Antioxidant Efficacy of Hwangryunhaedok-tang through Nrf2 and AMPK Signaling Pathway against Neurological Disorders In Vivo and In Vitro.

Alzheimer's disease (AD) is a representative cause of dementia and is caused by neuronal loss, leading to the accumulation of aberrant neuritic plaques and the formation of neurofibrillary tangles. Oxidative stress is involved in the impaired clearance of amyloid beta (Aß), and Aß-induced oxidative stress causes AD by inducing the formation of neurofibrillary tangles. Hwangryunhaedok-tang (HHT, Kracie K-09®), a traditional herbal medicine prescription, has shown therapeutic effects on various diseases. However, the studies of HHT as a potential treatment for AD are insufficient. Therefore, our study identified the neurological effects and mechanisms of HHT and its key bioactive compounds against Alzheimer's disease in vivo and in vitro. In a 5xFAD mouse model, our study confirmed that HHT attenuated cognitive impairments in the Morris water maze (MWM) test and passive avoidance (PA) test. In addition, the prevention of neuron impairment, reduction in the protein levels of Aß, and inhibition of cell apoptosis were confirmed with brain tissue staining. In HT-22 cells, HHT attenuates tBHP-induced cytotoxicity, ROS generation, and mitochondrial dysfunction. It was verified that HHT exerts a neuroprotective effect by activating signaling pathways interacting with Nrf2, such as MAPK/ERK, PI3K/Akt, and LKB1/AMPK. Among the components, baicalein, a bioavailable compound of HHT, exhibited neuroprotective properties and activated the Akt, AMPK, and Nrf2/HO-1 pathways. Our findings indicate a mechanism for HHT and its major bioavailable compounds to treat and prevent AD and suggest its potential.

Foreign Body Granuloma Following Hwangryunhaedok-tang Pharmacopuncture for Postherpetic Neuralgia.

Foreign body granuloma can be caused by endogenous compounds as well as various injectable materials. In oriental medicine, pharmacopuncture combining herbal medicine administration and injection is one of the commonly used procedures. Hwangryunhaedok-tang (HHT, a.k.a., Huang-Lian-Jie-Du-Tang in China), an oriental medicinal herb known to produce anti-inflammatory effects, has been recently made in pharmacopuncture products and commonly used for various disorders. An 88-year-old female presented with multiple tender nodules on the left parietal scalp and forehead. The diagnosis of foreign body granuloma caused by HHT pharmacopuncture was revealed by more detailed previous treatment history of postherpetic neuralgia and histopathologic examination. Herein, we report a foreign body granuloma as a delayed adverse reaction caused by non-standard administration of herbal extracts, considered biologically inert.

Two case reports of tapering sedative-hypnotic drugs through classical conditioning using herbal medicine (CARE-compliant).

RATIONALE: Sedative-hypnotic drugs (SHDs) used for insomnia are recommended for short-term use owing to concerns regarding abuse and dependence. Nevertheless, drug discontinuation is challenging owing to rebound insomnia that occurs when the SHD is ceased. Therefore, a strategy is required to reduce or discontinue SHDs, while minimizing rebound insomnia. PATIENT CONCERNS: The present report discusses the cases of a 72-year-old man and 27-year-old man with insomnia. Both patients had been taking SHDs for more than 2 months. DIAGNOSES: Both patients were diagnosed with primary insomnia. INTERVENTIONS AND OUTCOMES: After 2 weeks of classical conditioning (CC) using Hwangryunhaedok-tang (HHT), both patients succeeded in tapering SHD; the Korean version of the insomnia severity index of patient 1 dropped from 14 to 5, and that of patient 2 dropped from 28 to 11. In the final follow-up, the index dropped to 2 and 3, respectively. LESSONS: The CC, a concept emerging in psychology, could be a new strategy for tapering drugs. In this case, SHDs, which had been taken for a long time, were successfully reduced or discontinued through CC using HHT. Herbal medicine such as HHT is selected as a neutral stimulus for CC because the risk of rebound phenomenon is low as the compounds of herbal medicine have high structural similarities with human metabolites. Both patients in this study reported no particular challenges when HHT was discontinued. However, further studies are needed that will assist in tapering anticonvulsants or steroids as well as SHDs.

Efficacy of Hwangryunhaedok-tang (Huang-lian-jie-du-tang, Oren-gedoku-to) for patients with hyperlipidemia: a study protocol for a randomized, double-blind, placebo-controlled, parallel, investigator-initiated clinical trial.

BACKGROUND: The prevalence of hyperlipidemia continues to increase due to aging and lifestyle changes. Statins are currently used as the first choice for treating hyperlipidemia, but are limited by adverse reactions. Hwangryunhaedok-tang (HHT) has received attention as a promising intervention for hyperlipidemia through a few experimental and clinical trials. This study aims to explore the feasibility, effectiveness, and safety of HHT for hyperlipidemia treatment. METHODS: This is a study protocol for a randomized, double-blind, placebo-controlled, parallel, investigator-initiated, pilot clinical trial held in Daejeon, Republic of Korea. Thirty patients with hyperlipidemia will be randomly allocated to HHT or placebo granule groups in equal proportions. Participants will be administered HHT or placebo granules three times per day for 8 weeks and followed up for another 4 weeks. The primary outcome is low-density lipoprotein cholesterol at 8 weeks from the commencement of treatment. Other blood lipid parameters, biomarkers of atherosclerosis, the degree of arteriosclerosis, blood glucose parameters, blood pressure, anthropometric parameters, health-related quality of life, and the changes in the general symptoms of cold and hot patterns will be measured as secondary outcomes. Adverse events and laboratory test results will be investigated to assess the safety. Changes in the gut microbiome before and after intervention will also be assessed as an exploratory outcome through next-generation sequencing. Data will be recorded in electronic case report forms and analyzed using SAS® Version 9.4. DISCUSSION: This is a rigorously designed pilot clinical trial to explore the effect and safety of Hwangryunhaedok-tang compared to placebo control for patients with hyperlipidemia, thereby potentially facilitating better management of hyperlipidemia. The results of this pilot study could form the foundation for a future large-scale, confirmatory clinical trial. TRIAL REGISTRATION: Clinical Research Information Service KCT0004564 . Registered on December 18, 2019.

Hwangryunhaedok-Tang Exerts Neuropreventive Effect on Memory Impairment by Reducing Cholinergic System Dysfunction and Inflammatory Response in a Vascular Dementia Rat Model.

Hwangryunhaedok-tang (HRT) is a traditional oriental herbal formula used in Asian countries for treating inflammatory diseases and controlling fever. Our present study aimed to determine whether HRT has therapeutic effects for patients with vascular dementia (VaD) using a bilateral common carotid artery occlusion (BCCAO) rat model and assessing spatial memory impairment and activation of neuroinflammation. BCCAO was performed in male Sprague Dawley rats to induce VaD, and oral HRT was administered daily for 30 d. Our data showed that HRT ameliorated BCCAO-induced memory and cognitive impairment in behavioral tests. In addition, HRT reversed cholinergic dysfunction and neuronal damage in the hippocampus of BCCAO rats. Furthermore, HRT attenuated microglial activation and reduced the phosphorylation of p38 mitogen-activated protein kinase and c-Jun N-terminal kinase (JNK) induced by BCCAO. Simultaneous high-performance liquid chromatography analysis of HRT using index compounds from the herbal composition revealed that both HRT ethanol extract and commercial HRT granules primarily comprise geniposide, baicalin, and berberine. Our study showed that HRT administration resulted in the prevention of neuronal injury induced by BCCAO through improvement of cholinergic dysfunction and inhibition of neuroinflammatory responses, suggesting that HRT may have potential as a treatment for VaD.

Antiplatelet and Antithrombotic Activity of a Traditional Medicine, Hwangryunhaedok-Tang.

Platelet activation and accumulation at the site of vascular injury are central to thrombus formation resulted in thrombotic disorders. Medicinal herbs could be one of the most important pharmaceutical agents that ameliorate thrombotic disorders, such as unstable angina, myocardial infarction, stroke, and peripheral vascular diseases. Hwangryunhaedok-tang (HRT) is a traditional herbal medicine that displays multiple biological properties including anti-inflammatory abilities. However, its role in platelet activation has not been fully studied. Hence, we examined whether HRT has a potent inhibitory effect on platelet aggregation and thrombus formation. We demonstrated that HRT (30, 50, and 100 µg/ml) significantly impaired thrombin- and collagen-related peptide-induced platelet aggregation, granule secretion, thromboxane B2 generation, and intracellular Ca2+ mobilization. Biochemical studies revealed that HRT is involved in inhibiting the phosphorylation of phospholipase C and protein kinase B. The oral administration of HRT (30, 50, and 100 mg/kg once daily for 1 and/or 7 days) efficiently ameliorates ferric chloride induced arterial thrombus formation in vivo. Tail bleeding time was not significantly increased. The qualitative phytochemical constituents of the HRT extract were investigated using high-performance liquid chromatography. Our results demonstrated that HRT shows potential antiplatelet and antithrombotic effects without affecting hemostasis. Hence, HRT could be an effective therapeutic agent for the treatment of thrombotic diseases.

Hwangryunhaedok-tang induces the depolarization of pacemaker potentials through 5-HT3 and 5-HT4 receptors in cultured murine small intestine interstitial cells of Cajal.

AIM: To investigate the effects of a water extract of Hwangryunhaedok-tang (HHTE) on the pacemaker potentials of mouse interstitial cells of Cajal (ICCs). METHODS: We dissociated ICCs from small intestines and cultured. ICCs were immunologically identified using an anti-c-kit antibody. We used the whole-cell patch-clamp configuration to record the pacemaker potentials generated by cultured ICCs under the current clamp mode (I = 0). All experiments were performed at 30 °C-32 °C. RESULTS: HHTE dose-dependently depolarized ICC pacemaker potentials. Pretreatment with a 5-HT3 receptor antagonist (Y25130) or a 5-HT4 receptor antagonist (RS39604) blocked HHTE-induced pacemaker potential depolarizations, whereas pretreatment with a 5-HT7 receptor antagonist (SB269970) did not. Intracellular GDPßS inhibited HHTE-induced pacemaker potential depolarization and pretreatment with a Ca2+-free solution or thapsigargin abolished the pacemaker potentials. In the presence of a Ca2+-free solution or thapsigargin, HHTE did not depolarize ICC pacemaker potentials. In addition, HHTE-induced pacemaker potential depolarization was unaffected by a PKC inhibitor (calphostin C) or a Rho kinase inhibitor (Y27632). Of the four ingredients of HHT, Coptidis Rhizoma and Gardeniae Fructus more effectively inhibited pacemaker potential depolarization. CONCLUSION: These results suggest that HHTE dose-dependently depolarizes ICC pacemaker potentials through 5-HT3 and 5-HT4 receptors via external and internal Ca2+ regulation and via G protein-, PKC- and Rho kinase-independent pathways.

Effects of Hwangryunhaedok-tang on gastrointestinal motility function in mice.

AIM: To investigate the effects of Hwangryunhaedok-tang (HHT) on gastrointestinal (GI) motility in mice. METHODS: The effects of a boiling water extract of HHT (HHTE) on GI motility were investigated by calculating percent intestinal transit rates (ITR%) and gastric emptying (GE) values using Evans Blue and phenol red, respectively, in normal mice and in mice with experimentally induced GI motility dysfunction (GMD). In addition, the effects of the four components of HHT, that is, Gardeniae Fructus (GF), Scutellariae Radix (SR), Coptidis Rhizoma (CR), and Phellodendri Cortex (PC), on GI motility were also investigated. RESULTS: In normal ICR mice, ITR% and GE values were significantly and dose-dependently increased by the intragastric administration of HHTE (0.1-1 g/kg). The ITR% values of GMD mice were significantly lower than those of normal mice, and these reductions were significantly and dose-dependently inhibited by HHTE (0.1-1 g/kg). Additionally, GF, CR, and PC dose-dependently increased ITR% and GE values in normal and GMD mice. CONCLUSION: These results suggest that HHT is a novel candidate for the development of a gastroprokinetic agent for the GI tract.

Protective effect of HwangRyunHaeDok-Tang water extract against chronic obstructive pulmonary disease induced by cigarette smoke and lipopolysaccharide in a mouse model.

ETHNOPHARMACOLOGICAL RELEVANCE: Hwangryunhaedok-tang is an oriental herbal formula treated to cure inflammation and gastric disorders in China, Japan, and Korea. We explored the protective effects of Hwangryunhaedok-tang water extract (HRWE) against airway pathophysiological changes caused by cigarette smoke (CS) and lipopolysaccharide (LPS) in a mouse. MATERIALS AND METHODS: We performed quantitative analyses of five marker components, namely geniposide, baicalin, coptisine, plamatine, and berberine, using high-performance liquid chromatography. Animals were received CS exposure (1h per day) for 7 days. LPS was administered intranasally on day 4. Mice were received HRWE at dose of 100 or 200mg/kg for 1h before CS exposure. RESULTS: Treatment with HRWE significantly suppressed the increased inflammatory cell count induced by CS and LPS exposure. In addition, reduction in IL-6, TNF-α and IL-1ß in broncho-alveolar lavage fluid (BALF) was observed after HRWE treatment. HRWE not only decreased inflammatory cell infiltration in lung, but also decreased the expression of iNOS, NF-κB and matrix metallopeptidase (MMP)-9 in lung tissues. CONCLUSION: This study showed that HRWE can attenuate respiratory inflammation caused by CS and LPS exposure. Therefore, HRWE has potential for treating airway inflammatory disease.

Non-clinical safety assessment of Hwangryunhaedok-tang: 13-week toxicity in Crl:CD Sprague Dawley rats.

Hwangryunhaedok-tang (Huang-Lian-Jie-Du-Tang in Chinese, Oren-gedoku-to in Japanese) is a traditional herbal medicine with a long history of use for anti-inflammatory purposes. In this study, subchronic toxicity of daily oral administration of a Hwangryunhaedok-tang water extract (HHT) at 0, 250, 750, and 2000mg/kg for 13weeks was examined in rats. Mortality, clinical signs, and changes in body weight, food consumption, clinical signs, ophthalmological examination, urinalysis, hematology, serum biochemistry, gross observation, organ weight, and histopathology were monitored in accordance with Good Laboratory Practice and OECD guidelines. We found no mortality or abnormality in clinical signs, body weight, serum biochemistry, or organ weight in HHT-treated groups in either sex. However, there were significant changes in glucose, bilirubin, urobilinogen, protein (only male) in urine after 2000mg/kg/day HHT treatment for both sexes. In hematological examinations, we found a significant decreased number of red blood cells (RBC), whereas, an increased the mean corpuscular volume, number of platelets, and rate of reticulocyte (RET) after 2000mg/kg/day HHT treatment of male rats. In male and female rats, 750 and 2000mg/kg/day HHT treatment decreased the number of RBC and increased RET. Histopathological examinations revealed stomach mucosal erosion in female rats (2000mg/kg/day). No-observed-adverse-effect levels were established for 750mg/kg HHT in rats under the conditions of this study. However, other toxicological studies are necessary to evaluate the safety of HHT fully.

The alteration of components in the fermented Hwangryunhaedok-tang and its neuroprotective activity.

BACKGROUND: Hwangryunhaedok-tang is a traditional herbal prescription that has sedative activity, hypotensive and anti-bacterial effects. OBJECTIVE: In this study, we investigated the alteration of contents of components in Hwangryunhaedok-tang, antioxidant activity and neuroprotective activity by fermentation with Lactobacillus acidophilus KFRI 128. MATERIALS AND METHODS: Contents of three marker compounds (geniposide, berberine and palmatine) and unknown compounds in the Hwangryunhaedok-tang (HR) and the fermented Hwangryunhaedok-tang (FHR) were measured and compared using the established high-performance liqued chromatograph coupled with a photodiode (HPLC-DAD) method. The antioxidant activity of HR and FHR were determined by DPPH free radical and hydrogen peroxide (H(2)O(2)) scavenging assay. Also, the neuroprotective activities of HR and FHR against glutamate-induced oxidative stress in a mouse hippocampal cell line (HT22) were evaluated by MTT assay. RESULTS: The contents of geniposide and palmatine were decreased but the content of berberine was increased in the FHR. And the contents of unknown compounds (1), (2), (3), (4) and (5) in the HR were altered by fermentation. Electron donating activity (EDA, %) value of FHR was higher than HR for DPPH radical scavenging activity and H2O2 scavenging activity, respectively. In the MTT assay, FHR showed more potent neuroprotective activity than HR by 513.90%. CONCLUSION: The FHR using microorganism could convert compounds in HR and enhance the antioxidant and neuroprotective activity.