Heterogeneous immunoreactivity of axonal spheroids in focal symmetrical encephalomalacia produced by Clostridium perfringens type D epsilon toxin in sheep

Since axonal injury (AI) is an important component of many veterinary neurologic disorders, we assessed the relative ability of a panel of antibodies (amyloid precursor protein, 3 subunits of neurofilament protein, protein gene product 9.5, ubiquitin, and synaptophysin) to detect axonal swellings or spheroids. Abundant axonal spheroids found in necrotic internal capsule foci produced in 4 sheep by chronic Clostridium perfringens type D epsilon neurotoxicity provided a model system in which to evaluate this important diagnostic tool. There was heterogeneous labeling of subsets of spheroids by the respective antibodies, suggesting that, in order to detect the complete spectrum of AI in diagnostic cases, a range of antibodies should be used, not only when spheroids are plentiful but also when they are few in number or incompletely developed. The application of insufficient markers in the latter cases can potentially lead to the contribution of AI to lesion pathogenesis being underappreciated.

Human neocortical expansion involves glutamatergic neuron diversification

The neocortex is disproportionately expanded in human compared with mouse1,2, both in its total volume relative to subcortical structures and in the proportion occupied by supragranular layers composed of neurons that selectively make connections within the neocortex and with other telencephalic structures. Single-cell transcriptomic analyses of human and mouse neocortex show an increased diversity of glutamatergic neuron types in supragranular layers in human neocortex and pronounced gradients as a function of cortical depth3. Here, to probe the functional and anatomical correlates of this transcriptomic diversity, we developed a robust platform combining patch clamp recording, biocytin staining and single-cell RNA-sequencing (Patch-seq) to examine neurosurgically resected human tissues. We demonstrate a strong correspondence between morphological, physiological and transcriptomic phenotypes of five human glutamatergic supragranular neuron types. These were enriched in but not restricted to layers, with one type varying continuously in all phenotypes across layers 2 and 3. The deep portion of layer 3 contained highly distinctive cell types, two of which express a neurofilament protein that labels long-range projection neurons in primates that are selectively depleted in Alzheimer’s disease4,5. Together, these results demonstrate the explanatory power of transcriptomic cell-type classification, provide a structural underpinning for increased complexity of cortical function in humans, and implicate discrete transcriptomic neuron types as selectively vulnerable in disease.

Plasma biomarker profiles and the correlation with cognitive function across the clinical spectrum of Alzheimer’s disease

Background Plasma biomarkers showed a promising value in the disease diagnosis and management of Alzheimer’s disease (AD). However, profiles of the biomarkers and the associations with cognition across a spectrum of cognitive stages have seldom been reported. Methods We recruited 320 individuals with cognitive impairment and 131 cognitively normal participants from a memory clinic and a community cohort. Participants were classified into 6 groups based on their Clinical Dementia Rating (CDR) scores and clinical diagnosis, including AD, amnestic mild cognitive impairment (aMCI), and normal cognition (NC). A battery of neuropsychological tests was used to assess the global and domain-specific cognition. Plasma Aβ1-40, Aβ1-42, Aβ1-42/Aβ1-40, total tau (t-tau), neurofilament protein light chain (NfL), and phosphorylated tau at threonine 181 (p-tau181) were quantified using the single-molecule array (Simoa) platform. Results All the plasma markers (Aβ1-40, Aβ1-42, Aβ1-42/Aβ1-40, t-tau, NfL, p-tau181) showed certain discrepancies among NC, aMCI, and AD groups. The p-tau181 level showed a continuous escalating trend as the CDR scores increased from 0 (NC group) to 3 (severe AD). Compared with other biomarkers, p-tau181 had correlations with broader cognitive domains, covering global cognition (r = −0.536, P < 0.0001), memory (r = −0.481, P < 0.0001), attention (r = −0.437, P < 0.0001), visuospatial function (r = −0.385, P < 0.0001), and language (r = −0.177, P = 0.0003). Among participants with CDR ≥ 1, higher p-tau181 was correlated with worse global cognition (r = −0.301, P < 0.001). Conclusions Plasma p-tau181 had correlations with broader cognitive domains, suggesting its potential as a promising clinical-relevant blood-based biomarker.

Identification of sensory and motor nerve fascicles by immunofluorescence staining after peripheral nerve injury

Background Inappropriate matching of motor and sensory fibers after nerve repair or nerve grafting can lead to failure of nerve recovery. Identification of motor and sensory fibers is important for the development of new approaches that facilitate neural regeneration and the next generation of nerve signal-controlled neuro-prosthetic limbs with sensory feedback technology. Only a few methods have been reported to differentiate sensory and motor nerve fascicles, and the reliability of these techniques is unknown. Immunofluorescence staining is one of the most commonly used methods to distinguish sensory and motor nerve fibers, however, its accuracy remains unknown. Methods In this study, we aim to determine the efficacy of popular immunofluorescence markers for motor and sensory nerve fibers. We harvested the facial (primarily motor fascicles) and sural (primarily sensory fascicles) nerves in rats, and examined the immunofluorescent staining expressions of motor markers (choline acetyltransferase (ChAT), tyrosine kinase (TrkA)), and sensory markers [neurofilament protein 200 kDa (NF-200), calcitonin gene-related peptide (CGRP) and Transient receptor potential vanillic acid subtype 1 (TRPV1)]. Three methods, including the average area percentage, the mean gray value, and the axon count, were used to quantify the positive expression of nerve markers in the immunofluorescence images. Results Our results suggest the mean gray value method is the most reliable method. The mean gray value of immunofluorescence in ChAT (63.0 ± 0.76%) and TRKA (47.6 ± 0.43%) on the motor fascicles was significantly higher than that on the sensory fascicles (ChAT: 49.2 ± 0.72%, P < 0.001; and TRKA: 29.1 ± 0.85%, P < 0.001). Additionally, the mean gray values of TRPV1 (51.5 ± 0.83%), NF-200 (61.5 ± 0.62%) and CGRP (37.7 ± 1.22%) on the motor fascicles were significantly lower than that on the sensory fascicles respectively (71.9 ± 2.32%, 69.3 ± 0.46%, and 54.3 ± 1.04%) (P < 0.001). The most accurate cutpoint occurred using CHAT/CRCP ratio, where a value of 0.855 had 100% sensitivity and 100% specificity to identify motor and sensory nerve with an area under the ROC curve of 1.000 (P < 0.001). Conclusions A combination of ChAT and CGRP is suggested to distinguish motor and sensory nerve fibers.

Individual variability in the size and organization of the human arcuate nucleus of the medulla

The arcuate nucleus (Arc) of the medulla is found in almost all human brains and in a small percentage of chimpanzee brains. It is absent in the brains of other mammalian species including mice, rats, cats and macaque monkeys. The Arc is classically considered a precerebellar nucleus, presumably motor in function. However, several studies have found aplasia of the Arc in babies who died of SIDS (Sudden Infant Death Syndrome) suggesting a very different role for the Arc in the control of respiration. Aplasia of the Arc, however, has also been reported in adults suggesting that it is not critical for survival. We have examined the Arc in closely-spaced Nissl-stained sections in human cases to acquire a better understanding of the degree of variability of its size and location in adults. We have also examined immunostained sections to develop a neurochemical profile of this nucleus. Our data suggest that the Arc consists of rostral and caudal subdivisions. There is some degree of left-right asymmetry in Arc position, size and shape at all rostro-caudal levels. Neurons in the Arc express calretinin (CR), neuronal nitric oxide synthase (nNOS) and nonphosphorylated neurofilament protein (NPNFP) but not calbindin (CB) or parvalbumin (PV). There is also immunostaining for GAD and GABA receptors suggesting inhibitory input to Arc neurons. We support the classical idea of the Arc as a precerebellar nucleus; its absence in SIDS cases may not be the cause of death but rather a correlate of other brainstem abnormalities.

Diagnostic and pathogenic features of calcifying pseudoneoplasm of the neuraxis

Calcifying pseudoneoplasm of the neuraxis (CAPNON) is a rare tumefactive lesion with unclear pathogenesis. It is diagnosed by pathological findings of the typical histological features that include granular amorphous cores with palisading spindle to epithelioid cells, variable fibrous stroma, foreign-body reaction with giant cells, and calcification/ossification occasionally with psammoma bodies. However, its histopathology may be variable and currently immunohistochemistry plays a limited role in its diagnosis and understanding the pathogenesis. In this study, we examined 6 cases of CAPNONs including 3 intracranial and 3 spinal epidural lesions (age range: 59–69 years; 3 males and 3 females). Immunohistochemistry revealed that all CAPNON cores contain abundant positive deposits of neurofilament protein (NFP), which was supported by electron microscopy finding of filaments (8–13 nm in diameter). In comparison, no NFP positivity was found in 5 psammomatous/metaplastic meningiomas or 7 intervertebral tissue lesions with calcification/ossification. In addition, CAPNON cellular areas showed variable numbers of CD8+ cytotoxic T-cells with less CD4+ T-cells and a decreased ratio of CD4/CD8+ cells, versus the intervertebral tissue lesions without CD8+ or CD4+ cells. Our findings suggest that NFP may be a principal constituent of CAPNONs, and thus involved in the pathogenesis of CAPNON. Given the decreased CD4/CD8 ratio, the pathogenic process of CAPNON is possibly immune- mediated.LEARNING OBJECTIVESThe presentation will enable the learner to: 1.Discuss histopathological features of calcifying pseudoneoplasm of the neuraxis (CAPNON) with variation of non-core components.2.Explore diagnostic and pathogenic roles of immunohistochemical markers including neurofilament protein and CD4/CD8 in CAPNON.

AAV2-mediated and hypoxia response element-directed expression of bFGF in neural stem cells showed therapeutic effects on spinal cord injury in rats

Neural stem cell (NSCs) transplantation has been one of the hot topics in the repair of spinal cord injury (SCI). Fibroblast growth factor (FGF) is considered a promising nerve injury therapy after SCI. However, owing to a hostile hypoxia condition in SCI, there remains a challenging issue in implementing these tactics to repair SCI. In this report, we used adeno-associated virus 2 (AAV2), a prototype AAV used in clinical trials for human neuron disorders, basic FGF (bFGF) gene under the regulation of hypoxia response element (HRE) was constructed and transduced into NSCs to yield AAV2-5HRE-bFGF-NSCs. Our results showed that its treatment yielded temporally increased expression of bFGF in SCI, and improved scores of functional recovery after SCI compared to vehicle control (AAV2-5HRE-NSCs) based on the analyses of the inclined plane test, Basso–Beattie–Bresnahan (BBB) scale and footprint analysis. Mechanistic studies showed that AAV2-5HRE-bFGF-NSCs treatment increased the expression of neuron-specific neuronal nuclei protein (NeuN), neuromodulin GAP43, and neurofilament protein NF200 while decreased the expression of glial fibrillary acidic protein (GFAP) as compared to the control group. Further, the expressions of autophagy-associated proteins LC3-II and Beclin 1 were decreased, whereas the expression of P62 protein was increased in AAV2-5HRE-bFGF-NSCs treatment group. Taken together, our data indicate that AAV2-5HRE-bFGF-NSCs treatment improved the recovery of SCI rats, which is accompanied by evidence of nerve regeneration, and inhibition of SCI-induced glial scar formation and cell autophagy. Thus, this study represents a step forward towards the potential use of AAV2-5HRE-bFGF-NSCs for future clinical trials of SCI repair.

Plasma biomarker profiles and the correlation with cognitive function across the clinical spectrum of Alzheimer’s disease

BackgroundPlasma biomarkers showed a promising value in the disease diagnosis and management of Alzheimer’s disease (AD). However, profiles of the biomarkers and the association with cognitive domains along the spectrum of cognitive performance deterioration have seldom been reported.MethodsWe recruited 320 individuals with cognitive impairment and 131 cognitively normal participants from a memory clinic and a community cohort. Participants were classified into 6 groups based on their Clinical Dementia Rating (CDR) scores and clinical diagnosis of AD, amnestic mild cognitive impairment, and normal cognition (NC). Each participant was administered the neuropsychological tests assessing the global and domain-specific cognition. Plasma Aβ1-40, Aβ1-42, Aβ1-42/Aβ1-40, total tau (t-tau), neurofilament protein light chain (NfL), and phosphorylated tau at threonine 181 (p-tau181) were quantified using the Single molecule array platform.ResultsAlong with plasma Aβ1-40, Aβ1-42, Aβ1-42/Aβ1-40, t-tau, and NfL, p-tau181 significantly increased across the groups with the incremental CDR scores from NC (CDR = 0) to severe AD (CDR = 3). Compared with other biomarkers, p-tau181 had a stronger correlation with Global cognition (r = −0.494, P < 0.001), Memory (r = −0.417, P < 0.001), Attention (r = −0.388, P < 0.001), Visuospatial function (r = −0.328, P < 0.001), and Language (r = −0.123, P = 0.014). Among AD participants with CDR ≥ 1, higher p-tau181 was correlated with worse Global cognition (r = −0.295, P < 0.001), Memory (r = −0.172, P = 0.045), and Attention (r = −0.184, P = 0.031).ConclusionsPlasma p-tau181 had a stronger correlation with cognitive domains than other biomarkers, especially in late-stage AD. It could reflect the AD pathology in vivo and may be a promising blood-based biomarker in clinical settings.