bluuebunny/crossref_metadata_2025 huggingface dataset:
IGF‐1 deficiency impairs cerebral myogenic autoregulation in hypertensive mice (1079.2)
Authors: Toth Peter , Tucsek Zsuzsanna , Tarantini Stefano , Sosnowska Danuta , Gautam Tripti , Mitschelen Matthew , Koller Akos , Sonntag William , Csiszar Anna , Ungvari Zoltan
Published: June 2021

Aging impairs autoregulatory protection in the brain, exacerbating hypertension‐induced cerebromicrovascular injury, neuroinflammation and development of vascular cognitive impairment (VCI). Despite the paramount importance of the age‐related decline in circulating insulin‐like growth factor‐1 (IGF‐1) levels in brain and cerebrovascular aging, it is not well understood how IGF‐1 deficiency affects the functional adaptation of cerebral arteries to high blood pressure. To test the hypothesis that IGF‐1 deficiency impairs autoregulatory protection, hypertension was induced in control and IGF‐1 deficient mice (AAV knockdown of hepatic IGF‐1 [Igf1f/f + MUP‐iCre‐AAV8]) by chronic infusion of angiotensin II and changes in autoregulation of cerebral blood flow andmyogenic constriction of middle cerebral arteries (MCA) were assessed. In hypertensive control mice, the range of cerebral blood flow autoregulation was extended to higher pressure values and the pressure‐induced tone of MCA was increased. In hypertensive IGF‐1 deficient mice autoregulation was markedly disrupted, and MCAs did not show adaptive increases in myogenic tone. In control mice the mechanism of adaptation to hypertension involved up‐regulation of a TRPC channel‐dependent pathway and this mechanism was impaired in hypertensive IGF‐1 deficient mice. Downstream consequences of cerebrovascular autoregulatory dysfunction in hypertensive IGF‐1 deficient mice included exacerbated disruption of the blood‐brain barrier and neuroinflammation (microglia activation, up‐regulation of pro‐inflammatory cytokines and chemokines), which were associated with impaired hippocampal cognitive function. Collectively, IGF‐1 deficiency impairs autoregulatory protection in the brain of mice with angiotensin II‐induced hypertension, potentially exacerbating cerebromicrovascular injury and neuroinflammation mimicking the aging phenotype.

Grant Funding Source: Supported by the Nemzeti Fejlesztési Ügynökség and NIH
Cardiac myofilament Caa2+ sensitization disturbs regulation of coronary flow and leads to focal myocardial ischemia and arrhythmias during stress (1079.25)
Authors: Huke Sabine , Bennuri Sirish , Faria Thais , Knollmann Bjorn
Published: June 2021

Myofilament Ca2+ sensitization has been linked to increased arrhythmia susceptibility, but the underlying mechanism is not completely understood. Mice with the Ca2+ sensitizing TnT‐I79N mutation develop arrhythmias during stress, despite the absence of an overt anatomical substrate. Stress by fast pacing of isolated perfused TnT‐I79N hearts for 16 min elicited arrhythmias (63%) and caused the formation of hypoxic regions (14.1% of total tissue in I79N vs 2.6% in control, Hypoxyprobe) associated with focal cellular energy depletion (Connexin isoforms P0 and pT172‐AMP kinase both increased). The cause for the local energetic supply‐and‐demand mismatch is unclear and we tested if insufficient perfusion is at least in part responsible. Global coronary flow was not decreased in isolated TnT‐I79N hearts. To test if flow distribution was altered, we infused fluorescent 15 µm microspheres and compared the fraction of microspheres inside hypoxic areas with the fraction of hypoxic tissue. Microspheres inside hypoxic regions were drastically reduced, suggesting regional ischemia. These results were reproduced in control hearts treated with the Ca2+ sensitizing drug EMD57033. Next we tested if hypoxia also occurs naturally in TnT‐I79N mice in vivo. Despite expecting low sensitivity due to localized effects, the mRNA expression levels of genes up‐regulated by hypoxia (GLUT1, TUBB3) were increased (0.3fold, 3fold).

We conclude that myofilament Ca2+ sensitization disturbs regulation of coronary flow and leads to focal ischemia and arrhythmias during stress. This suggests that myofilament properties of cardiomyocytes importantly contribute to local regulation of coronary flow.

Grant Funding Source: supported by AHA 10SDG2640109 and NIH HL71670
Qualitative differences in reflex vagal control do exist controlling regional coronary flow resistance within and between mammalian species (1079.9)
Authors: White Saxon , Hamut Mutalip , Cottee David , Quail Anthony
Published: June 2021

The literature extensively documents inconsistencies in the vagal and acetylcholine control of coronary blood vessels between species. The reasons may relate to variation in experimental models, but reflex vagal control of coronary flow in man must remain uncertain. In awake canine species with paced hearts baroreflexes activate the vagus to dilate flow resistance vessels in the circumflex (CX) and right (R) coronary beds but have no apparent effect in the anterior descending (AD) bed. The effects therefore vary within one species. Seeking a unifying hypothesis of neural coronary control in 2010 we challenged awake sheep with identical protocols to those used in the dog, where, as in dog R serves only RV. We reported1 that the pattern of regional change between sheep and dog are similar for CX and AD, but the effects for R are opposite. In sheep as aortic perfusion pressure rises 28 mmHg, R flow does not change and the bed constricts (conductance 82%; P<0.001, c.f. dog dilates 118%; P<0.01). The effects in both species are blocked by methscopolamine. These data suggest that evolution of vagal control of the coronary circulation does vary qualitatively within, and, between species. The data also imply that the myocardium known for poor oxygen reserve at rest when facing rapid changes in cardiac work normally calls on beat‐by‐beat, rapid time‐constant hypoxia defence mechanisms. Emerging data2 suggest that myocardial cells may be normally hypoxia‐adapted, using low‐threshold cellular interacting glycolytic and hypoxia‐inducible transcriptional factors. Finally, the data imply a differential pattern of reflex vagal coronary dilator/constrictor effects will apply regionally to the heart of man. 1. Quail A et al, FASEB J. A1034.6, 2010 2. Songyue Yin et al, PLoS ONE 8(9):e74011, 2013

Grant Funding Source: Supported by project grants from NHMRC and HMRI Australia
Histone modification and exchange during gene transcription: signals and mechanisms (108.1)
Authors: Workman Jerry , Venkatesh Swaminathan , Smolle Mihaela
Published: June 2021

Maintenance of ordered chromatin structure over the body of genes is vital for the regulation of transcription. Increased access to the underlying DNA sequence results in the recruitment of RNA polymerase II to inappropriate, promoter‐like sites within genes, resulting in unfettered transcription. Our recent studies show how Set2‐mediated methylation of histone H3 on Lys36 (H3K36me) maintains chromatin structure by limiting histone dynamics over gene bodies, either by recruiting chromatin remodelers that preserve ordered nucleosomal distribution or by lowering the binding affinity of histone chaperones for histones, preventing their removal.
Adaptation to chronic hypoxia improves cardiac ischemic tolerance in spontaneously hypertensive rats (1080.3)
Authors: Neckar Jan , Brabcova Iveta , Weissova Romana , Zajickova Pavlina , Mandikova Petra , Silhavy Jan , Zurmanova Jitka , Novakova Olga , Kalous Martin , Pravenec Michal , Kolar Frantisek
Published: June 2021

The effect of adaptation to chronic continuous hypoxia (CCH) on cardiac ischemic tolerance and the expression of selected mitochondrial proteins, antioxidative enzymes and signalling molecules were analysed in left ventricles of spontaneously hypertensive rats (SHR) and in a novel conplastic strain SHR‐mtBN. These animals are characterized by a selective replacement of the mitochondrial genome of SHR with that of Brown Norway strain, which is more tolerant to ischemic injury. Rats were kept 21 days at CCH (inspired O2 fraction 0.1). CCH reduced the infarct size induced by coronary occlusion from 71±4% of the area at risk in normoxic SHR to 49±6 % in hypoxic SHR. It was accompanied by decreased protein levels of complex IV, cytosolic superoxide dismutase and mRNA levels of thioredoxin enzymes. On the other hand, mRNA levels of catalase, hexokinase 2 (HK2) and hypoxia‐inducible factor 1α (HIF‐1α) were increased in hypoxic SHR. As compared to SHR, CCH induced more pronounced infarct size‐limiting effect (by 19 %), elevated HK2 and HIF‐1α mRNA levels (by 36 and 25 %) and reduced complex IV proteins (by 13 %) in SHR‐mtBN. Overall, we demonstrate that CCH protects SHR hearts against acute ischemia/reperfusion injury and that mitochondrial genes can contribute to this protective effect.
Combination therapy of vagal nerve stimulation and left ventricular assist device maximally reduces the infarct size in ischemia reperfusion model (1080.4)
Authors: Kakino Takamori , Saku Keita , Sakamoto Takafumi , Akashi Takuya , Arimura Takahiro , Nishizaki Akiko , Oga Yasuhiro , Kishi Takuya , Ide Tomomi , Sunagawa Kenji
Published: June 2021

BACKGROUNDS: Despite the spread of early coronary reperfusion in the management of acute myocardial infarction (AMI), it remains difficult to fully salvage myocardium in the area at risk. We investigated the impact of VNS, LVAD and their combination (VNS+LVAD) on the infarct size in the IR model.

METHODS: We allocated 17 dogs into 4 groups, Control (n=5), VNS (n=4), LVAD (n=4) and VNS+LVAD (n=4). We occluded major branches of the left anterior descending coronary artery (90min) and reperfused for the following 300min. We titrated the amplitude of VNS to lower heart rate by 10‐20%. We introduced LVAD to unload LV totally and minimize myocardial oxygen consumption (MVO2).

RESULTS: Mean aortic pressure after reperfusion did not differ among 4 groups. Left atrial pressure was much lower in LVAD and VNS+LVAD (Control: 13±1.7, VNS:11±0.72, LVAD: ‐1.6±1.6, VNS/LVAD:‐0.4±1.3 mmHg, p<0.0001). VNS and LVAD reduced the infarct size (relative to Control) by 24% and 88%, respectively. VNS+LVAD reduced the infarct size below the detection threshold (0.0±0%, p<0.0001).

CONCLUSIONS: The VNS combined with mechanical LV unloading maximally reduces the infarct size and could serve as a powerful strategy to prevent post MI heart failure in the long term.
Combinational effects of apocynin and mitoquinone in reducing myocardial ischemia/ reperfusion injury (1080.7)
Authors: Chau William , Ondrasik Regina , Devine Issachar , Patel Harsh , Lepera Matthew , Chen Qian , Barsotti Robert , Young Lindon
Published: June 2021

Reactive oxygen species (ROS) is generated due to the influx of oxygen during MI/R and contributes to post‐reperfused cardiac contractile dysfunction and increased infarct size. Damaged mitochondria and NADPH oxidase activation are major sites of ROS in MI/R. In prior studies, apo, a NADPH oxidase inhibitor, and mitoQ, a mitochondrial‐targeted antioxidant, dose‐dependently improved post‐reperfused left ventricular developed pressure (LVDP) and reduced infarct size in rat hearts subjected to I(30min)/R(45min). This led us to question whether low doses of apo and mitoQ given together can act synergistically to improve post‐reperfused LVDP and reduce infarct size compared to either drug alone or control? Early data shows that the combination of apo (40μM) + mitoQ (1 μM ) (n=7) reduced infarct size to 27 ± 11% compared to mitoQ (1 μM) (n= 6) 54 ± 6% (p<0.05); apo (40 μM) (n=6) 40 ± 8% and control I/R hearts (n=7) 42 ± 5%. Reduced infarct size of the apo + mitoQ combination correlated with an improvement in LVDP (61 ± 4% of baseline) compared to control hearts (44 ± 10% of baseline) and apo 40 μM and mitoQ 1 μM (58 ± 9% and 44 ± 12% of baseline), respectively. Although the combination reduced infarct size and improved post‐reperfused LVDP more so then either drug alone or control, at these concentrations the combinations did not act synergistically. Other combinations will be tested to further establish the dose‐dependent effects.
Renal denervation prevents dendritic cell activation and renal T cell activation in mice in angiotensin II‐induced hypertension (1082.4)
Authors: Xiao Liang , Kirabo Annet , Wu Jing , Harrison David
Published: June 2021

Hypertension is associated with increased sympathetic outflow and subsequent activation of adaptive immunity. We hypothesized that renal sympathetic nerves activate dendritic cells, and that these in turn cause T cells to proliferate and accumulate in the kidney. To test this, we performed bilateral renal denervation (RDN) in C57BL/6 mice by applying phenol to the renal artery and cutting large visible nerves. One week later, mice received angiotensin II (490 ng/kg/min) subcutaneously for 14 days. RDN lowered the hypertensive response to angiotensin II (130±3 vs. 161±3 mmHg, p<0.001) as measured by telemetry. By flow cytometry, we found that angiotensin II increased expression of surface markers CD80 and CD86 on dendritic cells of the kidney by 2 to 3‐fold and that this is significantly attenuated by RDN. RDN also reduced accumulation of total T cells and CD44hi (memory) T cells in the kidney (See table). Moreover, angiotensin II increased IL‐1α, IL‐1β, and IL‐6 production from splenic dendritic cells by 4 to 6‐fold, and RDN eliminated this. In other studies, we found that adoptive transfer of dendritic cells from angiotensin II infused mice augmented the hypertensive response to low dose angiotensin II (140 ng/kg/min), while transfer of dendritic cells from mice that had received angiotensin II after RDN had no effect on the recipient mouse (systolic 156±10 vs 135±9 mmHg, p<0.05). These results indicate that renal sympathetic nerves cause dendritic cell activation, and that dendritic cells play a critical role in the genesis of hypertension.

Sham
Angiotensin
RDN+Angiotensin

Inflammatory cells in kidney (Ã—104 per kidney):

Leukocytes (CD45+)
10.5±1.6
16.8±3.0*
8.9±1.0â€

T cells (CD3+)
1.7±0.3
3.9±0.8*
1.7±0.2â€

Activated dendritic cells(Ã—103 per kidney):

CD80+
1.09±0.19
2.90±0.39*
2.00±0.42â€

CD86+
0.47±0.05
0.87±0.12*
0.59±0.12â€

Cytokines production in DCs (ng per 106 cells):

IL‐1Î±
0.76±0.10
5.04±2.20*
0.59±0.18â€

IL‐1Î²
3.18±0.32
23.99±13.59*
2.75±0.84â€

IL‐6
1.05±0.20
12.36±7.45*
1.02±0.34â€

*p<0.05 vs. sham; †p<0.05 vs. ANG
Interleukin‐15 contributes to decidual natural killer cell loss via interferon‐γ during early pregnancy in the pre‐eclamptic BPH/5 mouse model (1083.1)
Authors: Sones Jenny , Lob Heinrich , Isroff Catherine , Song Jiunn , Williamson Emilie , Davisson Robin
Published: June 2021

During early pregnancy dNK cells promote decidualization, which is critical for uterine angiogenesis and placentation. IL‐15 activates dNK cells, which release IFN‐γ to orchestrate vessel remodeling for proper placental perfusion. However, high levels of IFN‐ γ induce apoptosis of dNK cells at the maternal‐fetal interface. Reduced perfusion of the fetoplacental unit is a hallmark of PE. Our previous studies showed that BPH/5 mice, a rodent model of spontaneous PE, have increased peripheral inflammation. We hypothesize that peripheral inflammation in early pregnancy contributes to elevated IL‐15, which results in dNK cell apoptosis. Virgin BPH/5 mice had significantly elevated uterine IL‐15 mRNA as compared to C57 controls. Other pro‐inflammatory cytokines were unchanged. At e4.5 and e5.5, when decidualization first begins, we observed significantly increased IL‐15 mRNA in BPH/5 implantation sites compared to controls (60‐fold, 20‐fold respectively, p<0.05, n=4). Further, BPH/5 dNK cell mRNA expression was similar to controls at e4.5 but dramatically reduced at e5.5 (10‐fold, p<0.05, n=4). This finding was confirmed by flow cytometry (BPH/5 0.7 x 103 cells vs C57 9.7 x 103 cells, p<0.05, n=4), and was attributed to a 3‐fold increase in IFN‐γ mRNA. These findings support the hypothesis that early pregnancy events may lead to impaired uterine angiogenesis which precedes the development of PE in this model.

Grant Funding Source: #12POST11250010; HL63887, HL84207, 12SDG9160010
Inhibition of endoplasmic reticulum stress does not alter blood pressure in adult male and female spontaneously hypertensive rats (1083.2)
Authors: Crislip Gene , Williams Eric , Tipton Ashlee , Sullivan Jennifer
Published: June 2021

ER stress has recently been linked to the development of hypertension in male SHR. Little is known regarding ER stress in females, although blood pressure (BP) in female SHR is lower than in age‐matched males. This study was designed to test the hypothesis that female SHR have less ER stress than males, and consequently will have a smaller decrease in BP in response to an ER stress inhibitor. 11‐week old rats were treated with vehicle or the ER stress inhibitor phenylbutyric acid (PBA) for 2 weeks (100 mg/kg/day via drinking water; N=4) and BP was measured weekly via tail cuff. The ER stress markers glucose‐regulated protein (GRP78) and C/EBP homologous protein (CHOP) were measured in the renal cortex via Western blot analysis. Basal expression of CHOP was higher in females when compared to males (female: 2.7±0.4 vs. male: 1.3±0.2, P=0.007) as well as with GRP (female: 1.8±0.2 vs. male: 1.3±0.1, P=0.04), however these differences were abolished after treatment. BP did not change in either sex following PBA treatment (female: 169±8 to 174±6 vs. male: 182±4 to 183±5 mmHg, NS). In conclusion, regulation of blood pressure in adult SHR is not mediated by ER stress. Future studies will determine if inhibition of ER stress prior to the development in hypertension effects BP.
Pregnant rats treated with a high fat/pro‐oxidant western diet with angiotensin II and tumor necrosis factor α are resistant to elevations in blood pressure (BP) and have the normal fall in BP during late pregnancy (1084.1)
Authors: Cunningham Mark , West Crystal , Baylis Chris
Published: June 2021

Increased oxidative stress and inflammation are risk factors for hypertension in pregnancy. Here we examined the systolic BP (SBP) via telemetry in virgin (V) and pregnant (P) rats treated with a HFD, ANG II, and TNFα. Methods: Female Sprague Dawley rats were pretreated with a normal diet (ND) or a HFD for 8 weeks before mating. Day 6 P and age‐matched V rats were implanted with either a mini‐pump infusion of saline or ANGII (150 ng/kg/min) + TNFα (75ng/day) for 14 days and sacrificed at gestational day 21. Groups consisted of: V Con (ND + Saline, n=7); V Treatment (HFD + ANGII/TNFα, n=7); P Con (ND + Saline, n=6); P Treatment (HFD + ANGII/TNFα, n=8). Results: The total pressor response (area under the curve [AUC]) increased in V Treatment rats compared to V Con rats (AUC: 120±32 vs. ‐35±29 mmHg X 20 days, P< 0.05) and in P Treatment rats compared to P Con rats (AUC: 61±28 vs. 21±30 mmHg X 20 days, P< 0.05). However the pressor response was greater in V Treatment rats vs. P Treatment rats (120±32 vs. 21±30 mmHg X 20 days, P< 0.05). Both treated and untreated P rats had a normal decrease in SBP at day 20 of pregnancy. Conclusion: Pregnant rats subjected to chronic HFD + pro‐oxidant and pro‐inflammatory insults have a blunted pressor response compared to V Treatment rats. P Treatment rats also display the normal decrease in SBP at late pregnancy. Ongoing studies are investigating the mechanism by which the pregnancy milieu is protective.
Complement components C3a and C5a alter angiogenic balance in placental and endothelial cells (1084.11)
Authors: Gillham Haley , Banek Christopher , Needham Karen , Regal Jean , Gilbert Jeffrey
Published: June 2021

Recent studies have reported excessive activation of complement during pregnancy may contribute to the pathogenesis of preeclampsia, and inhibition of complement activation reduces blood pressure in an animal model of placental ischemia‐induced hypertension. As previous studies have shown a role for angiogenic balance in reducing blood pressure in placental ischemia induced hypertension and the anti‐angiogenic effects of complement activation products, we hypothesized complement activation products C3a and C5a would have anti‐angiogenic effects in human umbilical vein endothelial cells (HUVECs) and rat placental villi explants. Cells were cultured in physiologic normoxic (8%) or hypoxic (1%) conditions and treated with human C3a, human C5a (10 or 100 nmol/L) or vehicle for 24 hours. Experiments were run in triplicate. In placental explants, hypoxia stimulated secretion of vascular endothelial growth factor (VEGF) into the cell media vs. normoxia (1420 ± 75 vs. 1068 ± 52, P<0.05), and this was decreased (P<0.05) by 10nM (‐23%) and 100nM (‐22%) C3a. Secretion of soluble fms‐like tyrosine kinase‐1 (sFlt‐1) from explants was increased (P<0.05) by 10nM (18%) and 100nM (19%) C3a in normoxic but not hypoxic conditions compared to controls. C3a and C5a (100nM) increased sFlt‐1 secretion from HUVECs in normoxic conditions (P<0.05) when compared to controls (771 ± 22 vs. 776 ± 38 vs. 629 ± 14). Secretion of VEGF from HUVECs was decreased with C5a in both oxygen conditions compared to controls (P<0.05) and there was no effect of C3a. These data suggest that products of excessive complement activation have anti‐angiogenic effects in endothelial cells, impairing placental development and invasion and leading to poor placental perfusion. Further, complement activation products appear to disrupt regulation of angiogenic balance in placental cells, which in turn could contribute to continued placental dysfunction in later pregnancy.

Grant Funding Source: Supported by NIH RO1HL114096, 1R15HL109843 and AHA 10SDG2600040
Obese melanocortin‐4 receptor‐deficient rats exhibit attenuated adrenergic vasoconstriction (1084.2)
Authors: Spradley Frank , Palei Ana , Granger Joey
Published: June 2021

Preeclampsia is a hypertensive disorder of pregnancy associated with placental ischemia‐induced endothelial dysfunction and exaggerated vasoconstriction. Obesity is a major risk factor for the development of preeclampsia but the mechanisms are unclear. Intriguingly, we previously showed that obese MC4R heterozygous (+/‐) rats not subjected to placental ischemia are normotensive with augmented endothelial‐dependent relaxation and smooth muscle relaxation response to a nitric oxide donor. To further examine vascular function in obese pregnancy, we tested the hypothesis that obese MC4R +/‐ pregnant rats have attenuated vasoconstriction. At gestational day 19, third‐order mesenteric arteries were isolated from obese MC4R +/‐ (N=6) and lean MC4R +/+ (N=10) rats and mounted in a wire myograph. Sensitivity (logEC50) to vasoconstriction induced by phenylephrine (PE) was significantly (P<0.05) reduced in obese (‐4.9±0.3M) vs. lean (‐5.6±0.1M) pregnant rats whereas the maximum contractile response (% increase in force) to PE was similar between obese (241±57%) and lean (305±49%) pregnant groups. Vasoconstriction stimulated by depolarization to KCl was similar between obese vs. lean rats: EC50 (56.5±2.9mM vs. 48.1±1.3mM) or maximum response (150±13% vs. 168±13%). In conclusion, these data indicate that pregnancy in obese MC4R +/‐ rats is associated with reduced adrenergic vasoconstriction. However, we speculate that adrenergic vasoconstriction may be exaggerated in obese pregnant animals with placental ischemia‐induced endothelial dysfunction.

Grant Funding Source: Supported by HL105324, HL51971.
Antagonism of complement C5a receptor but not C3a receptor attenuates placental ischemia‐induced endothelial dysfunction in rat (1084.8)
Authors: Opacich Jonathan , Johnson Alex , Bauer Ashley , Gilbert Jeffrey , Regal Jean
Published: June 2021

Preeclampsia is characterized by new‐onset hypertension, abnormal placentation, endothelial dysfunction and increased activation of the complement system generating vasoactive products complement C3a and C5a. Previous studies in the reduced uterine perfusion pressure (RUPP) rat model of placental ischemia‐induced hypertension demonstrated impaired endothelial‐dependent relaxation in isolated blood vessels. In addition, inhibition of complement system activation inhibited the hypertension. Whether complement activation is responsible for the endothelial dysfunction following placental ischemia is unclear. Hypothesis: Complement activation products C3a and C5a mediate endothelial dysfunction following placental ischemia. On gestation day (GD) 14, rats underwent sham surgery or clip placement on ovarian arteries and abdominal aorta (RUPP) to cause placental ischemia and increase blood pressure. Rats were treated once daily with C5a receptor antagonist acetyl‐F‐[Orn‐P‐(D‐Cha)‐WR] (PMX53, 3 mg/kg SC), C3a receptor antagonist N2‐[(2,2‐Diphenylethoxy)acetyl]‐L‐arginine (SB290157, 5 mg/kg IV) or vehicle (10% ethanol/saline) from GD14‐18. On GD19, blood pressure was determined via carotid artery and mesenteric arteries removed. Mesenteric arteries (200‐400µm in diameter) were pre‐contracted with 0.44 µM thromboxane mimetic U46619. Endothelial‐dependent and independent relaxations were determined by fractional relaxation of the U46619 contraction to acetylcholine (ACh, 10 nM‐0.31 mM) and sodium nitroprusside (SNP, 7.7 nM‐0.23 mM), respectively. Maximum fractional relaxation to ACh in arteries from RUPP rats (0.68±0.06) was less than Sham (0.88±0.03; p<0.05) and RUPP rats treated with PMX53 (0.83±0.06) but not RUPP rats treated with SB290157 (0.76±0.07). Relaxation to SNP was not affected by the treatments. These data suggest that C5a, but not C3a, mediates endothelial dysfunction induced by placental ischemia.

Grant Funding Source: Supported by NIH HL109843
Early life stress enhances renal expression of T‐cell and B‐cell activation factors (1085.11)
Authors: Obi Ijeoma , Ho Dao , Loria Analia , Pollock Jennifer
Published: June 2021

Early life stress enhances renal expression of T‐cell and B‐cell activation factors Ijeoma Obi, Dao H. Ho, Analia S. Loria, and Jennifer S. Pollock Experimental Medicine, Georgia Regents University, Augusta, GA 30912 In humans, early life stress (ELS) increases cardiovascular disease (CVD) risk and elevates circulating pro‐inflammatory factors; yet the renal inflammatory consequences of ELS remains unclear. We previously showed that in a rat model of ELS, maternal separation (MatSep), baseline blood pressure and renal immune cell infiltration was similar to normally‐reared rats; however MatSep enhanced renal sensitization to the inflammatory and hypertensive effects of angiotensin II (ANGII). We hypothesized that MatSep promotes baseline expression of renal cortex T‐cell and B‐cell activation markers of adult male Wistar Kyoto rats. MatSep animals underwent separation from dams (3h/day) during postnatal day 2‐14; control (CON) littermates were undisturbed. Renal cortex expression of 84 genes representing T‐cell and B‐cell activation was assessed by real‐time qPCR. We determined that caspase 3, Fms‐related tyrosine kinase 3, interleukin 15, phosphoinositase‐3‐kinase, and prolactin receptor expression was significantly increased in MatSep (1.45, 1.44, 1.50, 1.65 and 2.07 fold change from CON, respectively; p < 0.05). Renal cortical T‐cell and macrophages as determined by histology was not different between groups. These data suggest that upregulation of immune cell activation preceeds the development of MatSep‐induced cardiovascular and renal disease.

1F32 HL116145; P01 HL69999

Grant Funding Source: 1F32 HL116145; P01 HL69999
Chronic placental ischemia alters expression of placental and fetal insulin‐like growth factor proteins in the rat (1085.12)
Authors: Beasley Kara , Needham Karen , Regal Jean , Lovering Andrew , Gilbert Jeffrey
Published: June 2021

Preeclampsia is often characterized by placental ischemia and dysregulation of angiogenic growth factors such as IGF and is a major cause of low birth weight and preterm deliveries. These offspring are at increased risk for chronic diseases such as bronchopulmonary dysplasia (BPD), a hypoplastic lung disease defined by preterm birth and neonatal O2 therapy. While O2 therapy and preterm birth clinically define BPD, current models of BPD employ hyperoxia in normally grown term neonates without considering the role of low birth weight and altered fetal endocrine milieu. To that end, we hypothesized chronic placental ischemia would decrease expression of insulin like growth factors (IGF) I and II and related proteins in placental and fetal tissues. Chronic placental ischemia was achieved using the reduced uterine perfusion pressure (RUPP; n 蠅7) model of preeclampsia and fetal growth restriction. Normal pregnant (NP; n 蠅8) rats served as controls and tissues were collected on day 19 of gestation. Placental IGF‐I was decreased in RUPP compared to NP (89.8 ± 10.9 vs. 133.0 ± 13.1ng/ml, p<0.05). Conversely, amniotic fluid IGF‐I (0.25 ± 0.04 vs. 0.16 ± 0.02pg/ml, p<0.05) and IGF‐II (62.7 ± 8.2 vs. 42.0 ± 5.5 pg/ml, p<0.05) were increased in RUPP vs. NP. Fetal hepatic IGF‐2 was increased in RUPP (155.6 ± 12.6 vs. 117.0 ± 6.3pg/ml, p<0.05). Finally, fetal lung IGF‐IRα was not different between the groups. These data indicate chronic placental ischemia alters expression of IGF proteins in placental and fetal tissues and could alter the developmental trajectory of programming susceptible organs such as the lung and kidney.

Grant Funding Source: Supported by NIH RO1HL114096 and AHA 10SDG2600040
Synergy of high salt and high fat diet on kidney injury and adiposity (1086.1)
Authors: Jin Chunhua , MacDonell Robert , Speed Joshua , Pollock David
Published: June 2021

The western‐style diet is a major risk factor for impaired kidney function and chronic kidney disease. The goal of the present study was to determine the relationship of high salt and high fat diet on kidney injury and adiposity. Male Sprague Dawley rats were placed on one of four specific diets for two weeks: (1) Vehicle (control, 0.4% NaCl); (2) high fat (HF, 36% fat); (3) high salt (HS, 4% NaCl); (4) high fat/high salt (HF/HS, 36% fat, 4% NaCl). At the end of the two‐week treatment, rats were placed in metabolic cages for 24 hour urine collection. HS treated rats had significantly higher protein excretion in compared to HF and HF/HS groups (HS: 35.8±6.7, HF: 11.8±1.9, HF/HS: 14.6±2.6 mg/day, p<0.05). Additionally, the HS group had significantly larger kidneys than control and HF groups (HS: 0.35±0.01, control: 0.30±0.01, HF: 0.29±0.01 g/100 g bwt, p<0.05). Furthermore, HF fed rats had significantly higher epididymal fat to body weight ratio than control or HS or HF/HS diet groups (HF: 0.75±0.05, control: 0.59+0.03, HS: 0.49±0.03, HF/HS: 0.60±0.04 g/100 g bwt, p<0.05). The HF diet is hypercaloric compared to the HS or control diet and so animals consumed less food when on the HF diet. In summary, a high fat consumption attenuated the effects of a high salt diet on renal hypertrophy and damage. In addition, a high salt diet appears to reduce high fat induced adiposity. These findings demonstrate a surprising synergy between these two aspects of the Western diet, fat and salt.

Grant Funding Source: PO1 HL‐95499
Resistance arteries from normally‐reared and maternal separation rats display similar vascular reactivity (1087.3)
Authors: Fox Brandon , Loria Analia , Pollock Jennifer
Published: June 2021

We previously reported that MatSep (3 hours/day from days 2‐14) induces exaggerated angiotensin II (AngII)‐induced vasoreactivity with decreased nitric oxide (NO) buffering capacity in aortae of adult rats compared to control rats (normally‐reared littermates). Also, chronic AngII infusion produced exaggerated hypertension in MatSep rats. The aim of this study was to investigate whether MatSep male and female rats display exaggerated vascular reactivity of resistance arteries. Third‐order mesenteric arteries from adult rats (n=5‐6 per group) were isolated and mounted on wire myographs. Concentration‐response curves were generated for phenylephrine or endothelin and normalized to maximal potassium chloride‐induced constriction. Additionally, we tested acetylcholine and sodium nitroprusside‐induced relaxation. No differences in EC50 or %maximum constriction/dilation were observed between MatSep and control rats. Curves in the presence of L‐NAME revealed similar NO buffering capacity in MatSep and control rats. No sex differences were observed across the study. These data suggest that the abnormal vascular phenotype due to MatSep involves large conduit arteries, sparing smaller resistance arteries. However, resistance arteries are known to play an important role in blood pressure control, and thus, further investigation of these arteries in response to chronic AngII infusion is warranted.

Grant Funding Source: Supported by NIH Grants: P01 HL69999, K99 HL111354
Changes in vascular reactivity caused by angiotensin II in heart resistance vessels and isolated vessels of rat aorta in a model of hypertension (1088.1)
Authors: Lopez‐Calderon Emilio , Acevedo‐Villavicencio Nallely , Guevara‐Balcazar Gustavo , Hong‐Chong Enrique , Castillo‐Hernandez Maria
Published: June 2021

Hypertension is a disease with high morbidity and mortality and is considered one of the main etiological factors of multiple pathologies. One of the regulatory mechanisms of blood pressure is the Renin Angiotensin System (RAS), so much so that the first‐line drugs in the treatment of hypertension are directed to some of the components of the RAS. One therapy that has been shown clinically to improve the condition of patients with hypertension is hyperbaric oxygenation (HBO). HBO has been shown to improve vascular smooth muscle relaxation. The aim of this study is analyze the effect of HBO on the RAS. Methods: hypertension was induced in male Wistar rats. Hypertensive rats received HBO and at the end of the treatment were sacrificed getting heart, thoracic and abdominal aorta; and then they were placed in an isolated organ system where we made concentration response curves to Ang II. Conclusion:HBO therapy reduces the reactivity in coronary resistance arteries under Ang II stimulation and reduced vascular reactivity in both segments of the hypertensive rat aorta. This suggests that HBO could possibly be used with hypertensive patients to reduce the use of drugs in therapy.

Grant Funding Source: COFAA / IPN
Molecular mechanisms associated to reversion of proteinuria by RAS antagonists in renovascular hypertensive rats (1088.4)
Authors: Correa Jose , Girardi Adriana , Salles Thiago , Boaro Karoline , Loredo Felipe , Yogi Alvaro , Callera Glaucia , Touyz Rhian , Bendhack Lusiane , Krieger Jose
Published: June 2021

The present work aimed to compare the effects of monotheraphy versus combined therapy of enalapril and losartan and investigate the mechanisms by which these antihypertensive agents revert proteinuria in 2K‐1C hypertensive rats. To this end, male Wistar rats were subjected to 2K‐1C surgery or sham‐operation (2K). After six weeks, rats were treated with losartan (30mg/Kg/day), enalapril (20mg/Kg/day), losartan+enalapril (20+30mg/Kg/day) or saline by gavage for 14 days. Systolic blood pressure (SBP, mmHg) was higher in 2K‐1C (247±5) than in 2K (129±2, P<0.0001). Losartan (197±6, P<0.05) and enalapril (205±7, n=8, P<0.05) progressively reduced 2K‐1C SBP, with an additional hypotensive effect in the group treated with both drugs (173±16mmHg, P<0.001). Plasma levels of Ang II and TBARS were similar in 2K and 2K‐1C rats and were not changed by the treatments. Albuminuria was remarkably increased in 2K‐1C control rats when compared to 2K control rats. Treatment of 2K‐1C rats with losartan and enalapril similarly reduced albumin excretion when compared to the untreated 2K‐1C. A more pronounced reduction of albuminuria was observed in rats treated with combined therapy. Lucigenin‐derived chemiluminescence was not different in the right and left kidneys of 2K and 2K‐1C rats. Treatments had no effect upon NADPH oxidase activity. The expression of glomerular proteins, including nephrin and podocin, and of proximal tubular protein such as megalin and the Cl/H+ exchanger ClC‐5 were much lower in 2K1C compared to 2K. Treatment with enalapril or losartan partially reverted whereas combined therapy completely restored the expression levels of nephrin and podocin. Taken together, these results suggest that the antiproteinuric effects induced by RAS inhibitors in 2K‐1C rats does not involve inhibition of ROS, but is associated with normalization of the expression of nephrin and podocin in the glomeruli.

Grant Funding Source: Supported by FAPESP, CAPES, CNPq, CIHR and Heart and Stroke Foundation of Canada.
Combination therapy of olmesartan/exenatide decreases renal ER stress in a rat model of metabolic syndrome (1088.5)
Authors: Duval Ruilova Steven , De Miguel Carmen , Ortiz Rudy , Pollock Jennifer
Published: June 2021

The OLETF rat is a model of metabolic syndrome, a condition associated with increased activation of RAS and endothelin (ET) systems, renal dysfunction, hypertension and impaired insulin signaling; however, the specific molecular mechanisms that lead to renal dysfunction during metabolic syndrome remain unclear. We hypothesized that this condition is associated with elevated renal ER stress, and that treatment with olmesartan (AT1 receptor blocker; ARB), exenatide (glucagon‐like peptide‐1 analog; EXE), or combination therapy (ARB+EXE), would decrease renal expression of ET, NOS and ER stress markers. OLETF rats and their controls (LETO) were subjected to individual/combined treatments of ARB (10ug/kg/day; n=4) and EXE (5ug/kg/day; n=7) for 6 weeks. Whole kidney tissue was analyzed by qRT‐PCR. Individual drug treatment of OLETF rats led to increased relative mRNA expression of ET system (ARB vs. EXE, fold increase; ET: 2.61 ± 0.5 vs. 3.66 ± 1.0; ETA: 1.58 ± 0.5 vs. 3.62 ± 1.1 and ETB: 1.29 ± 0.5 vs. 3.59 ± 0.9), ER stress (GRP78: 1.26 ± 0.6 vs. 2.05 ± 0.5; CHOP: 2.56 ± 0.8 vs. 4.05 ± 1.1 and spliced XBP‐1: 0.70 ± 0.3 vs. 2.12 ± 0.7), and NOS system (NOS1: 2.47 ± 1.1 vs. 3.89 ± 1.2; NOS3: 1.28 ± 0.6 vs. 2.98 ± 1.0). All markers were ameliorated after combined ARB+EXE treatment. In conclusion, these results suggest that a combination ARB+EXE therapy is a plausible preventive treatment for renal dysfunction associated with metabolic syndrome.
Endothelium‐targeted deletion of Nox4 attenuates hypoxia‐induced increases in right ventricular pressure (1089.19)
Authors: Sutliff Roy , Kleinhenz Jennifer , Ma Jing , Hart C.
Published: June 2021

The complex pathobiology of pulmonary hypertension (PH) involves reactive oxygen species (ROS) and increased pulmonary vascular resistance. NADPH oxidase 4 (Nox4) has been shown to be an important source of ROS in PH, and systemic administration of a Nox4 pharmacological inhibitor attenuated hypoxia‐induced PH. To further explore the role of pulmonary vascular Nox4 in hypoxia‐induced PH, a floxed Nox4 mouse model was crossed with mice that: 1) express Cre recombinase driven by the tie‐2 promoter to target deletion of endothelial Nox4 (eNox4), or 2) express Cre recombinase driven by the smooth muscle myosin heavy chain promoter to target deletion of smooth muscle Nox4 (smNox4). Littermate controls, eNox4 and smNox4 mice were exposed to normoxia (21% O2) or hypoxia (10% O2) for 3 weeks. PH development was assessed by measuring right ventricular systolic pressures (RVSP) and right ventricular hypertrophy (RVH). Hypoxia‐induced increases in RVSP were attenuated in the eNox4 model (N=5; p<0.05) whereas the RVSP in smNox4 knockout mice was not different than controls. In contrast, hypoxia‐induced RVH was no different than controls in either the eNox4 or smNox4 mice. These studies suggest that endothelial Nox4 promotes pulmonary vascular derangements involved in PH pathogenesis.

Grant Funding Source: Supported by VA Merit Review 1I01BX001910 (CMH) and NIH grant R01 HL102167 (CMH and RLS).
Augmentation area index: a novel approach to disease severity and outcome in patients with WHO group I and group III pulmonary hypertension (1089.2)
Authors: Shterental Sebastian , Alam Mian , Bachman Timothy , Corotto Paul , Gupta Shikha , Longhini Anthony , Nguyen Daniel , Sciurba Frank , Simon Marc , Champion Hunter
Published: June 2021

Background: Pulmonary hypertension (PH) has a poor outcome overwhelmingly related to right ventricular (RV) failure. Augmentation area index (AI) could capture hemodynamic effects of vessel stiffening on this processes. We studied AI’s relationship to other hemodynamic measures and to clinical outcomes.

Methods: This is a retrospective, exploratory clinical and hemodynamic data analysis of PH patients diagnosed with idiopathic pulmonary arterial hypertension (IPAH) or chronic obstructive pulmonary disease (COPD) who were catheterized at UPMC Presbyterian Hospital from 1/1/2006 to 3/26/2012. AI was calculated in a blinded fashion from RV pressure wave‐forms using validated image analysis software. We used linear regression to study the relationships between AI and hemodynamic parameters, multiple regression to study the primary outcome of time to death or transplantation, and logistic regression to study secondary outcome of being World Health Organization functional category 1 or 2 versus 3 or 4.

Results: Clinical and hemodynamic data were analyzed for 63 patients. Mean age was 56.3 years (SD 15.0) and 67% were female. There was a positive correlation between AI and PVR (R2=.16 t=2.85 p=0.007) and a negative correlation between AI and CI (R2=.07, t=‐2.04, p=0.046). When adjusted for calcium channel blocker treatment and pulse pressure, AI was a significant predictor of time to death or transplant (R2=0.67, F(3,16)=11, p<.001). Logistic regression testing of AI as the primary predictor of clinical function adjusted for systemic hypertension, tobacco use, prostacyclin treatment, body‐surface area and peripheral vascular resistance, yielded an odds ratio of 1.12 (p=.001, partial p= 0.014).

Conclusion: In patients with different etiologies of PH, AI could be a significant predictor of clinical outcome. A novel approach to the pulmonary system, it correlates with other hemodynamic values previously shown to be significant markers of disease progression.

Grant Funding Source: NIH TPPG
Phenotypic characteristics of pulmonary supernumerary arterial endothelium may be involved in the formation of plexiform lesions (1089.21)
Authors: Oshima Kaori , Matsumoto Yuri , O'Neill Kealan , McMurtry Ivan , Stevens Troy , Oka Masahiko
Published: June 2021

The plexiform lesion (PL), which comprises proliferative endothelial marker (EC)‐positive channel cells and chromatin‐rich core cells, is the histological hallmark of severe pulmonary arterial hypertension (PAH). PLs preferentially form in supernumerary arteries (SAs) in SU5416/hypoxia/normoxia‐exposed PAH rats, which supports clinical observations. However, the underlying pathological mechanisms of this observation are unknown. SAs are unique small pulmonary branches with specific anatomical and functional properties, such as the phenotype of the ECs, which may contribute to the formation of PLs. The EC phenotype in PLs seems to be shared with pulmonary microvascular ECs (PMVECs), which are highly proliferative. Therefore, we tested the hypothesis that the ECs in SAs share the same phenotype as PMVECs that predisposes to the development of PLs.

A lectin protein from Griffonia simplicifolia (GS), which selectively interacts with PMVECs was perfused through the pulmonary vasculature of control and PAH rats. Lung slices were prepared and examined by confocal fluorescence microscopy.

In normal rats, ECs in SAs interacted with GS. However, ECs in SAs and occlusive vascular lesions in PAH rats had no apparent interaction with GS.

The PMVEC‐like phenotype of ECs in SAs may contribute to the development of PLs, and there may be a phenotypic transformation of these cells during the progression of PAH.

Grant Funding Source: Supported by NIH HL66299 and HL60024 to T.S.
Preservation of Ca2+ spark activity during oxidative stress in pulmonary arterial myocytes of fetal sheep (1089.5)
Authors: Suarez Gema , Glasgow Shane , Romero Monica , Osman Noah , Puglisi Jose , Bers Donald , Izu Leighton , Longo Lawerence , Wilson Sean
Published: June 2021

Antenatal chronic hypoxia can lead to pulmonary hypertension in newborns or more severe vascular remodeling and dangerous elevations in pressure. We have previously shown that antenatal chronic hypoxia deregulates Ca2+ spark activity, which are important to pulmonary vasodilation at birth. Chronic hypoxia can also increase oxidative stress, which can impair Ca2+ spark activity. In this research, we examined the impact of oxidative stress on Ca2+ spark activity, which other laboratories have shown to increase Ca2+ spark activity. We exposed pulmonary arteries from normoxic fetal sheep to reactive oxygen species (ROS) to mimic oxidative stress such as might occur with acute hypoxic stress by treating them with tert‐butyl H2O2 and reduced ROS with the antioxidant N‐Acetyl‐L‐Cysteine (NAC). Using customized analysis software, roughly 7 % of control cells had Ca2+ sparks and the frequency was 0.07 sparks/100 μm/sec. Spark activity was maintained in the presence of tert‐butyl H2O2 and NAC. The spatial and temporal aspects to the Ca2+ sparks were then determined. Spark amplitudes were unaffected as was the width. However, Ca2+ spark decay was increased slightly by tert‐butyl H2O2. Overall, oxidative stress has only a mild influence on Ca2+ spark activity on fetal pulmonary arteries from normoxic sheep.

Grant Funding Source: Supported by NSF MRI 923559, NIH HD69746, P01HD31226, R01HD3807, LLUSOM, APS Frontiers in Physiology
cGMP amplification of pulmonary arterial myocyte Ca2+ waves is preferentially impaired in high altitude‐induced hypoxic fetal sheep (1089.7)
Authors: Paez Ricardo , Romero Monica , Blood Quintin , Osman Noah , Manjunath Chetas , Glasgow Shane , Francis Michael , Taylor Mark , Longo Lawerence , Wilson Sean
Published: June 2021

High altitude is a risk factor in the development of pulmonary hypertension (PH). Treatment often includes the use of phosphodiesterase (PDE) inhibitors of PDE3 (milrinone, cAMP) and PDE5 (sildenafil, cGMP), which increase cyclic nucleotides and promotes vasodilation in pulmonary arterial (PA) myocytes. With respect to the current studies, c‐AMP and c‐GMP enhance ryanodine receptor (RyR) and Sarco/Endoplasmic Reticulum Ca2+ ‐ATPase (SERCA) pump activity in myocytes, which regulate intracellular Ca2+. To test whether long term hypoxia (LTH) impairs, and maturity enhances the effects of c‐AMP and c‐GMP on myocyte Ca2+ waves, we selected 3‐isobutyl‐1‐methyl xanthine (IBMX), a PDE inhibitor, as well as 8‐Br‐c‐AMP and 8‐Br‐c‐GMP. We measured cytosolic Ca2+ in PA myocytes of fetal and adult sheep that lived at low (~353 m) or high altitude (~3801 m) via confocal imaging of fluo‐4. Maturation increased while IBMX, 8‐Br‐c‐AMP, and 8‐Br‐c‐GMP each failed to alter the percentage of responsive cells independent of altitude. LTH fetuses had reduced wave amplitude and area under the curve (AUC), but wave kinetics were unaltered. The AUC was maintained in LTH adults, but waves were faster and larger. c‐GMP increased AUC by lengthening waves in all groups except for hypoxic fetuses. Alterations of Ca2+ regulatory pathways including SERCAs and RyRs could contribute to LTH impairment in Ca2+ waves and c‐GMP amplified signals.

Grant Funding Source: Supported by NSF MRI 0923559, NIH HD‐069746, P01HD031226, , R01HD003807, 5P20 MD‐006988,LLUSOM