Cone-theoretic generalization of total positivity

This paper is devoted to the generalization of the theory of total positivity. We say that a linear operator A in R^n is generalized totally positive (GTP), if its jth exterior power preserves a proper cone K_j in the corresponding space for every j = 1, ..., n. We also define generalized strictly totally positive (GSTP) operators. We prove that the spectrum of a GSTP operator is positive and simple, moreover, its eigenvectors are localized in special sets. The existence of invariant cones of finite ranks is shown under some additional conditions. Some new insights and alternative proofs of the well-known results of Gantmacher and Krein describing the properties of TP and STP matrices are presented

Glutathione-S-transferase P promotes glycolysis in asthma in association with oxidation of pyruvate kinase M2

Background: Interleukin-1-dependent increases in glycolysis promote allergic airways disease in mice and disruption of pyruvate kinase M2 (PKM2) activity is critical herein. Glutathione-S-transferase P (GSTP) has been implicated in asthma pathogenesis and regulates the oxidation state of proteins via S-glutathionylation. We addressed whether GSTP-dependent S-glutathionylation promotes allergic airways disease by promoting glycolytic reprogramming and whether it involves the disruption of PKM2. Methods: We used house dust mite (HDM) or interleukin-1β in C57BL6/NJ WT or mice that lack GSTP. Airway basal cells were stimulated with interleukin-1β and the selective GSTP inhibitor, TLK199. GSTP and PKM2 were evaluated in sputum samples of asthmatics and healthy controls and incorporated analysis of the U-BIOPRED severe asthma cohort database. Results: Ablation of Gstp decreased total S-glutathionylation and attenuated HDM-induced allergic airways disease and interleukin-1β-mediated inflammation. Gstp deletion or inhibition by TLK199 decreased the interleukin-1β-stimulated secretion of pro-inflammatory mediators and lactate by epithelial cells. 13C-glucose metabolomics showed decreased glycolysis flux at the pyruvate kinase step in response to TLK199. GSTP and PKM2 levels were increased in BAL of HDM-exposed mice as well as in sputum of asthmatics compared to controls. Sputum proteomics and transcriptomics revealed strong correlations between GSTP, PKM2, and the glycolysis pathway in asthma. Conclusions: GSTP contributes to the pathogenesis of allergic airways disease in association with enhanced glycolysis and oxidative disruption of PKM2. Our findings also suggest a PKM2-GSTP-glycolysis signature in asthma that is associated with severe disease

Metabolic reprogramming identifies the most aggressive lesions at early phases of hepatic carcinogenesis

Metabolic changes are associated with cancer, but whether they are just bystander effects of deregulated oncogenic signaling pathways or characterize early phases of tumorigenesis remains unclear. Here we show in a rat model of hepatocarcinogenesis that early preneoplastic foci and nodules that progress towards hepatocellular carcinoma (HCC) are characterized both by inhibition of oxidative phosphorylation (OXPHOS) and by enhanced glucose utilization to fuel the pentose phosphate pathway (PPP). These changes respectively require increased expression of the mitochondrial chaperone TRAP1 and of the transcription factor NRF2 that induces the expression of the rate-limiting PPP enzyme glucose-6-phosphate dehydrogenase (G6PD), following miR-1 inhibition. Such metabolic rewiring exclusively identifies a subset of aggressive cytokeratin-19 positive preneoplastic hepatocytes and not slowly growing lesions. No such metabolic changes were observed during non-neoplastic liver regeneration occurring after two/third partial hepatectomy. TRAP1 silencing inhibited the colony forming ability of HCC cells while NRF2 silencing decreased G6PD expression and concomitantly increased miR-1; conversely, transfection with miR-1 mimic abolished G6PD expression. Finally, in human HCC patients increased G6PD expression levels correlates with grading, metastasis and poor prognosis. Our results demonstrate that the metabolic deregulation orchestrated by TRAP1 and NRF2 is an early event restricted to the more aggressive preneoplastic lesions

Phenotypic and functional analyses show stem cell-derived hepatocyte-like cells better mimic fetal rather than adult hepatocytes

Background & Aims: Hepatocyte-like cells (HLCs), differentiated from pluripotent stem cells by the use of soluble factors, can model human liver function and toxicity. However, at present HLC maturity and whether any deficit represents a true fetal state or aberrant differentiation is unclear and compounded by comparison to potentially deteriorated adult hepatocytes. Therefore, we generated HLCs from multiple lineages, using two different protocols, for direct comparison with fresh fetal and adult hepatocytes. Methods: Protocols were developed for robust differentiation. Multiple transcript, protein and functional analyses compared HLCs to fresh human fetal and adult hepatocytes. Results: HLCs were comparable to those of other laboratories by multiple parameters. Transcriptional changes during differentiation mimicked human embryogenesis and showed more similarity to pericentral than periportal hepatocytes. Unbiased proteomics demonstrated greater proximity to liver than 30 other human organs or tissues. However, by comparison to fresh material, HLC maturity was proven by transcript, protein and function to be fetal-like and short of the adult phenotype. The expression of 81% phase 1 enzymes in HLCs was significantly upregulated and half were statistically not different from fetal hepatocytes. HLCs secreted albumin and metabolized testosterone (CYP3A) and dextrorphan (CYP2D6) like fetal hepatocytes. In seven bespoke tests, devised by principal components analysis to distinguish fetal from adult hepatocytes, HLCs from two different source laboratories consistently demonstrated fetal characteristics. Conclusions: HLCs from different sources are broadly comparable with unbiased proteomic evidence for faithful differentiation down the liver lineage. This current phenotype mimics human fetal rather than adult hepatocytes

EFEK POLIMORFISME GENA GSTP-1 TERHADAP AKTIVITAS GLUTATION S-TRANSFERASE (GST) PADA INDIVIDU TERPAPAR LOGAM BERAT TIMBAL (Effect of GSTP-1 Gene Polymorphismson Glutation S- Transferase (GST) Activity in Heavy Metals Lead-Exposed Individual)

ABSTRAKGena GSTP-1 merupakan penghasil enzim glutation S- transferase (GST), yang berfungsi dalam proses detoksifikasi senyawa toksik di hati. Faktor keberadaan polimorfisme gena GSTP-1 akan menyebabkan penurunan ekspresi GST, sehingga proses detoksifikasi terhadap senyawa toksik akan terhambat. Kerentanan terhadap paparan senyawa toksik pada manusia akan meningkat apabila dijumpai polimorfisme gena. Salah satu senyawa toksik yang dapat menghambat aktivitas GST adalah timbal (Pb), terutama dalam bentuk tetra ethyl lead (TEL). Tujuan penelitian adalah untuk mengetahui pengaruh polimorfisme gena GSTP-1 terhadap aktivitas GST pada individu terpapar Pb, yang diwakili pekerja bengkel mobil. Faktor keberadaan polimorfisme gena individu ditentukan dengan metode PCR-RFLP dan enzim restriksi BsmA1. Parameter yang diukur adalah kadar Pb dan aktivitas GST. Analisis molekuler gena GSTP-1 dilakukan secara deskriptif. Data kadar Pb dan aktivitas GST dianalisis dengan uji t independent. Hasil analisis gena GSTP-1 dari 40 orang subyek kasus setelah dilakukan digesti dengan enzim BsmA1, ditemukan sebanyak 10 orang individu dengan polimorfisme Ile105Val gena GSTP 1 atau sekitar 25% dengan genotip Ile-Val, sedangkan 30 orang atau 75% ditemukan tanpa polimorfisme dengan genotip Ile-Ile. Pita DNA individu dengan polimorfisme terpotong menjadi 3 fragmen sepanjang 176, 91 dan 85 pp (mutan heterozygot), sedangkan tanpa polimorfisme terletak pada 176 bp. Subyek kasus dengan polimorfisme gena GSTP-1 memiliki kadar Pb lebih tinggi dan aktivitas GST lebih rendah dibandingkan individu non polimorfisme. Telah terbukti bahwa polimorfisme gena GSTP-1 menyebabkan penurunan ekspresi enzim GST. Pada individu terpapar Pb dengan polimorfisme gena GSTP-1 memiliki aktivitas GST lebih rendah dibandingkan individu tanpa polimorfisme.ABSTRACTGSTP-1 gene regulates the expression of gluthation S-transferase enzyme, which role in detoxification of toxicant on liver. If the polymorphisms gene is found in individual, the production of GST is decreased and the enzyme failed to eliminate toxicants. Lead is one of toxic agents that could inhibite GST activity especially tetra ethyl lead (TEL). The susceptibility to lead exposure will increase if the polymorphisms gene is found in population. The objective of this studies were to know the effect of gene GSTP-1 polymorphisms to GST activity on lead-exposed individual ie. autorepair workers. The genotype individu were analyzed by Polymerase Chain Reaction (PCR)-Restriction Fragment Length Polymorphisms with BsmA1 restriction enzyme followed by descriptived analyzed. Parameter recorded were blood lead and GST activity and data were analyzed by independent t-test. These result showed that 25% of 40 individual cases subject were detected by enzyme BsmA1 as polymorphisms individual of GSTP-1 gene, with Ile105Val genotype. As many as 75% were detected as non polymorphisms with Ile-Ile genotype. Three fragment DNA of polymorphisms individual of GSTP-1 is located on 176, 91 and 85 bp (heterozygote mutant) but non polymorphisms individual is only located on 176 bp. The Pb level of individual with polymorphisms GSTP-1 gene is higher than non polymorphisms individual but their GST activity was lower than non polymorphisms individual. It could be concluded that polymorphisms GSTP-1 gene could decrease the expression gene of GST enzyme and intoxication of lead-exposured could increased the decreasing of this activity

Role of glutathione s-transferase pi in neuronal protection under oxidative stress and proteasome inhibition relevance to Parkinson’s disease

Tese de doutoramento, Farmácia (Bioquímica), Universidade de Lisboa, Faculdade de Farmácia, 2012Although the molecular mechanisms underlying DA neuronal death in Parkinson’s disease (PD) are still not completely understood, solid evidence has accumulated implicating mitochondrial dysfunction, oxidative stress and failure of proteolytic pathways in the pathogenesis of the disease. Glutathione S-Transferase pi (GSTP), whose expression is regulated by the nuclear factor-erythroid 2-related factor 2 (Nrf2), has been shown to protect cells from reactive oxygen species by modulating S-glutathionylation of proteins, following oxidative and nitrosative stress, altering the levels of glutathione, and also by acting as a ligand-binding protein controlling the catalytic activity of kinases involved in cell-signaling pathways. Experimental models of PD based on the use of the selective dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) are widely used as they recapitulate the neurological features of the disease, triggering a cascade of deleterious events that culminate with DA neuronal demise. The main objective of the present work was to characterize the putative neuroprotective role of GSTP against MPTP-induced oxidative stress and to contribute to the elucidation of the role of proteasome inhibition in the MPTP Parkinson’s disease mouse model. Moreover with this work we aimed to further investigate the molecular mechanisms underlying MPTP-induced DA neuronal death. In the first part of this work we have demonstrated in vivo that GSTP is an endogenous regulator of the activity of c-Jun N-terminal kinase (JNK). We show that in mice brain, GSTP is able to bind JNK, through direct protein-protein interaction, inhibiting its activation, and consequently preventing its downstream effects on triggering cell death pathways. Moreover, we showed that GSTP knockout mice display increased susceptibility to MPTP neurotoxicity. We have further characterized GSTP neuroprotective function by demonstrating for the first time that, in vivo GSTP potentiates Kelch ECH associating protein 1 (Keap1) Abstract xxiv S-glutathionylation following MPTP administration, in mice brain. As Keap1 is the major endogenous regulator of Nrf2, Keap1 S-glutathionylation results in the subsequent Nrf2 pathway activation, and increased expression of GSTP, in a positive feedback regulatory loop. Finally, as the ubiquitin-proteasome system (UPS) is also a critical player in several processes involved in the regulation of neuronal survival vs neuronal death pathways, and specifically in the regulation of the Nrf2-elicited antioxidant response, we investigated the modifications in UPS function, upon MPTP-induced oxidative stress and proteasome inhibition, in wild-type and GSTP knockout mice brain. We concluded that different components of the UPS have different susceptibilities to oxidative stress and, importantly GSTP knockout mice display increased susceptibility to UPS damage and inactivation upon MPTP-induced oxidative stress. In conclusion, our results provided new insights into the molecular mechanisms involved in DA neuronal loss and we identified novel mechanisms involved in GSTP-elicited neuronal protection. Overall our results demonstrate important GSTP roles, other than detoxification, namely in the regulation of kinase signaling pathways and consequently in the modulation of cell death cascades, and in the potentiating of S-glutathionylation, thus enhancing antioxidant responses. Therefore the regulation of GSTP expression constitutes a promising therapeutic target in the development of novel therapeutic strategies for the treatment of neurodegenerative diseases and other diseases in which oxidative stress is an important factor of progression.A doença de Parkinson (PD) é uma doença neurodegenerativa caracterizada pela degenerescência progressiva dos neurónios dopaminérgicos da substantia nigra pars compacta (SNpc) e pela presença de inclusões citoplasmáticas constituídas por agregados proteicos, denominadas corpos de Lewy. A depleção de dopamina, resultante da morte dos neurónios dopaminérgicos, origina perda dos mecanismos de neurotransmissão entre a SNpc e o estriado, onde se situam os terminais nervosos destes neurónios, e conduz à incapacidade de controlo dos movimentos voluntários. Embora se conheçam as características neuroquímicas e neuropatológicas da PD, a sua etiologia bem como os mecanismos moleculares subjacentes à morte dos neurónios dopaminérgicos não estão ainda completamente elucidados. No entanto, resultados de numerosos estudos apontam para um importante papel de factores como o stress oxidativo, a disfunção mitocondrial e a desregulação de vias de degradação proteica, na patogénese das formas familiar e esporádica da PD. O desencadeamento da disfunção do metabolismo proteico, nas formas esporádicas da PD, poderá resultar do stress oxidativo mediado por espécies reactivas de oxigénio (ROS). A isoforma pi da Glutationo S-transferase (GSTP) é um enzima de destoxificação de fase II, que para além de catalisar a reacção de adição nucleofílica do glutationo reduzido a um elevado número de substratos, participando assim na eliminação das ROS, tem sido sugerido como protector em situações de stress oxidativo devido à sua capacidade de estabelecer interacções proteína-proteína, modulando vias de sinalização celulares. Os modelos experimentais baseados na administração da neurotoxina 1-metil-4- fenil-1,2,3,6-tetrahidropiridina (MPTP) a ratinhos mimetizam diversas características neuropatológicas da PD desencadeando uma cascata de eventos causadores de danos celulares, que culmina com a degenerescência selectiva dos neurónios dopaminérgicos. O presente trabalho teve como principal objectivo contribuir para a caracterização do potencial papel neuroprotector da GSTP, em resposta ao stress induzido pelo MPTP e Resumo xxvi à inibição do proteossoma. Mais especificamente foram estudados os mecanismos moleculares subjacentes à perda dos neurónios dopaminérgicos em situações de stress oxidativo induzido pelo MPTP e investigado o papel da inibição do proteossoma neste modelo de PD. Na fase inicial deste trabalho demonstrámos que a administração de MPTP a ratinhos C57BL/6 wild-type (wt) e GSTP knockout (ko) conduz à degenerescência selectiva dos neurónios dopaminérgicos da via nigro-estriado, com perda de densidade dos terminais nervosos no estriado e diminuição do número de neurónios dopaminérgicos no cérebro médio. Os nossos resultados mostraram que a degenerescência dos neurónios dopaminérgicos ocorre mais precocemente nos ratinhos GSTP ko do que nos wt, demonstrando que os ratinhos ko são mais susceptíveis à neurotoxicidade induzida pelo MPTP. Adicionalmente, observámos que o tratamento com MPTP conduz a um aumento da fosforilação da cinase N-terminal da c-Jun (JNK), bem como da sua actividade catalítica e consequentemente da fosforilação de c-Jun. A activação da cascata de sinalização da JNK despoleta vias de morte celular, pelo que os resultados obtidos indicam que esta será uma das vias envolvidas na morte dos neurónios dopaminérgicos induzida pelo MPTP. Os mecanismos moleculares subjacentes à regulação da actividade da JNK em condições de stress celular envolvem a activação de uma cascata de cinases bem como da regulação controlada de diversos repressores endógenos da JNK. Os nossos resultados demonstram que in vivo a GSTP actua como um regulador endógeno da resposta celular ao stress induzido pelo MPTP através da inibição directa da actividade da JNK, por interacção proteína-proteína. O factor de transcrição Nrf2 é um dos principais sensores e reguladores celulares da resposta ao stress oxidativo e a sua actividade é regulada maioritariamente pela proteína Keap1, que encaminha o Nrf2 para degradação pelo proteossoma. A indução da expressão de enzimas de fase II, nomeadamente a GSTP pelo factor de transcrição Nrf2, constitui um potencial mecanismo de defesa celular contra o stress oxidativo induzido pelo MPTP. Numa segunda parte deste trabalho procurámos por um lado clarificar de que forma a inibição do proteossoma, adicionalmente ao stress induzido pelo MPTP, influenciava a regulação da actividade do Nrf2 e quais as consequências na activação da GSTP, e por outro investigar o papel da GSTP na regulação dos mecanismos moleculares activados nestas condições experimentais. Resumo xxvii Os resultados obtidos demonstraram um aumento da formação de ROS bem como um aumento da acumulação de grupos carbonilo nas proteínas, indicando um aumento da quantidade de proteínas oxidadas no cérebro de ratinhos C57BL/6 wt tratados com MPTP e/ou com o inibidor do proteossoma, MG132. Verificou-se também um aumento da reactividade dos astrócitos e da microglia. Conjuntamente estes resultados sugerem uma activação inicial de vias de stress e inflamação, nas quais parece haver um efeito de sinergia resultante da administração simultânea dos dois compostos. Um dos marcadores de estado redox celular mais reconhecidos é o nível de glutationo intracelular, bem como o seu estado de oxidação. Os nossos resultados mostraram um decréscimo de glutationo, acompanhado de um aumento de actividade da peroxidase do glutationo e um decréscimo da actividade da reductase do glutationo, indicando um ambiente celular mais oxidante. No entanto não se verificou um aumento da forma oxidada livre do glutationo nestas condições, provavelmente porque o decréscimo observado nos níveis de glutationo livre se deve à sua conjugação com proteínas, visto que observámos um aumento dos níveis de proteínas S-glutationiladas nos ratinhos wt tratados com MPTP. Mais ainda, através de ensaios de co-imunoprecipitação verificámos a S-glutationilação específica da proteína Keap1, nos ratinhos wt mas não nos GSTP ko. A S-glutationilação conduz à dissociação do complexo entre a Keap1 e o Nrf2, evitando a degradação do Nrf2 pelo proteossoma e conduzindo à sua activação. Nos ratinhos wt tratados com MPTP, observámos ainda um aumento de expressão do Nrf2, bem como da sua translocação para o núcleo, indicativa da sua activação. Ainda de acordo com estes resultados, pudémos observar um aumento dos níveis de expressão da GSTP e da heme oxigenase (HO-1), ambas reguladas pelo Nrf2, nos ratinhos wt tratados com MPTP. No entanto, nos ratinhos ko não se verificou um aumento nos níveis de expressão para a HO-1. Tendo em conta que a GSTP foi sugerida como sendo um dos catalisadores da reacção de S-glutationilação, estes resultados indicam que, em resposta ao stress induzido pelo MPTP, a GSTP catalisa a reacção de S-glutationilação da Keap1, levando à dissociação do complexo Keap1-Nrf2 e à activação do Nrf2. O Nrf2 por sua vez é translocado para o núcleo onde regula a expressão de vários genes, nos quais se incluem GSTP e HO-1. Nos ratinhos GSTP ko, a ausência de GSTP inviabiliza este mecanismo. No seu conjunto, estes resultados demonstram a existência de um mecanismo de regulação por feedback positivo, em que a GSTP ao ser activada em condições de stress, poderá por sua vez deslocar o equilíbrio da reacção de S-glutationilação no sentido da Resumo xxviii glutationilação de diversas proteínas, nomeadamente da Keap1, levando à dissociação do complexo Keap1-Nrf2 e consequentemente a uma maior activação do Nrf2. O conjunto dos nossos resultados permitiu demonstrar pela primeira vez a S-glutationilação da Keap1 in vivo, e consequente activação do Nrf2, revelando um novo mecanismo de protecção neuronal pela GSTP. A disfunção das vias de degradação de proteínas é um dos factores envolvidos na degenerescência dopaminérgica na PD e em modelos experimentais. Por um lado a inibição das vias de degradação, nomeadamente do proteossoma, poderá causar a acumulação de proteínas danificadas por stress oxidativo, e por outro lado a acumulação de proteínas oxidadas poderá levar à sobrecarga e consequente inibição do proteossoma, criando assim uma potenciação cíclica dos danos oxidativos na célula. Na última parte deste trabalho procurámos clarificar de que forma a via da ubiquitina-proteossoma é alterada pela administração de MPTP e/ou MG132, e se existem diferenças entre ratinhos wt e GSTP ko no que diz respeito a esta via. Verificámos que, no cérebro de ratinhos wt e GSTP ko, diferentes componentes da via da ubiquitina proteossoma apresentam diferentes susceptibilidades ao stress induzido pelo MPTP ou ao tratamento com MG132. Mais importante, observámos que os ratinhos ko são mais susceptíveis à disfunção da via da ubiquitina proteossoma, após tratamento com MPTP, relativamente aos wt, apresentando menor capacidade de ubiquitinação e níveis de expressão do enzima de conjugação com a ubiquitina (E1) mais baixos. Uma vez que o aumento da proteólise pode de certo modo funcionar como um mecanismo antioxidante ao degradar proteínas oxidadas ou danificadas, impedindo a sua acumulação, o facto de os ratinhos GSTP ko terem défices nesta via pode contribuir para a sua maior susceptibilidade ao stress induzido pelo MPTP. Estes resultados sugerem que a GSTP pode também ter um papel na protecção/manutenção das vias de degradação proteica em condições de stress oxidativo. Em conclusão, com a realização deste trabalho contribuímos para a elucidação dos mecanismos implicados na degenerescência dos neurónios dopaminérgicos induzida pelo MPTP e identificámos alguns dos mecanismos subjacentes à neuroprotecção conferida pela GSTP. A expressão alterada de GSTP foi já descrita como estando relacionada com diversas patologias humanas. Os resultados aqui apresentados mostram que além de um importante papel na destoxificação, in vivo a GSTP tem a capacidade de regular a actividade catalítica da JNK, afectando a modulação de vias de morte celular, bem como de catalisar a S-glutationilação de proteínas envolvidas na resposta antioxidante. Assim, a regulação da expressão da GSTP revelou ser um importante alvo para o desenvolvimento de novas estratégias terapêuticas para o tratamento de doenças neurodegenerativas e outras doenças nas quais o stress oxidativo seja um importante factor de progressão.Fundação para a Ciência e a Tecnologia (FCT, SFRH/BD/39897/2007 e projetos PPCDT/SAU-FCF/58171/2004, PTDC/SAU-MMO/57216/2004, PEst-OE/SAU/UI4013/2011) e FEDER

Similarity of percolation thresholds on the hcp and fcc lattices

Extensive Monte-Carlo simulations were performed in order to determine the precise values of the critical thresholds for site ($p^{hcp}_{c,S} = 0.199 255 5 \pm 0.000 001 0$) and bond ($p^{hcp}_{c,B} = 0.120 164 0 \pm 0.000 001 0$) percolation on the hcp lattice to compare with previous precise measuremens on the fcc lattice. Also, exact enumeration of the hcp and fcc lattices was performed and yielded generating functions and series for the zeroth, first, and second moments of both lattices. When these series and the values of $p_c$ are compared to those for the fcc lattice, it is apparent that the site percolation thresholds are different; however, the bond percolation thresholds are equal within error bars, and the series only differ slightly in the higher order terms, suggesting the actual values are very close to each other, if not identical.Comment: 10 pages, 4 figures, submitted to J. Stat. Phy

Combining H-FABP and GFAP increases the capacity to differentiate between CT-positive and CT-negative patients with mild traumatic brain injury

Mild traumatic brain injury (mTBI) patients may have trauma-induced brain lesions detectable using CT scans. However, most patients will be CT-negative. There is thus a need for an additional tool to detect patients at risk. Single blood biomarkers, such as S100B and GFAP, have been widely studied in mTBI patients, but to date, none seems to perform well enough. In many different diseases, combining several biomarkers into panels has become increasingly interesting for diagnoses and to enhance classification performance. The present study evaluated 13 proteins individually—H-FABP, MMP-1, MMP-3, MMP-9, VCAM, ICAM, SAA, CRP, GSTP, NKDA, PRDX1, DJ-1 and IL-10—for their capacity to differentiate between patients with and without a brain lesion according to CT results. The best performing proteins were then compared and combined with the S100B and GFAP proteins into a CT-scan triage panel. Patients diagnosed with mTBI, with a Glasgow Coma Scale score of 15 and one additional clinical symptom were enrolled at three different European sites. A blood sample was collected at hospital admission, and a CT scan was performed. Patients were divided into two two-centre cohorts and further dichotomised into CT-positive and CT-negative groups for statistical analysis. Single markers and panels were evaluated using Cohort 1. Four proteins—H-FABP, IL-10, S100B and GFAP—showed significantly higher levels in CT-positive patients. The best-performing biomarker was H-FABP, with a specificity of 32% (95% CI 23–40) and sensitivity reaching 100%. The best-performing two-marker panel for Cohort 1, subsequently validated in Cohort 2, was a combination of H-FABP and GFAP, enhancing specificity to 46% (95% CI 36–55). When adding IL-10 to this panel, specificity reached 52% (95% CI 43–61) with 100% sensitivity. These results showed that proteins combined into panels could be used to efficiently classify CT-positive and CT-negative mTBI patients

Role of glutathionylation in infection and inflammation

Glutathionylation, that is, the formation of mixed disulfides between protein cysteines and glutathione (GSH) cysteines, is a reversible post-translational modification catalyzed by dierent cellular oxidoreductases, by which the redox state of the cell modulates protein function. So far, most studies on the identification of glutathionylated proteins have focused on cellular proteins, including proteins involved in host response to infection, but there is a growing number of reports showing that microbial proteins also undergo glutathionylation, with modification of their characteristics and functions. In the present review, we highlight the signaling role of GSH through glutathionylation, particularly focusing on microbial (viral and bacterial) glutathionylated proteins (GSSPs) and host GSSPs involved in the immune/inflammatory response to infection; moreover, we discuss the biological role of the process in microbial infections and related host responses