Influence of electromagnetic fields on the circadian rhythm: Implications for human health and disease.

Living organisms have evolved within the natural electromagnetic fields (EMFs) of the earth which comprise the global atmospheric electrical circuit, Schumann resonances (SRs) and the geomagnetic field. Research suggests that the circadian rhythm, which controls several physiological functions in the human body, can be influenced by light but also by the earth's EMFs. Cyclic solar disturbances, including sunspots and seasonal weakening of the geomagnetic field, can affect human health, possibly by disrupting the circadian rhythm and downstream physiological functions. Severe disruption of the circadian rhythm increases inflammation which can induce fatigue, fever and flu-like symptoms in a fraction of the population and worsen existing symptoms in old and diseased individuals, leading to periodic spikes of infectious and chronic diseases. Possible mechanisms underlying sensing of the earth's EMFs involve entrainment via electrons and electromagnetic waves, light-dependent radical pair formation in retina cryptochromes, and paramagnetic magnetite nanoparticles. Factors such as electromagnetic pollution from wireless devices, base antennas and low orbit internet satellites, shielding by non-conductive materials used in shoes and buildings, and local geomagnetic anomalies may also affect sensing of the earth's EMFs by the human body and contribute to circadian rhythm disruption and disease development.

Ganoderma lucidum stimulates autophagy-dependent longevity pathways in Caenorhabditis elegans and human cells.

The medicinal fungus Ganoderma lucidum is used as a dietary supplement and health tonic, but whether it affects longevity remains unclear. We show here that a water extract of G. lucidum mycelium extends lifespan of the nematode Caenorhabditis elegans. The G. lucidum extract reduces the level of fibrillarin (FIB-1), a nucleolar protein that correlates inversely with longevity in various organisms. Furthermore, G. lucidum treatment increases expression of the autophagosomal protein marker LGG-1, and lifespan extension is abrogated in mutant C. elegans strains that lack atg-18, daf-16, or sir-2.1, indicating that autophagy and stress resistance pathways are required to extend lifespan. In cultured human cells, G. lucidum increases concentrations of the LGG-1 ortholog LC3 and reduces levels of phosphorylated mTOR, a known inhibitor of autophagy. Notably, low molecular weight compounds (<10 kDa) isolated from the G. lucidum water extract prolong lifespan of C. elegans and the same compounds induce autophagy in human cells. These results suggest that G. lucidum can increase longevity by inducing autophagy and stress resistance.

Emerging use of senolytics and senomorphics against aging and chronic diseases.

Senescence is a state of cell cycle arrest that plays an important role in embryogenesis, wound healing and protection against cancer. Senescent cells also accumulate during aging and contribute to the development of age-related disorders and chronic diseases, such as atherosclerosis, type 2 diabetes, osteoarthritis, idiopathic pulmonary fibrosis, and liver disease. Molecules that induce apoptosis of senescent cells, such as dasatinib, quercetin, and fisetin, produce health benefits and extend lifespan in animal models. We describe here the mechanism of action of senolytics and senomorphics, many of which are derived from plants and fungi. We also discuss the possibility of using such compounds to delay aging and treat chronic diseases in humans.

Ectopic calcification and formation of mineralo-organic particles in arteries of diabetic subjects.

Vascular calcification occurs in various diseases including atherosclerosis, chronic kidney disease and type 2 diabetes but the mechanism underlying mineral deposition remains incompletely understood. Here we examined lower limb arteries of type 2 diabetes subjects for the presence of ectopic calcification and mineral particles using histology, electron microscopy and spectroscopy analyses. While arteries of healthy controls showed no calcification following von Kossa staining, arteries from 83% of diabetic individuals examined (19/23) revealed microscopic mineral deposits, mainly within the tunica media. Mineralo-organic particles containing calcium phosphate and proteins such as albumin, fetuin-A and apolipoprotein-A1 were detected in calcified arteries. Ectopic calcification and mineralo-organic particles were observed in a majority of diabetic patients and predominantly in arteries showing hyperplasia. While a low number of subjects was examined and information about disease severity and patient characteristics is lacking, these calcifications and mineralo-organic particles may represent signs of tissue dysfunction.

Hormetic Effects of Phytochemicals on Health and Longevity.

Caloric restriction, intermittent fasting, and exercise activate defensive cellular responses such as autophagy, DNA repair, and the induction of antioxidant enzymes. These processes improve health and longevity by protecting cells and organs against damage, mutations, and reactive oxygen species. Consuming a diet rich in vegetables, fruits, and mushrooms can also improve health and longevity. Phytochemicals such as alkaloids, polyphenols, and terpenoids found in plants and fungi activate the same cellular processes as caloric restriction, fasting, and exercise. Many of the beneficial effects of fruits and vegetables may thus be due to activation of stress resistance pathways by phytochemicals. A better understanding of the mechanisms of action of phytochemicals may provide important insights to delay aging and prevent chronic diseases.

Antrodia cinnamomea induces anti-tumor activity by inhibiting the STAT3 signaling pathway in lung cancer cells.

We examined the effects of an Antrodia cinnamomea ethanol extract (ACEE) on lung cancer cells in vitro and tumor growth in vivo. ACEE produced dose-dependent cytotoxic effects and induced apoptosis in Lewis lung carcinoma (LLC) cells. ACEE treatment increased expression of p53 and Bax, as well as cleavage of caspase-3 and PARP, while reducing expression of survivin and Bcl-2. ACEE also reduced the levels of JAK2 and phosphorylated STAT3 in LLC cells. In a murine allograft tumor model, oral administration of ACEE significantly inhibited LLC tumor growth and metastasis without affecting serum biological parameters or body weight. ACEE increased cleavage of caspase-3 in murine tumors, while decreasing STAT3 phosphorylation. In addition, ACEE reduced the growth of human tumor xenografts in nude mice. Our findings therefore indicate that ACEE inhibits lung tumor growth and metastasis by inducing apoptosis and by inhibiting the STAT3 signaling pathway in cancer cells.

Antiaging effects of bioactive molecules isolated from plants and fungi.

Aging is influenced by many lifestyle choices that are under human control, including nutrition and exercise. The most effective known antiaging intervention consists of calorie restriction (CR), which increases lifespan in yeasts, worms, fruit flies, mice, and nonhuman primates. CR also improves healthspan by preventing the development of various aging-related diseases such as cancer, cardiovascular disease, diabetes, and neurodegeneration. Many compounds isolated from plants and fungi prolong lifespan and prevent age-related diseases in model organisms. These plant and fungal compounds modulate the same cellular and physiological pathways as CR, including those involving insulin and insulin-like growth factor-1, mammalian target of rapamycin, and sirtuins. Modulation of these aging-related pathways results in the activation of various cellular processes such as autophagy, DNA repair, and neutralization of reactive oxygen species. Together, these cellular processes are believed to delay aging and prevent chronic diseases by improving bodily functions and stress resistance. We review here the mechanisms of action of plant and fungal molecules possessing antiaging properties and discuss the possibilities and challenges associated with the development of antiaging compounds isolated from natural products.

NK Cell-Derived IFN-γ Protects against Nontuberculous Mycobacterial Lung Infection.

In developed countries, pulmonary nontuberculous mycobacteria (NTM) infections are more prevalent than Mycobacterium tuberculosis infections. Given the differences in the pathogenesis of NTM and M. tuberculosis infections, separate studies are needed to investigate the pathological effects of NTM pathogens. Our previous study showed that anti-IFN-γ autoantibodies are detected in NTM-infected patients. However, the role of NK cells and especially NK cell-derived IFN-γ in this context has not been studied in detail. In the current study, we show that NK1.1 cell depletion increases bacterial load and mortality in a mouse model of pulmonary NTM infection. NK1.1 cell depletion exacerbates NTM-induced pathogenesis by reducing macrophage phagocytosis, dendritic cell development, cytokine production, and lung granuloma formation. Similar pathological phenomena are observed in IFN-γ-deficient (IFN-γ-/-) mice following NTM infection, and adoptive transfer of wild-type NK cells into IFN-γ-/- mice considerably reduces NTM pathogenesis. Injection of rIFN-γ also prevents NTM-induced pathogenesis in IFN-γ-/- mice. We observed that NK cells represent the main producers of IFN-γ in the lungs and production starts as soon as 1 d postinfection. Accordingly, injection of rIFN-γ into IFN-γ-/- mice 1 d (but not 2 wk) postinfection significantly improves immunity against NTM infection. NK cells also stimulate mycobacterial killing and IL-12 production by macrophages. Our results therefore indicate that IFN-γ production by NK cells plays an important role in activating and enhancing innate and adaptive immune responses at early stages of pulmonary NTM infection.

Antrodia cinnamomea produces anti-angiogenic effects by inhibiting the VEGFR2 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The medicinal mushroom Antrodia cinnamomea has been used to treat cancer but its anti-angiogenic effects have not been studied in detail. AIM OF THE STUDY: The main objective of this study was to determine the molecular mechanism of activity underlying the anti-angiogenic effects of A. cinnamomea. MATERIALS AND METHODS: The effects of an A. cinnamomea ethanol extract (ACEE) on cell migration and microvessel formation were investigated in endothelial cells in vitro and Matrigel plugs implanted into mice in vivo. Activation of intracellular signaling pathways was examined using Western blotting. Protein expression was assessed using immunohistochemistry in a mouse model of lung metastasis. RESULTS: We show that treatment with ACEE inhibits cell migration and tube formation in human umbilical vein endothelial cells (HUVECs). ACEE suppresses phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2) and expression of pro-angiogenic kinases in vascular endothelial growth factor (VEGF)-treated HUVECs, in addition to reducing expression of Janus kinase 2 (JAK2) and phosphorylation of signal transducer and activator of transcription 3 (STAT3). ACEE treatment inhibits VEGF-induced microvessel formation in Matrigel plugs in vivo. In addition, ACEE significantly reduces VEGFR2 expression in Lewis lung carcinoma cells and downregulates the expression of cluster of differentiation 31 (CD31) and VEGFR2 in murine lung metastases. CONCLUSION: These results indicate that A. cinnamomea produces anti-angiogenic effects by inhibiting the VEGFR2 signaling pathway.

Mineral particles stimulate innate immunity through neutrophil extracellular traps containing HMGB1.

Calcium phosphate-based mineralo-organic particles form spontaneously in the body and may represent precursors of ectopic calcification. We have shown earlier that these particles induce activation of caspase-1 and secretion of IL-1ß by macrophages. However, whether the particles may produce other effects on immune cells is unclear. Here, we show that these particles induce the release of neutrophil extracellular traps (NETs) in a size-dependent manner by human neutrophils. Intracellular production of reactive oxygen species is required for particle-induced NET release by neutrophils. NETs contain the high-mobility group protein B1 (HMGB1), a DNA-binding protein capable of inducing secretion of TNF-α by a monocyte/macrophage cell line and primary macrophages. HMGB1 functions as a ligand of Toll-like receptors 2 and 4 on macrophages, leading to activation of the MyD88 pathway and TNF-α production. Furthermore, HMGB1 is critical to activate the particle-induced pro-inflammatory cascade in the peritoneum of mice. These results indicate that mineral particles promote pro-inflammatory responses by engaging neutrophils and macrophages via signaling of danger signals through NETs.

Pleomorphic bacteria-like structures in human blood represent non-living membrane vesicles and protein particles.

Although human blood is believed to be a sterile environment, recent studies suggest that pleomorphic bacteria exist in the blood of healthy humans. These studies have led to the development of "live-blood analysis," a technique used by alternative medicine practitioners to diagnose various human conditions, including allergies, cancer, cardiovascular disease and septicemia. We show here that bacteria-like vesicles and refringent particles form in healthy human blood observed under dark-field microscopy. These structures gradually increase in number during incubation and show morphologies reminiscent of cells undergoing division. Based on lipid analysis and Western blotting, we show that the bacteria-like entities consist of membrane vesicles containing serum and exosome proteins, including albumin, fetuin-A, apolipoprotein-A1, alkaline phosphatase, TNFR1 and CD63. In contrast, the refringent particles represent protein aggregates that contain several blood proteins. 16S rDNA PCR analysis reveals the presence of bacterial DNA in incubated blood samples but also in negative controls, indicating that the amplified sequences represent contaminants. These results suggest that the bacteria-like vesicles and refringent particles observed in human blood represent non-living membrane vesicles and protein aggregates derived from blood. The phenomena observed during live-blood analysis are therefore consistent with time-dependent decay of cells and body fluids during incubation ex vivo.

Pinicolol B from Antrodia cinnamomea induces apoptosis of nasopharyngeal carcinoma cells.

ETHNOPHARMACOLOGICAL RELEVANCE: The medicinal mushroom Antrodia cinnamomea possesses anticancer properties but the active compounds responsible for these effects are mostly unknown. AIM OF THE STUDY: We aimed to identify novel A. cinnamomea compounds that produce cytotoxic effects on cancer cells. MATERIALS AND METHODS: Using ethanol extraction and chromatography, we isolated the lanostanoid compound lanosta-7,9(11),24-trien-3ß,15α,21-triol (1) from cultured A. cinnamomea mycelium. Cytotoxicity and pro-apoptotic effects of compound 1 were evaluated using the MTS assay and flow cytometry analysis, respectively. RESULTS: Compound 1 produced cytotoxic effects on the nasopharyngeal carcinoma cell lines TW02 and TW04, with IC50 values of 63.3 and 115.0µM, respectively. On the other hand, no cytotoxic effects were observed on non-tumorigenic nasopharyngeal epithelial cells (NP69). In addition, compound 1 induced apoptosis in TW02 and TW04 cells as revealed by flow cytometry analysis. CONCLUSIONS: Our results demonstrate for the first time the presence of pinicolol B in A. cinnamomea mycelium and suggest that this compound may contribute to the anticancer effects of A. cinnamomea.

Isolation, Culture and Characterization of Hirsutella sinensis Mycelium from Caterpillar Fungus Fruiting Body.

The caterpillar fungus Ophiocordyceps sinensis (previously called Cordyceps sinensis) has been used for centuries in Asia as a tonic to improve health and longevity. Recent studies show that O. sinensis produces a wide range of biological effects on cells, laboratory animals and humans, including anti-fatigue, anti-infection, anti-inflammatory, antioxidant, and anti-tumor activities. In view of the rarity of O. sinensis fruiting bodies in nature, cultivation of its anamorph mycelium represents a useful alternative for large-scale production. However, O. sinensis fruiting bodies harvested in nature harbor several fungal contaminants, a phenomenon that led to the isolation and characterization of a large number of incorrect mycelium strains. We report here the isolation of a mycelium from a fruiting body of O. sinensis and we identify the isolate as O. sinensis' anamorph (also called Hirsutella sinensis) based on multi-locus sequence typing of several fungal genes (ITS, nrSSU, nrLSU, RPB1, RPB2, MCM7, ß-tubulin, TEF-1α, and ATP6). The main characteristics of the isolated mycelium, including its optimal growth at low temperature (16°C) and its biochemical composition, are similar to that of O. sinensis fruiting bodies, indicating that the mycelium strain characterized here may be used as a substitute for the rare and expensive O. sinensis fruiting bodies found in nature.

Anti-obesogenic and antidiabetic effects of plants and mushrooms.

Obesity is reaching global epidemic proportions as a result of factors such as high-calorie diets and lack of physical exercise. Obesity is now considered to be a medical condition, which not only contributes to the risk of developing type 2 diabetes mellitus, cardiovascular disease and cancer, but also negatively affects longevity and quality of life. To combat this epidemic, anti-obesogenic approaches are required that are safe, widely available and inexpensive. Several plants and mushrooms that are consumed in traditional Chinese medicine or as nutraceuticals contain antioxidants, fibre and other phytochemicals, and have anti-obesogenic and antidiabetic effects through the modulation of diverse cellular and physiological pathways. These effects include appetite reduction, modulation of lipid absorption and metabolism, enhancement of insulin sensitivity, thermogenesis and changes in the gut microbiota. In this Review, we describe the molecular mechanisms that underlie the anti-obesogenic and antidiabetic effects of these plants and mushrooms, and propose that combining these food items with existing anti-obesogenic approaches might help to reduce obesity and its complications.

An iron detection system determines bacterial swarming initiation and biofilm formation.

Iron availability affects swarming and biofilm formation in various bacterial species. However, how bacteria sense iron and coordinate swarming and biofilm formation remains unclear. Using Serratia marcescens as a model organism, we identify here a stage-specific iron-regulatory machinery comprising a two-component system (TCS) and the TCS-regulated iron chelator 2-isocyano-6,7-dihydroxycoumarin (ICDH-Coumarin) that directly senses and modulates environmental ferric iron (Fe3+) availability to determine swarming initiation and biofilm formation. We demonstrate that the two-component system RssA-RssB (RssAB) directly senses environmental ferric iron (Fe3+) and transcriptionally modulates biosynthesis of flagella and the iron chelator ICDH-Coumarin whose production requires the pvc cluster. Addition of Fe3+, or loss of ICDH-Coumarin due to pvc deletion results in prolonged RssAB signaling activation, leading to delayed swarming initiation and increased biofilm formation. We further show that ICDH-Coumarin is able to chelate Fe3+ to switch off RssAB signaling, triggering swarming initiation and biofilm reduction. Our findings reveal a novel cellular system that senses iron levels to regulate bacterial surface lifestyle.

Translocation of mineralo-organic nanoparticles from blood to urine: a new mechanism for the formation of kidney stones?

Recent studies indicate that mineralo-organic nanoparticles form in various human body fluids, including blood and urine. These nanoparticles may form within renal tubules and increase in size in supersaturated urine, eventually leading to the formation of kidney stones. Here, we present observations suggesting that mineralo-organic nanoparticles found in blood may induce kidney stone formation via an alternative mechanism in which the particles translocate through endothelial and renal epithelial cells to reach urine. We propose that this alternative mechanism of kidney stone formation and the study of mineralo-organic nanoparticles in general may provide novel strategies for the early detection and treatment of ectopic calcifications and kidney stones.

Immunomodulatory properties of medicinal mushrooms: differential effects of water and ethanol extracts on NK cell-mediated cytotoxicity.

Medicinal mushrooms have been used for centuries in Asian countries owing to their beneficial effects on health and longevity. Previous studies have reported that a single medicinal mushroom may produce both stimulatory and inhibitory effects on immune cells, depending on conditions, but the factors responsible for this apparent dichotomy remain obscure. We show here that water and ethanol extracts of cultured mycelium from various species (Agaricus blazei Murrill, Antrodia cinnamomea, Ganoderma lucidum and Hirsutella sinensis) produce opposite effects on NK cells. Water extracts enhance NK cell cytotoxic activity against cancer cells, whereas ethanol extracts inhibit cytotoxicity. Water extracts stimulate the expression and production of cytolytic proteins (perforin and granulysin) and NKG2D/NCR cell surface receptors, and activate intracellular signaling kinases (ERK, JNK and p38). In contrast, ethanol extracts inhibit expression of cytolytic and cell surface receptors. Our results suggest that the mode of extraction of medicinal mushrooms may determine the nature of the immunomodulatory effects produced on immune cells, presumably owing to the differential solubility of stimulatory and inhibitory mediators. These findings have important implications for the preparation of medicinal mushrooms to prevent and treat human diseases.

Is the inflammasome relevant for epithelial cell function?

Inflammasomes are intracellular protein complexes that sense microbial components and damage of infected cells. Following activation by molecules released by pathogens or injured cells, inflammasomes activate caspase-1, allowing secretion of the pro-inflammatory cytokines IL-1ß and IL-18 from innate immune cells. Inflammasomes are also expressed in epithelial cells, where their function has attracted less attention. Nonetheless, depending on the tissue, epithelial inflammasomes can mediate inflammation, wound healing, and pain sensitivity. We review here recent findings on inflammasomes found in epithelial tissues, highlighting the importance of these protein complexes in the response of epithelial tissues to microbial infections.

Detection and characterization of mineralo-organic nanoparticles in human kidneys.

Ectopic calcification is associated with various human diseases, including atherosclerosis, cancer, chronic kidney disease, and diabetes mellitus. Although mineral nanoparticles have been detected in calcified blood vessels, the nature and role of these particles in the human body remain unclear. Here we show for the first time that human kidney tissues obtained from end-stage chronic kidney disease or renal cancer patients contain round, multilamellar mineral particles of 50 to 1,500 nm, whereas no particles are observed in healthy controls. The mineral particles are found mainly in the extracellular matrix surrounding the convoluted tubules, collecting ducts and loops of Henle as well as within the cytoplasm of tubule-delineating cells, and consist of polycrystalline calcium phosphate similar to the mineral found in bones and ectopic calcifications. The kidney mineral nanoparticles contain several serum proteins that inhibit ectopic calcification in body fluids, including albumin, fetuin-A, and apolipoprotein A1. Since the mineralo-organic nanoparticles are found not only within calcified deposits but also in areas devoid of microscopic calcifications, our observations indicate that the nanoparticles may represent precursors of calcification and renal stones in humans.