Insights into phenolic compounds from microalgae: structural variety and complex beneficial activities from health to nutraceutics.

Phenolic compounds (PCs) are a family of secondary metabolites with recognized biological activities making them attractive for the biomedical "red" biotechnology. The development of the eco-sustainable production of natural bioactive metabolites requires using easy cultivable organisms, such as microalgae, which represents one of the most promising sources for biotechnological applications. Microalgae are photosynthetic organisms inhabiting aquatic systems, displaying high levels of biological and functional diversities, and are well-known producers of fatty acids and carotenoids. They are also rich in other families of bioactive molecules e.g. phenolic compounds. Microalgal PCs however are less investigated than other molecular components. This study aims to provide a state-of-art picture of the actual knowledge on microalgal phenolic compounds, reviewing information on the PC content variety and chemodiversity in microalgae, their environmental modulation, and we aim to report discuss data on PC biosynthetic pathways. We report the challenges of promoting microalgae as a relevant source of natural PCs, further enhancing the interests of microalgal "biofactories" for biotechnological applications (i.e. nutraceutical, pharmacological, or cosmeceutical products).

MMP-9 and IL-1ß as Targets for Diatoxanthin and Related Microalgal Pigments: Potential Chemopreventive and Photoprotective Agents.

Photochemoprevention can be a valuable approach to counteract the damaging effects of environmental stressors (e.g., UV radiations) on the skin. Pigments are bioactive molecules, greatly attractive for biotechnological purposes, and with promising applications for human health. In this context, marine microalgae are a valuable alternative and eco-sustainable source of pigments that still need to be taken advantage of. In this study, a comparative in vitro photochemopreventive effects of twenty marine pigments on carcinogenic melanoma model cell B16F0 from UV-induced injury was setup. Pigment modulation of the intracellular reactive oxygen species (ROS) concentration and extracellular release of nitric oxide (NO) was investigated. At the cell signaling level, interleukin 1-ß (IL-1ß) and matrix metallopeptidase 9 protein (MMP-9) protein expression was examined. These processes are known to be involved in the signaling pathway, from UV stress to cancer induction. Diatoxanthin resulted the best performing pigment in lowering MMP-9 levels and was able to strongly lower IL-1ß. This study highlights the pronounced bioactivity of the exclusively aquatic carotenoid diatoxanthin, among the others. It is suggested increasing research efforts on this molecule, emphasizing that a deeper integration of plant ecophysiological studies into a biotechnological context could improve the exploration and exploitation of bioactive natural products.

In Vitro Evaluation of Antioxidant Potential of the Invasive Seagrass Halophila stipulacea.

Marine organisms with fast growth rates and great biological adaptive capacity might have biotechnological interests, since ecological competitiveness might rely on enhanced physiological or biochemical processes' capability promoting protection, defense, or repair intracellular damages. The invasive seagrass Halophila stipulacea, a non-indigenous species widespread in the Mediterranean Sea, belongs to this category. This is the premise to investigate the biotechnological interest of this species. In this study, we investigated the antioxidant activity in vitro, both in scavenging reactive oxygen species and in repairing damages from oxidative stress on the fibroblast human cell line WI-38. Together with the biochemical analysis, the antioxidant activity was characterized by the study of the expression of oxidative stress gene in WI-38 cells in presence or absence of the H. stipulacea extract. Concomitantly, the pigment pool of the extracts, as well as their macromolecular composition was characterized. This study was done separately on mature and young leaves. Results indicated that mature leaves exerted a great activity in scavenging reactive oxygen species and repairing damages from oxidative stress in the WI-38 cell line. This activity was paralleled to an enhanced carotenoids content in the mature leaf extracts and a higher carbohydrate contribution to organic matter. Our results suggest a potential of the old leaves of H. stipulacea as oxidative stress damage protecting or repair agents in fibroblast cell lines. This study paves the way to transmute the invasive H. stipulacea environmental threat in goods for human health.

Probing the Therapeutic Potential of Marine Phyla by SPE Extraction.

The marine environment is potentially a prolific source of small molecules with significant biological activities. In recent years, the development of new chromatographic phases and the progress in cell and molecular techniques have facilitated the search for marine natural products (MNPs) as novel pharmacophores and enhanced the success rate in the selection of new potential drug candidates. However, most of this exploration has so far been driven by anticancer research and has been limited to a reduced number of taxonomic groups. In this article, we report a test study on the screening potential of an in-house library of natural small molecules composed of 285 samples derived from 57 marine organisms that were chosen from among the major eukaryotic phyla so far represented in studies on bioactive MNPs. Both the extracts and SPE fractions of these organisms were simultaneously submitted to three different bioassays-two phenotypic and one enzymatic-for cytotoxic, antidiabetic, and antibacterial activity. On the whole, the screening of the MNP library selected 11 potential hits, but the distribution of the biological results showed that SPE fractionation increased the positive score regardless of the taxonomic group. In many cases, activity could be detected only in the enriched fractions after the elimination of the bulky effect due to salts. On a statistical basis, sponges and molluscs were confirmed to be the most significant source of cytotoxic and antimicrobial products, but other phyla were found to be effective with the other therapeutic targets.

Challenging microalgal vitamins for human health.

BACKGROUND: Vitamins' deficiency in humans is an important threat worldwide and requires solutions. In the concept of natural biofactory for bioactive compounds production, microalgae represent one of the most promising targets filling many biotechnological applications, and allowing the development of an eco-sustainable production of natural bioactive metabolites. Vitamins are probably one of the cutting edges of microalgal diversity compounds. MAIN TEXT: Microalgae can usefully provide many of the required vitamins in humans, more than terrestrial plants, for instance. Indeed, vitamins D and K, little present in many plants or fruits, are instead available from microalgae. The same occurs for some vitamins B (B12, B9, B6), while the other vitamins (A, C, D, E) are also provided by microalgae. This large panel of vitamins diversity in microalgal cells represents an exploitable platform in order to use them as natural vitamins' producers for human consumption. This study aims to provide an integrative overview on vitamins content in the microalgal realm, and discuss on the great potential of microalgae as sources of different forms of vitamins to be included as functional ingredients in food or nutraceuticals for the human health. We report on the biological roles of vitamins in microalgae, the current knowledge on their modulation by environmental or biological forcing and on the biological activity of the different vitamins in human metabolism and health protection. CONCLUSION: Finally, we critically discuss the challenges for promoting microalgae as a relevant source of vitamins, further enhancing the interests of microalgal "biofactory" for biotechnological applications, such as in nutraceuticals or cosmeceuticals.

New In Vitro Model of Oxidative Stress: Human Prostate Cells Injured with 2,2-diphenyl-1-picrylhydrazyl (DPPH) for the Screening of Antioxidants.

The antioxidant activity of natural compounds consists in their ability to modulate gene and protein expression, thus inducing an integrated cell protective response and repair processes against oxidative stress. New screening tools and methodologies are crucial for the actual requirement of new products with antioxidant activity to boost endogenous oxidative stress responsive pathways, Reactive Oxygen Species (ROS) metabolism and immune system activity, preserving human health and wellness. In this study, we performed and tested an integrated oxidative stress analysis, using DPPH assay and PNT2 cells injured with DPPH. We firstly investigated the mechanism of action of the oxidising agent (DPPH) on PNT2 cells, studying the variation in cell viability, oxidative stress genes, inflammatory mediator and ROS levels. The results reveal that DPPH activated ROS production and release of Prostaglandin E2 in PNT2 at low and intermediate doses, while cells switched from survival to cell death signals at high doses of the oxidising agent. This new in vitro oxidative stress model was validated by using Trolox, ß-carotene and total extract of the green microalga Testraselmis suecica. Only the T. suecica extract can completely counteract DPPH-induced injury, since its chemical complexity demonstrated a multilevel protecting and neutralising effect against oxidative stress in PNT2.

Diatom-Derived Polyunsaturated Aldehydes Activate Similar Cell Death Genes in Two Different Systems: Sea Urchin Embryos and Human Cells.

Programmed cell death, such as apoptosis and autophagy, are key processes that are activated early on during development, leading to remodelling in embryos and homeostasis in adult organisms. Genomic conservation of death factors has been largely investigated in the animal and plant kingdoms. In this study, we analysed, for the first time, the expression profile of 11 genes involved in apoptosis (extrinsic and intrinsic pathways) and autophagy in sea urchin Paracentrotus lividus embryos exposed to antiproliferative polyunsaturated aldehydes (PUAs), and we compared these results with those obtained on the human cell line A549 treated with the same molecules. We found that sea urchins and human cells activated, at the gene level, a similar cell death response to these compounds. Despite the evolutionary distance between sea urchins and humans, we observed that the activation of apoptotic and autophagic genes in response to cytotoxic compounds is a conserved process. These results give first insight on death mechanisms of P. lividus death mechanisms, also providing additional information for the use of this marine organism as a useful in vitro model for the study of cell death signalling pathways activated in response to chemical compounds.

Cardiovascular Active Peptides of Marine Origin with ACE Inhibitory Activities: Potential Role as Anti-Hypertensive Drugs and in Prevention of SARS-CoV-2 Infection.

Growing interest in hypertension-one of the main factors characterizing the cardiometabolic syndrome (CMS)-and anti-hypertensive drugs raised from the emergence of a new coronavirus, SARS-CoV-2, responsible for the COVID19 pandemic. The virus SARS-CoV-2 employs the Angiotensin-converting enzyme 2 (ACE2), a component of the RAAS (Renin-Angiotensin-Aldosterone System) system, as a receptor for entry into the cells. Several classes of synthetic drugs are available for hypertension, rarely associated with severe or mild adverse effects. New natural compounds, such as peptides, might be useful to treat some hypertensive patients. The main feature of ACE inhibitory peptides is the location of the hydrophobic residue, usually Proline, at the C-terminus. Some already known bioactive peptides derived from marine resources have potential ACE inhibitory activity and can be considered therapeutic agents to treat hypertension. Peptides isolated from marine vertebrates, invertebrates, seaweeds, or sea microorganisms displayed important biological activities to treat hypertensive patients. Here, we reviewed the anti-hypertensive activities of bioactive molecules isolated/extracted from marine organisms and discussed the associated molecular mechanisms involved. We also examined ACE2 modulation in sight of SARS2-Cov infection prevention.

An In Vitro Model to Investigate the Role of Helicobacter pylori in Type 2 Diabetes, Obesity, Alzheimer's Disease and Cardiometabolic Disease.

Helicobacter pylori (Hp) is a Gram-negative bacterium colonizing the human stomach. Nuclear Magnetic Resonance (NMR) analysis of intracellular human gastric carcinoma cells (MKN-28) incubated with the Hp cell filtrate (Hpcf) displays high levels of amino acids, including the branched chain amino acids (BCAA) isoleucine, leucine, and valine. Polymerase chain reaction (PCR) Array Technology shows upregulation of mammalian Target Of Rapamycin Complex 1 (mTORC1), inflammation, and mitochondrial dysfunction. The review of literature indicates that these traits are common to type 2 diabetes, obesity, Alzheimer's diseases, and cardiometabolic disease. Here, we demonstrate how Hp may modulate these traits. Hp induces high levels of amino acids, which, in turn, activate mTORC1, which is the complex regulating the metabolism of the host. A high level of BCAA and upregulation of mTORC1 are, thus, directly regulated by Hp. Furthermore, Hp modulates inflammation, which is functional to the persistence of chronic infection and the asymptomatic state of the host. Finally, in order to induce autophagy and sustain bacterial colonization of gastric mucosa, the Hp toxin VacA localizes within mitochondria, causing fragmentation of these organelles, depletion of ATP, and oxidative stress. In conclusion, our in vitro disease model replicates the main traits common to the above four diseases and shows how Hp may potentially manipulate them.

Bioinformatics for Marine Products: An Overview of Resources, Bottlenecks, and Perspectives.

The sea represents a major source of biodiversity. It exhibits many different ecosystems in a huge variety of environmental conditions where marine organisms have evolved with extensive diversification of structures and functions, making the marine environment a treasure trove of molecules with potential for biotechnological applications and innovation in many different areas. Rapid progress of the omics sciences has revealed novel opportunities to advance the knowledge of biological systems, paving the way for an unprecedented revolution in the field and expanding marine research from model organisms to an increasing number of marine species. Multi-level approaches based on molecular investigations at genomic, metagenomic, transcriptomic, metatranscriptomic, proteomic, and metabolomic levels are essential to discover marine resources and further explore key molecular processes involved in their production and action. As a consequence, omics approaches, accompanied by the associated bioinformatic resources and computational tools for molecular analyses and modeling, are boosting the rapid advancement of biotechnologies. In this review, we provide an overview of the most relevant bioinformatic resources and major approaches, highlighting perspectives and bottlenecks for an appropriate exploitation of these opportunities for biotechnology applications from marine resources.

Potent Cytotoxic Analogs of Amphidinolides from the Atlantic Octocoral Stragulum bicolor.

Amphidinolides are cytotoxic macrolides produced by symbiotic unicellular microalgae of the genus Amphidinium. Here we describe the identification of four related molecules belonging to this macrolide family isolated from the invertebrate Stragulum bicolor. The new molecules, named amphidinolide PX1-PX3 and stragulin A (1⁻4), show an unprecedented carbon skeleton whose complete stereochemistry has been determined by spectroscopic and computational methods. Differences in the structures of these molecules modulate their biological activity in a panel of tumor cell lines, but the opened derivative stragulin (4) shows a very potent and specific cytotoxic activity (IC50 0.18 µM) against the aggressive human melanoma cell A2058.

The Marine Dinoflagellate Alexandrium minutum Activates a Mitophagic Pathway in Human Lung Cancer Cells.

Marine dinoflagellates are a valuable source of bioactive molecules. Many species produce cytotoxic compounds and some of these compounds have also been investigated for their anticancer potential. Here, we report the first investigation of the toxic dinoflagellate Alexandrium minutum as source of water-soluble compounds with antiproliferative activity against human lung cancer cells. A multi-step enrichment of the phenol⁻water extract yielded a bioactive fraction with specific antiproliferative effect (IC50 = 0.4 µg·mL-1) against the human lung adenocarcinoma cells (A549 cell line). Preliminary characterization of this material suggested the presence of glycoprotein with molecular weight above 20 kDa. Interestingly, this fraction did not exhibit any cytotoxicity against human normal lung fibroblasts (WI38). Differential gene expression analysis in A549 cancer cells suggested that the active fraction induces specific cell death, triggered by mitochondrial autophagy (mitophagy). In agreement with the cell viability results, gene expression data also showed that no mitophagic event was activated in normal cells WI38.

On the Neuroprotective Role of Astaxanthin: New Perspectives?

Astaxanthin is a carotenoid with powerful antioxidant and anti-inflammatory activity produced by several freshwater and marine microorganisms, including bacteria, yeast, fungi, and microalgae. Due to its deep red-orange color it confers a reddish hue to the flesh of salmon, shrimps, lobsters, and crayfish that feed on astaxanthin-producing organisms, which helps protect their immune system and increase their fertility. From the nutritional point of view, astaxanthin is considered one of the strongest antioxidants in nature, due to its high scavenging potential of free radicals in the human body. Recently, astaxanthin is also receiving attention for its effect on the prevention or co-treatment of neurological pathologies, including Alzheimer and Parkinson diseases. In this review, we focus on the neuroprotective properties of astaxanthin and explore the underlying mechanisms to counteract neurological diseases, mainly based on its capability to cross the blood-brain barrier and its oxidative, anti-inflammatory, and anti-apoptotic properties.

Food Modulation Controls Astaxanthin Accumulation in Eggs of the Sea Urchin Arbacia lixula.

The carotenoid astaxanthin has strong antioxidant properties with beneficial effects for various degenerative diseases. This carotenoid is produced by some microalgae species when cultivated in particular conditions, and, interestingly, it is a predominant carotenoid in aquatic animals throughout a broad range of taxa. Recently, astaxanthin was detected in the eggs of the sea urchin Arbacia lixula in relevant concentrations when this organism was maintained in culture. These results have paved the way for deeper research into astaxanthin production by this species, particularly in regards to how astaxanthin production can be modulated by diet. Results showed that the highest content of astaxanthin in eggs was observed in sea urchins fed on a diet enriched with Spirulina platensis. This result was confirmed by the high antioxidant activity recorded in the egg extracts of these animals. Our results suggest that (i) the sea urchin A. lixula is able to synthesize astaxanthin from precursors obtained from food, and (ii) it is possible to modulate the astaxanthin accumulation in sea urchin eggs by modifying the proportions of different food ingredients provided in their diet. This study demonstrates the large potential of sea urchin cultivation for the eco-sustainable production of healthy supplements for nutraceutical applications.

The Sea Urchin Arbacia lixula: A Novel Natural Source of Astaxanthin.

Several echinoderms, including sea urchins, are valuable sources of bioactive compounds but their nutraceutical potential is largely unexplored. In fact, the gonads of some sea urchin species contain antioxidants including carotenoids and polyhydroxylated naphthoquinones (PHNQ's), such as echinochrome A. Astaxanthin is known to have particular bioactivity for the prevention of neurodegenerative diseases. This carotenoid is produced by microalgae, while several marine invertebrates can bioaccumulate or synthetize it from metabolic precursors. We determined the carotenoid content and analyzed the bioactivity potential of non-harvested Atlantic-Mediterranean sea urchin Arbacia lixula. The comparison of methanol crude extracts obtained from eggs of farmed and wild specimens revealed a higher bioactivity in farmed individuals fed with a customized fodder. HPLC-analysis revealed a high concentration of astaxanthin (27.0 µg/mg), which was the only pigment observed. This study highlights the potential of farmed A. lixula as a new source of the active stereoisomer of astaxanthin.

Development and Application of a Novel SPE-Method for Bioassay-Guided Fractionation of Marine Extracts.

The biological diversity of marine habitats is a unique source of chemical compounds with potential use as pharmaceuticals, cosmetics and dietary supplements. However, biological screening and chemical analysis of marine extracts pose specific technical constraints and require adequate sample preparation. Here we report an improved method on Solid Phase Extraction (SPE) to fractionate organic extracts containing high concentration of salt that hampers the recovery of secondary metabolites. The procedure uses a water suspension to load the extracts on a poly(styrene-divynylbenzene)-based support and a stepwise organic solvent elution to effectively desalt and fractionate the organic components. The novel protocol has been tested on MeOH-soluble material from three model organisms (Reniera sarai, Dendrilla membranosa and Amphidinium carterae) and was validated on a small panel of 47 marine samples, including sponges and protists, within discovery programs for identification of immuno-stimulatory and anti-infective natural products.

On the human health benefits of microalgal phytohormones: An explorative in silico analysis.

Phytohormones represent a group of secondary metabolites with different chemical structures, in which belong auxins, cytokinins, gibberellins, or brassinosteroids. In higher plants, they cover active roles in growth or defense function, while their potential benefits for human health protection were noted for some phytohormones and little explored for many others. In this study, we developed a target fishing strategy on fifty-three selected naturally occurring phytohormones covering different families towards proteins involved in key cellular functions related to human metabolism and health protection/disease. This in silico analysis strategy aims to screen the potential human health-driven bioactivity of more than fifty phytohormones through the analysis of their interactions with specific targets. From this analysis, twenty-eight human targets were recovered. Some targets e.g., the proteins mitochondrial glutamate dehydrogenase (GLUD1) or nerve growth factor (NGF) bound many phytohormones, highlighting their involvement in amino acid metabolism and/or in the maintenance or survival of neurons. Conversely, some phytohormones specifically interacted with some proteins, e.g., SPRY domain-containing SOCS box protein 2 (SPSB2) or Inosine-5'-monophosphate dehydrogenase 1 (IMPDH1), both involved in human immune response. They were then investigated with a molecular docking analysis approach. Our bioprospecting study indicated that many phytohormones may endow human health benefits, with potential functional role in multiple cellular processes including immune response and cell cycle progression.

Biotechnological response curve of the cyanobacterium Spirulina subsalsa to light energy gradient.

BACKGROUND: Microalgae represent a suitable and eco-sustainable resource for human needs thanks to their fast growth ability, together with the great diversity in species and intracellular secondary bioactive metabolites. These high-added-value compounds are of great interest for human health or animal feed. The intracellular content of these valuable compound families is tightly associated with the microalgal biological state and responds to environmental cues, e.g., light. Our study develops a Biotechnological response curve strategy exploring the bioactive metabolites synthesis in the marine cyanobacterium Spirulina subsalsa over a light energy gradient. The Relative Light energy index generated in our study integrates the red, green and blue photon flux density with their relative photon energy. The Biotechnological response curve combined biochemical analysis of the macromolecular composition (total protein, lipid, and carbohydrate content), total sterols, polyphenols and flavonoids, carotenoids, phenolic compounds, vitamins (A, B1, B2, B6, B9, B12, C, D2, D3, E, H, and K1), phycobiliproteins, together with the antioxidant activity of the biomass as well as the growth ability and photosynthesis. RESULTS: Results demonstrated that light energy significantly modulate the biochemical status of the microalga Spirulina subsalsa revealing the relevance of the light energy index to explain the light-induced biological variability. The sharp decrease of the photosynthetic rate at high light energy was accompanied with an increase of the antioxidant network response, such as carotenoids, total polyphenols, and the antioxidant capacity. Conversely, low light energy favorized the intracellular content of lipids and vitamins (B2, B6, B9, D3, K1, A, C, H, and B12) compared to high light energy. CONCLUSIONS: Results of the Biotechnological response curves were discussed in their functional and physiological relevance as well as for the essence of their potential biotechnological applications. This study emphasized the light energy as a relevant tool to explain the biological responses of microalgae towards light climate variability, and, therefore, to design metabolic manipulation of microalgae.

The multifaceted nature of IL-10: regulation, role in immunological homeostasis and its relevance to cancer, COVID-19 and post-COVID conditions.

Interleukin-10 (IL-10) is a pleiotropic cytokine that has a fundamental role in modulating inflammation and in maintaining cell homeostasis. It primarily acts as an anti-inflammatory cytokine, protecting the body from an uncontrolled immune response, mostly through the Jak1/Tyk2 and STAT3 signaling pathway. On the other hand, IL-10 can also have immunostimulating functions under certain conditions. Given the pivotal role of IL-10 in immune modulation, this cytokine could have relevant implications in pathologies characterized by hyperinflammatory state, such as cancer, or infectious diseases as in the case of COVID-19 and Post-COVID-19 syndrome. Recent evidence proposed IL-10 as a predictor of severity and mortality for patients with acute or post-acute SARS-CoV-2 infection. In this context, IL-10 can act as an endogenous danger signal, released by tissues undergoing damage in an attempt to protect the organism from harmful hyperinflammation. Pharmacological strategies aimed to potentiate or restore IL-10 immunomodulatory action may represent novel promising avenues to counteract cytokine storm arising from hyperinflammation and effectively mitigate severe complications. Natural bioactive compounds, derived from terrestrial or marine photosynthetic organisms and able to increase IL-10 expression, could represent a useful prevention strategy to curb inflammation through IL-10 elevation and will be discussed here. However, the multifaceted nature of IL-10 has to be taken into account in the attempts to modulate its levels.

The Carotenoid Diatoxanthin Modulates Inflammatory and Angiogenesis Pathways In Vitro in Prostate Cancer Cells.

Xanthophylls, a group of carotenoids, have attracted attention as human health benefit compounds thanks to their functionality and bioavailability. The great antioxidant and anti-inflammatory abilities of diatoxanthin (Dt), a photoprotective xanthophyll synthetized by diatoms, were recently documented. This study investigates the capacity of Dt to intercept prostate cancer progression in vitro on different human cell lines, exploring its role against cancer proliferation and angiogenesis. Our results highlighted the chemopreventive role of Dt already at low concentration (44.1 pM) and suggest that the Dt-induced cancer cell death occurred through oxidative stress mechanisms. This hypothesis was supported by variations on the expression of key genes and proteins. Oxidative stress cell deaths (e.g., ferroptosis) are recently described types of cell death that are closely related to the pathophysiological processes of many diseases, such as tumors. Nonetheless, the interest of Dt was further strengthened by its ability to inhibit angiogenesis. The results are discussed considering the actual progress and requirements in cancer therapy, notably for prostate cancer.