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- Identification of new targets of S-nitrosylation in neural stem cells by thiol redox proteomicsPublication . Santos, Ana Isabel; Lourenco, Ana S.; Simão, Sónia; Marques-da-Silva, Dorinda; Santos, Daniela F.; Carvalho, Ana Paula Onofre de; Pereira, Ana Catarina; Izquierdo-Álvarez, Alicia; Ramos, Elena; Morato, Esperanza; Marina, Anabel; Martínez-Ruiz, Antonio; Araújo, InêsNitric oxide (NO) is well established as a regulator of neurogenesis. NO increases the proliferation of neural stem cells (NSC), and is essential for hippocampal injury-induced neurogenesis following an excitotoxic lesion. One of the mechanisms underlying non-classical NO cell signaling is protein S-nitrosylation. This post-translational modification consists in the formation of a nitrosothiol group (R-SNO) in cysteine residues, which can promote formation of other oxidative modifications in those cysteine residues. S-nitrosylation can regulate many physiological processes, including neuronal plasticity and neurogenesis. In this work, we aimed to identify S-nitrosylation targets of NO that could participate in neurogenesis. In NSC, we identified a group of proteins oxidatively modified using complementary techniques of thiol redox proteomics. S-nitrosylation of some of these proteins was confirmed and validated in a seizure mouse model of hippocampal injury and in cultured hippocampal stem cells. The identified S-nitrosylated proteins are involved in the ERK/MAPK pathway and may be important targets of NO to enhance the proliferation of NSC.
- New fluorescent probes based on gallium(III) corrole complexes for the recognition of hydrogen sulfide: a journey from solution to intracellular sitePublication . Santos, Carla I.M.; Santiago, Ana M.; Araújo, Ana R.L.; Pinto, Sandra; Agostinho, Rafaela; Simão, Sónia; Azevedo, Tomás Pais de; Antunes, Catarina; Faustino, M. Amparo F.; Araujo, Ines; Neves, M. Graça P.M.S.; Martinho, José M.G.; Maçôas, Ermelinda M.S.In this work, three fluorescent probes for detection of hydrogen sulfide (H2S) where prepared based on gallium (III) corrole complexes bearing nitro groups at beta-pyrrolic positions. Two of the compounds selected, the 3-nitro5,10,15- tris(pentafluorophenyl)corrolatogallium(III)(pyridine) (CGa-NO2) and the 3,17-dinitro-5,10,15-tris (pentafluorophenyl)corrolatogallium(III)(pyridine) (CGa-2NO2) present one and two nitro groups directly linked to the beta-pyrrolic position. The third compound, the (E)-3-(2-nitroprop-1-en-1-yl)-5,10,15-tris(pentafluorophenyl)corrolatogallium(III)(pyridine) (CGa-EtNO2), has a carbon-carbon double bond spacer between the corrole unit and the nitro group. All these derivatives were obtained from 5,10,15-tris(pentafluorophenyl)corrolatogallium(III)(pyridine) (CGa). The precursor CGa and the derivative CGa-EtNO2 behaved as turn-OFF probes, while compound CGa-NO2 responded as a turn-ON probe in the presence of H2S in the pH range of 5-9. Mechanistic studies show that the interaction of H2S with the probes involves its coordination with gallium(III) and in some cases the reduction of the nitro group to a new aminated corrole. While the formation of the coordination complex with H2S is almost immediate, the kinetics of the reduction is slow. Interestingly, for CGaNO2 the two processes can be explored in a ratiometric sensing of H2S in a non-aqueous solution showing a good linearity over an extended concentration range (5-200 mu M). The response of the corroles to H2S in intracellular medium was studied in 2D cultured cells (HeLa).
- Rewired glycosylation activity promotes scarless regeneration and functional recovery in spiny mice after complete spinal cord transectionPublication . Nogueira-Rodrigues, Joana; Leite, Sérgio C.; Pinto-Costa, Rita; Sousa, Sara C.; Luz, Liliana L.; Sintra, Maria A.; Oliveira, Raquel; Monteiro, Ana C.; Pinheiro, Gonçalo; Vitorino, Marta; Silva, Joana A.; S, Simão; Vitor Fernandes, Dr; Provazník, Jan; Benes, Vladimir; Cruz, Célia D.; Safronov, Boris V.; Magalhães, Ana; Reis, Celso A.; Vieira, Jorge; Vieira, Cristina P.; Tiscórnia, Gustavo; Araujo, Ines; Sousa, Mónica M.Regeneration of adult mammalian central nervous system (CNS) axons is abortive, resulting in inability to recover function after CNS lesion, including spinal cord injury (SCI). Here, we show that the spiny mouse (Acomys) is an exception to other mammals, being capable of spontaneous and fast restoration of function after severe SCI, re-establishing hind limb coordination. Remarkably, Acomys assembles a scarless pro-regenerative tissue at the injury site, providing a unique structural continuity of the initial spinal cord geometry. The Acomys SCI site shows robust axon regeneration of multiple tracts, synapse formation, and electrophysiological signal propagation. Transcriptomic analysis of the spinal cord following transcriptome reconstruction revealed that Acomys rewires glycosylation biosynthetic pathways, culminating in a specific pro-regenerative proteoglycan signature at SCI site. Our work uncovers that a glycosylation switch is critical for axon regeneration after SCI and identifies beta 3gnt7, a crucial enzyme of keratan sulfate biosynthesis, as an enhancer of axon growth.
- Measuring healthy ageing: current and future toolsPublication . Silva, Nádia; Rajado, Ana Teresa; Esteves, Filipa; Brito, David V.C.; Apolónio, Joana; Roberto, Vânia; Binnie, Alexandra; Araújo, Inês Maria; Nóbrega, Clévio; Bragança, José; Castelo-Branco, PedroHuman ageing is a complex, multifactorial process characterised by physiological damage, increased risk of age-related diseases and inevitable functional deterioration. As the population of the world grows older, placing significant strain on social and healthcare resources, there is a growing need to identify reliable and easy-to-employ markers of healthy ageing for early detection of ageing trajectories and disease risk. Such markers would allow for the targeted implementation of strategies or treatments that can lessen suffering, disability, and dependence in old age. In this review, we summarise the healthy ageing scores reported in the literature, with a focus on the past 5 years, and compare and contrast the variables employed. The use of approaches to determine biological age, molecular biomarkers, ageing trajectories, and multi-omics ageing scores are reviewed. We conclude that the ideal healthy ageing score is multisystemic and able to encompass all of the potential alterations associated with ageing. It should also be longitudinal and able to accurately predict ageing complications at an early stage in order to maximize the chances of successful early intervention.
- Coronal brain atlas in stereotaxic coordinates of the African spiny mouse, acomys cahirinusPublication . Vitorino, Marta; S, Simão; Moreira, João B.; Nogueira‐Rodrigues, Joana; Silva, Joana A; Sofia Lourenço, Ana; Vitor Fernandes, Dr; Sousa, Monica M.; Tiscornia, Gustavo; Araújo, Inês MariaThe African spiny mouse (Acomys cahirinus) is an emerging model of mammalian epimorphic regeneration that has aroused the interest of the scientific community in the last decade. To date, studies on brain repair have been hindered by the lack of knowledge on the neuroanatomy of this species. Here, we present a coronal brain atlas in stereotaxic coordinates, which allows for three-dimensional identification and localization of the brain structures of this species. The brain of 12-week-old spiny mice was mapped in stereotaxic coordinates using cresyl violet-stained brain sections obtained from coronal cryosectioning of the brain after transcardial perfusion with fixative. The atlas is presented in 42 plates representing sections spaced 240 mu m apart. Stereotaxic coordinates were validated using both a model of Parkinsonian lesion of the striatum with 6-hydroxydopamine and labeling of the corticospinal tract in the spiny mouse spinal cord using AAV1/2-GFP intracortical injections. This work presents a new tool in A. cahirinus neurobiology and opens new avenues of research for the investigation of the regenerative ability of A. cahirinus in models of brain disorders.
- Regulation of Ras signaling by S-NitrosylationPublication . S, Simão; Agostinho, Rafaela; Martínez-Ruiz, Antonio; Araújo, Inês MariaRas are a family of small GTPases that function as signal transduction mediators and are involved in cell proliferation, migration, differentiation and survival. The significance of Ras is further evidenced by the fact that Ras genes are among the most mutated oncogenes in different types of cancers. After translation, Ras proteins can be targets of post-translational modifications (PTM), which can alter the intracellular dynamics of the protein. In this review, we will focus on how S-nitrosylation of Ras affects the way these proteins interact with membranes, its cellular localization, and its activity. S-Nitrosylation occurs when a nitrosyl moiety of nitric oxide (NO) is covalently attached to a thiol group of a cysteine residue in a target protein. In Ras, the conserved Cys118 is the most surface-exposed Cys and the preferable residue for NO action, leading to the initiation of transduction events. Ras transduces the mitogen-activated protein kinases (MAPK), the phosphoinositide-3 kinase (PI3K) and the RalGEF cellular pathways. S-Nitrosylation of elements of the RalGEF cascade remains to be identified. On the contrary, it is well established that several components of the MAPK and PI3K pathways, as well as different proteins associated with these cascades, can be modified by S-nitrosylation. Overall, this review presents a better understanding of Ras S-nitrosylation, increasing the knowledge on the dynamics of these proteins in the presence of NO and the underlying implications in cellular signaling.
- Reprogramming iPSCs to study age-related diseases: models, therapeutics, and clinical trialsPublication . Esteves, Filipa; Brito, David; Rajado, Ana Teresa; Silva, Nádia; Apolónio, Joana; Roberto, Vania Palma; Araújo, Inês Maria; Nóbrega, Clévio; Castelo-Branco, Pedro; Bragança, José; P. Andrade, Raquel; M. Calado, Sofia; Faleiro, L; Matos, Carlos A; Marques, Nuno; Marreiros, Ana; Nzwalo, Hipólito; Pais, Sandra; Palmeirim, Isabel; S, Simão; Joaquim, Natércia; Miranda, Rui; Pêgas, António; Raposo, Daniela Marques; Sardo, AnaThe unprecedented rise in life expectancy observed in the last decades is leading to a global increase in the ageing population, and age-associated diseases became an increasing societal, economic, and medical burden. This has boosted major efforts in the scientific and medical research communities to develop and improve therapies to delay ageing and age-associated functional decline and diseases, and to expand health span. The establishment of induced pluripotent stem cells (iPSCs) by reprogramming human somatic cells has revolutionised the modelling and understanding of human diseases. iPSCs have a major advantage relative to other human pluripotent stem cells as their obtention does not require the destruction of embryos like embryonic stem cells do, and do not have a limited proliferation or differentiation potential as adult stem cells. Besides, iPSCs can be generated from somatic cells from healthy individuals or patients, which makes iPSC technology a promising approach to model and decipher the mechanisms underlying the ageing process and age-associated diseases, study drug effects, and develop new therapeutic approaches. This review discusses the advances made in the last decade using iPSC technology to study the most common age-associated diseases, including age-related macular degeneration (AMD), neurodegenerative and cardiovascular diseases, brain stroke, cancer, diabetes, and osteoarthritis.
- Matrisomal components involved in regenerative wound healing in axolotl and acomys: implications for biomaterial developmentPublication . Avila-Martinez, Nancy; Gansevoort, Merel; Verbakel, Juul; Jayaprakash, Haarshaadri; Araujo, Ines; Vitorino, Marta; Tiscornia, Gustavo; van Kuppevelt, Toin H.; Daamen, Willeke F.Achieving regeneration in humans has been a long-standing goal of many researchers. Whereas amphibians like the axolotl (Ambystoma mexicanum) are capable of regenerating whole organs and even limbs, most mammals heal their wounds via fibrotic scarring. Recently, the African spiny mouse (Acomys sp.) has been shown to be injury resistant and capable of regenerating several tissue types. A major focal point of research with Acomys has been the identification of drivers of regeneration. In this search, the matrisome components related to the extracellular matrix (ECM) are often overlooked. In this review, we compare Acomys and axolotl skin wound healing and blastema-mediated regeneration by examining their wound healing responses and comparing the expression pattern of matrisome genes, including glycosaminoglycan (GAG) related genes. The goal of this review is to identify matrisome genes that are upregulated during regeneration and could be potential candidates for inclusion in pro-regenerative biomaterials. Research papers describing transcriptomic or proteomic coverage of either skin regeneration or blastema formation in Acomys and axolotl were selected. Matrisome and GAG related genes were extracted from each dataset and the resulting lists of genes were compared. In our analysis, we found several genes that were consistently upregulated, suggesting possible involvement in regenerative processes. Most of the components have been implicated in regulation of cell behavior, extracellular matrix remodeling and wound healing. Incorporation of such pro-regenerative factors into biomaterials may help to shift pro-fibrotic processes to regenerative responses in treated wounds.
- Oxidative stress and aging: synergies for age related diseasesPublication . Filipa Santos, Daniela; Simao, Sonia; Nóbrega, Clévio; Bragança, José; Castelo-Branco, Pedro; Pombinho de Araújo, Inês Maria; ALFA Score ConsortiumAging is characterized by a progressive decline in physiological function and underlies several disabilities, including the increased sensitivity of cells and tissues to undergo pathological oxidative stress. In recent years, efforts have been made to better understand the relationship between age and oxidative stress and further develop therapeutic strategies to minimize the impact of both events on age-related diseases. In this work, we review the impact of the oxidant and antioxidant systems during aging and disease development and discuss the crosstalk of oxidative stress and other aging processes, with a focus on studies conducted in elderly populations.