Insight into antioxidant-like activity and computational exploration of identified bioactive compounds in Talinum triangulare (Jacq.) aqueous extract as potential cholinesterase inhibitors.

BACKGROUND: Recent reports have highlighted the significance of plant bioactive components in drug development targeting neurodegenerative disorders such as Alzheimer's disease (AD). Thus, the current study assessed antioxidant activity and enzyme inhibitory activity of the aqueous extract of Talinum triangulare leave (AETt) as well as molecular docking/simulation of the identified phytonutrients against human cholinesterase activities. METHODS: In vitro assays were carried out to assess the 2,2- azinobis (3-ethyl-benzothiazoline-6-sulfonic acid) (ABTS) cation radicals and cholinesterase inhibitory activities of AETt using standard protocols. High performance liquid chromatography coupled with diode-array detection (HPLC-DAD) was employed to identify compounds in AETt. Also, for computational analysis, identified bioactive compounds from AETt were docked using Schrodinger's GLIDE against human cholinesterase obtained from the protein data bank ( https://www.rcsb.org/ ). RESULTS: The results revealed that AETt exhibited a significant concentration-dependent inhibition against ABTS cation radicals (IC50 = 308.26 ± 4.36 µg/ml) with butylated hydroxytoluene (BHT) as the reference. Similarly, AETt demonstrated a significant inhibition against acetylcholinesterase (AChE, IC50 = 326.49 ± 2.01 µg/ml) and butyrylcholinesterase (BChE, IC50 = 219.86 ± 4.13 µg/ml) activities with galanthamine as the control. Molecular docking and simulation analyses revealed rutin and quercetin as potential hits from AETt, having showed strong binding energies for both the AChE and BChE. In addition, these findings were substantiated by analyses, including radius of gyration, root mean square fluctuation, root mean square deviation, as well as mode similarity and principal component analyses. CONCLUSION: Overall, this study offers valuable insights into the interactions and dynamics of protein-ligand complexes, offering a basis for further drug development targeting these proteins in AD.

Impact of body mass index on the severity of bexarotene-associated hypertriglyceridemia: A post hoc analysis of an open-labeled clinical study of combined bexarotene and phototherapy in Japanese patients with cutaneous T-cell lymphoma.

Bexarotene is an effective oral drug for the treatment of cutaneous T-cell lymphoma, but careful management is required due to its various side effects. In particular, hypertriglyceridemia often requires a reduction or even suspension of bexarotene therapy. The risk factors of bexarotene-associated severe hypertriglyceridemia are not clear. Here, we conducted a post hoc analysis of the data from our previous clinical trial, which confirmed the efficacy and safety of combined bexarotene and phototherapy, to evaluate the effect of body mass index on bexarotene-associated hypertriglyceridemia. Twenty-five subjects were divided into two subgroups: normal and underweight (body mass index [BMI] <25 kg/m2 group) and overweight and obese (BMI ≥25 kg/m2 group) patients. The overall incidence of hypertriglyceridemia was 81.3% (13/16) in the BMI <25 kg/m2 group and 88.9% (8/9) in the BMI ≥25 kg/m2 group. The incidence of grade ≥3 hypertriglyceridemia (≥500 mg/dL) was 7.7% (1/13) in the BMI <25 kg/m2 group and 7/8 (87.5%) in the BMI ≥25 kg/m2 group (P < 0.001). Consequently, dose reduction in the BMI ≥25 kg/m2 group was larger than that in the BMI <25 kg/m2 group. The bexarotene-induced change in the serum triglyceride concentration was significantly increased in cutaneous T-cell lymphoma patients with a higher body mass index (ρ = 0.508, P = 0.009). The area under the curve was 0.886 (95% confidence interval 0.748-1.000, P = 0.002). With a body mass index cut-off of 24.85 kg/m2 , the sensitivity and specificity for identifying grade ≥3 hypertriglyceridemia were 0.875 and 0.882, respectively. The present findings suggest that BMI ≥25 kg/m2 is a risk factor for bexarotene-associated severe hypertriglyceridemia, therefore overweight and obese patients treated with bexarotene should receive lipid-lowering drugs prophylactically. Further studies for optimizing the initial bexarotene dose in such patients are required.

Effects of Green Tea Marinade in the Bioaccessibility of Tonalide and Benzophenone 3 in Cooked European Seabass.

Fish consumption is an indicator of human exposure to personal care products (PCP) such as tonalide (AHTN) and benzophenone 3 (BP3). Although most fish consumed is cooked, the impact of cooking procedures on PCP levels is difficult to evaluate. Hence, the aim of this work was to provide thorough information on the stability and bioaccessibility of AHTN and BP3 upon cooking and in vitro digestion. A green tea (Camellia sinensis) marinade, rich in polyphenol, was used as mitigating strategy to reduce these contaminants. Roasting and frying reduced AHTN and BP3 levels in European seabass (Dicentrarchus labrax) spiked samples. Additionally, the green tea marinade promoted a reduction of up to 47% AHTN and 35% BP3. Bioaccessibility of AHTN was higher (up to 45%), and increased with the use of green tea marinades. BP3 showed a bioaccessibility below 19% in all cooked samples. Overall, a decrease in PCP levels was observed after cooking; this decrease was even more pronounced when marination was previously used. However, this decrease is cancelled out by the fact that the bioaccessible fraction of the contaminants increases in an inverse way; therefore, none of these processes can be considered a mitigating alternative.

Inhibition of Integrin αVß3 Signaling Improves the Antineoplastic Effect of Bexarotene in Cutaneous T-Cell Lymphoma.

Bexarotene is a specific retinoid X receptor agonist that has been used for the treatment of cutaneous T-cell lymphoma (CTCL). Because bexarotene causes hypothyroidism, it requires the administration of levothyroxine. However, levothyroxine, in addition to its ubiquitous nuclear receptors, can activate the αVß3 integrin that is overexpressed in CTCL, potentially interfering the antineoplastic effect of bexarotene. We thus investigated the biological effect of levothyroxine in relation to bexarotene treatment. Although in isolated CTCL cells levothyroxine decreased, in an αVß3-dependent manner, the antineoplastic effect of bexarotene, levothyroxine supplementation in preclinical models was necessary to avoid suppression of lymphoma immunity. Accordingly, selective genetic and pharmacologic inhibition of integrin αVß3 improved the antineoplastic effect of bexarotene plus levothyroxine replacement while maintaining lymphoma immunity. Our results provide a mechanistic rationale for clinical testing of integrin αVß3 inhibitors as part of CTCL regimens based on bexarotene administration. TEASER: Inhibiting αVß3 integrin improves the antineoplastic effect of bexarotene while maintaining lymphoma immunity.

Aristolone in Nardostachys jatamansi DC. induces mesenteric vasodilation and ameliorates hypertension via activation of the KATP channel and PDK1-Akt-eNOS pathway.

BACKGROUND: Nardostachys jatamansi DC. is a common medicinal herb used to treat cardiovascular diseases, particularly hypertension. Previously, our lab characterized the chemical compounds of N. jatamansi. However, the bioactive compounds of N. jatamansi and their mechanisms of action on blood pressure and blood vessels are unknown. PURPOSE: The vasorelaxant effects of the methanolic extract (MeOH ext.) of the roots and rhizomes of N. jatamansi, its main compounds, and their underlying mode of action, were investigated. METHODS: The main compounds of N. jatamansi were isolated and identified using UHPLC-TOF MS. The antihypertensive effect of N. jatamansi extracts and (-)-aristolone were determined using spontaneously hypertensive rats. The extracts, fractions, and compounds were also evaluated for their vasorelaxant effects on U46619 contractile responses in isolated thoracic aortic and mesenteric arterial rings. The endothelial-dependent relaxation, as well as the regulatory pathways and targets of (-)-aristolone, were studied in-vitro and ex-vivo. Molecular docking and biophysical characterization (Surface plasmon resonance) studies were utilized to investigate the molecular interaction between (-)-aristolone and the target protein. RESULTS: MeOH ext. (200 mg/kg) reduces the systolic and diastolic blood pressure in spontaneously hypertensive rats. MeOH ext. and its ethyl acetate fraction (EtOAc Fr.), but not the H2O fraction, had a significant relaxing effect on the thoracic aorta. (-)-aristolone and kanshone H from EtOAc Fr. induced vasorelaxation of the thoracic aorta and mesenteric artery. In human umbilical vein endothelial cells, (-)-aristolone treatment upregulated phosphorylation of Akt (T308) and eNOS. Molecular docking and surface plasmon resonance experiments revealed an interaction between (-)-aristolone and phosphoinositide-dependent protein kinase 1 (PDK1), an upstream protein kinase that phosphorylates Akt at T308. Treatment with PDK1 inhibitor PHT-427 and eNOS inhibitor L-NAME consistently inhibited (-)-aristolone-induced vasorelaxation. In addition, KATP channel inhibitor glibenclamide dramatically inhibited the vasorelaxant effects of (-)-aristolone and kanshone H in the endothelium-denuded thoracic aorta. Finally, (-)-aristolone lowers hypertensive rats' systolic and diastolic blood pressure. CONCLUSIONS: The extracts of N. jatamansi promote vasorelaxation and alleviate hypertension. The essential chemicals responsible for producing vasorelaxation effects are (-)-aristolone and kanshone H, which activate the PDK1-Akt-eNOS-NO relaxing pathway and stimulate the opening of the KATP channel. These findings point to N. jatamansi and aristolone as possible antihypertensive agents.

Agathadiol, a labdane diterpenoid from juniper berries, is a positive allosteric modulator of CB1R.

The neutral fraction of a juniper (Juniperus communis L.) berries acetone extract could positively modulate the activity of type 1 - cannabinoid receptor (CB1R). Bioactivity-directed fractionation identified the labdane diterpenoid agathadiol (4) as a positive allosteric modulator of CB1R, while closely related analogues were inactive. Agathadiol (4) is a minor constituent of juniper, but could be more conveniently obtained by semisynthesis from agathic acid (8), a major constituent of Manila copal.

Selective estrogen receptor modulator lasofoxifene suppresses spondyloarthritis manifestation and affects characteristics of gut microbiota in zymosan-induced SKG mice.

Ankylosing spondylitis is a male-predominant disease and previous study revealed that estrogens have an anti-inflammatory effect on the spondyloarthritis (SpA) manifestations in zymosan-induced SKG mice. This study aimed to evaluate the effect of selective estrogen receptor modulator (SERM) lasofoxifene (Laso) on disease activity of SpA. Mice were randomized into zymosan-treated, zymosan + 17ß-estradiol (E2)-treated, and zymosan + Laso-treated groups. Arthritis was assessed by 18F-fluorodeoxyglucose (18F-FDG) small-animal positron emission tomography/computed tomography and bone mineral density (BMD) was measured. Fecal samples were collected and 16S ribosomal RNA gene sequencing was used to determine gut microbiota differences. Both zymosan + E2-treated mice and zymosan + Laso-treated mice showed lower arthritis clinical scores and lower 18F-FDG uptake than zymosan-treated mice. BMD was significantly higher in zymosan + E2-treated mice and zymosan + Laso-treated mice than zymosan-treated mice, respectively. Fecal calprotectin levels were significantly elevated at 8 weeks after zymosan injection in zymosan-treated mice, but it was not significantly changed in zymosan + E2-treated mice and zymosan + Laso-treated mice. Gut microbiota diversity of zymosan-treated mice was significantly different from zymosan + E2-treated mice and zymosan + Laso-treated mice, respectively. There was no significant difference in gut microbiota diversity between zymosan + E2-treated mice and zymosan + Laso -treated mice. Laso inhibited joint inflammation and enhanced BMD in SKG mice, a model of SpA. Laso also affected the composition and biodiversity of gut microbiota. This study provides new knowledge regarding that selected SpA patients could benefit from SERM treatment.

Dibenzoate esters of cis-tetralin-2,3-diol as analogs of (-)-epigallocatechin gallate: synthesis and crystal structure of anticancer drug candidates.

(-)-Epigallocatechin gallate (EGCG), the main component of green tea extract, displays multiple biological activities. However, it cannot be used as a drug due to its low cellular absorption, instability and metabolic degradation. Therefore, there is a need to provide analogs that can overcome the limitations of EGCG. In this work, six synthetic analogs of EGCG sharing a common tetralindiol dibenzoate core were synthesized and fully characterized by 1H NMR, 13C NMR, HRMS and IR spectroscopies, and X-ray crystallography. These are (2R,3S)-1,2,3,4-tetrahydronaphthalene-2,3-diyl bis[3,4,5-tris(benzyloxy)benzoate], C66H56O10, and the analogous esters bis(3,4,5-trimethoxybenzoate), C30H32O10, bis(3,4,5-trifluorobenzoate), C24H14F6O4, bis[4-(benzyloxy)benzoate], C38H32O6, bis(4-methoxybenzoate), C26H24O6, and bis(2,4,6-trifluorobenzoate), C24H14F6O4. Structural analysis revealed that the molecular shapes of these dibenzoate esters of tetralindiol are significantly different from that of previously reported dimandelate esters or monobenzoate esters, as the acid moieties extend far from the bicyclic system without folding back over the tetralin fragment. Compounds with small fluorine substituents take a V-shape, whereas larger methoxy and benzyloxy groups determine the formation of an L-shape or a cavity. Intermolecular interactions are dominated by π-π stacking and C-H...π interactions involving the arene rings in the benzoate fragment and the arene ring in the tetrahydronaphthalene moiety. All six crystal structures are determined in centrosymmetric space groups (either P-1, P21/n, C2/c or I2/a).

Nanoemulsions with oleoresin of Copaifera reticulata (Leguminosae) improve anthelmintic efficacy in the control of monogenean parasites when compared to oleoresin without nanoformulation.

This study compared the in vitro anthelmintic activity of Copaifera reticulata oleoresin (200, 400, 600, 800 and 1,000 mg/L) and of nanoemulsions prepared with this oleoresin (50, 100, 150, 200 and 250 mg/L) against monogeneans on the gills of Colossoma macropomum. The major compounds present in the oleoresin of C. reticulata were γ-macrocarpene (14.2%), α-bergamotene (13.6%), ß-selinene (13.4%) and ß-caryophyllene (11.7%). All concentrations of the nanoemulsion and the oleoresin without nanoformulation showed anthelmintic efficacy against monogeneans, and higher concentrations led to more rapid parasite mortality. Structural damages to the tegument of the parasites exposed to C. reticulata oleoresin were observed with scanning electron microscopy. At two hours of exposure, fish showed 100% tolerance to all nanoemulsion concentrations used in the in vitro assays, whereas 100% mortality was shown in the fish exposed to the oleoresin without nanoformulation after one hour. The results of this study suggest that nanoemulsions with oleoresin of C. reticulata have advantages in the control and treatment of monogenean infections in C. macropomum when compared to the oleoresin without nanoformulation. In addition, since nanoemulsions with the C. reticulata oleoresin are safe to control monogeneans, the efficacy of these nanoformulations may be assayed in therapeutic baths to treat C. macropomum infected by monogeneans.

Anticancer activity and molecular mechanisms of α-conidendrin, a polyphenolic compound present in Taxus yunnanensis, on human breast cancer cell lines.

α-Conidendrin is a polyphenolic compound found mainly in Taxus yunnanensis, as the source of chemotherapy drug paclitaxel, which has been used in traditional medicine for treatment of cancer. This study aimed to investigate the anticancer activity and molecular mechanisms of α-conidendrin on breast cancer cell lines. The results of the present study show that α-conidendrin possesses potent antiproliferative effects on breast cancer cell lines MCF-7 and MDA-MB-231. α-Conidendrin significantly induced apoptosis in breast cancer cells via reactive oxygen species generation, upregulation of p53 and Bax, downregulation of Bcl-2, depolarization of mitochondrial membrane potential (MMP), release of cytochrome c from mitochondria, and activation of caspases-3 and -9. α-Conidendrin remarkably inhibited the proliferation of breast cancer cells through induction of cell cycle arrest by upregulating p53 and p21 and downregulating cyclin D1 and CDK4. Unlike breast cancer cells, the antiproliferative effect of α-conidendrin on human foreskin fibroblast cells (normal cells) was very small. In normal cells, reactive oxygen species levels, loss of MMP, release of cytochrome c, mRNA expression of p53, p21, cyclin D1, CDK4, Bax, and Bcl-2 as well as mRNA expression and activity of caspases-3 and -9 were significantly less affected by α-conidendrin compared with cancer cells. These results suggest that α-conidendrin can be a promising agent for treatment of breast cancer with little or no toxicity against normal cells.

Metabolite profiling of Andrographis paniculata (Burm. f.) Nees. young and mature leaves at different harvest ages using 1H NMR-based metabolomics approach.

Andrographis paniculata (Burm. F.) Nees. is considered as the herb of the future due to its precious chemical compounds, andrographolide (ANDRO), neoandrographolide (NAG) and 14-deoxyandrographolide (DAG). This study aims to profile the metabolites in young and mature leaf at six different harvest ages using 1HNMR-based metabolomics combined with multivariate data analysis. Principal component analysis (PCA) indicated noticeable and clear discrimination between young and mature leaves. A comparison of the leaves stage indicated that young leaves were separated from mature leaves due to its larger quantity of ANDRO, NAG, DAG, glucose and sucrose. These similar metabolites are also responsible for the PCA separation into five clusters representing the harvest age at 14, 16, 18, 20, 22 weeks of leaves extract. Loading plots revealed that most of the ANDRO and NAG signals were present when the plant reached at the pre-flowering stage or 18 weeks after sowing (WAS). As a conclusion, A. paniculata young leaves at pre-flowering harvest age were found to be richer in ANDRO, NAG and DAG compared to mature leaves while glucose and choline increased with harvest age. Therefore, young leaves of A. paniculata should be harvested at 18 WAS in order to produce superior quality plant extracts for further applications by the herbal, nutraceutical and pharmaceutical industries.

High Concentration of C5a-Induced Mitochondria-Dependent Apoptosis in Murine Kidney Endothelial Cells.

Patients with a relapse of idiopathic nephrotic syndrome have significantly increased levels of serum complement component 5a (C5a), and proteinuria has been noted in mice treated with C5a via changes in permeability of kidney endothelial cells (KECs) in established animal models. However, the apoptosis of KECs treated with high concentrations of C5a has also been observed. As mitochondrial damage is known to be important in cell apoptosis, the aim of this study was to examine the association between C5a-induced mouse KEC apoptosis and mitochondrial damage. Mouse KECs were isolated and treated with different concentrations of C5a. Cell viability assays showed that a high-concentration mouse recombinant protein C5a (rmC5a) treatment reduced mouse KEC growth. Cell cycle phase analysis, including apoptosis (sub-G1 phase) showed an increased percentage of the subG1 phase with a high-concentration rmC5a treatment. Cytochrome c and caspase 3/9 activities were significantly induced in the mouse KECs after a high-dose rmC5a (50 ng/mL) treatment, and this was rescued by pretreatment with the C5a receptor (C5aR) inhibitor (W-54011) and N-acetylcysteine (NAC). Reactive oxygen species (ROS) formation was detected in C5a-treated mouse KECs; however, W-54011 or NAC pretreatment inhibited high-dose rmC5a-induced ROS formation and also reduced cytochrome c release, apoptotic cell formation, and apoptotic DNA fragmentation. These factors determined the apoptosis of mouse KECs treated with high-dose C5a through C5aR and subsequently led to apoptosis via ROS regeneration and cytochrome c release. The results showed that high concentrations of C5a induced mouse KEC apoptosis via a C5aR/ROS/mitochondria-dependent pathway. These findings may shed light on the potential mechanism of glomerular sclerosis, a process in idiopathic nephrotic syndrome causing renal function impairment.

Methods to Assess Activity and Potency of Rexinoids Using Rapid Luciferase-Based Assays: A Case Study with NEt-TMN.

This chapter outlines the materials, methods, and procedures for the in vitro biological evaluation of retinoid-X-receptor (RXR) agonists including 6-(ethyl(5,5,8,8-tetramethyl-5,6,7,8-tetrahydronaphthalen-2-yl)amino)nicotinic acid (NEt-TMN), as well as several NEt-TMN analog compounds recently reported by our group. These methods have general applicability beyond this NEt-TMN case study, and can be employed to characterize and biologically evaluate other putative RXR agonists (rexinoids), and benchmarked against perhaps the most common rexinoid known as bexarotene (Bex), a drug awarded FDA approval for the treatment of cutaneous T-cell lymphoma in 1999 but that is also prescribed for non-small cell lung cancer and continues to be explored in multiple human cancer types. The side-effect profile of Bex treatment includes hypothyroidism and hypertriglyceridemia arising from the inhibition or activation of additional nuclear receptors that partner with RXR. Because rexinoids often exhibit selectivity for RXR activation, versus activating the retinoic-acid-receptor (RAR), rexinoid treatment avoids the cutaneous toxicity commonly associated as a side effect with retinoids. There are many examples of other potent rexinoids, where biological evaluation has contributed useful insight into qSAR studies on these compounds, often also benchmarked to Bex, as potential treatments for cancer. Because of differential pleiotropy in other pathways, even closely related rexinoids display unique side-effect and activity profiles. Notable examples of potent rexinoids in addition to Bex and NEt-TMN include CD3254, LGD100268, and 9-cis-UAB30. Indeed, the methods described herein to evaluate NEt-TMN and analogous rexinoids are generally applicable to a wider variety of potent, moderate, and even weak RXR ligands.In terms of in vitro biological evaluation, methods for a rapid and preliminary assessment of rexinoid activity are described by employing a biologically relevant, RXR-responsive element (RXRE)-mediated transcription assay in mammalian cells. In addition, a second, more sensitive assay is also detailed that utilizes activation of RXR-RXR homodimers in the context of a mammalian two-hybrid (M2H) luciferase assay. Methods for applying the M2H assay at different rexinoid concentrations are further described for the determination of EC50 values for rexinoids from dose-response curves.

Quality Evaluation of Andrographis paniculata Capsules Based on Rapid and Accurate LC-ESI-MS/MS Assay of Three Diterpenoids.

Andrographis paniculata is an important traditional Chinese herbal material widely used for treatment of fever and diarrhea. The three diterpenoids [andrographolide (AP), dehydroandrographolide (DP) and neoandrographolide (NP)] in A. paniculata were characterized as the key active components for its extractive and related medicinal preparations. A rapid and accurate quantitative analysis of these three diterpenoids in A. paniculata capsules was accomplished by means of a valid and reliable high-performance liquid chromatography with tandem mass spectrometric (HPLC-MS/MS) analytical method. An optimized gradient elution method, which under the mobile phase consisting of methanol and water, was established successfully. An XDB C18 column (3.5 µm, 2.1 mm × 50 mm) was successfully applied to separate these three diterpenoids at the flow rate of 0.55 mL/min. A triple quadrupole mass spectrometer with multiple reaction monitoring (MRM) mode using an electrospray ionization source was served for analytical detection. The linearity, specificity, accuracy, recovery, precision, stability and repeatability were demonstrated to fully confirm the validity of this study. Good linear regression relationships for the three analytes were obtained over the range of 0.50-1000 ng/mL. Intra-day and inter-day precisions were evaluated with relative standard deviation < 7.62%, and the extraction recovery varied from 93.8% to 102.0%. The rapid and accurate HPLC-MS/MS analytical method was developed for the quality evaluation of A. paniculata capsules successfully. The content of AP, DP and NP obtained were 3.90-4.08, 4.77-5.04, 4.32-4.48 mg/g in A. paniculata capsules, respectively. Moreover, it is expected that this analytical technique can be applied to detect three effective components rapidly and accurately in other different medicinal preparations of A. paniculata as a method for their quality control.

Potential anti-neuroblastoma agents from Juniperus oblonga.

Neuroblastoma (NB) is a neuroendocrine tumor derived from neural crest cells. Approximately 90% of cases occur in children less than 5 years old. The amplification of MYCN correlates with high-risk neuroblastoma and patients with MYCN amplified showed poorer prognosis than those without MYCN amplification. In this study, three compounds isolated from Juniperus oblonga showed anti-proliferative activity against NB cell lines with and without tetracycline inducible MYCN over-expression which were identified as (-)-deoxypodophyllotoxin (1), (-)-matairesinol (2) and (+)-isocupressic acid (3). The effects of compounds 2 and 3 in NB cells included a decrease in NB cell viability and induction of apoptosis. Compound 1 was more effective in NB cells over-expressing MycN. Compound 1 also showed almost 2-fold induction of intracellular free calcium levels in M2(+) cells, which may indicate a different mechanism of action for this compound. Cytotoxicity studies against the human embryonic kidney cell (HEK-293) showed compounds 1, 2 and 3 were ineffective in the non-cancer cells at concentrations approximating their IC50 against the NB cell lines. These results may lead to safer and more effective treatment options for NB patients especially for those with high-risk NB.

Nirogacestat suppresses RANKL-Induced osteoclast formation in vitro and attenuates LPS-Induced bone resorption in vivo.

Bone resorption, initiated by osteoclasts (OCs), plays an essential role in bone homeostasis. The abnormalities of bone resorption may induce a series of diseases, including osteoarthritis, osteoporosis and aseptic peri-implant loosening. Nirogacestat (PF-03084014, PF), a novel gamma-secretase inhibitor, has been used in phase II clinical trial for treatment of desmoid tumor. However, whether it has the therapeutic effect on abnormal bone resorption remains to be evaluated. In this study, we investigated the role of PF in the regulation of receptor activator of nuclear factor-kB ligand (RANKL)-induced osteoclastogenesis in vitro, and the lipopolysaccharide (LPS)-induced bone resorption in vivo. It was found that PF could suppress the formation of osteoclasts from bone marrow macrophages (BMMs) without causing cytotoxicity, inhibit bone resorption and downregulate the mRNA level of osteoclast-specific markers, including calcitonin receptor (CTR), tartrate resistant acid phosphatase (TRAP), cathepsin K (CTSK), dendritic cell-specific transmembrane protein (Dc-stamp), Atp6v0d2 (V-ATPase d2) and nuclear factor of activated T-cells cytoplasmic 1 (NFATc1). Furthermore, Notch2 signaling, as well as RANKL-induced AKT signaling was significantly inhibited in BMMs. Consistent with in vitro observation, we found that PF greatly ameliorated LPS-induced bone resorption. Taken together, our study demonstrated that PF has a great potential to be used in management of osteolytic diseases.

Biodegradation of Tetralin: Genomics, Gene Function and Regulation.

Tetralin (1,2,3,4-tetrahydonaphthalene) is a recalcitrant compound that consists of an aromatic and an alicyclic ring. It is found in crude oils, produced industrially from naphthalene or anthracene, and widely used as an organic solvent. Its toxicity is due to the alteration of biological membranes by its hydrophobic character and to the formation of toxic hydroperoxides. Two unrelated bacteria, Sphingopyxis granuli strain TFA and Rhodococcus sp. strain TFB were isolated from the same niche as able to grow on tetralin as the sole source of carbon and energy. In this review, we provide an overview of current knowledge on tetralin catabolism at biochemical, genetic and regulatory levels in both strains. Although they share the same biodegradation strategy and enzymatic activities, no evidences of horizontal gene transfer between both bacteria have been found. Moreover, the regulatory elements that control the expression of the gene clusters are completely different in each strain. A special consideration is given to the complex regulation discovered in TFA since three regulatory systems, one of them involving an unprecedented communication between the catabolic pathway and the regulatory elements, act together at transcriptional and posttranscriptional levels to optimize tetralin biodegradation gene expression to the environmental conditions.

Melatonin MT1 and MT2 Receptors Exhibit Distinct Effects in the Modulation of Body Temperature across the Light/Dark Cycle.

Melatonin (MLT) is a neurohormone that regulates many physiological functions including sleep, pain, thermoregulation, and circadian rhythms. MLT acts mainly through two G-protein-coupled receptors named MT1 and MT2, but also through an MLT type-3 receptor (MT3). However, the role of MLT receptor subtypes in thermoregulation is still unknown. We have thus investigated the effects of selective and non-selective MLT receptor agonists/antagonists on body temperature (Tb) in rats across the 12/12-h light-dark cycle. Rectal temperature was measured every 15 min from 4:00 a.m. to 9:30 a.m. and from 4:00 p.m. to 9:30 p.m., following subcutaneous injection of each compound at either 5:00 a.m. or 5:00 p.m. MLT (40 mg/kg) had no effect when injected at 5 a.m., whereas it decreased Tb during the light phase only when injected at 5:00 p.m. This effect was blocked by the selective MT2 receptor antagonist 4P-PDOT and the non-selective MT1/MT2 receptor antagonist, luzindole, but not by the α1/MT3 receptors antagonist prazosin. However, unlike MLT, neither the selective MT1 receptor partial agonist UCM871 (14 mg/kg) nor the selective MT2 partial agonist UCM924 (40 mg/kg) altered Tb during the light phase. In contrast, UCM871 injected at 5:00 p.m. increased Tb at the beginning of the dark phase, whereas UCM924 injected at 5:00 a.m. decreased Tb at the end of the dark phase. These effects were blocked by luzindole and 4P-PDOT, respectively. The MT3 receptor agonist GR135531 (10 mg/kg) did not affect Tb. These data suggest that the simultaneous activation of both MT1 and MT2 receptors is necessary to regulate Tb during the light phase, whereas in a complex but yet unknown manner, they regulate Tb differently during the dark phase. Overall, MT1 and MT2 receptors display complementary but also distinct roles in modulating circadian fluctuations of Tb.

Sigma-2 receptor/TMEM97 agonist PB221 as an alternative drug for brain tumor.

BACKGROUND: There are limited effective drugs that can reach the brain to target brain tumors, in particular glioblastoma, which is one of the most difficult cancers to be cured from. Because the overexpression of the sigma-2 receptor is frequently reported in glioma clinical samples and associated with poor prognosis and malignancy, we herein studied the anti-tumor effect of the sigma-2 receptor agonist PB221 (4-cyclohexyl-1-[3-(5-methoxy-1,2,3,4-tetrahydronaphthalen-1-yl)propyl]piperidine) on an anaplastic astrocytoma tumor model based on previous encouraging results in pancreatic cancer and neuroblastoma SK-N-SH cells. METHODS: The expression of the sigma-2 receptor, transmembrane protein 97 (TMEM97), in ALTS1C1 and UN-KC6141 cell lines was measured by RT-PCR and quantitative RT-PCR. The binding of sigma-2 receptor fluorescent ligands PB385 (6-[5-[3-(4-cyclohexylpiperazin-1-yl)propyl]-5,6,7,8-tetrahydronaphthalen-5-yloxy]-N-(7-nitro-2,1,3-benzoxadiazol-4-yl)hexanamine) and NO1 (2-{6-[2-(3-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)propyl)-3,4-dihydroisoquinolin-1(2H)-one-5-yloxy]hexyl}-5-(dimethylamino)isoindoline-1,3-dione) was examined by flow cytometry and the fluorescent plate reader. The antitumor activity of PB221 was initially examined in the murine brain tumor cell line ALTS1C1 and then in the murine pancreatic cell line UN-KC6141. The potential therapeutic efficacy of PB221 for murine brain tumors was examined by in vitro migration and invasion assays and in vivo ectopic and orthotopic ALTS1C1 tumor models. RESULTS: The IC50 of PB221 for ALTS1C1 and UN-KC6141 cell lines was 10.61 ± 0.96 and 13.13 ± 1.15 µM, respectively. A low dose of PB221 (1 µM) significantly repressed the migration and invasion of ALTS1C1 cells, and a high dose of PB221 (20 µM) resulted in the apoptotic cell death of ALTS1C1 cells. These effects were reduced by the lipid antioxidant α-tocopherol, but not by the hydrophilic N-acetylcysteine, suggesting mitochondrial oxidative stress is involved. The in vivo study revealed that PB221 effectively retarded tumor growth to 36% of the control tumor volume in the ectopic intramuscular tumor model and increased the overall survival time by 20% (from 26 to 31 days) in the orthotopic intracerebral tumor model. CONCLUSIONS: This study demonstrates that the sigma-2 receptor agonist PB221 has the potential to be an alternative chemotherapeutic drug for brain tumors with comparable side effects as the current standard-of-care drug, temozolomide.

The genome of the medicinal plant Andrographis paniculata provides insight into the biosynthesis of the bioactive diterpenoid neoandrographolide.

Andrographis paniculata is a herbaceous dicot plant widely used for its anti-inflammatory and anti-viral properties across its distribution in China, India and other Southeast Asian countries. A. paniculata was used as a crucial therapeutic treatment during the influenza epidemic of 1919 in India, and is still used for the treatment of infectious disease in China. A. paniculata produces large quantities of the anti-inflammatory diterpenoid lactones andrographolide and neoandrographolide, and their analogs, which are touted to be the next generation of natural anti-inflammatory medicines for lung diseases, hepatitis, neurodegenerative disorders, autoimmune disorders and inflammatory skin diseases. Here, we report a chromosome-scale A. paniculata genome sequence of 269 Mb that was assembled by Illumina short reads, PacBio long reads and high-confidence (Hi-C) data. Gene annotation predicted 25 428 protein-coding genes. In order to decipher the genetic underpinning of diterpenoid biosynthesis, transcriptome data from seedlings elicited with methyl jasmonate were also obtained, which enabled the identification of genes encoding diterpenoid synthases, cytochrome P450 monooxygenases, 2-oxoglutarate-dependent dioxygenases and UDP-dependent glycosyltransferases potentially involved in diterpenoid lactone biosynthesis. We further carried out functional characterization of pairs of class-I and -II diterpene synthases, revealing the ability to produce diversified labdane-related diterpene scaffolds. In addition, a glycosyltransferase able to catalyze O-linked glucosylation of andrograpanin, yielding the major active product neoandrographolide, was also identified. Thus, our results demonstrate the utility of the combined genomic and transcriptomic data set generated here for the investigation of the production of the bioactive diterpenoid lactone constituents of the important medicinal herb A. paniculata.