Abstract. The automated laser-based hematology analyzer Sysmex XT-2000iV™ provides a 5-part differential count and specific cytograms that are of great interest for large veterinary laboratories. The aim of the study was to validate the Sysmex XT-2000iV compared to the laser-based hematology analyzer ADVIA® 2120 and manual differential in dogs, cats, and horses as well as the impact of anticoagulant (heparin, ethylenediamine tetra-acetic acid [EDTA], and citrate) and storage at 22°C and 4°C. Consecutive fresh K 3 -EDTA blood samples from 216 cats, 314 dogs, and 174 horses were included. The impact of anticoagulant and sample storage was assessed in specimens obtained from an additional 9 cats, 10 dogs, and 10 horses. Agreement between both analyzers was excellent to good except for monocytes and canine reticulocytes. Spearman rank correlation coefficients (r s ) between Sysmex XT-2000iV and manual differential were good to fair and ranged from 0.91 (cat lymphocytes) to 0.44 (cat monocytes). Hematocrit value (Hct), mean corpuscular hemoglobin (MCH), MCH concentration (MCHC; all: P < 0.001), and mean corpuscular volume (MCV; P < 0.01) were higher in canine citrated blood compared to heparin and EDTA. In cats, lymphocytes and monocytes were lower in heparinized blood compared to EDTA (P < 0.05), whereas in horses no significant effect was seen. Regarding storage time and temperature, white and red blood cell counts, hemoglobin, and MCH were stable. Hct, MCV, and MCHC were influenced by erythrocyte swelling. Differential count remained stable for 24 hr (22°C) and nearly 72 hr (4°C) except for monocytes. The overall performance of the Sysmex XT2000iV was excellent and compared favorably with that of the ADVIA 2120. A special strength was the excellent detection of feline eosinophils.

The automated laser-based hematology analyzer Sysmex XT-2000iV™ providing a complete blood cell count (CBC) and 5-part differential has been introduced in large veterinary laboratories. The aim of the current study was to determine precision, linearity, and accuracy of the Sysmex analyzer. Reference method for the accuracy study was the laser-based hematology analyzer ADVIA® 2120. For evaluation of accuracy, consecutive fresh blood samples from healthy and diseased cats (n = 216), dogs (n = 314), and horses (n = 174) were included. A low intra-assay coefficient of variation (CV) of approximately 1% was seen for the CBC except platelet count (PLT). An intra-assay CV ranging between 2% and 5.5% was evident for the differential count except for feline and equine monocytes (7.7%) and horse eosinophils (15.7%). Linearity was excellent for white blood cell count (WBC), hematocrit value, red blood cell count (RBC), and PLT. For all evaluated species, agreement was excellent for WBC and RBC, with Spearman rank correlation coefficients (r(s)) ranging from >0.99 to 0.98. Hematocrit value correlated excellently in cats and dogs, whereas for horses, a good correlation was evident. A good correlation between both analyzers was seen in feline and equine PLT (r(s) = 0.89 and 0.92, respectively), whereas correlation was excellent for dogs (r(s) = 0.93). Biases were close to 0 except for mean corpuscular hemoglobin concentration (4.11 to -7.25 mmol/l) and canine PLT (57 × 10(9)/l). Overall, the performance of the Sysmex analyzer was excellent and compared favorably with the ADVIA analyzer.

Focal cortical dysplasias (FCDs) are local malformations of the human neocortex with strong epileptogenic potential. To investigate the underlying pathomechanisms, we performed a whole human transcriptome screening to compare the gene expression pattern of dysplastic versus nondysplastic temporal neocortex. Tissue obtained from FCD IIIa cases (mean age 20.5 years) who had undergone surgical treatment, due to intractable epilepsy, was compared with nondysplastic specimens (mean age 19.9 years) by means of Affymetrix arrays covering 28 869 genes. We found 211 differentially expressed genes (DEX) among which mainly genes important for oligodendrocyte differentiation and myelination were downregulated in FCD IIIa. These findings were confirmed as functionally important by Database for Annotation, Visualization, and Integrated Discovery (DAVID) analysis. The reduced expression of myelin-associated transcripts was confirmed for FCD Ia, IIa, and IIIa by real-time RT-qPCR. In addition, we found that the density of myelin basic protein mRNA-expressing oligodendrocytes and of 2',3'-cyclic nucleotide 3'-phosphodiesterase-positive myelin fibers was significantly reduced in dysplastic cortex. Moreover, high-resolution confocal imaging and 3D reconstruction revealed that the myelin fiber network was severely disorganized in dysplastic neocortex, indicating a disturbance of myelin sheath formation and maintenance in FCD.

The PocH-100iV Diff is a suitable in-house instrument. A strength is its specific, but moderately sensitive, detection of feline eosinophils.

BackgroundFocal cortical dysplasias (FCD) are local disturbances of neocortical architecture and a common cause of pharmaco-resistant focal epilepsy. Little is known about the pathomechanisms leading to architectural abnormalities associated with FCD.ResultsIn the present study we compared 52 FCD cases originating from the frontal or temporal lobe with or without Ammon’s horn sclerosis (AHS) with regard to structural and molecular differences. We applied layer-specific (ER81, RORß, SMI32, TLE4) and interneuron (calbindin, parvalbumin) markers by means of immunohistochemistry, in situ hybridization (ISH), and real time RT-PCR and correlated our findings with clinical parameters. We found that: (1) Structural abnormalities were most prominent in layers III-VI including changed morphology of individual neurons or dispersion, blurring and thinning of layers. These alterations were most pronounced in isolated frontal FCD, whereas the most homogeneous group was FCD IIIa. (2) Numbers of calbindin- and parvalbumin-positive interneurons varied considerably within the different FCD groups, but were not generally reduced. A significant decrease was only found for calbindin-positive interneurons in frontal FCD, and for parvalbumin-positive interneurons in FCD IIIa. (3) Interestingly, FCD IIIa presented with significant changes in the numbers of calbindin- or TLE4-positive neurons when compared to isolated FCD or controls. (4) Correlations between clinical and cellular parameters strongly depended on FCD localisation and age of the patients.ConclusionsIn summary, our data suggest that late cortical development is disturbed in FCD, yet most likely by different causes depending on brain region, FCD type and FCD severity.

Surgical treatment of extratemporal epilepsy provides satisfying epileptological results with an acceptable morbidity. Best results can be achieved in younger patients with circumscribed MRI lesions, which can be resected completely.

Objectives: Focal cortical dysplasias (FCDs) are local malformations of the human neocortex and a leading cause of medically intractable epilepsy. FCDs are characterized by local architectural disturbances of the neocortex and often by a blurred gray-white matter boundary indicating abnormal white matter myelination. We have recently shown that myelination is also compromised in the gray matter of dysplastic areas, since transcripts encoding factors for oligodendrocyte differentiation and myelination are downregulated and myelin fibers appear fractured and disorganized. Methods: Here, we characterized the gray matter-associated myelination pathology in detail by in situ hybridization, immunohistochemistry, and electron microscopy with markers for myelin, mature oligodendrocytes, and oligodendrocyte precursor cells in tissue sections of FCD IIa and control cortices. In addition, we isolated oligodendrocyte precursor cells from resected dysplastic tissue and performed proliferation assays. Results: We show that the proportion of myelinated gray matter is similar in the dysplastic cortex to that in controls and myelinated fibers extend up to layer III.On the ultrastructural level, however, we found that the myelin sheaths of layer V axons are thinner in dysplastic specimens than in controls. In addition, the density of 172 | DONKELS Et aL. How to cite this article: Donkels C, Peters M, Fariña Núñez MT, et al. Oligodendrocyte lineage and myelination are compromised in the gray matter of focal cortical dysplasia type IIa. Epilepsia.

The application of a broad panel of markers defining layer-specific neuronal subpopulations revealed that in FCD2b neuronal diversity and a basic laminar arrangement are maintained despite the severe disturbance of cytoarchitecture. Moreover, it showed that Parvalbumin-positive, inhibitory interneurons are highly vulnerable in contrast to other interneuron subtypes, possibly related to the epileptic condition.