Capsaicin pretreatment attenuates salt-sensitive hypertension by alleviating AMPK/Akt/Nrf2 pathway in hypothalamic paraventricular nucleus.

Background: Long term hypertension seriously promotes target organ damage in the brain and heart, and has increasingly become serious public health problem worldwide. The anti-hypertensive effects of capsaicin has been reported, however, the role and mechanism of capsaicin within the brain on salt-induced hypertension have yet to be elucidated. This study aimed to verify the hypothesis that capsaicin attenuates salt-induced hypertension via the AMPK/Akt/Nrf2 pathway in hypothalamic paraventricular nucleus (PVN). Methods: Dahl salt-sensitive (Dahl S) rats were used as animal model for the present study. Rats were randomly divided into four groups based on their dietary regimen (0.3% normal salt diet and 8% high salt diet) and treatment methods (infusion of vehicle or capsaicin in the PVN). Capsaicin was chronically administered in the PVN throughout the animal experiment phase of the study that lasted 6 weeks. Results: Our results demonstrated that PVN pretreatment with capsaicin can slow down raise of the blood pressure elevation and heart rate (HR) of Dahl S hypertensive rats given high salt diet. Interestingly, the cardiac hypertrophy was significantly improved. Furthermore, PVN pretreatment with capsaicin induced decrease in the expression of mRNA expression of NADPH oxidase-2 (NOX2), inducible nitric oxide synthase (iNOS), NOX4, p-IKKß and proinflammatory cytokines and increase in number of positive cell level for Nrf2 and HO-1 in the PVN of Dahl S hypertensive rats. Additionally, the protein expressions of phosphatidylinositol 3-kinase (p-PI3K) and phosphorylated protein kinase-B (p-AKT) were decreased, phosphorylated adenosine monophosphate-activated protein kinase (p-AMPK) were increased after the PVN pretreatment with capsaicin. Conclusion: Capsaicin pretreatment attenuates salt-sensitive hypertension by alleviating AMPK/Akt/iNOS pathway in the PVN.

Testis specific protein, Y-encoded-like 2 activates JAK2/STAT3 pathway in hypothalamic paraventricular nucleus to sustain hypertension.

BACKGROUND: In the hypothalamic paraventricular nucleus (PVN) of spontaneously hypertensive rats (SHRs), the expression of Testis specific protein, Y-encoded-like 2 (TSPYL2) and the phosphorylation level of Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) are higher comparing with the normotensive Wistar-Kyoto rats (WKY). But how they are involved in hypertension remains unclear. TSPYL2 may interact with JAK2/STAT3 in PVN to sustain the high blood pressure during hypertension. METHODS: Knockdown of TSPYL2 via adeno-associated virus (AAV) carrying shRNA was conducted through bilateral micro-injection into the PVN of SHR and WKY rats. JAK2/STAT3 inhibition was achieved by intraperitoneally or PVN injection of AG490 into the SHRs. Blood pressure (BP), plasma norepinephrine (NE), PVN inflammatory response, and PVN oxidative stress were measured. RESULTS: TSPYL2 knock-down in the PVN of SHRs but not WKYs led to reduced BP and plasma NE, and deactivation of JAK2/STAT3, decreased expression of pro-inflammatory cytokine IL-1ß, and increased expression of anti-inflammatory cytokine IL-10 in the PVN. Meanwhile, AG490 administrated in both ways reduced the blood pressure in the SHRs and deactivated JAK2/STAT3 but failed to change the expression of TSPYL2 in PVN. AG490 also downregulated expression of IL-1ß and upregulated expression of IL-10. Both knockdown of TSPYL2 and inhibition of JAK2/STAT3 can reduce the oxidative stress in the PVN of SHRs. CONCLUSION: JAK2/STAT3 is regulated by TSPYL2 in the PVN of SHRs, and PVN TSPYL2/JAK2/STAT3 is essential for maintaining high blood pressure in the hypertensive rats, making it a potential therapeutic target for hypertension.

Central administration of AICAR attenuates hypertension via AMPK/Nrf2 pathway in the hypothalamic paraventricular nucleus of hypertensive rats.

BACKGROUND: Oxidative stress and inflammatory cytokines in the hypothalamus paraventricular nucleus (PVN) have been implicated in sympathetic nerve activity and the development of hypertension, but the specific mechanisms underlying their production in the PVN remains to be elucidated. Previous studies have demonstrated that activation of nuclear transcription related factor-2 (Nrf2) in the PVN reduced the production of reactive oxygen species (ROS) and inflammatory mediators. Moreover, AMP-activated protein kinase (AMPK), has been observed to decrease ROS and inflammatory cytokine production when activated in the periphery. 5-amino-1-ß-D-ribofuranosyl-imidazole-4-carboxamide (AICAR) is an AMPK agonist. However, little research has been conducted on the role of AMPK in the PVN during hypertension. Therefore, we hypothesized that AICAR in the PVN is involved in regulating AMPK/Nrf2 pathway, affecting ROS and inflammatory cytokine expression, influencing sympathetic nerve activity. METHODS: Adult male Sprague-Dawley rats were utilized to induce two-kidney, one-clip (2K1C) hypertension via constriction of the right renal artery. Bilateral PVN was microinjected with either artificial cerebrospinal fluid or AICAR once a day for 4 weeks. RESULTS: Compared to the SHAM group, the PVN of 2K1C hypertensive rats decreased p-AMPK and p-Nrf2 expression, increased Fra-Like, NAD(P)H oxidase (NOX)2, NOX4, tumor necrosis factor-α and interleukin (IL)-1ß expression, elevated ROS levels, decreased superoxide dismutase 1 and IL-10 expression, and elevated plasma norepinephrine levels. Bilateral PVN microinjection of AICAR significantly ameliorated these changes. CONCLUSION: These findings suggest that repeated injection of AICAR in the PVN suppresses ROS and inflammatory cytokine production through the AMPK/Nrf2 pathway, reducing sympathetic nerve activity and improving hypertension.

Roux-en-Y Gastric Bypass Improves Insulin Sensitivity in Obese Rats with Type 2 Diabetes Mellitus by Regulating the Grin3a/AMPK Signal Axis in Hypothalamic Arcuate Nucleus.

Objective: The objective of this study was to explore the effects and related mechanisms of Roux-en-Y gastric bypass (RYGB) on insulin sensitivity in obese rats with type 2 diabetes mellitus (T2DM). Methods: The obese T2DM rat model was constructed by feeding a high-fat diet and injecting streptozotocin (STZ), and treated with RYGB. Grin3a shRNA was injected into the bilateral hypothalamic arcuate nucleus (ARC) to knockdown the Grin3a expression on T2DM rats. Eight weeks after operation, the body weight, fasting blood glucose (FBG), fasting serum insulin (FSI), homeostatic model assessment of insulin resistance (HOMA-IR), and plasma triglyceride (TG) levels were assessed. Hematoxylin & eosin (H&E) staining was adopted to observe the white adipose tissue (WAT) of rats. Western blot and qRT-PCR were used to detect the expression of Grin3a, adenosine 5' monophosphate-activated protein kinase (AMPK) and p-AMPK in ARC of rats. Later, the plasmid over-expressing or knocking down Grin3a was transfected into differentiated 3T3-L1 adipocytes, and the TG level and the formation of lipid droplets in adipocyte were assessed by TG kit and oil red O staining. The expression of lipogenic transcription factors in cells was detected by qRT-PCR. Results: RYGB reduced FBG, FSI, HOMA-IR and plasma TG levels in T2DM rats while increasing Grin3a expression and p-AMPK/AMPK ratio in ARC. Knockdown of Grin3a not only reversed the decrease of FBG, FSI, HOMA-IR and plasma TG levels in T2DM rats induced by RYGB, but also reversed the up-regulation of p-AMPK/AMPK ratio in ARC affected by RYGB. Moreover, knocking down Grin3a significantly increased the TG level, promoted the formation of lipid droplets and up-regulated the expressions of lipogenic transcription factors in adipocytes. Conclusion: RYGB improved the insulin sensitivity, reduced the plasma TG level and lessens the fat accumulation in obese T2DM rats by regulating the Grin3a/AMPK signal in ARC.

Hypothalamic Paraventricular Nucleus Hydrogen Sulfide Exerts Antihypertensive Effects in Spontaneously Hypertensive Rats via the Nrf2 Pathway.

BACKGROUND: Hydrogen sulfide (H2S) is widely distributed throughout the nervous system with various antioxidant and anti-inflammatory properties. Hypertension involves an increase in reactive oxygen species (ROS) and inflammation in the hypothalamic paraventricular nucleus (PVN). However, it is unclear how H2S in PVN affects hypertension. METHODS: Our study used spontaneously hypertensive rats (SHR) and control Wistar Kyoto (WKY) rats, microinjected with adenovirus-associated virus (AAV)-CBS (cystathionine beta-synthase overexpression) or AAV-ZsGreen in bilateral PVN, or simultaneously injected with virus-carrying nuclear factor erythroid 2-related factor 2 (Nrf2)-shRNA for 4 weeks. Blood pressure (BP) and plasma noradrenaline level were detected, and the PVN was collected. Finally, levels of CBS, H2S, Nrf2, Fra-LI, ROS, gp91phox, p47phox, superoxide dismutase 1, interleukin (IL)-1ß, IL-6, IL-10, tumor necrosis factor-α, tyrosine hydroxylase, and glutamate decarboxylase 67 were measured. RESULTS: We found that AAV-CBS increased H2S in the PVN, and BP, neuronal activation, oxidative stress, and inflammation of PVN were substantially reduced. Furthermore, endogenous H2S in the PVN activated Nrf2 and corrected the PVN's imbalance of excitatory and inhibitory neurotransmitters. However, Nrf2 knockdown in the PVN was similarly observed to abolish the beneficial effect of H2S on hypertension. CONCLUSIONS: The findings imply that endogenous H2S in SHR PVN is reduced, and PVN endogenous H2S can alleviate hypertension via Nrf2-mediated antioxidant and anti-inflammatory effects.

Luteolin Attenuates Hypertension via Inhibiting NF-κB-Mediated Inflammation and PI3K/Akt Signaling Pathway in the Hypothalamic Paraventricular Nucleus.

BACKGROUND: Luteolin is widely distributed among a number of vegetal species worldwide. The pharmacological effects of luteolin are diverse and amongst antioxidant, free radical scavenging, and anti-inflammatory activities. Preliminary study showed that luteolin can ameliorate hypertension. However, the precise mechanism needs further investigation. There is no evidence that luteolin affects the paraventricular nucleus of the hypothalamus (PVN), a brain nucleus associated with a critical neural regulator of blood pressure. Our main aim was to explore the effect of luteolin on the PI3K/Akt/NF-κB signaling pathway within the PVN of hypertensive rats. METHODS: spontaneously hypertensive rats (SHRs) and corresponding normotensive control rats, the Wistar Kyoto (WKY) rats were divided into four groups and subsequently treated for 4 weeks with bilateral PVN injections of either luteolin (20 µg/0.11 µL, volume: 0.11 µL/h) or vehicle (artificial cerebrospinal fluid). RESULTS: luteolin infusion to the PVN significantly decreased some hemodynamic parameters including the mean arterial pressure (MAP), heart rate (HR), circulating plasma norepinephrine (NE) and epinephrine (EPI). Additionally, there was a decrease in the expressions of the phosphatidylinositol 3-kinase (p-PI3K) and phosphorylated protein kinase-B (p-AKT), levels of reactive oxygen species (ROS), NAD(P)H oxidase subunit (NOX2, NOX4) in the PVN of SHRs. Meanwhile, the expression of inflammatory cytokines and the activity of nuclear factor κB (NF-κB) p65 in the PVN of SHRs were lowered. Furthermore, immunofluorescence results showed that injection of luteolin in the PVN reduced the expression of tyrosine hydroxylase (TH), and increased that of superoxide dismutase (SOD1) and the 67-kDa isoform of glutamate decarboxylase (GAD67) in the PVN of SHRs. CONCLUSION: Our novel findings revealed that luteolin lowered hypertension via inhibiting NF-κB-mediated inflammation and PI3K/Akt signaling pathway in the PVN.

Resveratrol in the Hypothalamic Paraventricular Nucleus Attenuates Hypertension by Regulation of ROS and Neurotransmitters.

BACKGROUND: The hypothalamic paraventricular nucleus (PVN) is an important nucleus in the brain that plays a key role in regulating sympathetic nerve activity (SNA) and blood pressure. Silent mating-type information regulation 2 homolog-1 (sirtuin1, SIRT1) not only protects cardiovascular function but also reduces inflammation and oxidative stress in the periphery. However, its role in the central regulation of hypertension remains unknown. It is hypothesized that SIRT1 activation by resveratrol may reduce SNA and lower blood pressure through the regulation of intracellular reactive oxygen species (ROS) and neurotransmitters in the PVN. METHODS: The two-kidney one-clip (2K1C) method was used to induce renovascular hypertension in male Sprague-Dawley rats. Then, bilaterally injections of vehicle (artificial cerebrospinal fluid, aCSF, 0.4 µL) or resveratrol (a SIRT1 agonist, 160 µmol/L, 0.4 µL) into rat PVN were performed for four weeks. RESULTS: PVN SIRT1 expression was lower in the hypertension group than the sham surgery (SHAM) group. Activated SIRT1 within the PVN lowered systolic blood pressure and plasma norepinephrine (NE) levels. It was found that PVN of 2K1C animals injected with resveratrol exhibited increased expression of SIRT1, copper-zinc superoxide dismutase (SOD1), and glutamic acid decarboxylase (GAD67), as well as decreased activity of nuclear factor-kappa B (NF-κB) p65 and NAD(P)H oxidase (NOX), particularly NOX4. Treatment with resveratrol also decreased expression of ROS and tyrosine hydroxylase (TH). CONCLUSION: Resveratrol within the PVN attenuates hypertension via the SIRT1/NF-κB pathway to decrease ROS and restore the balance of excitatory and inhibitory neurotransmitters.

The Association of lncRNA and mRNA Changes in Adipose Tissue with Improved Insulin Resistance in Type 2 Obese Diabetes Mellitus Rats after Roux-en-Y Gastric Bypass.

Objective: Roux-en-Y gastric bypass (RYGB) has shown good effects in improving obesity and type II diabetes mellitus (T2DM), but the underlying mechanisms remain unclear. This study explored the changes of related lncRNAs, mRNAs, and signaling pathways in white adipose tissue of T2DM rats after RYGB based on RNA-Seq sequencing, with the aim to provide a theoretical basis for RYGB treatment. Methods: T2DM rat models were established by continuous feeding with a high-fat diet and injection of streptozotocin (STZ), after which they underwent RYGB or sham surgery. After the surgery, their body weight was measured weekly. Their fasting blood glucose (FBG) and fasting serum insulin (FSI) were also measured. A homeostasis model assessment of insulin resistance (HOMA-IR) was calculated at weeks 0, 8, and 12. Besides, white adipose tissue of T2DM rats was collected for RNA-Seq sequencing and validated by qRT-PCR. A series of bioinformatics analyses, such as differential expression genes (DEGs) screening, was performed. GO and KEGG functional enrichment analysis and protein-protein interaction (PPI) network construction were conducted based on the sequencing data. Results: RYGB surgery could significantly inhibit the weight growth rate and decrease the FBG, FSI, and HOMA-IR of T2DM rats. Bioinformatics analysis of RNA sequencing (RNA-Seq) results revealed that 87 DE- lncRNAs (49 upregulated and 38 downregulated) and 1,824 DEGs (896 upregulated and 928 downregulated) were present in between the RYGB group and Sham group. GO and KEGG analysis showed that the target genes of DEGs and differentially expressed lncRNAs (DE-lncRNAs) were mainly associated with amino acid metabolism, fatty acid metabolism, channel activity, and other processes. In addition, the PPI network diagram also displayed that genes such as Fasn, Grin3a, and Nog could be key genes playing a role after RYGB. qRT-PCR showed that the expression level of Grin3a in the RYGB group was significantly increased compared with the Sham group, while the expression of Fasn and Nog was significantly decreased, which was consistent with the sequencing results. Conclusion: Using RNA-Seq sequencing, this study revealed the changes of related lncRNAs, mRNAs, and signaling pathways in the white adipose tissue of T2DM rats after RYGB and identified Fasn, Grin3a, and Nog as potential key genes to function after RYGB.

Blockade of Microglial Activation in Hypothalamic Paraventricular Nucleus Improves High Salt-Induced Hypertension.

BACKGROUND: It has been shown that activated microglia in brain releasing proinflammatory cytokines (PICs) contribute to the progression of cardiovascular diseases. In this study, we tested the hypothesis that microglial activation in hypothalamic paraventricular nucleus (PVN), induced by high-salt diet, increases the oxidative stress via releasing PICs and promotes sympathoexcitation and development of hypertension. METHODS: High-salt diet was given to male Dahl salt-sensitive rats to induce hypertension. Those rats were bilaterally implanted with cannula for PVN infusion of minocycline, a selective microglial activation blocker, or artificial cerebrospinal fluid for 4 weeks. RESULTS: High-salt diet elevated mean arterial pressure of Dahl salt-sensitive rats. Meanwhile, elevations of renal sympathetic nerve activity and central prostaglandin E2, as well as increase of plasma norepinephrine, were observed in those hypertensive rats. Tumor necrosis factor-α, interleukin-1ß (IL-1ß), and IL-6 increased in the PVN of those rats, associated with a significant activation of microglia and prominent disruption of redox balance, which was demonstrated by higher superoxide and NAD(P)H oxidase 2 (NOX-2) and NAD(P)H oxidase 4 (NOX-4), and lower Cu/Zn superoxide dismutase in PVN. PVN infusion of minocycline attenuated all hypertension-related alterations described above. CONCLUSION: This study indicates that high salt leads to microglial activation within PVN of hypertensive rats, and those activated PVN microglia release PICs and trigger the production of reactive oxygen species, which contributes to sympathoexcitation and development of hypertension. Blockade of PVN microglial activation inhibits inflammation and oxidative stress, therefore attenuating the development of hypertension induced by high-salt diet.

Effects of Nrf1 in Hypothalamic Paraventricular Nucleus on Regulating the Blood Pressure During Hypertension.

The incidence rate and mortality of hypertension increase every year. Hypothalamic paraventricular nucleus (PVN) plays a critical role on the pathophysiology of hypertension. It has been demonstrated that the imbalance of neurotransmitters including norepinephrine (NE), glutamate (Glu) and γ-aminobutyric acid (GABA) are closely related to sympathetic overactivity and pathogenesis of hypertension. N-methyl-D-aspartate receptor (NMDAR), consisting of GluN1 and GluN2 subunits, is considered to be a glutamate-gated ion channel, which binds to Glu, and activates neuronal activity. Studies have found that the synthesis of respiratory chain enzyme complex was affected and mitochondrial function was impaired in spontaneously hypertensive rats (SHR), further indicating that mitochondria is associated with hypertension. Nuclear respiratory factor 1 (Nrf1) is a transcription factor that modulates mitochondrial respiratory chain and is related to GluN1, GluN2A, and GluN2B promoters. However, the brain mechanisms underlying PVN Nrf1 modulating sympathoexcitation and blood pressure during the development of hypertension remains unclear. In this study, an adeno-associated virus (AAV) vector carrying the shRNA targeting rat Nrf1 gene (shNrf1) was injected into bilateral PVN of male rats underwent two kidneys and one clip to explore the role of Nrf1 in mediating the development of hypertension and sympathoexcitation. Administration of shNrf1 knocked down the expression of Nrf1 and reduced the expression of excitatory neurotransmitters, increased the expression of inhibitory neurotransmitters, and reduced the production of reactive oxygen species (ROS), and attenuated sympathoexcitation and hypertension. The results indicate that knocking down Nrf1 suppresses sympathoexcitation in hypertension by reducing PVN transcription of NMDAR subunits (GluN1, GluN2A, and GluN2B), rebalancing PVN excitatory and inhibitory neurotransmitters, inhibiting PVN neuronal activity and oxidative stress, and attenuating sympathetic activity.

Acetylcholinesterase inhibition with Pyridostigmine attenuates hypertension and neuroinflammation in the paraventricular nucleus in rat model for Preeclampsia.

Preeclampsia (PE) is characterized by hypertension, autonomic imbalance and inflammation. The subfornical organ (SFO) reportedly relays peripheral inflammatory mediator's signals to the paraventricular nucleus (PVN), a brain autonomic center shown to mediate hypertension in hypertensive rat but not yet in PE rat models. Additionally, we previously showed that Pyridostigmine (PYR), an acetylcholinesterase inhibitor, attenuated placental inflammation and hypertension in PE models. In this study, we investigated the effect of PYR on the activities of these brain regions in PE model. PYR (20 mg/kg/day) was administered to reduced uterine perfusion pressure (RUPP) Sprague-Dawley rat from gestational day (GD) 14 to GD19. On GD19, the mean arterial pressure (MAP) was recorded and samples were collected for analysis. RUPP rats exhibited increased MAP (P = 0.0025), elevated circulating tumor necrosis factor-α (TNF-α, P = 0.0075), reduced baroreflex sensitivity (BRS), increased neuroinflammatory markers including TNF-α, interleukin-1ß (IL-1ß), microglial activation (P = 0.0039), oxidative stress and neuronal excitation within the PVN and the SFO. Changes in MAP, in molecular and cellular expression induced by RUPP intervention were improved by PYR. The ability of PYR to attenuate TNF-α mediated central effect was evaluated in TNF-α-infused pregnant rats. TNF-α infusion-promoted neuroinflammation in the PVN and SFO in dams was abolished by PYR. Collectively, our data suggest that PYR improves PE-like symptoms in rat by dampening placental ischemia and TNF-α-promoted inflammation and pro-hypertensive activity in the PVN. This broadens the therapeutical potential of PYR in PE.

miR-31-5p Promotes Oxidative Stress and Vascular Smooth Muscle Cell Migration in Spontaneously Hypertensive Rats via Inhibiting FNDC5 Expression.

Oxidative stress and the migration of vascular smooth muscle cells (VSMCs) are important for vascular remodeling in a variety of vascular diseases. miR-31-5p promotes cell migration in colorectal cancer cells but inhibits cell migration in renal cell carcinoma. However, whether miR-31-5p is involved in oxidative stress and VSMC migration remains unknown. This study shows the crucial roles of miR-31-5p in oxidative stress and VSMC migration, as well as underlying mechanisms. Experiments were carried out in primary VSMCs from aortic media of Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR), as well as the A7r5 cell line. Oxidative stress was assessed by NADPH oxidase (NOX) expression, NOX activity, and reactive oxygen species (ROS) production. Cell migration was evaluated with a Boyden chamber assay and a wound healing assay. The miR-31-5p mimic and inhibitor promoted and attenuated oxidative stress and cell migration in the VSMCs of SHR, respectively. A dual-luciferase reporter assay indicated that miR-31-5p targeted the 3'UTR domain of FNDC5. The miR-31-5p level was raised and FNDC5 expression was reduced in the VSMCs of SHR compared with those of WKY. The miR-31-5p mimic reduced FNDC5 expression in the A7r5 cells and the VSMCs of both WKY and SHR, while the miR-31-5p inhibitor only increased FNDC5 expression in the VSMCs of SHR. Exogenous FNDC5 attenuated not only the oxidative stress and VSMC migration in SHR but also the roles of the miR-31-5p mimic in inducing oxidative stress and VSMC migration. These results indicate that miR-31-5p promotes oxidative stress and VSMC migration in SHR via inhibiting FNDC5 expression. The increased miR-31-5p and reduced FNDC5 in the VSMCs of SHR contribute to enhanced oxidative stress and cell migration.

Paraventricular Nucleus Infusion of Oligomeric Proantho Cyanidins Improves Renovascular Hypertension.

Oxidative stress plays an important role in the pathogenesis of hypertension. Oligomeric proantho cyanidins (OPC) is the main polyphenol presents in grape seed and is known for its potent antioxidant and anti-inflammatory properties. In the present study, we hypothesize that OPC can attenuate oxidative stress in the paraventricular nucleus of hypothalamus (PVN), ameliorate neurotransmitter imbalance, decrease the blood pressure and sympathetic activity in renovascular hypertensive rats. After induction of renovascular hypertension by the two-kidney one-clip (2K-1C) method, male Sprague-Dawley rats received chronic bilateral PVN infusion of OPC (20 µg/h) or vehicle via osmotic minipump for 4 weeks. We found that hypertension induced by 2K-1C was associated with the production of reactive oxygen species (ROS) in the PVN. Infusion of OPC in the PVN significantly reduced the systolic blood pressure and norepinephrine in plasma of 2K-1C rats. In addition, PVN infusion of OPC decreased the level of ROS and the expression of stress-related nicotinamide adenine dinucleotide phosphate (NADPH) oxidases subunit NOX4, increased the levels of nuclear factor E2-related factor 2 (Nrf2) and antioxidant enzyme, balanced the content of cytokines, increased expression of glutamic acid decarboxylase and decreased the expression of tyrosine hydroxylase in the PVN of 2K-1C rats. Our findings provided strong evidence that PVN infusion of OPC inhibited the progression of renovascular hypertension through its potent anti-oxidative and anti-inflammatory function in the PVN.

Nrf1 Knock-Down in the Hypothalamic Paraventricular Nucleus Alleviates Hypertension Through Intervention of Superoxide Production-Removal Balance and Mitochondrial Function.

Oxidative stress in the hypothalamic paraventricular nucleus (PVN) contributes greatly to the development of hypertension. The recombinant nuclear respiratory factor 1 (Nrf1) regulates the transcription of several genes related to mitochondrial respiratory chain function or antioxidant expression, and thus may be involved in the pathogenesis of hypertension. Here we show that in the two-kidney, one-clip (2K1C) hypertensive rats the transcription level of Nrf1 was elevated comparing to the normotensive controls. Knocking down of Nrf1 in the PVN of 2K1C rats can significantly reduce their blood pressure and level of plasma norepinephrine (NE). Analysis revealed significant reduction of superoxide production level in both whole cell and mitochondria, along with up-regulation of superoxide dismutase 1 (Cu/Zn-SOD), NAD(P)H: quinone oxidoreductase 1 (NQO1), thioredoxin-dependent peroxiredoxin 3 (Prdx3), cytochrome c (Cyt-c) and glutathione synthesis rate-limiting enzyme (glutamyl-cysteine ligase catalytic subunit (Gclc) and modifier subunit (Gclm)), and down-regulation of cytochrome c oxidase subunit VI c (Cox6c) transcription after Nrf1 knock-down. In addition, the reduced ATP production and elevated mitochondrial membrane potential in the PVN of 2K1C rats were reinstated with Nrf1 knock-down, together with restored expression of peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α), mitochondrial transcription factor A (Tfam), coiled-coil myosin-like BCL2-interacting protein (Beclin1), and Mitofusin 1 (Mfn1), which are related to the mitochondrial biogenesis, fusion, and autophagy. Together, the results indicate that the PVN Nrf1 is associated with the development of 2K1C-induced hypertension, and Nrf1 knock-down in the PVN can alleviate hypertension through intervention of mitochondrial function and restorement of the production-removal balance of superoxide.

Antihypertensive effects of exercise involve reshaping of gut microbiota and improvement of gut-brain axis in spontaneously hypertensive rat.

Exercise (Ex) has long been recognized to produce beneficial effects on hypertension (HTN). This coupled with evidence of gut dysbiosis and an impaired gut-brain axis led us to hypothesize that reshaping of gut microbiota and improvement in impaired gut-brain axis would, in part, be associated with beneficial influence of exercise. Male spontaneously hypertensive rats (SHR) and Wistar Kyoto (WKY) rats were randomized into sedentary, trained, and detrained groups. Trained rats underwent moderate-intensity exercise for 12 weeks, whereas, detrained groups underwent 8 weeks of moderate-intensity exercise followed by 4 weeks of detraining. Fecal microbiota, gut pathology, intestinal inflammation, and permeability, brain microglia and neuroinflammation were analyzed. We observed that exercise training resulted in a persistent decrease in systolic blood pressure in the SHR. This was associated with increase in microbial α diversity, altered ß diversity, and enrichment of beneficial bacterial genera. Furthermore, decrease in the number of activated microglia, neuroinflammation in the hypothalamic paraventricular nucleus, improved gut pathology, inflammation, and permeability were also observed in the SHR following exercise. Interestingly, short-term detraining did not abolish these exercise-mediated improvements. Finally, fecal microbiota transplantation from exercised SHR into sedentary SHR resulted in attenuated SBP and an improved gut-brain axis. These observations support our concept that an impaired gut-brain axis is linked to HTN and exercise ameliorates this impairment to induce antihypertensive effects.

Interleukin-1ß in hypothalamic paraventricular nucleus mediates excitatory renal reflex.

Chemical stimulation of kidney causes sympathetic activation and pressor responses in rats. The excitatory renal reflex (ERR) is mediated by angiotensin type 1 receptor (AT1R) and superoxide anions in hypothalamic paraventricular nucleus (PVN). The aim of this study is to determine whether interleukin-1ß (IL-1ß) in the PVN mediates the ERR, and whether the IL-1ß production in the PVN is dependent on the AT1R-superoxide anion signaling. Experiments were performed in adult rats under anesthesia. The ERR was induced by renal infusion of capsaicin, and evaluated by the responses of the contralateral renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP). Inhibition of IL-1ß production with MCC950 in the PVN dose-dependently inhibited the capsaicin-induced ERR and sympathetic activation. The PVN microinjection of IL-1 receptor antagonist IL-1Ra or specific IL-1ß antibody abolished the capsaicin-induced ERR, while IL-1ß enhanced the ERR. Renal infusion of capsaicin promoted p65-NFκB phosphorylation and IL-1ß production in the PVN, which were prevented by PVN microinjection of NADPH oxidase inhibitor apocynin or the superoxide anion scavenger tempol. The PVN microinjection of NFκB inhibitor BMS-345541 abolished the capsaicin induced-ERR and IL-1ß production, but not the NADPH oxidase activation and superoxide anion production. Furthermore, capsaicin-induced p65-NFκB phosphorylation and IL-1ß production in the PVN were prevented by AT1R antagonist losartan, or angiotensin converting enzyme inhibitor captopril. These results indicate that capsaicin-induced ERR and sympathetic activation are mediated by IL-1ß in the PVN. The IL-1ß production in the PVN is dependent on the AT1R-mediated superoxide anion generation and NFκB activation.

MiR155-5p Inhibits Cell Migration and Oxidative Stress in Vascular Smooth Muscle Cells of Spontaneously Hypertensive Rats.

Migration of vascular smooth muscle cells (VSMCs) is essential for vascular reconstruction in hypertension and several vascular diseases. Our recent study showed that extracellular vesicles derived from vascular adventitial fibroblasts of normal rats inhibited VSMC proliferation by delivering miR155-5p to VSMCs. It is unknown whether miR155-5p inhibits cell migration and oxidative stress in VSMCs of spontaneously hypertensive rats (SHR) and in angiotensin II (Ang II)-treated VSMCs. The purpose of this study was to determine the role of miR155-5p in VSMC migration and its underlying mechanisms. Primary VSMCs were isolated from the aortic media of Wistar-Kyoto rats (WKY) and SHR. Wound healing assay and Boyden chamber assay were used to evaluate VSMC migration. A miR155-5p mimic inhibited, and a miR155-5p inhibitor promoted the migration of VSMC of SHR but had no significant effect on the migration of VSMC of WKY. The miR155-5p mimic inhibited angiotensin-converting enzyme (ACE) mRNA and protein expression in VSMCs. It also reduced superoxide anion production, NAD(P)H oxidase (NOX) activity, as well as NOX2, interleukin-1ß (IL-1ß), and tumor necrosis factor α (TNF-α) expression levels in VSMCs of SHR but not in VSMCs of WKY rats. Overexpression of miR155-5p inhibited VSMC migration and superoxide anion and IL-1ß production in VSMCs of SHR but had no impact on exogenous Ang II-induced VSMC migration and on superoxide anion and IL-1ß production in WKY rats and SHR. These results indicate that miR155-5p inhibits VSMC migration in SHR by suppressing ACE expression and its downstream production of Ang II, superoxide anion, and inflammatory factors. However, miR155-5p had no effects on exogenous Ang II-induced VSMC migration.

Angiotensin Type 1 Receptors and Superoxide Anion Production in Hypothalamic Paraventricular Nucleus Contribute to Capsaicin-Induced Excitatory Renal Reflex and Sympathetic Activation.

Chemical stimulation of the kidney increases sympathetic activity and blood pressure in rats. The hypothalamic paraventricular nucleus (PVN) is important in mediating the excitatory renal reflex (ERR). In this study, we examined the role of molecular signaling in the PVN in mediating the capsaicin-induced ERR and sympathetic activation. Bilateral PVN microinjections were performed in rats under anesthesia. The ERR was elicited by infusion of capsaicin into the cortico-medullary border of the right kidney. The reflex was evaluated as the capsaicin-induced changes in left renal sympathetic nerve activity and mean arterial pressure. Blockade of angiotensin type 1 receptors with losartan or inhibition of angiotensin-converting enzyme with captopril in the PVN abolished the capsaicin-induced ERR. Renal infusion of capsaicin significantly increased NAD(P)H oxidase activity and superoxide anion production in the PVN, which were prevented by ipsilateral renal denervation or microinjection of losartan into the PVN. Furthermore, either scavenging of superoxide anions or inhibition of NAD(P)H oxidase in the PVN abolished the capsaicin-induced ERR. We conclude that the ERR induced by renal infusion of capsaicin is mediated by angiotensin type 1 receptor-related NAD(P)H oxidase activation and superoxide anion production within the PVN.

Chemical Stimulation of Renal Tissue Induces Sympathetic Activation and a Pressor Response via the Paraventricular Nucleus in Rats.

Sympathetic activation and the kidney play critical roles in hypertension and chronic heart failure. The role of the kidney in sympathetic activation is still not well known. In this study, we revealed an excitatory renal reflex (ERR) in rats induced by chemical stimulation of the kidney that regulated sympathetic activity and blood pressure. The ERR was induced by renal infusion of capsaicin, and evaluated by the changes in renal sympathetic outflow, blood pressure, and heart rate. Renal infusion of capsaicin dose-dependently increased the contralateral renal sympathetic nerve activity, mean arterial pressure, and heart rate. Capsaicin in the cortico-medullary border had greater effects than in the cortex or medulla. Intravenous infusion of capsaicin had no significant effects. The effects of renal infusion of capsaicin were abolished by ipsilateral renal denervation, but were not affected by bilateral sinoaortic denervation. Renal infusion of capsaicin increased the ipsilateral renal afferent activity. The ERR was also induced by renal infusion of bradykinin, adenosine, and angiotensin II, but not by ATP. Renal infusion of capsaicin increased c-Fos expression in the paraventricular nucleus (PVN) of hypothalamus. Lesion of neurons in the PVN with kainic acid abolished the capsaicin-induced ERR. These findings indicate that chemical stimulation of kidney causes an excitatory reflex, leading to sympathetic activation, pressor response, and accelerated heart rate. The PVN is an important central nucleus in the pathway of the ERR.

Blockade of c-Src Within the Paraventricular Nucleus Attenuates Inflammatory Cytokines and Oxidative Stress in the Mechanism of the TLR4 Signal Pathway in Salt-Induced Hypertension.

Toll-like receptor 4 (TLR4) and cellular Src (c-Src) are closely associated with inflammatory cytokines and oxidative stress in hypertension, so we designed this study to explore the exact role of c-Src in the mechanism of action of the TLR4 signaling pathway in salt-induced hypertension. Salt-sensitive rats were given a high salt diet for 10 weeks to induce hypertension. This resulted in higher levels of TLR4, activated c-Src, pro-inflammatory cytokines, oxidative stress, and arterial pressure. Infusion of a TLR4 blocker into the hypothalamic paraventricular nucleus (PVN) decreased the activated c-Src, while microinjection of a c-Src inhibitor attenuated the PVN levels of nuclear factor-kappa B, pro-inflammatory cytokines, and oxidative stress. Our findings suggest that a long-term high-salt diet increases TLR4 expression in the PVN and this promotes the activation of c-Src, which upregulates the expression of pro-inflammatory cytokines and results in the overproduction of reactive oxygen species. Therefore, inhibiting central c-Src activity may be a new target for treating hypertension.