A triterpenoid-enriched extract of bitter melon leaves alleviates hepatic fibrosis by inhibiting inflammatory responses in carbon tetrachloride-treated mice.

Liver fibrosis is a progression of chronic liver disease characterized by excess deposition of fibrillary collagen. The aim of this study was to investigate the protective effect of a triterpenoid-enriched extract (TEE) from bitter melon leaves against carbon tetrachloride (CCl4)-induced hepatic fibrosis in mice. Male ICR mice received TEE (100 or 150 mg kg-1) by daily oral gavage for one week before starting CCl4 administration and throughout the entire experimental period. After intraperitoneal injection of CCl4 for nine weeks, serum and liver tissues of the mice were collected for biochemical, histopathological and molecular analyses. Our results showed that TEE supplementation reduced CCl4-induced serum aspartate aminotransferase and alanine aminotransferase activities. Histopathological examinations revealed that CCl4 administration results in hepatic fibrosis, while TEE supplementation significantly suppressed hepatic necroinflammation and collagen deposition. In addition, TEE supplementation decreased α-smooth muscle actin (α-SMA)-positive staining and protein levels of α-SMA and transforming growth factor-ß1. TEE-supplemented mice had lower mRNA expression levels of interleukin-6, tumor necrosis factor-α, and toll-like receptor 4. Moreover, TEE (150 mg kg-1) supplementation significantly reduced intrahepatic inflammatory Ly6C+ monocyte infiltration. We demonstrated that TEE could ameliorate hepatic fibrosis by regulating inflammatory cytokine secretion and α-SMA expression in the liver to reduce collagen accumulation.

Suppressive Effect of Two Cucurbitane-Type Triterpenoids from Momordica charantia on Cutibacterium acnes-Induced Inflammatory Responses in Human THP-1 Monocytic Cell and Mouse Models.

Cutibacterium acnes (formerly Propionibacterium acnes) is one of the major bacterial species responsible for acne vulgaris. Numerous bioactive compounds from Momordica charantia Linn. var. abbreviata Ser. have been isolated and examined for many years. In this study, we evaluated the suppressive effect of two cucurbitane-type triterpenoids, 5ß,19-epoxycucurbita-6,23-dien-3ß,19,25-triol (Kuguacin R; KR) and 3ß,7ß,25-trihydroxycucurbita-5,23-dien-19-al (TCD) on live C. acnes-stimulated in vitro and in vivo inflammatory responses. Using human THP-1 monocytes, KR or TCD suppressed C. acnes-induced production of interleukin (IL)-1ß, IL-6 and IL-8 at least above 56% or 45%, as well as gene expression of these three pro-inflammatory cytokines. However, a significantly strong inhibitory effect on production and expression of tumor necrosis factor (TNF)-α was not observed. Both cucurbitanes inhibited C. acnes-induced activation of the myeloid differentiation primary response 88 (MyD88) (up to 62%) and mitogen-activated protein kinases (MAPK) (at least 36%). Furthermore, TCD suppressed the expression of pro-caspase-1 and cleaved caspase-1 (p10). In a separate study, KR or TCD decreased C. acnes-stimulated mouse ear edema by ear thickness (20% or 14%), and reduced IL-1ß-expressing leukocytes and neutrophils in mouse ears. We demonstrated that KR and TCD are potential anti-inflammatory agents for modulating C. acnes-induced inflammation in vitro and in vivo.

In Vitro and In Vivo Screening of Wild Bitter Melon Leaf for Anti-Inflammatory Activity against Cutibacterium acnes.

Cutibacterium acnes (formerly Propionibacterium acnes) is a key pathogen involved in the development and progression of acne inflammation. The numerous bioactive properties of wild bitter melon (WBM) leaf extract and their medicinal applications have been recognized for many years. In this study, we examined the suppressive effect of a methanolic extract (ME) of WBM leaf and fractionated components thereof on live C. acnes-induced in vitro and in vivo inflammation. Following methanol extraction of WBM leaves, we confirmed anti-inflammatory properties of ME in C. acnes-treated human THP-1 monocyte and mouse ear edema models. Using a bioassay-monitored isolation approach and a combination of liquid-liquid extraction and column chromatography, the ME was then separated into n-hexane, ethyl acetate, n-butanol and water-soluble fractions. The hexane fraction exerted the most potent anti-inflammatory effect, suppressing C. acnes-induced interleukin-8 (IL-8) production by 36%. The ethanol-soluble fraction (ESF), which was separated from the n-hexane fraction, significantly inhibited C. acnes-induced activation of mitogen-activated protein kinase (MAPK)-mediated cellular IL-8 production. Similarly, the ESF protected against C. acnes-stimulated mouse ear swelling, as measured by ear thickness (20%) and biopsy weight (23%). Twenty-four compounds in the ESF were identified using gas chromatograph-mass spectrum (GC/MS) analysis. Using co-cultures of C. acnes and THP-1 cells, ß-ionone, a compound of the ESF, reduced the production of IL-1ß and IL-8 up to 40% and 18%, respectively. ß-ionone also reduced epidermal microabscess, neutrophilic infiltration and IL-1ß expression in mouse ear. We also found evidence of the presence of anti-inflammatory substances in an unfractionated phenolic extract of WBM leaf, and demonstrated that the ESF is a potential anti-inflammatory agent for modulating in vitro and in vivo C. acnes-induced inflammatory responses.

Ethanolic Extract of Origanum vulgare Suppresses Propionibacterium acnes-Induced Inflammatory Responses in Human Monocyte and Mouse Ear Edema Models.

Acne vulgaris (acne) is a common inflammatory skin disorder, and Propionibacterium acnes plays a major role in the development and progression of acne inflammation. Herbs possessing antimicrobial and anti-inflammatory activity have been applied as a medical option for centuries. In this study, we examined the suppressive effect of ethanolic oregano (Origanum vulgare) extract on live P. acnes-induced in vivo and in vitro inflammation. Following ethanol extraction of oregano leaves, four compounds with strong antioxidant activity, including rosmarinic acid, quercetin, apigenin, and carvacrol, were identified by high-performance liquid chromatography. Using the mouse ear edema model, we demonstrated that ethanol oregano extracts (EOE) significantly suppressed P. acnes-induced skin inflammation, as measured by ear thickness (32%) and biopsy weight (37%). In a separate study, using the co-culture of P. acnes and human THP-1 monocytes, EOE reduced the production of interleukin (IL)-8, IL-1ß and tumor necrosis factor (TNF)-α up to 40%, 37%, and 18%, respectively, as well as the expression of these three pro-inflammatory mediators at the transcriptional level. Furthermore, EOE inhibited the translocation of nuclear factor-kappa B (NF-κB) into the nucleus possibly by inactivating toll-like receptor-2 (TLR2). The suppressive effect of EOE on live P. acnes-induced inflammatory responses could be due, in part, to the anti-inflammatory and antioxidant properties, but not the anti-microbial effect of EOE.

Protective Effect of Caffeic Acid Derivatives on tert-Butyl Hydroperoxide-Induced Oxidative Hepato-Toxicity and Mitochondrial Dysfunction in HepG2 Cells.

Oxidative stress results in structural and functional abnormalities in the liver and is thought to be a crucial factor in liver diseases. The aim of this study was to investigate the cytoprotective and antioxidant effects of caffeic acid (CA) derivatives on tert-butyl hydroperoxide (t-BHP)-induced oxidative stress in HepG2 cells. Nine CA derivatives were synthesized, including N-phenylethyl caffeamide (PECA), N-(3-florophen)methyl caffeamide (FMCA), N-(4-methoxy-phen)methyl caffeamide (MPMCA), N-heptyl caffeamide (HCA), N-octyl caffeamide (OCA), octyl caffeate (CAOE), phenpropyl caffeate (CAPPE), phenethyl caffeate (CAPE), and phenmethyl caffeate (CAPME). The results showed that CA and its derivatives significantly inhibited t-BHP-induced cell death of HepG2 cells. The rank order of potency of the CA derivatives for cytoprotection was CAOE > HCA > OCA > FMCA > CAPPE > CAPME > CAPE > PECA > MPMCA > CA. Their cytoprotective activity was associated with lipophilicity. The antioxidant effect of these compounds was supported by the reduction in the levels of thiobarbituric acid reactive substrates, a biomarker of lipid peroxidation, in HepG2 cells. Pre-treatment of CA derivatives significantly prevented the depletion of glutathione, the most important water-soluble antioxidant in hepatocytes. Pre-treatment of CA derivatives before t-BHP exposure maintained mitochondrial oxygen consumption rate and ATP content in the injured HepG2 cells. CA derivatives except OCA and HCA significantly suppressed t-BHP-induced hypoxia-inducible factor-1α (HIF-1α) protein level. In addition, all of these CA derivatives markedly increased the nuclear factor erythroid 2-related factor 2 (Nrf2) accumulation in the nucleus, indicating that their cytoprotection may be mediated by the activation of Nrf2. Our results suggest that CA derivatives might be a hepatoprotective agent against oxidative stress.

Antilipotoxicity Activity of Osmanthus fragrans and Chrysanthemum morifolium Flower Extracts in Hepatocytes and Renal Glomerular Mesangial Cells.

The excess influx of free fatty acids (FFAs) into nonadipose tissues, such as those of liver and kidney, induces lipotoxicity leading to hepatic steatosis and renal dysfunction. The aim of this study was to investigate the protective effects of methanolic flower extracts of Osmanthus fragrans (OF) and Chrysanthemum morifolium (CM) against FFA-induced lipotoxicity in hepatocytes (human HepG2 cells) and renal glomerular mesangial cells (mouse SV40-Mes13 cells). The results showed that OF and CM significantly suppressed FFA-induced intracellular triacylglycerol accumulation via partially inhibiting the gene expression of sterol regulatory element-binding protein-1c (SREBP-1c) and glycerol-3-phosphate acyltransferase (GPAT) in HepG2 cells. Both extracts inhibited reactive oxygen species (ROS) generation by FFA-stimulated HepG2 cells. OF and CM also suppressed the mRNA expression of interleukin- (IL-) 1ß, IL-6, IL-8, tumor necrosis factor- (TNF-) α, and transforming growth factor- (TGF-) ß by HepG2 cells treated with conditioned medium derived from lipopolysaccharide-treated THP-1 monocytes. Furthermore, OF and CM effectively inhibited oleate-induced cellular lipid accumulation, TGF-ß secretion, and overexpression of fibronectin in mesangial cells. In conclusion, OF and CM possess hepatoprotective activity by inhibiting hepatic fat load and inflammation and renal protection by preventing FFA-induced mesangial extracellular matrix formation.

Wild Bitter Melon Leaf Extract Inhibits Porphyromonas gingivalis-Induced Inflammation: Identification of Active Compounds through Bioassay-Guided Isolation.

Porphyromonas gingivalis has been identified as one of the major periodontal pathogens. Activity-directed fractionation and purification processes were employed to identify the anti-inflammatory active compounds using heat-killed P. gingivalis-stimulated human monocytic THP-1 cells in vitro. Five major fractions were collected from the ethanol/ethyl acetate extract of wild bitter melon (Momordica charantia Linn. var. abbreviata Ser.) leaves and evaluated for their anti-inflammatory activity against P. gingivalis. Among the test fractions, Fraction 5 effectively decreased heat-killed P. gingivalis-induced interleukin (IL)-8 and was subjected to separation and purification by using chromatographic techniques. Two cucurbitane triterpenoids were isolated from the active fraction and identified as 5ß,19-epoxycucurbita-6,23-diene-3ß,19,25-triol (1) and 3ß,7ß,25-trihydroxycucurbita-5,23-dien-19-al (2) by comparing spectral data. Treatments of both compounds in vitro potently suppressed P. gingivalis-induced IL-8, IL-6, and IL-1ß levels and the activation of mitogen-activated protein kinase (MAPK) in THP-1 cells. Both compounds effectively inhibited the mRNA levels of IL-6, tumor necrosis factor (TNF)-α, and cyclooxygenase (COX)-2 in P. gingivalis-stimulated gingival tissue of mice. These findings imply that 5ß,19-epoxycucurbita-6,23-diene-3ß,19,25-triol and 3ß,7ß,25-trihydroxycucurbita-5,23-dien-19-al could be used for the development of novel therapeutic approaches against P. gingivalis infections.

Flavones Isolated from Scutellariae radix Suppress Propionibacterium Acnes-Induced Cytokine Production In Vitro and In Vivo.

Scutellariae radix, the root of Scutellaria baicalensis, has long been applied in traditional formulations and modern herbal medications. Propionibacterium acnes (P. acnes) in follicles can trigger inflammation and lead to the symptom of inflammatory acnes vulgaris. This study was aimed at evaluating the effect of Scutellariae radix extract and purified components isolated from it on inflammation induced by P. acnes in vitro and in vivo. The results showed the ethyl acetate (EA) soluble fraction from the partition of crude ethanolic extract from Scutellariae radix inhibited P. acnes-induced interleukin IL-8 and IL-1ß production in human monocytic THP-1 cells. Seven flavones were isolated from the EA fraction by repeated chromatographies, and identified as 5,7-dihydroxy-6-methoxyflavone (FL1, oroxylin), 5,7-dihydroxy-8-methoxyflavone (FL2, wogonin), 5-hydroxy-7,8-dimethoxyflavone (FL3, 7-O-methylwogonin), 5,6'-dihydroxy-6,7,8,2'-tetramethoxy flavone (FL4, skullcapflavone II), 5,7,4'-trihydroxy-8-methoxyflavone (FL5), 5,2',6'-trihydroxy-7,8-dimethoxyflavone (FL6, viscidulin II), and 5,7,2',5'-tetrahydroxy-8,6'-dimethoxyflavone (FL7, ganhuangenin). They all significantly suppressed P. acnes-induced IL-8 and IL-1ß production in THP-1 cells, and FL2 exerted the strongest effect with half maximal inhibition (IC50) values of 8.7 and 4.9 µM, respectively. Concomitant intradermal injection of each of the seven flavones (20 µg) with P. acnes effectively attenuated P. acnes-induced ear swelling, and decreased the production of IL-6 and tumor necrosis factor-α in ear homogenates. Our results suggested that all the seven flavones can be potential therapeutic agents against P. acnes-induced skin inflammation.

Inhibitory effects of wild bitter melon leaf extract on Propionibacterium acnes-induced skin inflammation in mice and cytokine production in vitro.

Propionibacterium acnes is a key pathogen involved in acne inflammation. Wild bitter melon (WBM, Momordica charantia L. var. abbreviate Seringe) is consumed as both a vegetable and as folk medicine in Taiwan. We examined the inhibitory activity of the total phenolic extract (TPE) of WBM leaf on P. acnes-induced inflammatory responses in vivo and in vitro. Our data showed that TPE significantly attenuated P. acnes-induced ear swelling in mice along with microabscess. Flow cytometry analysis revealed that TPE treatment significantly decreased the migration of neutrophils and interleukin (IL)-1ß(+) populations in vivo. In P. acnes-stimulated human monocytic THP-1 cells, TPE suppressed the mRNA levels and production of IL-8, IL-1ß, and tumor necrosis factor (TNF)-αin vitro. In addition, TPE suppressed P. acnes-induced matrix metalloproteinase-9 levels. TPE blocked nuclear factor-κB (NF-κB) activation and inactivated mitogen-activated protein kinases (MAPK); these actions may partially account for its inhibitory effect on cytokine production. The quantitative HPLC analysis revealed gallic, chlorogenic, caffeic, ferulic, and cinnamic acids, myricetin, quercetin, luteolin, apigenin, and thymol in TPE. All these phenolics significantly suppressed P. acnes-induced IL-8 production in vitro. Our results suggest that WBM leaf extract effectively inhibits P. acnes-induced inflammatory responses and may be useful to relieve the inflammation of acne.

Rosmarinus officinalis extract suppresses Propionibacterium acnes-induced inflammatory responses.

Propionibacterium acnes is a key pathogen involved in the progression of acne inflammation. The development of a new agent possessing antimicrobial and anti-inflammatory activity against P. acnes is therefore of interest. In this study, we investigated the inhibitory effect of rosemary (Rosmarinus officinalis) extract on P. acnes-induced inflammation in vitro and in vivo. The results showed that ethanolic rosemary extract (ERE) significantly suppressed the secretion and mRNA expression of proinflammatory cytokines, including interleukin (IL)-8, IL-1ß, and tumor necrosis factor-α in P. acnes-stimulated monocytic THP-1 cells. In an in vivo mouse model, concomitant intradermal injection of ERE attenuated the P. acnes-induced ear swelling and granulomatous inflammation. Since ERE suppressed the P. acnes-induced nuclear factor kappa-B (NF-κB) activation and mRNA expression of Toll-like receptor (TLR) 2, the suppressive effect of ERE might be due, at least partially, to diminished NF-κB activation and TLR2-mediated signaling pathways. Furthermore, three major constituents of ERE, carnosol, carnosic acid, and rosmarinic acid, exerted different immumodulatory activities in vitro. In brief, rosmarinic acid significantly suppressed IL-8 production, while the other two compounds inhibited IL-1ß production. Further study is needed to explore the role of bioactive compounds of rosemary in mitigation of P. acnes-induced inflammation.

Wild bitter melon (Momordica charantia Linn. var. abbreviata Ser.) extract and its bioactive components suppress Propionibacterium acnes-induced inflammation.

In this study, we aimed to evaluate the inhibitory effect of wild bitter melons (WBM; Momordica charantia Linn. var. abbreviata Ser.) on Propionibacterium acnes-induced inflammation and to identify the bioactive components. Our results showed that ethyl acetate (EA) extract of WBM fruit in vitro potently suppressed pro-inflammatory cytokine and matrix metalloproteinase (MMP)-9 levels in P. acnes-stimulated THP-1 cells. Furthermore, concomitant intradermal injection of WBM EA extract in mice effectively attenuated P. acnes-induced ear swelling and granulomatous inflammation. To further investigate the bioactive components, we found that both saponifiable (S) and nonsaponifiable (NS) fractions of WBM EA extract significantly suppressed pro-inflammatory cytokine and MMP-9 levels. Phytol and lutein, identified in the NS fraction, also inhibited cytokine production. Moreover, S and NS fractions of EA extract, phytol and lutein, activated peroxisome proliferator-activated receptor (PPAR) α and ß in the transactivation assay. Our results suggested that PPARα or PPARγ signalling may contribute, at least in part, to the anti-inflammatory activity of WBM.

Evaluation of NO-suppressing activity of several Mediterranean culinary spices.

Excess nitric oxide (NO) produced by inducible nitric oxide synthase (iNOS) is implicated in the development of a number of diseases. Due to the absence of any natural specific enzymatic defense system in vivo, the consumption of certain foods which exhibit selective suppressive ability as regards NO overproduction might boost the host's protective effects against NO-mediated toxicity. Spices, rich in phenolics, are speculated conceivably to act as potential NO-scavengers or iNOS suppressors. The relative NO-suppressing activity of methanol extracts deriving from nine Mediterranean culinary spices was determined by measuring their inhibitory effect upon NO production for lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. In addition, the specifics of the suppressing mechanism were further explored. All of the spices tested, with the exception of clove, displayed a rather linear dose-dependent NO-suppressing effect without there appearing to exist any effect upon cell viability. Furthermore, the NO-suppressing capacity of certain spices was able to be ranked based upon their IC(50) (the concentration of spice extracts is required to cause 50% inhibition of NO production by LPS-activated RAW 264.7 cells), the ranking appearing as: rosemary (0.031%)>tarragon (0.052%)>cinnamon (0.059%)>oregano (0.106%)>basil (0.162%)>marjoram (0.236%)>allspice (0.269%)>and thyme (0.270%). Only cinnamon displayed excellent NO-scavenging ability, whereas all of the other spices demonstrated moderate to poor activities in this regard. Moreover, the inhibitory effect of tested spices upon the iNOS protein level was almost equivalent to their suppressive effect upon NO production. It would appear that inhibition of iNOS expression was the primary mechanism of action of spices as regards their exerting NO-suppressing activity.