On the other hand, the increase in the levels of l -lactate dehydrogenase A chain LDH-A and decrease in LDH-B have been shown to contribute to high brain lactate levels, which are predictive of aging phenotypes Finally, CRYM and thyroid hormone levels may be key factors in the development of the pipecolate pathway in the brain 92 , especially that neonatal decrease in these hormones is well-known to influence brain development and cause mental retardation and neurological damage 93 , Interestingly, our proteomics data analysis also revealed alteration in a subset of proteins that have not been previously studied or tackled in TBI.
For instance, at both 1 and 7 days post-CCI, a number of proteins were found to be upregulated including elongation factor 2, alphainhibitor 3, and gamma-enolase Table 1 A. Eukaryotic elongation factor 2 is known to play a key role in regulating the protein translational machinery and controlling ribosomal movements across the mRNA, thus affecting the survival of neurons in cases of elevated oxidative stress Alphainhibitor 3, a protein of the alpha-macroglobulin family and an acute phase reactant protein, has been previously studied and shows a role in the early stages of the inflammatory response 96 , yet it has never been studied in TBI.
In addition, gamma-enolase, also known as neuron-specific enolase, possesses neuroprotective effects on cultured neurons from embryonic rat brain On the other hand, a subset of proteins was found to be downregulated at both time points including triosephosphate isomerase, protein DJ-1, and isoform 1 of NF Table 1 B. The importance of those proteins is that they may serve as potential biomarkers and therapeutic targets for TBI, where future studies may be conducted to assess their exact function in the context of TBI.
Moreover, other proteins that were found to be uniquely altered at each time point, such as alpha-enolase, complement inhibitory factor H, and zero beta-1 globin, and have never been studied in TBI, can also be assessed for their function in TBI Table 2. In order to scrutinize the differences in protein expression at a functional level, further analysis was carried forward using PANTHER software to identify enriched pathways and biological processes altered in TBI at different time points between acute and subacute states.
This software is a unique resource that classifies genes and proteins by their functions using published scientific experimental evidence and evolutionary relationships abstracted by curators with the goal of predicting function even in the absence of direct experimental evidence.
Proteins are classified into families and subfamilies of shared function, which are then categorized using a highly controlled vocabulary ontology terms. In our study, common pathways were altered including apoptosis, inflammatory response, oxidative stress, and autophagy both at 1 and 7 days post-CCI.
Yet, it is worthwhile to state that several cellular processes encompassing neuroprotective proteins were found to be enriched in subacute TBI, including cell regeneration, neurite outgrowth, axonogenesis, and cell growth. Interestingly, some pathways were found to be altered uniquely at 1 day post-CCI, such as cell migration, caspase, mitochondrial damage, neuron toxicity, and heat shock response, while others were found to be solely altered at 7 days post-CCI, including regeneration, axon guidance, axonogenesis, cell growth, and cell differentiation Figure 5.
This work builds on previous and current recent studies from our and other labs that have showed the capabilities of neuroproteomics in identifying putative markers of TBI UCH-L1, synaptotagmin, and spectrins utilizing the same CAX separation platform as the one applied here 26 , 27 , However, this work has a number of limitations that relates to the study experimental design, methodology, and finally complying with the recent recommendation of the NIH common data elements CDEs for preclinical TBI First, our work was based on an in silico bioinformatics approach lacking validation or confirmation steps via wet lab techniques; these experiments were kept for the follow-up functional analysis study where we selected few of the identified proteins to study their dynamic alteration.
Second, our work has considered only two time points 1 vs. Third, this work assessed only cortical regions, while the emphasis now is to look at different brain regions that can depict the global injury profile rather than being biased to the cortical areas. Therefore, future studies including multiple regions such as the hippocampus and thalamus will provide a better insight of the injury profile.
This will be gained in future studies through comparing our results to different models such as repetitive mild TBI, and consequently, analyzing clinical samples may be considered. As per the recommendation of the NIH CDEs for preclinical TBI, the use of female cohort should be included to highlight the diversity in injury profile and avoid any experimental bias; however, one should bear in mind that these neuroproteomics studies have their complexities and limitations in determining the number of samples to be used, the pooling consideration, and finally the time and cost they require.
Finally, it is of interest to mention and highlight the shortcomings of this proteomics and other high throughput approaches MRI, transcriptomics, gene arrays, etc. This will often lead to the missing of big chunk of extremely valuable data from any experimental study. To illustrate this point, it is estimated that the human genome contains around 20, protein-encoding genes, while the total number of proteins in human cells is estimated to range between , and one million. To conclude, this work utilizing systems biology concept has showed that it can highlight on several differential pathways and proteins, which can be of high prognostic and diagnostic value.
Several of the identified protein hits can be translated clinically as putative markers in clinical TBI. Collectively, data obtained from this work suggest that many of the differentially identified proteins in the acute and subacute phases of TBI may serve as potential neural biomarkers and therapeutic targets for TBI. Of interest, the identification of the acute phase protein C3 seems to be a promising candidate marker that is easily measured in human TBI biofluids. Recently, there has been increased interest in the inflammatory markers in neurotrauma that can be coupled with neural specific protein to constitute a panel of TBI markers; several of these inflammatory markers have shown to be elevated clinically after severe TBI FK and HB assisted in the systems biology data analysis.
OG has performed the TBI animal injury and sacrifice. All the authors assisted in the final assessment of data and reviewed the manuscript. All the authors have read and approved the final manuscript. The other authors declare no competing financial interests.
Naify Ramadan | Medarbetare
The opinions stated are the private views of the authors and are not to be construed as official views of the Department of the Army or the Department of Defense. Opinions, interpretations, conclusions, and recommendations are those of the authors and are not necessarily endorsed by the DoD. In conducting research using animals, the investigators adhere to the laws of the USA and regulations of the Department of Agriculture.
The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. Emergency department visits for traumatic brain injury in older adults in the United States: West J Emerg Med 13 3 — Epidemiology of adults receiving acute inpatient rehabilitation for a primary diagnosis of traumatic brain injury in the United States.
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The Centers for Disease Control and Prevention estimate that 1. Efforts continue to focus on elucidating the complex molecular mechanisms underlying TBI pathophysiology and defining sensitive and specific biomarkers that can aid in improving patient management and care. Recently, the area of neuroproteomics-systems biology is proving to be a prominent tool in biomarker discovery for central nervous system injury and other neurological diseases.
Rather than focusing on any one individual molecular entity, we used in silico systems biology approach to understand the global dynamics that govern proteins that are differentially altered post-injury. In addition, gene ontology analysis of the proteomic data was conducted in order to categorize the proteins by molecular function, biological process, and cellular localization.
Our study is among the first to assess temporal neuroproteome changes in the CCI model.
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Data presented here unveil potential neural biomarkers and therapeutic targets that could be used for diagnosis, for treatment and, most importantly, for temporal prognostic assessment following brain injury. Of interest, this work relies on in silico bioinformatics approach to draw its conclusion; further work is conducted for functional studies to validate and confirm the omics data obtained. Read Article at publisher's site. How does Europe PMC derive its citations network? Protein Interactions.
Protein Families. Nucleotide Sequences. Functional Genomics Experiments. Protein Structures.