Modelling glioblastoma tumour-host cell interactions using adult brain organotypic slice co-culture

Glioblastoma multiforme (GBM) is an aggressive incurable brain cancer. The cells that fuel the growth of tumours resemble neural stem cells found in the developing and adult mammalian forebrain. These are referred to as glioma stem cells (GSCs). Similar to neural stem cells, GSCs exhibit a variety of phenotypic states: dormant, quiescent, proliferative and differentiating. How environmental cues within the brain influence these distinct states is not well understood. Laboratory models of GBM can be generated using either genetically engineered mouse models, or via intracranial transplantation of cultured tumour initiating cells (mouse or human). Unfortunately, these approaches are expensive, time-consuming, low-throughput and ill-suited for monitoring live cell behaviours. Here, we explored whole adult brain coronal organotypic slices as an alternative model. Mouse adult brain slices remain viable in a serum-free basal medium for several weeks. GSCs can be easily microinjected into specific anatomical sites ex vivo, and we demonstrate distinct responses of engrafted GSCs to diverse microenvironments in the brain tissue. Within the subependymal zone – one of the adult neural stem cell niches – injected tumour cells could effectively engraft and respond to endothelial niche signals. Tumour-transplanted slices were treated with the antimitotic drug temozolomide as proof of principle of the utility in modelling responses to existing treatments. Engraftment of mouse or human GSCs onto whole brain coronal organotypic brain slices therefore provides a simplified, yet flexible, experimental model. This will help to increase the precision and throughput of modelling GSC-host brain interactions and complements ongoing in vivo studies. This article has an associated First Person interview with the first author of the paper

Long -term feeding with high plant protein based diets in gilthead seabream (Sparus aurata, L.) leads to changes in the inflammatory and immune related gene expression at intestinal level

[EN] Background: In order to ensure sustainability of aquaculture production of carnivourous fish species such as the gilthead seabream (Sparus aurata, L.), the impact of the inclusion of alternative protein sources to fishmeal, including plants, has been assessed. With the aim of evaluating long-term effects of vegetable diets on growth and intestinal status of the on-growing gilthead seabream (initial weight = 129 g), three experimental diets were tested: a strict plant protein-based diet (VM), a fishmeal based diet (FM) and a plant protein-based diet with 15% of marine ingredients (squid and krill meal) alternative to fishmeal (VM+). Intestines were sampled after 154 days. Besides studying growth parameters and survival, the gene expression related to inflammatory response, immune system, epithelia integrity and digestive process was analysed in the foregut and hindgut sections, as well as different histological parameters in the foregut. Results: There were no differences in growth performance (p = 0.2703) and feed utilization (p = 0.1536), although a greater fish mortality was recorded in the VM group (p = 0.0141). In addition, this group reported a lower expression in genes related to pro-inflammatory response, as Interleukine-1 beta (il1 beta, p = 0.0415), Interleukine-6 (il6, p = 0.0347) and cyclooxigenase-2 (cox2, p = 0.0014), immune-related genes as immunoglobulin M (igm, p = 0.0002) or bacterial defence genes as alkaline phosphatase (alp, p = 0.0069). In contrast, the VM+ group yielded similar survival rate to FM (p = 0.0141) and the gene expression patterns indicated a greater induction of the inflammatory and immune markers (il1 beta, cox2 and igm). However, major histological changes in gut were not detected. Conclusions: Using plants as the unique source of protein on a long term basis, replacing fishmeal in aqua feeds for gilthead seabream, may have been the reason of a decrease in the level of different pro-inflammatory mediators (il1 beta, il6 and cox2) and immune-related molecules (igm and alp), which reflects a possible lack of local immune response at the intestinal mucosa, explaining the higher mortality observed. Krill and squid meal inclusion in vegetable diets, even at low concentrations, provided an improvement in nutrition and survival parameters compared to strictly plant protein based diets as VM, maybe explained by the maintenance of an effective immune response throughout the assay.The research has been partially funded by Vicerrectorat d'Investigacio, Innovacio i Transferencia of the Universitat Politecnica de Valencia, which belongs to the project Aquaculture feed without fishmeal (SP20120603). The funders had no role in study design, data collection and analysis, decision to publish or preparation of the manuscript.Estruch-Cucarella, G.; Collado, MC.; Monge-Ortiz, R.; Tomas-Vidal, A.; Jover Cerdá, M.; Peñaranda, D.; Perez Martinez, G.... (2018). Long -term feeding with high plant protein based diets in gilthead seabream (Sparus aurata, L.) leads to changes in the inflammatory and immune related gene expression at intestinal level. BMC Veterinary Research. 14. https://doi.org/10.1186/s12917-018-1626-6S14Hardy RW. Utilization of plant proteins in fish diets: effects of global demand and supplies of fishmeal. Aquac Res. 2010;41:770–6.Martínez-Llorens S, Moñino AV, Vidal AT, Salvador VJM, Pla Torres M, Jover Cerdá M, et al. Soybean meal as a protein source in gilthead sea bream (Sparus aurata L.) diets: effects on growth and nutrient utilization. Aquac Res. 2007;38(1):82–90.Tacon AGJ, Metian M. Global overview on the use of fish meal and fish oil in industrially compounded aquafeeds: trends and future prospects. Aquaculture. 2008;285:146–58.Bonaldo A, Roem AJ, Fagioli P, Pecchini A, Cipollini I, Gatta PP. Influence of dietary levels of soybean meal on the performance and gut histology of gilthead sea bream (Sparus aurata L.) and European sea bass (Dicentrarchus labrax L.). Aquac Res. 2008;39(9):970–8.Kissil G, Lupatsch I. Successful replacement of fishmeal by plant proteins in diets for the gilthead seabream, Sparus Aurata L. Isr J Aquac – Bamidgeh. 2004;56(3):188–99.Monge-Ortíz R, Martínez-Llorens S, Márquez L, Moyano FJ, Jover-Cerdá M, Tomás-Vidal A. Potential use of high levels of vegetal proteins in diets for market-sized gilthead sea bream (Sparus aurata). Arch Anim Nutr. 2016;70(2):155–72.Santigosa E, Sánchez J, Médale F, Kaushik S, Pérez-Sánchez J, Gallardo MA. Modifications of digestive enzymes in trout (Oncorhynchus mykiss) and sea bream (Sparus aurata) in response to dietary fish meal replacement by plant protein sources. Aquaculture. 2008;282:68–74.Santigosa E, García-Meilán I, Valentin JM, Pérez-Sánchez J, Médale F, Kaushik S, et al. Modifications of intestinal nutrient absorption in response to dietary fish meal replacement by plant protein sources in sea bream (Sparus aurata) and rainbow trout (Onchorynchus mykiss). Aquaculture. 2011;317:146–54.Sitjá-Bobadilla A, Peña-Llopis S, Gómez-Requeni P, Médale F, Kaushik S, Pérez-Sánchez J. Effect of fish meal replacement by plant protein sources on non-specific defence mechanisms and oxidative stress in gilthead sea bream (Sparus aurata). Aquaculture. 2005;249:387–400.Martínez-Llorens S, Baeza-Ariño R, Nogales-Mérida S, Jover-Cerdá M, Tomás-Vidal A. Carob seed germ meal as a partial substitute in gilthead sea bream (Sparus aurata) diets: amino acid retention, digestibility, gut and liver histology. Aquaculture. 2012;338-341:124–33.Baeza-Ariño R, Martínez-Llorens S, Nogales-Mérida S, Jover-Cerda M, Tomás-Vidal A. Study of liver and gut alterations in sea bream, Sparus aurata L., fed a mixture of vegetable protein concentrates. Aquac Res. 2014;47(2):460–71.Estruch G, Collado MC, Peñaranda DS, Tomás Vidal A, Jover Cerdá M, Pérez Martínez G, et al. Impact of fishmeal replacement in diets for gilthead sea bream (Sparus aurata) on the gastrointestinal microbiota determined by pyrosequencing the 16S rRNA gene. PLoS One. 2015;10(8):e0136389. https://doi.org/10.1371/journal.pone.0136389 .Fekete SG, Kellems RO. Interrelationship of feeding with immunity and parasitic infection: a review. Vet Med. 2007;52(4):131–43.Kiron V. Fish immune system and its nutritional modulation for preventive health care. Anim Feed Sci Technol. 2012;173(1–2):111–33.Minghetti M, Drieschner C, Bramaz N, Schug H, Schirmer K. A fish intestinal epithelial barrier model established from the rainbow trout (Oncorhynchus mykiss) cell line, RTgutGC. Cell Biol Toxicol. 2017;33:539–55.Cerezuela R, Meseguer J, Esteban MÁ. Effects of dietary inulin, Bacillus subtilis and microalgae on intestinal gene expression in gilthead seabream (Sparus aurata L.). Fish Shellfish Immunol. 2013;34(3):843–8.Couto A, Kortner TM, Penn M, Bakke AM, Krogdahl O-TA, et al. Effects of dietary soy saponins and phytosterols on gilthead sea bream (Sparus aurata) during the on-growing period. Anim Feed Sci Technol. 2014;198:203–14.Estensoro I, Calduch-Giner JA, Kaushik S, Pérez-Sánchez J, Sitjá-Bobadilla A. Modulation of the IgM gene expression and IgM immunoreactive cell distribution by the nutritional background in gilthead sea bream (Sparus aurata) challenged with Enteromyxum leei (Myxozoa). Fish Shellfish Immunol. 2012;33(2):401–10.Pérez-Sánchez J, Estensoro I, Redondo MJ, Calduch-Giner JA, Kaushik S, Sitjà-Bobadilla A. Mucins as diagnostic and prognostic biomarkers in a fish-parasite model: transcriptional and functional analysis. PLoS One. 2013;8(6):e65457.Reyes-Becerril M, Guardiola F, Rojas M, Ascencio-Valle F, Esteban MÁ. Dietary administration of microalgae Navicula sp. affects immune status and gene expression of gilthead seabream (Sparus aurata). Fish Shellfish Immunol. 2013;35(3):883–9.Pérez-Sánchez J, Benedito-Palos L, Estensoro I, Petropoulos Y, Calduch-Giner JA, Browdy CL, et al. Effects of dietary NEXT ENHANCE ® 150 on growth performance and expression of immune and intestinal integrity related genes in gilthead sea bream (Sparus aurata L.). Fish Shellfish Immunol. 2015;44:117–28.Estensoro I, Ballester-Lozano G, Benedito-Palos L, Grammes F, Martos-Sitcha JA, Mydland L-T, et al. Dietary butyrate helps to restore the intestinal status of a marine teleost (Sparus aurata) fed extreme diets low in fish meal and fish oil. PLoS One. 2016;11(11):1–21.Torrecillas S, Caballero MJ, Mompel D, Montero D, Zamorano MJ, Robaina L, et al. Disease resistance and response against Vibrio anguillarum intestinal infection in European seabass (Dicentrarchus labrax) fed low fish meal and fish oil diets. Fish Shellfish Immunol. 2017;67:302–11.Schmittgen TD, Livak KJ. Analyzing real-time PCR data by the comparative C T method. Nat Protoc. 2008;3(6):1101–8.Omnes MH, Silva FCP, Moriceau J, Aguirre P, Kaushik S, Gatesoupe F-J. Influence of lupin and rapeseed meals on the integrity of digestive tract and organs in gilthead seabream (Sparus aurata L.) and goldfish (Carassius auratus L.) juveniles. Aquac Nutr. 2015;21:223–33.Francis G, Makkar HPS, Becker K. Antinutritional factors present in plant-derived alternate fish feed ingredients and their effects in fish. Aquaculture. 2001;199:197–227.Gatlin DM III, Barrows FT, Brown P, Dabrowski K, Gaylord TG, Hardy RW, et al. Expanding the utilization of sustainable plant products in aquafeeds: a review. Aquac Res. 2007;38:551–79.Kader MA, Bulbul M, Koshio S, Ishikawa M, Yokoyama S, Nguyen BT, et al. Effect of complete replacement of fishmeal by dehulled soybean meal with crude attractants supplementation in diets for red sea bream, Pagrus major. Aquaculture. 2012;350-353:109–16.Gómez-Requeni P, Mingarro M, Calduch-Giner JA, Médale F, Martin SAM, Houlihan DF, et al. Protein growth performance, amino acid utilisation and somatotropic axis responsiveness to fish meal replacement by plant protein sources in gilthead sea bream (Sparus aurata). Aquaculture. 2004;232(1–4):493–510.Kader MA, Koshio S, Ishikawa M, Yokoyama S, Bulbul M. Supplemental effects of some crude ingredients in improving nutritive values of low fishmeal diets for red sea bream, Pagrus major. Aquaculture. 2010;308(3–4):136–44.Mai K, Li H, Ai Q, Duan Q, Xu W, Zhang C, et al. Effects of dietary squid viscera meal on growth and cadmium accumulation in tissues of Japanese seabass, Lateolabrax japonicus (Cuvier 1828). Aquac Res. 2006;37(11):1063–9.Peres H, Oliva-Teles A. The optimum dietary essential amino acid profile for gilthead seabream (Sparus aurata) juveniles. Aquaculture. 2009;296(1–2):81–6.Cho CY, Slinger SJ, Bayley HS. Bioenergetics of salmonid fishes: energy intake, expenditure and productivity. Comp Biochem Physiol Part B. 1982;73(1):25–41.Venou B, Alexis MN, Fountoulaki E, Haralabous J. Effects of extrusion and inclusion level of soybean meal on diet digestibility , performance and nutrient utilization of gilthead sea bream ( Sparus aurata ). Aquaculture. 2006;261:343–56.Pfaffl MW, Tichopad A, Prgomet C, Neuvians TP. Determination of stable housekeeping genes, differentially regulated target genes and sample integrity: BestKeeper-excel-based tool using pair-wise correlations. Biotechnol Lett. 2004;26:509–15.Terova G, Robaina L, Izquierdo M, Cattaneo A, Molinari S, Bernardini G, et al. PepT1 mRNA expression levels in sea bream (Sparus aurata) fed different plant protein sources. Springerplus. 2013;2:17.Bates JM, Akerlund J, Mittge E, Guillemin K. Intestinal alkaline phosphatase detoxifies lipopolysaccharide and prevents inflammation in zebrafish in response to the gut microbiota. Cell Host Microbe. 2007;2(6):371–82.Adamidou S, Nengas I, Henry M, Grigorakis K, Rigos G, Nikolopoulou D, et al. Growth, feed utilization, health and organoleptic characteristics of European seabass (Dicentrarchus labrax) fed extruded diets including low and high levels of three different legumes. Aquaculture. 2009;293(3–4):263–71.Daprà F, Gai F, Costanzo MT, Maricchiolo G, Micale V, Sicuro B, et al. Rice protein-concentrate meal as a potential dietary ingredient in practical diets for blackspot seabream Pagellus bogaraveo: a histological and enzymatic investigation. J Fish Biol. 2009;74(4):773–89.Overland M, Sorensen M, Storebakken T, Penn M, Krogdahl A, Skrede A. Pea protein concentrate substituting fish meal or soybean meal in diets for Atlantic salmon (Salmo salar)-effect on growth performance, nutrient digestibility, carcass composition, gut health, and physical feed quality. Aquaculture. 2009;288(3–4):305–11.Penn MH, Bendiksen EA, Campbell P, Krogdahl AS. High level of dietary pea protein concentrate induces enteropathy in Atlantic salmon (Salmo salar L.). Aquaculture. 2011;310(3–4):267–73.Hedrera MI, Galdames JA, Jimenez-Reyes MF, Reyes AE, Avendaño-Herrera R, Romero J, et al. Soybean meal induces intestinal inflammation in zebrafish larvae. PLoS One. 2013;8(7):1–10.Kokou F, Sarropoulou E, Cotou E, Rigos G, Henry M, Alexis M. Effects of fish meal replacement by a soybean protein on growth, histology, selected immune and oxidative status markers of Gilthead Sea bream, Sparus aurata. J World Aquac Soc. 2015;46(2):115–28.Kokou F, Sarropoulou E, Cotou E, Kentouri M, Alexis M, Rigos G. Effects of graded dietary levels of soy protein concentrate supplemented with methionine and phosphate on the immune and antioxidant responses of gilthead sea bream (Sparus aurata L.). Fish Shellfish Immunol. 2017;64:111–21.Calduch-Giner JA, Sitjà-Bobadilla A, Davey GC, Cairns MT, Kaushik S, Pérez-Sánchez J. Dietary vegetable oils do not alter the intestine transcriptome of gilthead sea bream (Sparus aurata), but modulate the transcriptomic response to infection with Enteromyxum leei. BMC Genomics. 2012;13(1):470.Piazzon MC, Galindo-Villegas J, Pereiro P, Estensoro I, Calduch-Giner JA, Gómez-Casado E, et al. Differential modulation of IgT and IgM upon parasitic, bacterial, viral, and dietary challenges in a perciform fish. Front Immunol. 2016;7. Article 637. https://doi.org/10.3389/fimmu.2016.00637 .Salinas I, Zhang Y, Sunyer JO. Mucosal immunoglobulins and B cells of teleost fish. Dev Comp Immunol. 2011;35(12):1346–65.Krogdahl A, Bakke-McKellep AM, Roed KH, Baeverfjord G. Feeding Atlantic salmon Salmo salar L. soybean products: effects on disease resistance (furunculosis), and lysozyme and IgM levels in the intestinal mucosa. Aquac Nutr. 2000;6:77–84.Chasiotis H, Effendi JC, Kelly SP. Occludin expression in goldfish held in ion-poor water. J Comp Physiol B Biochem Syst Environ Physiol. 2009;179(2):145–54.Chen KT, Malo MS, Beasley-Topliffe LK, Poelstra K, Millan JL, Mostafa G, et al. A role for intestinal alkaline phosphatase in the maintenance of local gut immunity. Dig Dis Sci. 2011;56(4):1020–7.Vaishnava S, Hooper LV. Alkaline phosphatase: keeping the peace at the gut epithelial surface. Cell Host Microbe. 2007;2(6):365–7.Tort L. Stress and immune modulation in fish. Dev Comp Immunol [internet]. Elsevier Ltd. 2011;35(12):1366–75.Martin SAM, Król E. Nutrigenomics and immune function in fish: new insights from omics technologies. Dev Comp Immunol. 2017;75:86–98.Burrells C, Williams PD, Southgate PJ, Crampton VO. Immunological , physiological and pathological responses of rainbow trout (Oncorhynchus mykiss) to increasing dietary concentrations of soybean proteins. Vet Immunol Immunopathol. 1999;72:277–88.Sahlmann C, Sutherland BJG, Kortner TM, Koop BF, Krogdahl Å, Bakke AM. Early response of gene expression in the distal intestine of Atlantic salmon (Salmo salar L.) during the development of soybean meal induced enteritis. Fish Shellfish Immunol. 2013;34(2):599–609.Esteban MÁ, Cuesta A, Ortuño J, Meseguer J. Immunomodulatory effects of dietary intake of chitin on gilthead seabream ( Sparus aurata L .) innate immune system. Fish Shellfish Immunol. 2001;11:303–15.Storebakken T, Kvien IS, Shearer KD, Grisdale-Helland B, Helland SJ. Estimation of gastrointestinal evacuation rate in Atlantic salmon (Salmo salar) using inert markers and collection of faeces by sieving: evacuation of diets with fish meal, soybean meal or bacterial meal. Aquaculture. 1999;172(3–4):291–9.Olsen RE, Myklebust R, Ringø E, Mayhew TM. The influences of dietary linseed oil and saturated fatty acids on caecal enterocytes in Arctic char (Salvelinus alpinus L.): a quantitative ultrastructural study. Fish Physiol Biochem. 2000;22(3):207–16.Heikkinen J, Vielma J, Kemiläinen O, Tiirola M, Eskelinen P, Kiuru T, et al. Effects of soybean meal based diet on growth performance, gut histopathology and intestinal microbiota of juvenile rainbow trout (Oncorhynchus mykiss). Aquaculture. 2006;261(1):259–68.Krogdahl A, Bakke-McKellep AM, Baeverfjord G. Effects of graded levels of standard soybean meal on intestinal structure, mucosal enzyme activities, and pancreatic response in Atlantic salmon (Salmo salar L.). Aquac Nutr. 2003;9:361–71.Cerezuela R, Fumanal M, Tapia-Paniagua ST, Meseguer J, Moriñigo MA, Esteban MA. Changes in intestinal morphology and microbiota caused by dietary administration of inulin and Bacillus subtilis in gilthead sea bream (Sparus aurata L.) specimens. Fish Shellfish Immunol. 2013;34(5):1063–70.Cerezuela R, Fumanal M, Tapia-Paniagua ST, Meseguer J, Moriñigo MÁ, Esteban MÁ. Histological alterations and microbial ecology of the intestine in gilthead seabream (Sparus aurata L.) fed dietary probiotics and microalgae. Cell Tissue Res. 2012;350(3):477–89.Deplancke B, Gaskins HR. Microbial modulation of innate defense: goblet cells and the intestinal mucus layer. Am J Clin Nutr. 2001;73(suppl):1131S–41S.Kokou F, Rigos G, Henry M, Kentouri M, Alexis M. Growth performance, feed utilization and non-specific immune response of gilthead sea bream (Sparus aurata L.) fed graded levels of a bioprocessed soybean meal. Aquaculture. 2012;364-365:74–81

Growth, sensory and chemical characterization of Mediterranean yellowtail (Seriola dumerili) fed diets with partial replacement of fish meal by other protein sources

[EN] An 84-day trial was performed to assess the use of alternative protein sources in Seriola dumerili. Three diets were used, FM100 diet, as a control diet without fishmeal substitution, and FM66 and FM33 diets with a fishmeal replacement of 330 g/kg and 660 g/kg, respectively. At the end of experiment, fish fed the FM66 diet showed the no differences in growth, nutritional parameters and fatty acid composition. Heavy metals present some differences but are always lower than risk levels. In sensory analysis, differences between diets appeared in pH and color, and also in some texture parameters between FM33 and the other two diets. No differences appeared between diets related to flavor. In summary, long periods of feeding with high fish meal substitution diets, affects Seriola dumerili growth; despite this the quality of the fillet was not affected even with a 66 % of substitution.This project was financed by "Generalitat Valenciana. Ayudas para grupos de investigacion consolidables."Monge-Ortiz, R.; Martínez-Llorens, S.; Lemos-Neto, M.; Falco, S.; Pagán Moreno, MJ.; Godoy-Olmos, S.; Jover Cerda, M.... (2020). Growth, sensory and chemical characterization of Mediterranean yellowtail (Seriola dumerili) fed diets with partial replacement of fish meal by other protein sources. Aquaculture Reports. 18:1-10. https://doi.org/10.1016/j.aqrep.2020.100466S11018Abbas, K. A., Mohamed, A., Jamilah, B., & Ebrahimian, M. (2008). A Review on Correlations between Fish Freshness and pH during Cold Storage. American Journal of Biochemistry and Biotechnology, 4(4), 416-421. doi:10.3844/ajbbsp.2008.416.421Álvarez, A., García García, B., Garrido, M. D., & Hernández, M. D. (2008). The influence of starvation time prior to slaughter on the quality of commercial-sized gilthead seabream (Sparus aurata) during ice storage. Aquaculture, 284(1-4), 106-114. doi:10.1016/j.aquaculture.2008.07.025AMIARD, J., AMIARDTRIQUET, C., BARKA, S., PELLERIN, J., & RAINBOW, P. (2006). Metallothioneins in aquatic invertebrates: Their role in metal detoxification and their use as biomarkers. Aquatic Toxicology, 76(2), 160-202. doi:10.1016/j.aquatox.2005.08.015Baeverfjord, G., Refstie, S., Krogedal, P., & Åsgård, T. (2006). Low feed pellet water stability and fluctuating water salinity cause separation and accumulation of dietary oil in the stomach of rainbow trout (Oncorhynchus mykiss). Aquaculture, 261(4), 1335-1345. doi:10.1016/j.aquaculture.2006.08.033Baeza-Ariño, R., Martínez-Llorens, S., Nogales-Mérida, S., Jover-Cerda, M., & Tomás-Vidal, A. (2014). Study of liver and gut alterations in sea bream,Sparus aurataL., fed a mixture of vegetable protein concentrates. Aquaculture Research, 47(2), 460-471. doi:10.1111/are.12507Bell, J. G., McEvoy, J., Tocher, D. R., McGhee, F., Campbell, P. J., & Sargent, J. R. (2001). Replacement of Fish Oil with Rapeseed Oil in Diets of Atlantic Salmon (Salmo salar) Affects Tissue Lipid Compositions and Hepatocyte Fatty Acid Metabolism. The Journal of Nutrition, 131(5), 1535-1543. doi:10.1093/jn/131.5.1535Benedito-Palos, L., Navarro, J. C., Sitjà-Bobadilla, A., Gordon Bell, J., Kaushik, S., & Pérez-Sánchez, J. (2008). High levels of vegetable oils in plant protein-rich diets fed to gilthead sea bream (Sparus aurataL.): growth performance, muscle fatty acid profiles and histological alterations of target tissues. British Journal of Nutrition, 100(5), 992-1003. doi:10.1017/s0007114508966071Bjerkeng, B., Refstie, S., Fjalestad, K. T., Storebakken, T., Rødbotten, M., & Roem, A. J. (1997). Quality parameters of the flesh of Atlantic salmon (Salmo salar) as affected by dietary fat content and full-fat soybean meal as a partial substitute for fish meal in the diet. Aquaculture, 157(3-4), 297-309. doi:10.1016/s0044-8486(97)00162-2De Francesco, M., Parisi, G., Médale, F., Lupi, P., Kaushik, S. J., & Poli, B. M. (2004). Effect of long-term feeding with a plant protein mixture based diet on growth and body/fillet quality traits of large rainbow trout (Oncorhynchus mykiss). Aquaculture, 236(1-4), 413-429. doi:10.1016/j.aquaculture.2004.01.006DE FRANCESCO, M., PARISI, G., PÉREZ-SÁNCHEZ, J., GÓMEZ-RéQUENI, P., MÉDALE, F., KAUSHIK, S. J., … POLI, B. M. (2007). Effect of high-level fish meal replacement by plant proteins in gilthead sea bream (Sparus aurata) on growth and body/fillet quality traits. Aquaculture Nutrition, 13(5), 361-372. doi:10.1111/j.1365-2095.2007.00485.xEstruch, G., Collado, M. C., Peñaranda, D. S., Tomás Vidal, A., Jover Cerdá, M., Pérez Martínez, G., & Martinez-Llorens, S. (2015). Impact of Fishmeal Replacement in Diets for Gilthead Sea Bream (Sparus aurata) on the Gastrointestinal Microbiota Determined by Pyrosequencing the 16S rRNA Gene. PLOS ONE, 10(8), e0136389. doi:10.1371/journal.pone.0136389Estruch, G., Collado, M. C., Monge-Ortiz, R., Tomás-Vidal, A., Jover-Cerdá, M., Peñaranda, D. S., … Martínez-Llorens, S. (2018). Long-term feeding with high plant protein based diets in gilthead seabream (Sparus aurata, L.) leads to changes in the inflammatory and immune related gene expression at intestinal level. BMC Veterinary Research, 14(1). doi:10.1186/s12917-018-1626-6Estruch, G., Tomás-Vidal, A., El Nokrashy, A. M., Monge-Ortiz, R., Godoy-Olmos, S., Jover Cerdá, M., & Martínez-Llorens, S. (2018). Inclusion of alternative marine by-products in aquafeeds with different levels of plant-based sources for on-growing gilthead sea bream (Sparus aurata, L.): effects on digestibility, amino acid retention, ammonia excretion and enzyme activity. Archives of Animal Nutrition, 72(4), 321-339. doi:10.1080/1745039x.2018.1472408Estruch, G., Martínez-Llorens, S., Tomás-Vidal, A., Monge-Ortiz, R., Jover-Cerdá, M., Brown, P. B., & Peñaranda, D. S. (2020). Impact of high dietary plant protein with or without marine ingredients in gut mucosa proteome of gilthead seabream (Sparus aurata, L.). Journal of Proteomics, 216, 103672. doi:10.1016/j.jprot.2020.103672Fountoulaki, E., Vasilaki, A., Hurtado, R., Grigorakis, K., Karacostas, I., Nengas, I., … Alexis, M. N. (2009). Fish oil substitution by vegetable oils in commercial diets for gilthead sea bream (Sparus aurata L.); effects on growth performance, flesh quality and fillet fatty acid profile. Aquaculture, 289(3-4), 317-326. doi:10.1016/j.aquaculture.2009.01.023Francis, G., Makkar, H. P. ., & Becker, K. (2001). Antinutritional factors present in plant-derived alternate fish feed ingredients and their effects in fish. Aquaculture, 199(3-4), 197-227. doi:10.1016/s0044-8486(01)00526-9Korkmaz Görür, F., Keser, R., Akçay, N., & Dizman, S. (2012). Radioactivity and heavy metal concentrations of some commercial fish species consumed in the Black Sea Region of Turkey. Chemosphere, 87(4), 356-361. doi:10.1016/j.chemosphere.2011.12.022Hu, L., Yun, B., Xue, M., Wang, J., Wu, X., Zheng, Y., & Han, F. (2013). Effects of fish meal quality and fish meal substitution by animal protein blend on growth performance, flesh quality and liver histology of Japanese seabass (Lateolabrax japonicus). Aquaculture, 372-375, 52-61. doi:10.1016/j.aquaculture.2012.10.025Izquierdo, M. S., Obach, A., Arantzamendi, L., Montero, D., Robaina, L., & Rosenlund, G. (2003). Dietary lipid sources for seabream and seabass: growth performance, tissue composition and flesh quality. Aquaculture Nutrition, 9(6), 397-407. doi:10.1046/j.1365-2095.2003.00270.xIzquierdo, M. S., Montero, D., Robaina, L., Caballero, M. J., Rosenlund, G., & Ginés, R. (2005). Alterations in fillet fatty acid profile and flesh quality in gilthead seabream (Sparus aurata) fed vegetable oils for a long term period. Recovery of fatty acid profiles by fish oil feeding. Aquaculture, 250(1-2), 431-444. doi:10.1016/j.aquaculture.2004.12.001Jover, M., Garcı́a-Gómez, A., Tomás, A., De la Gándara, F., & Pérez, L. (1999). Growth of mediterranean yellowtail (Seriola dumerilii) fed extruded diets containing different levels of protein and lipid. Aquaculture, 179(1-4), 25-33. doi:10.1016/s0044-8486(99)00149-0Martínez-Llorens, S., Baeza-Ariño, R., Nogales-Mérida, S., Jover-Cerdá, M., & Tomás-Vidal, A. (2012). Carob seed germ meal as a partial substitute in gilthead sea bream (Sparus aurata) diets: Amino acid retention, digestibility, gut and liver histology. Aquaculture, 338-341, 124-133. doi:10.1016/j.aquaculture.2012.01.029MARTINS, D. A., VALENTE, L. M. P., & LALL, S. P. (2011). Partial replacement of fish oil by flaxseed oil in Atlantic halibut (Hippoglossus hippoglossus L.) diets: effects on growth, nutritional and sensory quality. Aquaculture Nutrition, 17(6), 671-684. doi:10.1111/j.1365-2095.2011.00869.xMatallanas, J., Casadevall, M., Carrasson, M., Bolx, J., & Fernandez, V. (1995). The Food of Seriola Dumerili (Pisces: Carangidae) in the Catalan Sea (Western Mediterranean). Journal of the Marine Biological Association of the United Kingdom, 75(1), 257-260. doi:10.1017/s0025315400015356Monge-Ortiz, R., Tomás-Vidal, A., Gallardo-Álvarez, F. J., Estruch, G., Godoy-Olmos, S., Jover-Cerdá, M., & Martínez-Llorens, S. (2018). Partial and total replacement of fishmeal by a blend of animal and plant proteins in diets for Seriola dumerili : Effects on performance and nutrient efficiency. Aquaculture Nutrition, 24(4), 1163-1174. doi:10.1111/anu.12655Monge-Ortiz, R., Tomás-Vidal, A., Rodriguez-Barreto, D., Martínez-Llorens, S., Pérez, J. A., Jover-Cerdá, M., & Lorenzo, A. (2017). Replacement of fish oil with vegetable oil blends in feeds for greater amberjack (Seriola dumerili) juveniles: Effect on growth performance, feed efficiency, tissue fatty acid composition and flesh nutritional value. Aquaculture Nutrition, 24(1), 605-615. doi:10.1111/anu.12595Mourente, G., & Bell, J. G. (2006). Partial replacement of dietary fish oil with blends of vegetable oils (rapeseed, linseed and palm oils) in diets for European sea bass (Dicentrarchus labrax L.) over a long term growth study: Effects on muscle and liver fatty acid composition and effectiveness of a fish oil finishing diet. Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology, 145(3-4), 389-399. doi:10.1016/j.cbpb.2006.08.012Nanton, D. A., Vegusdal, A., Rørå, A. M. B., Ruyter, B., Baeverfjord, G., & Torstensen, B. E. (2007). Muscle lipid storage pattern, composition, and adipocyte distribution in different parts of Atlantic salmon (Salmo salar) fed fish oil and vegetable oil. Aquaculture, 265(1-4), 230-243. doi:10.1016/j.aquaculture.2006.03.053O’Fallon, J. V., Busboom, J. R., Nelson, M. L., & Gaskins, C. T. (2007). A direct method for fatty acid methyl ester synthesis: Application to wet meat tissues, oils, and feedstuffs. Journal of Animal Science, 85(6), 1511-1521. doi:10.2527/jas.2006-491Olsen, R. L., & Toppe, J. (2017). Fish silage hydrolysates: Not only a feed nutrient, but also a useful feed additive. Trends in Food Science & Technology, 66, 93-97. doi:10.1016/j.tifs.2017.06.003De Paiva, E. L., Alves, J. C., Milani, R. F., Boer, B. S., Quintaes, K. D., & Morgano, M. A. (2016). Sushi commercialized in Brazil: Organic Hg levels and exposure intake evaluation. Food Control, 69, 115-123. doi:10.1016/j.foodcont.2016.04.029Panserat, S., Hortopan, G. A., Plagnes-Juan, E., Kolditz, C., Lansard, M., Skiba-Cassy, S., … Corraze, G. (2009). Differential gene expression after total replacement of dietary fish meal and fish oil by plant products in rainbow trout (Oncorhynchus mykiss) liver. Aquaculture, 294(1-2), 123-131. doi:10.1016/j.aquaculture.2009.05.013Piazzon, M. C., Calduch-Giner, J. A., Fouz, B., Estensoro, I., Simó-Mirabet, P., Puyalto, M., … Pérez-Sánchez, J. (2017). Under control: how a dietary additive can restore the gut microbiome and proteomic profile, and improve disease resilience in a marine teleostean fish fed vegetable diets. Microbiome, 5(1). doi:10.1186/s40168-017-0390-3Regost, C., Arzel, J., Robin, J., Rosenlund, G., & Kaushik, S. . (2003). Total replacement of fish oil by soybean or linseed oil with a return to fish oil in turbot (Psetta maxima). Aquaculture, 217(1-4), 465-482. doi:10.1016/s0044-8486(02)00259-4Robaina, L., Izquierdo, M. ., Moyano, F. ., Socorro, J., Vergara, J. ., & Montero, D. (1998). Increase of the dietary n−3/n−6 fatty acid ratio and addition of phosphorus improves liver histological alterations induced by feeding diets containing soybean meal to gilthead seabream, Sparus aurata. Aquaculture, 161(1-4), 281-293. doi:10.1016/s0044-8486(97)00276-7Serradell, A., Torrecillas, S., Makol, A., Valdenegro, V., Fernández-Montero, A., Acosta, F., … Montero, D. (2020). Prebiotics and phytogenics functional additives in low fish meal and fish oil based diets for European sea bass (Dicentrarchus labrax): Effects on stress and immune responses. Fish & Shellfish Immunology, 100, 219-229. doi:10.1016/j.fsi.2020.03.016Shimeno, S., Masumoto, T., Hujita, T., Mima, T., & Ueno, S. (1993). Protein Source for Fish Feed-V. Alternative Protein Sources for Fish Meal in Diets of Young Yellowtail. NIPPON SUISAN GAKKAISHI, 59(1), 137-143. doi:10.2331/suisan.59.137Sitjà-Bobadilla, A., Peña-Llopis, S., Gómez-Requeni, P., Médale, F., Kaushik, S., & Pérez-Sánchez, J. (2005). Effect of fish meal replacement by plant protein sources on non-specific defence mechanisms and oxidative stress in gilthead sea bream (Sparus aurata). Aquaculture, 249(1-4), 387-400. doi:10.1016/j.aquaculture.2005.03.031SLOTH, J. J., JULSHAMN, K., & LUNDEBYE, A.-K. (2005). Total arsenic and inorganic arsenic content in Norwegian fish feed products. Aquaculture Nutrition, 11(1), 61-66. doi:10.1111/j.1365-2095.2004.00334.xStergiou, K. I., & Karpouzi, V. S. (2001). Reviews in Fish Biology and Fisheries, 11(3), 217-254. doi:10.1023/a:1020556722822Thakur, D. P., Morioka, K., Itoh, N., Wada, M., & Itoh, Y. (2009). Muscle biochemical constituents of cultured amberjack Seriola dumerili and their influence on raw meat texture. Fisheries Science, 75(6), 1489-1498. doi:10.1007/s12562-009-0173-2TOMAS, A., DE LA GANDARA, F., GARCIA-GOMEZ, A., PEREZ, L., & JOVER, M. (2005). Utilization of soybean meal as an alternative protein source in the Mediterranean yellowtail, Seriola dumerili. Aquaculture Nutrition, 11(5), 333-340. doi:10.1111/j.1365-2095.2005.00365.xTomás‐Vidal, A., Monge‐Ortiz, R., Jover‐Cerdá, M., & Martínez‐Llorens, S. (2019). Apparent digestibility and protein quality evaluation of selected feed ingredients in Seriola dumerili. Journal of the World Aquaculture Society, 50(4), 842-855. doi:10.1111/jwas.12597Torstensen, B. E., Bell, J. G., Rosenlund, G., Henderson, R. J., Graff, I. E., Tocher, D. R., … Sargent, J. R. (2005). Tailoring of Atlantic Salmon (Salmo salar L.) Flesh Lipid Composition and Sensory Quality by Replacing Fish Oil with a Vegetable Oil Blend. Journal of Agricultural and Food Chemistry, 53(26), 10166-10178. doi:10.1021/jf051308iTurchini, G. M., Moretti, V. M., Mentasti, T., Orban, E., & Valfrè, F. (2007). Effects of dietary lipid source on fillet chemical composition, flavour volatile compounds and sensory characteristics in the freshwater fish tench (Tinca tinca L.). Food Chemistry, 102(4), 1144-1155. doi:10.1016/j.foodchem.2006.07.003Valente, L. M. P., Linares, F., Villanueva, J. L. R., Silva, J. M. G., Espe, M., Escórcio, C., … Peleteiro, J. B. (2011). Dietary protein source or energy levels have no major impact on growth performance, nutrient utilisation or flesh fatty acids composition of market-sized Senegalese sole. Aquaculture, 318(1-2), 128-137. doi:10.1016/j.aquaculture.2011.05.026Watanabe, K., Ura, K., Yada, T., Kiron, V., Satoh, S., & Watanabe, T. (2000). Energy and protein requirements of yellowtail for maximum growth and maintenance of body weight. Fisheries Science, 66(6), 1053-1061. doi:10.1046/j.1444-2906.2000.00168.

Single-Cell Transcriptional Analysis Reveals Novel Neuronal Phenotypes and Interaction Networks Involved in the Central Circadian Clock

Single-cell heterogeneity confounds efforts to understand how a population of cells organizes into cellular networks that underlie tissue-level function. This complexity is prominent in the mammalian suprachiasmatic nucleus (SCN). Here, individual neurons exhibit a remarkable amount of asynchronous behavior and transcriptional heterogeneity. However, SCN neurons are able to generate precisely coordinated synaptic and molecular outputs that synchronize the body to a common circadian cycle by organizing into cellular networks. To understand this emergent cellular network property, it is important to reconcile single-neuron heterogeneity with network organization. In light of recent studies suggesting that transcriptionally heterogeneous cells organize into distinct cellular phenotypes, we characterized the transcriptional, spatial, and functional organization of 352 SCN neurons from mice experiencing phase-shifts in their circadian cycle. Using the community structure detection method and multivariate analytical techniques, we identified previously undescribed neuronal phenotypes that are likely to participate in regulatory networks with known SCN cell types. Based on the newly discovered neuronal phenotypes, we developed a data-driven neuronal network structure in which multiple cell types interact through known synaptic and paracrine signaling mechanisms. These results provide a basis from which to interpret the functional variability of SCN neurons and describe methodologies toward understanding how a population of heterogeneous single cells organizes into cellular networks that underlie tissue-level function

Identifying niche mediated regulatory factors of stem cell phenotypic state: a systems biology approach

Understanding how the cellular niche controls the stem cell phenotype is often hampered due to the complexity of variegated niche composition, its dynamics, and nonlinear stem cell–niche interactions. Here, we propose a systems biology view that considers stem cell–niche interactions as a many‐body problem amenable to simplification by the concept of mean field approximation. This enables approximation of the niche effect on stem cells as a constant field that induces sustained activation/inhibition of specific stem cell signaling pathways in all stem cells within heterogeneous populations exhibiting the same phenotype (niche determinants). This view offers a new basis for the development of single cell‐based computational approaches for identifying niche determinants, which has potential applications in regenerative medicine and tissue engineering

LRIG1 is a gatekeeper to exit from quiescence in adult neural stem cells

Adult neural stem cells (NSCs) must tightly regulate quiescence and proliferation. Single-cell analysis has suggested a continuum of cell states as NSCs exit quiescence. Here we capture and characterize in vitro primed quiescent NSCs and identify LRIG1 as an important regulator. We show that BMP-4 signaling induces a dormant non-cycling quiescent state (d-qNSCs), whereas combined BMP-4/FGF-2 signaling induces a distinct primed quiescent state poised for cell cycle re-entry. Primed quiescent NSCs (p-qNSCs) are defined by high levels of LRIG1 and CD9, as well as an interferon response signature, and can efficiently engraft into the adult subventricular zone (SVZ) niche. Genetic disruption of Lrig1 in vivo within the SVZ NSCs leads an enhanced proliferation. Mechanistically, LRIG1 primes quiescent NSCs for cell cycle re-entry and EGFR responsiveness by enabling EGFR protein levels to increase but limiting signaling activation. LRIG1 is therefore an important functional regulator of NSC exit from quiescence

Decoding the Regulatory Logic of the Drosophila Male Stem Cell System

The niche critically controls stem cell behavior, but its regulatory input at the whole-genome level is poorly understood. We elucidated transcriptional programs of the somatic and germline lineages in the Drosophila testis and genome-wide binding profiles of Zfh-1 and Abd-A expressed in somatic support cells and crucial for fate acquisition of both cell lineages. We identified key roles of nucleoporins and V-ATPase proton pumps and demonstrate their importance in controlling germline development from the support side. To make our dataset publicly available, we generated an interactive analysis tool, which uncovered conserved core genes of adult stem cells across species boundaries. We tested the functional relevance of these genes in the Drosophila testis and intestine and found a high frequency of stem cell defects. In summary, our dataset and interactive platform represent versatile tools for identifying gene networks active in diverse stem cell type