This study reports on the effects of unmodified autohydrolyzed ethanol organosolv lignin (AH EOL) and modified autohydrolyzed ethanol organosolv lignin on the structural characteristics and antioxidant properties upon incorporation of p-hydroxyacetophenone (AHP EOL). The lignin samples isolated from black liquor of oil palm fronds (OPF) were evaluated and compared using various complementary analyses; FTIR, 1H and 13C NMR spectroscopy, 2D-NMR spectroscopy (HMBC and HSQC), CHN, GPC, HPLC and thermal analyses (TGA and DSC). Chemically modified organosolv lignin (AHP EOL) provided lignin with lower molecular weight (Mw), which has smaller fragments that leads to higher solubility rate in water in comparison to unmodified organosolv lignin, AH EOL (DAHP EOL: 19.8% > DAH EOL: 14.0%). It was evident that the antioxidant properties of modified organosolv lignin has better reducing power in comparison to the unmodified organosolv lignin. Therefore, the functionalization of lignin polymers enhanced their antioxidant properties and structural features towards a various alternative approach in lignin-based applications.
Paeonol is a naturally existing bioactive compound found in the root bark of Paeonia suffruticosa and it is traditionally used in Chinese medicine for the prevention and management of cardiovascular diseases. To date, a great deal of studies has been reported on the pharmacological effects of paeonol and its mechanisms of action in various diseases and conditions. In this review, the underlying mechanism of action of paeonol in cardiovascular disease has been elucidated. Recent studies have revealed that paeonol treatment improved endothelium injury, demoted inflammation, ameliorated oxidative stress, suppressed vascular smooth muscle cell proliferation, and repressed platelet activation. Paeonol has been reported to effectively protect the cardiovascular system either employed alone or in combination with other traditional medicines, thus, signifying it could be a hypothetically alternative or complementary atherosclerosis treatment. This review summarizes the biological and pharmacological activities of paeonol in the treatment of cardiovascular diseases and its associated underlying mechanisms for a better insight for future clinical practices.
The natural product molecule 2,4,6-trihydroxy-3-geranyl-acetophenone (tHGA) isolated from the medicinal plant Melicope ptelefolia was shown to exhibit potent lipoxygenase (LOX) inhibitory activity. It is known that LOX plays an important role in inflammatory response as it catalyzes the oxidation of unsaturated fatty acids, such as linoleic acid to form hydroperoxides. The search for selective LOX inhibitors may provide new therapeutic approach for inflammatory diseases. Herein, we report the synthesis of tHGA analogs using simple Friedel-Craft acylation and alkylation reactions with the aim of obtaining a better insight into the structure-activity relationships of the compounds. All the synthesized analogs showed potent soybean 15-LOX inhibitory activity in a dose-dependent manner (IC50 = 10.31-27.61 μM) where compound 3e was two-fold more active than tHGA. Molecular docking was then applied to reveal the important binding interactions of compound 3e in soybean 15-LOX binding site. The findings suggest that the presence of longer acyl bearing aliphatic chain (5Cs) and aromatic groups could significantly affect the enzymatic activity.
Transgenic oil palm (Elaeis guineensis Jacq.) plantlets are regenerated after Agrobacterium tumefaciens-mediated transformation of embryogenic calli derived from young leaves of oil palm. The calli are transformed with an Agrobacterium strain, LBA4404, harboring the plasmid pUBA, which carries a selectable marker gene (bar) for resistance to the herbicide Basta and is driven by a maize ubiquitin promoter. Modifications of the transformation method, treatment of the target tissues using acetosyringone, exposure to a plasmolysis medium, and physical injury via biolistics are applied. The main reasons for such modifications are to activate the bacterial virulence system and, subsequently, to increase the transformation efficiency. Transgenic oil palm cells are selected and regenerated on a medium containing herbicide Basta. Molecular analyses revealed the presence and integration of the introduced bar gene into the genome of the transformants.
Reactive oxygen species (ROS) such as superoxide (O2-) generated by NAD(P)H oxidases have emerged as important molecules in blood pressure regulation. This study investigated the effect of apocynin and catalase on blood pressure and renal haemodynamic and excretory function in an L-NAME induced hypertension model. Forty Male Wistar-Kyoto (WKY) rats (n=8 per group) were treated with either: vehicle (WKY-C); L-NAME (WKY-L, 15 mg/kg/day in drinking fluid); WKY-L given apocynin to block NAD(P)H oxidase (WKY-LApo, 73 mg/kg/day in drinking water.); WKY-L given catalase to enhance ROS scavenging (WKY-LCat, 10000 U/kg/day i.p.); and WKY-L receiving apocynin plus catalase (WKY-LApoCat) daily for 14 days. L-NAME elevated systolic blood pressure (SBP), 116+/-1 to 181±4 mmHg, reduced creatinine clearance, 1.69+/-0.26 to 0.97+/-0.05 ml/min/kg and fractional sodium excretion, 0.84+/-0.09 to 0.55+/-0.09 % at day 14. Concomitantly, plasma malondialdehyde (MDA) increased six fold, while plasma total superoxide dismutase (T-SOD), plasma nitric oxide (NO) and plasma total antioxidant capacity (T-AOC) were decreased by 60-70 % and Nox 4 mRNA expression was increased 2-fold. Treatment with apocynin and catalase attenuated the increase in SBP and improved renal function, enhanced antioxidative stress capacity and reduced the magnitude of Nox4 mRNAs expression in the L-NAME treated rats. This study demonstrated that apocynin and catalase offset the development of L-NAME induced hypertension, renal dysfunction and reduced oxidative stress status, possibly contributed by a reduction in Nox4 expression during NOS inhibition. These findings would suggest that antioxidant compounds such as apocynin and catalase have potential in treating cardiovascular diseases.
Paeonol is a phenolic compound isolated mainly from Moutan cortex, root bark of Chinese Peony tree. Moutan cortex holds a significant value in traditional Chinese medicine for alleviating various oxidative stress-related diseases mainly atherosclerosis and myocardial infarction. The present study seeks to identify the protective mechanisms of paeonol in oxidative stress-induced premature senescence in endothelial cells.
Asthma is associated with increased pulmonary inflammation and airway hyperresponsiveness. The current use of corticosteroids in the management of asthma has recently raised issues regarding safety and lack of responsiveness in 5-10% of asthmatic individuals. The aim of the present study was to investigate the therapeutic effect of a non-steroidal small molecule that has cysteinyl leukotriene (cysLT) inhibitory activity, upon attenuation of allergic lung inflammation in an acute murine model. Mice were sensitized with ovalbumin (OVA) and treated with several intraperitoneal doses (100, 20, 2 and 0.2mg/kg) of 2,4,6,-trihydroxy-3-geranylacetophenone (tHGA). Bronchoalveolar lavage was performed, blood and lung samples were obtained and respiratory function was measured. OVA sensitization increased pulmonary inflammation and pulmonary allergic inflammation was significantly reduced at doses of 100, 20 and 2mg/kg with no effect at the lowest dose of 0.2mg/kg. The beneficial effects in the lung were associated with reduced eosinophilic infiltration and reduced secretion of Th2 cytokines and cysLTs. Peripheral blood reduction of total IgE was also a prominent feature. Treatment with tHGA significantly attenuated altered airway hyperresponsiveness as measured by the enhanced pause (Penh) response to incremental doses of methacholine. These data demonstrate that tHGA, a synthetic non-steroidal small molecule, can prevent acute allergic inflammation. This proof of concept opens further avenues of research and development of tHGA as an additional option to the current armamentarium of anti-asthma therapeutics.
Endoplasmic reticulum (ER) stress leads to endothelial dysfunction which is commonly associated in the pathogenesis of several cardiovascular diseases. We explored the vascular protective effects of chronic treatment with paeonol (2'-hydroxy-4'-methoxyacetophenone), the major compound from the root bark of Paeonia suffruticosa on ER stress-induced endothelial dysfunction in mice. Male C57BL/6J mice were injected intraperitoneally with ER stress inducer, tunicamycin (1 mg/kg/week) for 2 weeks to induce ER stress. The animals were co-administered with or without paeonol (20 mg/kg/oral gavage), reactive oxygen species (ROS) scavenger, tempol (20 mg/kg/day) or ER stress inhibitor, tauroursodeoxycholic acid (TUDCA, 150 mg/kg/day) respectively. Blood pressure and body weight were monitored weekly and at the end of treatment, the aorta was isolated for isometric force measurement. Protein associated with ER stress (GRP78, ATF6 and p-eIF2α) and oxidative stress (NOX2 and nitrotyrosine) were evaluated using Western blotting. Nitric oxide (NO) bioavailability were determined using total nitrate/nitrite assay and western blotting (phosphorylation of eNOS protein). ROS production was assessed by en face dihydroethidium staining and lucigenin-enhanced chemiluminescence assay, respectively. Our results revealed that mice treated with tunicamycin showed an increased blood pressure, reduction in body weight and impairment of endothelium-dependent relaxations (EDRs) of aorta, which were ameliorated by co-treatment with either paeonol, TUDCA and tempol. Furthermore, paeonol reduced the ROS level in the mouse aorta and improved NO bioavailability in tunicamycin treated mice. These beneficial effects of paeonol observed were comparable to those produced by TUDCA and tempol, suggesting that the actions of paeonol may involve inhibition of ER stress-mediated oxidative stress pathway. Taken together, the present results suggest that chronic treatment with paeonol preserved endothelial function and normalized blood pressure in mice induced by tunicamycin in vivo through the inhibition of ER stress-associated ROS.
L-arginine is a substrate for nitric oxide synthase (NOS) responsible for the production of NO. This investigation studied the effect of apocynin, an NADPH oxidase inhibitor and catalase, an H2O2 scavenger on L-arginine induced oxidative stress and hypotension. Forty Wistar-Kyoto rats were treated for 14 days with vehicle, L-arginine (12.5mg/ml p.o.), L-arginine+apocynin (2.5mmol/L p.o.), L-arginine+catalase (10000U/kg/day i.p.) and L-arginine plus apocynin+catalase respectively. Weekly renal functional and hemodynamic parameters were measured and kidneys harvested at the end of the study for histopathological and renal NADPH oxidase 4 (Nox4) assessments. L-arginine administration in normotensive rats decreased systolic blood pressure (120±2 vs 91±2mmHg) and heart rate (298±21 vs 254±15b/min), enhanced urinary output (21.5±4.2 vs 32±1.9ml/24h , increased creatinine clearance (1.72±0.56 vs 2.62±0.40ml/min/kg), and fractional sodium excretion (0.88±0.16 vs 1.18±0.16 %), caused proteinuria (28.10±1.93 vs 35.26±1.69mg/kg/day) and a significant decrease in renal cortical blood perfusion (292±3 vs 258±5bpu) and pulse wave velocity (3.72±0.20 vs 2.84±0.13m/s) (all P<0.05). L-arginine increased plasma malondialdehyde (by ~206 % P<0.05) and NO (by~51 %, P<0.05) but decreased superoxide dismutase (by~31 %, P<0.05) and total antioxidant capacity (by~35 %, P<0.05) compared to control. Renal Nox4 mRNA activity was approximately 2.1 fold higher (P<0.05) in the L-arginine treated rats but was normalized by apocynin and apocynin plus catalase treatment. Administration of apocynin and catalase, but not catalase alone to rats fed L-arginine, restored the deranged renal function and structure, prevented hypotension and enhanced the antioxidant capacity and suppressed Nox4 expression. These findings suggest that apocynin and catalase might be used prophylactically in states of oxidative stress.
Inflammatory injury of the endothelium leads to apoptosis and endothelial dysfunction. The current study explored the effect and mechanisms of paeonol in inflammation-induced apoptosis and endothelial dysfunction induced by lipopolysaccharides (LPSs). The effects of paeonol on LPS-induced inflammatory injury were assessed by Western blotting, flow cytometry and reactive oxygen species (ROS) measurement in human umbilical vein endothelial cells (HUVECs) and C57BL/6J mice. Vascular reactivity of isolated mouse aortae was examined using wire myographs. The exposure of HUVECs to LPS increased the protein presence of Toll-like receptor 4 (TLR4), bone morphogenic protein 4 (BMP4), BMP receptor type 1A, nicotinamide adenine dinucleotide phosphate oxidase subunit 2, mitogen-activated protein kinase (MAPK), inducible nitric oxide synthase (iNOS), and cleaved caspase 3, as well as decreased it in phosphorylated endothelial nitric oxide synthase; these effects were prevented by treatment with paeonol. Similarly, cotreatment with paeonol reversed BMP4-induced apoptosis in HUVECs. Relaxation in response to the endothelium-dependent vasodilator acetylcholine were impaired in mouse aortae after exposure to LPSs; this endothelial dysfunction was reversed by cotreatment with paeonol, noggin (a BMP4 inhibitor), TAK242 (TLR4 antagonist), apocynin (an ROS scavenger), MAPK inhibitors, and AG (an iNOS inhibitor). BMP4 small interfering RNAs (siRNAs) abolished LPS-induced upregulation of BMP4 and cleaved caspase 3 protein, but not in cells treated with TLR4 siRNA and vice versa. The silencing of TLR4 and BMP4 abolished the inhibitory effects of paeonol on LPS-induced activation of cleaved caspase 3. The present results demonstrate that paeonol reduces LPS-induced endothelial dysfunction and apoptosis by inhibiting TLR4 and BMP4 signaling independently.
tHGA, a geranyl acetophenone compound originally isolated from a local shrub called Melicope ptelefolia, has been previously reported to prevent ovalbumin-induced allergic airway inflammation in a murine model of allergic asthma by targeting cysteinyl leukotriene synthesis. Mast cells are immune effector cells involved in the pathogenesis of allergic diseases including asthma by releasing cysteinyl leukotrienes. The anti-asthmatic properties of tHGA could be attributed to its inhibitory effect on mast cell degranulation. As mast cell degranulation is an important event in allergic responses, this study aimed to investigate the anti-allergic effects of tHGA in cellular and animal models of IgE-mediated mast cell degranulation. For in vitro model of IgE-mediated mast cell degranulation, DNP-IgE-sensitized RBL-2H3 cells were pre-treated with tHGA before challenged with DNP-BSA to induce degranulation. For IgE-mediated passive systemic anaphylaxis, Sprague Dawley rats were sensitized by intraperitoneal injection of DNP-IgE before challenged with DNP-BSA. Both in vitro and in vivo models showed that tHGA significantly inhibited the release of preformed mediators (β-hexosaminidase and histamine) as well as de novo mediators (interleukin-4, tumour necrosis factor-α, prostaglandin D2 and leukotriene C4). Pre-treatment of tHGA also prevented IgE-challenged RBL-2H3 cells and peritoneal mast cells from undergoing morphological changes associated with mast cell degranulation. These findings indicate that tHGA possesses potent anti-allergic activity via attenuation of IgE-mediated mast cell degranulation and inhibition of IgE-mediated passive systemic anaphylaxis. Thus, tHGA may have the potential to be developed as a mast cell stabilizer for the treatment of allergic diseases in the future.
Protein kinase C (PKC) has been implicated in carcinogenesis and displays variable expression profiles during cancer progression. Studies of dietary phytochemicals on cancer signalling pathway regulation have been conducted to search for potent signalling regulatory agents. The present study was designed to evaluate any suppressive effect of maslinic acid on PKC expression in human B-lymphoblastoid cells (Raji cells), and to identify the PKC isoforms expressed. Effects of maslinic acid on PKC activity were determined using a PepTag assay for non-radioactive detection of PKC. The highest expression in Raji cells was obtained at 20 nM PMA induced for 6 hours. Suppressive effects of maslinic acid were compared with those of four PKC inhibitors (H- 7, rottlerin, sphingosine, staurosporine) and two triterpenes (oleanolic acid and ursolic acid). The IC₅₀ values achieved for maslinic acid, staurosporine, H-7, sphingosine, rottlerin, ursolic acid and oleanolic acid were 11.52, 0.011, 0.767, 2.45, 5.46, 27.93 and 39.29 μM, respectively. Four PKC isoforms, PKC βI, βII, δ, and ζ, were identified in Raji cells via western blotting. Maslinic acid suppressed the expression of PKC βI, δ, and ζ in a concentration-dependent manner. These preliminary results suggest promising suppressive effects of maslinic acid on PKC activity in Raji cells. Maslinic acid could be a potent cancer chemopreventive agent that may be involved in regulating many downstream signalling pathways that are activated through PKC receptors.
A bioassay-guided investigation of Melicope ptelefolia Champ ex Benth (Rutaceae) resulted in the identification of an acyphloroglucinol, 2,4,6-trihydroxy-3-geranylacetophenone or tHGA, as the active principle inhibiting soybean 15-LOX. The anti-inflammatory action was also demonstrated on human leukocytes, where the compound showed prominent inhibitory activity against human PBML 5-LOX, with an IC(50) value of 0.42 μM, very close to the effect produced by the commonly used standard, NDGA. The compound concentration-dependently inhibited 5-LOX product synthesis, specifically inhibiting cysteinyl leukotriene LTC(4) with an IC(50) value of 1.80 μM, and showed no cell toxicity effects. The anti-inflammatory action does not seem to proceed via redox or metal chelating mechanism since the compound tested negative for these bioactivities. Further tests on cyclooxygenases indicated that the compound acts via a dual LOX/COX inhibitory mechanism, with greater selectivity for 5-LOX and COX-2 (IC(50) value of 0.40 μM). The molecular features that govern the 5-LOX inhibitory activity was thus explored using in silico docking experiments. The residues Ile 553 and Hie 252 were the most important residues in the interaction, each contributing significant energy values of -13.45 (electrostatic) and -5.40 kcal/mol (electrostatic and Van der Waals), respectively. The hydroxyl group of the phloroglucinol core of the compound forms a 2.56Å hydrogen bond with the side chain of the carboxylate group of Ile 553. Both Ile 553 and Hie 252 are crucial amino acid residues which chelate with the metal ion in the active site. Distorting the geometry of these ligands could be the reason for the inhibition activity shown by tHGA. The molecular simulation studies supported the bioassay results and served as a good model for understanding the way tHGA binds in the active site of human 5-LOX enzyme.
Oxidised low density lipoprotein (oxLDL) is thought to be a significant contributor to the death of macrophage cells observed in advanced atherosclerotic plaques. Using human-derived U937 cells we have examined the effect of cytotoxic oxLDL on oxidative stress and cellular catabolism. Within 3h of the addition of oxLDL, there was a rapid, concentration dependent rise in cellular reactive oxygen species followed by the loss of cellular GSH, and the enzyme activity of both glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and aconitase. The loss of these catabolic enzymes was accompanied by the loss of cellular ATP and lower lactate generation. Addition of the macrophage antioxidant 7,8-dihydroneopterin inhibited the ROS generation, glutathione loss and catabolic inactivation. NOX was shown to be activated by oxLDL addition while apocynin inhibited the loss of GSH and cell viability. The data suggests that oxLDL triggers an excess of ROS production through NOX activation, and catabolic failure through thiol oxidation resulting in cell death.
An efficient system for shoot regeneration and Agrobacterium tumefaciens-mediated transformation of Brassica oleracea cv. Green Marvel cultivar is described. This study focuses on developing shoot regeneration from hypocotyl explants of broccoli cv. Green Marvel using thidiazuron (TDZ), zeatin, and kinetin, the optimization of factors affecting Agrobacterium-mediated transformation of the hypocotyl explants with heat-resistant cDNA, followed by the confirmation of transgenicity of the regenerants. High shoot regeneration was observed in 0.05-0.1 mg dm(-3) TDZ. TDZ at 0.1 mg dm(-3) produced among the highest percentage of shoot regeneration (96.67 %) and mean number of shoot formation (6.17). The highest percentage (13.33 %) and mean number (0.17) of putative transformant production were on hypocotyl explants subjected to preculture on shoot regeneration medium (SRM) with 200 µM acetosyringone. On optimization of bacterial density and inoculation time, the highest percentage and mean number of putative transformant production were on hypocotyl explants inoculated with a bacterial dilution of 1:5 for 30 min. Polymerase chain reaction (PCR) assay indicated a transformation efficiency of 8.33 %. The luciferase assay showed stable integration of the Arabidopsis thaliana HSP101 (AtHSP101) cDNA in the transgenic broccoli regenerants. Three out of five transgenic lines confirmed through PCR showed positive hybridization bands of the AtHSP101 cDNA through Southern blot analysis. The presence of AtHSP101 transcripts in the three transgenic broccoli lines indicated by reverse transcription-PCR (RT-PCR) confirmed the expression of the gene. In conclusion, an improved regeneration system has been established from hypocotyl explants of broccoli followed by successful transformation with AtHSP101 for resistance to high temperature.
Alveolar epithelial barrier dysfunction contributes to lung edema and can lead to acute lung injury (ALI). The features include increased epithelial permeability, upregulation of inflammatory mediators and downregulation of junctional complex molecules; these changes are often induced by inflammation. tHGA is an acetophenone analogue with therapeutic potential in asthma. Its therapeutic potential in ALI is presently unknown. Herein, the effects of tHGA on epithelial barrier dysfunction were determined in TNF-α-induced human alveolar epithelial cells. The anti-inflammatory properties of tHGA were assessed by monocyte adhesion assay and analysis of MCP-1 and ICAM-1 expression. The epithelial barrier function was assessed by paracellular permeability and transepithelial electrical resistance (TEER) assays, and analysis of junctional complex molecules expression. To elucidate the mechanism of action, the effects of tHGA on the NF-κB and MAPK pathways were determined. Gene and protein expression were analyzed by RT-PCR and Western blotting or ELISA, respectively. tHGA suppressed leukocyte adhesion to TNF-α-induced epithelium and reduced MCP-1 and ICAM-1 gene expression and secretion. tHGA also increased TEER readings, reduced epithelial permeability and enhanced expression of junctional complex molecules (zona occludens-1, occludin and E-cadherin) in TNF-α-induced cells. Correspondingly, the NF-κB, ERK and p38 MAPK pathways were also inhibited by tHGA. These findings suggest that tHGA is able to preserve alveolar epithelial barrier function in response to acute inflammation, via its anti-inflammatory activity and stabilization of epithelial barrier integrity, mediated by NF-κB, ERK and p38 MAPK signaling.
Increased airway smooth muscle (ASM) mass is a prominent hallmark of airway remodeling in asthma. Inhaled corticosteroids and long-acting beta2-agonists remain the mainstay of asthma therapy, however are not curative and ineffective in attenuating airway remodeling. The geranyl acetophenone 2,4,6-trihydroxy-3-geranyl acetophenone (tHGA), an in-house synthetic non-steroidal compound, attenuates airway hyperresponsiveness and remodeling in murine models of asthma. The effect of tHGA upon human ASM proliferation, migration and survival in response to growth factors was assessed and its molecular target was determined. Following serum starvation and induction with growth factors, proliferation and migration of human bronchial smooth muscle cells (hBSMCs) treated with tHGA were significantly inhibited without any significant effects upon cell survival. tHGA caused arrest of hBSMC proliferation at the G1 phase of the cell cycle with downregulation of cell cycle proteins, cyclin D1 and diminished degradation of cyclin-dependent kinase inhibitor (CKI), p27Kip1. The inhibitory effect of tHGA was demonstrated to be related to its direct inhibition of AKT phosphorylation, as well as inhibition of JNK and STAT3 signal transduction. Our findings highlight the anti-remodeling potential of this drug lead in chronic airway disease.
Mast cells (MCs), a type of immune effector cell, have recently become recognized for their ability to cause vascular leakage during dengue virus (DENV) infection. Although MC stabilizers have been reported to attenuate DENV induced infection in animal studies, there are limited in vitro studies on the use of MC stabilizers against DENV induced MC degranulation. 2,4,6-trihydroxy-3-geranyl acetophenone (tHGA) has been reported to be a potential MC stabilizer by inhibiting IgE-mediated MC activation in both cellular and animal models. The present study aims to establish an in vitro model of DENV3-induced RBL-2H3 cells using ketotifen fumarate as a control drug, as well as to determine the effect of tHGA on the release of MC mediators upon DENV infection. Our results demonstrated that the optimal multiplicities of infection (MOI) were 0.4 × 10-2 and 0.8 × 10-2 focus forming units (FFU)/cell. Ketotifen fumarate was proven to attenuate DENV3-induced RBL-2H3 cells degranulation in this in vitro model. In contrast, tHGA was unable to attenuate the release of both β-hexosaminidase and tumor necrosis factor (TNF)-α. Nonetheless, our study has successfully established an in vitro model of DENV3-induced RBL-2H3 cells, which might be useful for the screening of potential MC stabilizers for anti-dengue therapies.
2,4,6-Trihydroxy-3-geranyl acetophenone (tHGA) is a synthetic compound that is naturally found in Melicope ptelefolia. We had previously demonstrated that parenteral administration of tHGA reduces pulmonary inflammation in OVA-sensitized mice. In this study, we evaluated the effect of orally administered tHGA upon airway remodeling in a murine model of chronic asthma. Female BALB/C mice were sensitized intraperitoneally with ovalbumin (OVA) on day 0, 7 and 14, followed by aerosolized 1% OVA 3 times per week for 6 weeks. Control groups were sensitized with saline. OVA sensitized animals were either treated orally with vehicle (saline with 1% DMSO and Tween 80), tHGA (80, 40, 20mg/kg) or zileuton (30mg/kg) 1h prior to each aerosolized OVA sensitization. On day 61, mice underwent methacholine challenge to determine airway hyperresponsiveness prior to collection of bronchoalveolar lavage (BAL) fluid and lung samples. BAL fluid inflammatory cell counts and cytokine concentrations were evaluated while histological analysis and extracellular matrix protein concentrations were determined on collected lung samples. Oral tHGA treatment attenuated airway hyperresponsiveness and inhibited airway remodeling in a dose-dependent fashion. tHGA's effect on airway remodeling could be attributed to the reduction of inflammatory cell infiltration and decreased expression of cytokines associated with airway remodeling. Oral administration of tHGA attenuates airway hyperresponsiveness and remodeling in OVA-induced BALB/c mice. tHGA is an interesting compound that should be evaluated further for its possible role as an alternative non-steroidal pharmacological approach in the management of asthma.
Endoplasmic reticulum (ER) stress in endothelial cells often leads to endothelial dysfunction which underlies the pathogenesis of cardiovascular diseases. Paeonol, a major phenolic component extracted from Moutan Cortex, possesses various medicinal benefits which have been used extensively in traditional Chinese medicine. The present study investigated the protective mechanism of paeonol against tunicamycin-induced ER stress in isolated mouse aortas and human umbilical vein endothelial cells (HUVECs). Vascular reactivity in aorta was measured using a wire myograph. The effects of paeonol on protein expression of ER stress markers, reactive oxygen species (ROS) production, nitric oxide (NO) bioavailability and peroxisome proliferator-activated receptor δ (PPARδ) activity in the vascular wall were assessed by Western blot, dihydroethidium fluorescence (DHE) or lucigenin enhanced-chemiluminescence, 4-amino-5-methylamino-2',7'-difluorofluorescein (DAF-FM DA) and dual luciferase reporter assay, respectively. Ex vivo treatment with paeonol (0.1μM) for 16h reversed the impaired endothelium-dependent relaxations in C57BJ/6J and PPARδ wild type (WT) mouse aortas following incubation with tunicamycin (0.5μg/mL). Elevated ER stress markers, oxidative stress and reduction of NO bioavailability induced by tunicamycin in HUVECs, C57BJ/6J and PPARδ WT mouse aortas were reversed by paeonol treatment. These beneficial effects of paeonol were diminished in PPARδ knockout (KO) mouse aortas. Paeonol increased the expression of 5' adenosine monophosphate-activated protein kinase (AMPK) and PPARδ expression and activity while restoring the decreased phosphorylation of eNOS. The present study delineates that paeonol protects against tunicamycin-induced vascular endothelial dysfunction by inhibition of ER stress and oxidative stress, thus elevating NO bioavailability via the AMPK/PPARδ signaling pathway.