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Head of Institute: Prof. Oren Froy

Administrative manager: Ms. Yael Fruchter

Office Address:
Institute of Biochemistry, Food Science and Nutrition,
Robert H. Smith Faculty of Agriculture, Food and Environment,
The Hebrew University of Jerusalem, 
P.O.Box 12, Rehovot 7610001, ISRAEL

Tel: +972 - (0)8-9489385
Fax: +972 - (0)8-9363208
Email Address: yaelf@savion.huji.ac.il

Publications

2018
Goldstein, I. ; Hager, G. L. The Three Ds of Transcription Activation by Glucagon: Direct, Delayed, and Dynamic. Endocrinology 2018, 159, 206-216.Abstract
Upon lowered blood glucose occurring during fasting, glucagon is secreted from pancreatic islets, exerting various metabolic effects to normalize glucose levels. A considerable portion of these effects is mediated by glucagon-activated transcription factors (TFs) in liver. Glucagon directly activates several TFs via immediate cyclic adenosine monophosphate (cAMP)- and calcium-dependent signaling events. Among these TFs, cAMP response element-binding protein (CREB) is a major factor. CREB recruits histone-modifying enzymes and cooperates with other TFs on the chromatin template to increase the rate of gene transcription. In addition to direct signal transduction, the transcriptional effects of glucagon are also influenced by dynamic TF cross talk. Specifically, assisted loading of one TF by a companion TF leads to increased binding and activity. Lastly, transcriptional regulation by glucagon is also exerted by TF cascades by which a primary TF induces the gene expression of secondary TFs that bring about their activity a few hours after the initial glucagon signal. This mechanism of a delayed response may be instrumental in establishing the temporal organization of the fasting response by which distinct metabolic events separate early from prolonged fasting. In this mini-review, we summarize recent advances and critical discoveries in glucagon-dependent gene regulation with a focus on direct TF activation, dynamic TF cross talk, and TF cascades.
Froy, O. ; Garaulet, M. The Circadian Clock in White and Brown Adipose Tissue: Mechanistic, Endocrine, and Clinical Aspects. Endocrine reviews 2018, 39, 261 - 273. Publisher's VersionAbstract
Obesity is a major risk factor for the development of illnesses, such as insulin resistance and hypertension, and has become a serious public health problem. Mammals have developed a circadian clock located in the hypothalamic suprachiasmatic nuclei (SCN) that responds to the environmental light-dark cycle. Clocks similar to the one located in the SCN are found in peripheral tissues, such as the kidney, liver, and adipose tissue. The circadian clock regulates metabolism and energy homeostasis in peripheral tissues by mediating activity and/or expression of key metabolic enzymes and transport systems. Knockouts or mutations in clock genes that lead to disruption of cellular rhythmicity have provided evidence to the tight link between the circadian clock and metabolism. In addition, key proteins play a dual role in regulating the core clock mechanism, as well as adipose tissue metabolism, and link circadian rhythms with lipogenesis and lipolysis. Adipose tissues are distinguished as white, brown, and beige (or brite), each with unique metabolic characteristics. Recently, the role of the circadian clock in regulating the differentiation into the different adipose tissues has been investigated. In this review, the role of clock proteins and the downstream signaling pathways in white, brown, and brite adipose tissue function and differentiation will be reviewed. In addition, chronodisruption and metabolic disorders and clinical aspects of circadian adiposity will be addressed.
Christ, P. ; Sowa, A. S. ; Froy, O. ; Lorentz, A. The Circadian Clock Drives Mast Cell Functions in Allergic Reactions. Frontiers in immunology 2018, 9 1526 - 1526. Publisher's VersionAbstract
Allergic diseases are known to vary in the severity of their symptoms throughout the day/night cycle. This rhythmicity is also observed in mast cell function and responsiveness. Mast cells are key effector cells of allergic reactions and release cytokines, chemokines, and important inflammatory mediators such as histamine, which have been shown to display diurnal variation. Recent research clarified that mast cells are controlled by their internal clock-which is regulated by a specific set of clock genes-as well as external factors such as light sensed by the suprachiasmatic nuclei, hormonal status, or diet. Here, we give an overview of the connections between circadian clock, mast cells, and allergic disease. Further work aimed at studying the role of chronotherapy/chronomedicine should take into account this rhythmic nature of not only mast cells but also the immune responses generated by mast cell signaling.
Weintraub, Y. ; Cohen, S. ; Dotan, I. ; Tauman, R. ; Chapnik, N. ; Froy, O. P334 Does the circadian clock have a role in the pathogenesis of inflammatory bowel disease (IBD)?. Journal of Crohn's and Colitisecco-jcc 2018, 12, S270 - S271. Publisher's VersionAbstract
Sleep dysfunction modifies the immune system and has been implicated as a potential trigger of IBD flares. Sleep dysfunction also alters the synchrony among clock genes leading to disruption of overall circadian regulation. Specifically, in the intestine, it is manifested by increased gut cellular permeability. We hypothesised that changes in mucosal immune balance may be reflected by alterations in the circadian clock and constitute an unattended pathogenic mechanism of IBD. Our aim was to investigate intestinal and systemic clock gene expression (in patients with newly diagnosed IBD and in healthy controls).Patients and controls were recruited upon diagnostic endoscopic evaluation. Demographics, familial medical history, sleep questionnaires, disease activity indices, and endoscopic scores were recorded. Anthropometric parameters, C-reactive protein (CRP), albumin, haemoglobin (Hb), and fecal calprotectin (Fcal) were measured as well. Peripheral blood and tissue samples were analysed for clock gene (Clock, Bmal1, Cry1, Cry2, Per1, and Per2) expression.Of the 32 participants recruited (age 8–25 years, median: 16.1), 14 had newly diagnosed IBD and 18 were healthy controls. Age, gender, sleep questionnaire scores, and time of endoscopy were not statistically different between the groups. Hb, CRP, and Fcal levels were significantly higher in the IBD compared with the healthy controls group (p < 0.05), while albumin was significantly lower (p < 0.05). Clock gene expression (Clock, Cry1, Cry2, Per1, and Per2) in WBC was decreased in newly diagnosed IBD patients compared with health controls (p < 0.05). Similarly, the expression level of the aforementioned genes was lower in inflamed intestinal tissues (p < 0.05). Interestingly, similar reduction in clock gene expression was seen even in healthy (non-inflamed) intestinal tissue from IBD patients (p < 0.05).Clock gene expression is reduced in both inflamed and non-inflamed intestinal tissue in patients with newly diagnosed IBD. Moreover, IBD patients show a systemic reduction in clock gene expression. Our findings may lead to new therapeutic approaches and strategies as well as serve as diagnostic tools in IBD.
Froy, O. Circadian rhythms, nutrition and implications for longevity in urban environments. Proceedings of the Nutrition Society 2018, 77, 216-222. Publisher's VersionAbstract
Presently, about 12% of the population is 65 years or older and by the year 2030 that figure is expected to reach 21%. In order to promote the well-being of the elderly and to reduce the costs associated with health care demands, increased longevity should be accompanied by ageing attenuation. Energy restriction, which limits the amount of energy consumed to 60–70% of the daily intake, and intermittent fasting, which allows the food to be available ad libitum every other day, extend the life span of mammals and prevent or delay the onset of major age-related diseases, such as cancer, diabetes and cataracts. Recently, we have shown that well-being can be achieved by resetting of the circadian clock and induction of robust catabolic circadian rhythms via timed feeding. In addition, the clock mechanism regulates metabolism and major metabolic proteins are key factors in the core clock mechanism. Therefore, it is necessary to increase our understanding of circadian regulation over metabolism and longevity and to design new therapies based on this regulation. This review will explore the present data in the field of circadian rhythms, ageing and metabolism.
Adar, C. ; Sirotinskaya, V. ; Bar Dolev, M. ; Friehmann, T. ; Braslavsky, I. Falling water ice affinity purification of ice-binding proteins. Scientific Reports 2018, 8 11046. Publisher's VersionAbstract
Ice-binding proteins (IBPs) permit their hosts to thrive in the presence of ice. The ability of IBPs to control ice growth makes them potential additives in industries ranging from food storage and cryopreservation to anti-icing systems. For IBPs to be used in commercial applications, however, methods are needed to produce sufficient quantities of high-quality proteins. Here, we describe a new method for IBP purification, termed falling water ice affinity purification (FWIP). The method is based on the affinity of IBPs for ice and does not require molecular tags. A crude IBP solution is allowed to flow over a chilled vertical surface of a commercial ice machine. The temperature of the surface is lowered gradually until ice crystals are produced, to which the IBPs bind but other solutes do not. We found that a maximum of 35 mg of IBP was incorporated in 1 kg of ice. Two rounds of FWIP resulted in >95% purity. An ice machine that produces 60 kg of ice per day can be used to purify one gram of IBP per day. In combination with efficient concentration of the protein solution by tangential flow filtration the FWIP method is suitable for the purification of grams of IBPs for research purposes and applications.
Braslavsky, I. ; Stavans, J. Application of Algebraic Topology to Homologous Recombination of DNA. iScience 2018, 4 64 - 67. Publisher's VersionAbstract
SummaryBrouwer's fixed point theorem, a fundamental theorem in algebraic topology proved more than a hundred years ago, states that given any continuous map from a closed, simply connected set into itself, there is a point that is mapped unto itself. Here we point out the connection between a one-dimensional application of Brouwer's fixed point theorem and a mechanism proposed to explain how extension of single-stranded DNA substrates by recombinases of the RecA superfamily facilitates significantly the search for homologous sequences on long chromosomes.
Bahari, L. ; Bein, A. ; Yashunsky, V. ; Braslavsky, I. Directional freezing for the cryopreservation of adherent mammalian cells on a substrate. PLOS ONE 2018, 13, e0192265 -. Publisher's VersionAbstract
Successfully cryopreserving cells adhered to a substrate would facilitate the growth of a vital confluent cell culture after thawing while dramatically shortening the post-thaw culturing time. Herein we propose a controlled slow cooling method combining initial directional freezing followed by gradual cooling down to -80°C for robust preservation of cell monolayers adherent to a substrate. Using computer controlled cryostages we examined the effect of cooling rates and dimethylsulfoxide (DMSO) concentration on cell survival and established an optimal cryopreservation protocol. Experimental results show the highest post-thawing viability for directional ice growth at a speed of 30 μm/sec (equivalent to freezing rate of 3.8°C/min), followed by gradual cooling of the sample with decreasing rate of 0.5°C/min. Efficient cryopreservation of three widely used epithelial cell lines: IEC-18, HeLa, and Caco-2, provides proof-of-concept support for this new freezing protocol applied to adherent cells. This method is highly reproducible, significantly increases the post-thaw cell viability and can be readily applied for cryopreservation of cellular cultures in microfluidic devices.
Mangiagalli, M. ; Sarusi, G. ; Kaleda, A. ; Bar Dolev, M. ; Nardone, V. ; Vena, V. F. ; Braslavsky, I. ; Lotti, M. ; Nardini, M. Structure of a bacterial ice binding protein with two faces of interaction with ice. The FEBS Journal 2018, 285, 1653 - 1666. Publisher's VersionAbstract
Ice-binding proteins (IBPs) contribute to the survival of many living beings at subzero temperature by controlling the formation and growth of ice crystals. This work investigates the structural basis of the ice-binding properties of EfcIBP, obtained from Antarctic bacteria. EfcIBP is endowed with a unique combination of thermal hysteresis and ice recrystallization inhibition activity. The three-dimensional structure, solved at 0.84 Å resolution, shows that EfcIBP belongs to the IBP-1 fold family, and is organized in a right-handed ?-solenoid with a triangular cross-section that forms three protein surfaces, named A, B, and C faces. However, EfcIBP diverges from other IBP-1 fold proteins in relevant structural features including the lack of a ?capping? region on top of the ?-solenoid, and in the sequence and organization of the regions exposed to ice that, in EfcIBP, reveal the presence of threonine-rich ice-binding motifs. Docking experiments and site-directed mutagenesis pinpoint that EfcIBP binds ice crystals not only via its B face, as common to other IBPs, but also via ice-binding sites on the C face. Database Coordinates and structure factors have been deposited in the Protein Data Bank under accession number 6EIO.
2017
Oren, T. ; Nimri, L. ; Yehuda-Shnaidman, E. ; Staikin, K. ; Hadar, Y. ; Friedler, A. ; Amartely, H. ; Slutzki, M. ; Di Pizio, A. ; Niv, M. Y. ; et al. Recombinant ostreolysin induces brown fat-like phenotype in HIB-1B cells. Mol Nutr Food Res 2017, 61.Abstract
SCOPE: Brown adipose tissue (BAT) is the main regulator of thermogenesis by increasing energy expenditure through the uncoupling of oxidative metabolism from ATP synthesis. There is a growing body of evidence for BAT being the key responsible organ in combating obesity and its related disorders. Herein we propose the fungal protein ostreolysin (Oly), which has been previously shown to bind to cholesterol-enriched raft-like membrane domains (lipid rafts) of mammalian cells, as a suitable candidate for interaction with brown preadipocytes. The aim of the present study was therefore to characterize the mechanism by which a recombinant version of ostreolysin (rOly) induces brown adipocyte differentiation. METHODS AND RESULTS: Primary isolated brown preadipocytes or HIB-1B brown preadipocyte cells were treated with rOly and the effects on morphology, lipid accumulation, respiration rate, and associated gene and protein expression were measured. rOly upregulated mRNA and protein levels of factors related to brown adipocyte differentiation, induced lipid droplet formation, and increased cellular respiration rate due to expression of uncoupling protein 1. rOly also upregulated β-tubulin expression, and therefore microtubules might be involved in its mechanism of action. CONCLUSION: rOly promotes brown adipocyte differentiation, suggesting a new mechanism for rOly's contribution to the battle against obesity.
Hatting, M. ; Rines, A. K. ; Luo, C. ; Tabata, M. ; Sharabi, K. ; Hall, J. A. ; Verdeguer, F. ; Trautwein, C. ; Puigserver, P. Adipose Tissue CLK2 Promotes Energy Expenditure during High-Fat Diet Intermittent Fasting. Cell Metab 2017, 25, 428-437.Abstract
A promising approach to treating obesity is to increase diet-induced thermogenesis in brown adipose tissue (BAT), but the regulation of this process remains unclear. Here we find that CDC-like kinase 2 (CLK2) is expressed in BAT and upregulated upon refeeding. Mice lacking CLK2 in adipose tissue exhibit exacerbated obesity and decreased energy expenditure during high-fat diet intermittent fasting. Additionally, tissue oxygen consumption and protein levels of UCP1 are reduced in CLK2-deficient BAT. Phosphorylation of CREB, a transcriptional activator of UCP1, is markedly decreased in BAT cells lacking CLK2 due to enhanced CREB dephosphorylation. Mechanistically, CREB dephosphorylation is rescued by the inhibition of PP2A, a phosphatase that targets CREB. Our results suggest that CLK2 is a regulatory component of diet-induced thermogenesis in BAT through increased CREB-dependent expression of UCP1.
Sharabi, K. ; Lin, H. ; Tavares, C. D. J. ; Dominy, J. E. ; Camporez, J. P. ; Perry, R. J. ; Schilling, R. ; Rines, A. K. ; Lee, J. ; Hickey, M. ; et al. Selective Chemical Inhibition of PGC-1α Gluconeogenic Activity Ameliorates Type 2 Diabetes. Cell 2017, 169, 148-160.e15.Abstract
Type 2 diabetes (T2D) is a worldwide epidemic with a medical need for additional targeted therapies. Suppression of hepatic glucose production (HGP) effectively ameliorates diabetes and can be exploited for its treatment. We hypothesized that targeting PGC-1α acetylation in the liver, a chemical modification known to inhibit hepatic gluconeogenesis, could be potentially used for treatment of T2D. Thus, we designed a high-throughput chemical screen platform to quantify PGC-1α acetylation in cells and identified small molecules that increase PGC-1α acetylation, suppress gluconeogenic gene expression, and reduce glucose production in hepatocytes. On the basis of potency and bioavailability, we selected a small molecule, SR-18292, that reduces blood glucose, strongly increases hepatic insulin sensitivity, and improves glucose homeostasis in dietary and genetic mouse models of T2D. These studies have important implications for understanding the regulatory mechanisms of glucose metabolism and treatment of T2D.
Myers, E. F. ; Trostler, N. ; Varsha, V. ; Voet, H. Insights from the Diabetes in India Nutrition Guidelines Study: Adopting Innovations Using a Knowledge Transfer Model. Topics in Clinical Nutrition 2017, 32, 69-86. Publisher's VersionAbstract
This 12-month prospective randomized cluster trial of 20 dietitians in India compared usual care (UC) and evidence-based nutrition practice guideline (EBNPG) care for patients with type 2 diabetes mellitus. Baseline, 6-month, and 12-month data from 238 patients were analyzed. EBNPG implementation was evaluated using the Ottawa Model for Knowledge Transfer. EBNPG and UC groups achieved significant hemoglobin A1C improvements. EBNPG-treated participants were significantly more likely to meet low-density lipoprotein, high-density lipoprotein, and triglyceride goals at 6 or 12 months. Dietitian dropout, implementation barriers, and undetermined EBNPG intervention fidelity are limitations. Future research should assess barriers/supports and degree of EBNPG use. © 2017 Wolters Kluwer Health, Inc.
Swan, W. I. ; Vivanti, A. ; Hakel-Smith, N. A. ; Hotson, B. ; Orrevall, Y. ; Trostler, N. ; Beck Howarter, K. ; Papoutsakis, C. Nutrition Care Process and Model Update: Toward Realizing People-Centered Care and Outcomes Management. Journal of the Academy of Nutrition and Dietetics 2017, 117, 2003-2014. Publisher's Version
Soon, J. M. ; Saguy, I. Crowdsourcing: A new conceptual view for food safety and quality. Trends in Food Science and Technology 2017, 66, 63-72. Publisher's VersionAbstract
Background Crowdsourcing is a new tool offered mainly over the internet for obtaining ideas, content, funding by seeking contributions from a large group of people and especially from the online community rather than from traditional employees or suppliers. Crowdsourcing is widespread in numerous food applications (e.g., technology, entrepreneurial projects, start-ups funding, innovative product developments). Scope and approach Although the use of crowdsourcing has increased rapidly, there is still much untapped potential in harnessing its vast innovative potential in food quality and safety solutions. This paper aims to review recent utilization of crowdsourcing practices in the food domain. Additionally, to furnish a conceptual view on possible application where crowdsourcing can be harnessed in enhancing food quality, safety and reducing risks. Key findings and conclusions It argues that crowdsourcing initiative is potentially a very useful tool as a part of the big data by utilizing the crowd's data in shelf-life monitoring, inventory control, foodborne illness surveillance, identification of contaminated products and to improve food businesses’ hygiene, enhance food safety, communication and allergen management and minimizing risk. The limitations include the number of reports and data generated may overwhelm the food industry or authority due to lack of internal resources i.e. time and technical expert to process the information. There is also risk of lack of crowd participation and loss of control. Hence, a mechanism to facilitate, evaluate and process the data should be in place. © 2017 Elsevier Ltd
Belkin, S. ; Yagur-Kroll, S. ; Kabessa, Y. ; Korouma, V. ; Septon, T. ; Anati, Y. ; Zohar-Perez, C. ; Rabinovitz, Z. ; Nussinovitch, A. ; Agranat, A. J. Remote detection of buried landmines using a bacterial sensor. Nature Biotechnology 2017, 35, 308-310. Publisher's Version
Rachman-Elbaum, S. ; Stark, A. H. ; Kachal, J. ; Johnson, T. W. ; Porat-Katz, B. S. A New System of Documentation to Improve Dietitian-Physician Crosstalk. Isr Med Assoc J 2017, 19, 360-364.Abstract
BACKGROUND: Standardization of the dietetic care process allows for early identification of malnutrition and metabolic disorders, interdisciplinary collaboration among the medical team, and improved quality of patient care. Globally, dietitians are adopting a nutrition care model that integrates national regulations with professional scope of practice. Currently, Israel lacks a standardized dietetic care process and documentation terminology. OBJECTIVES: To assess the utilization of a novel sectoral documentation system for nutrition care in Israel. METHODS: Seventy dietitians working in 63 geriatric facilities completed an online training program presenting the proposed patient-sectoral-model. Training was followed by submission of sample case studies from clinical practice or completion of a case simulation. Application of the proposed model was assessed by measuring the frequency participants implemented different sections of the model and responses to an approval questionnaire. RESULTS: Fifty-four participants (77%) provided completed cases. Over 80% of participants reported each step of the proposed dietary care process with 100% reporting the "nutrition diagnosis". Fifty-one dietitians (72.8%) completed the approval survey with the section on nutrition diagnosis receiving a highly favorable response (95%), indicating that the new documentation system was beneficial. Over 80% of participants rated the model useful in clinical practice. CONCLUSIONS: A sectoral approach for documenting dietetic care may be the ideal model for dietitians working in specific patient populations with the potential for improving interdisciplinary collaboration in patient care.
Nimri, L. ; Spivak, O. ; Tal, D. ; Schälling, D. ; Peri, I. ; Graeve, L. ; Salame, T. M. ; Yarden, O. ; Hadar, Y. ; Schwartz, B. A recombinant fungal compound induces anti-proliferative and pro-apoptotic effects on colon cancer cells. Oncotarget 2017, 8 28854-28864.Abstract
Finding intracellular pathways and molecules that can prevent the proliferation of colon cancer cells can provide significant bases for developing treatments for this disease. Ostreolysin (Oly) is a protein found in the mushroom Pleurotus ostreatus, and we have produced a recombinant version of this protein (rOly).We measured the viability of several colon cancer cells treated with rOly. Xenografts and syngeneic colon cancer cells were injected into in vivo mouse models, which were then treated with this recombinant protein.rOly treatment induced a significant reduction in viability of human and mouse colon cancer cells. In contrast, there was no reduction in the viability of normal epithelial cells from the small intestine. In the search for cellular targets of rOly, we showed that it enhances the anti-proliferative activity of drugs targeting cellular tubulin. This was accompanied by a reduction in the weight and volume of tumours in mice injected with rOly as compared to their respective control mice in two in vivo models.Our results advance the functional understanding of rOly as a potential anti-cancer treatment associated with pro-apoptotic activities preferentially targeting colon cancer cells.
Bein, A. ; Zilbershtein, A. ; Golosovsky, M. ; Davidov, D. ; Schwartz, B. LPS Induces Hyper-Permeability of Intestinal Epithelial Cells. Journal of Cellular Physiology 2017, 232, 381-390. Publisher's VersionAbstract
Necrotizing Enterocolitis (NEC) is a severe inflammatory disorder leading to high morbidity and mortality rates. A growing body of evidence demonstrate the key role of the Toll like receptor 4 (TLR4) in NEC. This membranal receptor recognizes lipopolysaccharides (LPS) from the bacterial wall and triggers an inflammatory response. The aim of the present study was to elucidate the effect of LPS on paracellular permeability known to be severely affected in NEC. IEC-18 cells were treated with LPS and the effects on morphology, paracellular permeability and their associated gene and protein expressions were measured. Our results show that LPS down regulated the expression of occludin and ZO-1 mRNAs while up regulating Cdkn1a. In addition LPS caused a significant increase in paracellular permeability and epithelial barrier damage. Finally ZO-1 protein was found to be spatially disarrayed in the intercellular junctions in response to LPS. We conclude that LPS adversely affected the functionality of the intestinal epithelial barrier suggesting a new mechanism by which bacterial infection may contribute to the development of NEC. J. Cell. Physiol. 232: 381–390, 2017. © 2016 Wiley Periodicals, Inc.
Avni, S. ; Ezove, N. ; Hanani, H. ; Yadid, I. ; Karpovsky, M. ; Hayby, H. ; Gover, O. ; Hadar, Y. ; Schwartz, B. ; Danay, O. Olive Mill Waste Enhances α-Glucan Content in the Edible Mushroom Pleurotus eryngii. Int J Mol Sci 2017, 18.Abstract
Mushroom polysaccharides are edible polymers that have numerous reported biological functions; the most common effects are attributed to β-glucans. In recent years, it became apparent that the less abundant α-glucans also possess potent effects in various health conditions. Here we explore several species for their total, β and α-glucan content. was found to have the highest total glucan concentrations and the highest α-glucans proportion. We also found that the stalks (stipe) of the fruit body contained higher glucan content then the caps (pileus). Since mushrooms respond markedly to changes in environmental and growth conditions, we developed cultivation methods aiming to increase the levels of α and β-glucans. Using olive mill solid waste (OMSW) from three-phase olive mills in the cultivation substrate. We were able to enrich the levels mainly of α-glucans. Maximal total glucan concentrations were enhanced up to twice when the growth substrate contained 80% of OMSW compared to no OMSW. Taking together this study demonstrate that can serve as a potential rich source of glucans for nutritional and medicinal applications and that glucan content in mushroom fruiting bodies can be further enriched by applying OMSW into the cultivation substrate.