RESUMEN
STUDY DESIGN: This is a narrative review of the literature on neurochemical biomarkers in spinal cord injury (SCI). OBJECTIVES: The objective was to summarize the literature on neurochemical biomarkers in SCI and describe their use in facilitating clinical trials for SCI. Clinical trials in spinal cord injury (SCI) have been notoriously difficult to conduct, as exemplified by the paucity of definitive prospective randomized trials that have been completed, to date. This is related to the relatively low incidence and the complexity and heterogeneity of the human SCI condition. Given the increasing number of promising approaches that are emerging from the laboratory which are vying for clinical evaluation, novel strategies to help facilitate clinical trials are needed. METHODS: A literature review was conducted, with a focus on neurochemical biomarkers that have been described in human neurotrauma. RESULTS: We describe advances in our understanding of neurochemical biomarkers as they pertain to human SCI. The application of biomarkers from serum and cerebrospinal fluid (CSF) has been led by efforts in the human traumatic brain injury (TBI) literature. A number of promising biomarkers have been described in human SCI whereby they may assist in stratifying injury severity and predicting outcome. CONCLUSIONS: Several time-specific biomarkers have been described for acute SCI and for chronic SCI. These appear promising for stratifying injury severity and potentially predicting outcome. The subsequent application within a clinical trial will help to demonstrate their utility in facilitating the study of novel approaches for SCI.
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Biomarcadores/sangre , Biomarcadores/líquido cefalorraquídeo , Traumatismos de la Médula Espinal/líquido cefalorraquídeo , Humanos , Pronóstico , Recuperación de la Función , Traumatismos de la Médula Espinal/sangreRESUMEN
Molecular markers associated with CNS injury are of diagnostic interest. Mechanical trauma generates cellular deformation associated with membrane permeability with unknown molecular consequences. We used an in vitro model of stretch-injury and proteomic analyses to determine protein changes in murine astrocytes and their surrounding fluids. Abrupt pressure-pulse stretching resulted in the rapid release of 59 astrocytic proteins with profiles reflecting cell injury and cell death, i.e., mechanoporation and cell lysis. This acute trauma-release proteome was overrepresented with metabolic proteins compared with the uninjured cellular proteome, bearing relevance for post-traumatic metabolic depression. Astrocyte-specific deletion of signal transducer and activator of transcription 3 (STAT3-CKO) resulted in reduced stretch-injury tolerance, elevated necrosis and increased protein release. Consistent with more lysed cells, more protein complexes, nuclear and transport proteins were released from STAT3-CKO versus nontransgenic astrocytes. STAT3-CKO astrocytes had reduced basal expression of GFAP, lactate dehydrogenase B (LDHB), aldolase C (ALDOC), and astrocytic phosphoprotein 15 (PEA15), and elevated levels of tropomyosin (TPM4) and α actinin 4 (ACTN4). Stretching caused STAT3-dependent cellular depletion of PEA15 and GFAP, and its filament disassembly in subpopulations of injured astrocytes. PEA15 and ALDOC signals were low in injured astrocytes acutely after mouse spinal cord crush injury and were robustly expressed in reactive astrocytes 1 day postinjury. In contrast, α crystallin (CRYAB) was present in acutely injured astrocytes, and absent from uninjured and reactive astrocytes, demonstrating novel marker differences among postinjury astrocytes. These findings reveal a proteomic signature of traumatically-injured astrocytes reflecting STAT3-dependent cellular survival with potential diagnostic value.
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Astrocitos/metabolismo , Factor de Transcripción STAT3/metabolismo , Traumatismos de la Médula Espinal/patología , Transcriptoma/genética , Animales , Apolipoproteínas E/metabolismo , Células Cultivadas , Corteza Cerebral/citología , Modelos Animales de Enfermedad , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Espectrometría de Masas , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fragmentos de Péptidos/metabolismo , Proteómica , Factor de Transcripción STAT3/genética , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Estrés MecánicoRESUMEN
BACKGROUND: Neurotrauma or injuries to the central nervous system (CNS) are a serious public health problem worldwide. Approximately 75% of all traumatic brain injuries (TBIs) are concussions or other mild TBI (mTBI) forms. Evaluation of concussion injury today is limited to an assessment of behavioral symptoms, often with delay and subject to motivation. Hence, there is an urgent need for an accurate chemical measure in biofluids to serve as a diagnostic tool for invisible brain wounds, to monitor severe patient trajectories, and to predict survival chances. Although a number of neurotrauma marker candidates have been reported, the broad spectrum of TBI limits the significance of small cohort studies. Specificity and sensitivity issues compound the development of a conclusive diagnostic assay, especially for concussion patients. Thus, the neurotrauma field currently has no diagnostic biofluid test in clinical use. CONTENT: We discuss the challenges of discovering new and validating identified neurotrauma marker candidates using proteomics-based strategies, including targeting, selection strategies and the application of mass spectrometry (MS) technologies and their potential impact to the neurotrauma field. SUMMARY: Many studies use TBI marker candidates based on literature reports, yet progress in genomics and proteomics have started to provide neurotrauma protein profiles. Choosing meaningful marker candidates from such 'long lists' is still pending, as only few can be taken through the process of preclinical verification and large scale translational validation. Quantitative mass spectrometry targeting specific molecules rather than random sampling of the whole proteome, e.g., multiple reaction monitoring (MRM), offers an efficient and effective means to multiplex the measurement of several candidates in patient samples, thereby omitting the need for antibodies prior to clinical assay design. Sample preparation challenges specific to TBI are addressed. A tailored selection strategy combined with a multiplex screening approach is helping to arrive at diagnostically suitable candidates for clinical assay development. A surrogate marker test will be instrumental for critical decisions of TBI patient care and protection of concussion victims from repeated exposures that could result in lasting neurological deficits.
RESUMEN
Multi-modal biomarkers (e.g., imaging, blood-based, physiological) of unique traumatic brain injury (TBI) endophenotypes are necessary to guide the development of personalized and targeted therapies for TBI. Optimal biomarkers will be specific, sensitive, rapidly and easily accessed, minimally invasive, cost effective, and bidirectionally translatable for clinical and research use. For both uses, understanding how TBI biomarkers change over time is critical to reliably identify appropriate time windows for an intervention as the injury evolves. Biomarkers that enable researchers and clinicians to identify cellular injury and monitor clinical improvement, inflection, arrest, or deterioration in a patient's clinical trajectory are needed for precision healthcare. Prognostic biomarkers that reliably predict outcomes and recovery windows to assess neurodegenerative change and guide decisions for return to play or duty are also important. TBI biomarkers that fill these needs will transform clinical practice and could reduce the patient's risk for long-term symptoms and lasting deficits. This article summarizes biomarkers currently under investigation and outlines necessary steps to achieve short- and long-term goals, including how biomarkers can advance TBI treatment and improve care for patients with TBI.
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Lesiones Traumáticas del Encéfalo , Biomarcadores , Lesiones Traumáticas del Encéfalo/diagnóstico , Lesiones Traumáticas del Encéfalo/genética , Lesiones Traumáticas del Encéfalo/terapia , Humanos , PronósticoRESUMEN
Traumatic brain injury (TBI) is an expanding public health epidemic with pathophysiology that is difficult to diagnose and thus treat. TBI biomarkers should assess patients across severities and reveal pathophysiology, but currently, their kinetics and specificity are unclear. No single ideal TBI biomarker exists. We identified new candidates from a TBI CSF proteome by selecting trauma-released, astrocyte-enriched proteins including aldolase C (ALDOC), its 38kD breakdown product (BDP), brain lipid binding protein (BLBP), astrocytic phosphoprotein (PEA15), glutamine synthetase (GS) and new 18-25kD-GFAP-BDPs. Their levels increased over four orders of magnitude in severe TBI CSF. First post-injury week, ALDOC levels were markedly high and stable. Short-lived BLBP and PEA15 related to injury progression. ALDOC, BLBP and PEA15 appeared hyper-acutely and were similarly robust in severe and mild TBI blood; 25kD-GFAP-BDP appeared overnight after TBI and was rarely present after mild TBI. Using a human culture trauma model, we investigated biomarker kinetics. Wounded (mechanoporated) astrocytes released ALDOC, BLBP and PEA15 acutely. Delayed cell death corresponded with GFAP release and proteolysis into small GFAP-BDPs. Associating biomarkers with cellular injury stages produced astroglial injury-defined (AID) biomarkers that facilitate TBI assessment, as neurological deficits are rooted not only in death of CNS cells, but also in their functional compromise.
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Astrocitos/patología , Biomarcadores/análisis , Lesiones Traumáticas del Encéfalo/líquido cefalorraquídeo , Proteínas Reguladoras de la Apoptosis , Astrocitos/química , Conmoción Encefálica , Lesiones Traumáticas del Encéfalo/diagnóstico , Células Cultivadas , Proteína de Unión a los Ácidos Grasos 7/sangre , Fructosa-Bifosfato Aldolasa/sangre , Humanos , Péptidos y Proteínas de Señalización Intracelular/sangre , Cinética , Fosfoproteínas/sangre , Proteoma/análisis , Proteínas Supresoras de Tumor/sangreRESUMEN
Protocols are presented describing a unique in vitro injury model and how to culture and mature mouse, rat, and human astrocytes for its use. This injury model produces widespread injury and astrocyte reactivity that enable quantitative measurements of morphological, biochemical, and functional changes in rodent and human reactive astrocytes. To investigate structural and molecular mechanisms of reactivity in vitro, cultured astrocytes need to be purified and then in vitro "matured" to reach a highly differentiated state. This is achieved by culturing astrocytes on deformable collagen-coated membranes in the presence of adult-derived horse serum (HS), followed by its stepwise withdrawal. These in vitro matured, process-bearing, quiescent astrocytes are then subjected to mechanical stretch injury by an abrupt pressure pulse from a pressure control device that briefly deforms the culture well bottom. This inflicts a measured reproducible, widespread strain that induces reactivity and injury in rodent and human astrocytes. Cross-species comparisons are possible because mouse, rat, and human astrocytes are grown using essentially the same in vitro treatment regimen. Human astrocytes from fetal cerebral cortex are compared to those derived from cortical biopsies of epilepsy patients (ages 1-12 years old), with regard to growth, purity, and differentiation. This opens a unique opportunity for future studies on glial biology, maturation, and pathology of human astrocytes. Prototypical astrocyte proteins including GFAP, S100, aquaporin4, glutamate transporters, and tenascin are expressed in mouse, rat, and human in vitro matured astrocyte. Upon pressure-stretching, rodent and human astrocytes undergo dynamic morphological, gene expression, and protein changes that are characteristic for trauma-induced reactive astrogliosis.