Determining the number of synapses that are present in different brain regions is crucial to understand brain connectivity as a whole. Membrane-associated guanylate kinases (MAGUKs) are a family of ...scaffolding proteins that are expressed in excitatory glutamatergic synapses. We used genetic labeling of two of these proteins (PSD95 and SAP102), and Spinning Disc confocal Microscopy (SDM), to estimate the number of fluorescent puncta in the CA1 area of the hippocampus. We also used FIB-SEM, a three-dimensional electron microscopy technique, to calculate the actual numbers of synapses in the same area. We then estimated the ratio between the three-dimensional densities obtained with FIB-SEM (synapses/µm
) and the bi-dimensional densities obtained with SDM (puncta/100 µm
). Given that it is impractical to use FIB-SEM brain-wide, we used previously available SDM data from other brain regions and we applied this ratio as a conversion factor to estimate the minimum density of synapses in those regions. We found the highest densities of synapses in the isocortex, olfactory areas, hippocampal formation and cortical subplate. Low densities were found in the pallidum, hypothalamus, brainstem and cerebellum. Finally, the striatum and thalamus showed a wide range of synapse densities.
The medulla oblongata is the caudal portion of the vertebrate hindbrain. It contains major ascending and descending fiber tracts as well as several motor and interneuron populations, including neural ...centers that regulate the visceral functions and the maintenance of bodily homeostasis. In the avian embryo, it has been proposed that the primordium of this region is subdivided into five segments or crypto-rhombomeres (r7–r11), which were defined according to either their parameric position relative to intersomitic boundaries (Cambronero and Puelles, in J Comp Neurol 427:522–545,
2000
) or a stepped expression of Hox genes (Marín et al., in Dev Biol 323:230–247,
2008
). In the present work, we examine the implied similar segmental organization of the mouse medulla oblongata. To this end, we analyze the expression pattern of Hox genes from groups 3 to 8, comparing them to the expression of given cytoarchitectonic and molecular markers, from mid-gestational to perinatal stages. As a result of this approach, we conclude that the mouse medulla oblongata is segmentally organized, similarly as in avian embryos. Longitudinal structures such as the nucleus of the solitary tract, the dorsal vagal motor nucleus, the hypoglossal motor nucleus, the descending trigeminal and vestibular columns, or the reticular formation appear subdivided into discrete segmental units. Additionally, our analysis identified an internal molecular organization of the migrated pontine nuclei that reflects a differential segmental origin of their neurons as assessed by Hox gene expression.
(A) Correlation matrix of unsupervised co‐regulated genes, based on the 208 genes included in the NanoString platform. Some of the clusters of co‐regulated genes corresponded to the following: ...Inflammatory cells; Epstein–Barr virus; B‐cells; Cytotoxic T‐cells; T‐cells; and Proliferation. (B) Analysis of genomic alterations by targeted sequencing. Distribution of mutations in the 62 analyzed genes. Rows correspond to sequenced genes, columns represent individual patients. Color coding: green, missense; blue, synonymous; pink, frameshift; violet, Indel; red, stop gained; yellow, UTR.
Gene profiling using a customized NanoString platform can be applied to routine mycosis fungoides (MF) samples, revealing their specific molecular features. This study shows that MF samples carry a ...signature derived from individual molecular features found in consecutive biopsies.
Peripheral T-cell lymphoma (PTCL) is a clinically aggressive disease, with a poor response to therapy and a low overall survival rate of approximately 30% after 5 years. We have analyzed a series of ...105 cases with a diagnosis of PTCL using a customized NanoString platform (NanoString Technologies, Seattle, WA) that includes 208 genes associated with T-cell differentiation, oncogenes and tumor suppressor genes, deregulated pathways, and stromal cell subpopulations. A comparative analysis of the various histological types of PTCL (angioimmunoblastic T-cell lymphoma AITL; PTCL with T follicular helper TFH phenotype; PTCL not otherwise specified NOS) showed that specific sets of genes were associated with each of the diagnoses. These included TFH markers, cytotoxic markers, and genes whose expression was a surrogate for specific cellular subpopulations, including follicular dendritic cells, mast cells, and genes belonging to precise survival (NF-κB) and other pathways. Furthermore, the mutational profile was analyzed using a custom panel that targeted 62 genes in 76 cases distributed in AITL, PTCL-TFH, and PTCL-NOS. The main differences among the 3 nodal PTCL classes involved the RHOAG17V mutations (P < .0001), which were approximately twice as frequent in AITL (34.09%) as in PTCL-TFH (16.66%) cases but were not detected in PTCL-NOS. A multivariate analysis identified gene sets that allowed the series of cases to be stratified into different risk groups. This study supports and validates the current division of PTCL into these 3 categories, identifies sets of markers that can be used for a more precise diagnosis, and recognizes the expression of B-cell genes as an IPI-independent prognostic factor for AITL.
•Gene expression and mutational analysis confirm the differences among the 3 peripheral TCL subclasses: AITL, PTCL-NOS, and PTCL-TFH.•The expression of a gene set, including B-cell genes, is an IPI-independent prognostic factor for AITL cases.
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Neurodevelopmental disorders of genetic origin delay the acquisition of normal abilities and cause disabling phenotypes. Nevertheless, spontaneous attenuation and even complete amelioration of ...symptoms in early childhood and adolescence can occur in many disorders, suggesting that brain circuits possess an intrinsic capacity to overcome the deficits arising from some germline mutations. We examined the molecular composition of almost a trillion excitatory synapses on a brain-wide scale between birth and adulthood in mice carrying a mutation in the homeobox transcription factor Pax6, a neurodevelopmental disorder model. Pax6 haploinsufficiency had no impact on total synapse number at any age. By contrast, the molecular composition of excitatory synapses, the postnatal expansion of synapse diversity and the acquisition of normal synaptome architecture were delayed in all brain regions, interfering with networks and electrophysiological simulations of cognitive functions. Specific excitatory synapse types and subtypes were affected in two key developmental age-windows. These phenotypes were reversed within 2-3 weeks of onset, restoring synapse diversity and synaptome architecture to the normal developmental trajectory. Synapse subtypes with rapid protein turnover mediated the synaptome remodeling. This brain-wide capacity for remodeling of synapse molecular composition to recover and maintain the developmental trajectory of synaptome architecture may help confer resilience to neurodevelopmental genetic disorders.
Möbius syndrome (MBS) is a neurological disorder that is characterized by paralysis of the facial nerves and variable other congenital anomalies. The aetiology of this syndrome has been enigmatic ...since the initial descriptions by von Graefe in 1880 and by Möbius in 1888, and it has been debated for decades whether MBS has a genetic or a non-genetic aetiology. Here, we report de novo mutations affecting two genes, PLXND1 and REV3L in MBS patients. PLXND1 and REV3L represent totally unrelated pathways involved in hindbrain development: neural migration and DNA translesion synthesis, essential for the replication of endogenously damaged DNA, respectively. Interestingly, analysis of Plxnd1 and Rev3l mutant mice shows that disruption of these separate pathways converge at the facial branchiomotor nucleus, affecting either motoneuron migration or proliferation. The finding that PLXND1 and REV3L mutations are responsible for a proportion of MBS patients suggests that de novo mutations in other genes might account for other MBS patients.
Diffuse large B‐cell lymphoma (DLBCL), the most frequent non‐Hodgkin's lymphoma subtype, is characterized by strong biological, morphological, and clinical heterogeneity, but patients are treated ...with immunochemotherapy in a relatively homogeneous way. Here, we have used a customized NanoString platform to analyze a series of 197 homogeneously treated DLBCL cases. The platform includes the most relevant genes or signatures known to be useful for predicting response to R‐CHOP (Rituximab, Cyclophosphamide, Doxorubicin, Vincristine, and Prednisone) in DLBCL cases. We generated a risk score that combines the International Prognostic Index with cell of origin and double expression of MYC/BCL2, and stratified the series into three groups, yielding hazard ratios from 0.15 to 5.49 for overall survival, and from 0.17 to 5.04 for progression‐free survival. Group differences were highly significant (p < 0.0001), and the scoring system was applicable to younger patients (<60 years of age) and patients with advanced or localized stages of the disease. Results were validated in an independent dataset from 166 DLBCL patients treated in two distinct clinical trials. This risk score combines clinical and biological data in a model that can be used to integrate biological variables into the prognostic models for DLBCL cases.
Hereditary congenital facial paresis (HCFP) consists of the paralysis or weakness of facial muscles caused by a maldevelopment of the facial branchiomotor (FBM) nucleus and its nerve. Linkage ...analyses have related this disorder to two loci, HCFP1 and HCFP2, placed respectively in human chromosomes 3q21.2-q22.1 and 10q21.3-q22.1, but the causative genes are still unknown. In this work we aimed to identify which genes from these loci are expressed in the developing hindbrain and particularly in the FBM nucleus. To this end, we retrieved from the ENSEMBL genomic database the list of these genes as well as their respective mouse orthologs. Subsequently we examined their respective expression patterns in the mouse embryo by using the GenePaint gene expression database. As a result of this screening, we found a new gene (Mgll) from the HCFP1 locus that has strong and specific expression in the developing FBM nucleus. In its turn, the HCFP2 locus appeared as a large gene-desert region, flanked by two genes, Reep3, with specific expression in the FBM nucleus, and Lrrtm3, broadly expressed in the brainstem, including the same nucleus. The concurrence of genomic position and neural expression pattern makes these genes new potential candidates for HCFP.
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