iBET

Brain research

The development of drugs for human central nervous system (CNS) diseases has traditionally relied on genetically engineered mouse models (GEMMs). However, the CNS is highly complex and also, GEMMs’ phenotypes often diverge from that of human diseases. These factors represent the main obstacle to the translation of CNS drugs pre-clinical to the clinical trials.

Therefore, 3D in vitro models are useful complementary tools towards more accurate evaluation of drug candidates in pre-clinical stages, as they present an intermediate degree of complexity in terms of cell-cell and cell-matrix interactions, between the traditional 2D monolayer culture conditions and the complex brain and can be a better starting point for the analysis of the in vivo situation. We have been developing brain models using primary cultures of rat brain cells and cultures of differentiated human embryonal carcinoma cell lines and human neural stem cells.

Furthermore, we are exploring our 3D brain cell models to reveal additional layers of information, usually concealed by the high complexity of interactions that occur in vivo. Using metabolic flux analysis tools and data on 13C enrichment into intra and extracellular metabolites (NMR and GC-MS analysis) we have been quantifying the metabolic effects caused by specific insults which frequently occur in the brain.

Selected Publications

  1. Materne EM, Ramme AP, Terrasso AP, Serra M, Alves PM, Brito C, Sakharov DA, Tonevitsky AG, Lauster R, Marx U. 2015. A multi-organ chip co-culture of neurospheres and liver equivalents for long-term substance testing. J Biotechnol. Feb 9. pii: S0168-1656(15)00045-0. 
  2. Terrasso AP, Pinto C, Serra M, Filipe A, Almeida S, Ferreira AL, Pedroso P, Brito C, Alves PM. Novel scalable 3D cell based model for in vitro neurotoxicity testing: Combining human differentiated neurospheres with gene expression and functional endpoints. J Biotechnol. 2015 Jan 5. pii: S0168-1656(14)01047-5. 
  3. Simao D, Pinto C, Piersanti S, Weston A, Peddie CJ, Bastos AEP, Licursi V, Schwarz, S. C., Collinson, L. M., Salinas S, Serra M, Teixeira AP, Saggio I, Lima PA, Kremer EJ, Schiavo G, Brito C & Alves PM. 2015. Modeling Human Neural Functionality In Vitro: Three-Dimensional Culture for Dopaminergic Differentiation.Tissue Engineering Part A 21, 654-668. 
  4. Gualda EJ, Simão D, Pinto C, Alves PM, Brito C. 2014. Imaging of human differentiated 3D neural aggregates using light sheet fluorescence
    Frontiers in Cellular Neuroscience. 8:221
  5. Almeida AS, Queiroga CS, Sousa MF, Alves PM, Vieira HL, 2012. Carbon monoxide modulates apoptosis by reinforcing oxidative metabolism in astrocytes: role of BCL-2. J Biol Chem, 287 (14): 10761-70. 
  6. Brito C, Simão D, Costa I, Malpique R, Pereira CI, Fernandes P, Serra M, Schwarz SC, Schwarz J, Kremer EJ, Alves PM, 2012. Generation and genetic modification of 3D cultures of human dopaminergic neurons derived from neural progenitor cells. Methods, 56(3):452-60. 
  7. Queiroga CS, Tomasi S, Wideroe M, Alves PM, Vercelli A, Vieira HL, 2012. Preconditioning triggered by Carbon Monoxide (CO) provides neuronal protection following perinatal hypoxia-ischemia. Plos One, 7(8): e42632. 
  8. Vieira HL, Alves PM, Vercelli A, 2011. Modulation of neuronal stem cell differentiation by hypoxia and reactive oxygen species. Prog Neurobiol, 93:444-55. 
  9. Santos SS, Sonnewald U, Carrondo MJ, Alves PM, 2011. The role of glia in neuronal recovery following anoxia: in vitro evidence of neuronal adaptation. Neurochem Int, 58:665-75. 
  10. Amaral AI , Teixeira AP, Sonnewald U, Alves PM, 2011. Estimation of intracellular fluxes in cerebellar neurons after hypoglycemia: importance of the pyruvate recycling pathway and glutamine oxidation. J Neurosci Res, 89(5)-700:10. 1
  11. Amaral IA, Teixeira AP, Håkonsen BI, Sonnewald U, Alves PM, 2011. A comprehensive metabolic profile of cultured astrocytes using isotopic transient metabolic flux analysis and 13C-labelled glucose. Front Neuroenerg, 3:5. 
  12. Amaral AI, Teixeira AP, Martens S, Sousa MF, Bernal V, Alves PM, 2010. Metabolic alterations induced by ischemia in primary cultures of astrocytes: merging 13C NMR spectroscopy and metabolic flux analysis. J Neurochem, 113(3):735-48. 
  13. Serra M, Brito C, Costa E, Sousa MF, Alves PM, 2009. Integrating human stem cell expansion and neuronal differentiation in bioreactors. BMC Biotechnol, 9:82. 
  14. Vieira HL, Queiroga CS, Alves PM, 2008. Preconditioning induced by Carbon Monoxide provides neuronal protection against apoptosis. J Neurochem, 107:375-84. 
  15. Teixeira AP, Santos SS, Carinhas N, Oliveira R, Alves PM, 2008. Combining metabolic flux analysis tools and 13C-NMR to estimate intracellular fluxes of cultured astrocytes. Neurochem Int, 52:478-86. 
  16. Santos SS, Leite SB, Sonnewald U, Carrondo MJ, Alves PM, 2007. Stirrer vessel cultures of rat brain cells aggregates: characterization of major metabolic pathways and cell population dynamics. J Neurosci Res, 85(15):3386-97. 
  17. Serra M, Leite SB, Brito C, Costa J, Carrondo MJ, Alves PM, 2007. Novel culture strategy for human stem cell proliferation and neuronal differentiation. J Neurosci Res, 85(16):3557-66. 
  18. Santos SS, Gibson GE, Cooper AJ, Denton TT, Thompson CM, Bunik VI, Alves PM, Sonnewald U, 2006. Inhibitors of the a-ketoglutarate dehydrogenase complex alter [1-13C]Glucose and [U-13C]Glutamate metabolism in cerebellar granule neurons. J Neurosci Res, 83:450-8. 
  19. Fonseca LL, Monteiro MA, Alves PM, Carrondo MJ, Santos H, 2005. Cultures of rat astrocytes challenged with a steady supply of glutamate: a new model to study flux distribution in the glutamate-glutamine cycle. Glia, 51:286-96. 
  20. Santos SS, Fonseca LL, Monteiro MA, Carrondo MJ, Alves PM,  2005. Culturing primary brain astrocytes under fully controlled environment in a novel bioreactor. J Neurosci Res, 79:26-32. 
  21. Alves PM, Fonseca LL, Peixoto CC, Almeida AC, Carrondo MJ, Santos H, 2000. NMR studies on energy metabolism of immobilized primary neurons and astrocytes during hypoxia, ischemia and hypoglycemia. NMR Biomed, 13:438-48. 
  22. Alves PM, Nunes R, Zhang C, Maycock CD, Sonnewald U, Carrondo MJ, Santos H, 2000. Metabolism of 3-[13C]malate in primary cultures of mouse astrocytes. Dev Neurosci, 22:456-62. 
  23. Alves PM, Moreira JL, Rodrigues JM, Aunins JG, Carrondo MJ, 1996. Immobilization of primary astrocytes and neurons for on-line monitoring of biochemical processes by NMR. Dev Neurosci, 18:478-83.
  24. Alves PM, MC Mckenna, U Sonnewald, 1995. Lactate metabolism in mouse brain astrocytes studied by [13C]NMR spectroscopy. NeuroReport, 6:2201-04. 
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