Asiaticasics A-O, structurally intriguing coumarins from Toddalia asiatica with potential inflammatory inhibitory activity.

Ethyl acetate fraction of Toddalia asiatica was fractionated to yield fifteen previously undescribed prenylated coumarins, asiaticasics A-O (1-15) along with nine (16-24) known derivatives. The structures of these undescribed coumarins were established by spectroscopic analysis and reference data. Biological activity evaluation showed that compound 3 with the IC50 value of 2.830 µM and compound 12 with the IC50 value of 0.682 µM owned anti-inflammatory activity by detecting the rate of lactate dehydrogenase release in pyroptosis J774A.1 cells. The results showed that the expression of Caspase-1 and IL-1ß was decreased in a dose-dependent manner in the compound 12 treatment group, suggesting that compound 12 may reduce pyroptosis by inhibiting NLRP3 inflammasome. To further determine that compound 12 treatment can inhibit macrophage pyroptosis, morphological observation was performed and the results were consistent with the bioactivity evaluation.

16,17-dinor-abietane diterpenoids from Casearia kurzii.

Eleven undescribed 16,17-dinor-abietane diterpenoids, caseazins A-K (1-11), and ten known diterpenoids (12-21) were isolated from the twigs and leaves of Casearia kurzii (Flacourtiaceae). Caseazins A-K were the first abietane -type dinorditerpenoids to have been isolated from the plant of Casearia kurzii. Their chemical structures were elucidated using a combination of 1D and 2D NMR spectroscopy and mass spectrometry. The absolute configurations of 5 and 10 were established by electronic circular dichroism calculations. Moreover, compounds 2, 3, 13, 14, and 18 exhibited anti-inflammatory activity with IC50 values of 0.17, 0.36, 6.55, 1.30, and 4.53 µM, respectively. IL-1ß and caspase-1 analyses suggested that compound 14 inhibited NLRP3 inflammasome activation and blocked macrophage pyroptosis.

Nudifloids A-N, structurally diverse 3,4-seco-labdane diterpenoids from Callicarpa nudiflora with inflammatory inhibitory activity.

Fourteen undescribed seco-type diterpenoids, named nudifloids A-N, together with ten known analogs, were isolated from the leaves of Callicarpa nudiflora. Nudifloids A-N had a characteristic 3,4-seco-labdane-type diterpenoid skeleton, whereas nudifloids A-C and K-N were 3,4-seco-norditerpenoids. Nudifloid A was the first example of a 3,4-seco-12,13,14,15,16-quartnor-labdane diterpenoid, with a seven-membered lactone ring formed through esterification between C-3 and C-11. Nudifloids B and C were a pair of highly modified 3,4-seco-labdane nor-diterpenoid epimers, of which C-2 and C-18 were cyclized together to form a cyclohexene fragment. The structures of these undescribed diterpenoids were established by spectroscopic analysis and reference data. The anti-inflammatory activity of diterpenoids in rich yield was evaluated by analyzing the inhibition of lipopolysaccharide plus nigericin-induced pyroptosis in J774A.1 cells. Nudifloids D and E exhibited prominent anti-NLRP3 inflammasome activity, with IC50 values of 1.80 and 1.59 µM, respectively. Cell permeability assays revealed that nudifloid D inhibited pyroptosis, which could ameliorate inflammation by blocking the activation of the NLRP3 inflammasome.

Diterpenoids inhibit ox-LDL-induced foam cell formation in RAW264.7 cells by promoting ABCA1 mediated cholesterol efflux.

Introduction: Atherosclerosis is the main cause of many cardiovascular diseases and contributes to morbidity and mortality worldwide. The formation of macrophage-derived foam cells plays a critical role in the early stage of atherosclerosis pathogenesis. Diterpenoids found in the flowers of Callicarpa rubella Lindl., a traditional Chinese medicine, have been reported to have anti-inflammatory activity. However, little is known about the effects of these diterpenoids on macrophage foam cell formation. Methods: A macrophage-derived foam cell formation model was established by treating RAW264.7 cells with oxidized low-density lipoprotein (ox-LDL) for 24 h. Oil red O staining were used to detect the intracellular lipids. The cholesterol efflux capacity was assayed by labeling cells with 22-NBD-cholesterol. Western blots and real-time PCRs were performed to quantify protein and mRNA expressions. Results: Two diterpenoid molecules, 14α-hydroxyisopimaric acid (C069002) and isopimaric acid (C069004), extracted from the flowers of Callicarpa rubella Lindl., significantly attenuated ox-LDL-induced foam cell formation in RAW264.7 macrophages. Further investigation showed that these two diterpenoids could promote cholesterol efflux from RAW264.7 macrophages to apolipoprotein A-I or high-density lipoproteins, which was associated with upregulated expression of ATP-binding cassette A1/G1 (ABCA1/G1), liver X receptor-α (LXRα), and peroxisome proliferator-activated receptor-γ (PPARγ). Unexpectedly, the diterpenoids C069002 and C069004 failed to enhance the mRNA transcription of the ABCG1 gene in macrophage-derived foam cells induced by ox-LDL. To evaluate the effects of diterpenoids on macrophage foam cell formation and determine the underlying mechanism, two drugs (lovastatin and rosiglitazone) were used as positive controls. Although both drugs could reduce macrophage foam cell formation and promote cholesterol efflux, they each had distinctive abilities to modulate the expression of cholesterol efflux-related genes. In contrast to lovastatin, rosiglitazone showed a similar influence on the expression of cholesterol efflux-related genes (including ABCA1, LXRα, and PPARγ) as the diterpenoids regardless of the presence or absence of ox-LDL, implying a similar mechanism by which they may exert atheroprotective effects. Conclusion: Our research indicates that diterpenoids effectively inhibit ox-LDL-induced macrophage foam cell formation by promoting cholesterol efflux from macrophages via the PPARγ-LXRα-ABCA1 pathway. Further investigation of diterpenoids as potential drugs for the treatment of atherosclerosis is warranted.

Caseatardies A-K, eleven undescribed clerodane diterpenoids isolated from Casearia tardieuae and their anti-inflammatory activity.

A phytochemical investigation to obtain bioactive substances as lead compounds or agents for anti-inflammatory led to the obtainment of eleven previously undescribed clerodane diterpenoids, named caseatardies A-K (1-11), and four known clerodane diterpenoids (12-15) from the twigs and leaves of Casearia tardieuae. The structural elucidation of these clerodane diterpenoids was based on 1D and 2D-NMR spectroscopy (COSY, HSQC, HMBC and ROESY) as well as high resolution mass spectrometry (HR-ESI-MS). The relative configurations were defined by ROESY correlations. The anti-inflammatory activity of all the isolated compounds was screened and compound 15 decreased LDH level in a dose-dependent manner, showing IC50 value of 2.89 µM.

Euphzycopias A-I, macrocyclic diterpenes with NLRP3 inflammasome inhibitory activity from Euphorbia helioscopia L.

A phytochemical investigation was conducted on Euphorbia helioscopia, resulting in the isolation of thirteen compounds, including nine undescribed diterpenoids, Euphzycopias A - I (1-9), of which the skeletons of compounds 1-4 were found in E. helioscopia L. Compounds 1-3 had 5/7/6 cyclic systems, while compound 4 had a 4/11 polycyclic system with a 4,7-cyclic ether between C-4 and C-7. The anti-inflammasome test using the isolated compounds (1-6, 8-13) showed that the diterpenes from E. helioscopia L. had a strong inhibitory effect on NLRP3 inflammasomes with IC50 values of 3.34-14.92 µM.

Exploring the mechanism of Buxue Yimu Pill on hemorrhagic anemia through molecular docking, network pharmacology and experimental validation.

Buxue Yimu Pill (BYP) is a classic gynecological medicine in China, which is composed of Angelica sinensis (Oliv.) Diels, Leonurus japonicus Houtt, Astragalus membranaceus (Fisch.) Bunge, Colla corii asini and Citrus reticulata Blanco. It has been widely used in clinical therapy with the function of enriching Blood, nourishing Qi, and removing blood stasis. The current study was designed to determine the bioactive molecules and therapeutic mechanism of BYP against hemorrhagic anemia. Herein, GC-MS and UPLC/Q-TOF-MS/MS were employed to identify the chemical compounds from BYP. The genecards database (https: //www.genecards.org/) was used to obtain the potential target proteins related to hemorrhagic anemia. Autodock/Vina was adopted to evaluate the binding ability of protein receptors and chemical ligands. Gene ontology and KEGG pathway enrichment analysis were conducted using the ClusterProfiler. As a result, a total of 62 candidate molecules were identified and 152 targets related to hemorrhagic anemia were obtained. Furthermore, 34 active molecules and 140 targets were obtained through the virtual screening experiment. The data of molecular-target (M-T), target-pathway (T-P), and molecular-target-pathway (M-T-P) network suggested that 32 active molecules enhanced hematopoiesis and activated the immune system by regulating 57 important targets. Pharmacological experiments showed that BYP significantly increased the counts of RBC, HGB, and HCT, and significantly down-regulated the expression of EPO, IL-6, CSF3, NOS2, VEGFA, PDGFRB, and TGFB1. The results also showed that leonurine, leonuriside B, leosibiricin, ononin, rutin, astragaloside I, riligustilide and levistolide A, were the active molecules closely related to enriching Blood. In conclusion, based on molecular docking, network pharmacology and validation experiment results, the enriching blood effect of BYP on hemorrhagic anemia may be associated with hematopoiesis, anti-inflammation, and immunity enhancement.

Gene therapy via canalostomy approach preserves auditory and vestibular functions in a mouse model of Jervell and Lange-Nielsen syndrome type 2.

Mutations in voltage-gated potassium channel KCNE1 cause Jervell and Lange-Nielsen syndrome type 2 (JLNS2), resulting in congenital deafness and vestibular dysfunction. We conducted gene therapy by injecting viral vectors using the canalostomy approach in Kcne1-/- mice to treat both the hearing and vestibular symptoms. Results showed early treatment prevented collapse of the Reissner's membrane and vestibular wall, retained the normal size of the semicircular canals, and prevented the degeneration of inner ear cells. In a dose-dependent manner, the treatment preserved auditory (16 out of 20 mice) and vestibular (20/20) functions in mice treated with the high-dosage for at least five months. In the low-dosage group, a subgroup of mice (13/20) showed improvements only in the vestibular functions. Results supported that highly efficient transduction is one of the key factors for achieving the efficacy and maintaining the long-term therapeutic effect. Secondary outcomes of treatment included improved birth and litter survival rates. Our results demonstrated that gene therapy via the canalostomy approach, which has been considered to be one of the more feasible delivery methods for human inner ear gene therapy, preserved auditory and vestibular functions in a dose-dependent manner in a mouse model of JLNS2.

Diterpenoids from Callicarpa rubella and their in vitro anti-NLRP3 inflammasome activity.

Nine new diterpenoids, Rubellacrns A - I (1-9), including five isopimaranes (1-4, 9), four pimaranes (5-8), together with five known isopimarane analogues (10-14), were isolated from Callicarpa rubella. The structures of these compounds were unambiguously established by HR-ESIMS and NMR spectroscopic data, the absolute configurations of compounds 5 and 9 were determined by ECD. All the isolated compounds were tested for their anti-inflammatory effects and compounds 2 and 11-14 showed NLRP3-inflammasome inhibitory activity with IC50 values ranging from 7.02 to 14.38 µM.

Toonaolides A-X, limonoids from Toona ciliata: Isolation, structural elucidation, and bioactivity against NLRP3 inflammasome.

Twenty-four new limonoids, toonaolides A-X (1-24), characterized with an α,ß-unsaturated-γ-lactone A-ring were isolated from the twigs of Toona ciliata. Their structures and absolute configurations were elucidated by spectroscopic data, X-ray diffraction crystallography, and quantum chemistry calculations. Most of the isolated compounds (except 9, 18, and 24 which possessed the maleimide ring) featured the rare 21-hydroxybutenolide or 23-hydroxybutenolide moieties. In particular, compound 1 has an unprecedented limonoid architecture with 6/6 cis-fused A/B ring system and 2 has an unusual tetrahydrofuran ring B skeleton, featuring a 7/5/6/5 ring system. The biological evaluation showed that compounds 9, 11, 12, 14, and 18 exhibited significantly anti-NLRP3 inflammasome activity with IC50 values ranging from 3.2 to 9.7 µM. Analysis of IL-1ß and caspase-1 expression revealed that compounds 11 and 12 are selective inhibitors of NLRP3 inflammasome, which could ameliorate cell pyroptosis by blocking NLRP3 inflammasome activation.

Multivariate analysis of prognostic factors for idiopathic sudden sensorineural hearing loss treated with adjuvant hyperbaric oxygen therapy.

The objective of this study is to evaluate possible prognostic factors of idiopathic sudden sensorineural hearing loss (ISSNHL) treated with adjuvant hyperbaric oxygen therapy (HBOT) using univariate and multivariate analyses. From January 2008 to October 2016, records of 178 ISSNHL patients treated with auxiliary hyperbaric oxygen therapy were reviewed to assess hearing recovery and evaluate associated prognostic factors (gender, age, localization, initial hearing threshold, presence of tinnitus, vertigo, ear fullness, hypertension, diabetes, onset of HBOT, number of HBOT, and audiogram), by using univariate and multivariate analyses. The overall recovery rate was 37.1%, including complete recovery (19.7%) and partial recovery (17.4%). According to multivariate analysis, later onset of HBOT and higher initial hearing threshold were associated with a poor prognosis in ISSNHL patients treated with HBOT. HBOT is a safe and beneficial adjuvant therapy for ISSNHL patients. 20 sessions of HBOT is possibly enough to show its therapeutic effect. Earlier HBOT onset and lower initial hearing threshold is associated with favorable hearing recovery.

Tetrastigma hemsleyanum (Sanyeqing) root tuber extracts induces apoptosis in human cervical carcinoma HeLa cells.

AIM OF THE STUDY: Tetrastigma hemsleyanum (Sanyeqing) is traditionally used as a folk medicine for the treatment of cancer. However, the underlying mechanisms remain unclear. The purpose of this study was to investigate the possible mechanisms by which petroleum ether fraction (PEF) of Sanyeqing has anti-tumor activity on HeLa cells. METHODS: The chemical components of PEF were analyzed by gas chromatography-mass spectrometry. The cytotoxicity of PEF on HeLa cells was measured by MTT assay. Apoptosis was evaluated by phosphatidylserine translocation, mitochondrial membrane potential (Δψm) changes and the activation of caspase-3, caspase-8 and caspase-9. The levels in T-SOD, CAT, GSH-PX and MDA were measured. RESULTS: PEF of Sanyeqing inhibited the growth and induced apoptosis of HeLa cells in dose- and time-dependent manner. PEF triggered intrinsic apoptotic pathway indicated by the loss of mitochondrial membrane potential and the activation of caspase-9 and caspase-3. In addition, PEF activated extrinsic apoptotic pathway indicated by the activation of caspase-8. Furthermore, PEF decreased T-SOD, CAT, GSH-PX activities and increased MDA level. Chemical analysis revealed the presence of fatty acids and phytosterol in PEF. CONCLUSIONS: PEF of Tetrastigma hemsleyanum Diels et. Gilg (Sanyeqing) exhibits cytotoxic effects, triggers both extrinsic and intrinsic apoptotic pathways, and augments oxidative stress in cervical carcinoma HeLa cells. Sanyeqing has strong potential to be developed as an agent for the treatment of cervical cancer.