Joo Yun Jun

Brain Microglial Cytokines in Neurogenic Hypertension

Accumulating evidence indicates a key role of inflammation in hypertension and cardiovascular disorders. However, the role of inflammatory processes in neurogenic hypertension remains to be determined. Thus, our objective in the present study was to test the hypothesis that activation of microglial cells and the generation of proinflammatory cytokines in the paraventricular nucleus (PVN) contribute to neurogenic hypertension. Intracerebroventricular infusion of minocycline, an anti-inflammatory antibiotic, caused a significant attenuation of mean arterial pressure, cardiac hypertrophy, and plasma norepinephrine induced by chronic angiotensin II infusion. This was associated with decreases in the numbers of activated microglia and mRNAs for interleukin (IL) 1&bgr;, IL-6, and tumor necrosis factor-&agr;, and an increase in the mRNA for IL-10 in the PVN. Overexpression of IL-10 induced by recombinant adenoassociated virus-mediated gene transfer in the PVN mimicked the antihypertensive effects of minocycline. Furthermore, acute application of a proinflammatory cytokine, IL-1&bgr;, into the left ventricle or the PVN in normal rats resulted in a significant increase in mean arterial pressure. Collectively, this indicates that angiotensin II induced hypertension involves activation of microglia and increases in proinflammatory cytokines in the PVN. These data have significant implications on the development of innovative therapeutic strategies for the control of neurogenic hypertension.

The angiotensin-converting enzyme 2/angiogenesis-(1-7)/Mas axis confers cardiopulmonary protection against lung fibrosis and pulmonary hypertension.

RATIONALE An activated vasoconstrictive, proliferative, and fibrotic axis of the renin angiotensin system (angiotensin-converting enzyme [ACE]/angiotensin [Ang]II/AngII type 1 receptor) has been implicated in the pathophysiology of pulmonary fibrosis (PF) and pulmonary hypertension (PH). The recent discovery of a counterregulatory axis of the renin angiotensin system composed of ACE2/Ang-(1-7)/Mas has led us to examine the role of this vasoprotective axis on such disorders. OBJECTIVES We hypothesized that Ang-(1-7) treatment would exert protective effects against PF and PH. METHODS Lentiviral packaged Ang-(1-7) fusion gene or ACE2 cDNA was intratracheally administered into the lungs of male Sprague Dawley rats. Two weeks after gene transfer, animals received bleomycin (2.5 mg/kg). In a subsequent study, animals were administered monocrotaline (MCT, 50 mg/kg). MEASUREMENTS AND MAIN RESULTS In the PF study, bleomycin administration resulted in a significant increase in right ventricular systolic pressure, which was associated with the development of right ventricular hypertrophy. The lungs of these animals also exhibited excessive collagen deposition, decreased expression of ACE and ACE2, increased mRNA levels for transforming growth factor β and other proinflammatory cytokines, and increased protein levels of the AT₁R. Overexpression of Ang-(1-7) significantly prevented all the above-mentioned pathophysiological conditions. Similar protective effects were also obtained with ACE2 overexpression. In the PH study, rats injected with MCT developed elevated right ventricular systolic pressure, right ventricular hypertrophy, right ventricular fibrosis, and pulmonary vascular remodeling, all of which were attenuated by Ang-(1-7) overexpression. Blockade of the Mas receptor abolished the beneficial effects of Ang-(1-7) against MCT-induced PH. CONCLUSIONS Our observations demonstrate a cardiopulmonary protective role for the ACE2/Ang-(1-7)/Mas axis in the treatment of lung disorders.

ACE2 activation promotes antithrombotic activity.

The aim of the present study was to test the hypothesis that the activation of the angiotensin-converting enzyme (ACE)2/angiotensin-(1–7)/Mas receptor axis by use of a novel ACE2 activator (XNT) would protect against thrombosis. Thrombi were induced in the vena cava of spontaneously hypertensive rats (SHR) and Wistar Kyoto (WKY) rats, and ACE2 and ACE activity in the thrombus was determined. Real-time thrombus formation was viewed through intravital microscopy of vessels in nude mice. Thrombus weight was 40% greater in the SHR (4.99 ± 0.39 versus 7.04 ± 0.66 mg). This weight increase was associated with a 20% decrease in ACE2 activity in the thrombus. In contrast, there were no differences between the WKY and SHR in ACE2 protein and ACE activity in the thrombi. ACE2 inhibition (DX600; 0.1 µmol/L/kg) increased thrombus weight by 30% and XNT treatment (10 mg/kg) resulted in a 30% attenuation of thrombus formation in the SHR. Moreover, XNT reduced platelet attachment to injured vessels, reduced thrombus size, and prolonged the time for complete vessel occlusion in mice. Thus, a decrease in thrombus ACE2 activity is associated with increased thrombus formation in SHR. Furthermore, ACE2 activation attenuates thrombus formation and reduces platelet attachment to vessels. These results suggest that ACE2 could be a novel target for the treatment of thrombogenic diseases.

Altered Inflammatory Response Is Associated With an Impaired Autonomic Input to the Bone Marrow in the Spontaneously Hypertensive Rat

Autonomic nervous system dysfunction, exaggerated inflammation, and impaired vascular repair are all hallmarks of hypertension. Considering that bone marrow (BM) is a major source of the inflammatory cells (ICs) and endothelial progenitor cells (EPCs), we hypothesized that impaired BM–autonomic nervous system interaction contributes to dysfunctional BM activity in hypertension. In the spontaneously hypertensive rat (SHR), we observed a >30% increase in BM and blood ICs (CD4.8+) and a >50% decrease in EPCs (CD90+.CD4.5.8–) when compared with the normotensive Wistar–Kyoto rat. Increased tyrosine hydroxylase (70%) and norepinephrine (160%) and decreased choline acetyl transferase (30%) and acetylcholine esterase (55%) indicated imbalanced autonomic nervous system in SHR BM. In Wistar–Kyoto rat, night time–associated elevation in sympathetic nerve activity (50%) and BM norepinephrine (41%) was associated with increased ICs (50%) and decreased EPCs (350%) although BM sympathetic denervation decreased ICs (25%) and increased EPCs (40%). In contrast, these effects were blunted in SHR, possibly because of chronic downregulation of BM adrenergic receptor &agr;2a (by 50%–80%) and &bgr;2 (30%–45%). Application of norepinephrine resulted in increased BM IC activation/release, which was prevented by preadministration of acetylcholine. Electrophysiological recordings of femoral sympathetic nerve activity showed a more robust femoral sympathetic nerve activity in SHR when compared with Wistar–Kyoto rat, peaking earlier in the respiratory cycle, indicative of increased sympathetic tone. Finally, manganese-enhanced MRI demonstrated that presympathetic neuronal activation in SHR was associated with an accelerated retrograde transport of the green fluorescent protein–labeled pseudorabies virus from the BM. These observations demonstrate that a dysfunctional BM autonomic nervous system is associated with imbalanced EPCs and ICs in hypertension.

Nucleus of the solitary tract (pro)renin receptor-mediated antihypertensive effect involves nuclear factor-κB-cytokine signaling in the spontaneously hypertensive rat.

The importance of the (pro)renin receptor (PRR) in the function of the central nervous system is increasingly evident because PRR seems to play a role in neuronal control of cardiovascular function. PRR expression is elevated in the nucleus of the solitary tract (NTS) of spontaneously hypertensive rats (SHR). In this study, we tested the hypothesis that altered activity of PRR in the NTS is linked to hypertension. Eight weeks of chronic knockdown of the NTS PRR, using recombinant adeno-associated virus type 2 (AAV2)-PRR-small hairpain RNA (shRNA)–mediated gene transduction, caused a significant increase in mean arterial pressure (MAP) in the SHR (shRNA, 173±5; Control, 151±6 mm Hg) but not in Wistar Kyoto rats (shRNA, 108±7; Control, 106±6 mm Hg). The MAP elevation in the SHR was associated with decreased inflammatory markers tumor necrosis factor-&agr;, interleukin-6, C-C motif ligand 5, and their transcription factor, nuclear factor-&kgr;B. Consistent with the pressor effects of the PRR knockdown, acute bilateral NTS injection of human renin (2 pmol/side) decreased MAP and heart rate (HR) in SHR (&Dgr;MAP, −38±4 mm Hg; &Dgr;heart rate, −40±10 bpm), with negligible responses in Wistar Kyoto rats (&Dgr;MAP, −4±3 mm Hg; &Dgr;heart rate, −12±7 bpm). These effects in SHR were attenuated (80%) by prorenin handle region peptide but were not affected by angiotensin II type 1 or angiotensin II type 2 receptor blockers. Finally, PRR activation in SHR neuronal cultures by prorenin activated nuclear factor-&kgr;B and increased mRNA levels of interleukin-1&bgr; (250-fold), tumor necrosis factor-&agr; (32-fold), interleukin-6 (35-fold), C-C motif ligand 5 (12-fold), and interleukin-10 (7-fold) in a nuclear factor-&kgr;B–dependent but angiotensin II type 1 receptor–independent manner. Therefore, NTS PRR mediates antihypertensive effects via an angiotensin II–independent mechanism in SHR, which involves stimulation of the nuclear factor-&kgr;B–cytokine signaling pathway.

Chronic Knockdown of the Nucleus of the Solitary Tract AT1 Receptors Increases Blood Inflammatory-Endothelial Progenitor Cell Ratio and Exacerbates Hypertension in the Spontaneously Hypertensive Rat

AT1 receptor subtype a (AT1Ra) expression is increased in the nucleus of the solitary tract (NTS) in spontaneously hypertensive rat (SHR) compared with Wistar Kyoto controls. However, the chronic role of AT1Ra in the NTS for cardiovascular control is not well understood. In this study, we investigated the hypothesis that the NTS AT1Ra is involved in the neural regulation of the peripheral inflammatory status and linked with hypertension. Transduction of brain neuronal cultures with recombinant adeno-associated virus type 2 (AAV2)-AT1R-small hairpin RNA (shRNA) resulted in a 72% decrease in AT1Ra mRNA and attenuated angiotensin II–induced increase in extracellular signal–regulated kinase 1/2 phosphorylation and neuronal firing. Specific NTS microinjection of AAV2-AT1R-shRNA vector in the SHR resulted in a ≈30 mm Hg increase in the mean arterial pressure compared with control vector–injected animals (Sc-shRNA: 154±4 mm Hg; AT1R-shRNA: 183±10 mm Hg) and induced a resetting of the baroreflex control of heart rate to higher mean arterial pressure. In addition, AAV2-AT1R-shRNA–treated SHRs exhibited a 74% decrease in circulating endothelial progenitor cells (CD90+, CD4−/CD5−/CD8−) and a 300% increase in the circulating inflammatory cells, including CD4+ +CD8+, CD45+/3+ T lymphocytes, and macrophages (CD68+). As a result, the endothelial progenitor cell/inflammatory cells ratio was decreased by 8- to 15-fold in the AT1R-shRNA–treated SHR. However, identical injection of AAV2-AT1R-shRNA into the NTS of Wistar Kyoto rats had no effect on mean arterial pressure and inflammatory cells. These observations suggest that increased expression of the AT1Ra in SHR NTS may present a counterhypertensive mechanism involving inflammatory/angiogenic cells.AT1 receptor subtype a (AT1R1a) expression is increased in the nucleus of the solitary tract (NTS) in Spontaneously Hypertensive Rat (SHR) compared to Wistar Kyoto (WKY) controls. However, the chronic role of AT1Ra in the NTS for cardiovascular control is not well understood. In this study, we investigated the hypothesis that the NTS AT1Ra is involved in neural regulation of the peripheral inflammatory status, and linked with hypertension. Transduction of brain neuronal cultures with AAV2-AT1R-shRNA resulted in a 72% decrease in AT1Ra mRNA, and attenuated AngII-induced increase in ERK1/2 phosphorylation and neuronal firing. Specific NTS microinjection of AAV2-AT1R-shRNA vector in the SHR resulted in a ~30 mmHg increase in the mean arterial pressure (MAP) compared to control vector injected animals (Sc-shRNA: 154±4; AT1R-shRNA: 183±10 mmHg), and induced a resetting of the baroreflex control of heart rate to higher MAP. In addition, AAV2-AT1R-shRNA-treated SHRs exhibited a 74% decrease in circulating endothelial progenitor cells (EPC, CD90+, CD4−/5−/8−), and a 300% increase in the circulating inflammatory cells (IC) including CD4+/CD8+, CD45+/3+ T lymphocytes, and macrophages (CD68+). As a result, the EPC/IC ratio was decreased by 8~15 fold in the AT1R-shRNA-treated SHR. However, identical injection of AAV2-AT1R-shRNA into the NTS of WKY had no effect on MAP and ICs. These observations suggest that increased expression of the AT1Ra in SHR NTS may present a counter-hypertensive mechanism involving inflammatory/angiogenic cells.

Chronic Knockdown of the Nucleus of the Solitary Tract AT1 Receptors Increases Blood Inflammatory-Endothelial Progenitor Cell Ratio and Exacerbates Hypertension in the Spontaneously Hypertensive RatNovelty and Significance

AT1 receptor subtype a (AT1Ra) expression is increased in the nucleus of the solitary tract (NTS) in spontaneously hypertensive rat (SHR) compared with Wistar Kyoto controls. However, the chronic role of AT1Ra in the NTS for cardiovascular control is not well understood. In this study, we investigated the hypothesis that the NTS AT1Ra is involved in the neural regulation of the peripheral inflammatory status and linked with hypertension. Transduction of brain neuronal cultures with recombinant adeno-associated virus type 2 (AAV2)-AT1R-small hairpin RNA (shRNA) resulted in a 72% decrease in AT1Ra mRNA and attenuated angiotensin II–induced increase in extracellular signal–regulated kinase 1/2 phosphorylation and neuronal firing. Specific NTS microinjection of AAV2-AT1R-shRNA vector in the SHR resulted in a ≈30 mm Hg increase in the mean arterial pressure compared with control vector–injected animals (Sc-shRNA: 154±4 mm Hg; AT1R-shRNA: 183±10 mm Hg) and induced a resetting of the baroreflex control of heart rate to higher mean arterial pressure. In addition, AAV2-AT1R-shRNA–treated SHRs exhibited a 74% decrease in circulating endothelial progenitor cells (CD90+, CD4−/CD5−/CD8−) and a 300% increase in the circulating inflammatory cells, including CD4+ +CD8+, CD45+/3+ T lymphocytes, and macrophages (CD68+). As a result, the endothelial progenitor cell/inflammatory cells ratio was decreased by 8- to 15-fold in the AT1R-shRNA–treated SHR. However, identical injection of AAV2-AT1R-shRNA into the NTS of Wistar Kyoto rats had no effect on mean arterial pressure and inflammatory cells. These observations suggest that increased expression of the AT1Ra in SHR NTS may present a counterhypertensive mechanism involving inflammatory/angiogenic cells.AT1 receptor subtype a (AT1R1a) expression is increased in the nucleus of the solitary tract (NTS) in Spontaneously Hypertensive Rat (SHR) compared to Wistar Kyoto (WKY) controls. However, the chronic role of AT1Ra in the NTS for cardiovascular control is not well understood. In this study, we investigated the hypothesis that the NTS AT1Ra is involved in neural regulation of the peripheral inflammatory status, and linked with hypertension. Transduction of brain neuronal cultures with AAV2-AT1R-shRNA resulted in a 72% decrease in AT1Ra mRNA, and attenuated AngII-induced increase in ERK1/2 phosphorylation and neuronal firing. Specific NTS microinjection of AAV2-AT1R-shRNA vector in the SHR resulted in a ~30 mmHg increase in the mean arterial pressure (MAP) compared to control vector injected animals (Sc-shRNA: 154±4; AT1R-shRNA: 183±10 mmHg), and induced a resetting of the baroreflex control of heart rate to higher MAP. In addition, AAV2-AT1R-shRNA-treated SHRs exhibited a 74% decrease in circulating endothelial progenitor cells (EPC, CD90+, CD4−/5−/8−), and a 300% increase in the circulating inflammatory cells (IC) including CD4+/CD8+, CD45+/3+ T lymphocytes, and macrophages (CD68+). As a result, the EPC/IC ratio was decreased by 8~15 fold in the AT1R-shRNA-treated SHR. However, identical injection of AAV2-AT1R-shRNA into the NTS of WKY had no effect on MAP and ICs. These observations suggest that increased expression of the AT1Ra in SHR NTS may present a counter-hypertensive mechanism involving inflammatory/angiogenic cells.

Chronic Knockdown of the NTS AT1R Increases Blood Inflammatory-Endothelial Progenitor Cells Ratio and Exacerbates Hypertension in the SHR

AT1 receptor subtype a (AT1Ra) expression is increased in the nucleus of the solitary tract (NTS) in spontaneously hypertensive rat (SHR) compared with Wistar Kyoto controls. However, the chronic role of AT1Ra in the NTS for cardiovascular control is not well understood. In this study, we investigated the hypothesis that the NTS AT1Ra is involved in the neural regulation of the peripheral inflammatory status and linked with hypertension. Transduction of brain neuronal cultures with recombinant adeno-associated virus type 2 (AAV2)-AT1R-small hairpin RNA (shRNA) resulted in a 72% decrease in AT1Ra mRNA and attenuated angiotensin II–induced increase in extracellular signal–regulated kinase 1/2 phosphorylation and neuronal firing. Specific NTS microinjection of AAV2-AT1R-shRNA vector in the SHR resulted in a ≈30 mm Hg increase in the mean arterial pressure compared with control vector–injected animals (Sc-shRNA: 154±4 mm Hg; AT1R-shRNA: 183±10 mm Hg) and induced a resetting of the baroreflex control of heart rate to higher mean arterial pressure. In addition, AAV2-AT1R-shRNA–treated SHRs exhibited a 74% decrease in circulating endothelial progenitor cells (CD90+, CD4−/CD5−/CD8−) and a 300% increase in the circulating inflammatory cells, including CD4+ +CD8+, CD45+/3+ T lymphocytes, and macrophages (CD68+). As a result, the endothelial progenitor cell/inflammatory cells ratio was decreased by 8- to 15-fold in the AT1R-shRNA–treated SHR. However, identical injection of AAV2-AT1R-shRNA into the NTS of Wistar Kyoto rats had no effect on mean arterial pressure and inflammatory cells. These observations suggest that increased expression of the AT1Ra in SHR NTS may present a counterhypertensive mechanism involving inflammatory/angiogenic cells.AT1 receptor subtype a (AT1R1a) expression is increased in the nucleus of the solitary tract (NTS) in Spontaneously Hypertensive Rat (SHR) compared to Wistar Kyoto (WKY) controls. However, the chronic role of AT1Ra in the NTS for cardiovascular control is not well understood. In this study, we investigated the hypothesis that the NTS AT1Ra is involved in neural regulation of the peripheral inflammatory status, and linked with hypertension. Transduction of brain neuronal cultures with AAV2-AT1R-shRNA resulted in a 72% decrease in AT1Ra mRNA, and attenuated AngII-induced increase in ERK1/2 phosphorylation and neuronal firing. Specific NTS microinjection of AAV2-AT1R-shRNA vector in the SHR resulted in a ~30 mmHg increase in the mean arterial pressure (MAP) compared to control vector injected animals (Sc-shRNA: 154±4; AT1R-shRNA: 183±10 mmHg), and induced a resetting of the baroreflex control of heart rate to higher MAP. In addition, AAV2-AT1R-shRNA-treated SHRs exhibited a 74% decrease in circulating endothelial progenitor cells (EPC, CD90+, CD4−/5−/8−), and a 300% increase in the circulating inflammatory cells (IC) including CD4+/CD8+, CD45+/3+ T lymphocytes, and macrophages (CD68+). As a result, the EPC/IC ratio was decreased by 8~15 fold in the AT1R-shRNA-treated SHR. However, identical injection of AAV2-AT1R-shRNA into the NTS of WKY had no effect on MAP and ICs. These observations suggest that increased expression of the AT1Ra in SHR NTS may present a counter-hypertensive mechanism involving inflammatory/angiogenic cells.