributes to microglial cell activation in the PVN of hypertensive rats
Authors: Biancardi Vinicia Campana , Stern Javier E
Published: June 2021

Accumulating evidence supports microglia‐mediated inflammatory pathways within the brain to contribute to cardiovascular disorders including hypertension. Yet, the precise mechanisms remain unknown. Here, we start to test the hypothesis of a pathophysiological link between angiotensin II (AngII) and microglial cells in the hypothalamic paraventricular nucleus (PVN) of hypertensive rats, via activation of toll like receptor 4 (TLR4). In a first series of immunohistochemical experiments, we found a significant increase in microglia (IBA1) as well as TLR4 immunoreactivity in SHR compared to WKY rats (71% and 106%). Importantly, a high degree of colocalization between both markers was observed in SHRs. In vitro experiments performed in acute hypothalamic slices showed that exposure of slices to AngII (1μM) significantly increased the density of activated microglial cells in the PVN of WKY rats (64%). Compared to WKYs, basal microglial cell density was significantly higher in SHRs (45%), in which AngII failed to further increase microglial staining. Ongoing experiments are being performed in C3H/HeJ mutant mice to determine the contribution of TLR4 to AngII‐mediated microglial activation in the PVN. Taken together, our results support the notion that AngII contributes to microglial cell activation in the PVN of SHRs, likely via activation and/or upregulation TLR4 receptors. Supported by RO1 HL 090948.
Involvement of nuclear factor‐kappa B in superoxide‐lowered protein expression of voltage‐gated sodium channels in nodose ganglia from heart failure rats
Authors: Tu Huiyin , Liu Jinxu , Zhang Dongze , Muelleman Robert , Li Yulong
Published: June 2021

Our previous study has shown that chronic heart failure (CHF) elevates mitochondria‐derived superoxide level and reduces protein expression of voltage‐gated sodium (Nav) channels in rat nodose neurons. In the present study, we investigated the involvement of mitochondrial superoxide and nuclear factor‐kappa B (NF‐kB) in CHF‐decreased Nav1.7 channel expression in rat nodose ganglia. CHF was induced by left coronary ligation. Adenoviral MnSOD (Ad.MnSOD) gene (4 × 1010 PFU/ml, 1 μl) or lentiviral NF‐kB (p65) shRNA (5 × 106 IFU/ml, 1 μl) was injected into the nodose ganglia. Mitochondrial superoxide was detected using mitochondria‐targeted superoxide sensitive flurogenic probe MitoSOX. MnSOD, NF‐kB (p65) and Nav1.7 expression were measured by western blot. NF‐κB binding to the promoter of Nav1.7 channels was detected with chromosome immunoprecipitation assay (ChIP). We found that: 1) HF decreased Nav1.7 expression, increased NF‐kB (p65) phosphorylation and NF‐kB (p65) binding with Nav1.7 promoter; 2) NF‐kB (p65) shRNA reduced total and phosphorylated NF‐kB (p65) and increased Nav1.7 expression in CHF nodese neurons; 3) Ad.MnSOD transfection elevated MnSOD protein, reduced the elevation of mitochondrial superoxide level and phosphorylated NF‐kB (p65), and increased Nav1.7 expression in the nodose ganglia from CHF rat. These results indicate that mitochondrial superoxide overproduction triggered NF‐kB activation and subsequently depressed Nav1.7 expression in CHF nodose neurons.
Exercise training and hypocaloric diet improves sympathetic arterial baroreflex control in patients with metabolic syndrome and obstructive sleep apnea
Authors: Toschi‐Dias Edgar , Trombetta Ivani C , Silva Valdo J D , Maki‐Nunes Cristiane , Cepeda Felipe X , Alves Maria Janieire N N , Carvalho Glauce , Drager Luciano , Lorenzi‐Filho Geraldo , Negrao Carlos E , Rondon Maria Urbana P B
Published: June 2021

Metabolic syndrome (MetS) decreases arterial baroreflex control of muscle sympathetic nerve activity (ABRMSNA). And, the obstructive sleep apnea (OSA) exacerbates this autonomic dysfunction. We tested the hypothesis that exercise training and hypocaloric diet (ET+D) would restore ABRMSNA in patients with MetS and OSA. Forty‐four MetS patients were allocated into four groups: Sedentary with OSA (MetS+OSA Sed, n=10) or without OSA (MetS‐OSA Sed, n=10), and ET+D with OSA (MetS+OSA ET+D, n=11) or without OSA (MetS‐OSA ET+D, n=13). The ET+D groups were submitted to −500 kcal/day and 40 min bicycle exercise for 4‐months. OSA was deferred as apnea‐hypopnea index>;15 events/hour. MSNA (microneurography), blood pressure (beat‐to‐beat, noninvasive) and spontaneous ABRMSNA (gain, sensitivity and time delay, latency) were evaluated. ET+D decreased MSNA (P<0.05) and increased ABRMSNA gain in both MetS+OSA (13±1 vs. 24±2 ms/mmHg, P=0.01) and MetS‐OSA (27±3 vs. 37±3ms/mmHg, P=0.03) groups. The time delay of ABRMSNA was reduced by ET+D only in MetS+OSA group (4.1±0.2 vs. 2.8±0.3 s, P=0.04). No changes were observed in the sedentary groups. In conclusion, the ET+D improve ABRMSNA sensitivity in patients with MetS regardless of OSA. However, this effect of ET+D is more pronounced in patients with MetS+OSA.

Support: CNPq and FAPESP#2011/17533–6
Effect of diet on the progression of diabetic nephropathy in db/db mice
Authors: Kandlikar Sachin S , Joly Kristin M , Horn Todd R , Glascock Chris G , Plato Craig F
Published: June 2021

Diabetic nephropathy (DN) is the leading cause of end stage renal disease. Severity of diabetes and its complications are influenced by dietary intake while reduced renal mass (uninephrectomy, UNX) accelerates progression and worsens severity of DN. We hypothesized that high‐energy intake (HE) exacerbates the progression of DN in db/db mice. We studied long‐term interaction of HE and UNX on DN in db/db mice. Baseline water intake and urine volume were measured and assayed in male db/db and control mice (7–8 wks age). Depending on the diet [Purina 5008 (HE) or Harlan 8640 (normal chow, NC)] and surgical treatment (Sham or UNX), db/db mice were then functionally matched (10 wks age) into following five groups: 1) Control+NC, 2) Sham+NC, 3) UNX+NC, 4) Sham+HE, and 5) UNX+HE. Urine and blood samples, collected every 2 weeks, showed significant elevation in fasting blood glucose, HbA1c, and serum creatinine in all db/db compared to controls while urine creatinine was increased only in animals on HE. Water intake and urine excretion increased with age and were significantly higher in all db/db groups compared to controls. The fold change in 24‐hour urine albumin excretion compared to controls at 26 wks were 21.9 ± 8.6, 20.7 ± 5.4, 45.0 ± 7.5, and 60.2 ± 14.8 in groups 2, 3, 4, and 5 respectively. We conclude that HE combined with UNX exacerbates progression and severity of DN as indexed by albuminuria in db/db mice.
RAS Blockade Inhibits Bcl‐2 Modifying Factor (Bmf) Expression and Prevents Hypertension and Tubular Apoptosis in Diabetic Akita Mouse Kidneys
Authors: Maachi Hasna , Lo Chao‐Sheng , Abdo Shaaban , Chenier Isabelle , Zhang Shao‐Ling , Chan John S D
Published: June 2021

Background

We reported that pro‐apoptotic protein Bmf is highly expressed in diabetic mouse renal proximal tubular cells (RPTCs) and induces RPTC apoptosis in vitro (Diabetes 2012). The regulation of Bmf expression in RPTCs, however, is not well understood. We investigated whether renin‐angiotensin system (RAS) blockade could prevent Bmf expression, hypertension and RPTC apoptosis in Akita mice (a model of type 1 diabetes) in vivo and in vitro.

Methods

Adult male Akita mice (11 weeks) were treated ± RAS blockers (losartan, 30 mg.Kg−1.day−1 and perindopril, 4 mg.Kg− 1.day−1 in drinking water) until 16 weeks. Controls were untreated non‐Akita littermates. Plasma glucose, systolic blood pressure (SBP) and albuminuria were monitored weekly. Kidneys were processed for histology and apoptosis studies (TUNEL assay). RPTC gene and protein expression were quantified by real time‐qPCR and Western blotting, respectively. We also examined the effect of angiotensin II (Ang II) on Bmf gene expression in cultured rat immortalized RPTCs.

Results

Akita mice developed hyperglycemia and significantly higher SBPs, kidney/body weight ratios and albuminuria, cf. to non‐Akita littermates. Kidneys from Akita mice displayed renal hypertrophy and tubular apoptosis cf. to non‐Akita littermates. Increased expression of Agt, Bmf and active caspase‐3 were evident in RPTCs of Akita mice. Treatment with RAS blockers inhibited Agt and Bmf expression, prevented hypertension and albuminuria and attenuated tubular apoptosis in Akita mice. Finally, Ang II stimulated Bmf mRNA expression in RPTCs and its effect was abolished in the presence of losartan.

Conclusion

Ang II up‐regulates Bmf expression and RAS blockade effectively inhibits Bmf expression and prevents hypertension and tubular apoptosis in diabetic kidneys.

Supported by Canadian Institutes of Health Research.
H2S Therapy Improves MMP‐9 and NMDA Receptor Mediated Diabetic Renovascular Remodeling
Authors: Kundu Sourav , Pushpakumar Sathnur , Tyagi Suresh C , Sen Utpal
Published: June 2021

Previously we reported that matrix metalloproteinase‐9 (MMP‐9) plays an important role in extracellular matrix (ECM) remodeling in diabetic kidney due to decreased H2S production by downregulation of CBS/CSE. Altered ECM resulted from NMDA‐R induction and dysregulation of connexins (Cxs). In this study, we determined whether H2S treatment reduces MMP‐9 induced ECM remodeling in diabetic nephropathy. Wild type (C57BL/6J), diabetic (Akita, C57BL/6J‐Ins2Akita), MMP‐9 knockout (MMP‐9−/−, M9KO) and double KO of Akita/MMP‐9−/− (DKO) mice were treated without or with 0.05 g/L of NaHS in drinking water. Cortex blood flow, renal function, vascularity, MMP‐9, NMDA‐R, Cxs, collagen and elastin were measured. Diabetic mice showed decreased cortical blood flow and vascularity which improved with H2S treatment. DKO mice showed improved blood flow with H2S treatment. Akita and DKO mice showed low levels of CBS/CSE and thus H2S production. The levels of MMP‐9, NMDA‐R and Cxs‐40 and ‐43 were increased in Akita which normalized with H2S supplementation. DKO mice showed a decreasing trend for NMDA‐R and Cxs but the levels normalized only after H2S treatment. We conclude that H2S treatment reduces adverse diabetic renal remodeling by downregulation of MMP‐9 and NMDA‐R.
Differential roles for tissue‐infiltrating T cells during angiotensin II‐induced hypertension
Authors: Vinh Antony , Wei Zihui , Diep Henry , Welungoda Iresha , Widdop Robert E
Published: June 2021

Hypertension is associated with elevated T cell infiltration into blood pressure‐controlling organs, such as the aorta and kidney. Since the function of these infiltrates remains speculative and undefined, the current study investigated the relative T cell derived‐cytokine and ROS production between hypertensive and normotensive mice. Splenic, blood, aortic, kidney and brain T cells were isolated from C57BL/6J mice following 14‐day vehicle or angiotensin (Ang) II (490ng/kg/min, s.c.) infusion. Cytokine analysis in isolated tissue‐infiltrating T cells indicated an overall Th1 pro‐inflammatory phenotype with a similar proportion (flow cytometry) and quantity (cytometric bead array) of IFN‐γ and TNF‐α released from T cells in vehicle‐ and Ang II‐treated groups. Strikingly, elevated T cell‐derived production of a chemokine, CCL2, was observed in aorta (~7‐fold), kidney and brain in response to Ang II (n=6–8), but not in spleen or blood. Moreover, T cell‐derived ROS production in kidney and brain was elevated (~2‐fold and ~3‐fold) in Ang II‐treated mice (n = 6–8; P <0.01). In conclusion, while Ang II‐induced hypertension does not modulate the overall infiltrating‐T cell cytokine profile, elevated T cell‐derived ROS production and/or leukocyte recruitment due to increased CCL2 may represent the functional contribution of tissue‐infiltrating T cells during hypertension.
Exercise‐induced electrical remodeling of endocardial and epicardial tissue in perfused rat hearts
Authors: Mark Laura , Nelson Cassandra , Fitts Robert H
Published: June 2021

Exercise‐training (ET) prolongs the epicardial monophasic action potential (MAP) of female rats at rest but changes at higher heart rates (HR), and gender differences are unknown. To assess this, female and male Sprague‐Dawley rats were randomly assigned to either voluntary wheel running (TRN) or control group (SED). After six weeks, rats were anesthetized, hearts excised, and cannulated on a Langendorff apparatus. MAP's were recorded using 0.25 mm chlorided silver wires from base and apex, areas representative of ENDO and EPI tissue, respectively. The sinoatrial node was stimulated at frequencies from 200 to 400 BPM. Initial spike duration (SD) and total action potential duration (APD) were measured. SED males and females had comparable SD and APD at all HR except 400 BPM, where ENDO APD was shorter in females (p<0.05). The SD and APD were unaltered by ET in males, while in females the ENDO APD at 350 and 400 BPM was significantly shortened (p<0.05). Further, TRN females showed a prolonged EPI SD at 400 BPM (p<0.05). Results suggest that the cardiac electrical remodeling seen with exercise training is gender‐specific and dependent on HR. Supported by Marquette University Way Klinger Fellowship.
Do exercise and estrogens interact to alter stress responses in rats?
Authors: Jones Alexis Brooke , Gupton Rebecca , Curtis Kathleen S
Published: June 2021

The beneficial effects of exercise in coping with stress/anxiety may be particularly important for women, as anxiety and depression increase after menopause. The present experiments examined anxiety‐like behaviors and corticosterone (CORT) levels induced by acute stress in female rats with or without estradiol benzoate (EB) after a single bout of exercise vs. 3 days of exercise before and after stress.

Adult female Sprague Dawley rats were ovariectomized, recovered for 7 days, given EB or OIL for 2 consecutive days, and tested during the next 2 days. In Expt. 1, rats were stressed by being placed in a plastic restrainer for 40 min, then were returned to a novel cage with a running wheel attached. Software recorded distance, speed, and time spent on wheels for 14 hrs. The next morning, anxiety‐like behaviors were assessed using the Elevated Plus Maze (EPM). In Expt. 2, exercising rats had access to running wheels beginning on day 1 of EB/OIL treatment and ran throughout the experiment, including the 14 hrs after restraint stress.

Exercise prior to restraint stress reduced CORT levels in EB‐treated rats compared to rats that ran after stress. However, 3 days of exercise increased anxiety‐like behaviors in both groups compared to rats exercised for only 1 day, with or without restraint stress.

Thus, exercise duration affects stress hormones in EB‐treated rats, but CORT levels may not accurately predict anxiety‐like behaviors.
Intermittent respiratory stimulation with doxapram induces phrenic motor plasticity
Authors: Sandhu Milap Singh , Lee Kun‐Ze , Reier Paul J , Fuller David D
Published: June 2021

Intermittent hypoxia can induce a persistent increase in phrenic motor output via a central neural mechanism. Here we explored a pharmacological alterative to hypoxia as a tool for triggering phrenic motor plasticity. Specifically, we hypothesized that persistent increases in efferent phrenic burst activity can be evoked via systemic delivery of a low‐dose of doxapram – a drug which can robustly stimulate phrenic activity due to activation of carotid chemoafferents. Anesthetized, vagotomized, paralyzed and mechanically‐ventilated rats were given intravenous doxapram either intermittently (3 × 2mg/kg) or as a single dose (2mg/kg or 6mg/kg). A persistent increase in phrenic burst amplitude was evoked by intermittent doxapram (161 ± 27% baseline at 60 min post‐injection, p<0.05 vs. sham injection and time control), but not by the single dose (104 ± 13% baseline). The acute phrenic response was substantially reduced in carotid‐denervated rats (120 ± 3% baseline compared to 165 ± 9% baseline in animals with intact carotid nerves, p<0.05) confirming that doxapram acts primarily via the carotid chemoreceptors. Since doxapram is an FDA approved drug and commonly used as a respiratory stimulant, it may be a viable alternative or supplement to hypoxia for targeted induction of respiratory neuroplasticity in neurological conditions such as spinal cord injury.

Support: Craig H. Neilsen Foundation #220521 (MSS), 1R01NS080180–01A1 (DDF), NIH1R21HL104294–01 (DDF)
Ouabain increases association between Na‐K ATPase (Na‐K) and NHE1 through N‐terminal domain of Na‐K
Authors: Khundmiri Syed J , Xie Zijian , Lederer Eleanor D
Published: June 2021

Low dose ouabain increases Na‐K activity in kidney basolateral membranes in an NHE1 dependent manner. TIRF and FRET microscopy suggested that 10 pM ouabain increases association between Na‐K and NHE1. The aim of the study was to identify the molecular domains on Na‐K and NHE1 required for association. Using mouse (MRPT) and human (HK11) kidney proximal tubule cells co‐transfected with full‐length GFP or RFP‐NHE1, and full length mCherry‐Na‐K a‐subunit or N‐terminal deletion mutant of YFP‐Na‐K, FRET and TIRFM were performed before and after ouabain treatment. We also measured the effect of ouabain on Na‐K activity in HK2 cells transfected with WT or scaffold domain mutated NHE1. Ouabain increased association between Na‐K and NHE1. NHE1 and Na‐K association was abolished in cells expressing N‐terminal deletion mutant Na‐K. Ouabain increased Na‐K activity in HK2 cells expressing full length NHE1 but not in cells expressing scaffold domain mutated NHE1. These data suggest that low concentrations of ouabain increase Na‐K association with NHE1 and this association involves N‐terminal domain of Na‐K and the scaffolding domain of NHE1.

Supported by AHA and VA
Quantification of tendon cell nuclei in rat Achilles tendon after 12 weeks of running
Authors: Couppé Christian , Thomsen Emilie Wøjdemann , Svensson René , Heinemeier Katja , Kongsgaard Mads , Kjaer Michael , Magnusson S. Peter , Scherling Peter
Published: June 2021

Introduction

It remains unknown why tendons improve mechanical properties with loading. However, tendon cell nuclei (TCN) have been claimed to induce these mechanical adaptations, although this has not been previously investigated.

Purpose

To quantify the number of TCN in rat Achilles tendon after 12 wk of running.

Methods

8 adult male Sprague‐Dawley rats ran (RR) on a treadmill with 10° incline, 1 h/day, 5 days/wk (17–20 m/min) for 12 wk (which led to improved tendon mechanical properties) and were compared with 11 control rats (CR). Tissue‐Tek embedded serial sections (10 μm) from the mid region of the Achilles tendon were cut longitudinally on a cryostat. Sections were stained with PicroSirius Red and hematoxylin staining. One investigator counted the number of TCN 2–3 times in three separate parts of the mid longitudinal tendon section with fields of 390 μm ×280 μm in a blinded fashion. Unpaired t‐test was used for the statistical analysis (Mean±SE).

Results

Typical Error % for the three parts was 5.3%: There was no difference between running rats vs. control rats (TCN per image (≈ 105μm): RR, 152±9; CR, 146±8, P=0.642).

Conclusion

This new method provided reproducible determination of tendon cell nuclei density. However, even though the tendon mechanical properties were improved, no difference in tendon cell nuclei density between running rats vs. control rats could be detected. Research support: Danish Association of Rheumatism.
Function and molecular regulation of WNT‐5A expression by TGF‐β
Authors: Kumawat Kuldeep , Menzen Mark H , Halayko Andrew J , Schmidt Martina , Gosens Reinoud
Published: June 2021

Disruption of WNT‐5A homeostasis has been linked to various disorders like fibrosis, cancer and inflammation with recent evidence implicating it in asthma. Here, we have investigated the function and molecular regulation of WNT‐5A expression in airway smooth muscle (ASM) cells by TGF‐β.

Immortalized human ASM cells were used to measure mRNA by qRT‐PCR and protein abundance by immunoblotting.

ASM cells expressed a variety of WNT ligands and Frizzled receptors which were differentially modulated by TGF‐β. WNT‐5A was the most abundant ligand expressed and showed ~4–16 fold upregulation in response to TGF‐β (2 ng/mL). Knockdown using siRNA demonstrated WNT‐5A requirement for TGF‐β‐induced collagen and fibronectin expression. TGF‐β induction of WNT‐5A appeared to require TGF‐β activated kinase (TAK)‐1 activity, as pharmacological inhibition of TAK‐1 with LL‐Z1640–2 (500 nM) prevented WNT5A expression. Interestingly, inhibition of ERK1/2 (U0126, 3μM), Smad3 (SIS3, 3μM) and β‐catenin/TCF (PKF115–584, 100 nM) augmented WNT‐5A expression suggesting a negative regulatory role.

In conclusion, TGF‐β‐induced WNT‐5A expression mediates extracellular matrix expression by human ASM cells. WNT5A induction occurs via complex signaling cascades with TAK‐1 acting as a positive regulator, whereas ERK1/2, Smad3 and β‐catenin/TCF mediate a negative constraint.

This study is funded by University of Groningen.
Small litter size at birth is associated with insulin resistance and low energy expenditure in adulthood
Authors: Tavares Neuziane Kloos Amorim , de Jesus Aguiar Fernanda , Furukawa Luzia Naôko , Heimann Joel Claudio
Published: June 2021

Objective

To evaluate the effect of litter size standardization at birth on adult metabolic parameters.

Methods

Female Wistar rats became pregnant at 12 weeks of age. Offspring were divided into control(C), G8(8) and G4(4 newborns/litter) groups. At 12 weeks of age, food intake (FI,g/day), glucose (G,mg/dL), insulin (I,pM), cholesterol (Chol,mg/dL), triacylglycerols (TAG,mg/dL) levels, white and brown adipose tissue (BAT) masses (g/100g) adiposity index (AI,% body weight) were determined.

Results

(mean±SEM p<0.05, n=6 to 32):

Male
C
G8
G4

AI
4±0.1
4±0.1
5±0.51

FI
26±0.7
24±0.9
17±21

BAT
0.13±0.01
0.11±0.003
0.09±0.011

G
109±3
100±3
120±41

I
129±13
126±10
203±361

Chol
81±32
66±3
69±5

TAG
50±4
67±53
67±43

BP
127±44
101±6
121±124

p<0.05 vs C and G8;

p<0.05 vs G8 and G4;

p<0.05 vs C;

p<0.05 vs G8

Female
C
G8
G4

AI
5±0.3
5±0.3
7±0.71

FI
17±0.82
15±0.8
16.6±0.92

BAT
0.16±0.022
0.13±0.007
0.11±0.0051

G
92±42
89±2
104±41

I
50±62
71±9
106±161

Chol
57±3
62±4
66±53

TAG
46±5
63±53
65±73

BP
125±74
99±6
106±82

p<0.05 vs C and G8;

p<0.05 vs G8;

p<0.05 vs C;

p<0.05 vs G8 and SG4

Conclusion

Results suggest that changes in litter size is associated with energy balance, carbohydrate and lipid disorders, insulin resistance and alterations in blood pressure in adult male and female.

Supported by FAPESP
The ontogeny of GnIH and KISS‐1 in the female rat hypothalamus
Authors: Maze TD , Lovvorn Jennifer , Smith Tara
Published: June 2021

The effects of kisspeptin on reproductive development have been well‐documented in the past decade. More recently, a mammalian orthologue of Gonadotrophin Inhibitory Hormone (GnIH), known as RF‐amide‐related‐peptide‐3 (RFRP3), has also been found to affect reproductive function. It has been proposed that GnIH antagonizes the effect of KISS‐1that upregulates the reproductive system during puberty. In the current study, differential expression of kisspeptin and RFRP3 were examined in the hypothalamic of female rats at various stages of postnatal development. Hypothalamic samples from rats at days 10, 20, 30, 40, 50, and 60 were examined for gene expression of GnRH, GnIH, KISS‐1 and the receptors GPR147 and GPR54. GnRH was expressed very early and its expression was maintained throughout the experiment. GnIH and KISS‐1 expression was greatest on day 30 and then declined during the transition into puberty. These data suggest that perhaps GnIH is up‐regulated once the hypothalamus has fully developed and is prepared for reproduction but is inhibiting reproductive function until other signals from the reproductive axis provide feedback that oogenesis is also complete. This research was supported by the Lander University Foundation Grant.
Targeted expression of SLC26A4 rescues hearing and balance in Slc26a4Δ/Δ mice
Authors: Wangemann Philine , Li Xiangming , Sanneman Joel , Zhou Fei , Harbidge Donald G. , Nelson Raoul D. , Chambrey Régine , Eladari Dominique , Miesner Tracy , Marcus Daniel C.
Published: June 2021

Mutations of SLC26A4 are a common cause for deafness associated with enlarged vestibular aqueduct. SLC26A4 encodes pendrin, an anion exchanger expressed in a variety of epithelial cells in the cochlea and the vestibular labyrinth. Slc26a4Δ/Δ mice develop a severe enlargement of the membranous labyrinth, fail to acquire hearing and balance, and thereby provide a stark model for the human phenotype. Here we generated a transgenic mouse line that expresses SLC26A4 under the promoter of ATP6V1B1. Crossing this transgene into the Slc26a4Δ/Δ line restored the temporal and spatial expression pattern of pendrin in the endolymphatic sac without inducing expression in the cochlea or the vestibular sensory organs. The transgene prevented enlargement and restored normal hearing and balance. Our study demonstrates that a limited restoration of pendrin is sufficient to restore normal inner ear function. These data in conjunction with notion that pendrin is required for the development but not the maintenance of hearing (Choi et al., 2011) opens the prospect that a spatially and temporally limited therapy may restore a life‐time of normal hearing in Slc26a4Δ/Δ mice, and possibly humans with SLC26A4 mutations.

Supported by NIH‐R01‐DC012151, NIH‐P20‐RR 17686, KINBRE, Kansas State University, ANR BLANC 2010‐R10164DD
PULSE Partnerships for Change: Moving from “Vision” to “Change” in Undergraduate Life Science Education
Authors: Schlegel Whitney M , McFarland Jenny L , Marley Katherine E
Published: June 2021

What will it take to bring about the necessary transformation of STEM higher education described in Vision and Change (2011)? PULSE (Partnership for Undergraduate Life Sciences Education) is a collaborative effort by National Science Foundation (NSF), National Institutes of Health (NIH) and Howard Hughes Medical Institute (HHMI) to stimulate systemic change within biology departments at all types of post‐secondary educational institutions, based upon the 2011 report Vision and Change in Undergraduate Biology Education and other calls for transformation of life sciences education. As life science educators we understand the change mandate (the problem). We have sought the evidence to support change (the solutions). We now must implement and institutionalize, specifically at the departmental level, these evidence‐based best practices. The PULSE Leadership Fellows are tasked with facilitating partnerships to foster change in undergraduate life science education. This poster will share the PULSE action agenda and our work to date, enhance communication with the broader scientific community and engage the American Physiological Society (APS) in the PULSE community as we cultivate change.
Practical Approach to Molecular Biology (PAMB): Sequential Experiments to Teach Basic Laboratory Skills
Authors: Appleyard Caroline B , Rivera Elizabeth , Sanchez Maria del C , Torres Lillith , Rivera‐Amill Vanessa
Published: June 2021

Students pursuing graduate degrees in the biomedical sciences require specific laboratory skills. We designed a course to teach these skills, ensuring uniformity and faster adaption in their education.

Aim

To test whether the course improved the technical ability and theoretical knowledge of first‐year graduate students.

Methods

The experiments were done sequentially—the results of each experiment leading to the next—and supplemented with lectures. Students learned about working safely in a laboratory, preparing solutions, keeping a laboratory notebook, and preparing a laboratory report, among other things; journal club discussion sessions and presentations were also included.

Results

Pre– and post– self‐evaluations showed that students felt that the course addressed various deficiencies and enhanced their skills, an assessment supported by the course coordinator and supporting personnel, all of whom observed and subsequently evaluated student performance.

Conclusions

The PAMB course provides us with a tool to help incoming students better adapt to the PhD program, thereby improving the laboratory experience. This course generates data suitable for a written report, thus reinforcing presentation and scientific writing skills. We expect that this course will enhance the productivity and increase the success of the students in our PhD program. R25GM082406 & 5G12RR003050/8G12MD007579
Evolution of Phenotypic Responses Characterizing Fetal Alcohol Spectrum Disorders (FASD) Among Vertebrate Model Organisms
Authors: Meers Mason B. , Chinea Angel
Published: June 2021

Model organism research on fetal alcohol spectrum disorders (FASD) crosses a diverse array of taxa without regard to evolutionary relationships. Responses to fetal alcohol exposure can consequently be expected to range widely as the affected genotypes represent taxa divergent for about 600 million years. In this study, we examine the symptoms of fetal alcohol exposure in a phylogenetic, evolutionary context known as character optimization or sometimes, character mapping. Our results indicate that of the 112 reported FASD symptoms, many neurological and craniofacial manifestations appear to characterize Mammalia, though significant autapomorphic responses may characterize humans. Still other symptoms seem to characterize Vertebrata and Animalia. Non‐neurological and craniofacial characters are less clearly congruent with phylogeny, suggesting that these responses to fetal alcohol exposure may be convergent in nature, lacking underlying anatomical bases. While this research suggests limitations on the utility of some taxa as model organisms, it also provides opportunity to define homologous responses to disease processes, as well as provides an evolutionary rationale for maximizing ethical decisions in the use and development of animal models in biomedical research.