Traumatic brain injury (TBI) is a heterogeneous condition, associated with diverse etiologies, clinical presentations and degrees of severity, and may result in chronic neurobehavioral sequelae. The field of TBI biomarkers is rapidly evolving to address the many facets of TBI pathology and improve its clinical management. Recent years have witnessed a marked increase in the number of publications and interest in the role of extracellular vesicles (EVs), which include exosomes, cell signaling, immune responses, and as biomarkers in a number of pathologies. Exosomes have a well-defined lipid bilayer with surface markers that reflect the cell of origin and an aqueous core that contains a variety of biological material including proteins (e.g., cytokines and growth factors) and nucleic acids (e.g., microRNAs). The presence of proteins associated with neurodegenerative changes such as amyloid-β, α-synuclein and phosphorylated tau in exosomes suggests a role in the initiation and propagation of neurological diseases. However, mechanisms of cell communication involving exosomes in the brain and their role in TBI pathology are poorly understood. Exosomes are promising TBI biomarkers as they can cross the blood-brain barrier and can be isolated from peripheral fluids, including serum, saliva, sweat, and urine. Exosomal content is protected from enzymatic degradation by exosome membranes and reflects the internal environment of their cell of origin, offering insights into tissue-specific pathological processes. Challenges in the clinical use of exosomal cargo as biomarkers include difficulty in isolating pure exosomes, variable yields of the isolation processes, quantification of vesicles, and lack of specificity of exosomal markers. Moreover, there is no consensus regarding nomenclature and characteristics of EV subtypes. In this review, we discuss current technical limitations and challenges of using exosomes and other EVs as blood-based biomarkers, highlighting their potential as diagnostic and prognostic tools in TBI.

DNA interstrand cross-links are complex lesions that covalently bind complementary strands of DNA and whose mechanism of repair remains poorly understood. In , several gene products have been proposed to be involved in cross-link repair based on the hypersensitivity of mutants to cross-linking agents. However, cross-linking agents induce several forms of DNA damage, making it challenging to attribute mutant hypersensitivity specifically to interstrand cross-links. To address this, we compared the survival of UVA-irradiated repair mutants in the presence of 8-methoxypsoralen-which forms interstrand cross-links and monoadducts-to that of angelicin-a congener forming only monoadducts. We show that incision by nucleotide excision repair is not required for resistance to interstrand cross-links. In addition, neither RecN nor DNA polymerases II, IV, or V is required for interstrand cross-link survival, arguing against models that involve critical roles for double-strand break repair or translesion synthesis in the repair process. Finally, estimates based on Southern analysis of DNA fragments in alkali agarose gels indicate that lethality occurs in wild-type cells at doses producing as few as one to two interstrand cross-links per genome. These observations suggest that may lack an efficient repair mechanism for this form of damage.

Psoralen DNA interstrand cross-links are highly toxic lesions with antimicrobial and anticancer properties. Despite the lack of effective mechanisms for repair, cells can become resistant to cross-linking agents through mechanisms that remain poorly defined.

Abstract8-methoxypsoralen is a DNA-intercalating agent, which can photoreact with pyrimidine bases on opposing DNA strands, to form an interstrand crosslink. These lesions completely block replication and transcription, and are widely used in chemotherapies; yet how these lesions are processed in the cell remains poorly understood and insight into these processes could lead to better therapies that evade resistance. Previous studies isolated an Escherichia coli mutant demonstrating hyper-resistance to interstrand crosslink-inducing agents. The mutation was mapped to 57.2 minutes on the chromosome, and potentially encoded a 55-kDa protein induced as part of the SOS response. Although these genes remain unidentified, hscA and hscB map to this location, have a similar size, and are SOS-inducible. To determine if these or other genes might confer interstrand crosslink resistance in E. coli, we characterized how cells survived 8-methoxypsoralen-UVA treatment in the absence of HscAB, and when these gene products were overexpressed. In a second approach, we developed a selection system to isolate hyper-resistant strains through the sequential growth and exposure of wild-type cultures to 8-methoxypsoralen-UVA. We found no effect on cell survival in the hscAB mutant compared to its wild-type parent, suggesting that HscAB may not contribute to interstrand crosslink resistance as previously hypothesized. However, due to the significant cytotoxicity of plasmids containing hscAB even in the absence of 8-methoxypsoralen-UVA treatment, we could not determine whether overexpression of these gene products provided cellular protection. Using iteratively 8-methoxypsoralen-UVA treated cells, we isolated strains that were >10 4 -fold more resistant to this interstrand crosslink-inducing agent compared to the parent strain. This result suggests that E.coli possess mechanisms of interstrand crosslink repair or tolerance and could serve as a model system for understanding the development of drug resistance in human cells.3