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Dehydroepiandrosterone (DHEA) is frequently integrated as an adjuvant in over a quarter of controlled ovarian hyperstimulation (COH) protocols, despite the ongoing debate regarding its impact. This study aimed to evaluate the efficacy and mechanism of action of DHEA on ovarian follicular development and ovarian response in rats with varying ovarian reserves. The study involved 75 rats categorized into 15 distinct groups. The ovarian tissues of rats in both the normal ovarian reserve group and the premature ovarian insufficiency (POI) group, induced by 4-vinylcyclohexene diepoxide (VCD) injection, were subjected to histomorphological and biochemical analyses following the administration of DHEA, either alone or in combination with COH. Follicle counting was performed on histological sections obtained from various tissues. Serum concentrations of anti-Müllerian hormone (AMH) and the quantification of specific proteins in ovarian tissue, including phosphatase and tensin homolog of chromosome 10 (PTEN), phosphoinositide 3-kinase (PI3K), phosphorylated protein kinase B (pAKT), cyclooxygenase 2 (COX-2), caspase-3, as well as assessments of total antioxidant status and total oxidant status, were conducted employing the ELISA method. The impact of DHEA exhibited variability based on ovarian reserve. In the POI model, DHEA augmented follicular development and ovarian response to the COH protocol by upregulating the PTEN/PI3K/AKT signaling pathway, mitigating apoptosis, inflammation, and oxidative stress, contrary to its effects in the normal ovarian reserve group. In conclusion, it has been determined that DHEA may exert beneficial effects on ovarian stimulation response by enhancing the initiation of primordial follicles and supporting antral follicle populations.

Magnoliae Flos (MF) is a medicinal herb widely employed in traditional medicine for relieving sinusitis, allergic rhinitis, headaches, and toothaches. Here, we investigated the potential preventive effects of MF extract (MFE) against 4-vinylcyclohexene diepoxide (VCD)-induced ovotoxicity in ovarian cells and a mouse model of premature ovarian insufficiency (POI). The cytoprotective effects of MFE were assessed using CHO-K1 or COV434 cells. In vivo, B6C3F1 female mice were intraperitoneally injected with VCD for two weeks to induce POI, while MFE was orally administered for four weeks, beginning one week before VCD administration. VCD led to a significant decline in the viabilities of CHO-K1 and COV434 cells and triggered excessive reactive oxygen species (ROS) production and apoptosis specifically in CHO-K1 cells. However, pretreatment with MFE effectively prevented VCD-induced cell death and ROS generation, while also activating the Akt signaling pathway. In vivo, MFE increased relative ovary weights, follicle numbers, and serum estradiol and anti-Müllerian hormone levels versus controls under conditions of ovary failure. Collectively, our results demonstrate that MFE has a preventive effect on VCD-induced ovotoxicity through Akt activation. These results suggest that MFE may have the potential to prevent and manage conditions such as POI and diminished ovarian reserve.

Risk of cardiovascular disease mortality rises in women after menopause. While increased cardiovascular risk is largely attributed to postmenopausal declines in estrogens, the molecular changes in the heart that contribute to risk are poorly understood. Disruptions in intracellular calcium handling develop in ovariectomized mice and have been implicated in cardiac dysfunction. Using a mouse model of menopause in which ovarian failure occurs over 120 days, we sought to determine if perimenopause impacted calcium removal mechanisms in the heart and identify the molecular mechanisms. Mice were injected with 4-vinylcyclohexene diepoxide (VCD) to induce ovarian failure over 120 days, mimicking perimenopause. Hearts were removed at 60 and 120 days after VCD injections, representing the middle and end of perimenopause. SERCA2a function was significantly diminished at the end of perimenopause. Neither SERCA2a nor phospholamban expression changed at either time point, but phospholamban phosphorylation at S16 and T17 was dynamically altered. Intrinsic SERCA inhibitors sarcolipin and myoregulin increased >4-fold at day 60, as did the native activator DWORF. At the end of perimenopause, sarcolipin and myoregulin returned to baseline levels while DWORF was significantly reduced below controls. Sodium-calcium exchanger expression was significantly increased at the end of perimenopause. These results show that the foundation for increased cardiovascular disease mortality develops in the heart during perimenopause and that regulators of calcium handling exhibit significant fluctuations over time. Understanding the temporal development of cardiovascular risk associated with menopause and the underlying mechanisms is critical to developing interventions that mitigate the rise in cardiovascular mortality that arises after menopause.

Background: Cancer is a fatal disease that severely affects humans. Designing new anticancer strategies and understanding the mechanism of action of anticancer agents is imperative., Hypothesis/purpose: In this study, we evaluated the utility of metformin and D-limonene, alone or in combination, as potential anticancer therapeutics using the human liver and breast cancer cell lines HepG2 and MCF-7., Study Design: An integrated systems pharmacology approach is presented for illustrating the molecular interactions between metformin and D-limonene., Methods: We applied a systems-based analysis to introduce a drug-target-pathway network that clarifies different mechanisms of treatment. The combination treatment of metformin and D-limonene induced apoptosis in both cell lines compared with single drug treatments, as indicated by flow cytometric and gene expression analysis., Results: The mRNA expression of Bax and P53 genes were significantly upregulated while Bcl-2, iNOS, and Cox-2 were significantly downregulated in all treatment groups compared with normal cells. The percentages of late apoptotic HepG2 and MCF-7 cells were higher in all treatment groups, particularly in the combination treatment group. Calculations for the combination index (CI) revealed a synergistic effect between both drugs for HepG2 cells (CI = 0.14) and MCF-7 cells (CI = 0.22)., Conclusion: Our data show that metformin, D-limonene, and their combinations exerted significant antitumor effects on the cancer cell lines by inducing apoptosis and modulating the expression of apoptotic genes., (© 2024. The Author(s).)

The abrupt cessation of ovarian hormone release is associated with declines in muscle contractile function, yet the impact of gradual ovarian failure on muscle contractility across peri-, early- and late-stage menopause remains unclear. In this study, a 4-vinylcyclohexene diepoxide (VCD)-induced ovarian failure mouse model was used to examine time course changes in muscle mechanical function. Plantar flexors of female mice (VCD: n = 10; CON: n = 8) were assessed at 40 (early perimenopause), 80 (late perimenopause), 120 (menopause onset) and 176 (late menopause) days post-initial VCD injection. A torque-frequency relationship was established across a range of frequencies (10-200 Hz). Isotonic dynamic contractions were elicited against relative loads (10-80% maximal isometric torque) to determine the torque-velocity-power relationship. Mice then performed a fatigue task using intermittent 100 Hz isometric contractions until torque dropped by 60%. Recovery of twitch, 10 Hz and 100 Hz torque were tracked for 10 min post-task failure. Additionally, intact muscle fibres from the flexor digitorum brevis underwent a fatigue task (50 repetitions at 70 Hz), and 10 and 100 Hz tetanic [Ca 2+ ] were monitored for 10 min afterward. VCD mice exhibited 16% lower twitch torque than controls across all time points. Apart from twitch torque, 10 Hz torque and 10 Hz tetanic [Ca 2+ ], where VCD showed greater values relative to pre-fatigue during recovery, no significant differences were observed between control and VCD mice during recovery. These results indicate that gradual ovarian failure has minimal detriments to in vivo muscle mechanical function, with minor alterations observed primarily for low-frequency stimulation during recovery from fatigue., (© 2024 The Authors. Experimental Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Background: Sarcoptic mange is a serious animal welfare concern in bare-nosed wombats (Vombatus ursinus). Fluralaner (Bravecto ® ) is a novel acaricide that has recently been utilised for treating mange in wombats. The topical 'spot-on' formulation of fluralaner can limit treatment delivery options in situ, but dilution to a volume for 'pour-on' delivery is one practicable solution. This study investigated the in vitro acaricidal activity of Bravecto, a proposed essential oil-based diluent (Orange Power ® ), and two of its active constituents, limonene and citral, against Sarcoptes scabiei., Methods: Sarcoptes scabiei were sourced from experimentally infested pigs. In vitro assays were performed to determine the lethal concentration (LC 50 ) and survival time of the mites when exposed to varying concentrations of the test solutions., Results: All compounds were highly effective at killing mites in vitro. The LC 50 values of Bravecto, Orange Power, limonene and citral at 1 h were 14.61 mg/ml, 4.50%, 26.53% and 0.76%, respectively. The median survival times of mites exposed to undiluted Bravecto, Orange Power and their combination were 15, 5 and 10 min, respectively. A pilot survival assay of mites collected from a mange-affected wombat showed survival times of < 10 min when exposed to Bravecto and Orange Power and 20 min when exposed to moxidectin., Conclusions: These results confirm the acaricidal properties of Bravecto, demonstrate acaricidal properties of Orange Power and support the potential suitability of Orange Power and its active constituents as a diluent for Bravecto. As well as killing mites via direct exposure, Orange Power could potentially enhance the topical delivery of Bravecto to wombats by increasing drug penetration in hyperkeratotic crusts. Further research evaluating the physiochemical properties and modes of action of Orange Power and its constituents as a formulation vehicle would be of value., (© 2024. The Author(s).)

Herein, (-)-galiellalactone 1 congeners responsible for the nuclear factor erythroid 2-related factor 2 (Nrf2)-activating neuroprotective effects were elucidated. Additionally, novel congener-based Nrf2 activators were identified using a drug repositioning strategy. (-)-Galiellalactone, which comprises a tricyclic lactone skeleton, significantly activates antioxidant response element (ARE)-mediated transcription in neuroblastoma SH-SY5Y cells. Interestingly, two cyclohexene-truncated [3.3] bicyclic lactone analogs, which possess an exocyclic α-methylene-γ-butyrolactone moiety, exhibited higher Nrf2/ARE transcriptional activities than the parent (-)-galiellalactone. We confirmed that the cyclohexene moiety embedding the [3.3] bicyclic lactone congener does not play the essential role of (-)-galiellalactone for Nrf2/ARE activation. Nrf2/ARE activation by novel analogs resulted in the upregulation of downstream antioxidative and phase II detoxifying enzymes, heme oxygenase-1, and NAD(P)H quinone oxidoreductase 1, which are closely related to the cytoprotective effects on neurodegenerative diseases. (-)-Galiellalactone and its [3.3] bicyclic variants 3l and 3p increased the expression of antioxidant genes and exhibited neuroprotective effects against 6-hydroxydopamine-mediated neurotoxicity in the neuroblastoma SH-SY5Y cell line., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Objective: 2-Dodecyl-6-Methoxycyclohexa-2, 5-Diene-1, 4-Dione (DMDD) isolated from Averrhoa carambola L. root, has been proven therapeutic effects on diabetic kidney disease (DKD). This research aims to assess DMDD's effects on DKD and to investigate its underlying mechanisms, to establish DMDD as a novel pharmaceutical agent for DKD treatment., Methods: The human renal tubular epithelial (HK-2) cells were induced by high glucose (HG) to mimic DKD and followed by DMDD treatment. The cytotoxicity of DMDD was assessed using the Cell Counting Kit-8 (CCK-8) assay. The migratory capacity of HK-2 cells was evaluated through transwell and scratch-wound assays. To investigate the effect of Smad7 and miR-21-5p, lentiviral transfection was employed in HK-2 cells. Additionally, the expression of proteins related to epithelial-mesenchymal transition (EMT) and TGFβ1/Smad2/3 pathway was checked by QRT-PCR, Western blot, and immunofluorescence techniques., Results: This study has shown that DMDD significantly suppresses cell migration and the expression of Vimentin, α-SMA, TGFβ1, and p-Smad2/3 in HK-2 cells under HG conditions. Concurrently, DMDD enhances the protein expression of E-cadherin and Smad7. Intriguingly, the therapeutic effect of DMDD was abrogated upon Smad7 silencing. Further investigations revealed that DMDD effectively inhibits miR-21-5p expression, which is upregulated by HG. Downregulation of miR-21-5p inhibits the activation of the TGFβ1/Smad2/3 pathway and EMT induced by HG. In contrast, overexpression of miR-21-5p negates DMDD's therapeutic benefits., Conclusion: DMDD mitigates EMT in HG-induced HK-2 cells by modulating the miR-21-5p/Smad7 pathway, thereby inhibiting renal fibrosis in DKD. These findings suggest that DMDD holds promise as a potential therapeutic agent for DKD., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Background/aim: Ischemia-reperfusion (IR) injury to a part of the body can cause damage to distant organs such as the kidney and heart. This study investigated the protective effects of safranal against IR-induced renal injury., Materials and Methods: Used in this study were 24 Wistar Albino male rats, which were divided into 3 equal and randomised groups. The sham group underwent laparotomy only. In the IR group, the infrarenal aorta was clamped for 1 h, and then reperfused for 2 h. In the IR-safranal group, safranal was administered 30 min before the procedure and IR injury was induced in the same way as in the IR group. After the procedure, blood and tissue samples were collected from the rats for biochemical and histopathological analyses. Antioxidant capacity and proinflammatory cytokine analyses were performed on the blood samples. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining was performed to determine the number of cells undergoing apoptosis in the kidney tissue., Results: The estimated glomerular filtration rate, an indicator of renal function, was lower in the IR group (p1 = 0.024 vs. p3 = 0.041, respectively) compared to the other groups, while creatinine levels were higher in the IR group compared to the other groups (p1 = 0.032 vs. p2 = 0.044, respectively). The blood urea nitrogen level was higher in the IR group than in the other groups (p1 = 0.001vs p2 = 0.035, respectively). The total antioxidant and total oxidant status, indicating tissue oxidative stress, did not differ between groups (p = 0.914 vs. p = 0.184, respectively). Among the proinflammatory cytokines, the interleukin-1β (IL-1β) and IL-6 levels were significantly higher in the IR group (p = 0.034 vs. p = 0.001, respectively), but the tumour necrosis factor-α (p = 0.19), and interferon-γ (p = 0.311) levels did not differ between groups. Histopathological examination showed significantly less damage to glomerular and tubular cells in the IR-safranal group (p < 0.001). The number of TUNEL-positive cells was higher in the IR group compared to the other groups (p < 0.001)., Conclusion: Safranal may have protective effects against kidney damage caused by distant ischemia-reperfusion injury., Competing Interests: Conflict of interest: The authors declare that they have no proprietary or commercial interest in any of the products mentioned or concepts discussed within the article. There are no conflicts of interest to declare., (© TÜBİTAK.)

Dyslipidemia is a lipid metabolism disorder associated with the loss of the physiological homeostasis that ensures safe levels of lipids in the organism. This metabolic disorder can trigger pathological conditions such as atherosclerosis and cardiovascular diseases. In this regard, statins currently represent the main pharmacological therapy, but their contraindications and side effects limit their use. This is stimulating the search for new therapeutic strategies. In this work, we investigated in HepG2 cells the hypolipidemic potential of a picrocrocin-enriched fraction, analyzed by high-resolution 1 H NMR and obtained from a saffron extract, the stigmas of Crocus sativus L., a precious spice that has already displayed interesting biological properties. Spectrophotometric assays, as well as expression level of the main enzymes involved in lipid metabolism, have highlighted the interesting hypolipidemic effects of this natural compound; they seem to be exerted through a non-statin-like mechanism. Overall, this work provides new insights into the metabolic effects of picrocrocin, thus confirming the biological potential of saffron and paving the way for in vivo studies that could validate this spice or its phytocomplexes as useful adjuvants in balancing blood lipid homeostasis.

D-limonene (4-isopropenyl-1-methylcyclohexene) is an important compound in several citrus essential oils (such as orange, lemon, tangerine, lime, and grapefruit). It has been used as a flavoring agent and as a food preservative agent, with generally recognized as safe (GRAS) status. D-limonene has been well-studied for its anti-inflammatory, antioxidant, anti-cancer, and antibacterial properties. The antibacterial activity of D-limonene against food-borne pathogens was investigated in this study by preparing a D-limonene nanoemulsion. The D-limonene solution and nanoemulsion have been prepared in six concentrations, 0.04%, 0.08%, 0.1%, 0.2%, 0.4%, and 0.8% ( v / v ), respectively, and the antibacterial activity was tested against four food-borne pathogens ( Staphylococcus aureus , Listeria monocytogenes , Salmonella enterica , and Escherichia coli ). The results showed that the D-limonene nanoemulsion had good nanoscale and overall particle size uniformity, and its particle size was about 3~5 nm. It has been found that the D-limonene solution and nanoemulsion have a minimal inhibitory concentration of 0.336 mg/mL, and that they could inhibit the growth of microorganisms efficiently. The data indicate that the D-limonene nanoemulsion has more antibacterial ability against microorganisms than the D-limonene essential oil. After bananas are treated with 1.0% and 1.5% D-limonene nanoemulsion coatings, the water loss of the bananas during storage and the percentage of weight loss are reduced, which can inhibit the activity of pectinase. The application of a biocoating provides a good degree of antibacterial activity and air and moisture barrier properties, which help with extending the shelf life of bananas.

Mitochondrial oxidative phosphorylation (OXPHOS) has become an attractive target in anti-cancer studies in recent years. In this study, we found that a small molecule phenylbutenoid dimer NMac1 (Nm23-H1 activator 1), (±)-trans-3-(3,4-dimethoxyphenyl)-4-[(E)-3,4-dimethoxystyryl]cyclohex-1-ene, a previously identified anti-metastatic agent, has novel anti-proliferative effect only under glucose starvation in metastatic breast cancer cells. NMac1 causes significant activation of AMPK by decreasing ATP synthesis, lowers mitochondrial membrane potential (MMP, ΔΨm), and inhibits oxygen consumption rate (OCR) under glucose starvation. These effects of NMac1 are provoked by a consequence of OXPHOS complex I inhibition. Through the structure-activity relationship (SAR) study of NMac1 derivatives, NMac24 was identified as the most effective compound in anti-proliferation. NMac1 and NMac24 effectively suppress cancer cell proliferation in 3D-spheroid in vivo-like models only under glucose starvation. These results suggest that NMac1 and NMac24 have the potential as anti-cancer agents having cytotoxic effects selectively in glucose restricted cells., (© 2021. The Author(s).)

Hops contain flavonoids that have sedative and sleep-promoting activities such as α-acid, β-acid, and xanthohumol. In this study, the sleep-enhancing activity of a Saaz-Saphir hops mixture was measured. In the caffeine-induced insomnia model, the administration of a Saaz-Saphir mixture increased the sleep time compared to Saaz or Saphir administration alone, which was attributed to the increase in NREM sleep time by the δ-wave increase. Oral administration of the Saaz-Saphir mixture for 3 weeks increased the γ-amino butyric acid (GABA) content in the brain and increased the expression of the GABA A receptor. As the GABA antagonists picrotoxin and bicuculline showed a decrease in sleep activity, it was confirmed that the GABA A receptor was involved in the Saaz-Saphir mixture activity. In addition, the GABA A receptor antagonist also reduced the sleep activity induced by xanthohumol and humulone contained in the Saaz-Saphir mixture. Therefore, xanthohumol and humulone contained in the Saaz-Saphir mixture showed sleep-promoting activity mediated by the GABA A receptors. The mixture of the Saaz and Saphir hop varieties may thus help mitigate sleep disturbances compared to other hop varieties.

A secondary metabolites investigation on Biscogniauxia sp. 71-10-1-1 was carried out, which led to the obtention of nine new diisoprenyl-cyclohexene/ane-type meroterpenoids ( 1 - 9 ) and two new isoprenylbenzoic acid-type meroterpeniods ( 10 - 11 ). The structures of these isolates were established on the basis of multispectroscopic analyses, ECD, and 13 C chemical shifts calculations, and single-crystal X-ray diffraction. Among them, biscognin A ( 1 ) is the first diisoprenyl-cyclohexene-type meroterpenoid with a unique 2-isopropyl-6'-methyloctahydro-1' H -spiro[cyclopropane-1,2'-naphthalene] skeleton. Biscognienyne F ( 5 ) is the first diisoprenyl-cyclohexene-type meroterpenoid with a cyclic carbonate. The anti-inflammatory assays of the majority of compounds were evaluated, which exhibited that compounds 3 and 5 can obviously inhibit pro-inflammatory cytokines TNF-α and IL-6 productions. This is the first report for diisoprenyl-cyclohexene-type meroterpenoids with anti-inflammatory activity. Moreover, the possible biogenetic pathways of the majority of compounds ( 1 - 5 ) are proposed.

Glycoside hydrolases (GH) are a large family of hydrolytic enzymes found in all domains of life. As such, they control a plethora of normal and pathogenic biological functions. Thus, understanding selective inhibition of GH enzymes at the atomic level can lead to the identification of new classes of therapeutics. In these studies, we identified a 4-⍺-glucoside of valienamine (8) as an inhibitor of Streptomyces coelicolor (Sco) GlgE1-V279S which belongs to the GH13 Carbohydrate Active EnZyme family. The results obtained from the dose-response experiments show that 8 at a concentration of 1000 µM reduced the enzyme activity of Sco GlgE1-V279S by 65%. The synthetic route to 8 and a closely related 4-⍺-glucoside of validamine (7) was achieved starting from readily available D-maltose. A key step in the synthesis was a chelation-controlled addition of vinylmagnesium bromide to a maltose-derived enone intermediate. X-ray structures of both 7 and 8 in complex with Sco GlgE1-V279S were solved to resolutions of 1.75 and 1.83 Å, respectively. Structural analysis revealed the valienamine derivative 8 binds the enzyme in an E 2 conformation for the cyclohexene fragment. Also, the cyclohexene fragment shows a new hydrogen-bonding contact from the pseudo-diaxial C(3)-OH to the catalytic nucleophile Asp 394 at the enzyme active site. Asp 394, in fact, forms a bidentate interaction with both the C(3)-OH and C(7)-OH of the inhibitor. In contrast, compound 7 disrupts the catalytic sidechain interaction network of Sco GlgE1-V279S via steric interactions resulting in a conformation change in Asp 394. These findings will have implications for the design other aminocarbasugar-based GH13-inhibitors and will be useful for identifying more potent and selective inhibitors.

Introduction: Asthma is a chronic and recurring airway disease, which related to mast cell activation. Many compounds derived from Chinese herbal medicine has promising effects on stabilizing mast cells and decreasing inflammatory mediator production. Safranal, one of the active compounds from Crocus sativus , shows many anti-inflammatory properties. In this study, we evaluated the effect of safranal in ovalbumin (OVA)-induced asthma model. Furthermore, we investigate the effectiveness of safranal on stabilizing mast cell and inhibiting the production of inflammatory mediators in passive systemic anaphylaxis (PSA) model., Methods: OVA-induced asthma and PSA model were used to evaluate the effect of safranal in vivo. Lung tissues were collected for H&E, TB, IHC, and PAS staining. ELISA were used to determine level of IgE and chemokines (IL-4, IL-5, TNF-α, and IFN-γ). RNA sequencing was used to uncovers genes that safranal regulate. Bone marrow-derived mast cells (BMMCs) were used to investigate the inhibitory effect and mechanism of safranal. Cytokine production (IL-6, TNF-α, and LTC 4 ) and NF-κB and MAPKs signaling pathway were assessed., Results: Safranal reduced the level of serum IgE, the number of mast cells in lung tissue were decreased and Th1/Th2 cytokine levels were normalized in OVA-induced asthma model. Furthermore, safranal inhibited BMMCs degranulation and inhibited the production of LTC 4 , IL-6, and TNF-α. Safranal inhibits NF-κB and MAPKs pathway protein phosphorylation and decreases NF-κB p65, AP-1 nuclear translocation. In the PSA model, safranal reduced the levels of histamine and LTC 4 in serum., Conclusions: Safranal alleviates OVA-induced asthma, inhibits mast cell activation and PSA reaction. The possible mechanism occurs through the inhibition of the MAPKs and NF-κB pathways., Competing Interests: Authors XZ and WP were employed by company Shanghai Traditional Chinese Medicine Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Lertnimitphun, Zhang, Fu, Yang, Zheng, Yuan, Zhou, Zhang, Pei, Lu and Xu.)

A series of mono- and di-methylenecyclohexenone derivatives, 3a-f and 4a-f, respectively, were designed and synthesized from piperlongumine (PL) and their in vitro and in vivo pharmacological properties were evaluated. A majority of the compounds exhibited a potent antiproliferative effect on five human cancer cell lines, especially those causing breast cancer. Compound 4f showed the highest antiproliferative potency among all of the compounds, almost a 10-fold higher inhibitory potency against thioredoxin reductase (TrxR) compared with PL in cells causing breast cancer. In addition, 4f was found to increase the levels of reactive oxygen species (ROS), thus leading to more potent antiproliferative effects. More importantly, the suppression assays of migration and invasion revealed that compound 4f could reverse the epithelial-mesenchymal transition induced by the transforming growth factor β1, and exhibit prominent anti-metastasis effects. Compound 4f also showed strong inhibition potency toward solid tumors of breast cancer in vivo. Our findings show that compound 4f is a promising therapeutic candidate in the treatment of breast cancer, which, however, needs further research to be proved., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Terpenes constitute one of the largest groups of natural products. They exhibit a wide range of biological activities including antioxidant, anticancer, and drug resistance modulating properties. Saffron extract and its terpene constituents have been demonstrated to be cytotoxic against various types of cancer cells, including breast, liver, lung, pancreatic, and colorectal cancer. In the present work, we have studied anticancer properties of TMPE, a newly synthesized monoterpene derivative of β-cyclocitral-the main volatile produced by the stigmas of unripe crocuses. TMPE presented selective cytotoxic activity to doxorubicin-resistant colon cancer cells and was identified to be an effective MDR modulator in doxorubicin-resistant cancer cells. Synergy between this derivative and doxorubicin was observed. Most probably, TMPE inhibited transport activity of ABCB1 protein without affecting its expression level. Analysis of TMPE physicochemical parameters suggested it was not likely to be transported by ABCB1. Molecular modeling showed TMPE being more reactive molecule than the parental compound-β-cyclocitral. Analysis of electrostatic potential maps of both compounds prompted us to hypothesize that reduced reactivity as well as susceptibility to electrophilic attack were related to the lower general toxicity of β-cyclocitral. All of the above pointed to TMPE as an interesting candidate molecule for MDR reversal in cancer cells.

Anxiety and depression have high prevalence in the general population, affecting millions of people worldwide, but there is still a need for effective and safe treatments. Nutritional supplements have recently received a lot of attention, particularly saffron. Thus, several pre-clinical studies support a beneficial role for bioactive compounds, such as saffron, in anxiety and depression. Here we used an animal model of depression based on social isolation to assess the effects of affron ® , a standardized saffron extract containing ≥3.5% of total bioactive compounds safranal and crocin isomers. Affron ® was administered both through the oral and the intraperitoneal routes, and several tasks related to anxiety and depression, such as the elevated plus maze, the forced swimming test or the sucrose preference test, were assessed. These tasks model key features of depressive states and anxious states relating to fear, behavioral despair or anhedonia, the lack of motivation and/or pleasure from everyday activities, respectively. Animals receiving oral affron ® displayed behaviors congruent with improvements in their anxious/depressive state, showing the enhanced consumption of a sweet solution, as well as an increase in certain escape responses in the forced swimming test. Our data support a beneficial role for oral saffron in anxious/depressive states.

Background: 2-Dodecyl-6-methoxycyclohexa-2,5-diene-1,4-dione (DMDD) has been reported to inhibit a variety of cancer cell lines. The purpose of this study was to investigate the effects of DMDD on 4T1 breast cancer cells and the effects of DMDD on 4T1 breast cancer in mice and its molecular mechanisms., Methods: 4T1 breast cancer cells were treated with different concentrations of DMDD, and their proliferation, apoptosis, cell-cycle distribution, migration, and invasion were detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT, Acridine orange and ethidium bromide dual staining analysis (AO/EB) dual staining, flow cytometry, scratch test, and the Transwell assay. Relative quantitative real-time qPCR analysis and Western blot were applied to examine the expression levels of related genes and proteins. In animal experiments, we established a xenograft model to assess the anti-breast cancer effects of DMDD by evaluating the inhibition rate. The apoptotic activity of DMDD was evaluated by hematoxylin-eosin (HE) staining, transmission electron microscope (TEM) analysis and TdT-mediated dUTP nick end labeling (TUNEL) assays. The mRNA expression levels of MAPK pathway components were detected by relative quantitative real-time qPCR. In addition, the protein expression levels of MAPK pathway components were assessed through immunohistochemical assays and Western blotting., Results: Experiments showed that DMDD could inhibit the proliferation, migration, invasion of 4T1 cells and induce cellular apoptosis and G1 cell cycle arrest. Moreover, DMDD down-regulated the mRNA expressions of raf1, mek1, mek2, erk1, erk2, bcl2, and up-regulated the mRNA expression of bax. DMDD reduced the protein expressions of p-raf1, p-mek, p-erk, p-p38, Bcl2, MMP2, MMP9 and increased the protein expressions of Bax and p-JNK. The results showed that DMDD can effectively reduce the tumor volume and weight of breast cancer in vivo, up-regulate the expression of IL-2, down-regulate the expression of IL-4 and IL-10, induce the apoptosis of breast cancer cells in mice, and regulate the expression of genes and proteins of the MAPK pathway., Conclusion: Our study indicates that DMDD can inhibit proliferation, migration, and invasion and induces apoptosis and cell-cycle arrest of 4T1 breast cancer cells. Also, our findings indicate that DMDD induces the apoptosis of breast cancer cells and inhibits the growth in mice. Its mechanism may be related to the MAPK pathway., Competing Interests: The authors state that there is no conflict of interest in this work., (© 2020 Zhou et al.)

Safranal (SFR) is the major constituent of saffron. The purpose of this study was to observe the effect of SFR on myocardial ischemia induced by isoprenaline (ISO) and to explore its possible mechanism. The myocardial ischemia rat model was established by subcutaneous injection of ISO (85 mg/kg/d) on the 8th and 9th day of the experiment. Serum creatine kinase (CK), lactate dehydrogenase (LDH), malondialdehyde (MDA) and superoxide dismutase (SOD) were measured, as were changes in calcium concentration, reactive oxygen species (ROS) and cardiac morphology of the myocardial tissue. The effects of SFR on cell contraction, Ca 2+ transient and L-type Ca 2+ current (I Ca-L ) in isolated rat myocardial cells were measured using the Ion Optix detection system and the whole-cell patch-clamp technique. SFR can decrease the activity of serum CK, LDH and MDA, and increase the activity of serum SOD, reduce intracellular calcium concentration and the manufacture of ROS. In addition, SFR can improve changes in heart morphology. SFR can significantly inhibit contraction, Ca 2+ transients and I Ca-L in isolated ventricular myocytes. SFR has a cardioprotective role in ISO-induced MI rats, and the underling mechanism is related to the inhibition of oxidative stress, myocardial contractility, I Ca-L and the regulation of Ca 2+ homeostasis., Competing Interests: Conflict of interest The authors declare that there are no conflicts of interest., (Copyright © 2020 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

The chemical analysis of the sponge Dysidea avara afforded the known sesquiterpene quinone avarone, along with its reduced form avarol. To further explore the role of the thiazinoquinone scaffold as an antiplasmodial, antileishmanial and antischistosomal agent, we converted the quinone avarone into the thiazinoquinone derivative thiazoavarone. The semisynthetic compound, as well as the natural metabolites avarone and avarol, were pharmacologically investigated in order to assess their antiparasitic properties against sexual and asexual stages of P lasmodium falciparum , larval and adult developmental stages of Schistosoma mansoni (eggs included), and also against promastigotes and amastigotes of Leishmania infantum and Leishmania tropica . Furthermore, in depth computational studies including density functional theory (DFT) calculations were performed. A toxic semiquinone radical species which can be produced starting both from quinone- and hydroquinone-based compounds could mediate the anti-parasitic effects of the tested compounds., Competing Interests: The authors declare no conflict of interest.

Dementia and cognitive decline are global public health problems. Moderate consumption of alcoholic beverages reduces the risk of dementia and cognitive decline. For instance, resveratrol, a polyphenolic compound found in red wine, has been well studied and reported to prevent dementia and cognitive decline. However, the effects of specific beer constituents on cognitive function have not been investigated in as much detail. In the present review, we discuss the latest reports on the effects and underlying mechanisms of hop-derived bitter acids found in beer. Iso-α-acids (IAAs), the main bitter components of beer, enhance hippocampus-dependent memory and prefrontal cortex-associated cognitive function via dopamine neurotransmission activation. Matured hop bitter acids (MHBAs), oxidized components with β-carbonyl moieties derived from aged hops, also enhance memory functions via norepinephrine neurotransmission-mediated mechanisms. Furthermore, the effects of both IAAs and MHBAs are attenuated by vagotomy, suggesting that these bitter acids enhance cognitive function via vagus nerve stimulation. Moreover, supplementation with IAAs attenuates neuroinflammation and cognitive impairments in various rodent models of neurodegeneration including Alzheimer's disease. Daily supplementation with hop-derived bitter acids (e.g., 35 mg/day of MHBAs) may be a safe and effective strategy to stimulate the vagus nerve and thus enhance cognitive function.

We present a study that uses a laser trapping technique for measurement of radiation sensitivity of untreated and chemo-treated cancer cells. We used a human mammary tumor cell line (4T1) treated by an antitumor compound, 2-Dodecyl-6-methoxycyclohexa-2, 5-diene-1,4-dione (DMDD), which was extracted from the root of Averrhoa carambola L. The untreated control group, and both 2-hour and 24-hour treated groups of 4T1 cells were used in this study. The absorbed threshold ionization energy (TIE) and the threshold radiation dose (TRD) were determined using a high-power infrared laser (at 1064 nm) trap by single and multiple cells trapping and ionization. The results were analyzed using descriptive and t-statistics. The relation of the TIE and TRD to the mass of the individual cells were also analyzed for different hours of treatment in comparison with the control group. Both TIE and TRD decrease with increasing treatment periods. However, the TRD decreases with mass regardless of the treatment. Analyses of the TRD for single vs multiple cells ionizations within each group have also consistently showed this same behavior regardless of the treatment. The underlying factors for these observed relations are explained in terms of radiation, hyperthermia, and chemo effects.

DNA mismatch repair (MMR) deficiency has been associated with a higher risk of developing colorectal, endometrial, and ovarian cancer, and confers resistance in conventional chemotherapy. In addition to the lack of treatment options that work efficaciously on these MMR-deficient cancer patients, there is a great need to discover new drug leads for this purpose. In this study, we screened through a library of commercial and semisynthetic natural compounds to identify potential synthetic lethal drugs that may selectively target MLH1 mutants using MLH1 isogenic colorectal cancer cell lines and various cancer cell lines with known MLH1 status. We identified a novel diarylpentanoid analogue, 2-benzoyl-6-(2,3-dimethoxybenzylidene)-cyclohexenol, coded as AS13, that demonstrated selective toxicity toward MLH1-deficient cancer cells. Subsequent analysis suggested AS13 induced elevated levels of oxidative stress, resulting in DNA damage where only the proficient MLH1 cells were able to be repaired and hence escaping cellular death. While AS13 is modest in potency and selectivity, this discovery has the potential to lead to further drug development that may offer better treatment options for cancer patients with MLH1 deficiency.

Many medicinal plant species are currently threatened in their natural habitats because of the growing demand for phytochemicals worldwide. A sustainable alternative for the production of bioactive plant compounds are plant biofactories based on cell cultures and organs. In addition, plant extracts from biofactories have significant advantages over those obtained from plants, since they are free of contamination by microorganisms, herbicides and pesticides, and they provide more stable levels of active ingredients. In this context, we report the establishment of Satureja khuzistanica cell cultures able to produce high amounts of rosmarinic acid (RA). The production of this phytopharmaceutical was increased when the cultures were elicited with coronatine and scaled up to a benchtop bioreactor. S. khuzistanica extracts enriched in RA were found to reduce the viability of cancer cell lines, increasing the sub-G0/G1 cell population and the activity of caspase-8 in MCF-7 cells, which suggest that S. khuzistanica extracts can induce apoptosis of MCF-7 cells through activation of the extrinsic pathway. In addition, our findings indicate that other compounds in S. khuzistanica extracts may act synergistically to potentiate the anticancer activity of RA.

Safranal, the main volatile chemical of Saffron ( Crocus sativus ) was studied to estimate its allelopathic effects on the photosynthetic pigment chlorophyll, leaf electrolyte leakage, fresh weight, catalase (CAT), and peroxidase (POX) activity of the test plant Lettuce ( Lactuca sativa ). In this study, the effective concentration (EC 50 ) of safranal on CAT was estimated to be 6.12 µg/cm 3 . CAT activity was inhibited in a dose-dependent manner by the increase in the safranal concentration while POX activity was increased. Moreover, Safranal caused significant physiological changes in chlorophyll content, leaf electrolyte leakage, and fresh weight of several weed species with Lolium multiflorum being the most sensitive. Furthermore, 5 µM Safranal showed significant inhibitory activity against dicotyledonous in comparison to the monocotyledons under greenhouse conditions. The inhibition of the CAT by safranal was similar to those of uncompetitive inhibitors, and therefore the decline in carbon fixation by plants might be the mechanism behind the inhibitory activity of safranal., Competing Interests: The authors declare no conflict of interest.

Background: Hyperglycemia stimulated epithelial-mesenchymal transition (EMT) plays a critical role in initiating and progressing renal fibrosis in diabetic kidney disease (DKD). It is crucial to explore novel renal protective drugs for the treatment of DKD., Objective: The present study is to confirm our hypothesis and to accumulate the information for the application of DMDD (2-Dodecyl-6-methoxycyclohexa-2,5-diene-1,4-dione) as a novel therapeutic agent to potentially inhibit renal fibrogenesis and EMT in the DKD., Methods: High glucose induced renal proximal tubular epithelial cell line (HK-2 cells) was cultured and treated with DMDD. The cell viability and DMDD cytotoxicity were assessed by CCK8. Immunofluorescence was used for detection of TLR4 and downstream protein in normal and high glucose induced HK-2 cells. HK-2 cells were transfected with lentivirus codifying for BAMBI (BMP and activin membrane bound inhibitor) and interfering RNA for determination of the effect of BAMBI over-expression and silencing, respectively. TLR4-BAMBI-Smad2/3 pathway was analyzed by means of RT-PCR and western blot., Results: A high concentration (60mM) of glucose induced significant EMT process and TLR4 expression was increased obviously in this circumstance. DMDD inhibited high expressions of TLR4 and Smad2/3 in HG induced cells and decreased the expression of BAMBI. In addition, the effects of decreased BAMBI expression and increased Smad2/3 expression in HG cultured cells were reversed in the cells of TAK-242 (TLR4 signaling inhibitor) intervention. BAMBI gene silencing dramatically increased EMT process and the over-expression of BAMBI was opposite in HK-2 cells with HG condition. These observations of EMT were ameliorated when the HK-2 cells were pre-treated with DMDD., Conclusions: Our study demonstrates that DMDD treatment improves EMT in the HG induced HK-2 cells. In addition, DMDD significantly inhibits EMT by TLR4-BAMBI-Smad2/3 pathway, which hints that DMDD may be an alternative approach in diabetic renal injury., (Copyright © 2019. Published by Elsevier Masson SAS.)

The chemical composition and biological activity of essential oils isolated from the leaves of Siparuna aspera , Siparuna macrotepala , Piper leticianum , Piper augustum and the rhizome of Hedychium coronarium were evaluated. These species are used medicinally in different ways by the Amazonian communities that live near the Kutukú mountain range. Chemical studies revealed that the main components for the two Siparuna species were germacrene D, bicyclogermacrene, α-pinene, δ-cadinene, δ-elemene, α-copaene and β-caryophyllene; for the two Piper species β-caryophyllene, germacrene D, α-( E,E )-farnesene, β-elemene, bicyclogermacrene, δ-cadinene and for H. coronarium 1,8-cineole, β-pinene, α-pinene and α-terpineol. The antioxidant activity of all essential oils was evaluated by 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), photochemiluminescence (PCL) quantitative assays, and DPPH and ABTS bioautographic profiles, with different results for each of them. Antimicrobial activity studies were carried out on three yeasts, six Gram positive and four Gram negative bacteria, by means of the disc diffusion method. The essential oil of H. coronarium showed the most relevant results on L. grayi , K. oxytoca and S. mutans , P. augustum and P. leticianum on S. mutans . An antibacterial bioautographic test for H. coronarium was also carried out and highlighted the potential activity of terpinen-4-ol and 1,8-cineole.

Essential oils (EOs) have been shown to have a diversity of beneficial human health effects. Clausena is a large and highly diverse genus of plants with medicinal and cosmetic significance. The aim of this study was to analyze the composition of Clausena lansium EOs and to investigate their potential antifungal effects. The chemical compositions of Clausena lansium EOs obtained by hydrodistillation were analyzed by gas chromatography-mass spectrometry (GC-MS). A total of 101 compounds were identified among the diverse extracts of C. lansium . EOs of leaves and pericarps from different cultivars (Hainan local wampee and chicken heart wampee) collected in Hainan (China) were classified into four clusters based on their compositions. These clusters showed different antifungal activities against five Candida species ( C. albicans , C. tropicalis , C. glabrata , C. krusei and C. parapsilosis ) using the disc diffusion method. Clausena lansium EOs of pericarps displayed noteworthy antifungal activitives against all the tested Candida strains with inhibition zone diameters in the range of 11.1–23.1 mm. EOs of leaves showed relatively low antifungal activities with inhibition zone diameters in the range of 6.5–22.2 mm. The rank order of antifungal activities among the four EO clusters was as follows: Cluster IV> Cluster III > Cluster I ≥ Cluster II. These results represent the first report about the correlation between chemical composition of C. lansium EOs and antifungal activity. Higher contents of β-phellandrene, β-sesquiphellandrene and β-bisabolene in EOs of pericarps were likely responsible for the high antifungal activity of Cluster IV EOs. Taken together, our results demonstrate the chemical diversity of Clausena lansium EOs and their potential as novel antifungal agents for candidiasis caused by Candida spp. Furthermore, the obtained results showing a wide spectrum of antifungal activities provide scientific evidence for the traditional use of these plants.

Safranal, contained in Crocus sativus L., exerts anti-inflammatory and analgesic effects. However, the underlying mechanisms for such effects are poorly understood. We explored whether safranal targets the transient receptor potential ankyrin 1 (TRPA1) channel, which in nociceptors mediates pain signals. Safranal by binding to specific cysteine/lysine residues, stimulates TRPA1, but not the TRP vanilloid 1 and 4 channels (TRPV1 and TRPV4), evoking calcium responses and currents in human cells and rat and mouse dorsal root ganglion (DRG) neurons. Genetic deletion or pharmacological blockade of TRPA1 attenuated safranal-evoked release of calcitonin gene-related peptide (CGRP) from rat and mouse dorsal spinal cord, and acute nociception in mice. Safranal contracted rat urinary bladder isolated strips in a TRPA1-dependent manner, behaving as a partial agonist. After exposure to safranal the ability of allyl isothiocyanate (TRPA1 agonist), but not that of capsaicin (TRPV1 agonist) or GSK1016790A (TRPV4 agonist), to evoke currents in DRG neurons, contraction of urinary bladder strips and CGRP release from spinal cord slices in rats, and acute nociception in mice underwent desensitization. As previously shown for other herbal extracts, including petasites or parthenolide, safranal might exert analgesic properties by partial agonism and selective desensitization of the TRPA1 channel., (© 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Diarrhea is one of the leading causes of infant death in the world accounting for high child mortality rate. It is also present in different pathophysiologies related to several etiological agents. The aim of this study is to investigate the antidiarrheal effect of α -Terpineol (α-TPN) in different diarrhea models in rodents. The antidiarrheal effect of α-TPN in the treatment of acute diarrhea and enteropooling induced by castor oil or PGE 2 in Swiss mice pretreated orally with saline (NaCl 0.9%), Loperamide (5 mg/kg) and α-TPN (6.25, 12.5, 25 and 50 mg/kg) was analyzed. Additionally, parameters of severity, total weight of faeces and post-treatment for 4 h were evaluated. Modulation of the opioid and cholinergic pathways was performed and intestinal transit model using activated charcoal as marker was also used. The effect of α-TPN on secretory diarrhea was investigated using the model of fluid secretion in intestinal loops isolated from cholera toxin-treated mice. α-TPN showed antidiarrheal effect (*p < 0.05), reducing the total stool amount (*55%, *48%, *44%, *24%) and diarrheal (*47%, *66%; *56%, 10%) respectively for the doses tested. All doses investigated in the enteropooling test presented significant changes (*46%, *78%, *66%, *41% respectively) in relation to the control. α-TPN through the muscarinic pathway reduced the gastrointestinal transit (*31%), besides inhibiting PGE 2 -induced diarrhea (*39%). α-TPN also reduced fluid formation and loss of Cl - ions, by interacting directly with GM1 receptors and cholera toxin, thus increasing the uptake of intestinal fluids. The results suggest an anti-diarrheal activity of α-TPN due to its anticholinergic action, ability to block PGE 2 and GM1 receptors and interaction with cholera toxin in secretory diarrhea, making it a promising candidate drug for the treatment of diarrheal diseases., (Copyright © 2018 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Interleukin (IL)-33 is a member of the IL-1 family and is able to act cardioprotective. The aim of this study was to investigate the regulation of IL-33 by 3-hydroxy-3-methylglutaryl-coenzyme-A (HMG-CoA) reductase inhibitors (statins) and bisphosphonates (BPs) in human cardiac tissue. The lipophilic fluvastatin, simvastatin, atorvastatin, and lovastatin as well as the nitrogenous BPs alendronate and ibandronate, but not hydrophilic pravastatin increased IL-33 mRNA and intracellular IL-33 protein levels in both human adult cardiac myocytes (HACM) and fibroblasts (HACF). Additionally, fluvastatin reduced soluble ST2 secretion from HACM. IL-33 was also up-regulated by the general inhibitor of prenylation perillic acid, a RhoA kinase inhibitor Y-27632, and by latrunculin B, but statin-induced IL-33 expression was inhibited by mevalonate, geranylgeranyl pyrophosphate (GGPP) and RhoA activator U-46619. The IL-33 promoter was 2.3-fold more accessible in statin-treated HACM compared to untreated cells (P = 0.037). In explanted hearts of statin-treated patients IL-33 protein was up-regulated as compared with the hearts of non-statin-treated patients (P = 0.048). As IL-33 was previously shown to exert cardioprotective effects, one could speculate that such up-regulation of IL-33 expression in human cardiac cells, which might happen mainly through protein geranylgeranylation, could be a novel mechanism contributing to known cardioprotective effects of statins and BPs., (© 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Context: Safranal (SAF) is verified to have potential effects in promoting nerve growth., Objectives: This study verifies the role of SAF in promoting dopaminergic neurons growth in vitro and in vivo., Material and Methods: Rat neural stem cells (NSC) were treated with 1, 20, or 100 ng/mL of SAF, and the expression levels of tyrosine hydroxylase (TH) and dopamine transporter (DAT) were assayed by flow cytometry and real-time PCR and the secretion of dopamine (DA) was assayed by ELISA. Then, 2 × 10 6  cells of SAF-treated NSC was administrated into PD rat models induced by 6-OHDA. The differentiation and survival of dopaminergic neurons was identified by fluorescence microscope and TH + cells by immunostaining and DA secretion by ELISA at week 2 and week 4, respectively., Results: After being treated with SAF at 20 and 100 ng/mL for 1 week, TH and DAT positive rates increased 1.4- and 1.7-fold (p < 0.01, respectively). TH and DAT mRNA also increased 8.05- and 4.41-fold, respectively. And the release of DA statistically increased 1.5-fold (p < 0.01). In vivo, the number of rotations decreased to 4.33 ± 0.97 rpm (p < 0.01) and the survival rates increased to 77.66 ± 7.87% (p < 0.05) at week 4 after transplantation of SAF-treated NSC. Moreover, the transplanted cells increased three-fold, TH fluorescence density increased four-fold and DA releases increased 1.4-fold (p < 0.01) at week 4 after transplantation., Conclusions: SAF promoted the production of functional DA cells and alleviated PD, which may contribute to a new therapy for PD patients.

Non-alcoholic fatty liver disease (NAFLD) is considered to be the hepatic manifestation of the metabolic syndrome. Iso-alpha acids (IAAs), hop-derived bitter compounds in beer, have been shown to beneficially affect different components of the metabolic syndrome such as insulin resistance and dyslipidemia. However, IAAs have not yet been studied in the context of chronic liver disease. Here we analyzed the effect of IAA on the pathogenesis of NAFLD. Once, we applied IAA to mice in combination with a NAFLD-inducing Western-type diet (WTD), and observed that IAA significantly inhibited WTD-induced body weight gain, glucose intolerance, and hepatic steatosis. Fitting to this, IAA dose-dependently inhibited cellular lipid accumulation in primary human hepatocytes (PHH) in vitro. Reduced expression of PPAR-gamma and key enzymes of lipid synthesis as well as increased expression of PPAR-alpha, indicative for increased lipid combustion, were identified as underlying mechanisms of reduced hepatocellular steatosis in vitro and in vivo. Analysis of hepatic HMOX1 expression indicated reduced oxidative stress in IAA-treated mice, which was paralleled by reduced activation of the JNK pathway and pro-inflammatory gene expression and immune cell infiltration. Furthermore, IAA reduced hepatic stellate cell (HSC) activation and pro-fibrogenic gene expression. Similarly, IAA also dose-dependently reduced oxidative stress and JNK activation in steatotic PHH, inhibited HSC activation, and reduced proliferation and pro-fibrogenic gene expression in already activated HSC in vitro. In conclusion, IAAs inhibit different pathophysiological steps of disease progression in NAFLD. Together with previous studies, which demonstrated the safety of even long-term application of IAA in humans, our data suggest IAA as promising therapeutic agent for the prevention and treatment of (non)alcoholic (fatty) liver disease.

Hop-derived compounds have been subjected to numerous biomedical studies investigating their impact on a wide range of pathologies. Isomerised bitter acids (isoadhumulone, isocohumulone and isohumulone) from hops, used in the brewing process of beer, are known to inhibit members of the aldo-keto-reductase superfamily. Aldo-keto-reductase 1B10 (AKR1B10) is upregulated in various types of cancer and has been reported to promote carcinogenesis. Inhibition of AKR1B10 appears to be an attractive means to specifically treat RAS-dependent malignancies. However, the closely related reductases AKR1A1 and AKR1B1, which fulfil important roles in the detoxification of endogenous and xenobiotic carbonyl compounds oftentimes crossreact with inhibitors designed to target AKR1B10. Accordingly, there is an ongoing search for selective AKR1B10 inhibitors that do not interact with endogeneous AKR1A1 and AKR1B1-driven detoxification systems. In this study, unisomerised α-acids (adhumulone, cohumulone and n -humulone) were separated and tested for their inhibitory potential on AKR1A1, AKR1B1 and AKR1B10. Also AKR1B10-mediated farnesal reduction was effectively inhibited by α-acid congeners with K i -values ranging from 16.79 ± 1.33 µM (adhumulone) to 3.94 ± 0.33 µM ( n -humulone). Overall, α-acids showed a strong inhibition with selectivity (115⁻137 fold) for AKR1B10. The results presented herein characterise hop-derived α-acids as a promising basis for the development of novel and selective AKR1B10-inhibitors.

Poor prognoses remain the most challenging aspect of hepatocellular carcinoma (HCC) therapy. Consequently, alternative therapeutics are essential to control HCC. This study investigated the anticancer effects of safranal against HCC using in vitro, in silico, and network analyses. Cell cycle and immunoblot analyses of key regulators of cell cycle, DNA damage repair and apoptosis demonstrated unique safranal-mediated cell cycle arrest at G2/M phase at 6 and 12 h, and at S-phase at 24 h, and a pronounced effect on DNA damage machinery. Safranal also showed pro-apoptotic effect through activation of both intrinsic and extrinsic initiator caspases; indicating ER stress-mediated apoptosis. Gene set enrichment analysis provided consistent findings where UPR is among the top terms of up-regulated genes in response to safranal treatment. Thus, proteins involved in ER stress were regulated through safranal treatment to induce UPR in HepG2 cells.

The cytotoxic and antiprotozoal activities of the phytoquinoide, jacaranone, and related compounds have been an ongoing topic in recent drug discovery. Starting from the natural product-derived cyclohexadienone scaffold, a series of nitrogen-containing derivatives were synthesized and subsequently evaluated for their antiproliferative and antiprotozoal activity. Anticancer potency was analyzed using different types of cancer cell lines: MDA-MB-231 breast cancer, CCRF-CEM leukemia, HCT-116 colon cancer, U251 glioblastoma, and, in addition, non-tumorigenic MRC-5 lung fibroblasts. Antiproliferative activities at micromolar concentrations could be shown. Antiprotozoal activity was assessed against Plasmodium falciparum NF54 and Trypanosoma brucei rhodesiense STIB900. For all compounds, selectivity indices (SI) were calculated based on assessed cytotoxicity towards L6 cells. In addition, the structure-activity-relationships and physicochemical parameters of these compounds are discussed.

Safranal is one of saffron constituents and has antioxidant and neuroprotective properties. Metformin is used as an anti-diabetic drug. This study was planned to investigate the separate and combined treatment effects of safranal and metformin on diabetes-induced learning and memory impairments by behavioral and hippocampal histopathological and biochemical evaluations. Diabetes was induced by intraperitoneal injection of streptozotocin (STZ), treatments with safranal (0.025, 0.1 and 0.4 mg/kg), metformin (50 and 200 mg/kg), and a combination of low doses of this chemicals were initiated after confirmation of diabetes and continued for 37 days. Blood glucose concentration was measured before and on days 15, 25 and 35 after injection of streptozotocin. Learning and memory tested using Morris Water Maze (MWM) on days 40-45 and on day 45 hippocampal specimens were collected for determination of malodialdehyde (MDA), tumor necrosis factor-alpha (TNF-α) and Caspase-3 levels and superoxide dismutase (SOD) activity. The hippocampus was also designed for light microscopy evaluation. Hyperglycemia, spatial learning and memory impairments, hippocampal neuron loss, increase of hippocampal MDA, TNF-α and caspase-3 levels and decrease of SOD activity were observed in diabetic rats. Safranal (0.1 and 0.4 mg/kg), metformin (200 mg/kg) and safranal (0.025 mg/kg) with metformin (50 mg/kg) improved the above-mentioned behavioral, histopathological and biochemical changes. Safranal and metformin and their combination improved learning and memory impairments in STZ-induced diabetic rats. Antioxidant, anti-inflammatory and antiapoptotic mechanisms might be involved. It is recommended that safranal be considered for diabetes management., (Copyright © 2018. Published by Elsevier Masson SAS.)

The bioactive natural compound from marine origin, (+)-aeroplysinin-1, has been shown to exhibit potent anti-inflammatory and anti-angiogenic effects . The aim of the present study was to identify new targets for (+)-aeroplysinin-1 in endothelial cells. The sequential use of 2D-electrophoresis and MALDI-TOF-TOF/MS allowed us to identify several differentially expressed proteins. Four of these proteins were involved in redox processes and were validated by Western blot. The effects of (+)-aeroplysinin-1 were further studied by testing the effects of the treatment with this compound on the activity of several anti- and pro-oxidant enzymes, as well as on transcription factors involved in redox homeostasis. Finally, changes in the levels of total reactive oxygen species and mitochondrial membrane potential induced by endothelial cell treatments with (+)-aeroplysinin-1 were also determined. Taken altogether, these findings show that (+)-aeroplysinin-1 has multiple targets involved in endothelial cell redox regulation.

α-Terpineol (TP) is present in a wide range of essential oils of the genus Eucalyptus, with recognized potential for a range of biological effects, such as analgesic. Hence, our study aimed to investigate the effect of TP on cancer pain induced by sarcoma 180 in Swiss mice. Our results showed that TP reduced significantly mechanical hyperalgesia and spontaneous and palpation-induced nociception, improved paw use without reducing tumor growth and grip strength. Importantly, no evident biochemical and hematological toxicity was oberved. Furthermore, TP increased the tissue antioxidant capacity due to ferric-reducing antioxidant power (FRAP) and glutathione (GSH). TP also reduced inducible nitric oxide synthase (iNOS) immunocontent in the tumors. Molecular docking estimated that TP binds within the same range of iNOS regions (other iNOS inhibitors), such as N-Nitroarginine methyl ester (L-NAME). These data provide strong evidence that TP may be an interesting candidate for the development of new safe analgesic drugs that are effective for cancer pain control., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Chios mastic products are well-known for their broad applications in food industry, cosmetics, and healthcare since the antiquity. Given our recent finding that Chios mastic water (CMW) exerts antigenotoxic action, in the present study, we evaluated the genotoxic as well as the antigenotoxic potential of the four major compounds of CMW, namely, verbenone, α-terpineol, linalool, and trans-pinocarveol. The cytokinesis block micronucleus (CBMN) assay in cultured human lymphocytes and the Drosophila Somatic Mutation And Recombination Test (SMART), also known as the wing spot test, were employed. None of the four major CMW's constituents or their mixtures showed genotoxic or recombinogenic activity in either of the assays used. Co-treatment of each of the constituents with MMC revealed that all except trans-pinocarveol exerted antigenotoxic potential. Moreover, co-administration of verbenone with linalool or α-terpineol presented statistically significant reduction of MMC-induced mutagenicity. In conclusion, the major CMW constituents were shown to be free of genotoxic effects, while some exerted antigenotoxic activity either alone or in combinations, suggesting synergistic phenomena. Our results provide evidence on the key antigenotoxicity effectors of the plant extract CMW.

The Apiaceae family encompasses aromatic plants of economic importance employed in foodstuffs, beverages, perfumery, pharmaceuticals and cosmetics. Apiaceae are rich sources of essential oils because of the wealth of secretory structures (ducts and vittae) they are endowed with. The Apiaceae essential oils are available on an industrial level because of the wide cultivation and disposability of the bulky material from which they are extracted as well as their relatively cheap price. In the fight against protozoal infections, essential oils may represent new therapeutic options. In the present work, we focused on a panel of nine Apiaceae species (Siler montanum, Sison amomum, Echinophora spinosa, Kundmannia sicula, Crithmum maritimum, Helosciadium nodiflorum, Pimpinella anisum, Heracleum sphondylium and Trachyspermum ammi) and their essential oils as a model for the identification of trypanocidal compounds to be used as alternative/integrative therapies in the treatment of Human African trypanosomiasis (HAT) and as starting material for drug design. The evaluation of inhibitory effects of the Apiaceae essential oils against Trypanosoma brucei showed that some of them (E. spinosa, S. amomum, C. maritimum and H. nodiflorum) were active, with EC 50 in the range 2.7-10.7 μg/mL. Most of these oils were selective against T. brucei, except the one from C. maritimum that was highly selective against the BALB/3T3 mammalian cells. Testing nine characteristic individual components (α-pinene, sabinene, α-phellandrene, p-cymene, limonene, β-ocimene, γ-terpinene, terpinolene, and myristicin) of these oils, we showed that some of them had much higher selectivity than the oils themselves. Terpinolene was particularly active with an EC 50 value of 0.035 μg/mL (0.26 µM) and a selectivity index (SI) of 180. Four other compounds with EC 50 in the range 1.0-6.0 μg/mL (7.4-44 µM) had also good SI: α-pinene (>100), β-ocimene (>91), limonene (>18) and sabinene (>17). In conclusion, these results highlight that the essential oils from the Apiaceae family are a reservoir of substances to be used as leading compounds for the development of natural drugs for the treatment of HAT., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Purpose: Melanoma is one of the lethal types of skin malignancies and is responsible for significant morbidity and mortality across the globe. In this study the anticancer effects of picrocrocin on the human SK-MEL-2 malignant melanoma cell line were evaluated along with its mode of action., Methods: The effect of picrocrocin was evaluated on SKMEL- 2 cells by MTT assay. Apoptosis was determined by DAPI and annexin V/PI staining. The effects on reactive oxygen species (ROS), mitochondrial membrane potential (MMP) and cell cycle analysis were performed by flow cytometry. Expression of the proteins (JAK/STAT5) was estimated by western blotting., Results: Picrocrocin exerted growth inhibitory effects on the SK-MEL-2 melanoma cells. The IC50 of picrocrocin against the SK-MEL-2 cells was 20 μM at 24-h incubation. The antiproliferative activity of picrocrocin was due to apoptosis and cell cycle arrest. In addition, picrocrocin enhanced the ROS levels and decreased the MMP levels of SK-MEL-2 cells. Finally, picrocrocin inhibited the JAK/STAT5 signalling pathway in the SK-MEL-2 melanoma cells., Conclusions: Taken together, these results indicate that picrocrocin can prove to be an important molecule in the search of more efficient therapies for melanoma.

The present study used isometric tension recording to investigate the vasorelaxant effect of limonene (LM), carveol (CV), and perillyl alcohol (POH) on contractility parameters of the rat aorta, focusing in particular on the structure-activity relationship. LM, CV, and POH showed a reversible inhibitory effect on the contraction induced by electromechanical and pharmacomechanical coupling. In the case of LM, but not CV and POH, this effect was influenced by preservation of the endothelium. POH and CV but not LM exhibited greater pharmacological potency on BayK-8644-induced contraction and on electromechanical coupling than on pharmacomechanical coupling. In endothelium-denuded preparations, the order of pharmacological potency on electrochemical coupling was LM < CV < POH. These compounds inhibited also, with grossly similar pharmacological potency, the contraction induced by phorbol ester dibutyrate. The present results suggest that LM, CV and POH induced relaxant effect on vascular smooth muscle by means of different mechanisms likely to include inhibition of PKC and IP3 pathway. For CV and POH, hydroxylated compounds, it was in electromechanical coupling that the greater pharmacological potency was observed, thus suggesting a relative specificity for a mechanism likely to be important in electromechanical coupling, for example, blockade of voltage-dependent calcium channel.

Xerostomia (dry mouth) is the most common side effect of radiation therapy in patients with head and neck cancer and causes difficulty speaking and swallowing. Since aldehyde dehydrogenase 3A1 (ALDH3A1) is highly expressed in mouse salivary stem/progenitor cells (SSPCs), we sought to determine the role of ALDH3A1 in SSPCs using genetic loss-of-function and pharmacologic gain-of-function studies. Using DarkZone dye to measure intracellular aldehydes, we observed higher aldehyde accumulation in irradiated Aldh3a1 -/- adult murine salisphere cells and in situ in whole murine embryonic salivary glands enriched in SSPCs compared with wild-type glands. To identify a safe ALDH3A1 activator for potential clinical testing, we screened a traditional Chinese medicine library and isolated d-limonene, commonly used as a food-flavoring agent, as a single constituent activator. ALDH3A1 activation by d-limonene significantly reduced aldehyde accumulation in SSPCs and whole embryonic glands, increased sphere-forming ability, decreased apoptosis, and improved submandibular gland structure and function in vivo after radiation. A phase 0 study in patients with salivary gland tumors showed effective delivery of d-limonene into human salivary glands following daily oral dosing. Given its safety and bioavailability, d-limonene may be a good clinical candidate for mitigating xerostomia in patients with head and neck cancer receiving radiation therapy., Competing Interests: The authors declare no conflict of interest., (Copyright © 2018 the Author(s). Published by PNAS.)

Background: Limonene is a cyclic monoterpene (CTL) found in citrus fruits and many plant kingdoms. It has attracted attention as potential molecule due to its diverse biological activities. However, molecular mechanism involved in the osteogenic induction of CTL in C2C12 skeletal muscle cells remain unclear., Purpose: Skeletal development maintains the bone homeostasis through bone remodeling process. It coordinated between the osteoblast and osteoblast process. Osteoporosis is one of the most common bone diseases caused by a systemic reduction in bone mass. Recent osteoporosis treatment is based on the use of anti-resorptive and bone forming drugs. However, long term use of these drugs is associated with serious side effects and strategies on the discovery of lead compounds from natural products for osteoblast differentiation are urgently needed. Therefore, we planned to find out the role of CTL on osteoblast differentiation and glucose uptake in C2C12 cells and its effect on signaling pathways., Methods: Cell proliferation, alkaline phosphatase (ALP) activity, calcium deposition, genes, and proteins associated with osteoblast activation and glucose utilization were analysed., Results: CTL did not affect the cell viability. CTL significantly increased ALP activity, calcium depositions and the expression of osteogenic specific genes such as Myogenin, Myogenic differentiation 1 (MyoD), ALP, Run-related transcription factor 2(RUNX2), osteocalcin (OCN). In addition, CTL induced the mRNA expression of bone morphogenetic proteins (BMP-2 BMP-4 BMP-6 BMP-7 BMP-9). CTL treatment enhanced 2-Deoxy-d-glucose (2DG) uptake. Moreover, CTL stimulated the activation of p38 mitogen activated protein kinase (p38MAPK), Protein kinase B (Akt), Extracellular signal related kinase (ERKs) by increasing phosphorylation. CTL treatment abolished p38 inhibitor (SB203580) mediated inhibition of osteoblast differentiation, but no effect was noted by ERKs specific inhibitor (PD98059)., Conclusion: These results suggest that limonene induces osteoblast differentiation and glucose uptake through activating p38MAPK and Akt signaling pathways, confirming the molecular basis of the osteoblast differentiation by limonene in C2C12 skeletal muscle cells., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

The knowledge of aggregate exposure to different types of products is paramount in the risk assessment. The aim of this study was to compare the relative contribution of essential oils compared to cosmetics on the daily dermal exposure to limonene, an ubiquitous fragrance compound that can be an allergen depending on its degree of oxidation. Aggregate daily exposure to limonene was calculated among a panel of French volunteers using both essential oils and cosmetics, for 4 different specific zones, i.e. face and neck, chest, upper limbs and lower limbs. Calculations were made using a probabilistic Monte Carlo method and sensitivity analysis. The main strength of this work was the inclusion of essential oils in addition to cosmetics in the model. For the first time, the generated data could be used to compare the contribution of these two products in dermal exposure. Essential oils appear to be significant contributors to exposure to limonene particularly for the face. This work is a first step that will permit to determine the exposure to other fragrance compounds with sensitizing potential. These data will be useful for risk managers to consider the inclusion of essential oils in the overall burden of this pathology., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The effects of MC-3129, a synthetic cyclohexene derivative, on cell viability and apoptosis have been investigated in human leukemia cells. Exposure of leukemia cells to MC-3129 led to the inhibition of cell viability and induction of apoptosis through the dephosphorylation and mitochondrial translocation of cofilin. A mechanistic study revealed that interruption of the RhoA/ROCK1/PTEN/PI3K/Akt signaling pathway plays a crucial role in the MC-3129-mediated dephosphorylation and mitochondrial translocation of cofilin and induction of apoptosis. Our in vivo study also showed that the MC-3129-mediated inhibition of the tumor growth in a mouse leukemia xenograft model is associated with the interruption of ROCK1/PTEN/PI3K/Akt signaling and apoptosis. Molecular docking suggested that MC-3129 might activate the RhoA/ROCK1 pathway by targeting LPAR2. Collectively, these findings suggest a hierarchical model, in which the induction of apoptosis by MC-3129 primarily results from the activation of RhoA/ROCK1/PTEN and inactivation of PI3K/Akt, leading to the dephosphorylation and mitochondrial translocation of cofilin, and culminating in cytochrome c release, caspase activation, and apoptosis. Our study reveals a novel role for RhoA/ROCK1/PTEN/PI3K/Akt signaling in the regulation of mitochondrial translocation of cofilin and apoptosis and suggests MC-3129 as a potential drug for the treatment of human leukemia.

Over 10% of pheochromocytoma and paraganglioma (PPGL) patients have malignant disease at their first presentation in the clinic. Development of malignancy and the underlying molecular pathways in PPGLs are poorly understood and efficient treatment strategies are missing. Marine sponges provide a natural source of promising anti-tumorigenic and anti-metastatic agents. We evaluate the anti-tumorigenic and anti-metastatic potential of Aeroplysinin-1 and Isofistularin-3, two secondary metabolites isolated from the marine sponge Aplysina aerophoba , on pheochromocytoma cells. Aeroplysinin-1 diminished the number of proliferating cells and reduced spheroid growth significantly. Beside these anti-tumorigenic activity, Aeroplysinin-1 decreased the migration ability of the cells significantly ( p = 0.01), whereas, the invasion capacity was not affected. Aeroplysinin-1 diminished the high adhesion capacity of the MTT cells to collagen ( p < 0.001) and, furthermore, reduced the ability to form spheroids significantly. Western Blot and qRT-PCR analysis showed a downregulation of integrin β1 that might explain the lower adhesion and migration capacity after Aeroplysinin-1 treatment. Isofistularin-3 showed only a negligible influence on proliferative and pro-metastatic cell properties. These in vitro investigations show promise for the application of the sponge-derived marine drug, Aeroplysinin-1 as anti-tumorigenic and anti-metastatic agent against PPGLs for the first time., Competing Interests: The authors declare no conflict of interest.