Sikyungbanha-Tang Suppressing Acute Lung Injury in Mice Is Related to the Activation of Nrf2 and TNFAIP3.

Sikyungbanha-Tang (SKBHT) is a Chinese traditional medicine popularly prescribed to patients with respiratory inflammatory symptoms in Korea. Although the Korea Food and Drug Administration approved SKBHT as a therapeutics for relieving the symptoms, experimental evidence for SKBHT suppressing inflammation is scarce. Here, we presented evidence that SKBHT can suppress inflammation in an acute lung injury (ALI) mouse model and explored the possible underlying mechanisms of SKBHT's anti-inflammatory activity. Single intratracheal (i.t.) injection of SKBHT (1 mg/kg or 10 mg/kg body weight) into mouse lungs decreased prototypic features of lung inflammation found in ALI, such as a high level of proinflammatory cytokines, neutrophil infiltration, and the formation of hyaline membrane, which were induced by a single i.t. LPS (2 mg/kg body weight). When added to a murine macrophage RAW 264.7 cells, SKBHT activated an anti-inflammatory factor Nrf2, increasing the expression of genes regulated by Nrf2. SKBHT suppressed the ubiquitination of Nrf2, suggesting that SKBHT increases the level of and thus activates Nrf2 by blunting the ubiquitin-dependent degradation of Nrf2. SKBHT induced the expression of tumor necrosis factor α-induced protein 3 (TNFAIP3), an ubiquitin-modulating protein that suppresses various cellular signals to NF-κB. Concordantly, SKBHT suppressed NF-κB activity and the expression of inflammatory cytokine genes regulated by NF-κB. Given that Nrf2 and TNFAIP3 are involved in regulating inflammation, our results suggest that SKBHT suppresses inflammation in the lung, the effect of which is related to SKBHT activating Nrf2 and TNFAIP3.

The methanol extract of Guettarda speciosa Linn. Ameliorates acute lung injury in mice.

BACKGROUND: Guettarda speciosa is mainly found in tropical areas in Asia. Although G. speciosa is traditionally used to treat some of the inflammatory disorders, the experimental evidence supporting the anti-inflammatory effect of G. speciosa is limited. Here, we sought to obtain evidence that G. speciosa has anti-inflammatory activity using an acute lung injury (ALI) mouse model and to explore possible underlying mechanisms for the activity. METHODS: The methanol extract of G. speciosa Linn. (MGS) was fingerprinted by HPLC. Cytotoxicity was determined by MTT and flow cytometer. As for an ALI mouse model, C57BL/6 mice received an intratracheal (i.t.) injection of lipopolysaccharide (LPS). The effects of MGS on lung inflammation in the ALI mice were assessed by differential cell counting and FACS of inflammatory cells and hematoxylin and eosin staining of lung tissue. Proteins were analyzed by immunoprecipitation and immunoblotting, and gene expression was by real-time qPCR. Neutrophil elastase activity was measured by ELISA. RESULTS: MGS did not cause metabolic disarray or produce reactive oxygen species that could induce cytotoxicity. Similar to ALI patients, C57BL/6 mice that received an i.t. LPS developed a high level of neutrophils, increased pro-inflammatory cytokines, and inflicted tissue damage in the lung, which was suppressed by i.t. MGS administered at 2 h after LPS. Mechanistically, MGS activated Nrf2, which was related to MGS interrupting the ubiquitin-dependent degradation of Nrf2. MGS suppressed the nuclear localization of NF-κB induced by LPS, suggesting the inhibition of NF-κB activity. Furthermore, MGS inhibited the enzymatic activity of neutrophil elastase. CONCLUSION: MGS could suppress lung inflammation in an ALI mouse model, the effect of which could be attributed to multiple mechanisms, including the activation of Nrf2 and the suppression of NF-κB and neutrophil elastase enzymatic activity by MGS.

White Ginseng Ameliorates Depressive Behavior and Increases Hippocampal 5-HT Level in the Stressed Ovariectomized Rats.

Postmenopausal depression is closely associated with depletion of estrogen which modulates transmission of 5-HT, a key neurotransmitter that regulates stress-managing circuits in the brain. In this study, antidepressive efficacy of white ginseng (Panax gingseng Meyer, WG) was evaluated in stressed ovariectomized rats. Female Sprague Dawley rats were ovariectomized and repeatedly restraint stressed for 2 weeks (2h/day). Thirty minutes before restraint stress, rats were administered saline (control), WG 200 mg/kg (p.o.), WG 400 mg/kg (p.o.), or fluoxetine (PC, 10 mg/kg, i.p.). Tail suspension test (TST) and forced swimming test (FST) were performed to assess antidepressant effect of WG. After behavioral tests, levels of serum corticosterone (CORT) and hippocampal 5-HT were measured. Significant decrease of immobility time in TST and FST was shown in rats administered with PC or WG 400 compared to the control. WG200-treated rats showed remarkable reduction in immobility time of TST. PC, WG 200, or WG 400-administred group exhibited significant reduction of CORT compared to the control. PC or WG-treated rats exhibited remarkable increase in hippocampal 5-HT concentration compared to the control. Hippocampal 5-HT levels in WG groups were higher than those in the PC group. The present study demonstrated that WG had antidepressant efficacy in an animal model of menopausal depression. Treatment with WG enhanced hippocampal 5-HT level while suppressing depressive symptom and serum CORT level. These results provide evidence that WG plays an important role in activating serotonergic neurons in stressful situation, suggesting that WG might be a reliable natural alternative of antidepressant drugs to treat menopausal depression.

Suppression of lung inflammation by the ethanol extract of Chung-Sang and the possible role of Nrf2.

BACKGROUND: Asian traditional herbal remedies are typically a concoction of a major and several complementary herbs. While balancing out any adverse effect of the major herb, the complementary herbs could dilute the efficacy of the major herb, resulting in a suboptimal therapeutic effect of an herbal remedy. Here, we formulated Chung-Sang (CS) by collating five major herbs, which are used against inflammatory diseases, and tested whether an experimental formula composed of only major herbs is effective in suppressing inflammation without significant side effects. METHODS: The 50% ethanol extract of CS (eCS) was fingerprinted by HPLC. Cytotoxicity to RAW 264.7 cells was determined by an MTT assay and a flow cytometer. Nuclear NF-κB and Nrf2 were analyzed by western blot. Ubiquitinated Nrf2 was similarly analyzed following immunoprecipitation of Nrf2. Acute lung inflammation and sepsis were induced in C57BL/6 mice. The effects of eCS on lung disease were measured by HE staining of lung sections, a differential cell counting of bronchoalveolar lavage fluid, a myeloperoxidase (MPO) assay, a real-time qPCR, and Kaplan-Meier survival of mice. RESULTS: eCS neither elicited cytotoxicity nor reactive oxygen species. While not suppressing NF-κB, eCS activated Nrf2, reduced the ubiquitination of Nrf2, and consequently induced the expression of Nrf2-dependent genes. In an acute lung inflammation mouse model, an intratracheal (i.t.) eCS suppressed neutrophil infiltration, the expression of inflammatory cytokine genes, and MPO activity. In a sepsis mouse model, a single i.t. eCS was sufficient to significantly decrease mouse mortality. CONCLUSIONS: eCS could suppress severe lung inflammation in mice. This effect seemed to associate with eCS activating Nrf2. Our findings suggest that herbal remedies consisting of only major herbs are worth considering.

Hominis placenta Suppresses Acute Lung Inflammation by Activating Nrf2.

Hominis placenta (HP), a dried human placenta, has been known to target liver, lung, or kidney meridians, improving the functions associated with these meridians in traditional Chinese or Asian medicine (TCM). Since recent studies implicate an HP extract in suppressing inflammation, we investigated whether an aqueous HP extract can ameliorate inflammation that occurred in the lungs. When administered with a single intratracheal lipopolysaccharide (LPS), C57BL/6 mice developed an acute neutrophilic lung inflammation along with an increased expression of pro-inflammatory cytokine genes. However, this was diminished by the administration HP extract via an intraperitoneal route 2 h after LPS treatment. Western blot and semi-quantitative RT-PCR analyses revealed that while suppressing the activity of a proinflammatory factor NF-[Formula: see text]B marginally, the HP extract strongly activated an anti-inflammatory factor Nrf2, with concomitant expression of Nrf2-dependent genes. Mechanistically, the HP extract suppressed the ubiquitin-mediated degradation of Nrf2, functioning similarly to a 26S proteasome inhibitor, MG132. Collectively, these results suggest that the HP extract suppresses inflammation in mouse lungs, which is in part related to the HP extract perturbing the ubiquitin-dependent degradation of Nrf2 and thus increasing the function of Nrf2.

Electroacupuncture ameliorates poloxamer 407-induced hyperlipidemia through suppressing hepatic SREBP-2 expression in rats.

AIMS: Acupuncture, particularly electroacupuncture (EA) has been shown to have the lipid-lowering effects, but not completely investigated. The present study was aimed to examine whether EA could attenuate poloxamer-407 (P-407)-induced hyperlipidemia in the rats and to investigate its potential mechanisms. MAIN METHODS: Rats received P-407 (0.4 g/kg, i.p.) to induce hyperlipidemia. EA was performed at ST36 and ST40 acupoints a total of three times with 12 h-interval starting 1 h before the P-407 injection at 0.6 mA intensity and 2 Hz frequency for 10 min. KEY FINDINGS: In P-407-induced hyperlipidemic rats, EA stimulation at ST36 and ST40 acupoints significantly lowered the serum levels of triglycerides, total cholesterol, LDL-cholesterol and atherogenic index, while markedly increasing the serum HDL-cholesterol levels. Meanwhile, hyperlipidemic rats had significantly higher expression of sterol regulatory element-binding protein (SREBP)-2, without any difference in SREBP-1 expression in the liver, as compared with normal ones. EA significantly attenuated the expression of SREBP-2 with a subsequent decrease in 3-hydroxy-3-methylglutaryl coenzyme A reductase and an increase in low-density lipoprotein receptor at both mRNA and protein levels in the liver of hyperlipidemic rats. These changes did not occur after electrical stimulation at a non-acupoint. SIGNIFICANCE: Taken together, our findings indicate that EA stimulation to P-407-induced hyperlipidemic rats improves the lipid abnormalities, which may be associated with regulation of the expression of key enzymes of cholesterol synthesis in the liver through modulation of SREBP-2.

Differential Regulation of NF-κB and Nrf2 by Bojungikki-Tang Is Associated with Suppressing Lung Inflammation.

Bojungikki-tang (BT), an Asian herbal remedy, has been prescribed to increase the vitality of debilitated patients. Since a compromised, weakened vitality often leads to illness, BT has been widely used to treat various diseases. However, little is known about the mechanism by which BT exerts its effect. Given that BT ameliorates inflammatory pulmonary diseases including acute lung injury (ALI), we investigated whether BT regulates the function of key inflammatory factors such as NF-κB and Nrf2, contributing to suppressing inflammation. Results show that BT interrupted the nuclear localization of NF-κB and suppressed the expression of the NF-κB-dependent genes in RAW 264.7 cells. In similar experiments, BT induced the nuclear localization of Nrf2 and the expression of the Nrf2-dependent genes. In a lipopolysaccharide-induced ALI mouse model, a single intratracheal administration of BT to mouse lungs ameliorated alveolar structure and suppressed the expression of proinflammatory cytokine genes and neutrophil infiltration to mouse lungs. Therefore, our findings suggest that suppression of NF-κB and activation of Nrf2, by which BT suppresses inflammation, are ways for BT to exert its effect.

Effects of Electroacupuncture at BL60 on Formalin-Induced Pain in Rats.

Acupuncture was used to treat symptoms of pain in the ancient orient. The present study was conducted to determine the effects of electroacupuncture (EA) at the BL60 acupoint on male Sprague-Dawley rats. Each rat received EA at BL60 acupoint before formalin injection. Behavioral responses were recorded using a video camera and c-Fos immunohistochemistry was performed thereafter. Treatment of EA at BL60 significantly inhibited flinching behavior and c-fos expression induced by formalin injection into the paw, compared to a control group. These results suggest that electroacupuncture at BL60 acupoint may be effective in relieving inflammatory pain.