HPLC- and NMR-Based Chemical Profiling, Wound-Healing Potential, Anti-Inflammatory and Antibacterial Activities of Satureja pilosa (Lamiaceae), a Neglected Medicinal-Aromatic Herb.

Satureja pilosa Velen. (Lamiaceae) is a perennial and melliferous aromatic-medicinal subshrub which is range-restricted in adjacent parts of Greece and Bulgaria and locally in Italy, known in Northern Greece as wild oregano ("agriorigani") and traditionally collected from the wild for culinary purposes. Since the ethnopharmacological data and modern biological activities of Satureja spp. suggest promising applications in skin conditions, the present study aimed to investigate the hitherto unknown phenolic content of cultivated S. pilosa and its potential biological activities, focusing mainly on wound-healing and anti-inflammatory effects. An HPLC-PDA-MS-targeted phytochemical investigation, along with NMR, allowed for the isolation and characterization of the main constituents, resulting in 18 compounds. Representative extracts and purified compounds were tested for wound-healing activity on NIH/3T3 fibroblasts. The butanol extract exhibited a significantly higher cell migration rate (73.4%) compared to aqueous (50.6%) and methanolic (49.6%) ones, enhancing the cell migration more rapidly at both concentration levels, whilst rosmarinic acid was the most potent among the isolated compounds, with a migration rate of 64.0% at the concentration level of 10-5 mg/mL, followed by 3,4-dihydrophenyllactic acid (54.7%). Moreover, potential effects on endothelial activation processes were explored, including the leukocyte-endothelial cell interaction during inflammatory processes and the migratory capacity during angiogenic actions, since these processes are commonly associated with skin diseases. Finally, extracts and purified compounds demonstrated weak antibacterial potential against two important pathogens (Staphylococcus aureus and Pseudomonas aeruginosa), suggesting that further investigation is warrented.

The Pharmacological Potential of Novel Melittin Variants from the Honeybee and Solitary Bees against Inflammation and Cancer.

The venom of honeybees is composed of numerous peptides and proteins and has been used for decades as an anti-inflammatory and anti-cancer agent in traditional medicine. However, the bioactivity of specific biomolecular components has been evaluated for the predominant constituent, melittin. So far, only a few melittin-like peptides from solitary bee species have been investigated, and the molecular mechanisms of bee venoms as therapeutic agents remain largely unknown. Here, the preclinical pharmacological activities of known and proteo-transcriptomically discovered new melittin variants from the honeybee and more ancestral variants from phylogenetically older solitary bees were explored in the context of cancer and inflammation. We studied the effects of melittin peptides on cytotoxicity, second messenger release, and inflammatory markers using primary human cells, non-cancer, and cancerous cell lines. Melittin and some of its variants showed cytotoxic effects, induced Ca2+ signaling and inhibited cAMP production, and prevented LPS-induced NO synthesis but did not affect the IP3 signaling and pro-inflammatory activation of endothelial cells. Compared to the originally-described melittin, some phylogenetically more ancestral variants from solitary bees offer potential therapeutic modalities in modulating the in vitro inflammatory processes, and hindering cancer cell viability/proliferation, including aggressive breast cancers, and are worth further investigation.

Benefits of Ginger and Its Constituent 6-Shogaol in Inhibiting Inflammatory Processes.

Ginger (Zingiber officinale Roscoe) is widely used as medicinal plant. According to the Committee on Herbal Medicinal Products (HMPC), dried powdered ginger rhizome can be applied for the prevention of nausea and vomiting in motion sickness (well-established use). Beyond this, a plethora of pre-clinical studies demonstrated anti-cancer, anti-oxidative, or anti-inflammatory actions. 6-Shogaol is formed from 6-gingerol by dehydration and represents one of the main bioactive principles in dried ginger rhizomes. 6-Shogaol is characterized by a Michael acceptor moiety being reactive with nucleophiles. This review intends to compile important findings on the actions of 6-shogaol as an anti-inflammatory compound: in vivo, 6-shogaol inhibited leukocyte infiltration into inflamed tissue accompanied with reduction of edema swelling. In vitro and in vivo, 6-shogaol reduced inflammatory mediator systems such as COX-2 or iNOS, affected NFκB and MAPK signaling, and increased levels of cytoprotective HO-1. Interestingly, certain in vitro studies provided deeper mechanistic insights demonstrating the involvement of PPAR-γ, JNK/Nrf2, p38/HO-1, and NFκB in the anti-inflammatory actions of the compound. Although these studies provide promising evidence that 6-shogaol can be classified as an anti-inflammatory substance, the exact mechanism of action remains to be elucidated. Moreover, conclusive clinical data for anti-inflammatory actions of 6-shogaol are largely lacking.

Natural products as drugs and tools for influencing core processes of eukaryotic mRNA translation.

Eukaryotic protein synthesis is the highly conserved, complex mechanism of translating genetic information into proteins. Although this process is essential for cellular homoeostasis, dysregulations are associated with cellular malfunctions and diseases including cancer and diabetes. In the challenging and ongoing search for adequate treatment possibilities, natural products represent excellent research tools and drug leads for new interactions with the translational machinery and for influencing mRNA translation. In this review, bacterial-, marine- and plant-derived natural compounds that interact with different steps of mRNA translation, comprising ribosomal assembly, translation initiation and elongation, are highlighted. Thereby, the exact binding and interacting partners are unveiled in order to accurately understand the mode of action of each natural product. The pharmacological relevance of these compounds is furthermore assessed by evaluating the observed biological activities in the light of translational inhibition and by enlightening potential obstacles and undesired side-effects, e.g. in clinical trials. As many of the natural products presented here possess the potential to serve as drug leads for synthetic derivatives, structural motifs, which are indispensable for both mode of action and biological activities, are discussed. Evaluating the natural products emphasises the strong diversity of their points of attack. Especially the fact that selected binding partners can be set in direct relation to different diseases emphasises the indispensability of natural products in the field of drug development. Discovery of new, unique and unusual interacting partners again renders them promising tools for future research in the field of eukaryotic mRNA translation.

BNO 1095, a Standardized Dry Extract from the Fruits of Vitex agnus-castus, Impairs Angiogenesis-related Endothelial Cell Functions In Vitro.

BNO 1095, a standardized dry extract from the fruits of Vitex agnus-castus, represents an approved herbal medicinal product for the treatment of premenstrual syndrome. Angiogenesis, the formation of new blood vessels from pre-existing capillaries, plays a major role in physiological situations, such as wound healing or tissue growth in female reproductive organs, but it is also of great importance in pathophysiological conditions such as chronic inflammatory diseases or cancer. Angiogenesis is a highly regulated multi-step process consisting of distinct key events that can be influenced pharmacologically. Few studies suggested anti-angiogenic actions of V. agnus-castus fruit extracts in in vivo and ex vivo models. Here, we provide for the first time profound in vitro data on BNO 1095-derived anti-angiogenic effects focusing on distinct angiogenesis-related endothelial cell functions that are inevitable for the process of new blood vessel formation. We found that V. agnus-castus extract significantly attenuated undirected and chemotactic migration of primary human endothelial cells. Moreover, the extract efficiently inhibited endothelial cell proliferation and reduced the formation of tube-like structures on Matrigel. Of note, the treatment of endothelial cell spheroids almost blocked endothelial sprouting in a 3D collagen gel. Our data present new and detailed insights into the anti-angiogenic actions of BNO 1095 and, therefore, suggest a novel scope of potential therapeutic applications of the extract for which these anti-angiogenic properties are required.

l-Thyroxin and the Nonclassical Thyroid Hormone TETRAC Are Potent Activators of PPARγ.

Thyroid hormones (THs) operate numerous physiological processes through modulation of the nuclear thyroid hormone receptors and several other proteins. We report direct activation of the nuclear peroxisome proliferator-activated receptor gamma (PPARγ) and retinoid X receptor (RXR) by classical and nonclassical THs as another molecular activity of THs. The T4 metabolite TETRAC was the most active TH on PPARγ with nanomolar potency and binding affinity. We demonstrate that TETRAC promotes PPARγ/RXR signaling in cell-free, cellular, and in vivo settings. Simultaneous activation of the heterodimer partners PPARγ and RXR resulted in high dimer activation efficacy. Compared to fatty acids as known natural ligands of PPARγ and RXR, TETRAC differs markedly in its molecular structure and the PPARγ-TETRAC complex revealed a distinctive binding mode of the TH. Our observations suggest a potential connection of TH and PPAR signaling through overlapping ligand recognition and may hold implications for TH and PPAR pharmacology.

Best practice in research - Overcoming common challenges in phytopharmacological research.

BACKGROUND: The pharmacology, toxicology and pharmacokinetics of bioactive preparations derived from natural sources has become a flourishing field of research. However, researching complex extracts and natural products faces numerous challenges. More broadly in recent years the critique of pharmacological research, and specifically its design, the methods used and reporting has intensified. AIMS: This consensus document provides a perspective on what constitutes best practice in pharmacological research on bioactive preparations derived from natural sources, providing a perspective of what the leading specialist journals in the field consider as the core characteristics of good research. APPROACH ('METHODS'): The editors-in-chief of seven journals developed this best practice statement in an iterative process. A first draft of the guidelines (prepared by MH) was then discussed and amended by the other editors. OUTCOMES: Core to this contribution is a table which provides detailed advice including simple points like a use of appropriate controls and the full taxonomic validity of the material under investigation (see also below), to the relevance of the model for the question being researched (e.g., can specific in silico or in vitro models really address the species anti-inflammatory activity?). Therefore, obviously, researchers must pay detailed attention to reporting and discussing such studies. This information must be discussed critically (as much as it is possible based on the published papers) in terms of their scientific quality and validity. While these points are obvious, as editors, we are aware that they are often not properly implemented. CONCLUSION: We call for an approach which incorporates a careful design, meticulous execution and a detailed reporting of studies focusing on the pharmacology/bioactivity of bioactive preparations. Clearly testable research questions must be developed and investigated experimentally. As the founder of pharmacology Claude Bernard put it already in 1865: '…. either the experimenter's hypothesis will be disproved or it will be proved by experiment. When experiment disproves its preconceived ideas, the experimenter must discard or modify it.'

Pharmaco-epidemiological research on herbal medicinal products in the paediatric population: data from the PhytoVIS study.

In paediatrics, clinical study data are limited, especially on herbal medicinal products. To address this gap, 2063 datasets from the paediatric population were evaluated in the PhytoVIS data base. By screening for paediatric data, information on indication, gender, treatment, co-medication and tolerability were evaluated. The majority of patients was treated because of common cold, fever, digestive complaints, skin diseases, sleep disturbances and anxiety. The perceived effect of the therapy was rated in 84% of the patients as very good or good without adverse events. The data shed light on a still neglected field of phyto-pharmacotherapy by giving information on the use of herbal medicines in an unselected cohort of paediatric patients. The results confirm the good clinical effects and safety of herbal medicinal products in this patient population and show that they are widely used in Germany.What is Known:• In Germany, about 85% of children receive one or more herbal medicinal products per year.• Despite international initiatives to promote clinical research in paediatrics, there are still many gaps of knowledge in the use of drugs in paediatrics.What is New:• The PhytoVIS project evaluated 2063 data sets from the paediatric population using herbal medicinal products.• The majority of patients was treated because of common cold, fever, digestive complaints, skin diseases, sleep disturbances and anxiety, and 84% of the patients rated the therapy as very good or good without adverse events.

ELOM-080 as Add-On Treatment for Respiratory Tract Diseases - A Review of Clinical Studies Conducted in China.

ELOM-080, also known as Myrtol, represents a distillate of a mixture of 4 rectified essential oils: eucalyptus oil, sweet orange oil, myrtle oil, and lemon oil. ELOM-080 is an approved herbal medicinal product for the secretolytic therapy and facilitation of expectoration in acute and chronic bronchitis and for the secretolytic therapy of sinusitis. Its clinical efficacy has been reported by several randomized control trials. Interestingly, in the recent past, a considerable number of clinical studies on the use of ELOM-080 as add-on treatment of different respiratory tract diseases has been conducted and published in China. As these publications were only available in Chinese, the international attention in the literature was limited. Based on the translation of these studies into English, this review aims to provide a brief overview of the studies' major results, which contribute to the knowledge on the efficacy of ELOM-080 in the treatment of respiratory tract diseases: ELOM-080 was shown to be of great value as add-on treatment not only for the well-established indications bronchitis and sinusitis, but also for pharyngitis, asthma, chronic obstructive pulmonary disease, and, most importantly, otitis media. Besides this clinical evidence, this review also summarizes the great progress in deciphering the mode of action of ELOM-080 that has been made by Chinese publications.

Identification of Narciclasine as an in Vitro Anti-Inflammatory Component of Cyrtanthus contractus by Correlation-Based Metabolomics.

In this study, an extract from the bulbs of Cyrtanthus contractus showed strong anti-inflammatory activity in vitro. The extract was partially separated into 14 fractions and analyzed by ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry metabolomics, and the correlation coefficients were calculated between biological activities and metabolite levels. As a result, the top-scoring metabolite narciclasine (1) is proposed as the active principle of C. contractus. This was confirmed by comparing the biological effect of crude extract with that of an authentic standard.

New Knowledge About Old Drugs: The Anti-Inflammatory Properties of Cardiac Glycosides.

In the 19th century, cardio-active steroid glycosides, shortly cardiac glycosides, were scientifically established as drugs against heart failure. Their in vivo, cellular, and molecular actions as well as their predominant target, Na+-K+-ATPase, have been comprehensively investigated in the 20th century and the discovery of endogenous cardiac glycosides has fostered this research field. In the last years, however, results from clinical trials and meta-analyses have questioned their therapeutic value due to efficacy and safety issues. This has led to a considerable decline of their usage. Beyond the cardiovascular system, cardiac glycosides have been increasingly recognized as antitumor compounds and Na+-K+-ATPase has evolved into a promising drug target in oncology. A wealth of review articles exists that intensively discuss these topics. Surprisingly, the anti-inflammatory actions of cardiac glycosides, which were discovered in the 1960s, have so far hardly been perceived and have not yet been summarized. This review provides an overview of the in vivo and in vitro actions of cardiac glycosides on inflammatory processes and of the signaling mechanisms responsible for these effects: cardiac glycosides have been found to decrease inflammatory symptoms in different animal models of acute and chronic inflammation. Regarding the underlying mechanisms most research has focused on leukocytes. In these cells, cardiac glycosides primarily inhibit cell proliferation and the secretion of proinflammatory cytokines.

The Dual Edema-Preventing Molecular Mechanism of the Crataegus Extract WS 1442 Can Be Assigned to Distinct Phytochemical Fractions.

The hawthorn (Crataegus spp.) extract WS 1442 is used against mild forms of chronic heart failure. This disease is associated with endothelial barrier dysfunction and edema formation. We have recently shown that WS 1442 protects against this dysfunction by a dual mechanism: it both promotes endothelial barrier integrity by activation of a barrier-enhancing pathway (cortactin activation) and inhibits endothelial hyperpermeability by blocking a barrier disruptive pathway (calcium signaling). In this study, we aimed to identify the bioactive compounds responsible for these actions by using a bioactivity-guided fractionation approach. From the four fractions generated from WS 1442 by successive elution with water, 95 % ethanol, methanol, and 70 % acetone, only the water fraction was inactive, whereas the other three triggered a reduction of endothelial hyperpermeability. Analyses of intracellular calcium levels and cortactin phosphorylation were used as readouts to estimate the bioactivity of subfractions and isolated compounds. Interestingly, only the ethanolic fraction interfered with the calcium signaling, whereas only the methanolic fraction led to an activation of cortactin. Thus, the dual mode of action of WS 1442 could be clearly assigned to two distinct fractions. Although the identification of the calcium-active substance(s) was not successful, we could exclude an involvement of phenolic compounds. Cortactin activation, however, could be clearly attributed to oligomeric procyanidins with a distinct degree of polymerization. Taken together, our study provides the first approach to identify the active constituents of WS 1442 that address different cellular pathways leading to the inhibition of endothelial barrier dysfunction.

Narciclasine - an Amaryllidaceae Alkaloid with Potent Antitumor and Anti-Inflammatory Properties.

The isocarbostyril alkaloid narciclasine, also known as lycoricidinol, was discovered in Narcissus species (Amaryllidaceae) in 1967. A few years later, the 60S subunit of ribosomes, and thus protein biosynthesis, were shown to be directly targeted by narciclasine. Due to its selective and highly potent cytotoxic action on cancer cells, narciclasine was intensively investigated as an antitumor compound both in vitro and in vivo. However, narciclasine did not show a strong pharmacological activity in animal tumor models. During the last decade, new fascinating actions, mechanisms, and targets of narciclasine have emerged. This review intends to present a brief but comprehensive overview of these novel insights. Beneficial therapeutical actions have been reported particularly in brain tumor models. The translation elongation factor eEF1A, which does not only participate in protein biosynthesis but also in the regulation of the actin cytoskeleton, was discovered as new direct target. Moreover, narciclasine was found to trigger actin stress fiber formation via the activation of the small GTPase RhoA. Progress has also been made regarding the pharmacokinetic characterization of the alkaloid. The synthesis of a great number of narciclasine derivatives led to a substantial understanding of its pharmacophore and of the structure-activity relationships. However, an optimized compound did not result from these efforts. Most importantly, a new field of indication has emerged: Narciclasine was proven to exert profound anti-inflammatory actions in vivo. Taken together, there has been a strong advance in the preclinical knowledge about the alkaloid. Nevertheless, narciclasine has not been tested in human clinical trials up to now.

Revisited anti-inflammatory activity of matricine in vitro: Comparison with chamazulene.

The proazulene matricine (1) is present in chamomile flower heads and has been proven to exhibit strong in vivo anti-inflammatory activity. In contrast to other secondary metabolites in chamomile preparations like its degradation product chamazulene (2), no plausible targets have been found to explain this activity. Therefore we revisited 1 regarding its in vitro anti-inflammatory activity in cellular and molecular studies. Using ICAM-1 as a marker for NF-κB activation, it was shown that ICAM-1 protein expression induced by TNF-α and LPS, but not by IFN-γ, was remarkably inhibited by 1 in endothelial cells (HMEC-1). Inhibition was concentration-dependent in a micromolar range (10-75 µM) and did not involve cytotoxic effects. At 75 µM expression of the adhesion molecule ICAM-1 was down to 52.7 ± 3.3% and 20.4 ± 1.8% of control in TNF-α and LPS-stimulated HMEC-1, respectively. In contrast, 2 showed no activity. Quantitative RT-PCR experiments revealed that TNF-α-induced expression of the ICAM-1 gene was also reduced by 1 in a concentration-dependent manner, reaching 32.3 ± 6.2% of control at 100 µM matricine. Additional functional assays (NF-κB promotor activity and cytoplasm to nucleus translocation) confirmed the inhibitory effect of 1 on NF-κB signaling. Despite the fact that 1 lacks an α,ß-unsaturated carbonyl and is thus not able to act via a Michael reaction with electron rich SH groups of functional biological molecules, data gave strong evidence that 1 inhibits NF-κB transcriptional activity in endothelial cells by an hitherto unknown mechanism and this may contribute to its well-known anti-inflammatory activity in vivo.

Evidence-Based Phytotherapy in Europe: Where Do We Stand?

Medicinal plants represent the oldest source of pharmacotherapy used by mankind. A considerable number of traditional systems of medicine (folk medicine) have emerged over the last millennia under different cultural conditions. Even nowadays, the majority of people in less developed countries have to rely on herbal remedies as primary health care. Based on scientific and technical progress, the options to produce high quality herbal medicinal products have been largely improved in the last decades. The acceptance of phytotherapy as a "natural and mild alternative" to synthetic drugs is very high within the general public in developed countries and, from a global perspective, sales figures of herbal medicines are constantly rising. However, we still face many issues in this field. In contrast to the popularity of herbal medicinal products, physicians and their respective societies often have a very critical view of them. Besides dogmatic obstacles, this is based on the frequently missing clinical trials that clearly demonstrate their efficacy and/or safety. This perspective discusses the reasons and implications of the lack of scientific evidence and also of the wrong understanding of the principles of rational phytotherapy.