Chronic Upregulation of Cleaved-Caspase-3 Associated with Chronic Myelin Pathology and Microvascular Reorganization in the Thalamus after Traumatic Brain Injury in Rats.

Traumatic brain injury (TBI) is associated with long-term disabilities and devastating chronic neurological complications including problems with cognition, motor function, sensory processing, as well as behavioral deficits and mental health problems such as anxiety, depression, personality change and social unsuitability. Clinical data suggest that disruption of the thalamo-cortical system including anatomical and metabolic changes in the thalamus following TBI might be responsible for some chronic neurological deficits following brain trauma. Detailed mechanisms of these pathological processes are not completely understood. The goal of this study was to evaluate changes in the thalamus following TBI focusing on cleaved-caspase-3, a specific effector of caspase pathway activation and myelin and microvascular pathologies using immuno- and histochemistry at different time points from 24 h to 3 months after controlled cortical impact (CCI) in adult Sprague-Dawley rats. Significant increases in cleaved-caspase-3 immunoreactivity in the thalamus were observed starting one month and persisting for at least three months following experimental TBI. Further, the study demonstrated an association of cleaved-caspase-3 with the demyelination of neuronal processes and tissue degeneration in the gray matter in the thalamus, as reflected in alterations of myelinated fiber integrity (luxol fast blue) and decreases in myelin basic protein (MBP) immunoreactivity. The immunofluorescent counterstaining of cleaved-caspase-3 with endothelial barrier antigen (EBA), a marker of blood-brain barrier, revealed limited direct and indirect associations of cleaved caspase-3 with blood-brain barrier damage. These results demonstrate for the first time a significant chronic upregulation of cleaved-caspase-3 in selected thalamic regions associated with cortical regions directly affected by CCI injury. Further, our study is also the first to report that significant upregulation of cleaved-caspase-3 in selected ipsilateral thalamic regions is associated with microvascular reorganization reflected in the significant increases in the number of microvessels with blood-brain barrier alterations detected by EBA staining. These findings provide new insights into potential mechanisms of TBI cell death involving chronic activation of caspase-3 associated with disrupted cortico-thalamic and thalamo-cortical connectivity. Moreover, this study offers the initial evidence that this upregulation of activated caspase-3, delayed degeneration of myelinated nerve fibers and microvascular reorganization with impaired blood-brain barrier integrity in the thalamus might represent reciprocal pathological processes affecting neuronal networks and brain function at the chronic stages of TBI.

Interpretable deep learning of myelin histopathology in age-related cognitive impairment.

Age-related cognitive impairment is multifactorial, with numerous underlying and frequently co-morbid pathological correlates. Amyloid beta (Aß) plays a major role in Alzheimer's type age-related cognitive impairment, in addition to other etiopathologies such as Aß-independent hyperphosphorylated tau, cerebrovascular disease, and myelin damage, which also warrant further investigation. Classical methods, even in the setting of the gold standard of postmortem brain assessment, involve semi-quantitative ordinal staging systems that often correlate poorly with clinical outcomes, due to imperfect cognitive measurements and preconceived notions regarding the neuropathologic features that should be chosen for study. Improved approaches are needed to identify histopathological changes correlated with cognition in an unbiased way. We used a weakly supervised multiple instance learning algorithm on whole slide images of human brain autopsy tissue sections from a group of elderly donors to predict the presence or absence of cognitive impairment (n = 367 with cognitive impairment, n = 349 without). Attention analysis allowed us to pinpoint the underlying subregional architecture and cellular features that the models used for the prediction in both brain regions studied, the medial temporal lobe and frontal cortex. Despite noisy labels of cognition, our trained models were able to predict the presence of cognitive impairment with a modest accuracy that was significantly greater than chance. Attention-based interpretation studies of the features most associated with cognitive impairment in the top performing models suggest that they identified myelin pallor in the white matter. Our results demonstrate a scalable platform with interpretable deep learning to identify unexpected aspects of pathology in cognitive impairment that can be translated to the study of other neurobiological disorders.

Cuprizone Intoxication Results in Myelin Vacuole Formation.

Myelin damage is a histopathological hallmark of multiple sclerosis lesions. Results of post mortem studies suggest that impaired myelin-axon interaction characterized by focal myelin detachments is an early event during lesion genesis. In this study, we investigated the ultrastructural changes of the axon-myelin interface in the cuprizone model using serial block face scanning electron microscopy and immunohistochemistry. We show that non-inflammatory injury of oligodendrocytes by cuprizone intoxication results in myelin vacuole formation and axonal swellings, paralleled by early alterations of the node of Ranvier cytoarchitecture. This remarkable resemblance of ultrastructural myelin characteristics in multiple sclerosis and the cuprizone animal model suggests that the cuprizone model is a valuable tool to study early pathologies during lesion formation.

Myelin Pathology: Involvement of Molecular Chaperones and the Promise of Chaperonotherapy.

The process of axon myelination involves various proteins including molecular chaperones. Myelin alteration is a common feature in neurological diseases due to structural and functional abnormalities of one or more myelin proteins. Genetic proteinopathies may occur either in the presence of a normal chaperoning system, which is unable to assist the defective myelin protein in its folding and migration, or due to mutations in chaperone genes, leading to functional defects in assisting myelin maturation/migration. The latter are a subgroup of genetic chaperonopathies causing demyelination. In this brief review, we describe some paradigmatic examples pertaining to the chaperonins Hsp60 (HSPD1, or HSP60, or Cpn60) and CCT (chaperonin-containing TCP-1). Our aim is to make scientists and physicians aware of the possibility and advantages of classifying patients depending on the presence or absence of a chaperonopathy. In turn, this subclassification will allow the development of novel therapeutic strategies (chaperonotherapy) by using molecular chaperones as agents or targets for treatment.

Anillin facilitates septin assembly to prevent pathological outfoldings of central nervous system myelin.

Myelin serves as an axonal insulator that facilitates rapid nerve conduction along axons. By transmission electron microscopy, a healthy myelin sheath comprises compacted membrane layers spiraling around the cross-sectioned axon. Previously we identified the assembly of septin filaments in the innermost non-compacted myelin layer as one of the latest steps of myelin maturation in the central nervous system (CNS) (Patzig et al., 2016). Here we show that loss of the cytoskeletal adaptor protein anillin (ANLN) from oligodendrocytes disrupts myelin septin assembly, thereby causing the emergence of pathological myelin outfoldings. Since myelin outfoldings are a poorly understood hallmark of myelin disease and brain aging we assessed axon/myelin-units in Anln-mutant mice by focused ion beam-scanning electron microscopy (FIB-SEM); myelin outfoldings were three-dimensionally reconstructed as large sheets of multiple compact membrane layers. We suggest that anillin-dependent assembly of septin filaments scaffolds mature myelin sheaths, facilitating rapid nerve conduction in the healthy CNS.