Discovery and mechanistic study of Imperatorin that inhibits HBsAg expression and cccDNA transcription.

Chronic hepatitis B virus (HBV) infection remains a significant global health challenge due to its link to severe conditions like HBV-related cirrhosis and hepatocellular carcinoma (HCC). Although current treatments effectively reduce viral levels, they have limited impact on certain HBV elements, namely hepatitis B surface antigen (HBsAg) and covalently closed circular DNA (cccDNA). This highlights the urgent need for innovative pharmaceutical and biological interventions that can disrupt HBsAg production originating from cccDNA. In this study, we identified a natural furanocoumarin compound, Imperatorin, which markedly inhibited the expression of HBsAg from cccDNA, by screening a library of natural compounds derived from Chinese herbal medicines using ELISA assay and qRT-PCR. The pharmacodynamics study of Imperatorin was explored on HBV infected HepG2-NTCP/PHHs and HBV-infected humanized mouse model. Proteome analysis was performed on HBV infected HepG2-NTCP cells following Imperatorin treatment. Molecular docking and bio-layer interferometry (BLI) were used for finding the target of Imperatorin. Our findings demonstrated Imperatorin remarkably reduced the level of HBsAg, HBV RNAs, HBV DNA and transcriptional activity of cccDNA both in vitro and in vivo. Additionally, Imperatorin effectively restrained the actions of HBV promoters responsible for cccDNA transcription. Mechanistic study revealed that Imperatorin directly binds to ERK and subsequently interfering with the activation of CAMP response element-binding protein (CREB), a crucial transcriptional factor for HBV and has been demonstrated to bind to the PreS2/S and X promoter regions of HBV. Importantly, the absence of ERK could nullify the antiviral impact triggered by Imperatorin. Collectively, the natural compound Imperatorin may be an effective candidate agent for inhibiting HBsAg production and cccDNA transcription by impeding the activities of HBV promoters through ERK-CREB axis.

Vasorelaxant and Blood Pressure-Lowering Effects of Cnidium monnieri Fruit Ethanol Extract in Sprague Dawley and Spontaneously Hypertensive Rats.

Cnidium monnieri (L.) Cusson, a member of the Apiaceae family, is rich in coumarins, such as imperatorin and osthole. Cnidium monnieri fruit (CM) has a broad range of therapeutic potential that can be used in anti-bacterial, anti-cancer, and sexual dysfunction treatments. However, its efficacy in lowering blood pressure through vasodilation remains unknown. This study aimed to assess the potential therapeutic effect of CM 50% ethanol extract (CME) on hypertension and the mechanism of its vasorelaxant effect. CME (1-30 µg/mL) showed a concentration-dependent vasorelaxation on constricted aortic rings in Sprague Dawley rats induced by phenylephrine via an endothelium-independent mechanism. The vasorelaxant effect of CME was inhibited by blockers of voltage-dependent and Ca2+-activated K+ channels. Additionally, CME inhibited the vascular contraction induced by angiotensin II and CaCl2. The main active compounds of CM, i.e., imperatorin (3-300 µM) and osthole (1-100 µM), showed a concentration-dependent vasorelaxation effect, with half-maximal effective concentration values of 9.14 ± 0.06 and 5.98 ± 0.06 µM, respectively. Orally administered CME significantly reduced the blood pressure of spontaneously hypertensive rats. Our research shows that CME is a promising treatment option for hypertension. However, further studies are required to fully elucidate its therapeutic potential.

Furanocoumarins as Enhancers of Antitumor Potential of Sorafenib and LY294002 toward Human Glioma Cells In Vitro.

Furanocoumarins are naturally occurring compounds in the plant world, characterized by low molecular weight, simple chemical structure, and high solubility in most organic solvents. Additionally, they have a broad spectrum of activity, and their properties depend on the location and type of attached substituents. Therefore, the aim of our study was to investigate the anticancer activity of furanocoumarins (imperatorin, isoimperatorin, bergapten, and xanthotoxin) in relation to human glioblastoma multiforme (T98G) and anaplastic astrocytoma (MOGGCCM) cell lines. The tested compounds were used for the first time in combination with LY294002 (PI3K inhibitor) and sorafenib (Raf inhibitor). Apoptosis, autophagy, and necrosis were identified microscopically after straining with Hoechst 33342, acridine orange, and propidium iodide, respectively. The levels of caspase 3 and Beclin 1 were estimated by immunoblotting and for the blocking of Raf and PI3K kinases, the transfection with specific siRNA was used. The scratch test was used to assess the migration potential of glioma cells. Our studies showed that the anticancer activity of furanocoumarins strictly depended on the presence, type, and location of substituents. The obtained results suggest that achieving higher pro-apoptotic activity is determined by the presence of an isoprenyl moiety at the C8 position of the coumarin skeleton. In both anaplastic astrocytoma and glioblastoma, imperatorin was the most effective in induction apoptosis. Furthermore, the usage of imperatorin, alone and in combination with sorafenib or LY294002, decreased the migratory potential of MOGGCCM and T98G cells.

Detection and identification of imperatorin metabolites in rat, dog, monkey, and human liver microsomes by ultra-high-performance liquid chromatography combined with high-resolution mass spectrometry and Compound Discoverer software.

Imperatorin, a furanocoumarin that widely exists in many umbelliferous herbs, has been demonstrated to have a variety of pharmacological effects, including anti-inflammatory, antiosteoporosis, and antitumor activities. The purpose of this study was to investigate the metabolism of imperatorin using liver microsomes. The metabolites were generated by individually incubating imperatorin with rat, dog, monkey, and human liver microsomes. To trap the reactive metabolites during microsomal metabolism, glutathione (GSH) was included in the incubation. A LC technique coupled with benchtop orbitrap MS with full mass/data-dependent tandem mass spectrometry acquisition mode was used to detect and identify the generated metabolites. The possible structures of the metabolites were characterized according to their accurate masses and fragment ions. Under the current conditions, a total of 10 metabolites, including four GSH adducts, were identified. The results indicated that imperatorin underwent extensive metabolic reactions including hydroxylation, oxidation, glucuronidation, and GSH conjugation. This study provides essential data on the metabolism of imperatorin, which will be helpful for us to understand the safety and efficacy of this bioactive compound.

Imperatorin inhibits LPS-induced bone marrow-derived macrophages activation by decreased NF-κB p65 phosphorylation.

BACKGROUND: Imperatorin (IMP) is a secondary metabolite of plants and is the most abundant in Angelica dahurica. Previous studies showed that IMP exhibited anti-inflammatory activity in RAW264.7 cell line. Here, we aim to investigate the roles and mechanisms of IMP in bone marrow-derived macrophages (BMDMs), in view of the difference between primary macrophages and cell lines. METHODS: BMDMs were stimulated with LPS for the inflammation model. Flow cytometry was performed with BMDMs treated with different doses of IMP (0-20mg/L) within staining Annexin V-APC for 5 min. The cytokines and inflammatory mediators were detected by RT-PCR or ELISA. RNA-seq was performed in IMP-treated BMDMs or control, stimulated with LPS for 6h. Western blotting is carried out to determine the phosphorylation of p65, ERK1/2, JNK1, p38, and Akt. RESULTS: Our results showed that IMP inhibited IL-12p40, IL-6, TNF-α and IL-1ß in LPS-stimulated BMDMs. RNA-seq analysis suggested that IMP inhibits Toll-like receptor signaling pathway (KEGG), TNF signaling pathway (KEGG), NF-κB signaling pathway (KEGG), Inflammatory Response (GO). In addition, IMP inhibited myd88, tpl2, cxcl1, ptgs2(COX-2) expression in mRNA level. Finally, we found decreased phosphorylation of NF-κB p65 in IMP-treated BMDMs, after stimulated with LPS. CONCLUSION: IMP inhibits IL-12p40, IL-6, TNF-α, and IL-1ß expression in LPS-stimulated BMDMs. IMP inhibits macrophage activation, which maybe resulted in decreased phosphorylation of NF-κB p65. Furthermore, IMP may protect against the progress of inflammatory-related diseases.

The Assessment of the Efficacy of Imperatorin in Reducing Overactive Bladder Symptoms.

Overactive bladder syndrome (OAB) is a prevalent condition that affects the elderly population in particular and significantly impairs quality of life. Imperatorin, a naturally occurring furocoumarin, possesses diverse pharmacological properties that warrant consideration for drug development. The aim of this study was to investigate the potential of imperatorin (IMP) to attenuate the cystometric and biochemical changes typically associated with retinyl acetate-induced overactive bladder (OAB) and to assess its viability as a pharmacological intervention for OAB patients. A total of 60 rats were divided into four groups: I-control, II-rats with rapamycin (RA)-induced OAB, III-rats administered IMP at a dose of 10 mg/kg/day, and IV-rats with RA-induced OAB treated with IMP. IMP or vehicle were injected intraperitoneally for 14 days. The cystometry and assessment of bladder blood flow were performed two days after the last dose of IMP. The rats were then placed in metabolic cages for 24 h. Urothelial thickness measurements and biochemical analyses were performed. Intravesical infusion of RA induced OAB. Notably, intraperitoneal administration of imperatorin had no discernible effect on urinary bladder function and micturition cycles in normal rats. IMP attenuated the severity of RA-induced OAB. RA induced increases in urothelial ATP, calcitonin gene-related peptide (CGRP), organic cation transporter 3 (OCT3), and vesicular acetylcholine transporter (VAChT), as well as significant c-Fos expression in all micturition areas analyzed, which were attenuated by IMP. Furthermore, elevated levels of Rho kinase (ROCK1) and VAChT were observed in the detrusor, which were reversed by IMP in the context of RA-induced OAB in the urothelium, detrusor muscle, and urine. Imperatorin has a mitigating effect on detrusor overactivity. The mechanisms of action of IMP in the bladder appear to be diverse and complex. These findings suggest that IMP may provide protection against RA-induced OAB and could potentially develop into an innovative therapeutic strategy for the treatment of OAB.

Imperatorin Improves Obesity-Induced Cardiac Sympathetic Nerve Injury Mediated by P2X4 Receptor in Stellate Sympathetic Ganglion.

Obesity can activate the inflammatory signal pathway, induce in the body a state of chronic inflammation, and increase the excitability of the sympathetic nervous system, which may induce sympathetic neuropathic injury. The stellate sympathetic ganglia (SG) can express the P2X4 receptor, and the abnormal expression of the P2X4 receptor is related to inflammation. Imperatorin (IMP) is a kind of furan coumarin plant which has anti-inflammatory effects. This project aimed to investigate whether IMP can affect the expression of P2X4 receptors in the SG of obese rats to display a protective effect from high-fat-triggered cardiac sympathetic neuropathic injury. Molecular docking through homology modelling revealed that IMP had good affinity for the P2X4 receptor. Our results showed that compared with the normal group, the administration of IMP or P2X4 shRNA decreased sympathetic excitement; reduced the serum levels of triglyceride, total cholesterol, and lactate dehydrogenase; downregulated the expression of P2X4 receptors in SG; and inhibited the expression of inflammatory factors in the SG and serum of obese rats significantly. In addition, the expression of factors associated with the cell pyroptosis GSDMD, caspase-1, NLRP-3, and IL-18 in obese rats were significantly higher than those of the normal rats, and such effects were decreased after treatment with IMP or P2X4 shRNA. Furthermore, IMP significantly reduced the ATP-activated currents in HEK293 cells transfected with P2X4 receptor. Thus, the P2X4 receptor may be a key target for the treatment of obesity-induced cardiac sympathetic excitement. IMP can improve obesity-induced cardiac sympathetic excitement, and its mechanism of action may be related to the inhibition of P2X4 receptor expression and activity in the SG, suppression of cellular pyroptosis in the SG, and reduction of inflammatory factor levels.

Ameliorative Effect of Imperatorin on Dermatophagoides pteronyssinus-Induced Allergic Asthma by Suppressing the Th2 Response in Mice.

Imperatorin is a furanocoumarin derivative and an effective ingredient in several Chinese medicinal herbs. It has favorable expectorant, analgesic, and anti-inflammatory effects. In this study, we investigated whether imperatorin has protective effects against Dermatophagoides pteronyssinus (Der p)-induced asthma in mice. Lung and bronchial tissues were histopathologically examined through hematoxylin-eosin staining. The concentrations of immunoglobin E (IgE), IgG1, IgG2a in serum and those of T helper 1 (Th1) and two cytokines and eosinophil-activated chemokines in bronchoalveolar lavage fluid (BALF) were detected using an enzyme immunoassay. Histological examination revealed that imperatorin reduced inflammatory cell infiltration, mucus hypersecretion, and endothelial cell hyperplasia. The examination also indicated that imperatorin could reduce the inflammatory cell count in BALF as well as IgE and IgG1 expression in serum, but IgG2a expression was significantly increased. Imperatorin reduced the production of interleukin (IL)-4, IL-5, and IL-13 by Th2, promoted the production of interferon-γ and IL-12 by Th1, and increased the production of IL-10 in bronchoalveolar lavage fluid. These findings suggest that imperatorin has a considerable anti-inflammatory effect on Der p-induced allergic asthma in mice.

Imperatorin Influences Depressive-like Behaviors: A Preclinical Study on Behavioral and Neurochemical Sex Differences.

Imperatorin, a naturally derived furanocoumarin, exerts promising neuropharmacological properties. Therefore, it might be applicable in the treatment of brain diseases such as depression. In the present project, we aimed to investigate the sex-dependent effects of imperatorin (1, 5, and 10 mg/kg) on behavior and neurochemistry associated with antidepressant effects. The depressive-like behaviors of male and female Swiss mice were investigated in a forced swim test (FST). Subsequently, High-Performance Liquid Chromatography (HPLC) was used to evaluate the level of serotonin, its metabolite, 5-HIAA, and noradrenaline, in mouse brains. The study revealed that only males responded to imperatorin (1 and 5 mg/kg) treatment and caused an antidepressant effect, such as with respect to depressive-like behaviors, lowering immobility time and increasing immobility latency. The HPLC analysis demonstrated that serotonin levels in the prefrontal cortex of females decreased with the middle dose of imperatorin (5 mg/kg), while in the male prefrontal cortex, the lower dose (1 mg/kg) boosted serotonin levels. There were no evident changes observed with respect to noradrenaline and serotonin metabolite levels in the male hippocampus. To conclude, we propose that imperatorin has antidepressant potential, seemingly only in males, influencing brain serotonin level, but the direct mechanism of action requires further investigation.

Imperatorin Positively Regulates Melanogenesis through Signaling Pathways Involving PKA/CREB, ERK, AKT, and GSK3ß/ß-Catenin.

The present study investigated the melanogenic effects of imperatorin and isoimperatorin and the underlying mechanisms of imperatorin using a mouse melanoma B16F10 model. Interestingly, treatment with 25 µM of either imperatorin or isoimperatorin, despite their structural differences, did not produce differences in melanin content and intracellular tyrosinase activity. Imperatorin also activated the expression of melanogenic enzymes, such as tyrosinase (TYR) and tyrosinase-related proteins TYRP-1 and TYRP-2. Mechanistically, imperatorin increases melanin synthesis through the cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA)/cAMP-responsive element-binding protein (CREB)-dependent upregulation of microphthalmia-associated transcription factor (MITF), which is a key transcription factor in melanogenesis. Furthermore, imperatorin exerted melanogenic effects by downregulating extracellular signal-regulated kinase (ERK) and upregulating phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT)/glycogen synthesis kinase-3ß (GSK-3ß). Moreover, imperatorin increased the content of ß-catenin in the cell cytoplasm and nucleus by reducing the content of phosphorylated ß-catenin (p-ß-catenin). Finally, we tested the potential of imperatorin in topical application through primary human skin irritation tests. These tests were performed on the normal skin (upper back) of 31 volunteers to determine whether 25 or 50 µM of imperatorin had irritation or sensitization potential. During these tests, imperatorin did not induce any adverse reactions. Taken together, these findings suggest that the regulation of melanogenesis by imperatorin can be mediated by signaling pathways involving PKA/CREB, ERK, AKT, and GSK3ß/ß-catenin and that imperatorin could prevent the pathogenesis of pigmentation diseases when used as a topical agent.

Chemosensitizing effect and mechanism of imperatorin on the anti-tumor activity of doxorubicin in tumor cells and transplantation tumor model.

Multidrug resistance of tumors has been a severe obstacle to the success of cancer chemotherapy. The study wants to investigate the reversal effects of imperatorin (IMP) on doxorubicin (DOX) resistance in K562/DOX leukemia cells, A2780/Taxol cells and in NOD/SCID mice, to explore the possible molecular mechanisms. K562/DOX and A2780/Taxol cells were treated with various concentrations of DOX and Taol with or without different concentrations of IMP, respectively. K562/DOX xenograft model was used to assess anti-tumor effect of IMP combined with DOX. MTT assay, Rhodamine 123 efflux assay, RT-PCR, and Western blot analysis were determined in vivo and in vitro. Results showed that IMP significantly enhanced the cytotoxicity of DOX and Taxol toward corresponding resistance cells. In vivo results illustrated both the tumor volume and tumor weight were significantly decreased after 2-week treatment with IMP combined with DOX compared to the DOX alone group. Western blotting and RT-PCR analyses indicated that IMP downregulated the expression of P-gp in K562/DOX xenograft tumors in NOD/SCID mice. We also evaluated glycolysis and glutamine metabolism in K562/DOX cells by measuring glucose consumption and lactate production. The results revealed that IMP could significantly reduce the glucose consumption and lactate production of K562/DOX cells. Furthermore, IMP could also remarkably repress the glutamine consumption, α-KG and ATP production of K562/DOX cells. Thus, IMP may sensitize K562/DOX cells to DOX and enhance the anti-tumor effect of DOX in K562/DOX xenograft tumors in NOD/SCID mice. IMP may be an adjuvant therapy to mitigate the multidrug resistance in leukemia chemotherapy.

[Identification of metabolites of imperatorin in rats: based on UHPLC-Q-Exactive Orbitrap MS].

This study aims to identify and analyze the metabolites of imperatorin in rats by UHPLC-Q-Exactive Orbitrap MS. Specifically, after rats were treated(ig) with imperatorin, the plasma, urine, and feces were collected, and the samples were processed by solid phase extraction. Then, UHPLC-Q-Exactive Orbitrap MS was performed. In MS, 0.1% formic acid water(A)-acetonitrile(B) was applied as mobile phase for gradient elution and the data of MS in both positive and negative ion modes were collected. The metabolites of imperatorin in blood, urine, and feces of rats were analyzed to explore the metabolic pathways of imperatorin in rats. According to accurate molecular weight, multistage MS data, MS fragmentation rule of the standard substance, and previous reports, a total of 51 metabolites were identified, with 35, 40, and 16 from plasma, urine, and feces, separately. The main metabolic pathways were oxidization, glucuronidation, isopentenyl removal, sulphation, carboxylation, among others. The conclusion in this study is expected to serve as a reference for the further development and the further pharmacodynamics study of imperatorin.

Imperatorin induces autophagy and G0/G1 phase arrest via PTEN-PI3K-AKT-mTOR/p21 signaling pathway in human osteosarcoma cells in vitro and in vivo.

BACKGROUND: Osteosarcoma is the third most common cancer in adolescence and the first common primary malignant tumor of bone. The long-term prognosis of osteosarcoma still remains unsatisfactory in the past decades. Therefore, development of novel therapeutic agents which are effective to osteosarcoma and are safe to normal tissue simultaneously is quite essential and urgent. METHODS: Firstly, MTT assay, cell colony formation assay, cell migration and invasion assays were conducted to evaluate the inhibitory effects of imperatorin towards human osteosarcoma cells. RNA-sequence assay and bioinformatic analysis were then performed to filtrate and assume the potential imperatorin-induced cell death route and signaling pathway. Moreover, quantitative real-time PCR assay, western blot assay and rescue experiments were conducted to confirm the assumptions of bioinformatic analysis. Finally, a subcutaneous tumor-transplanted nude mouse model was established and applied to evaluate the internal effect of imperatorin on osteosarcoma by HE and immunohistochemistry staining. RESULTS: Imperatorin triggered time-dependent and dose-dependent inhibition of tumor growth mainly by inducing autophagy promotion and G0/G1 phase arrest in vitro and in vivo. Besides, imperatorin treatment elevated the expression level of PTEN and p21, down-regulated the phosphorylation of AKT and mTOR. In contrast, the inhibition of PTEN using Bpv (HOpic), a potential and selective inhibitor of PTEN, concurrently rescued imperatorin-induced autophagy promotion, cell cycle arrest and inactivation of PTEN-PI3K-AKT-mTOR/p21 pathway. CONCLUSIONS: This work firstly revealed that imperatorin induced autophagy and cell cycle arrest through PTEN-PI3K-AKT-mTOR/p21 signaling pathway by targeting and up-regulating PTEN in human osteosarcoma cells. Hence, imperatorin is a desirable candidate for clinical treatments of osteosarcoma.

Imperatorin alleviates cancer cachexia and prevents muscle wasting via directly inhibiting STAT3.

Skeletal muscle wasting is the most remarkable phenotypic feature of cancer cachexia that increases the risk of morbidity and mortality. Imperatorin (IMP), a main bioactive component of Angelica dahurica Radix, has been reported to possess several pharmacological effects including potential anti-colitis, anti-arthritis and anti-tumor activities. In this work, we demonstrated that IMP is a promising agent for the treatment of muscle wasting in cancer cachexia. IMP (5-20 µM) dose-dependently attenuated TCM-induced C2C12 myotube atrophy and prevented the induction of E3 ubiquitin ligases muscle RING-finger containing protein-1 (MuRF1) and muscle atrophy Fbox protein (Atrogin-1/MAFbx). Moreove, IMP administration significantly improved chief features of cancer cachexia in vivo, with significant prevention of the loss of body weight and deleterious wasting of multiple tissues, including skeletal muscle, fat and kidney and decreased expression of MuRF1 and Atrogin-1 in cachectic muscles. Cellular signaling pathway analysis showed that IMP selectively inhibited the phosphorylation of signal transducer and activator of transcription 3 (STAT3) in vitro and in vivo, and surface plasmon resonance (SPR) affinity experiments further demonstrated IMP bound to STAT3 in a concentration-dependent resonance manner. Molecular docking results revealed that IMP binds to the SH2 domain of STAT3, forming a hydrogen bond interaction with Arg-609, and a Sigma-Pi interaction with Lys-591. Mechanism analysis demonstrated that STAT3 overexpression markedly weakens the improvements of IMP on myotube atrophy and muscle wasting of cancer cachexia, indicating that STAT3 mediated the therapeutic effect of IMP. All these favorable results indicated that IMP is a new potential therapeutic candidate for cancer cachexia.

Imperatorin alleviates ROS-mediated airway remodeling by targeting the Nrf2/HO-1 signaling pathway.

In this study, we investigated the role and mechanism of imperatorin (IMP) in chronic inflammation and airway remodeling. The levels of TNF-α, IL-1ß, IL-6, IL-8, VEGF, α-SMA, and ROS were detected by ELISA, immunohistochemistry (IHC), immunofluorescence, and Western blot. In addition, we evaluated the effect of IMP on MAPK, PI3K/Akt, NF-κB, and Nrf2/HO-1 signaling pathways. IMP treatment obviously attenuated the production of inflammatory cytokines and inflammatory cells in bronchoalveolar lavage fluid of OVA-induced airway remodeling model. Meanwhile, it significantly inhibited inflammatory cell infiltration, goblet cell hyperplasia, collagen deposition, VEGF production, α-SMA, and ROS expression. Our study has shown that IMP could regulate the signaling pathways including MAPK, PI3K/Akt, NF-κB, and Nrf2/HO-1 to release the inflammatory responses. IMP might attenuate airway remodeling by the down-regulation of Nrf2/HO-1/ROS/PI3K/Akt, Nrf2/HO-1/ROS/MAPK, and Nrf2/HO-1/ROS/NF-κB signaling pathways.

Imperatorin ameliorates learning and memory deficits through BDNF/TrkB and ERK/CaMKIIα/CREB signaling in prenatally-stressed female offspring.

Prenatal stress (PS) can lead to impaired spatial learning and memory in offspring. Imperatorin (IMP) is a naturally occurring furanocoumarin with many pharmacological properties. However, the effects of IMP on cognitive impairment induced by PS and the underlying molecular mechanisms remain unclear. We investigated the protective effect of IMP treatment after PS on learning and memory deficits in female offspring at postnatal 60 days. After treating prenatally-stressed offspring with IMP (15 and 30 mg/kg) for 28 days, we found that IMP increased body weight and ameliorated spatial learning and memory and working memory deficits in female offspring rats. Meanwhile, hippocampal Glu and serum corticosterone levels in prenatally-stressed offspring were significantly decreased after IMP administration. Additionally, IMP treatment significantly increased BDNF, TrkB, CaMKII, and CREB mRNA expression in the hippocampus of offspring rats. Furthermore, PS-mediated induction of RKIP protein and mRNA expression and glucocorticoid receptor protein expression in the hippocampus of offspring rats were significantly decreased by IMP treatment, and the protein expression of BDNF and TrkB and relative levels of p-EKR/ERK, p-CaMKIIα/CaMKIIα, and p-CREB/CREB were remarkably increased after IMP treatment. Taken together, IMP can ameliorate PS-induced learning and memory deficits through BDNF/TrkB and ERK/CaMKIIα/CREB signaling pathway and hypothalamic-pituitary-adrenal axis.

Anti-nociceptive and anti-inflammatory effect of imperatorin: evidences for involvement of COX-2, iNOS, NFκB and inflammatory cytokines.

Aim: The objective of the current investigation was to explore the analgesic effect of naturally occurring furanocoumarin, imperatorin and the involvement of inducible cyclooxygenase (COX-2), inducible nitric oxide synthase (iNOS), NFκB and cytokines in the observed effect.Materials and methods: Anti-nociceptive effect was explored by inducing chemical hyperalgesia using acetic acid and formalin in mice. ED50 of imperatorin was calculated in acetic acid model. Modulation of cyclooxygenase and nitric oxide pathway by imperatorin was examined by stimulator/precursor challenge with substance P and L-arginine, respectively and quantification of COX-2, iNOS and NFκB expression by immunohistochemical analysis in spinal tissues. Involvement of inflammatory cytokines TNF-α and IL-1ß was investigated using LPS challenge and subsequent ELISA analysis of these inflammatory mediators in serum. Carrageenan inflicted paw edema was employed to explore the anti-inflammatory activity of imperatorin.Results: A significant reduction in the nociceptive behaviour was observed with imperatorin treatment in acetic acid and formalin test. ED50 of imperatorin was found to be 4.53 mg/kg. Pre-treatment with substance P and L-arginine significantly attenuated the anti-nociceptive activity of imperatorin in formalin test. Immunohistochemical findings revealed marked decrease in spinal COX-2, iNOS and NFκB expression. Imperatorin administration significantly reduced LPS induced rise in level of TNF-α and IL-1ß dose dependently. In carrageenan-induced paw edema test, maximum possible anti-inflammatory effect of imperatorin was evident after 240 min of carrageenan administration.Conclusion: Current investigation revealed that anti-nociceptive and anti-inflammatory potential of imperatorin is probably mediated through the attenuation of COX-2, iNOS, NFκB activity and reduction in circulatory cytokines.

Imperatorin as a Promising Chemotherapeutic Agent Against Human Larynx Cancer and Rhabdomyosarcoma Cells.

Naturally occurring coumarins are bioactive compounds widely used in Asian traditional medicine. They have been shown to inhibit proliferation, induce apoptosis, and/or enhance the cytotoxicity of currently used drugs against a variety of cancer cell types. The aim of our study was to examine the antiproliferative activity of different linear furanocoumarins on human rhabdomyosarcoma, lung, and larynx cancer cell lines, and dissolve their cellular mechanism of action. The coumarins were isolated from fruits of Angelica archangelica L. or Pastinaca sativa L., and separated using high-performance counter-current chromatography (HPCCC). The identity and purity of isolated compounds were confirmed by HPLC-DAD and NMR analyses. Cell viability and toxicity assessments were performed by means of methylthiazolyldiphenyl-tetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays, respectively. Induction of apoptosis and cell cycle progression were measured using flow cytometry analysis. qPCR method was applied to detect changes in gene expression. Linear furanocoumarins in a dose-dependent manner inhibited proliferation of cancer cells with diverse activity regarding compounds and cancer cell type specificity. Imperatorin (IMP) exhibited the most potent growth inhibitory effects against human rhabdomyosarcoma and larynx cancer cell lines owing to inhibition of the cell cycle progression connected with specific changes in gene expression, including CDKN1A. As there are no specific chemotherapy treatments dedicated to laryngeal squamous cell carcinoma and rhabdomyosarcoma, and IMP seems to be non-toxic for normal cells, our results could open a new direction in the search for effective anti-cancer agents.

Design, synthesis, and evaluation of new series of Imperatorin analogs with potential vasodilatory activity.

Two series of imperatorin analogs were synthesized based on our previous research and evaluated for their vasodilatation activities on in vitro rat mesenteric artery, basilar artery, and renal artery ring models. Target compounds were characterized by infrared, 1H NMR, and mass spectra. Most derivatives possessed significant vasodilatory activity on the mesenteric artery, and compound 3a exhibited favorable and broad vasodilatation activities on three kinds of rat artery ring models. The pharmacological results indicated that introducing nitrogen-contained ring in side chain or large steric hindrance at the distal end could increase the vasodilatory activity. Further, replacement of oxygen atom (-O-) in the skeleton of furocoumarin derivatives with nitrogen (-NH-) could cause the decrease of vasodilatory activity. The molecular docking also indicated that compound 3a showed a best affinity with α-1C receptor (PDB ID: 3G43). All these results suggested compound 3a would be a potential vasodilatory agent for hypertension.

Furocoumarin Content of Fennel-Below the Safety Threshold.

Furocoumarins are known for their phototoxic and potential carcinogenic effects. These types of compounds have previously been reported from fennel (Foeniculum vulgare Mill.), a widely used medicinal plant and spice; however, no reliable quantitative data are available on the occurrence of these compounds in fennel fruits. For the first time, we report a comprehensive analysis of fennel fruit samples of different origins, representing a wide range of accessions for their furocoumarin content. Psoralene, 5-methoxypsoralene (bergapten), and imperatorin contents of 33 fennel samples were analyzed using a sensitive liquid chromatography-mass spectrometry (LC-MS) method. When applied at the highest therapeutic dose described in the monograph issued by the European Medicines Agency, the furocoumarin content of the fruits ranged up to 1.22 µg/d, which is below the most restrictive recommendations. Based on our findings, fennel consumption can be considered as safe, at least based on its low furocoumarin content.