Total Chemical Synthesis of Correctly Folded Disulfide-Rich Proteins Using a Removable O-Linked ß-N-Acetylglucosamine Strategy.

Disulfide-rich proteins are useful as drugs or tool molecules in biomedical studies, but their synthesis is complicated by the difficulties associated with their folding. Here, we describe a removable glycosylation modification (RGM) strategy that expedites the chemical synthesis of correctly folded proteins with multiple or even interchain disulfide bonds. Our strategy comprises the introduction of simple O-linked ß-N-acetylglucosamine (O-GlcNAc) groups at the Ser/Thr sites that effectively improve the folding of disulfide-rich proteins by stabilization of their folding intermediates. After folding, the O-GlcNAc groups can be efficiently removed using O-GlcNAcase (OGA) to afford the correctly folded proteins. Using this strategy, we completed the synthesis of correctly folded hepcidin, an iron-regulating hormone bearing four pairs of disulfide-bonds, and the first total synthesis of correctly folded interleukin-5 (IL-5), a 26 kDa homodimer cytokine responsible for eosinophil growth and differentiation.

Comparison of different strategies towards the chemical synthesis of long-chain scorpion toxin AaH-II.

Long-chain scorpion toxin AaH-II isolated from Androctonus australis Hector can selectively inhibit mammalian voltage-gated sodium ion channel Nav 1.7 responsible for pain sensation. Efficient chemical synthesis of AaH-II and its derivatives is beneficial to the study of the function and mechanism of Nav 1.7 and the development of potential peptide inhibitors. Herein, we compared three different strategies, namely, direct solid-phase peptide synthesis, hydrazide-based two-segment native chemical ligation, and hydrazide-based three-segment native chemical ligation for the synthesis of AaH-II. The hydrazide-based two-segment native chemical ligation affords the target toxin with the optimal efficiency, which provides a practically robust procedure for the preparation of tool molecules derived from AaH-II to study the biological functions and modulation of Nav 1.7. Our work highlights the importance of selecting suitable segment condensation approach in the chemical synthesis of protein toxins.

Early Use of Corticosteroid May Prolong SARS-CoV-2 Shedding in Non-Intensive Care Unit Patients with COVID-19 Pneumonia: A Multicenter, Single-Blind, Randomized Control Trial.

BACKGROUND: There is still no clinical evidence available to support or to oppose corticosteroid treatment for coronavirus disease 2019 (COVID-19) pneumonia. OBJECTIVE: To investigate the efficacy and safety of corticosteroid given to the hospitalized patients with COVID-19 pneumonia. METHODS: This was a prospective, multicenter, single-blind, randomized control trial. Adult patients with COVID-19 pneumonia who were admitted to the general ward were randomly assigned to either receive methylprednisolone or not for 7 days. The primary end point was the incidence of clinical deterioration 14 days after randomization. RESULTS: We terminated this trial early because the number of patients with COVID-19 pneumonia in all the centers decreased in late March. Finally, a total of 86 COVID-19 patients underwent randomization. There was no difference of the incidence of clinical deterioration between the methylprednisolone group and control group (4.8 vs. 4.8%, p = 1.000). The duration of throat viral RNA detectability in the methylprednisolone group was 11 days (interquartile range, 6-16 days), which was significantly longer than that in the control group (8 days [2-12 days], p = 0.030). There were no significant differences between the 2 groups in other secondary outcomes. Mass cytometry discovered CD3+ T cells, CD8+ T cells, and NK cells in the methylprednisolone group which were significantly lower than those in the control group after randomization (p < 0.05). CONCLUSIONS: From this prematurely closed trial, we found that the short-term early use of corticosteroid could suppress the immune cells, which may prolong severe acute respiratory syndrome coronavirus 2 shedding in patients with COVID-19 pneumonia. TRIAL REGISTRATION: ClinicalTrials.gov, NCT04273321.

Effects of Glycosylation and d-Amino Acid Substitution on the Antitumor and Antibacterial Activities of Bee Venom Peptide HYL.

Glycosylation is a promising strategy for modulating the physicochemical properties of peptides. However, the influence of glycosylation on the biological activities of peptides remains unknown. Here, we chose the bee venom peptide HYL as a model peptide and 12 different monosaccharides as model sugars to study the effects of glycosylation site, number, and monosaccharide structure on the biochemical properties, activities, and cellular selectivities of HYL derivatives. Some analogues of HYL showed improvement not only in cell selectivity and proteolytic stability but also in antitumor and antimicrobial activity. Moreover, we found that the helicity of glycopeptides can affect its antitumor activity and proteolytic stability, and the α-linked d-monosaccharides can effectively improve the antitumor activity of HYL. Therefore, it is possible to design peptides with improved properties by varying the number, structure, and position of monosaccharides. What's more, the glycopeptides HYL-31 and HYL-33 show a promising prospect for antitumor and antimicrobial drugs development, respectively. In addition, we found that the d-lysine substitution strategy can significantly improve the proteolytic stability of HYL. Our new approach provides a reference or guidance for the research of novel antitumor and antimicrobial peptide drugs.

Total synthesis and cyclization strategy of samoamide A, a cytotoxic cyclic octapeptide rich in proline and phenlalanine isolated from marine cyanobacterium.

Samoamide A is a cyclic octapeptide rich in proline and phenylalanine residues isolated from an American Samoa marine cyanobacterium, which exhibits potent activity against H460 human non-small-cell lung cancer cells (IC50 of 1.1 µM). The first total synthesis of samoamide A was achieved by employing a strategy of a solid-phase peptide synthesis (SPPS) and a head-to-tail cyclization selecting free steric-hinrance connection sites. Then the final product was purified and identified. This strategy not only provides a basis in producing potent cytotoxic agents for drug discovery, but also provides a reference to the total synthesis of proline-rich peptides.

The opioid receptor triple agonist DPI-125 produces analgesia with less respiratory depression and reduced abuse liability.

Opioid analgesics remain the first choice for the treatment of moderate to severe pain, but they are also notorious for their respiratory depression and addictive effects. This study focused on the pharmacology of a novel opioid receptor mixed agonist DPI-125 and attempted to elucidate the relationship between the δ-, µ- and κ-receptor potency ratio and respiratory depression and abuse liability. Five diarylmethylpiperazine compounds (DPI-125, DPI-3290, DPI-130, KUST202 and KUST13T02) were selected for this study. PKA fluorescence redistribution assays in CHO cells individually expressing δ-, µ- or κ-receptors were used to measure the agonist potency. The respiratory safety profiles were estimated in rats by the ratio of ED50 (pCO2 increase)/ED50 (antinociception). The abuse liability of DPI-125 was evaluated with a self-administration model in rhesus monkeys. The observed agonist potencies of DPI-125 for δ-, µ- and κ-opioid receptors were 4.29±0.36, 11.10±3.04, and 16.57±4.14 nmol/L, respectively. The other four compounds were also mixed agonists with varying potencies. DPI-125 exhibited a high respiratory safety profile, clearly related to its high δ-receptor potency. The ratio of the EC50 potencies for the µ- and δ-receptors was found to be positively correlated with the respiratory safety ratio. DPI-125 has similar potencies for µ- and κ-receptors, which is likely the reason for its reduced abuse potential. Our results demonstrate that the opioid receptor mixed agonist DPI-125 is safer and less addictive than traditional µ-agonist analgesics. These findings suggest that the development of δ>µâˆ¼κ opioid receptor mixed agonists is feasible, and such compounds could represent a promising class of potent analgesics with wider therapeutic windows.

Solitary primary pulmonary synovial sarcoma: A case report.

BACKGROUND: Synovial sarcoma (SS) is an uncommon and highly malignant soft tissue sarcoma in the clinic, with primary pulmonary SS (PPSS) being extremely rare. Here, we describe the clinical characteristics, diagnosis, and treatment of a solitary PPSS case confirmed via surgical resection and fluorescence in situ hybridization (FISH). CASE SUMMARY: A 33-year-old man was admitted because of intermittent coughing and hemoptysis for one month, with lung shadows observed for two years. Whole-body positron emission tomography-computed tomography (PET-CT) revealed a solitary mass in the upper lobe of the right lung, with uneven radioactivity uptake and a maximum standardized uptake value of 5.6. The greyish-yellow specimen obtained following thoracoscopic resection was covered with small multi-nodulated structures and consisted of soft tissue. Hematoxylin and eosin staining revealed spindle-shaped malignant tumor cells. Immunohistochemistry indicated these tumor cells were CD99 and BCL-2-positive. Furthermore, the FISH test revealed synovial sarcoma translocation genetic reassortment, which confirmed the diagnosis of SS. CONCLUSION: PPSS is extremely rare and tends to be misdiagnosed as many primary pulmonary diseases. PET-CT, histologic analysis, and FISH tests can be used to differentiate PPSS from other diseases. Surgical resection is regularly recommended for the treatment of solitary PPSS and is helpful for improving the prognosis.

Pharmacological mechanisms underlying the antinociceptive and tolerance effects of the 6,14-bridged oripavine compound 030418.

AIM: To investigate possible pharmacological mechanisms underlying the antinociceptive effect of and tolerance to N-methyl-7α-[(R)-1-hydroxy-1-methyl-3-(thien-3-yl)-propyl]-6,14-endo-ethanotetrahydronororipavine (030418), a derivative of thienorphine. METHODS: The binding affinity and efficacy of 030418 were determined using receptor binding and guanosine 5'-O-(3-[(35)S]thio)triphosphate ([(35)S]GTPγS) assays in CHO-µ, CHO-κ, CHO-δ, and CHO-ORL1 cell membranes. The analgesic activity of and tolerance to 030418 were evaluated in thermal nociceptive tests in mice. The effects of 030418 on opioid receptors were further investigated using in vivo pharmacological antagonist blockade and in vitro tissue preparations. RESULTS: The compound 030418 displayed high binding affinity to all subtypes of opioid receptors with K(i) values in the nanomolar range. In [(35)S]GTPγS binding assay, the maximal stimulation of 030418 to µ-, κ-, δ-receptors and the ORL1 receptor was 89%, 86%, 67% and 91%, respectively. In hot-plate test, the antinociceptive effect of 030418 was more potent and longer than morphine. The nonselective opioid receptor antagonist naloxone could completely block 030418-induced antinociception, while both the µ-opioid receptor antagonist ß-FNA and the κ-opioid receptor antagonist nor-BNI attenuated 030418-induced antinociception. In contrast, the ORL1 receptor antagonist J-113397 enhanced the antinociceptive effect of 030418. Additionally, chronic treatment with 030418 resulted in a dramatic development of tolerance that could not be effectively prevented by J-113397. In guinea pig ileum preparation, the existing action of 030418 could be removed with difficulty after prolonged washing. CONCLUSION: The compound 030418 is a novel agonist of opioid receptors with high efficiency, long-lasting effect and liability to tolerance, which may be closely correlated with the methyl group at the N(17) position and the high hydrophobicity of the C(7)-thiophene group in its chemical structure.

[Primary investigation on heterodimerization of kappa-opioid receptor and ORL1 receptor].

This study investigates whether kappa-opioid receptor and ORL1 receptor may interact to form a heterodimer. In immunofluorescence and co-immunoprecipitation experiments, differentially epitope-tagged receptors, colocalization and heterodimerization of kappa-opioid receptor and ORL1 receptor were used and examined in primary culturing rat neurons, Chinese hamster ovary (CHO) or human embryonic kidney 293 (HEK293) cells. The results show that fluorescence of both kappa-opioid receptor and ORL1 receptor were overlapping in primary culturing hippocampal and cortical neurons. Similarly in co-expressing CHO or HEK293 cells, HA-KOR and Myc-ORL1 were almost exclusively confined to the membranes, revealing extensive colocalization. When Flag-KOR and Myc-ORL1 were co-expressing in CHO cells, heterodimerization was identified to have the ability to co-immunoprecipitate ORL1-receptors with kappa-opioid receptor and vice versa. In the current study, further evidence was provided for the direct interaction of two subtypes of opioid receptors, kappa-opioid receptor and ORL1-receptor, to form the heterodimerization. The finding represents the novel pharmacological mechanism for modulation of opioid receptor function as well as diversity of G protein-coupled receptors.

Flumazenil-Insensitive Benzodiazepine Effects in Recombinant αß and Neuronal GABAA Receptors.

Gamma-aminobutyric acid, type A (GABAA) receptors are complex heterogeneous pentamers with various drug binding sites. Several lines of evidence suggest that benzodiazepines modulate certain GABAA receptors in a flumazenil-insensitive manner, possibly via binding sites other than the classical ones. However, GABAA receptor subtypes that contain non-classical benzodiazepine binding sites are not systemically studied. The present study investigated the high-concentration effects of three benzodiazepines and their sensitivity to flumazenil on different recombinant (α1ß2, α2ß2, α3ß2, α4ß2, α5ß2 and α1ß3) and native neuronal GABAA receptors using the whole-cell patch-clamp electrophysiology technique. The classical benzodiazepine diazepam (200 µmol/L) and midazolam (200 µmol/L) produced flumazenil-insensitive effects on α1ß2 receptor, whereas the imidazopyridine zolpidem failed to modulate the receptor. Flumazenil-insensitive effects of diazepam were also observed on the α2ß2, α3ß2 and α5ß2, but not α4ß2 receptors. Unlike ß2-containing receptors, the α1ß3 receptor was insensitive to diazepam. Moreover, the diazepam (200 µmol/L) effects on some cortical neurons could not be fully antagonized by flumazenil (200 µmol/L). These findings suggested that the non-classical (flumazenil-insensitive) benzodiazepine effects depended on certain receptor subtypes and benzodiazepine structures and may be important for designing of subtype- or binding site- specific drugs.

Wnt1/ß-catenin signaling upregulates spinal VGLUT2 expression to control neuropathic pain in mice.

Vesicular glutamate transporter 2 (VGLUT2)-which uptakes glutamate into presynaptic vesicles-is a fundamental component of the glutamate neurotransmitter system. Although several lines of evidence from genetically modified mice suggest a possible association of VGLUT2 with neuropathic pain, the specific role of VGLUT2 in the spinal cord during neuropathic pain, and its regulatory mechanism remain elusive. In this study, we report that spared nerve injury induced an upregulation of VGLUT2 in the spinal cord, and intrathecal administration of small hairpin RNAs (shRNA) against VGLUT2 before or after surgery attenuated mechanical allodynia, and pathologically-enhanced glutamate release. Meanwhile, nerve injury activated the Wnt1/ß-catenin signaling pathway in a quick-onset and sustained manner, and blocking the Wnt1 signaling with a Wnt1 targeting antibody attenuated neuropathic pain. In naïve mice, administration of a Wnt agonist or Wnt1 increased spinal VGLUT2 protein levels. Moreover, intrathecal administration of the Wnt/ß-catenin inhibitor, XAV939 attenuated mechanical allodynia, and this effect was concurrent with that of VGLUT2 downregulation. Pretreatment with VGLUT2 shRNAs abolished the allodynia induced by the Wnt agonist or Wnt1. These findings reveal a novel mechanism wherein there is Wnt1/ß-catenin-dependent VGLUT2 upregulation in neuropathic pain, thus potentiating the development of new therapeutic strategies in pain management.

Hospitalization and Critical Care of 109 Decedents with COVID-19 Pneumonia in Wuhan, China.

Rationale: The current outbreak of coronavirus disease (COVID-19) pneumonia caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in Wuhan, China, spreads across national and international borders. The overall death rate of COVID-19 pneumonia in the Chinese population was 4%.Objectives: To describe the process of hospitalization and critical care of patients who died of COVID-19 pneumonia.Methods: This was a multicenter observational study of 109 decedents with COVID-19 pneumonia from three hospitals in Wuhan. Demographic, clinical, laboratory, and treatment data were collected and analyzed, and the final date of follow-up was February 24, 2020.Results: The mean age of 109 decedents with COVID-19 pneumonia was 70.7 years, 35 patients (32.1%) were female, and 85 patients (78.0%) suffered from one or more underlying comorbidities. Multiple organ failure, especially respiratory failure and heart failure, appeared in all patients even at the early stage of disease. Overall, the mean time from onset of symptoms to death was 22.3 days. All 109 hospitalized patients needed admission to an intensive care unit (ICU); however, because of limited availability, only 51 (46.8%) could be admitted. The period from hospitalization to death in the ICU group and non-ICU group was 15.9 days (standard deviation = 8.8 d) and 12.5 days (8.6 d, P = 0.044), respectively.Conclusions: Mortality due to COVID-19 pneumonia was concentrated in patients above the age of 65 years, especially those with major comorbidities. Patients who were admitted to the ICU lived longer than those who were not. Our findings should aid in the recognition and clinical management of such infections, especially with regard to ICU resource allocation.

[Effect of spinal glutamate transporter 1 on chronic constriction injury of sciatic nerve and morphine tolerance of rats].

In order to investigate the role of spinal glutamate transporter 1 (GLT-1) in the neuropathic pain and morphine tolerance, rat chronic constriction injury (CCI) of sciatic nerve was performed, and the mechanical allodynia was evaluated by mechanical withdrawal threshold (MWT), the expression of GLT-1 was measured by real-time PCR and Western blotting analysis. The results showed that compared to sham group, the MWT of CCI group had decreased approximately 80%. Administration of morphine alone could develop tolerance rapidly in initial two days, and then had no significant difference with CCI group, the expression of GLT-1 was down-regulated. Ceftriaxone sodium alone could improve mechanical allodynia. Co-administration of ceftriaxone sodium with morphine attenuated morphine tolerance and up-regulated GLT-1 expression, and the MWT remained at high level after 6 days. In conclusion, change of spinal GLT-1 expression and function has close correlation with the development of neuropathic pain and morphine tolerance.

The Impact and Mechanism of a Novel Allosteric AMPA Receptor Modulator LCX001 on Protection Against Respiratory Depression in Rodents.

Analgesics and sedative hypnotics in clinical use often give rise to significant side effects, particularly respiratory depression. For emergency use, specific antagonists are currently administered to counteract respiratory depression. However, antagonists are often short-lasting and eliminate drug generated analgesia. To resolve this issue, novel positive AMPA modulators, LCX001, was tested to alleviate respiratory depression triggered by different drugs. The acetic acid writhing and hot-plate test were conducted to evaluate analgesic effect of LCX001. Binding assay, whole-cell recording, live cell imaging, and Ca2+ imaging were used to clarify mechanism and impact of LCX001 on respiratory protection. Results showed that LCX001 effectively rescued and prevented opioid (fentanyl and TH-030418), propofol, and pentobarbital-induced respiratory depression by strengthening respiratory frequency and minute ventilation. The acetic acid writhing test and hot-plate test revealed potent anti-nociceptive efficacy of LCX001, in contrast to other typical ampakines that did not affect analgesia. Furthermore, LCX001 potentiated [3H]AMPA and L-glutamate binding affinity to AMPA receptors, and facilitated glutamate-evoked inward currents in HEK293 cells stably expressing GluA2(R). LCX001 had a typical positive modulatory impact on AMPAR-mediated function. Importantly, application of LCX001 generated a significant increase in GluA2(R) surface expression, and restrained opioid-induced abnormal intracellular Ca2+ load, which might participate in breathing modulation. Our study improves therapeutic interventions for the treatment of drug induced respiratory depression, and increases understanding of potential mechanism of AMPA receptor modulators.

Effects of taurine on rat behaviors in three anxiety models.

In our previous studies using an elevated plus-maze test in mice, taurine was shown to present an anxiolytic-like effect after single and repeated administration. The aim of the present study was to investigate the anxiolytic and behavioral effects of taurine on rats in the open field, hole-board, and social interaction test compared to the positive control diazepam. Taurine (14, 42, and 126 mg/kg, i.p.) was administered 30 min before the tests. In the social interaction and hole-board tests, taurine (42 mg/kg) significantly increased social interaction time and the number and duration of head-dipping. In the open field test, taurine (126 mg/kg, i.p.) presented anxiolytic-like effects by increasing the number of center entries, time spent in the central area and the anti-thigmotactic score while having no effect on the locomotor activity. Results from these experiments suggest that taurine produces an anxiolytic-like effect in these animal models and may act as a modulator or anti-anxiety agent in the central nervous system.

Anxiolytic-like effect of asiaticoside in mice.

The putative anxiolytic activity of asiaticoside was examined in male mice by using a number of experimental paradigms of anxiety, with diazepam being as a positive anxiolytic control. In the elevated plus-maze test, diazepam (1 and 2 mg/kg) or asiaticoside (5 or 10 mg/kg) increased the percentage of entries into open arms and of time spent on open arms. In the light/dark test, as with 1 mg/kg diazepam, asiaticoside (10 and 20 mg/kg) increased the time spent in the light area and the movement in the light area without altering the total locomotor activity of the animals. In the hole-board test, asiaticoside at 10 mg/kg significantly increased head-dipping counts and duration as well as diazepam (0.3 mg/kg). Thus, these findings indicated that asiaticoside exhibited an anxiolytic-like effect. Further studies will be required to assess the generality of present findings to other species and behavioural paradigms.

Resolving the α-glycosidic linkage of arginine-rhamnosylated translation elongation factor P triggers generation of the first ArgRha specific antibody.

A previously discovered posttranslational modification strategy - arginine rhamnosylation - is essential for elongation factor P (EF-P) dependent rescue of polyproline stalled ribosomes in clinically relevant species such as Pseudomonas aeruginosa and Neisseria meningitidis. However, almost nothing is known about this new type of N-linked glycosylation. In the present study we used NMR spectroscopy to show for the first time that the α anomer of rhamnose is attached to Arg32 of EF-P, demonstrating that the corresponding glycosyltransferase EarP inverts the sugar of its cognate substrate dTDP-ß-l-rhamnose. Based on this finding we describe the synthesis of an α-rhamnosylated arginine containing peptide antigen in order to raise the first anti-rhamnosyl arginine specific antibody (anti-ArgRha). Using ELISA and Western Blot analyses we demonstrated both its high affinity and specificity without any cross-reactivity to other N-glycosylated proteins. Having the anti-ArgRha at hand we were able to visualize endogenously produced rhamnosylated EF-P. Thus, we expect the antibody to be not only important to monitor EF-P rhamnosylation in diverse bacteria but also to identify further rhamnosyl arginine containing proteins. As EF-P rhamnosylation is essential for pathogenicity, our antibody might also be a powerful tool in drug discovery.

Behavioral effects of sinomenine in murine models of anxiety.

The present study was designed to investigate the putative anxiolytic-like effect of sinomenine in three experimental models of anxiety in male rats and mice. Use of the elevated plus-maze test revealed that sinomenine (20 and 40 mg/kg, p.o.) increased the percentage of open arm entries and diazepam (2 mg/kg, p.o.) increased the percentage of open arm entries, the percentage of time spent on open arms and total arm entries in mice. In the light/dark transition test, sinomenine (20 and 40 mg/kg, p.o.) increased time spent in the light area and diazepam (2 mg/kg, p.o.) increased time spent in the light area and the overall movements in mice. In the social interaction test, the sinomenine-treated animals significantly increased social interaction time in low light unfamiliar (7 mg/kg, p.o.) and high light unfamiliar conditions (7 and 14 mg/kg, p.o.) as well as diazepam (3 mg/kg, p.o.). Sinomenine (28 mg/kg, p.o.) can also decrease squares entered in rats in social interaction test under low light unfamiliar condition. In the open-field test, sinomenine (160 mg/kg) decreased squares entered in mice. Thus, these findings indicated that sinomenine exhibited anxiolytic-like effect.

Anxiolytic-like effect of paeonol in mice.

The present study in mice compared the putative anxiolytic-like effect of paeonol, a phenolic component from the root bark of Paeonia moutan, with the benzodiazepine diazepam in the elevated plus maze and the light/dark box-test. The comparison was also with regard to locomotor activity (open-field test) and myorelaxant potential (inclined plane test). As with 2 mg/kg diazepam, paeonol (at 17.5 mg/kg) increased the percentage of time spent on open arms in the elevated plus maze and increased the time spent in the light area of the light/dark box (at 8.75 and 17.5 mg/kg). Since paeonol, in contrast to diazepam, had no effect on either the number of squares entered in the open-field test or in the inclined plane test, its side-effect profile is considered as superior to the benzodiazepine.

The effects of angelica essential oil in social interaction and hole-board tests.

In our previous studies, we have demonstrated the anxiolytic effects of angelica essential oil in three anxiety models using mice. This study aimed to characterize the similar behavior effects of angelica essential oil in the social interaction test of anxiety and the hole-board test of exploration and locomotor activity in rats. These results indicate that angelica essential oil possessed a wide range of anxiolytic properties. In the social interaction test, angelica essential oil decreased aggressive behaviors at the doses of 21 and 42 mg/kg, while the doses of 21 and 42 mg/kg significantly increased social interaction time of the high light, unfamiliar test condition and 21 mg/kg could also prolong social interaction time of the high light, familiar test condition. In the hole-board test, angelica essential oil at 10.5 mg/kg significantly increased head-dipping counts and duration. Thus, our findings suggest the potential usefulness of angelica essential oil against various types of anxiety-related disorders and social failure.