Exploring the Bioactive Potential of Pisolithus (Basidiomycota): Comprehensive Insights into Antimicrobial, Anticancer, and Antioxidant Properties for Innovative Applications.

Addressing pressing health concerns, modern medical research seeks to identify new antimicrobials to combat drug resistance, novel molecules for cancer treatment, and antioxidants for inflammation-related diseases. Pisolithus (Basidiomycota) is a ubiquitous and widely distributed fungal genus in forest ecosystems, known for establishing ectomycorrhizal associations with a range of host plants, enhancing their growth, and conferring protection against biotic and abiotic stresses. Beyond ecological applications, Pisolithus yields bioactive compounds with medicinal potential. This comprehensive review explores the transversal biological activity of Pisolithus fungi, aiming to provide a thorough overview of their antimicrobial, anticancer, and antioxidant potential. The focus is on elucidating bioactive compounds within Pisolithus to trigger further research for innovative applications. Compounds from Pisolithus displayed antimicrobial activity against a broad spectrum of microorganisms, including antibiotic-resistant bacteria. The efficacy of Pisolithus-derived compounds matched established medications, emphasizing their therapeutic potential. In anticancer research, the triterpene pisosterol stood out with documented cytotoxicity against various cancer cell lines, showcasing promise for novel anticancer therapies. Pisolithus was also recognized as a potential source of antioxidants, with basidiocarps exhibiting high antioxidant activity. In vivo validation and comprehensive studies on a broader range of compounds, together with mechanistic insights into the mode of action of Pisolithus-derived compounds, are compelling areas for future research.

Reduction of neutral lipid reservoirs, bioconversion and untargeted metabolomics reveal distinct roles for vitamin K isoforms on lipid metabolism.

Vitamin K isoforms are known as co-factors for the synthesis of blood-clotting proteins, but several other bioactivities were reported. In this work, we isolated a vitamin K1-analogue (OH-PhQ) from the cyanobacterium Tychonema sp. LEGE 07196 with lipid reducing activity. OH-PhQ reduced neutral lipid reservoirs with an EC50 value of 31 µM after 48 h exposure in zebrafish larvae, while other vitamin K isoforms had EC50 values of 21.1 µM (K2) and 1.2 µM (K3). No lipid reducing activity was observed for K1 up to 50 µM. The presence of vitamin K isoforms was studied in zebrafish after exposure (OH-PhQ, K1, K2 and K3), and a clear preference for bioconversion was observed to retain K1 and OH-PhQ. Untargeted metabolomics revealed different biological effects for vitamin K isoforms on the subclass and metabolite level, but similarities were present on the compound class level, particularly on the regulation of glycerophospholipids. Our data showed for the first time a lipid reducing activity of OH-PhQ and performed a comparative analysis of vitamin K isoforms, which could be important for the development of future nutraceuticals or food supplements.

Tracing the Path between Mushrooms and Alzheimer's Disease-A Literature Review.

Alzheimer's disease (AD) is well-known among neurodegenerative diseases for the decline of cognitive functions, making overall daily tasks difficult or impossible. The disease prevails as the most common form of dementia and remains without a well-defined etiology. Being considered a disease of multifactorial origin, current targeted treatments have only managed to reduce or control symptoms, and to date, only two drugs are close to being able to halt its progression. For decades, natural compounds produced by living organisms have been at the forefront of research for new therapies. Mushrooms, which are well-known for their nutritional and medicinal properties, have also been studied for their potential use in the treatment of AD. Natural products derived from mushrooms have shown to be beneficial in several AD-related mechanisms, including the inhibition of acetylcholinesterase (AChE) and ß-secretase (BACE 1); the prevention of amyloid beta (Aß) aggregation and neurotoxicity; and the prevention of Tau expression and aggregation, as well as antioxidant and anti-inflammatory potential. Several studies in the literature relate mushrooms to neurodegenerative diseases. However, to the best of our knowledge, there is no publication that summarizes only AD data. In this context, this review aims to link the therapeutic potential of mushrooms to AD by compiling the anti-AD potential of different mushroom extracts or isolated compounds, targeting known AD-related mechanisms.

Corrigendum: Antiproliferative effects of the natural oxadiazine Nocuolin A are associated with impairment of mitochondrial oxidative phosphorylation.

[This corrects the article DOI: 10.3389/fonc.2019.00224.].

A Multiplex Molecular Cell-Based Sensor to Detect Ligands of PPARs: An Optimized Tool for Drug Discovery in Cyanobacteria.

Cyanobacteria produce a wealth of secondary metabolites. Since these organisms attach fatty acids into molecules in unprecedented ways, cyanobacteria can serve as a novel source for bioactive compounds acting as ligands for Peroxisome Proliferator-Activated Receptors (PPAR). PPARs (PPARα, PPARß/δ and PPARγ) are ligand-activated nuclear receptors, involved in the regulation of various metabolic and cellular processes, thus serving as potential drug targets for a variety of pathologies. Yet, given that PPARs' agonists can have pan-, dual- or isoform-specific action, some controversy has been raised over currently approved drugs and their side effects, highlighting the need for novel molecules. Here, we expand and validate a cell-based PPAR transactivation activity biosensor, and test it in a screening campaign to guide drug discovery. Biosensor upgrades included the use of different reporter genes to increase signal intensity and stability, a different promoter to modulate reporter gene expression, and multiplexing to improve efficiency. Sensor's limit of detection (LOD) ranged from 0.36-0.89 nM in uniplex and 0.89-1.35 nM in multiplex mode. In triplex mode, the sensor's feature screening, a total of 848 fractions of 96 cyanobacteria extracts were screened. Hits were confirmed in multiplex mode and in uniplex mode, yielding one strain detected to have action on PPARα and three strains to have dual action on PPARα and -ß.

Discovery and Heterologous Expression of Microginins from Microcystis aeruginosa LEGE 91341.

Microginins are a large family of cyanobacterial lipopeptide protease inhibitors. A hybrid polyketide synthase/non-ribosomal peptide synthetase biosynthetic gene cluster (BGC) found in several microginin-producing strains─mic─was proposed to encode the production of microginins, based on bioinformatic analysis. Here, we explored a cyanobacterium, Microcystis aeruginosa LEGE 91341, which contains a mic BGC, to discover 12 new microginin variants. The new compounds contain uncommon amino acids, namely, homophenylalanine (Hphe), homotyrosine (Htyr), or methylproline, as well as a 3-aminodecanoic acid (Ada) residue, which in some variants was chlorinated at its terminal methyl group. We have used direct pathway cloning (DiPaC) to heterologously express the mic BGC from M. aeruginosa LEGE 91341 in Escherichia coli, which led to the production of several microginins. This proved that the mic BGC is, in fact, responsible for the biosynthesis of microginins and paves the way to accessing new variants from (meta)genome data or through pathway engineering.

Uncovering the Bioactive Potential of a Cyanobacterial Natural Products Library Aided by Untargeted Metabolomics.

The Blue Biotechnology and Ecotoxicology Culture Collection (LEGE-CC) holds a vast number of cyanobacteria whose chemical richness is still largely unknown. To expedite its bioactivity screening we developed a natural products library. Sixty strains and four environmental samples were chromatographed, using a semiautomatic HPLC system, yielding 512 fractions that were tested for their cytotoxic activity against 2D and 3D models of human colon carcinoma (HCT 116), and non-cancerous cell line hCMEC/D3. Six fractions showed high cytotoxicity against 2D and 3D cell models (group A), and six other fractions were selected by their effects on 3D cells (group B). The metabolome of each group was organized and characterized using the MolNetEnhancer workflow, and its processing with MetaboAnalyst allowed discrimination of the mass features with the highest fold change, and thus the ones that might be bioactive. Of those, mass features without precedented identification were mostly found in group A, indicating seven possible novel bioactive molecules, alongside in silico putative annotation of five cytotoxic compounds. Manual dereplication of group B tentatively identified nine pheophytin and pheophorbide derivatives. Our approach enabled the selection of 7 out of 60 cyanobacterial strains for anticancer drug discovery, providing new data concerning the chemical composition of these cyanobacteria.

Discovery of Cyanobacterial Natural Products Containing Fatty Acid Residues*.

In recent years, extensive sequencing and annotation of bacterial genomes has revealed an unexpectedly large number of secondary metabolite biosynthetic gene clusters whose products are yet to be discovered. For example, cyanobacterial genomes contain a variety of gene clusters that likely incorporate fatty acid derived moieties, but for most cases we lack the knowledge and tools to effectively predict or detect the encoded natural products. Here, we exploit the apparent absence of a functional ß-oxidation pathway in cyanobacteria to achieve efficient stable-isotope-labeling of their fatty acid derived lipidome. We show that supplementation of cyanobacterial cultures with deuterated fatty acids can be used to easily detect natural product signatures in individual strains. The utility of this strategy is demonstrated in two cultured cyanobacteria by uncovering analogues of the multidrug-resistance reverting hapalosin, and novel, cytotoxic, lactylate-nocuolin A hybrids-the nocuolactylates.

Exploitation of Filamentous and Picoplanktonic Cyanobacteria for Cosmetic Applications: Potential to Improve Skin Structure and Preserve Dermal Matrix Components.

The use of natural products in skin care formulations gained interest as a concern for modern societies. The undesirable side effects of synthetic COMPOUNDS, as well as the associated environmental hazards, have driven investigation on photosynthetic organisms as sustainable sources of effective and environmentally friendly ingredients. The use of natural extracts in cosmetics has been highlighted and, along with plants and algae, cyanobacteria have come into focus. Due to their low culture demands, high grow rates and ability to produce a wide variability of bioactive metabolites, cyanobacteria emerged as an economic and sustainable base for the cosmetic industry. In this study, we evaluated the potential of ethanol extracts of picocyanobacteria strains of the genera Cyanobium and Synechocystis and filamentous strains of the genera Nodosilinea, Phormidium and Tychonema for skin applications, with focus in the field of anti-aging. The extracts were analyzed for their pigment profile, phenolic content, antioxidant potential, cytotoxicity against keratinocytes (HaCat), fibroblasts (3T3L1), endothelial cells (hCMEC/D3) and capacity to inhibit hyaluronidase (HAase). The total carotenoid content ranged from 118.69 to 383.89 µg g-1 of dry biomass, and the total phenolic content from 1.07 to 2.45 mg GAE g-1. Identified carotenoids consisted of zeaxanthin, lutein, canthaxanthin, echinenone and ß-carotene, with zeaxanthin and lutein being the most representative (49.82 and 79.08 µg g-1, respectively). The highest antioxidant potential was found for Phormidium sp. LEGE 05292 and Tychonema sp. LEGE 07196 for superoxide anion radical (O2•-) scavenging (IC50 of 822.70 and 924 µg mL-1, respectively). Low or no cytotoxicity was registered. Regarding HAase inhibition, Tychonema sp. LEGE 07196 and Cyanobium sp. LEGE 07175 showed the best IC50 (182.74 and 208.36 µg mL-1, respectively). In addition, an increase in fibroblast proliferation was registered with these same strains. From this work, the ethanol extracts of the species Tychonema sp. and Cyanobium sp. are particularly interesting for their potential application in anti-aging formulations, once they stimulated fibroblast proliferation and inhibit hyaluronic acid digestion.

Antiproliferative Effects of the Natural Oxadiazine Nocuolin A Are Associated With Impairment of Mitochondrial Oxidative Phosphorylation.

Natural products are interesting sources for drug discovery. The natural product oxadiazine Nocuolin A (NocA) was previously isolated from the cyanobacterial strain Nodularia sp. LEGE 06071 and here we examined its cytotoxic effects against different strains of the colon cancer cell line HCT116 and the immortalized epithelial cell line hTERT RPE-1. NocA was cytotoxic against colon cancer cells and immortalized cells under conditions of exponential growth but was only weakly active against non-proliferating immortalized cells. NocA induced apoptosis by mechanism(s) resistant to overexpression of BCL family members. Interestingly, NocA affected viability and induced apoptosis of HCT116 cells grown as multicellular spheroids. Analysis of transcriptome profiles did not match signatures to any known compounds in CMap but indicated stress responses and induction of cell starvation. Evidence for autophagy was observed, and a decrease in various mitochondrial respiration parameter within 1 h of treatment. These results are consistent with previous findings showing that nutritionally compromised cells in spheroids are sensitive to impairment of mitochondrial energy production due to limited metabolic plasticity. We conclude that the antiproliferative effects of NocA are associated with effects on mitochondrial oxidative phosphorylation.

Gloeothece sp. as a Nutraceutical Source-An Improved Method of Extraction of Carotenoids and Fatty Acids.

The nutraceutical potential of microalgae boomed with the exploitation of new species and sustainable extraction systems of bioactive compounds. Thus, a laboratory-made continuous pressurized solvent extraction system (CPSE) was built to optimize the extraction of antioxidant compounds, such as carotenoids and PUFA, from a scarcely studied prokaryotic microalga, Gloeothece sp. Following "green chemical principles" and using a GRAS solvent (ethanol), biomass amount, solvent flow-rate/pressure, temperature and solvent volume-including solvent recirculation-were sequentially optimized, with the carotenoids and PUFA content and antioxidant capacity being the objective functions. Gloeothece sp. bioactive compounds were best extracted at 60 °C and 180 bar. Recirculation of solvent in several cycles (C) led to an 11-fold extraction increase of ß-carotene (3C) and 7.4-fold extraction of C18:2 n6 t (5C) when compared to operation in open systems. To fully validate results CPSE, this system was compared to a conventional extraction method, ultrasound assisted extraction (UAE). CPSE proved superior in extraction yield, increasing total carotenoids extraction up 3-fold and total PUFA extraction by ca. 1.5-fold, with particular extraction increase of 18:3 n3 by 9.6-fold. Thus, CPSE proved to be an efficient and greener extraction method to obtain bioactive extract from Gloeothece sp. for nutraceutical purposes-with low levels of resources spent, while lowering costs of production and environmental impacts.

Cytotoxicity of portoamides in human cancer cells and analysis of the molecular mechanisms of action.

Portoamides are cyclic peptides produced and released by the cyanobacterial strain Phormidium sp. presumably to interfere with other organisms in their ecosystems ("allelopathy"). Portoamides were previously demonstrated to have an antiproliferative effect on human lung carcinoma cells, but the underlying mechanism of this activity has not been described. In the present work, the effects of portoamides on proliferation were examined in eight human cancer cell lines and two non-carcinogenic cell lines, and major differences in sensitivities were observed. To generate hypotheses with regard to molecular mechanisms of action, quantitative proteomics using 2D gel electrophoresis and MALDI-TOF/TOF were performed on the colon carcinoma cell line HT-29. The expression of proteins involved in energy metabolism (mitochondrial respiratory chain and pentose phosphate pathway) was found to be affected. The hypothesis of altered energy metabolism was tested in further experiments. Exposure to portoamides resulted in reduced cellular ATP content, likely due to decreased mitochondrial energy production. Mitochondrial hyperpolarization and reduced mitochondrial reductive capacity was observed in treated cells. Furthermore, alterations in the expression of peroxiredoxins (PRDX4, PRDX6) and components of proteasome subunits (PSB4, PSA6) were observed in portoamide-treated cells, but these alterations were not associated with detectable increases in oxidative stress. We conclude that the cytotoxic activity of portoamides is associated with disturbance of energy metabolism, and alterations in mitochondrial structure and function.

Obesity: The Metabolic Disease, Advances on Drug Discovery and Natural Product Research.

Obesity is a global health threat. OECD reported that more than half (52%) of the adult population in the European Union is overweight or obese. Obesity and obesity-related co-morbidities have deep negative effects on morbidity, mortality, professional and personal quality of life. Healthcare costs represent a negative impact of this disease, with an associated economic cost of 100 billion US$ per year in the United States. The most prescribed drugs for obesity treatment worldwide are orlistat, and phentermine/topiramate extended release, while the major prescribed drug for the same disease in the US are exenatide and dapagliflozin. The so far developed drugs, targeting weight loss, have a long history of malignant secondary effects. There is still a lack of efficient and safe drugs to treat obesity and related metabolic complications since in many cases cure cannot be reached by bariatric surgery or healthy lifestyle habits. Terrestrial and aquatic organisms are a promising source of valuable, bioactive compounds, often with interest for human health. Some of the natural compounds or organisms have been used for centuries by humans as traditional medicine foods. In this review, we give insights into the adipose tissue function and development, and the progress in traditional anti-obesity pharmacotherapy. A major focus is to highlight the state of the art of natural compounds with anti-obesity properties and their potential as candidates for drug development; an overview is given about natural compounds derived from different marine animal sources, cyanobacteria, marine phytoplankton, fungus or plants.

The conifer biomarkers dehydroabietic and abietic acids are widespread in Cyanobacteria.

Terpenes, a large family of natural products with important applications, are commonly associated with plants and fungi. The diterpenoids dehydroabietic and abietic acids are defense metabolites abundant in resin, and are used as biomarkers for conifer plants. We report here for the first time that the two diterpenoid acids are produced by members of several genera of cyanobacteria. Dehydroabietic acid was isolated from two cyanobacterial strains and its identity was confirmed spectroscopically. One or both of the diterpenoids were detected in the cells of phylogenetically diverse cyanobacteria belonging to four cyanobacterial 'botanical orders', from marine, estuarine and inland environments. Dehydroabietic acid was additionally found in culture supernatants. We investigated the natural role of the two resin acids in cyanobacteria using ecologically-relevant bioassays and found that the compounds inhibited the growth of a small coccoid cyanobacterium. The unexpected discovery of dehydroabietic and abietic acids in a wide range of cyanobacteria has implications for their use as plant biomarkers.

Cytotoxicity Induced by Extracts of Pisolithus tinctorius Spores on Human Cancer and Normal Cell Lines-Evaluation of the Anticancer Potential.

Fungi have been considered a potential source of natural anticancer drugs. However, studies on these organisms have mainly focused on compounds present in the sporocarp and mycelium. The aim of this study was to assess the anticancer potential of fungal spores using a bioassay-guided fractionation with cancer and normal cell lines. Crude extracts from spores of the basidiomycetous fungus Pisolithus tinctorius were prepared using five solvents/solvent mixtures in order to select the most effective crude extraction procedure. A dichloromethane/methanol (DCM/MeOH) mixture was found to produce the highest extraction yield, and this extract was fractionated into 11 fractions. Crude extracts and fractions were assayed for cytotoxicity in the human osteocarcinoma cell line MG63, the human breast carcinoma cell line T47D, the human colon adenocarcinoma cell line RKO, and the normal human brain capillary endothelial cell line hCMEC/D3. Cytotoxicity was assessed by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) reduction assay. The results showed a reduction in cancer cell viability of approximately 95% with 4 of 11 fractions without a significant reduction in viability of hCMEC/D3 cells. Data demonstrated that spores of P. tinctorius might serve as an interesting source of compounds with potential anticancer properties.

Structural insight on the activity of type 1 angiotensin II peptide antagonists using MD simulations.

Angiotensin II (AngII) is an octapeptide hormone, which plays a very important role in the blood pressure control mechanism. The excess production of this hormone is one of the main causes of hypertension illness. The antagonists for AngII At1 receptor constitute some of the most effective antihypertension drugs. In this work, both tested type1 AngII antagonists as well as new modeled antagonists (obtained by substitution of nonspecific amino acids by noncode residues (Sarcosine (Sar) and several Calpha, Calpha-dialkylglycines)) were simulated in dimethyl sulfoxide (DMSO) using molecular dynamics (MD). A number of common structural characteristics were identified on the active (and potentially active) simulated analogs, which seem to be correlated with their antagonistic activity. Two of the designed analogs were proposed as possible antagonists.

Molecular dynamics simulations of angiotensin II in aqueous and dimethyl sulfoxide environments.

Angiotensin II (Ang II) is an octapeptidic hormone, which plays an important role in the mechanisms of blood pressure control. In this work, extensive molecular dynamics (MD) simulations have been carried out on this peptide, both in aqueous and in dimethyl sulfoxide (DMSO) environments. Experimentally proposed models for the structure of angiotensin II in both environments are not consensual and the results obtained have provided some further insight about the structural properties of this hormone. In these simulations, the N-terminus of Ang II in the aqueous environment has been associated with a considerable larger flexibility than the correspondent C-terminus, but this was not found in the case of the DMSO environment. This is consistent with the assumption that the biological activity of Ang II is associated with its C-terminal residues embedded in a hydrophobic environment of its AT1 receptor. Other features detected in DMSO environment were an H(His6 imidazole)-O(Phe8 carboxylate) hydrogen bond and a salt-bridge structure involving the Asp1 and Arg2 side chains. An additional important conformational feature is the spatial proximity between Tyr4 and His6 in both water and DMSO environments. This molecular feature may trigger the interest for the synthetic chemists to apply rational design for the synthesis of novel AT1 antagonists.