[A preliminary study on the construction of the triad medical experimental system].

The traditional medical experiment based on animal studies fails to reflect competency-oriented goal, and is not closely combined with clinical and scientific research, which does not meet the need for early clinical and scientific training. In order to cultivate the first-class medical talents, medical experimental teaching should conform to the trend of modern medical education, innovate teaching ideas and models, and update the hardware and software in time. Therefore, our teaching center adopts the triad medical experimental system which consists of "animal experiments, human functional experiments, and electronic standardized patient (ESP)-based virtual simulation experiments", and uses one system to integrate basic and clinical medicine, practice and virtual learning, teaching and scientific training. The system retains the core content of traditional animal experiments, and includes the most mature and widely used human physiological experiments to increase students' learning experience. With medical simulation experiment, it explains the specific physiological and pathophysiological processes of human body to improve students' cognitive and thinking ability. Here, we provide a systematic description on our triad medical experimental system, and discuss the experience to establish this novel system.

[Building virtual simulation teaching platform based on electronic standardized patient].

Informatization is an effective way to promote the reform and innovation of higher education and improve its quality. Virtual simulation teaching is indispensable in the educational informatization. Here, we describe the development and current situation of virtual simulation teaching, and introduce electronic standardized patient (ESP) based-virtual human body system powered by the real-time human physiological parameters. We also discuss how to build an ESP-based community in the teaching of human physiology, preclinical integrated case learning and other teaching projects. These ESP-based virtual simulation projects display the advantages of interdisciplinary fusion and the combination of basic and clinical knowledge, and open up the third type of functional experiments. Therefore, ESP-based virtual simulation teaching platform presumably becomes a considerable option for the first-class course construction in physiology.

MiR155-5p in adventitial fibroblasts-derived extracellular vesicles inhibits vascular smooth muscle cell proliferation via suppressing angiotensin-converting enzyme expression.

Proliferation of vascular smooth muscle cells (VSMCs) plays crucial roles in vascular remodelling and stiffening in hypertension. Vascular adventitial fibroblasts are a key regulator of vascular wall function and structure. This study is designed to investigate the roles of adventitial fibroblasts-derived extracellular vesicles (EVs) in VSMC proliferation and vascular remodelling in normotensive Wistar-Kyoto rat (WKY) and spontaneously hypertensive rat (SHR), an animal model of human essential hypertension. EVs were isolated from aortic adventitial fibroblasts of WKY (WKY-EVs) and SHR (SHR-EVs). Compared with WKY-EVs, miR155-5p content was reduced, while angiotensin-converting enzyme (ACE) content was increased in SHR-EVs. WKY-EVs inhibited VSMC proliferation of SHR, which was prevented by miR155-5p inhibitor. SHR-EVs promoted VSMC proliferation of both strains, which was enhanced by miR155-5p inhibitor, but abolished by captopril or losartan. Dual luciferase reporter assay showed that ACE was a target gene of miR155-5p. MiR155-5p mimic or overexpression inhibited VSMC proliferation and ACE upregulation of SHR. WKY-EVs reduced ACE mRNA and protein expressions while SHR-EVs only increased ACE protein level in VSMCs of both strains. However, the SHR-EVs-derived from the ACE knockdown-treated adventitial fibroblasts lost the roles in promoting VSMC proliferation and ACE upregulation. Systemic miR155-5p overexpression reduced vascular ACE, angiotensin II and proliferating cell nuclear antigen levels, and attenuated hypertension and vascular remodelling in SHR. Repetitive intravenous injection of SHR-EVs increased blood pressure and vascular ACE contents, and promoted vascular remodelling in both strains, while WKY-EVs reduced vascular ACE contents and attenuated hypertension and vascular remodelling in SHR. We concluded that WKY-EVs-mediated miR155-5p transfer attenuates VSMC proliferation and vascular remodelling in SHR via suppressing ACE expression, while SHR-EVs-mediated ACE transfer promotes VSMC proliferation and vascular remodelling.

FNDC5 inhibits foam cell formation and monocyte adhesion in vascular smooth muscle cells via suppressing NFκB-mediated NLRP3 upregulation.

Foam cell formation and monocytes adhesion are key events in pathogenesis of atherosclerosis. Vascular smooth muscle cells (VSMCs) are an important origin of foam cells besides macrophages. Fibronectin type III domain containing protein 5 (FNDC5) is a protein, which induces browning of fat and attenuates glucose/lipid metabolic derangements in obese mice. The present study was designed to determine the roles of FNDC5 in inhibiting foam cell formation and monocyte adhesion in VSMCs and its underlying mechanisms. Oxidized low-density lipoprotein (oxLDL) was used to induce foam cell formation and monocyte adhesion in human aortic VSMCs. Foam cell formation was evaluated by intracellular lipid droplets, cholesterol contents, and mRNA levels of acyl-coenzyme A: cholesterol acyltransferase 1 (ACAT-1) and ATP binding cassette transporter A-1 (ABCA-1). Monocyte adhesion was evaluated by the number of monocytes adhered to VSMCs and mRNA levels of monocyte chemotactic protein-1 (MCP-1) and vascular cell adhesion molecule-1 (VCAM-1). FNDC5 inhibited oxLDL-induced foam cell formation, monocyte adhesion, ABCA-1 mRNA downregulation, and ACAT-1, MCP-1 and VCAM-1 mRNA upregulation in VSMCs. It inhibited oxLDL-induced p65-NFκB nuclear translocation, NLRP3 upregulation, caspase-1 and IL-1ß production. Inhibition of NFκB with BMS-345541 or inhibition of NLRP3 inflammasome with MCC950 showed similar effects to FNDC5 in attenuating the oxLDL-induced foam cell formation, monocyte adhesion, and caspase-1 and IL-1ß production. The oxLDL-induced NLRP3 upregulation was prevented by BMS-345541 rather than MCC950. These results indicate that FNDC5 inhibits oxLDL-induced foam cell formation and monocyte adhesion in VSMCs via suppressing NFκB-mediated NLRP3 upregulation and IL-1ß production.

FNDC5 attenuates adipose tissue inflammation and insulin resistance via AMPK-mediated macrophage polarization in obesity.

BACKGROUND: Obesity-induced chronic inflammation is critical in the pathogenesis of insulin resistance, and the recruitment and proinflammatory activation of adipose tissue macrophages (ATMs) is important for the development of this process. Here, we examined the effects of fibronectin type III domain-containing 5 (FNDC5) on inflammation and insulin resistance in high-fat diet-induced obese mice. MATERIALS AND METHODS: Male wild-type (WT) and FNDC5-/- mice were fed with standard chow (Ctrl) or high fat diet (HFD) for 20 weeks to induce obesity and insulin resistance. Firstly, effects of FNDC5 gene deletion on obesity, insulin resistance, macrophage accumulation and polarization and adipose tissue inflammation were determined in mice. Secondly, the macrophage polarity shift was further examined with flow cytometry in isolated stromal vascular fraction (SVF). Thirdly, the effects of exogenous FNDC5 on lipopolysaccharide (LPS)-induced macrophage polarization, inflammation and the underlying signaling mechanism were investigated in RAW264.7 macrophages and primary mouse peritoneal cavity macrophages (PMs). Finally, the therapeutic effects of FNDC5 overexpression were examined in HFD-induced obese WT and FNDC5-/- mice. RESULTS: FNDC5 gene deletion aggravated obesity, insulin resistance, fat accumulation and inflammation accompanied with enhanced AMPK inhibition, macrophages recruitment and M1 polarization in mice fed with HFD. Exogenous FNDC5 inhibited LPS-induced M1 macrophage polarization and inflammatory cytokine production via AMPK phosphorylation in both RAW264.7 macrophages and PMs. FNDC5 overexpression attenuated insulin resistance, AMPK inhibition, M1 macrophage polarization and inflammatory cytokine production in adipose tissue of obese WT and FNDC5-/- mice. CONCLUSIONS: FNDC5 attenuates adipose tissue inflammation and insulin resistance via AMPK-mediated macrophage polarization in HFD-induced obesity. FNDC5 plays several beneficial roles in obesity and may be used as a therapeutic regimen for preventing inflammation and insulin resistance in obesity and diabetes.

BCL6 attenuates renal inflammation via negative regulation of NLRP3 transcription.

Renal inflammation contributes to the pathogeneses of hypertension. This study was designed to determine whether B-cell lymphoma 6 (BCL6) attenuates renal NLRP3 inflammasome activation and inflammation and its underlying mechanism. Male spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY) were used in the present study. Angiotensin (Ang) II or lipopolysaccharides (LPS) was used to induce inflammation in HK-2 cells, a human renal tubular epithelial (RTE) cell line. NLRP3 inflammasome was activated and BCL6 was downregulated in the kidneys of SHR. Either Ang II or LPS suppressed BCL6 expression in HK-2 cells. BCL6 overexpression in HK-2 cells attenuated Ang II-induced NLRP3 upregulation, inflammation and cell injury. The inhibitory effects of BCL6 overexpression on NLRP3 expression and inflammation were also observed in LPS-treated HK-2 cells. BCL6 inhibited the NLRP3 transcription via binding to the NLRP3 promoter. BCL6 knockdown with shRNA increased NLRP3 and mature IL-1ß expression levels in both PBS- or Ang II-treated HK-2 cells but had no significant effects on ASC, pro-caspase-1 and pro-IL-1ß expression levels. BCL6 overexpression caused by recombinant lentivirus expressing BCL6 reduced blood pressure in SHR. BCL6 overexpression prevented the upregulation of NLRP3 and mature IL-1ß expression levels in the renal cortex of SHR. The results indicate that BCL6 attenuates Ang II- or LPS-induced inflammation in HK-2 cells via negative regulation of NLRP3 transcription. BCL6 overexpression in SHR reduced blood pressure, NLRP3 expression and inflammation in the renal cortex of SHR.

FNDC5 Alleviates Hepatosteatosis by Restoring AMPK/mTOR-Mediated Autophagy, Fatty Acid Oxidation, and Lipogenesis in Mice.

Fibronectin type III domain-containing 5 (FNDC5) protein induces browning of subcutaneous fat and mediates the beneficial effects of exercise on metabolism. However, whether FNDC5 is associated with hepatic steatosis, autophagy, fatty acid oxidation (FAO), and lipogenesis remains unknown. Herein, we show the roles and mechanisms of FNDC5 in hepatic steatosis, autophagy, and lipid metabolism. Fasted FNDC5-/- mice exhibited severe steatosis, reduced autophagy, and FAO, and enhanced lipogenesis in the liver compared with wild-type mice. Energy deprivation-induced autophagy, FAO, and AMPK activity were attenuated in FNDC5-/- hepatocytes, which were restored by activating AMPK with 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR). Inhibition of mammalian target of rapamycin (mTOR) complex 1 with rapamycin enhanced autophagy and FAO and attenuated lipogenesis and steatosis in FNDC5-/- livers. FNDC5 deficiency exacerbated hyperlipemia, hepatic FAO and autophagy impairment, hepatic lipogenesis, and lipid accumulation in obese mice. Exogenous FNDC5 stimulated autophagy and FAO gene expression in hepatocytes and repaired the attenuated autophagy and palmitate-induced steatosis in FNDC5-/- hepatocytes. FNDC5 overexpression prevented hyperlipemia, hepatic FAO and autophagy impairment, hepatic lipogenesis, and lipid accumulation in obese mice. These results indicate that FNDC5 deficiency impairs autophagy and FAO and enhances lipogenesis via the AMPK/mTOR pathway. FNDC5 deficiency aggravates whereas FNDC5 overexpression prevents the HFD-induced hyperlipemia, hepatic lipid accumulation, and impaired FAO and autophagy in the liver.

ß-aminoisobutyric acid attenuates hepatic endoplasmic reticulum stress and glucose/lipid metabolic disturbance in mice with type 2 diabetes.

ß-aminoisobutyric acid (BAIBA) is a nature thymine catabolite, and contributes to exercise-induced protection from metabolic diseases. Here we show the therapeutical effects of BAIBA on hepatic endoplasmic reticulum (ER) stress and glucose/lipid metabolic disturbance in diabetes. Type 2 diabetes was induced by combined streptozotocin (STZ) and high-fat diet (HFD) in mice. Oral administration of BAIBA for 4 weeks reduced blood glucose and lipids levels, hepatic key enzymes of gluconeogenesis and lipogenesis expressions, attenuated hepatic insulin resistance and lipid accumulation, and improved insulin signaling in type 2 diabetic mice. BAIBA reduced hepatic ER stress and apoptosis in type 2 diabetic mice. Furthermore, BAIBA alleviated ER stress in human hepatocellular carcinoma (HepG2) cells with glucosamine-induced insulin resistance. Hepatic AMPK phosphorylation was reduced in STZ/HFD mice and glucosamine-treated HepG2 cells, which were restored by BAIBA treatment. The suppressive effects of BAIBA on glucosamine-induced ER stress were reversed by knockdown of AMPK with siRNA. In addition, BAIBA prevented thapsigargin- or tunicamycin-induced ER stress, and tunicamycin-induced apoptosis in HepG2 cells. These results indicate that BAIBA attenuates hepatic ER stress, apoptosis and glucose/lipid metabolic disturbance in mice with type 2 diabetes. AMPK signaling is involved to the role of BAIBA in attenuating ER stress.

Relaxin in paraventricular nucleus contributes to sympathetic overdrive and hypertension via PI3K-Akt pathway.

Relaxin is recognized as an ovarian polypeptide hormone. Abundant relaxin binding sites are observed in hypothalamic paraventricular nucleus (PVN). This study was conducted to determine the roles and underlying mechanisms of relaxin in the PVN in sympathetic activation and hypertension in spontaneously hypertensive rats (SHR). Experiments were performed in normotensive Wistar-Kyoto rats (WKY) and SHR. Relaxin and its RXFP1 receptors in PVN were up-regulated in SHR. Relaxin-positive neurons existed in both parvocellular and magnocellular parts of the PVN. Presympathetic neurons and AVP neurons in the PVN expressed RXFP1, but not relaxin. Bilateral PVN microinjection of human relaxin-2 increased but anti-relaxin IgG reduced renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP), plasma norepinephrine (NE) and arginine vasopressin (AVP) levels in SHR. The effects of relaxin-2 on RSNA and MAP were abolished by intravenous infusion of ganglionic blocker hexamethonium, and attenuated by AVP V1 receptor antagonist AAVP. Akt phosphorylation was enhanced in SHR, and relaxin-2 stimulated Akt phosphorylation and p85α subunit of PI3K expression. PI3K inhibitor LY294002 or Akt inhibitor MK-2206 abolished the effects of relaxin-2 on the RSNA, MAP and plasma NE, and attenuated the relaxin-2-induced AVP secretion. STAT5a and polymerase II (Pol II) binding to relaxin-promoter were significantly increased in SHR. Chronic PVN infusion of relaxin-2 with osmotic pumps in normal rats induced sympathetic activation, AVP secretion and hypertension accompanied with cardiovascular remodeling. Relaxin in the PVN contributes to sympathetic overdrive and hypertension via PI3K-Akt pathway.

GABA in Paraventricular Nucleus Regulates Adipose Afferent Reflex in Rats.

BACKGROUND: Chemical stimulation of white adipose tissue (WAT) induces adipose afferent reflex (AAR), and thereby causes a general sympathetic activation. Paraventricular nucleus (PVN) is important in control of sympathetic outflow. This study was designed to investigate the role of γ-aminobutyric acid (GABA) in PVN in regulating the AAR. METHODOLOGY/PRINCIPAL FINDINGS: Experiments were carried out in anesthetized rats. Renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were continuously recorded. AAR was evaluated by the RSNA and MAP responses to electrical stimulation of the right epididymal WAT (eWAT) afferent nerve. Electrical stimulation of eWAT afferent nerve increase RSNA. Bilateral microinjection of the GABAA receptor agonist isoguvacine or the GABAB receptor agonist baclofen attenuated the AAR. The effect of isoguvacine on the AAR was greater than that of baclofen. The GABAA receptor antagonist gabazine enhanced the AAR, while the GABAB receptor antagonist CGP-35348 had no significant effect on the AAR. Bilateral PVN microinjection of vigabatrin, a selective GABA-transaminase inhibitor, to increase endogenous GABA levels in the PVN abolished the AAR. The inhibitory effect of vigabatrin on the AAR was attenuated by the pretreatment with gabazine or CGP-35348. Pretreatment with combined gabazine and CGP-35348 abolished the effects of vigabatrin. CONCLUSIONS: Activation of GABAA or GABAB receptors in the PVN inhibits the AAR. Blockade of GABAA receptors in the PVN enhances the AAR. Endogenous GABA in the PVN plays an important role in regulating the AAR.

FNDC5 overexpression and irisin ameliorate glucose/lipid metabolic derangements and enhance lipolysis in obesity.

Irisin is a cleaved and secreted fragment of fibronectin type III domain containing 5 (FNDC5), and contributes to the beneficial effects of exercise on metabolism. Here we report the therapeutical effects of FNDC5/irisin on metabolic derangements and insulin resistance in obesity, and show the lipolysis effect of irisin and its signal molecular mechanism. In obese mice, lentivirus mediated-FNDC5 overexpression enhanced energy expenditure, lipolysis and insulin sensitivity, and reduced hyperlipidemia, hyperglycemia, hyperinsulinism, blood pressure and norepinephrine levels; it increased hormone-sensitive lipase (HSL) expression and phosphorylation, and reduced perilipin level and adipocyte diameter in adipose tissues. Subcutaneous perfusion of irisin reduced hyperlipidemia and hyperglycemia, and improved insulin resistance. Either FNDC5 overexpression or irisin perfusion only induced a tendency toward a slight decrease in body weight in obese mice. In 3T3-L1 adipocytes, irisin enhanced basal lipolysis rather than isoproterenol-induced lipolysis, which were prevented by inhibition of adenylate cyclase or PKA; irisin increased the HSL and perilipin phosphorylation; it increased PKA activity, and cAMP and HSL mRNA levels, but reduced perilipin expression. These results indicate that FNDC5/irisin ameliorates glucose/lipid metabolic derangements and insulin resistance in obese mice, and enhances lipolysis via cAMP-PKA-HSL/perilipin pathway. FNDC5 or irisin can be taken as an effective therapeutic strategy for metabolic disorders.

Salusin-ß contributes to vascular remodeling associated with hypertension via promoting vascular smooth muscle cell proliferation and vascular fibrosis.

Vascular smooth muscle cell (VSMC) proliferation and vascular fibrosis are closely linked with hypertension and atherosclerosis. Salusin-ß is a bioactive peptide involved in the pathogenesis of atherosclerosis. However, it is still largely undefined whether salusin-ß is a potential candidate in the VSMC proliferation and vascular fibrosis. Experiments were carried out in human vascular smooth muscle cells (VSMCs) and in rats with intravenous injection of lentivirus expressing salusin-ß. In vitro, salusin-ß promoted VSMCs proliferation, which was attenuated by adenylate cyclase inhibitor SQ22536, PKA inhibitor Rp-cAMP, epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor AG1478, ERK inhibitor U0126 or cAMP response element binding protein (CREB) inhibitor KG501. It promoted the phosphorylation of ERK1/2, CREB and EGFR, which were abolished by SQ22536 or Rp-cAMP. Furthermore, epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor AG1478 diminished the salusin-ß-evoked ERK1/2 and CREB phosphorylation. On the other hand, salusin-ß increased collagen-I, collagen-III, fibronectin and connective tissue growth factor (CTGF) mRNA and phosphorylation of Smad2/3, which were prevented by ALK5 inhibitor A83-01. In vivo, salusin-ß overexpression increased the media thickness, media/lumen ratio coupled with ERK1/2, CREB, EGFR and Smad2/3 phosphorylation, as well as the mRNA of collagen-I, collagen-III, fibronectin, transforming growth factor-ß1 (TGF-ß1) and CTGF in arteries. Moreover, salusin-ß overexpression in rats caused severe hypertension. Intravenous injection of salusin-ß dose-relatedly increased blood pressure, but excessive salusin-ß decreased blood pressure and heart rate. These results indicate that salusin-ß promotes VSMC proliferation via cAMP-PKA-EGFR-CREB/ERK pathway and vascular fibrosis via TGF-ß1-Smad pathway. Increased salusin-ß contributes to vascular remodeling and hypertension.

Superoxide anions in paraventricular nucleus modulate adipose afferent reflex and sympathetic activity in rats.

BACKGROUND: Adipose afferent reflex (AAR) is a sympatho-excitatory reflex induced by chemical stimulation of white adipose tissue (WAT). Ionotropic glutamate receptors including NMDA receptors (NMDAR) and non-NMDA receptors (non-NMDAR) in paraventricular nucleus (PVN) mediate the AAR. Enhanced AAR contributes to sympathetic activation and hypertension in obesity rats. This study was designed to investigate the role and mechanism of superoxide anions in PVN in modulating the AAR. METHODOLOGY/PRINCIPAL FINDINGS: Renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded in anesthetized rats. AAR was evaluated by the RSNA and MAP responses to injections of capsaicin into four sites of right inguinal WAT (8.0 nmol in 8.0 µl for each site). Microinjection of polyethylene glycol-superoxide dismutase (PEG-SOD), the superoxide anion scavenger tempol or the NAD(P)H oxidase inhibitor apocynin into the PVN decreased the baseline RSNA and MAP, and attenuated the AAR. Unilateral WAT injection of capsaicin increased superoxide anions in bilateral PVN, which was prevented by the WAT denervation. WAT injection of capsaicin increased superoxide anion level and NAD(P)H oxidase activity in the PVN, which was abolished by the PVN pretreatment with the combined NMDAR antagonist AP5 and non-NMDAR antagonist CNQX. Microinjection of the NMDAR agonist NMDA or the non-NMDAR agonist AMPA increased superoxide anion level and NAD(P)H oxidase activity in the PVN. CONCLUSIONS: NAD(P)H oxidase-derived superoxide anions in the PVN contributes to the tonic modulation of AAR. Activation of ionotropic glutamate receptors in the PVN is involved in the AAR-induced production of superoxide anions in the PVN.

Angiotensin-(1-7) in paraventricular nucleus modulates sympathetic activity and cardiac sympathetic afferent reflex in renovascular hypertensive rats.

BACKGROUND: Excessive sympathetic activity contributes to the pathogenesis and progression of hypertension. Enhanced cardiac sympathetic afferent reflex (CSAR) is involved in sympathetic activation. This study was designed to determine the roles of angiotensin (Ang)-(1-7) in paraventricular nucleus (PVN) in modulating sympathetic activity and CSAR and its signal pathway in renovascular hypertension. METHODOLOGY/PRINCIPAL FINDINGS: Renovascular hypertension was induced with two-kidney, one-clip method. Renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded in sinoaortic-denervated and cervical-vagotomized rats with anesthesia. CSAR was evaluated with the RSNA and MAP responses to epicardial application of capsaicin. PVN microinjection of Ang-(1-7) and cAMP analogue db-cAMP caused greater increases in RSNA and MAP, and enhancement in CSAR in hypertensive rats than in sham-operated rats, while Mas receptor antagonist A-779 produced opposite effects. There was no significant difference in the angiotensin-converting enzyme 2 (ACE2) activity and Ang-(1-7) level in the PVN between sham-operated rats and hypertensive rats, but the Mas receptor protein expression in the PVN was increased in hypertensive rats. The effects of Ang-(1-7) were abolished by A-779, adenylyl cyclase inhibitor SQ22536 or protein kinase A (PKA) inhibitor Rp-cAMP. SQ22536 or Rp-cAMP reduced RSNA and MAP in hypertensive rats, and attenuated the CSAR in both sham-operated and hypertensive rats. CONCLUSIONS: Ang-(1-7) in the PVN increases RSNA and MAP and enhances the CSAR, which is mediated by Mas receptors. Endogenous Ang-(1-7) and Mas receptors contribute to the enhanced sympathetic outflow and CSAR in renovascular hypertension. A cAMP-PKA pathway is involved in the effects of Ang-(1-7) in the PVN.

Enhanced adipose afferent reflex contributes to sympathetic activation in diet-induced obesity hypertension.

We recently found that adipose afferent reflex (AAR) induced by chemical stimulation of white adipose tissue (WAT) increased sympathetic outflow and blood pressure in normal rats. The study was designed to test the hypothesis that AAR contributes to sympathetic activation in obesity hypertension. Male rats were fed with a control diet (12% kcal as fat) or high-fat diet (42% kcal as fat) for 12 weeks to induce obesity hypertension. Stimulation of WAT with capsaicin increased renal sympathetic nerve activity and mean arterial pressure. Both AAR and WAT afferent activity were enhanced in obesity hypertension (OH) compared with obesity nonhypertension (ON) and in ON compared with obesity-resistant or control diet rats. WAT sensory denervation induced by resiniferatoxin caused greater decreases in renal sympathetic nerve activity and mean arterial pressure in OH than ON and in ON than obesity-resistant or control. The depressor effect of resiniferatoxin lasted ≥ 3 weeks in OH. Leptin antagonist in WAT reduced renal sympathetic nerve activity and mean arterial pressure in OH. WAT injection of capsaicin increased plasma renin, angiotensin II, and norepinephrine levels in OH and caused more c-fos expression in paraventricular nucleus in OH than ON and in ON than obesity-resistant or control rats. Inhibiting paraventricular nucleus neurons with lidocaine attenuated renal sympathetic nerve activity in OH and ON, decreased mean arterial pressure in OH, and abolished the capsaicin-induced AAR in all groups. The results indicate that enhanced AAR contributes to sympathetic activation in OH, and paraventricular nucleus plays an important role in the enhanced AAR and sympathetic activation in OH.

Endothelin-1 in paraventricular nucleus modulates cardiac sympathetic afferent reflex and sympathetic activity in rats.

BACKGROUND: Cardiac sympathetic afferent reflex (CSAR) is a positive-feedback, sympathoexcitatory reflex. Paraventricular nucleus (PVN) is an important component of the central neurocircuitry of the CSAR. The present study is designed to determine whether endothelin-1 (ET-1) in the PVN modulates the CSAR and sympathetic activity, and whether superoxide anions are involved in modulating the effects of ET-1 in the PVN in rats. METHODOLOGY/PRINCIPAL FINDINGS: In anaesthetized Sprague-Dawley rats with cervical vagotomy and sinoaortic denervation, renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded. The CSAR was evaluated by the responses of the RSNA and MAP to epicardial application of capsaicin. Microinjection of ET-1 into the bilateral PVN dose-dependently enhanced the CSAR, increased the baseline RSNA and MAP. The effects of ET-1 were blocked by PVN pretreatment with the ET(A) receptor antagonist BQ-123. However, BQ-123 alone had no significant effects on the CSAR, the baseline RSNA and MAP. Bilateral PVN pretreatment with either superoxide anion scavenger tempol or polyethylene glycol-superoxide dismutase (PEG-SOD) inhibited the effects of ET-1 on the CSAR, RSNA and MAP. Microinjection of ET-1 into the PVN increased the superoxide anion level in the PVN, which was abolished by PVN pretreatment with BQ-123. Epicardial application of capsaicin increased superoxide anion level in PVN which was further enhanced by PVN pretreatment with ET-1. CONCLUSIONS: Exogenous activation of ET(A) receptors with ET-1 in the PVN enhances the CSAR, increases RSNA and MAP. Superoxide anions in PVN are involved in the effects of ET-1 in the PVN.

Involvement of enhanced cardiac sympathetic afferent reflex in sympathetic activation in early stage of diabetes.

Cardiac sympathetic afferent reflex (CSAR) is involved in sympathetic activation. The present study was designed to investigate the contribution of enhanced CSAR to sympathetic activation in the early stage of diabetes and the involvement of AT(1) receptors in the paraventricular nucleus (PVN). Diabetes was induced by a single intravenous injection of streptozotocin in rats. Acute experiments were carried out under anesthesia after 3 wk. The CSAR was evaluated by the responses of renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) to epicardial application of capsaicin or bradykinin. Sympathetic activity and CSAR were enhanced in diabetic rats. Plasma norepinephrine and angiotensin II were increased, but the transient receptor potential vanilloid 1 (TRPV1) in the left ventricle wall was not significantly increased in diabetic rats. Pericardial injection of resiniferatoxin to desensitize cardiac afferents or PVN microinjection of lidocaine attenuated the CSAR and decreased the RSNA and MAP in diabetic rats. The AT(1) receptor expression in the PVN increased in diabetic rats. Angiotensin II in the PVN caused greater increases in the RSNA and MAP and enhancement in the CSAR in diabetic rats, which were abolished by the losartan pretreatment. Losartan decreased the RSNA and MAP and attenuated the CSAR in diabetic rats but not in control rats. These results indicate that the CSAR is enhanced in the early stage of diabetic rats, which contributes to the sympathetic activation. AT(1) receptors in the PVN are involved in the enhanced CSAR in diabetic rats.

Sympathetic activation by chemical stimulation of white adipose tissues in rats.

Injection of leptin into white adipose tissue (WAT) increases sympathetic outflow. The present study was designed to determine the effects of capsaicin and other chemicals in WAT on the sympathetic outflow and blood pressure and the roles of WAT afferents and hypothalamic paraventricular nucleus (PVN) in the adipose afferent reflex (AAR). The AAR was induced by injection of capsaicin, bradykinin, adenosine, adenosine triphosphate (ATP), or leptin into inguinal WAT (iWAT) or retroperitoneal WAT (rWAT) in anesthetized rats. The iWAT injection of capsaicin increased the renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) but not the heart rate. Bradykinin, adenosine, or leptin but not ATP in the iWAT caused similar effects to capsaicin on the RSNA and MAP. Intravenous, intramuscular, or intradermal injection of capsaicin had no significant effects on the RSNA and MAP. The effects of capsaicin in rWAT were similar to that in iWAT on the RSNA and MAP. Furthermore, injection of capsaicin into the iWAT increased the WAT afferent nerve activities, WAT efferent nerve activity, and brown adipose tissue efferent nerve activity. The iWAT denervation or chemical lesion of the PVN neurons with kainic acid abolished the AAR induced by the iWAT injection of capsaicin. These results indicate that the stimulation of iWAT afferents with capsaicin, bradykinin, adenosine, or leptin reflexly increases the RSNA and blood pressure. The iWAT afferents and the PVN are involved in the AAR induced by capsaicin in the iWAT.

SOD1 overexpression in paraventricular nucleus improves post-infarct myocardial remodeling and ventricular function.

Excessive sympathetic activation contributes to the progression of chronic heart failure. Reactive oxygen species in paraventricular nucleus (PVN) play an important role in the enhanced sympathetic outflow. This study was designed to determine whether superoxide dismutase 1 (SOD1) overexpression in the PVN attenuated the sympathetic activation and cardiac dysfunction in rats after an episode of myocardial infarction (MI). Adenoviral vectors containing human SOD1 (Ad-SOD) or null adenoviral vectors (Ad-null) were immediately microinjected into the PVN of rats with coronary artery ligation or sham operation. At the eighth week, the SOD1 protein level and activity in the PVN increased while the superoxide anions in the PVN decreased in Ad-SOD rats. The SOD1 overexpression in the PVN prevented the increases in left ventricular end-diastolic pressure and volume, and the decreases in ejection fraction and peak velocities of contraction in MI rats. In addition, there was an attenuation of renal sympathetic nerve activity, cardiac sympathetic afferent reflex and plasma norepinephrine level in MI rats. Furthermore, the SOD1 overexpression in the PVN reduced cardiomyocyte size, collagen deposition and the TUNEL-positive cardiomyocytes in MI rats. These results indicate that the SOD1 overexpression in the PVN attenuates the excessive sympathetic activation, myocardial remodeling, cardiomyocyte apoptosis and ventricular dysfunction in MI rats.

Inhibition of cardiac sympathetic afferent reflex and sympathetic activity by baroreceptor and vagal afferent inputs in chronic heart failure.

BACKGROUND: Cardiac sympathetic afferent reflex (CSAR) contributes to sympathetic activation and angiotensin II (Ang II) in paraventricular nucleus (PVN) augments the CSAR in vagotomized (VT) and baroreceptor denervated (BD) rats with chronic heart failure (CHF). This study was designed to determine whether it is true in intact (INT) rats with CHF and to determine the effects of cardiac and baroreceptor afferents on the CSAR and sympathetic activity in CHF. METHODOLOGY/PRINCIPAL FINDINGS: Sham-operated (Sham) or coronary ligation-induced CHF rats were respectively subjected to BD+VT, VT, cardiac sympathetic denervation (CSD) or INT. Under anesthesia, renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded, and the CSAR was evaluated by the RSNA and MAP responses to epicardial application of capsaicin. Either CSAR or the responses of RSNA, MAP and CSAR to Ang II in PVN were enhanced in CHF rats treated with BD+VT, VT or INT. Treatment with VT or BD+VT potentiated the CSAR and the CSAR responses to Ang II in both Sham and CHF rats. Treatment with CSD reversed the capsaicin-induced RSNA and MAP changes and the CSAR responses to Ang II in both Sham and CHF rats, and reduced the RSNA and MAP responses to Ang II only in CHF rats. CONCLUSIONS: The CSAR and the CSAR responses to Ang II in PVN are enhanced in intact CHF rats. Baroreceptor and vagal afferent activities inhibit CSAR and the CSAR responses to Ang II in intact Sham and CHF rats.