Medicinal Tiger Milk Mushroom Lignosus rhinocerus TM02® (Agaricomycetes) Sclerotia Supplementation Mitigates Hypertension and Alleviates Vascular Dysfunction Partly through Oxidative Stress Modulation in Spontaneously Hypertensive Rats.

Hypertension is a risk factor for cardiovascular diseases such as coronary artery disease, heart failure, and stroke. Lignosus rhinocerus (Cooke) Ryvarden (also known as tiger milk mushroom), has been reported to exhibit a range of pharmacological effects, such as anti-inflammatory, anti-proliferative, antioxidative, immunomodulatory and anti-asthmatic activities. Thus far, there is limited research that has explored its ability to mediate vascular effects in vivo. Therefore, this study investigated the antihypertensive and vascular protective effects of L. rhinocerus TM02® sclerotia supplementation in spontaneously hypertensive rats (SHR). Wistar Kyoto (WKY) rats served as a normotensive control group. SHR were orally administered with L. rhinocerus TM02® sclerotia (100 mg/kg and 300 mg/kg, respectively) for 8 weeks, and blood pressure was monitored every 2 weeks. Vascular function was evaluated using an organ bath (aorta) and wire myograph (renal artery) at the treatment endpoint. The levels of reactive oxygen species (ROS) and nitric oxide (NO) in the aorta and renal artery were evaluated using dihydroethidium (DHE) and difluoro fluorescein acetate (DAF-FM) fluorescence assays, respectively. Total plasma nitrate/nitrite and tumor necrosis factor alpha (TNF-α) levels were evaluated via colorimetric assays. In vivo treatment with L. rhinocerus TM02® sclerotia significantly attenuated the increase in systolic blood pressure (SBP). It also alleviated vascular dysfunction and decreased elevated ROS in the aorta and renal arteries of the treated SHRs. Moreover, L. rhinocerus TM02® sclerotia attenuated plasma TNF-α level but increased total plasma nitrate/nitrite, albeit slightly, coupled with significantly increased NO at the vascular level. Collectively, the present study demonstrated that L. rhinocerus TM02® sclerotia supplementation exerted blood pressure lowering effects, partly attributed to improvements in vascular function via reduction in vascular oxidative stress.

Mechanisms of retinal photoreceptor loss in spontaneously hypertensive rats.

Retinal neurodegenerative diseases, including hypertensive retinopathy, involve progressive damage to retinal neurons, leading to visual impairment. In this study, we investigated the pathological mechanisms underlying retinal neurodegeneration in spontaneously hypertensive rats (SHR), using Wistar Kyoto (WKY) rats as normotensive controls. We observed that SHR exhibited significantly higher blood pressure and decreased retinal thickness, indicating retinal neurodegeneration. Molecular tests including quantitative real-time polymerase chain reaction, immunoblot, and immunofluorescent staining showed elevated levels of the pro-inflammatory cytokine tumor necrosis factor-α, apoptotic markers (Fas, FasL, caspase-8, active caspase-3, and cleaved poly (ADP-ribose) polymerase), and necroptotic markers (receptor-interacting protein kinase-1 and -3) in SHR retinas. Additionally, we found elevated transforming growth factor-ß (TGF-ß) levels in the retinal pigment epithelium (RPE) of SHR, with a decrease in lecithin retinol acyltransferase (LRAT), which regulates retinoid metabolism and photoreceptor health. In human RPE cells (ARPE-19), TGF-ß administration suppressed mRNA and protein levels of LRAT; and vactosertib, a selective inhibitor of TGF-ß receptor kinase type 1, reversed the effect of TGF-ß. These findings suggest that hypertension-induced retinal neurodegeneration involves inflammation, apoptosis, necroptosis, and disrupted retinoid metabolism, providing potential therapeutic targets for hypertensive retinopathy.

Supplementation with Lactobacillus helveticus NS8 alleviated behavioral, neural, endocrine, and microbiota abnormalities in an endogenous rat model of depression.

Introduction: Major depressive disorder is a condition involving microbiota-gut-brain axis dysfunction. Increasing research aims to improve depression through gut microbiota regulation, including interventions such as probiotics, prebiotics, and fecal microbiota transplants. However, most research focuses on exogenous depression induced by chronic stress or drugs, with less attention given to endogenous depression. Additionally, research on gut mycobiota in depression is significantly less than that on gut bacteria. Methods: In the present study, Wistar-Kyoto rats were used as an endogenous depression and treatment-resistant depression model, while Wistar rats served as controls. Differences between the two rat strains in behavior, gut bacteria, gut mycobiota, nervous system, endocrine system, immune system, and gut barrier were evaluated. Additionally, the effects of Lactobacillus helveticus NS8 supplementation were investigated. Results: Wistar-Kyoto rats demonstrated increased depressive-like behaviors in the forced swimming test, reduced sucrose preference in the sucrose preference test, and decreased locomotor activity in the open field test. They also exhibited abnormal gut bacteria and mycobiota, characterized by higher bacterial α-diversity but lower fungal α-diversity, along with increased butyrate, L-tyrosine, and L-phenylalanine biosynthesis from bacteria. Furthermore, these rats showed dysfunction in the microbiota-gut-brain axis, evidenced by a hypo-serotonergic system, hyper-noradrenergic system, defective hypothalamic-pituitary-adrenal axis, compromised gut barrier integrity, heightened serum inflammation, and diminished gut immunity. A 1-month L. helveticus NS8 intervention increased the fecal abundance of L. helveticus; reduced the abundance of Bilophila and Debaryomycetaceae; decreased immobility time but increased climbing time in the forced swimming test; reduced hippocampal corticotropin-releasing hormone levels; decreased hypothalamic norepinephrine levels; increased hippocampal glucocorticoid receptor, brain-derived neurotrophic factor dopamine, and 5-hydroxyindoleacetic acid content; and improved the gut microbiota, serotonergic, and noradrenergic system. Conclusion: The depressive phenotype of Wistar-Kyoto rats is not only attributed to their genetic context but also closely related to their gut microbiota. Abnormal gut microbiota and a dysfunctional microbiota-gut-brain axis play important roles in endogenous depression, just as they do in exogenous depression. Supplementing with probiotics such as L. helveticus NS8 is likely a promising approach to improve endogenous depression and treatment-resistant depression.

Renal denervation improves mitochondrial oxidative stress and cardiac hypertrophy through inactivating SP1/BACH1-PACS2 signaling.

BACKGROUND: Renal denervation (RDN) has been proved to relieve cardiac hypertrophy; however, its detailed mechanisms remain obscure. This study investigated the detailed protective mechanisms of RDN against cardiac hypertrophy during hypertensive heart failure (HF). METHODS: Male 5-month-old spontaneously hypertension (SHR) rats were used in a HF rat model, and male Wistar-Kyoto (WKY) rats of the same age were used as the baseline control. Myocardial hypertrophy and fibrosis were evaluated by hematoxylin-eosin (HE) staining and Masson staining. The expression of target molecule was analyzed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), Western blot, immunohistochemical and immunofluorescence, respectively. Cardiomyocyte hypertrophy was induced by norepinephrine (NE) in H9c2 cells in vitro and evaluated by brain natriuretic peptide (BNP), atrial natriuretic peptide (ANP), ß-myosin heavy chain (ß-MHC), and α-myosin heavy chain (α-MHC) levels. Oxidative stress was determined by malondialdehyde (MDA) level, superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) enzyme activities. Mitochondrial function was measured by mitochondrial membrane potential, adenosine triphosphate (ATP) production, mitochondrial DNA (mtDNA) number, and mitochondrial complex I-IV activities. Molecular mechanism was assessed by dual luciferase reporter and chromatin immunoprecipitation (ChIP) assays. RESULTS: RDN decreased sympathetic nerve activity, attenuated myocardial hypertrophy and fibrosis, and improved cardiac function in the rat model of HF. In addition, RDN ameliorated mitochondrial oxidative stress in myocardial tissues as evidenced by reducing MDA and mitochondrial reactive oxygen species (ROS) levels, and enhancing SOD and GSH-Px activities. Moreover, phosphofurin acid cluster sorting protein 2 (PACS-2) and broad-complex, tramtrak and bric à brac (BTB) domain and cap'n'collar (CNC) homolog 1 (BACH1) were down-regulated by RDN. In NE-stimulated H9c2 cells, PACS-2 and BACH1 levels were markedly elevated, and knockdown of them could suppress NE-induced oxidative stress, cardiomyocyte hypertrophy, fibrosis, as well as mitochondrial dysfunction. Transforming growth factor beta1(TGFß1)/SMADs signaling pathway was inactivated by RDN in the HF rats, which sequentially inhibited specificity protein 1 (SP1)-mediated transcription of PACS2 and BACH1. CONCLUSION: Collectively, these data demonstrated that RDN improved cardiac hypertrophy and sympathetic nerve activity of HF rats via repressing BACH1 and PACS-2-mediated mitochondrial oxidative stress by inactivating TGF-ß1/SMADs/SP1 pathway, which shed lights on the cardioprotective mechanism of RDN in HF.

[Effects of acupuncture on endothelial active factors and related autonomic neurotransmitters in spontaneous hypertension rats].

OBJECTIVE: To observe the effects of acupuncture at "antihypertensive acupoint prescription" on endothelial active factors and related autonomic neurotransmitters in spontaneous hypertension rats, and explore the vascular regulation and central regulation mechanisms of acupuncture for anti-hypertension. METHODS: Thirty SPF grade male spontaneous hypertension rats were randomly divided into a model group (15 rats) and an acupuncture group (15 rats). Besides, 15 Wistar Kyoto rats were collected as a blank control group (normal group). In the acupuncture group, acupuncture was delivered at the "antihypertensive acupoint prescription" (bilateral "Renying" [ST 9], "Quchi" [LI 11], "Zusanli" [ST 36], "Taichong" [LR 3] and "Neiguan" [PC 6]), with needles retained for 30 min, once daily. The duration of intervention was 28 days. Every week, using the the irritation scale, the sign of sympathetic irritation was evaluated dynamically. The arterial blood pressure of the rats tail was determined, using non-invasive blood pressure measurement system. ELISA was adopted to detect the levels of calcitonin gene-related peptide (CGRP), nitric oxide (NO), endothelin-1 (ET-1), neuropeptide Y (NPY) in the serum. DAB chromogenic in situ hybridization (CISH) was provided to detect the mRNA expression of endothelial nitric oxide synthase (eNOS) in the internal carotid artery and the arcuate nucleus (ARC), and that of CGRP in the paraventricular nucleus posterior (PVP) and the ventrolateral medulla (VLM). Liquid chromatography-mass spectrometry (LC-MS) was used to detect the levels of epinephrine (E) and norepinephrine (NE) in the paraventricular nucleus anterior (PVA). RESULTS: Compared with the normal group, the irritation scores, systolic blood pressure and diastolic blood pressure were increased at each time point in the model group (P<0.05). When compared with the model group, the irritation scores after the intervention for 3 and 4 weeks, and systolic and diastolic blood pressure after intervention for 2, 3 and 4 weeks were reduced in the acupuncture group (P<0.05). In comparison with the normal group, the serum CGRP and NO levels of the rats were decreased (P<0.05), and the serum ET-1 and NPY levels, as well as E and EN levels in PVA were increased (P<0.05) in the model group. The levels of serum CGRP and NO were elevated (P<0.05), and the serum ET-1 and NPY levels, as well as E and EN levels of PVA were reduced (P<0.05) in the acupuncture group when compared with those of the model group. In the model group, the media of internal carotid artery exhibited thickening and remodeling, while the neuron volume in ARC was small. In the acupuncture group, every layer of internal carotid artery was acceptably arranged, and the parvicellular neuron of ARC was moderate in volume. For the in situ hybridization of eNOS mRNA for the rats of each group, the smooth muscle cells were predominantly expressed in each layer of the internal carotid artery, whereas the expression of parvicellular neurons was dominated in ARC. In the model group, the large and small neurosecretory cells were distributed sparsely in the nerves of PVP; in the acupuncture group, the cells of these two species were distributed regularly; and there were few species of glial cell in the VLM of either the model group or the acupuncture group. In each group, for the in situ hybridization of CGRP mRNA, the small neurosecretory cells were expressed predominately in the PVP, while, the expression of glial cell nuclei and the cell cytoplasm was dominated in the VLM. Compared with the normal group, the mRNA expression of eNOS in the internal carotid artery and ARC and that of CGRP mRNA in the PVP and VLM was decreased in the model group (P<0.05). In the acupuncture group, when compared with the model group, the mRNA expression of eNOS in the internal carotid artery and ARC and that of CGRP in the PVP and VLM was increased in the acupuncture group (P<0.05). CONCLUSION: Acupuncture at "antihypertensive acupoint prescription" can upregulate the level of vascular relaxing factors, downregulate the level of contracting factors, enhance the response of relaxing factors in targeting blood vessels and regulating the center. The mechanism may be related to the modulation of the sympathetic-adrenergic autonomic neurotransmitters in the paraventricular nucleus in spontaneous hypertension rats.

Hypotensive and Cardioprotective Potential of Yellow Bedstraw Extract-Based Oral Liquid in Spontaneously Hypertensive Rats.

This study aimed to prepare, characterize and assess the antioxidant activity of yellow bedstraw extracts (YBEs), focusing on identifying extracts with high antioxidant capacity. The selected extract was loaded into an oral liquid formulation and further investigated for its therapeutic potential in reducing blood pressure and associated complications in spontaneously hypertensive Wistar kyoto rats (SHR). Rats were divided into untreated SHR and SHR treated with a YBE-based oral formulation over four weeks. After treatment, blood pressure was measured, and cardiac function was assessed using the Langendorff technique to simulate ex vivo ischemic conditions. Prooxidant levels were assessed in plasma while antioxidant activity was evaluated in red blood cells. Histological analyses of heart, kidney, and liver samples were conducted to assess pathological changes induced by hypertension. Our results showed that the oral formulation loaded with ethanol YBE effectively reduced blood pressure, preserved myocardial function under ischemic stress, and decreased oxidative stress markers in blood. Importantly, our formulation with YBE demonstrated potential in attenuating structural kidney damage associated with hypertension. Overall, these findings suggest a cardioprotective effect of orally administered YBE formulation, highlighting its potential as an herbal supplement. However, clinical studies are warranted to validate these findings and explore the extract's suitability for clinical use.

Differential gene expression in the kidneys of SHR and WKY rats after intravenous administration of Akkermansia muciniphila-derived extracellular vesicles.

Although Akkermansia muciniphila (Am) plays a beneficial role as a probiotic in the treatment of metabolic syndrome, the mechanisms remain elusive. We tested the hypothesis that Am extracellular vesicles (AmEVs) protect against hypertension through modulation of gene expression in the kidneys of spontaneously hypertensive rats (SHRs). Extracellular vesicles purified from anaerobically cultured Am (1.0 × 108 or 1.0 × 109 particles/kg) or vehicles were injected into the tail veins of Wistar-Kyoto rats (WKYs) and SHRs weekly for 4 weeks. Renal cortical tissues isolated from both rat strains were analyzed by trichrome stain and RT-qPCR. AmEVs protect against the development of hypertension in SHRs without a serious adverse reaction. AmEVs increased the expression of vasocontracting Agt and At1ar as well as vasodilating At2r, Mas1 and Nos2 in the kidneys of both strains. These results indicate that AmEVs have a protective effect against hypertension without a serious adverse reaction. Therefore, it is foreseen that AmEVs may be utilized as a novel therapeutic for the treatment of hypertension.

MiR-196a-5p hinders vascular smooth muscle cell proliferation and vascular remodeling via repressing BACH1 expression.

Hyperproliferation of vascular smooth muscle cells (VSMCs) is a driver of hypertensive vascular remodeling. This study aimed to uncover the mechanism of BTB and CNC homology 1 (BACH1) and microRNAs (miRNAs) in VSMC growth and hypertensive vascular remodeling. With the help of TargetScan, miRWalk, miRDB, and miRTarBase online database, we identified that BACH1 might be targeted by miR-196a-5p, and overexpressed in VSMCs and aortic tissues from spontaneously hypertensive rats (SHRs). Gain- and loss-of-function experiments demonstrated that miR-196a-5p suppressed VSMC proliferation, oxidative stress and hypertensive vascular remodeling. Double luciferase reporter gene assay and functional verification showed that miR-196a-5p cracked down the transcription and translation of BACH1 in both Wistar Kyoto rats (WKYs) and SHRs. Silencing BACH1 mimicked the actions of miR-196a-5p overexpression on attenuating the proliferation and oxidative damage of VSMCs derived from SHRs. Importantly, miR-196a-5p overexpression and BACH1 knockdown cooperatively inhibited VSMC proliferation and oxidative stress in SHRs. Furthermore, miR-196a-5p, if knocked down in SHRs, aggravated hypertension, upregulated BACH1 and promoted VSMC proliferation, all contributing to vascular remodeling. Taken together, targeting miR-196a-5p to downregulate BACH1 may be a promising strategy for retarding VSMC proliferation and hypertensive vascular remodeling.

More efficient adaptation of cardiovascular response to repeated restraint in spontaneously hypertensive rats: the role of autonomic nervous system.

We hypothesized that sympathetic hyperactivity and parasympathetic insuficiency in spontaneously hypertensive rats (SHR) underlie their exaggerated cardiovascular response to acute stress and impaired adaptation to repeated restraint stress exposure compared to Wistar-Kyoto rats (WKY). Cardiovascular responses to single (120 min) or repeated (daily 120 min for 1 week) restraint were measured by radiotelemetry and autonomic balance was evaluated by power spectral analysis of systolic blood pressure variability (SBPV) and heart rate variability (HRV). Baroreflex sensitivity (BRS) was measured by the pharmacological Oxford technique. Stress-induced pressor response and vascular sympathetic activity (low-frequency component of SBPV) were enhanced in SHR subjected to single restraint compared to WKY, whereas stress-induced tachycardia was similar in both strains. SHR exhibited attenuated cardiac parasympathetic activity (high-frequency component of HRV) and blunted BRS compared to WKY. Repeated restraint did not affect the stress-induced increase in blood pressure. However, cardiovascular response during the post-stress recovery period of the 7th restraint was reduced in both strains. The repeatedly restrained SHR showed lower basal heart rate during the dark (active) phase and slightly decreased basal blood pressure during the light phase compared to stress-naive SHR. SHR subjected to repeated restraint also exhibited attenuated stress-induced tachycardia, augmented cardiac parasympathetic activity, attenuated vascular sympathetic activity and improved BRS during the last seventh restraint compared to single-stressed SHR. Thus, SHR exhibited enhanced cardiovascular and sympathetic responsiveness to novel stressor exposure (single restraint) compared to WKY. Unexpectedly, the adaptation of cardiovascular and autonomic responses to repeated restraint was more effective in SHR.

Vasohibin inhibition improves myocardial relaxation in a rat model of heart failure with preserved ejection fraction.

Heart failure with preserved ejection fraction (HFpEF) is a complex syndrome associated with increased myocardial stiffness and cardiac filling abnormalities. Prior studies implicated increased α-tubulin detyrosination, which is catalyzed by the vasohibin enzymes, as a contributor to increased stabilization of the cardiomyocyte microtubule network (MTN) and stiffness in failing human hearts. We explored whether increased MTN detyrosination contributed to impaired diastolic function in the ZSF1 obese rat model of HFpEF and designed a small-molecule vasohibin inhibitor to ablate MTN detyrosination in vivo. Compared with ZSF1 lean and Wistar Kyoto rats, obese rats exhibited increased tubulin detyrosination concomitant with diastolic dysfunction, left atrial enlargement, and cardiac hypertrophy with a preserved left ventricle ejection fraction, consistent with an HFpEF phenotype. Ex vivo myocardial phenotyping assessed cardiomyocyte mechanics and contractility. Vasohibin inhibitor treatment of isolated cardiomyocytes from obese rats resulted in reduced stiffness and faster relaxation. Acute in vivo treatment with vasohibin inhibitor improved diastolic relaxation in ZSF1 obese rats compared with ZSF1 lean and Wistar Kyoto rats. Vasohibin inhibition also improved relaxation in isolated human cardiomyocytes from both failing and nonfailing hearts. Our data suggest the therapeutic potential for vasohibin inhibition to reduce myocardial stiffness and improve relaxation in HFpEF.

Gstp1 negatively regulates blood pressure in hypertensive rat via promoting APLNR ubiquitination degradation mediated by Nedd4.

Hypertension is a leading risk factor for disease burden worldwide. Vascular contraction and remodeling contribute to the development of hypertension. Glutathione S-transferase P1 (Gstp1) plays several critical roles in both normal and neoplastic cells. In this study, we investigated the effect of Gstp1 on hypertension as well as on vascular smooth muscle cell (VSMC) contraction and phenotypic switching. We identified the higher level of Gstp1 in arteries and VSMCs from hypertensive rats compared with normotensive rats for the first time. We then developed Adeno-associated virus 9 (AAV9) mediated Gstp1 down-regulation and overexpression in rats and measured rat blood pressure by using the tail-cuff and the carotid catheter method. We found that the blood pressure of spontaneously hypertensive rats (SHR) rose significantly with Gstp1 down-regulation and reduced apparently after Gstp1 overexpression. Similar results were obtained from the observations of 2-kidney-1-clip renovascular (2K1C) hypertensive rats. Gstp1 did not influence blood pressure of normotensive Wistar-Kyoto (WKY) rats and Sprague-Dawley (SD) rats. Further in vitro study indicated that Gstp1 knockdown in SHR-VSMCs promoted cell proliferation, migration, dedifferentiation and contraction, while Gstp1 overexpression showed opposite effects. Results from bioinformatic analysis showed that the Apelin/APLNR system was involved in the effect of Gstp1 on SHR-VSMCs. The rise in blood pressure of SHR induced by Gstp1 knockdown could be reversed by APLNR antagonist F13A. We further found that Gstp1 enhanced the association between APLNR and Nedd4 E3 ubiquitin ligases to induce APLNR ubiquitination degradation. Thus, in the present study, we discovered a novel anti-hypertensive role of Gstp1 in hypertensive rats and provided the experimental basis for designing an effective anti-hypertensive therapeutic strategy.

Protective effects of the R-(+)-thioctic acid treatment: possible anti-inflammatory activity on heart of hypertensive rats.

BACKGROUND: In cardiovascular disease, high blood pressure is associated with oxidative stress, promoting endothelial dysfunction, vascular remodeling, and inflammation. Clinical trials are discordant that the most effective treatment in the management of hypertension seems to be the administration of anti-hypertensive drugs with antioxidant properties. The study aims to evaluate the effects of the eutomer of thioctic acid on oxidative stress and inflammation in the heart of spontaneously hypertensive rats compared to normotensive Wistar Kyoto rats. METHODS: To study the oxidative status, the malondialdehyde and 4-hydroxynonenal concentration, protein oxidation were measured in the heart. Morphological analysis were performed. Immunohistochemistry and Western blot were done for alpha-smooth muscle actin and transforming growth factor beta to assess fibrosis; cytokines and nuclear factor kappaB to assess inflammatory processes. RESULTS: Spontaneously hypertensive rats were characterized by hypertension with increased malondialdehyde levels in the heart. OxyBlot in the heart of spontaneously hypertensive rats showed an increase in proteins' oxidative status. Cardiomyocyte hypertrophy and fibrosis in the ventricles were associated with an increased expression of alpha-smooth muscle actin and pro-inflammatory cytokines, reduced by the eutomer of thioctic acid supplementation. CONCLUSIONS: Based on this evidence, eutomer of thioctic acid could represent an appropriate antioxidant molecule to reduce oxidative stress and prevent inflammatory processes on the cardiomyocytes and cardiac vascular endothelium.

Acupuncture modulation of the ACE/Ang II/AT1R and ACE2/Ang(1-7)/MasR pathways in the rostral ventrolateral medulla reduces sympathetic output and prevents cardiac injury caused by SHR hypertension.

Acupuncture can reduce blood pressure, heart rate (HR), and ameliorate cardiac damage by modulating the excitability of the sympathetic nervous system, but the exact mechanism of this effect remains unclear. This study investigated the potential mechanisms of acupuncture in the treatment of cardiac damage in hypertension. Spontaneously hypertensive rats (SHR) were used as the hypertension model with Wistar-Kyoto rats as the control. Manual acupuncture, electroacupuncture, and metoprolol were used as interventions. Systolic and diastolic blood pressure (SBP, DBP) plus HR were monitored with cardiac structure determined using Masson staining. Angiotensin II (Ang II) and norepinephrine in myocardium were detected with ELISA as was Ang(1-7) and gamma aminobutyric acid (GABA) in the rostral ventrolateral medulla (RVLM). Expression of mRNA for collagen type I (Col-I), Col-III, actin α1 (ACTA1), and thrombospondin 4 (THBS4) in myocardium was detected using real-time PCR. Expression of angiotensin converting enzyme (ACE), Ang II, angiotensin II type 1 receptor (AT1R), ACE2, and Mas receptor (MasR) proteins in RVLM was monitored using western blot. After manual acupuncture and electroacupuncture treatment, SHRs showed decreased SBP, DBP and HR, reduced myocardial damage. There was decreased expression of the ACE/Ang II/AT1R axis, and increased expression of the ACE2/Ang(1-7)/MasR axis within the RVLM. GABA levels were increased within the RVLM and norepinephrine levels were decreased in myocardial tissue. Metoprolol was more effective than either manual acupuncture or electroacupuncture. Acupuncture directed against hypertensive cardiac damage may be associated with regulation of ACE/Ang II/AT1R and the ACE2/Ang(1-7)/MasR pathway within the RLVM to reduce cardiac sympathetic excitability.

BK Channels in Tail Artery Vascular Smooth Muscle Cells of Normotensive (WKY) and Hypertensive (SHR) Rats Possess Similar Calcium Sensitivity But Different Responses to the Vasodilator Iloprost.

It has been reported that, in the spontaneously hypertensive rat (SHR) model of hypertension, different components of the G-protein/adenylate cyclase (AC)/Calcium-activated potassium channel of high conductance (BK) channel signaling pathway are altered differently. In the upstream part of the pathway (G-protein/AC), a comparatively low efficacy has been established, whereas downstream BK currents seem to be increased. Thus, the overall performance of this signaling pathway in SHR is elusive. For a better understanding, we focused on one aspect, the direct targeting of the BK channel by the G-protein/AC pathway and tested the hypothesis that the comparatively low AC pathway efficacy in SHR results in a reduced agonist-induced stimulation of BK currents. This hypothesis was investigated using freshly isolated smooth muscle cells from WKY and SHR rat tail artery and the patch-clamp technique. It was observed that: (1) single BK channels have similar current-voltage relationships, voltage-dependence and calcium sensitivity; (2) BK currents in cells with a strong buffering of the BK channel activator calcium have similar current-voltage relationships; (3) the iloprost-induced concentration-dependent increase of the BK current is larger in WKY compared to SHR; (4) the effects of activators of the PKA pathway, the catalytic subunit of PKA and the potent and selective cAMP-analogue Sp-5,6-DCl-cBIMPS on BK currents are similar. Thus, our data suggest that the lower iloprost-induced stimulation of the BK current in freshly isolated rat tail artery smooth muscle cells from SHR compared with WKY is due to the lower efficacy of upstream elements of the G-Protein/AC/BK channel pathway.

Levilactobacillus brevis SG031 modulates mood-related behaviors and attenuates stress-related sleep disturbance and autonomic dysfunction via gut microbiota modulation in Wistar-Kyoto rats.

AIMS: The probiotic bacterium Levilactobacillus brevis (L. brevis) has been proposed as a potential solution to manage mood disorders and alleviate stress-related sleep disturbances. However, the underlying mechanisms of its effects have not been fully elucidated. The aim of this study was to explore the impact and potential mechanisms of L. brevis SG031 supplementation on anxiety/depression-like behaviors and stress-induced changes in sleep patterns and sleep-related autonomic function. MAIN METHODS: Male Wistar-Kyoto rats were administered low, medium, or high doses of L. brevis SG031 or a vehicle for 4 weeks, followed by behavioral tests to evaluate anxiety and depression. After an additional 2 weeks of SG031 or vehicle administration, a cage-exchange paradigm was performed with 24-hour physiological signal measurements under different stress conditions. Fecal samples were collected to construct a 16S rRNA library and assess fecal short-chain fatty acids (SCFAs). KEY FINDINGS: High-dose SG031 administration yielded reduced depression-like responses and enhanced social interaction in behavioral tests. It also exhibited a protective effect against stress-induced sleep disturbance characterized by decreased sleep time, increased awake time, and autonomic dysfunction during sleep. Fecal examination indicated that high-dose SG031 administration exerted beneficial effects on gut health by maintaining the gut microbial abundance, preserving stability of the microbial composition, and enriching the gut with SCFAs, which were associated with improvements in sleep and autonomic function. SIGNIFICANCE: These findings collectively underscore the multifaceted potential of SG031 in addressing mental health and stress-related sleep challenges through the modulation of the gut microbiota.

miR-193b-3p and miR-346 Exert Antihypertensive Effects in the Rostral Ventrolateral Medulla.

BACKGROUND: Rostral ventrolateral medulla (RVLM) neuron hyperactivity raises sympathetic outflow, causing hypertension. MicroRNAs (miRNAs) contribute to diverse biological processes, but their influence on RVLM neuronal excitability and blood pressure (BP) remains widely unexplored. METHODS AND RESULTS: The RVLM miRNA profiles in spontaneously hypertensive rats were unveiled using RNA sequencing. Potential effects of these miRNAs in reducing neuronal excitability and BP and underlying mechanisms were investigated through various experiments. Six hundred thirty-seven miRNAs were identified, and reduced levels of miR-193b-3p and miR-346 were observed in the RVLM of spontaneously hypertensive rats. Increased miR-193b-3p and miR-346 expression in RVLM lowered neuronal excitability, sympathetic outflow, and BP in spontaneously hypertensive rats. In contrast, suppressing miR-193b-3p and miR-346 expression in RVLM increased neuronal excitability, sympathetic outflow, and BP in Wistar Kyoto and Sprague-Dawley rats. Cdc42 guanine nucleotide exchange factor (Arhgef9) was recognized as a target of miR-193b-3p. Overexpressing miR-193b-3p caused an evident decrease in Arhgef9 expression, resulting in the inhibition of neuronal apoptosis. By contrast, its downregulation produced the opposite effects. Importantly, the decrease in neuronal excitability, sympathetic outflow, and BP observed in spontaneously hypertensive rats due to miR-193b-3p overexpression was greatly counteracted by Arhgef9 upregulation. CONCLUSIONS: miR-193b-3p and miR-346 are newly identified factors in RVLM that hinder hypertension progression, and the miR-193b-3p/Arhgef9/apoptosis pathway presents a potential mechanism, highlighting the potential of targeting miRNAs for hypertension prevention.

Acid-sensing ion channel 1 in nucleus tractus solitarii neurons contributes to the enhanced CO2-stimulated cardiorespiratory effect in spontaneously hypertensive rats.

AIMS: Activation of central respiratory chemoreceptors provides excitatory drive to both respiratory and sympathetic outputs. The enhanced respiratory-sympathetic coupling contributes to the onset and development of hypertension. However, the specific central targets and molecular mechanisms involved in this process remain elusive. This study aimed to investigate the role of acid-sensing ion channel 1 (ASIC1) in nucleus tractus solitarii (NTS) neurons in CO2-stimulated cardiorespiratory effects in spontaneously hypertensive rats (SHRs). MAIN METHODS: Respiration and blood pressure of conscious rats were recorded by whole-body plethysmography and telemetry, respectively. Western blot was used to detect the expression difference of ASIC1 protein in NTS region between Wistar-Kyoto (WKY) rats and SHRs. Excitability of NTS neurons were assessed by extracellular recordings. KEY FINDINGS: Compared to WKY rats, the enhanced CO2-stimulated cardiopulmonary effect and up-regulation of ASIC1 in the NTS were already observed in 4-week-old prehypertensive SHRs. Furthermore, specific blockade of ASIC1 effectively attenuated the CO2-stimulated increase in firing rate of NTS neurons in anesthetized adult SHRs. Intracerebroventricular injections of the ASIC1a blocker PcTx1 or knockdown Asic1 in NTS neurons significantly reduced the heightened CO2-stimulated ventilatory response, and diminished the CO2-stimulated increase in arterial pressure and heart rate in adult SHRs. SIGNIFICANCE: These findings showed that dysregulated ASIC1 signaling in the NTS contribute to the exaggerated CO2-stimulated cardiorespiratory effects observed in SHRs.

The antihypertensive effect of Alizarin is achieved by activating VEGFR2/eNOS pathway, attenuating oxidative stress-induced mitochondrial damage and premature senescence.

The primary and initial manifestations of hypertension encompass arterial hypoelasticity and histiocyte senescence. Oxidative stress plays a pivotal role in the progression of senescence. Elevated intracellular oxidative stress levels will directly induce cell damage, disrupt normal physiological signal transduction, which can cause mitochondrial dysfunction to accelerate the process of senescence. Alizarin, an anthraquinone active ingredient isolated from Rubia cordifolia L., has a variety of pharmacological effects, including antioxidant, anti-inflammatory and anti-platelet. Nevertheless, its potential in lowering blood pressure (BP) and mitigating hypertension-induced vascular senescence remains uncertain. In this study, we used spontaneously hypertensive rats (SHR) and human umbilical vein endothelial cells (HUVECs) to establish a model of vascular senescence in hypertension. Our aim was to elucidate the mechanisms underpinning the vascular protective effects of Alizarin. By assessing systolic blood pressure (SBP) and diastolic blood pressure (DBP), H&E staining, SA-ß-Gal staining, vascular function, oxidative stress levels, calcium ion concentration and mitochondrial membrane potential, we found that Alizarin not only restored SBP and increased endothelium-dependent relaxation (EDR) in SHR, but also inhibited oxidative stress-induced mitochondrial damage and significantly delayed the vascular senescence effect in hypertension, and the mechanism may be related to the activation of VEGFR2/eNOS signaling pathway.

[Effects of acupuncture on fecal short-chain fatty acids and TLR4/MyD88/NF-κB signaling pathway in spontaneously hypertensive rats].

OBJECTIVE: To observe the effects of acupuncture on blood pressure, fecal short-chain fatty acids (SCFAs) and toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88)/nuclear factor-κB (NF-κB) signaling pathway in spontaneously hypertensive rats (SHR), and to explore the mechanism of acupuncture for anti-hypertension. METHODS: Twenty-four male SHR of SPF grade were randomly divided into a model group, a western medication group, an acupuncture group and a sham acupuncture group, with 6 rats in each group, and 6 male Wistar-Kyoto rats were selected as the blank group additionally. Hydrochlorothiazide solution was given by gavage in the western medication group; acupuncture was applied at bilateral "Renying" (ST 9) and "Zusanli" (ST 36) in the acupuncture group, 20 min a time; acupuncture was applied at the non-meridian and non-acupoint points close to bilateral "Renying" (ST 9) and "Zusanli" (ST 36) in the sham acupuncture group, 20 min a time. The intervention was adopted once a day for 4 weeks continuously in each group. The systolic blood pressure (SBP) of the caudal artery was measured before intervention and after 1, 2, 3 and 4 weeks of intervention. After intervention, the morphology of colonic tissue was observed by HE staining; the fecal level of SCFAs was detected by gas chromatography; the serum levels of interleukin (IL)-6, IL-1ßand tumor necrosis factor-α (TNF-α) were detected by ELISA; the protein expression of TLR4, MyD88 and NF-κB p65 in the mesenteric artery was detected by Western blot. RESULTS: Compared with the blank group, in the model group, the SBP was increased (P<0.05), significant pathological changes could be found in the colonic tissue, the fecal SCFAs level was decreased (P<0.05), the serum levels of IL-6, IL-1ß and TNF-α were increased (P<0.05), the protein expression of TLR4, MyD88 and NF-κB p65 in the mesenteric artery was increased (P<0.05). Compared with the model group, the SBP after 2, 3 and 4 weeks of intervention was decreased (P<0.05), the serum levels of IL-6, IL-1ß and TNF-α were decreased (P<0.05) in the acupuncture group and the western medication group; the mucosal epithelium of colonic tissue was intact, the number of intestinal glands was abundant, the fecal SCFAs level was increased (P<0.05), and the protein expression of TLR4, MyD88 and NF-κB p65 in the mesenteric artery was decreased (P<0.05) in the acupuncture group. Compared with the sham acupuncture group, the SBP after 2, 3 and 4 weeks of intervention was decreased (P<0.05), the fecal SCFAs level was increased (P<0.05), the serum levels of IL-6, IL-1ß and TNF-α were decreased (P<0.05), the protein expression of TLR4, MyD88 and NF-κB p65 in the mesenteric artery was decreased (P<0.05) in the acupuncture group. CONCLUSION: Acupuncture at bilateral "Renying" (ST 9) and "Zusanli" (ST 36) can effectively play an anti-hypertensive role in SHR. Its mechanism may be related to regulating fecal SCFAs level and inhibiting the TLR4/MyD88/NF-κB signaling pathway.

Alcohol withdrawal and amphetamine co-use in an animal model for attention deficit hyperactivity disorder.

Background: Non-medical use of amphetamine and other stimulants prescribed for treatment of attention deficit/hyperactivity disorder (ADHD) is of special concern when combined with alcohol consumption. In a previous study, we modeled chronic ethanol-amphetamine co-use in adolescent Long-Evans (LE) rats and provided evidence that amphetamine attenuates alcohol withdrawal symptoms.Objectives: This project modeled co-use of amphetamine with alcohol in adolescents with ADHD-like symptoms by examining ethanol-amphetamine administration in adolescent Spontaneously Hypertensive Rats (SHR), an experimental model for the study of ADHD. Withdrawal symptoms were compared among SHR and two control rat strains, LE and Wistar Kyoto (WKY).Methods: At postnatal day 32, parallel groups of 12-24 male SHR, WKY and LE rats were administered a liquid diet containing ethanol (3.6%) and/or amphetamine (20 mg/L). Following administration periods up to 26 days, rats were withdrawn from their treatment and tested for overall severity of alcohol withdrawal symptoms, general locomotor activity, and anxiety-like behavior.Results: Overall withdrawal severity was lower for SHR than for LE (p < .001) or WKY (p = .027). Co-consumption of amphetamine decreased withdrawal severity for LE (p = .033) and WKY (p = .011) but not SHR (p = .600). Only WKY showed increased anxiety-like behavior during withdrawal (p = .031), but not after amphetamine co-administration (p = .832).Conclusion: Alcohol withdrawal severity may be attenuated when co-used with amphetamine. However, as a model for ADHD, SHR adolescents appeared resistant to developing significant signs of alcohol withdrawal following alcohol consumption. Whether alcohol withdrawal symptoms are attenuated or absent, potential consequences could include a decreased awareness of an emerging problem with alcohol use.