Rapid Detection of SARS-CoV-2 Variants of Concern by Genomic Surveillance Techniques.

This chapter describes the application of genomic, transcriptomic, proteomic, and metabolomic methods in the study of SARS-CoV-2 variants of concern. We also describe the important role of machine learning tools to identify the most significant biomarker signatures and discuss the latest point-of-care devices that can be used to translate these findings to the physician's office or to bedside care. The main emphasis is placed on increasing our diagnostic capacity and predictability of disease outcomes to guide the most appropriate treatment strategies.

Antibody-Based Affinity Capture Combined with LC-MS Analysis for Identification of COVID-19 Disease Serum Biomarkers.

Blood serum or plasma proteins are potentially useful in COVID-19 research as biomarkers for risk prediction, diagnosis, stratification, and treatment monitoring. However, serum protein-based biomarker identification and validation is complicated due to the wide concentration range of these proteins, which spans more than ten orders of magnitude. Here we present a combined affinity purification-liquid chromatography mass spectrometry approach which allows identification and quantitation of the most abundant serum proteins along with the nonspecifically bound and interaction proteins. This led to the reproducible identification of more than 100 proteins that were not specifically targeted by the affinity column. Many of these have already been implicated in COVID-19 disease.

Liquid Chromatography-Mass Spectrometry Analysis of Peripheral Blood Mononuclear Cells from SARS-CoV-2 Infected Patients.

COVID-19 disease is caused by infection with the SARS-CoV-2 virus and is associated with a cytokine storm effect in some patients. This can lead to decreased ability of the host to cope with the infection and result in severe disease outcomes. Here, we present a protocol for isolation of peripheral blood mononuclear cells (PBMCs) from COVID-19 patients followed by liquid chromatography-mass spectrometry (LC-MS) profiling to identify the affected molecules and molecular pathways. It is hoped that this will lead to the identification of potential biomarkers for monitoring the disease as well as treatment responses. This approach could also be used in the study of other respiratory viruses.

COVID-19 Detection Using the NHS Lateral Flow Test Kit.

Approximately one in three people infected with the SARS-CoV-2 virus have mild symptoms or are asymptomatic. However, these individuals can still spread the virus. Regular self-testing can help to detect these individuals and thereby slow the spread and protect the more vulnerable members of society. Here, we present a protocol for use of the COVID-19 rapid antigen test which was made freely available to residents of the United Kingdom in April of this year. This using the lateral flow technique for detection of antigens and is amenable to multiplexing.

Point-of-Care Device for Assessment of Blood Coagulation Status in COVID-19 Patients.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent driving the current COVID-19 pandemic. The acute respiratory distress that occurs in some severe COVID-19 cases has been linked with hypercoagulation or thrombotic events as well as a worse prognosis and increased risk of death. Thus, point-of-care devices that can be used for early detection of coagulation abnormalities would assist in COVID-19 management. This chapter describes the use of the Roche Diagnostics CoaguChek® XS test kit for potential use in COVID-19 personalized medicine approaches.

Proteomic Analysis of Rat Hippocampus for Studies of Cognition and Memory Loss with Aging.

This chapter describes a protocol for proteomic profiling of the rat hippocampal proteome using a combination of liquid chromatography tandem mass spectrometry (LC-MS/MS) and data analysis to determine the cellular location of the identified proteins. In the example given, many of these proteins were localized in the plasma membrane and nucleus. These could be of interest in further studies of neurological and neurodegenerative disorders linked with hippocampal dysfunction, such as aging, major depression, and Alzheimer's disease.

Characterization of the db/db Mouse Model of Type 2 Diabetes.

This chapter presents characterization of the db/db mouse model of type 2 diabetes. The protocol describes generation of the mice along with measurements of physical characteristics such as body weight and body composition, and behaviours including food and water consumption. Such a procedure can also be used for omics-based analyses of brain tissues to investigate the potential use of db/db mouse model in neuropsychiatric research. In addition, we present measurement of blood glucose and plasma insulin levels which tend to be elevated in this model and can modulated post treatment with anti-diabetic agents. All of these characteristics can be used for studying the pathophysiology and for monitoring response to potential therapeutics.

Blood Sampling and Preparation Procedures for Proteomic Biomarker Studies of Psychiatric Disorders.

A major challenge in proteomic biomarker discovery and validation for psychiatric diseases is the inherent biological complexity underlying these conditions. There are also many technical issues which hinder this process such as the lack of standardization in sampling, processing and storage of bio-samples in preclinical and clinical settings. This chapter describes a reproducible procedure for sampling blood serum and plasma that is specifically designed for maximizing data quality output in two-dimensional gel electrophoresis, multiplex immunoassay and mass spectrometry profiling studies.

Development of an Assay for Measuring Proprotein-Conversion Activity on a Multiplex Magnetic Bead-Based Array Platform.

This chapter describes a protocol for measuring prolyl oligopeptidase (POP) activity using a biotinylated peptide substrate coupled to magnetic microspheres. The complex is incubated in the presence of a pituitary extract and activity can be detected by loss of the biotin label. The assay can be multiplexed for measuring multiple proprotein-cleaving enzymes simultaneously and can be used in analyses of neuropsychiatric diseases in which proteolytic cleavage of biologically active peptides is known to play a role.

Blood Bio-Sampling Procedures for Multiplex Biomarkers Studies.

A major challenge in single or panel of biomarker discovery and validation is the inherent biological complexity underlying disease heterogeneity and inconsistent responses to treatment. Moreover, the lack of standardization in the sampling, processing, and storage of biological fluids such as plasma and serum disrupts the discovery and validation of blood-based biomarker tests in preclinical and clinical settings. This chapter presents a reproducible sample collection and handling procedure that aims to enhance analyte stability and ensure compatibility with the corresponding multiplex biomarker profiling platforms. The importance of defining bio-sample acquisition and processing, study design, and profiling platform guidelines for blood-based biomarker measurements is paramount for the success of personalized healthcare strategy and development of companion diagnostics.

Testes sanguíneos de biomarcadores para diagnóstico e tratamento de desordens mentais: foco em esquizofrenia / Biomarker blood tests for diagnosis and management of mental disorders: focus on schizophrenia

A descoberta e a aplicação clínica de biomarcadores para desordens mentais são confrontadas com muitos desafios. Em geral, os atuais métodos de descoberta e validação de biomarcadores não produziram os resultados que foram inicialmente aguardados depois da finalização do Projeto Genoma Humano. Isso se deve principalmente à falta de processos padronizados conectando a descoberta de marcadores com tecnologias para a validação e a tradução para uma plataforma que ofereça precisão e fácil uso em clínica. Como consequência, a maior parte dos psiquiatras e praticantes em geral são relutantes em aceitar que testes de biomarcadores pode suplementar ou substituir os métodos de diagnóstico utilizados baseados em entrevista. Apesar disso, agências regulatórias concordam agora que melhoras nos correntes métodos são essenciais. Além disso, essas agências estipularam que biomarcadores são importantes para o desenvolvimento de futuras drogas e iniciaram esforços no sentido de modernizar métodos e técnicas para suportar esses esforços. Aqui revisamos os desafios encontrados por essa tentativa do ponto de vista de psiquiatras, praticantes em geral, agências reguladoras e cientistas de biomarcadores. Também descrevemos o desenvolvimento de um novo teste sanguíneo molecular para esquizofrenia como um primeiro passo a esse objetivo

The discovery and clinical application of biomarkers for mental disorders is faced with many challenges. In general, the current methods for discovery and validation of biomarkers have not produced the results which were first anticipated after completion of the human genome project. This is mostly due to the lack of a standardized pipeline connecting marker discovery with technologies for validation and translation to a platform that offers accuracy and ease of use in a clinical setting. As a consequence, most psychiatrists and general practitioners are still reluctant to accept that biomarker tests can supplement or replace the long standing interview-based methods for diagnosis. Despite this, the regulatory agencies now agree that improvements over the current methods are essential. Furthermore, these agencies stipulate that biomarkers are important for future drug development and have initiated efforts to modernize methods and techniques to support these efforts. Here, we review the challenges faced by this endeavour from the point of view of psychiatrists, general practitioners, the regulatory agencies and biomarker scientists. We also describe the development of a novel molecular blood-test for schizophrenia as a first promising step towards achieving this goal

Os efeitos do estresse na função do eixo hipotalâmico-pituitário-adrenal em indivíduos com esquizofrenia / The effects of stress on hypothalamic-pituitary-adrenal (HPA) axis function in subjects with schizophrenia

Nas últimas décadas, têm surgido evidências sugerindo que a patogênese de desordens psiquiátricas, tais como a esquizofrenia, pode envolver perturbações no eixo hipotalâmico-pituitário-adrenal (HPA). Variações na manifestação desses efeitos poderiam estar relacionadas a diferenças em sintomas clínicos entre os indivíduos afetados, assim como a diferenças na resposta ao tratamento. Tais efeitos podem também ser originados de complexas interações entre genes e fatores ambientais. Aqui, revisamos os efeitos do estresse maternal em anormalidades na regulação do eixo HPA e desenvolvimento de desordens psiquiátricas, incluindo a esquizofrenia. Estudos nessa área podem gerar o aumento do nosso entendimento da natureza multidimensional da esquizofrenia. Posterior pesquisa nesse campo poderia, em última instância, levar ao desenvolvimento de melhores diagnósticos e novas abordagens terapêuticas para essa debilitante condição psiquiátrica

Over the last few decades, evidence has been emerging that the pathogenesis of psychiatric disorders such as schizophrenia can involve perturbations of the hypothalamic-pituitary-adrenal (HPA) axis. Variations in the manifestation of these effects could be related to the differences in clinical symptoms between affected individuals as well as to differences in treatment response. Such effects can also arise from the complex interaction between genes and environmental factors. Here, we review the effects of maternal stress on abnormalities in HPA axis regulation and the development of psychiatric disorders including schizophrenia. Studies in this area may prove critical for increasing our understanding of the multi-dimensional nature of schizophrenia. Further research in this area could ultimately lead to the development of improved diagnostics and novel therapeutic approaches for treating this debilitating psychiatric condition

Explorando o componente inflamatório da esquizofrenia / Targeting the inflammatory component of schizophrenia

A esquizofrenia é uma doença heterogênea caracterizada por um conjunto de manifestações clínicas. Um grande número de estudos ao longo dos últimos 20 anos apontou para anormalidades no sistema imune em pacientes que sofrem dessa condição. Em adição, tem sido mostrado que a psicose e a disfunção cognitiva associadas com a esquizofrenia estão ligadas a doenças autoimunes. Aqui, revisamos a evidência que sugere que um status pró-inflamatório do sistema imune induz sintomas psicopatológicos e pode estar envolvido na fisiopatologia dessa principal doença mental. Também propomos que futuros estudos pré-clínicos e clínicos deveriam levar em conta tais causas predefinidas e o status do componente inflamatório. Estratificação de pacientes e estratégias de medicina personalizadas baseadas no direcionamento ao componente inflamatório da doença poderiam ajudar na redução de sintomas e da progressão da doença. Por fim, isso poderia levar a novos conceitos na identificação de alvos moleculares em esquizofrenia e estratégias de descoberta de drogas

Schizophrenia is a heterogeneous disease characterised by an array of clinical manifestations. A large number of studies over the last 20 years have pointed towards immune system abnormalities in patients suffering from this condition. In addition, the psychosis and cognitive dysfunction associated with schizophrenia have been shown to be linked with autoimmune diseases. Here, we review the evidence, which suggests that a pro-inflammatory status of the immune system induces psychopathologic symptoms and may be involved in the pathophysiology of this major mental illness. We also propose that future preclinical and clinical studies should take such pre-defined causes and the dynamic status of the inflammatory component into account. Patient stratification and personalised medicine strategies based on targeting the inflammatory component of the disease could help in alleviation of symptoms and slowing disease progression. Ultimately, this could also lead to novel concepts in schizophrenia target/molecular identification and drug discovery strategies