Ester Pagano

Novel Insights into the Pharmacology of Flavonoids

Flavonoids are widely distributed secondary metabolites and currently consumed in large amounts in the daily diet. In this article, some of the most recent developments in flavonoid – and related polyphenolic compounds – pharmacology are discussed, with particular emphasis on very recent data, most of which are published in Phytotherapy Research, which highlight new aspects in flavonoid anti‐inflammatory, antilipidemic, antihyperglycemic, antiviral, hepatoprotective, gastric antiulcer, cardioprotective, neuroprotective, antioxidant and anticancer actions. These updated data confirm the well‐established diverse beneficial pharmacological actions and might support the perspective for a therapeutic use. Copyright

Beneficial effect of the non-psychotropic plant cannabinoid cannabigerol on experimental inflammatory bowel disease

Inflammatory bowel disease (IBD) is an incurable disease which affects millions of people in industrialized countries. Anecdotal and scientific evidence suggests that Cannabis use may have a positive impact in IBD patients. Here, we investigated the effect of cannabigerol (CBG), a non-psychotropic Cannabis-derived cannabinoid, in a murine model of colitis. Colitis was induced in mice by intracolonic administration of dinitrobenzene sulphonic acid (DNBS). Inflammation was assessed by evaluating inflammatory markers/parameters (colon weight/colon length ratio and myeloperoxidase activity), by histological analysis and immunohistochemistry; interleukin-1β, interleukin-10 and interferon-γ levels by ELISA, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) by western blot and RT-PCR; CuZn-superoxide dismutase (SOD) activity by a colorimetric assay. Murine macrophages and intestinal epithelial cells were used to evaluate the effect of CBG on nitric oxide production and oxidative stress, respectively. CBG reduced colon weight/colon length ratio, myeloperoxidase activity, and iNOS expression, increased SOD activity and normalized interleukin-1β, interleukin-10 and interferon-γ changes associated to DNBS administration. In macrophages, CBG reduced nitric oxide production and iNOS protein (but not mRNA) expression. Rimonabant (a CB1 receptor antagonist) did not change the effect of CBG on nitric oxide production, while SR144528 (a CB2 receptor antagonist) further increased the inhibitory effect of CBG on nitric oxide production. In conclusion, CBG attenuated murine colitis, reduced nitric oxide production in macrophages (effect being modulated by the CB2 receptor) and reduced ROS formation in intestinal epithelial cells. CBG could be considered for clinical experimentation in IBD patients.

Colon carcinogenesis is inhibited by the TRPM8 antagonist cannabigerol, a Cannabis-derived non-psychotropic cannabinoid

Cannabigerol (CBG) is a safe non-psychotropic Cannabis-derived cannabinoid (CB), which interacts with specific targets involved in carcinogenesis. Specifically, CBG potently blocks transient receptor potential (TRP) M8 (TRPM8), activates TRPA1, TRPV1 and TRPV2 channels, blocks 5-hydroxytryptamine receptor 1A (5-HT1A) receptors and inhibits the reuptake of endocannabinoids. Here, we investigated whether CBG protects against colon tumourigenesis. Cell growth was evaluated in colorectal cancer (CRC) cells using the 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide and 3-amino-7-dimethylamino-2-methylphenazine hydrochloride assays; apoptosis was examined by histology and by assessing caspase 3/7 activity; reactive oxygen species (ROS) production by a fluorescent probe; CB receptors, TRP and CCAAT/enhancer-binding protein homologous protein (CHOP) messenger RNA (mRNA) expression were quantified by reverse transcription-polymerase chain reaction; small hairpin RNA-vector silencing of TRPM8 was performed by electroporation. The in vivo antineoplastic effect of CBG was assessed using mouse models of colon cancer. CRC cells expressed TRPM8, CB1, CB2, 5-HT1A receptors, TRPA1, TRPV1 and TRPV2 mRNA. CBG promoted apoptosis, stimulated ROS production, upregulated CHOP mRNA and reduced cell growth in CRC cells. CBG effect on cell growth was independent from TRPA1, TRPV1 and TRPV2 channels activation, was further increased by a CB2 receptor antagonist, and mimicked by other TRPM8 channel blockers but not by a 5-HT1A antagonist. Furthermore, the effect of CBG on cell growth and on CHOP mRNA expression was reduced in TRPM8 silenced cells. In vivo, CBG inhibited the growth of xenograft tumours as well as chemically induced colon carcinogenesis. CBG hampers colon cancer progression in vivo and selectively inhibits the growth of CRC cells, an effect shared by other TRPM8 antagonists. CBG should be considered translationally in CRC prevention and cure.

Palmitoylethanolamide, a naturally occurring lipid, is an orally effective intestinal anti‐inflammatory agent

Palmitoylethanolamide (PEA) acts via several targets, including cannabinoid CB1 and CB2 receptors, transient receptor potential vanilloid type‐1 (TRPV1) ion channels, peroxisome proliferator‐activated receptor alpha (PPAR α) and orphan G protein‐coupled receptor 55 (GRR55), all involved in the control of intestinal inflammation. Here, we investigated the effect of PEA in a murine model of colitis.

Palmitoylethanolamide normalizes intestinal motility in a model of post‐inflammatory accelerated transit: involvement of CB1 receptors and TRPV1 channels

Palmitoylethanolamide (PEA), a naturally occurring acylethanolamide chemically related to the endocannabinoid anandamide, interacts with targets that have been identified in peripheral nerves controlling gastrointestinal motility, such as cannabinoid CB1 and CB2 receptors, TRPV1 channels and PPARα. Here, we investigated the effect of PEA in a mouse model of functional accelerated transit which persists after the resolution of colonic inflammation (post‐inflammatory irritable bowel syndrome).

Phytotherapy of Benign Prostatic Hyperplasia. A Minireview

Benign prostate hyperplasia (BPH) is a common condition affecting older men, with an incidence that is age‐dependent. Histological BPH, which typically develops after the age of 40 years, ranges in prevalence from >50% at 60 years to as high as 90% by 85 years of age. Typical symptoms include increased frequency of urination, nocturia, urgency, hesitancy, and weak urine stream. Conventional medicines used for the treatment of BPH include alpha blockers and 5‐alpha reductase inhibitors. This articles review the mode of action, the efficacy, and the safety, including herb‐drug interactions of the most common botanicals (Serenoa repens, Pygeum africanum, Urtica dioica, and Cucurbita pepo) and nutraceuticals (isoflavones, lycopene, selenium, and β‐Sitosterol) in controlling the lower urinary tract symptoms associated to BPH. Copyright

Inhibition of colon carcinogenesis by a standardized Cannabis sativa extract with high content of cannabidiol.

PURPOSE Colon cancer is a major public health problem. Cannabis-based medicines are useful adjunctive treatments in cancer patients. Here, we have investigated the effect of a standardized Cannabis sativa extract with high content of cannabidiol (CBD), here named CBD BDS, i.e. CBD botanical drug substance, on colorectal cancer cell proliferation and in experimental models of colon cancer in vivo. METHODS Proliferation was evaluated in colorectal carcinoma (DLD-1 and HCT116) as well as in healthy colonic cells using the MTT assay. CBD BDS binding was evaluated by its ability to displace [(3)H]CP55940 from human cannabinoid CB1 and CB2 receptors. In vivo, the effect of CBD BDS was examined on the preneoplastic lesions (aberrant crypt foci), polyps and tumours induced by the carcinogenic agent azoxymethane (AOM) as well as in a xenograft model of colon cancer in mice. RESULTS CBD BDS and CBD reduced cell proliferation in tumoral, but not in healthy, cells. The effect of CBD BDS was counteracted by selective CB1 and CB2 receptor antagonists. Pure CBD reduced cell proliferation in a CB1-sensitive antagonist manner only. In binding assays, CBD BDS showed greater affinity than pure CBD for both CB1 and CB2 receptors, with pure CBD having very little affinity. In vivo, CBD BDS reduced AOM-induced preneoplastic lesions and polyps as well as tumour growth in the xenograft model of colon cancer. CONCLUSIONS CBD BDS attenuates colon carcinogenesis and inhibits colorectal cancer cell proliferation via CB1 and CB2 receptor activation. The results may have some clinical relevance for the use of Cannabis-based medicines in cancer patients.

Genotoxicity assessment of some cosmetic and food additives

α-Hexylcinnamaldehyde (HCA) and p-tert-butyl-alpha-methylhydrocinnamic aldehyde (BMHCA) are synthetic aldehydes, characterized by a typical floral scent, which makes them suitable to be used as fragrances in personal care (perfumes, creams, shampoos, etc.) and household products, and as flavouring additives in food and pharmaceutical industry. The aldehydic structure suggests the need for a safety assessment for these compounds. Here, HCA and BMHCA were evaluated for their potential genotoxic risk, both at gene level (frameshift or base-substitution mutations) by the bacterial reverse mutation assay (Ames test), and at chromosomal level (clastogenicity and aneuploidy) by the micronucleus test. In order to evaluate a primary and repairable DNA damage, the comet assay has been also included. In spite of their potential hazardous chemical structure, a lack of mutagenicity was observed for both compounds in all bacterial strains tested, also in presence of the exogenous metabolic activator, showing that no genotoxic derivatives were produced by CYP450-mediated biotransformations. Neither genotoxicity at chromosomal level (i.e. clastogenicity or aneuploidy) nor single-strand breaks were observed. These findings will be useful in further assessing the safety of HCA and BMHCA as either flavour or fragrance chemicals.

The chemopreventive action of bromelain, from pineapple stem (Ananas comosus L.), on colon carcinogenesis is related to antiproliferative and proapoptotic effects

SCOPE Colorectal cancer is an important health problem across the world. Here, we investigated the possible antiproliferative/proapoptotic effects of bromelain (from the pineapple stem Ananas comosus L., family Bromeliaceae) in a human colorectal carcinoma cell line and its potential chemopreventive effect in a murine model of colon cancer. METHODS AND RESULTS Proliferation and apoptosis were evaluated in human colon adenocarcinoma (Caco-2) cells by the (3) H-thymidine incorporation assay and caspase 3/7 activity measurement, respectively. Extracellular signal-related kinase (ERK) and Akt expression were evaluated by Western blot analysis, reactive oxygen species production by a fluorimetric method. In vivo, bromelain was evaluated using the azoxymethane murine model of colon carcinogenesis. Bromelain reduced cell proliferation and promoted apoptosis in Caco-2 cells. The effect of bromelain was associated to downregulation of pERK1/2/total, ERK, and pAkt/Akt expression as well as to reduction of reactive oxygen species production. In vivo, bromelain reduced the development of aberrant crypt foci, polyps, and tumors induced by azoxymethane. CONCLUSION Bromelain exerts antiproliferative and proapoptotic effects in colorectal carcinoma cells and chemopreventive actions in colon carcinogenesis in vivo. Bromelain-containing foods and/or bromelain itself may represent good candidates for colorectal cancer chemoprevention.

Intestinal antispasmodic effects of Helichrysum italicum (Roth) Don ssp. italicum and chemical identification of the active ingredients

ETHNOPHARMACOLOGICAL RELEVANCE In the Mediterranean Area, the flowers of Helichrysum italicum ssp. italicum are a traditional remedy for the treatment of intestinal complaints and are used as herbal tea for curing digestive, stomachic and intestinal diseases. In order to find scientific evidence for the traditional utilization of this plant, the effect of an ethanolic extract of Helichrysum italicum was investigated by using in vivo and in vitro experimental models. Then, through bioassay-guided fractionation procedures, active component(s) were identified. MATERIAL AND METHODS Contractility in vitro was evaluated by stimulating the isolated ileum, in an organ bath, with acetylcholine and barium chloride; motility in vivo was evaluated by measuring upper gastrointestinal transit, both in control mice and in mice with experimental intestinal inflammation induced by croton oil. Chromatographic separation techniques such as HPLC and silica gel columns have yielded the active principles of Helichrysum italicum. RESULTS We found that the ethanolic extract of Helichrysum italicum ssp. italicum flowers elicited antispasmodic actions in the isolated mouse ileum and inhibited transit preferentially in the inflamed gut. A bioassay guided fractionation of the extract yielded the known compounds 12-acetoxytremetone (1) and 2,3-dihydro-2-[1-(hydroxymethyl)ethenyl]-5-benzofuranyl]-ethanone (2). CONCLUSION Present study supported the traditional use of Helichrysum italicum ssp. italicum flowers for intestinal complaints and through bioassay-guided fractionation procedures from the crude extract we showed that 12-acetoxytremetone (1) and 2,3-dihydro-2-[1-(hydroxymethyl)ethenyl]-5-benzofuranyl]-ethanone (2) acted in a synergistic way to produce an intestinal antispasmodic effect.