Influence of co-solvents on properties of terpene-based eutectic mixtures: Implications for skin delivery.

Terpene-based eutectic mixtures (EMs) are attractive platforms for transdermal delivery due to their solubilizing potential and ability to alter the barrier function of the stratum corneum (SC). Despite this, little is known about the effect of diluting EMs with co-solvents (CSs) on their solubility- and permeation-enhancing properties. Furthermore, insufficient attention has been paid to comparing these platforms with traditional solvents, such as propylene glycol (PG) or ethanol (EtOH). To address this gap, the present study investigates the impact of the CS content in EM:CS blends on the transdermal delivery of clotrimazole (CLOT). Two CSs, PG and EtOH, and two terpene-based EMs, menthol:thymol and thymol:ß-citronellol, were used. Each of the EMs was investigated at two different molar ratios between the terpenes, with one being their eutectic point, to explore its potential benefit for skin permeation. At each step, properties of the blends were compared with those of pure CSs. The EM:CS blends showed a better solubilizing potential for CLOT than EMs or CSs on their own. A higher content of CSs in the blends resulted in a higher skin permeation and retention of CLOT, and a lower degree of disarrangement of the SC structure. Furthermore, the blends of EMs at their EPs led to overall poorer permeation profiles, implying that the permeation rate is more affected by the properties of the individual terpenes than by the specific ratio at the eutectic point between them. In conclusion, addition of CSs to the EMs promotes permeation and retention of CLOT, while reducing the skin impairment caused by the terpenes.

Medicinal cannabis tea contains variable doses of cannabinoids and no terpenes.

Tea is a recommended way of administration of prescribed cannabis plant products in Denmark. We aimed to investigate the cannabinoid and terpene doses contained in different teas. We analysed tetrahydrocannabinol (THC), tetrahydrocannabinolic acid (THCA), cannabidiol (CBD), cannabidiolic acid (CBDA), and terpene concentrations in three repeated preparations of each type of tea, and in plant material. In standard tea, concentrations of THC were [median (min-max)] 9.5 (2.3-15), 19 (13-34), and 36 (26-57) µg/mL for products with a labelled content of 6.3%, 14%, and 22% total THC (THC + THCA), respectively. The CBD concentration in tea from a product labelled with 8% total CBD (CBD + CBDA) was 7.5 (1.9-10) µg/mL. Based on this, the recommended starting amount of 0.2 L of the different teas would contain between 0.46 and 11.3 mg THC, and 0.38 to 2.0 mg CBD. Adding creamer before, but not after boiling, increased the THC and CBD concentration 2.3-4.4 and 2.1-fold, respectively. Terpenes were detected in plant material, but not in tea. The study elucidates THC and CBD doses in different teas, which may assist the clinician's choice of cannabis product. Moreover, it underscores the need for caution as administration as tea can result in exposure to different doses, even when the same cannabis product is used.

Encapsulation of Citrus sinensis essential oil and R-limonene in lipid nanocarriers: A potential strategy for the treatment of leishmaniasis.

Leishmaniases, a group of neglected tropical diseases caused by an intracellular parasite of the genus Leishmania, have significant impacts on global health. Current treatment options are limited due to drug resistance, toxicity, and high cost. This study aimed to develop nanostructured lipid carriers (NLCs) for delivering Citrus sinensis essential oil (CSEO) and its main constituent, R-limonene, against leishmaniasis. The influence of surface-modified NLCs using chitosan was also examined. The NLCs were prepared using a warm microemulsion method, and surface modification with chitosan was achieved through electrostatic interaction. These nanocarriers were characterized by differential scanning calorimetry (DSC), X-ray diffraction (XRD), transmission electron microscopy, and dynamic light scattering (DLS). In vitro cytotoxicity was assessed in L929 and RAW 264.7 cells, and leishmanicidal activity was evaluated against promastigote and amastigote forms. The NLCs were spherical, with particle sizes ranging from 97.9 nm to 111.3 nm. Chitosan-coated NLCs had a positive surface charge, with zeta potential values ranging from 45.8 mV to 59.0 mV. Exposure of L929 cells to NLCs resulted in over 70 % cell viability. Conversely, surface modification significantly reduced the viability of promastigotes (93 %) compared to free compounds. Moreover, chitosan-coated NLCs presented a better IC50 against the amastigote forms than uncoated NLCs. Taken together, these findings demonstrate the feasibility of using NLCs to overcome the limitations of current leishmaniasis treatments, warranting further research.

HSPiP, Computational, and Thermodynamic Model-Based Optimized Solvents for Subcutaneous Delivery of Tolterodine Tartrate and GastroPlus­Based In Vivo Prediction in Humans: Part II.

In part I, we reported Hansen solubility parameters (HSP, HSPiP program), experimental solubility at varied temperatures for TOTA delivery. Here, we studied dose volume selection, stability, pH, osmolality, dispersion, clarity, and viscosity of the explored combinations (I-VI). Ex vivo permeation and deposition studies were performed to observe relative diffusion rate from the injected site in rat skin. Confocal laser scanning microscopy (CLSM) study was conducted to support ex vivo findings. Moreover, GastroPlus predicted in vivo parameters in humans and the impact of various critical factors on pharmacokinetic parameters (PK). Immediate release product (IR) contained 60% of PEG400 whereas controlled release formulation (CR) contained PEG400 (60%), water (10%) and d-limonene (30%) to deliver 2 mg of TOTA. GastroPlus predicted the plasma drug concentration of weakly basic TOTA as function of pH (from pH 2.0 to 9). The cumulative drug permeation and drug deposition were found to be in the order as B-VI˃ C-VI˃A-VI across rat skin. This finding was further supported with CLSM. Moreover, IR and CR were predicted to achieve Cmax of 0.0038 µg/ mL and 0.00023 µg/mL, respectively, after sub-Q delivery. Added limonene in CR extended the plasma drug concentration over period of 12 h as predicted in GastroPlus. Parameters sensitivity analysis (PSA) assessment predicted that sub-Q blood flow rate is the only factor affecting PK parameters in IR formulation whereas this was insignificant for CR. Thus, sub-Q delivery CR would be promising alternative with ease of delivery to children and aged patient.

The effect of daily usage of Listerine Cool Mint mouthwash on the oropharyngeal microbiome: a substudy of the PReGo trial.

Introduction. ListerineÒ is a bactericidal mouthwash widely used to prevent oral health problems such as dental plaque and gingivitis. However, whether it promotes or undermines a healthy oral microbiome is unclear.Hypothesis/Gap Statement. We hypothesized that the daily use of Listerine Cool Mint would have a significant impact on the oropharyngeal microbiome.Aim. We aimed to assess if daily usage of Listerine Cool Mint influenced the composition of the pharyngeal microbiome.Methodology. The current microbiome substudy is part of the Preventing Resistance in Gonorrhoea trial. This was a double-blind single-centre, crossover, randomized controlled trial of antibacterial versus placebo mouthwash to reduce the incidence of gonorrhoea/chlamydia/syphilis in men who have sex with men (MSM) taking HIV pre-exposure prophylaxis (PrEP). Fifty-nine MSM taking HIV PrEP were enrolled. In this crossover trial, participants received 3 months of daily Listerine followed by 3 months of placebo mouthwash or vice versa. Oropharyngeal swabs were taken at baseline and after 3 months use of each mouthwash. DNA was extracted for shotgun metagenomic sequencing (Illumina Inc.). Non-host reads were taxonomically classified with MiniKraken and Bracken. The alpha and beta diversity indices were compared between baseline and after each mouthwash use. Differentially abundant bacterial taxa were identified using ANOVA-like differential expression analysis.Results. Streptococcus was the most abundant genus in most samples (n = 103, 61.7 %) with a median relative abundance of 31.5% (IQR 20.6-44.8), followed by Prevotella [13.5% (IQR 4.8-22.6)] and Veillonella [10.0% (IQR 4.0-16.8)]. Compared to baseline, the composition of the oral microbiome at the genus level (beta diversity) was significantly different after 3 months of Listerine (P = 0.006, pseudo-F = 2.29) or placebo (P = 0.003, pseudo-F = 2.49, permutational multivariate analysis of variance) use. Fusobacterium nucleatum and Streptococcus anginosus were significantly more abundant after Listerine use compared to baseline.Conclusion. Listerine use was associated with an increased abundance of common oral opportunistic bacteria previously reported to be enriched in periodontal diseases, oesophageal and colorectal cancer, and systemic diseases. These findings suggest that the regular use of Listerine mouthwash should be carefully considered.

Therapeutic potential of limonene-based syringic acid nanoemulsion: Enhanced ex-vivo cutaneous deposition and clinical anti-psoriatic efficacy.

Nanoemulsions have carved their position in topical delivery owing to their peculiar features of forming a uniform film on the skin and conquering stratum corneum barrier and hence fostering dermal penetration and retention. The present work developed syringic acid nanoemulsion (SA-NE) by spontaneous emulsification as an anti-psoriatic remedy via the dermal route. SA-NE were prepared with either lauroglycol90, limonene or their combination (oil phase) and tween80 (surfactant) with variable concentrations. The physicochemical characteristics of SA-NE were assessed together with Ex-vivo skin deposition and dermal toxicity. The effectiveness of optimal formula in psoriatic animal model and psoriatic patients was investigated using PASI scoring and dermoscope examination. Results showed that, SA-NE containing mixture of lauroglycol 90, limonene and 10 % tween80 (F5), was selected as the optimal formula presenting stable nanoemulsion for 2-month period, showing droplet size of 177.6 ± 13.23 nm, polydispersity index of 0.16 ± 0.06, zeta potential of -21.23 ± 0.41 mV. High SA% in different skin strata and no dermal irritation was noticed with limonene-based SA-NE also it showed high in-vitro anti- inflammatory potential compared to the blank and control formulations. A preclinical study demonstrated that limonene-based SA-NE is effective in alleviating psoriasis-like skin lesions against imiquimod-induced psoriasis in rats. Clinically, promising anti-psoriatic potential was asserted as all patients receiving F5 experienced better clinical improvement and response to therapy, achieving ≥ 50 % reduction in PASI scores versus only 35 % responders in the Dermovate® cream group. Collectively, the practical feasibility of limonene-based SA-NE topical delivery can boost curative functionality in the treatment of psoriatic lesions.

The Solution Is a Solution.

BACKGROUND: Infections in neonates with herpes simplex virus 1 (HSV-1) following circumcision due to Metzitzah Be'Peh (MBP) performed by a Mohel occur each year in small numbers. One solution to this problem is the use of a mucus extractor device instead of MBP, which has been authorized by some rabbis. Yet, using a mucus extractor remains controversial among ultra-Orthodox Jews; thus, creating a need for additional solutions. OBJECTIVES: To seek to reduce HSV-1 infection of neonates due to MBP. METHODS: We tested several oral rinse solutions for their ability to destroy virus infectivity following incubation for 30 seconds and using plaque reduction assays. RESULTS: Corsodyl, Decapinol, and Listerine® all destroyed plaques formation of spiked virus, while Gengigel and Tantum Verde were found to be less effective. We focused specifically on Listerine® due to its efficacy in eliminating contagious HSV-1 from saliva after a 30-second oral rinse. Five different products of Listerine® reduced the infectivity of a spiked virus by more than 4 orders of magnitude in 30 seconds. We also showed that Listerine (up to 7% v/v) can stay in the mouth but did not harm living cells and therefore will not cause any damage to the injured tissue. CONCLUSIONS: Significant reduction in cases of infection with HSV-1 due to MBP can be achieved if Mohalim consistently adopt the practice of careful mouth washing with Listerine® just before performing MBP.

Cytotoxic terpenoids from Tripterygium hypoglaucum against human pancreatic cancer cells SW1990 by increasing the expression of Bax protein.

ETHNOPHARMACOLOGICAL RELEVANCE: Tripterygium hypoglaucum (Kunmingshanhaitang in Chinese) is a plant of the genus Tripterygium which have been used as anti-tumor folk medicines in Yi and Bai ethnic groups in Yunnan province, China for hundreds of years. Terpenoids from T. hypoglaucum presented therapeutic effects on multiple tumors. But there were few studies about pancreatic cancer treatment of these terpenoids. Pancreatic cancer is an aggressive malignancy and lacked of specific drugs. Currently, anti-tumor drugs have poor therapeutic effect and prognosis for pancreatic cancer. AIM OF THE STUDY: This study aimed to elucidate the terpenoids from T. hypoglaucum and illuminate their anti-pancreatic cancer bioactivities. MATERIAL AND METHODS: Terpenoids were obtained through sequential chromatographic methods including silica gel, MCI gel, Sephadex LH-20, and preparative HPLC. Their structures were determined by HRESIMS, 1D and 2D NMR spectroscopic analysis. The absolute configurations of some new diterpenoids were assigned through comparison of experimental and calculated circular dichroism spectra. The cytotoxicity of isolates was measured using the MTT method on human pancreatic cancer cells SW1990. The effects on expressions of AKT, Erk1/2, p-AKT, p-Erk1/2, and Bax proteins in human pancreatic cancer cells SW1990 of these compounds were determined by western blotting assays. RESULTS: Eleven new (compounds 1∼11) and fourteen known terpenoids (compounds 12∼25) were isolated from the underground parts of T. hypoglaucum. These compounds were belonged to abietane diterpenoids, isoprimara diterpenoids, ent-kaurane diterpenoids, oleanane triterpenoids, and friedelane triterpenoids. Compounds 5, 7, 8, 9, 16, 18, 22, 24, and 25 possessed significant cytotoxicity against SW1990 cells with IC50 values of 19.28 ± 4.39, 9.91 ± 2.23, 27.32 ± 5.89, 56.43 ± 6.92, 0.16 ± 0.05, 0.58 ± 0.15, 0.81 ± 0.04, 0.48 ± 0.11, and 10.01 ± 1.39 µM respectively. After compounds 16, 22, and 24 been treated with the pancreatic cancer cells in medium and high doses, the protein expressions of AKT, p-AKT, Erk, and p-Erk were not remarkably reduced and the expressions of Bax protein were significantly increased. CONCLUSION: This study indicated that terpenoids from T. hypoglaucum could inhibit human pancreatic cancer cells SW1990. Especially, compounds 16, 22, and 24 possessed significant cytotoxicity against SW1990 cells with low IC50 values and could increase the expressions of Bax protein. These compounds shared a wide variety of structural characteristics which provided us more candidate molecules for the development of anti-pancreatic cancer drugs and further prompted us to investigate their anti-pancreatic mechanisms.

Safranal Alleviated OVA-Induced Asthma Model and Inhibits Mast Cell Activation.

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 LTC4) 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 LTC4, 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 LTC4 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.

The influence of dietary inclusion of wood vinegar supplementation on growth performance, nutrient digestibility, and meat quality in grower-finisher pigs.

The current research aimed to evaluate the effects of dietary inclusion of wood vinegar on growth performance, nutrient digestibility, and meat quality of grower-finisher pigs. In total, 132 crossbred ({Landrace × Yorkshire × Duroc}) grower-finisher pigs with an initial average body weight 30.48±4.23 kg (11 replications/treatment; 4 pigs/pen) were used in a 16-week trial. Based on the body weight and sex the pigs were randomly assigned to three treatments. Dietary treatments consisted of the basal diet (CON) or the basal diet supplemented with 0.05% and 0.1% wood vinegar. The inclusion of dietary wood vinegar supplementation significantly improved the body weight gain (BWG) and average daily gain (ADG) (P=0.0521; 0.043) of pigs at week 16. The total track nutrient digestibility of dry matter and nitrogen was linearly increased in pigs fed with an increased amount of wood vinegar. In addition, dietary supplementation of wood vinegar linearly improved longissimus muscle area, yellowness (b*) of the meat color, and carcass weight (P<0.05) and a tendency in linear reduction was observed for water holding capacity (P=0.068), and drip loss at d5 and d7 (P=0.091, 0.069). However, there was no significant difference found for lean meat percentage and backfat thickness in this experiment. In summary, dietary inclusion of wood vinegar supplementation enhanced growth performance and total track digestibility of nutrients and had no effects on lean meat percentage and backfat thickness of grower-finisher pigs.

Iguratimod ameliorates nephritis by modulating the Th17/Treg paradigm in pristane-induced lupus.

OBJECTIVE: Iguratimod, an anti-rheumatic drug, has been widely used in the treatment of rheumatoid arthritis, but is still at an investigative stage for treatment of systemic lupus erythematosus (SLE). We examined the therapeutic effects of iguratimod and the mechanism underlying the efficacy in murine lupus model. METHODS: Pristane-induced lupus model of BALB/c mice (PI mice) were treated with iguratimod and mycophenolate mofetil. Proteinuria, anti-dsDNA antibodies and immunoglobulins production were measured. Renal pathology was evaluated. The percentage of Th17 and Treg cells in spleen and the expression of cytokines and mRNAs related to Th17 and Treg cells was analyzed. RESULTS: Iguratimod attenuated the severity of nephritis in PI mice in a dose-dependent manner. Proteinuria was continuously decreased and pathology of glomerulonephritis and tubulonephritis was significantly reduced along with reduction of glomerular immune complex deposition. Also, serum anti-dsDNA and total IgG and IgM levels were reduced by iguratimod in mice. It is worth mentioning that the efficacy of the 30 mg/kg/d iguratimod dose is comparable to, or even better than, 100 mg kg/d of mycophenolate mofetil. Furthermore, the percentage of Th17 cells was found decreased and the percentage of Treg cells increased. ROR-γt mRNA and serum cytokines (IL-17A and IL-22) of Th17 cells decreased accordingly. By contrast, Foxp3 mRNA and cytokines (TGF-ß and IL-10) of Treg cells increased. CONCLUSION: Iguratimod ameliorates nephritis of SLE and modulates the Th17/Treg ratio in murine nephritis of SLE, suggesting that Iguratimod could be an effective drug in treatment of SLE.

Terpenoids from Curcumae Rhizoma: Their anticancer effects and clinical uses on combination and versus drug therapies.

Cancer is a fatal disease with high mortality and low survival rate worldwide. At present, there is still no known cure for most cancers. Traditional Chinese medicine (TCM) represents a noteworthy reservoir for anticancer agents in drug discovery and development. Curcumae Rhizoma (called Ezhu in Chinese) is widely prescribed in TCM for anticancer therapy owing to its broad-spectrum antineoplastic activities. Especially, the terpenoids isolated from the essential oil of Curcumae Rhizoma form an integral part of cancer research and are well established as a potential anticancer agent. For example, ß-elemene has been developed into a new drug for the treatment of solid tumors in China, and is currently undergoing clinical trials in the United States. The review aims to systematically summarize the recent advances on the anticancer effects and related molecular mechanisms of Curcumae Rhizoma, and its terpenoids (ß-elemene, Furanodiene, Furanodienone, Germacrone, Curcumol, Curdione). In addition, we evaluated and compared the anticancer efficacy and clinical use of the terpenoids with combination therapies and traditional therapies. Therefore, this review provides sufficient evidence for the anticancer therapeutic potential of Curcumae Rhizoma and its terpenoids, and will contribute to the development of potential anticancer drugs.

Preclinical Testing of Viral Therapeutic Efficacy in Pristane-Induced Lupus Nephritis and Diffuse Alveolar Hemorrhage Mouse Models.

Systemic lupus erythematosus (SLE) is a multifactorial and heterogeneous autoimmune disease involving multiple organ systems and tissues. Lupus nephritis occurs in approximately 60% of patients with SLE and is the leading cause of morbidity. Diffuse alveolar hemorrhage (DAH) is a rare but very serious complication of SLE with a greater than 50% associated mortality. The etiology of SLE is unclear but has proposed genetic, hormonal, and environmental aspects. Pristane is a saturated terpenoid alkane and has become the most popular laboratory model for inducing lupus in mice. The pristane model of SLE has the capacity to reproduce many components of the human presentation of the disease. Previous studies have demonstrated that virus-derived immune-modulating proteins have the potential to control inflammatory and autoimmune disorders. Serp-1, a 55 kDa secreted and highly glycosylated immune modulator derived from myxoma virus (MYXV), has potent immunomodulatory activity in models of vasculitis, viral sepsis, collagen-induced arthritis, and transplant rejection. This chapter describes the mouse preclinical pristane lupus model as a method to examine virus-derived protein efficacy for treating autoimmune diseases and specifically lupus nephritis and DAH.

Hop Bitter Acids Increase Hippocampal Dopaminergic Activity in a Mouse Model of Social Defeat Stress.

As daily lifestyle is closely associated with mental illnesses, diet-based preventive approaches are receiving attention. Supplementation with hop bitter acids such as iso-α-acids (IAA) and mature hop bitter acids (MHBA) improves mood states in healthy older adults. However, the underlying mechanism remains unknown. Since acute oral consumption with IAA increases dopamine levels in hippocampus and improves memory impairment via vagal nerve activation, here we investigated the effects of chronic administration of hop bitter acids on the dopaminergic activity associated with emotional disturbance in a mouse model of repeated social defeat stress (R-SDS). Chronic administration of IAA and MHBA significantly increased dopaminergic activity based on the dopamine metabolite to dopamine ratio in the hippocampus and medial prefrontal cortex following R-SDS. Hippocampal dopaminergic activity was inversely correlated with the level of R-SDS-induced social avoidance with or without IAA administration. Therefore, chronic treatment with hop bitter acids enhances stress resilience-related hippocampal dopaminergic activity.

Topical Administration of Terpenes Encapsulated in Nanostructured Lipid-Based Systems.

Terpenes are a group of phytocompounds that have been used in medicine for decades owing to their significant role in human health. So far, they have been examined for therapeutic purposes as antibacterial, anti-inflammatory, antitumoral agents, and the clinical potential of this class of compounds has been increasing continuously as a source of pharmacologically interesting agents also in relation to topical administration. Major difficulties in achieving sustained delivery of terpenes to the skin are connected with their low solubility and stability, as well as poor cell penetration. In order to overcome these disadvantages, new delivery technologies based on nanostructures are proposed to improve bioavailability and allow controlled release. This review highlights the potential properties of terpenes loaded in several types of lipid-based nanocarriers (liposomes, solid lipid nanoparticles, and nanostructured lipid carriers) used to overcome free terpenes' form limitations and potentiate their therapeutic properties for topical administration.

Combined Treatment of Heteronemin and Tetrac Induces Antiproliferation in Oral Cancer Cells.

BACKGROUND: Heteronemin, a marine sesterterpenoid-type natural product, possesses an antiproliferative effect in cancer cells. In addition, heteronemin has been shown to inhibit p53 expression. Our laboratory has demonstrated that the thyroid hormone deaminated analogue, tetrac, activates p53 and induces antiproliferation in colorectal cancer. However, such drug mechanisms are still to be studied in oral cancer cells. METHODS: We investigated the antiproliferative effects by Cell Counting Kit-8 and flow cytometry. The signal transduction pathway was measured by Western blotting analyses. Quantitative PCR was used to evaluate gene expression regulated by heteronemin, 3,3',5,5'-tetraiodothyroacetic acid (tetrac), or their combined treatment in oral cancer cells. RESULTS: Heteronemin inhibited not only expression of proliferative genes and Homo Sapiens Thrombospondin 1 (THBS-1) but also cell proliferation in both OEC-M1 and SCC-25 cells. Remarkably, heteronemin increased TGF-ß1 expression in SCC-25 cells. Tetrac suppressed expression of THBS-1 but not p53 expression in both cancer cell lines. Furthermore, the synergistic effect of tetrac and heteronemin inhibited ERK1/2 activation and heteronemin also blocked STAT3 signaling. Combined treatment increased p53 protein and p53 activation accumulation although heteronemin inhibited p53 expression in both cancer cell lines. The combined treatment induced antiproliferation synergistically more than a single agent. CONCLUSIONS: Both heteronemin and tetrac inhibited ERK1/2 activation and increased p53 phosphorylation. They also inhibited THBS-1 expression. Moreover, tetrac suppressed TGF-ß expression combined with heteronemin to further enhance antiproliferation and anti-metastasis in oral cancer cells.

Antihyperglycemic Effects of Annona diversifolia Safford and Its Acyclic Terpenoids: α-Glucosidase and Selective SGLT1 Inhibitiors.

Annona diversifolia Safford and two acyclic terpenoids were evaluated to determine their antihyperglycemic activity as potential α-glucosidase and selective SGLT-1 inhibitiors. Ethanolic extract (EEAd), chloroformic (CHCl3Fr), ethyl acetate (EtOAcFr), aqueous residual (AcRFr), secondary 5 (Fr5) fractions, farnesal (1), and farnesol (2) were evaluated on normoglycemic and streptozocin-induced diabetic mice. EEAd, CHCl3Fr, Fr5, (1) and (2) showed antihyperglycemic activity. The potential as α-glucosidase inhibitors of products was evaluated with oral sucrose and lactose tolerance (OSTT and OLTT, respectively) and intestinal sucrose hydrolysis (ISH) tests; the potential as SGLT-1 inhibitors was evaluated using oral glucose tolerance (OGTT), intestinal glucose absorption (IGA), and urinary glucose excretion (UGE) tests. In OSTT and OLTT, all treatments showed significant activity at two and four hours. In ISH, half maximal effective concentrations (CE50) of 565, 662 and 590 µg/mL, 682 and 802 µM were calculated, respectively. In OGTT, all treatments showed significant activity at two hours. In IGA, CE50 values of 1059, 783 and 539 µg/mL, 1211 and 327 µM were calculated, respectively. In UGE Fr5, (1) and (2) showed significant reduction of the glucose excreted compared with canagliflozin. These results suggest that the antihyperglycemic activity is mediated by α-glucosidase and selective SGLT-1 inhibition.

Correlation of the levels of DNA-binding inhibitor Id3 and regulatory T cells with SLE disease severity.

E proteins, a subset of basic helix-loop-helix (bHLH) proteins, are transcription activators and their functions are inhibited by DNA-binding inhibitor (Id) 1-4. Studies have shown that Treg levels are decreased in Id3 knockout mice. Mice over-expressing Id1 in CD4 T cells possessed a greater number of regulatory T cells (Treg) and exhibited attenuated experimental autoimmune encephalomyelitis (EAE). The significance of Id proteins in human systemic lupus erythematosus (SLE) remains unclear. In this study, we systematically analyzed Id transcription in naïve, memory CD4 cells and regulatory T cells in peripheral blood mononuclear cells (PBMCs) in patients with active or inactive SLE. In parallel, Treg subsets in PBMCs were analyzed using different strategies. Id expression levels were correlated with Treg numbers as well as clinical indicators. We found that Id genes expressed in human peripheral CD4 cells were mainly Id2 and Id3. Id3 levels were significantly elevated in CD4+CD25hi T cells of patients with active SLE. Likewise, Id3 levels were positively correlated with increased CD4+FoxP3+ and CD4+Helios+FoxP3+ Treg cells in these patients. Id3 levels were found to be positively correlated with erythrocyte sedimentation rate (ESR), lupus anticoagulant (LAC), ribosomal antibody and SLE Disease Activity Index (SLEDAI) in patients with active SLE. Mice overexpressing Id1 in CD4+ T cells possessed significantly higher Treg levels in spleen and lower autoantibody concentrations in serum. Our results suggest that during the pathogenesis of SLE, up-regulation of Id3 can promote Treg differentiation to play an inhibitory effect on autoimmune responses.

Novel Combinatorial Regimen of Garcinol and Curcuminoids for Non-alcoholic Steatohepatitis (NASH) in Mice.

Non-alcoholic steatohepatitis (NASH) is a progressive form of Non-alcoholic fatty liver disease (NAFLD), a chronic liver disease with a significant unmet clinical need. In this study, we examined the protective effects of Garcinia indica extract standardized to contain 20% w/w of Garcinol (GIE) and 95% Curcuminoids w/w from Curcuma longa (Curcuminoids) in a Stelic animal model (STAM) of NASH. The STAM mice developed steatosis, hepatocyte ballooning, and inflammation, which were significantly reduced by the combination of GIE and Curcuminoids, resulting in a lower NAFLD activity score. The treatment reduced fibrosis as observed by Sirius red staining, liver hydroxyproline content and mRNA levels of TGF- ß and collagen in the liver. Immunostaining with alpha-smooth muscle actin (α SMA) revealed a significant reduction in hepatic stellate cells. Intriguingly, the combination regimen markedly decreased the mRNA levels of MCP1 and CRP and both mRNA and protein levels of TNF-α. NF-kB, reduced the hepatic and circulating FGF21 levels and altered the nonenzymatic (glutathione) and enzymatic antioxidant markers (Glutathione peroxidase, and superoxide dismutase). Our results suggest that the combination of GIE and Curcuminoids can reduce the severity of NASH by reducing steatosis, fibrosis, oxidative stress, and inflammation. The results suggest that the combinatorial regimen could be an effective supplement to prevent the progression of liver steatosis to inflammation and fibrosis in NASH.

Palbinone alleviates diabetic retinopathy in STZ-induced rats by inhibiting NLRP3 inflammatory activity.

Diabetic retinopathy (DR) is the primary cause of blindness and visual impairment in diabetes patients worldwide. However, laser and surgical therapies at DR have short-term effectiveness and cause side effects. Treatment with natural products is a reasonable alternative treatment for DR. The main objective of this investigation is to explore the efficacy of a bioactive compound such as palbinone (PB) in DR. Experimental rats were injected intraperitoneally with streptozotocin (STZ, 65 mg/kg), and these established experimental rats were treated with PB (20 mg/kg/bw) for 42 days. The observed results showed that PB considerably reduced the proinflammatory cytokine (interleukin-18 [IL-18] and IL-1ß) production as well as improved the activities of antioxidant enzymes (superoxide dismutase, catalase, and glutathione peroxidase) particularly in the retinal region of STZ-induced DR rats. In addition, PB treatment improved nuclear factor erythroid 2-related factor 2 (Nrf2) accumulation and enhanced the heme oxygenase-1 expression, and major antioxidants downregulated Nrf2 in the damaged retina. Also, the expression levels of nod-like receptor family pyrin domain containing 3 (NLRP3), cleaved-caspase-1, IL-1ß, and apoptosis-associated speck-like protein containing CARD in the retinal region were notably upregulated in STZ-induced DR, which was eliminated by PB interference. PB administration exerted efficient antioxidant activities, Nrf2 pathway activation, and inhibition of NLRP3 inflammasome. This current investigation concluded that PB considerably reduced the retinal inflammation and oxidative stress stimulated via high glucose, and also activated the antioxidative Nrf2 pathway and inhibited the NLRP3 inflammasome formation in rats.