Hemodynamic changes in patients with SARS-CoV-2 infection presenting for cesarean delivery under spinal anesthesia: a retrospective case-control study.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is associated with adverse maternal and neonatal outcomes. Early studies suggested that COVID-19 was associated with a higher incidence of hypotension following neuraxial anesthesia in parturients. We explored the hemodynamic response to spinal anesthesia for cesarean delivery in pregnant severe respiratory distress syndrome-coronavirus-2 (SARS-CoV-2) positive patients, using a retrospective case-control design. METHODS: We searched our electronic medical records for patients who received spinal anesthesia for cesarean delivery, and were SARS-CoV-2 positive or recovered at delivery, and used historical and SARS-CoV-2 negative controls from two tertiary care hospitals. We compared the demographic, clinical, and hemodynamic variables between patients who were SARS-CoV-2 positive at delivery, those who were positive during pregnancy and recovered before delivery, and controls. Analyses were stratified by normotensive versus hypertensive status of the patients at delivery. RESULTS: We identified 22 SARS-CoV-2 positive, 73 SARS-CoV-2 recovered, and 1517 controls. The SARS-CoV-2 positive, and recovered pregnant patients, had on average 5.6 and 2.2 mmHg, respectively, higher post-spinal mean arterial pressures (MAPs) than control patients, adjusting for covariates. Additionally, the lowest post-spinal MAP was negatively correlated with the number of daysbetween the onset of COVID-19 symptoms and delivery in patients with hypertension (correlation -0.55, 95% CI -0.81 to -0.09). CONCLUSIONS: Patients with SARS-CoV-2 infection during pregnancy exhibit less spinal hypotension than non-infected patients. While the clinical significance of this finding is unknown, it points to important cardiovascular effects of the virus.

AUTS2 Regulates RNA Metabolism and Dentate Gyrus Development in Mice.

Human AUTS2 mutations are linked to a syndrome of intellectual disability, autistic features, epilepsy, and other neurological and somatic disorders. Although it is known that this unique gene is highly expressed in developing cerebral cortex, the molecular and developmental functions of AUTS2 protein remain unclear. Using proteomics methods to identify AUTS2 binding partners in neonatal mouse cerebral cortex, we found that AUTS2 associates with multiple proteins that regulate RNA transcription, splicing, localization, and stability. Furthermore, AUTS2-containing protein complexes isolated from cortical tissue bound specific RNA transcripts in RNA immunoprecipitation and sequencing assays. Deletion of all major functional isoforms of AUTS2 (full-length and C-terminal) by conditional excision of exon 15 caused breathing abnormalities and neonatal lethality when Auts2 was inactivated throughout the developing brain. Mice with limited inactivation of Auts2 in cerebral cortex survived but displayed abnormalities of cerebral cortex structure and function, including dentate gyrus hypoplasia with agenesis of hilar mossy neurons, and abnormal spiking activity on EEG. Also, RNA transcripts that normally associate with AUTS2 were dysregulated in mutant mice. Together, these findings indicate that AUTS2 regulates RNA metabolism and is essential for development of cerebral cortex, as well as subcortical breathing centers.

Intermediate Progenitor Cohorts Differentially Generate Cortical Layers and Require Tbr2 for Timely Acquisition of Neuronal Subtype Identity.

Intermediate progenitors (IPs) amplify the production of pyramidal neurons, but their role in selective genesis of cortical layers or neuronal subtypes remains unclear. Using genetic lineage tracing in mice, we find that IPs destined to produce upper cortical layers first appear early in corticogenesis, by embryonic day 11.5. During later corticogenesis, IP laminar fates are progressively limited to upper layers. We examined the role of Tbr2, an IP-specific transcription factor, in laminar fate regulation using Tbr2 conditional mutant mice. Upon Tbr2 inactivation, fewer neurons were produced by immediate differentiation and laminar fates were shifted upward. Genesis of subventricular mitoses was, however, not reduced in the context of a Tbr2-null cortex. Instead, neuronal and laminar differentiation were disrupted and delayed. Our findings indicate that upper-layer genesis depends on IPs from many stages of corticogenesis and that Tbr2 regulates the tempo of laminar fate implementation for all cortical layers.

Neuropathology of brain and spinal malformations in a case of monosomy 1p36.

Monosomy 1p36 is the most common subtelomeric chromosomal deletion linked to mental retardation and seizures. Neuroimaging studies suggest that monosomy 1p36 is associated with brain malformations including polymicrogyria and nodular heterotopia, but the histopathology of these lesions is unknown. Here we present postmortem neuropathological findings from a 10 year-old girl with monosomy 1p36, who died of respiratory complications. The findings included micrencephaly, periventricular nodular heterotopia in occipitotemporal lobes, cortical dysgenesis resembling polymicrogyria in dorsolateral frontal lobes, hippocampal malrotation, callosal hypoplasia, superiorly rotated cerebellum with small vermis, and lumbosacral hydromyelia. The abnormal cortex exhibited "festooned" (undulating) supragranular layers, but no significant fusion of the molecular layer. Deletion mapping demonstrated single copy loss of a contiguous 1p36 terminal region encompassing many important neurodevelopmental genes, among them four HES genes implicated in regulating neural stem cell differentiation, and TP73, a monoallelically expressed gene. Our results suggest that brain and spinal malformations in monosomy 1p36 may be more extensive than previously recognized, and may depend on the parental origin of deleted genes. More broadly, our results suggest that specific genetic disorders may cause distinct forms of cortical dysgenesis.

The protomap is propagated to cortical plate neurons through an Eomes-dependent intermediate map.

The cortical area map is initially patterned by transcription factor (TF) gradients in the neocortical primordium, which define a "protomap" in the embryonic ventricular zone (VZ). However, mechanisms that propagate regional identity from VZ progenitors to cortical plate (CP) neurons are unknown. Here we show that the VZ, subventricular zone (SVZ), and CP contain distinct molecular maps of regional identity, reflecting different gene expression gradients in radial glia progenitors, intermediate progenitors, and projection neurons, respectively. The "intermediate map" in the SVZ is modulated by Eomes (also known as Tbr2), a T-box TF. Eomes inactivation caused rostrocaudal shifts in SVZ and CP gene expression, with loss of corticospinal axons and gain of corticotectal projections. These findings suggest that cortical areas and connections are shaped by sequential maps of regional identity, propagated by the Pax6 → Eomes → Tbr1 TF cascade. In humans, PAX6, EOMES, and TBR1 have been linked to intellectual disability and autism.

Dystroglycan on radial glia end feet is required for pial basement membrane integrity and columnar organization of the developing cerebral cortex.

Interactions between the embryonic pial basement membrane (PBM) and radial glia (RG) are essential for morphogenesis of the cerebral cortex because disrupted interactions cause cobblestone malformations. To elucidate the role of dystroglycan (DG) in PBM-RG interactions, we studied the expression of DG protein and Dag1 mRNA (which encodes DG protein) in developing cerebral cortex and analyzed cortical phenotypes in Dag1 CNS conditional mutant mice. In normal embryonic cortex, Dag1 mRNA was expressed in the ventricular zone, which contains RG nuclei, whereas DG protein was expressed at the cortical surface on RG end feet. Breaches of PBM continuity appeared during early neurogenesis in Dag1 mutants. Diverse cellular elements streamed through the breaches to form leptomeningeal heterotopia that were confluent with the underlying residual cortical plate and contained variably truncated RG fibers, many types of cortical neurons, and radial and intermediate progenitor cells. Nevertheless, layer-specific molecular expression seemed normal in heterotopic neurons, and axons projected to appropriate targets. Dendrites, however, were excessively tortuous and lacked radial orientation. These findings indicate that DG is required on RG end feet to maintain PBM integrity and suggest that cobblestone malformations involve disturbances of RG structure, progenitor distribution, and dendrite orientation, in addition to neuronal "overmigration."

Multiple autism-like behaviors in a novel transgenic mouse model.

Autism spectrum disorder (ASD) diagnoses are behaviorally based with no defined universal biomarkers, occur at a 1:110 ratio in the population, and predominantly affect males compared to females at approximately a 4:1 ratio. One approach to investigate and identify causes of ASD is to use organisms that display abnormal behavioral responses that model ASD-related impairments. This study describes a novel transgenic mouse, MALTT, which was generated using a forward genetics approach. It was determined that the transgene integrated within a non-coding region on the X chromosome. The MALTT line exhibited a complete repertoire of ASD-like behavioral deficits in all three domains required for an ASD diagnosis: reciprocal social interaction, communication, and repetitive or inflexible behaviors. Specifically, MALTT male mice showed deficits in social interaction and interest, abnormalities in pup and juvenile ultrasonic vocalization communications, and exhibited a repetitive stereotypy. Abnormalities were also observed in the domain of sensory function, a secondary phenotype prevalently associated with ASD. Mapping and expression studies suggested that the Fam46 gene family may be linked to the observed ASD-related behaviors. The MALTT line provides a unique genetic model for examining the underlying biological mechanisms involved in ASD-related behaviors.

Genetic variation in adiponectin (ADIPOQ) and the type 1 receptor (ADIPOR1), obesity and prostate cancer in African Americans.

Adiponectin is a protein derived from adipose tissue suspected to have an important role in prostate carcinogenesis. Variants in the adiponectin gene (ADIPOQ) and its type 1 receptor (ADIPOR1) have been recently linked to risk of both breast and colorectal cancer. Therefore, we set out to examine the relationship between polymorphisms in these genes, obesity and prostate cancer in study of African-American men. Ten single-nucleotide polymorphisms (SNPs) in ADIPOQ and ADIPOR1 were genotyped in DNA samples from 131 African-American prostate cancer cases and 344 controls participating in the Flint Men's Health Study. Logistic regression was then used to estimate their association with prostate cancer and obesity. While no significant associations were detected between any of the tested SNPs and prostate cancer, the rs1501299 SNP in ADIPOQ was significantly associated with body mass (P=0.03). Genetic variation in ADIPOQ and ADIPOR1 did not predict risk of prostate cancer in this study of African-American men. However, the rs1501299 SNP in ADIPOQ was associated with obesity. Further investigation is warranted to determine if racial differences exist in the influence of the adiponectin pathway on prostate cancer risk.

Tbr1 regulates regional and laminar identity of postmitotic neurons in developing neocortex.

Areas and layers of the cerebral cortex are specified by genetic programs that are initiated in progenitor cells and then, implemented in postmitotic neurons. Here, we report that Tbr1, a transcription factor expressed in postmitotic projection neurons, exerts positive and negative control over both regional (areal) and laminar identity. Tbr1 null mice exhibited profound defects of frontal cortex and layer 6 differentiation, as indicated by down-regulation of gene-expression markers such as Bcl6 and Cdh9. Conversely, genes that implement caudal cortex and layer 5 identity, such as Bhlhb5 and Fezf2, were up-regulated in Tbr1 mutants. Tbr1 implements frontal identity in part by direct promoter binding and activation of Auts2, a frontal cortex gene implicated in autism. Tbr1 regulates laminar identity in part by downstream activation or maintenance of Sox5, an important transcription factor controlling neuronal migration and corticofugal axon projections. Similar to Sox5 mutants, Tbr1 mutants exhibit ectopic axon projections to the hypothalamus and cerebral peduncle. Together, our findings show that Tbr1 coordinately regulates regional and laminar identity of postmitotic cortical neurons.

Absence of truncating BRIP1 mutations in chromosome 17q-linked hereditary prostate cancer families.

BACKGROUND: In a genome-wide scan (GWS) of 175 multiplex prostate cancer (PCa) families from the University of Michigan Prostate Cancer Genetics Project (PCGP), linkage was observed to markers on chromosome 17q21-24, a region that includes two breast cancer susceptibility genes, BRCA1 and BRIP1. BRIP1 is a Fanconi anaemia gene (FANCJ) that interacts with the BRCT domain of BRCA1 and has a role in DNA damage repair. Protein truncating mutations in BRIP1 have been identified in hereditary breast and ovarian cancer families, and a recent report suggested that a recurrent truncating mutation (R798X) may have a role in PCa susceptibility. METHODS: We examined the role of BRIP1 mutations in hereditary PCa through sequence analysis of 94 individuals from PCGP families showing linkage to 17q. RESULTS: A total of 24 single-nucleotide polymorphisms, including 7 missense variants but no protein truncating mutations, were observed. CONCLUSION: The data presented here suggest that BRIP1 truncating mutations are uncommon in PCa cases and do not account for the linkage to chromosome 17q observed in our GWS. Additional investigation is needed to determine the significance, if any, of the observed BRIP1 missense variants in hereditary PCa.

(R)-3-hydroxyhexan-2-one is a major pheromone component of Anelaphus inflaticollis (Coleoptera: Cerambycidae).

We report the identification and field bioassays of a major component of the male-produced aggregation pheromone of Anelaphus inflaticollis Chemsak, an uncommon desert cerambycine beetle. Male A. inflaticollis produced a sex-specific blend of components that included (R)-3-hydroxyhexan-2-one, (S)-2-hydroxyhexan-3-one, 2,3-hexanedione, and (2R,3R)- and (2R,3S)-2,3-hexanediols. Field trials with baited bucket traps determined that the reconstructed synthetic pheromone blend and (R)-3-hydroxyhexan-2-one alone attracted adult A. inflaticollis of both sexes, with significantly more beetles being attracted to the blend. We conclude that (R)-3-hydroxyhexan-2-one is a major pheromone component of A. inflaticollis, and our results suggest that one or more of the minor components may further increase attraction of conspecifics. Scanning electron microscopy showed that male A. inflaticollis have pores on the prothorax that are consistent in structure with sex-specific pheromone gland pores in related species. Males also displayed stereotyped calling behavior similar to that observed in other cerambycine species. This study represents the first report of volatile pheromones for a cerambycine species in the tribe Elaphidiini.

Intermediate neuronal progenitors (basal progenitors) produce pyramidal-projection neurons for all layers of cerebral cortex.

The developing cerebral cortex contains apical and basal types of neurogenic progenitor cells. Here, we investigated the cellular properties and neurogenic output of basal progenitors, also called intermediate neuronal progenitors (INPs). We found that basal mitoses expressing transcription factor Tbr2 (an INP marker) were present throughout corticogenesis, from embryonic day 10.5 through birth. Postnatally, Tbr2(+) progenitors were present in the dentate gyrus, subventricular zone (SVZ), and posterior periventricle (pPV). Two morphological subtypes of INPs were distinguished in the embryonic cortex, "short radial" in the ventricular zone (VZ) and multipolar in the SVZ, probably corresponding to molecularly defined INP subtypes. Unexpectedly, many short radial INPs appeared to contact the apical (ventricular) surface and some divided there. Time-lapse video microscopy suggested that apical INP divisions produced daughter INPs. Analysis of neurogenic divisions (Tis21-green fluorescent protein [GFP](+)) indicated that INPs may produce the majority of projection neurons for preplate, deep, and superficial layers. Conversely, proliferative INP divisions (Tis21-GFP(-)) increased from early to middle corticogenesis, concomitant with SVZ growth. Our findings support the hypothesis that regulated amplification of INPs may be an important factor controlling the balance of neurogenesis among different cortical layers.

Organotypic slice culture of embryonic brain tissue.

INTRODUCTIONThis protocol describes how to dissect, assemble, and cultivate mouse embryonic (E) brain tissue from age E11.5 to E18.5 (days) for organotypic slice culture. These preparations can be used for a variety of assays and studies including coculture of different brain regions, cell migration assays, axon guidance assays, and DNA electroporation experiments. During electroporation, an electric current is applied to the surface of a specific target area of the brain slice in order to open holes in the plasma membrane and introduce a plasmid of coding DNA. The floating slice-on-membrane construct helps to preserve the structural integrity of the brain slices, while maintaining easy experimental access and optimal viability. Experiments can be monitored in living slices (e.g., with confocal imaging), and further studies can be completed using slices that have been fixed and cryosectioned at the end of the experiment. Any region of embryonic brain or spinal tissue can be used in this protocol.

Unipolar brush cells of the cerebellum are produced in the rhombic lip and migrate through developing white matter.

Unipolar brush cells (UBCs) are glutamatergic interneurons in the cerebellar cortex and dorsal cochlear nucleus. We studied the development of UBCs, using transcription factor Tbr2/Eomes as a marker for UBCs and their progenitors in embryonic and postnatal mouse cerebellum. Tbr2+ UBCs appeared to migrate out of the upper rhombic lip via two cellular streams: a dorsal pathway into developing cerebellar white matter, where the migrating cells dispersed widely before entering the internal granular layer, and a rostral pathway along the cerebellar ventricular zone toward the brainstem. Ablation of the rhombic lip in organotypic slice cultures substantially reduced the production of Tbr2+ UBCs. In coculture experiments, Tbr2+ UBCs migrated from rhombic lip explants directly into the developing white matter of adjacent cerebellar slices. The origin of Tbr2+ UBCs was confirmed by colocalization with beta-galactosidase expressed from the Math1 locus, a molecular marker of rhombic lip lineages. Moreover, the production of Tbr2+ UBCs was Math1 dependent, as Tbr2+ UBCs were severely reduced in Math1-null cerebellum. In reeler mutant mice, Tbr2+ UBCs accumulated near the rhombic lip, consistent with impaired migration through developing white matter. Our results suggest that UBCs arise from the rhombic lip and migrate via novel pathways to their final destinations in the cerebellum and dorsal cochlear nucleus. Our findings support a model of cerebellar neurogenesis, in which glutamatergic and GABAergic neurons are produced from separate progenitor pools located mainly in the rhombic lip and the cerebellar ventricular zone, respectively.

Prohibitin mutations are uncommon in prostate cancer families linked to chromosome 17q.

BACKGROUND: Linkage studies have provided evidence for a prostate cancer susceptibility locus on chromosome 17q. The mitochondrial protein prohibitin (PHB) is a plausible candidate gene based on its chromosomal location (17q21) and known function. METHODS: All coding regions and intron/exon junctions of the PHB gene were sequenced in 32 men from families participating in the University of Michigan Prostate Cancer Genetics Project that demonstrated evidence of linkage to 17q markers. RESULTS: Although a number of nucleotide variants were identified, no coding region substitutions were identified in any of the 32 men with prostate cancer from 32 unrelated multiplex prostate cancer families. CONCLUSIONS: PHB mutations do not appear to account for the linkage signal on 17q21-22 detected in PCGP families. Fine mapping of this region is in progress to refine the candidate region and highlight additional candidate prostate cancer susceptibility genes for sequence analysis.

Transcription factors in glutamatergic neurogenesis: conserved programs in neocortex, cerebellum, and adult hippocampus.

Glutamatergic, pyramidal-projection neurons are produced in the embryonic cerebral cortex by a series of genetically programmed fate choices, implemented in large part by developmental transcription factors. Our work has focused on Pax6, Tbr2/Eomes, NeuroD, and Tbr1, which are expressed sequentially during the neurogenesis of pyramidal-projection neurons. Recently, we have found that the same transcription factors are expressed, in the same order, during glutamatergic neurogenesis in the adult dentate gyrus, and (with modifications) in the developing cerebellum. While the precise functional significance of this transcription factor expression sequence is unknown, its common appearance in embryonic and adult neurogenesis, and in different brain regions, suggests it is part of a conserved genetic program that specifies general properties of glutamatergic neurons in these regions. Subtypes of glutamatergic neurons (e.g., layer-specific fates in the cortex) are further determined by combinations of transcription factors, superimposed on general sequential programs. These new perspectives on neurogenesis add to the conceptual framework for strategies to engineer neural stem cells for the repair of specific brain circuits.

Development of the deep cerebellar nuclei: transcription factors and cell migration from the rhombic lip.

The deep cerebellar nuclei (DCN) are the main output centers of the cerebellum, but little is known about their development. Using transcription factors as cell type-specific markers, we found that DCN neurons in mice are produced in the rhombic lip and migrate rostrally in a subpial stream to the nuclear transitory zone (NTZ). The rhombic lip-derived cells express transcription factors Pax6, Tbr2, and Tbr1 sequentially as they enter the NTZ. A subset of rhombic lip-derived cells also express reelin, a key regulator of Purkinje cell migrations. In organotypic slice cultures, the rhombic lip was necessary and sufficient to produce cells that migrate in the subpial stream, enter the NTZ, and express Pax6, Tbr2, Tbr1, and reelin. In later stages of development, the subpial stream is replaced by the external granular layer, and the NTZ organizes into distinct DCN nuclei. Tbr1 expression persists to adulthood in a subset of medial DCN projection neurons. In reeler mutant mice, which have a severe cerebellar malformation, rhombic lip-derived cells migrated to the NTZ, despite reelin deficiency. Studies in Tbr1 mutant mice suggested that Tbr1 plays a role in DCN morphogenesis but is not required for reelin expression, glutamatergic differentiation, or the initial formation of efferent axon pathways. Our findings reveal underlying similarities in the transcriptional programs for glutamatergic neuron production in the DCN and the cerebral cortex, and they support a model of cerebellar neurogenesis in which glutamatergic and GABAergic neurons are produced from separate progenitor compartments.

Polymorphisms in the prostate-specific antigen gene promoter do not predict serum prostate-specific antigen levels in African-American men.

A major problem with the use of serum prostate-specific antigen (PSA) in predicting prostate cancer risk is the considerable variability of such measurements. Cramer et al. identified a set of single-nucleotide polymorphisms (SNPs) in the upstream regulatory region of the PSA gene that were each associated with increased promoter activity and serum PSA, further suggesting that genotyping these SNPs could be useful in improving the predictive value of PSA screening. In order to replicate this finding, DNA samples from 475 African-American men were genotyped for the same SNPs and no association was observed with either serum PSA level or prostate cancer diagnosis.

Pax6, Tbr2, and Tbr1 are expressed sequentially by radial glia, intermediate progenitor cells, and postmitotic neurons in developing neocortex.

The developing neocortex contains two types of progenitor cells for glutamatergic, pyramidal-projection neurons. The first type, radial glia, produce neurons and glia, divide at the ventricular surface, and express Pax6, a homeodomain transcription factor. The second type, intermediate progenitor cells, are derived from radial glia, produce only neurons, and divide away from the ventricular surface. Here we show that the transition from radial glia to intermediate progenitor cell is associated with upregulation of Tbr2, a T-domain transcription factor, and downregulation of Pax6. Accordingly, Tbr2 expression in progenitor compartments (the subventricular zone and ventricular zone) rises and falls with cortical plate neurogenesis. The subsequent transition from intermediate progenitor cell to postmitotic neuron is marked by downregulation of Tbr2 and upregulation of Tbr1, another T-domain transcription factor. These findings delineate the transcription factor sequence Pax6 --> Tbr2 --> Tbr1 in the differentiation of radial glia --> intermediate progenitor cell --> postmitotic projection neuron. This transcription factor sequence is modified in preplate neurons, in which Tbr2 is transiently coexpressed with Tbr1, and in the direct differentiation pathway from radial glia --> postmitotic projection neuron, in which Tbr2 is expressed briefly or not at all.

Beyond laminar fate: toward a molecular classification of cortical projection/pyramidal neurons.

Cortical projection neurons exhibit diverse morphological, physiological, and molecular phenotypes, but it is unknown how many distinct types exist. Many projection cell phenotypes are associated with laminar fate (radial position), but each layer may also contain multiple types of projection cells. We have investigated two hypotheses: (1) that different projection cell types exhibit characteristic molecular expression profiles and (2) that laminar fates are determined primarily by molecular phenotype. We found that several transcription factors were differentially expressed by projection neurons, even within the same layer: Otx1 and Er81, for example, were expressed by different neurons in layer 5. Retrograde tracing showed that Er81 was expressed in corticospinal and corticocortical neurons. In contrast, Otx1 has been detected only in corticobulbar neurons [Weimann et al., Neuron 1999;24:819-831]. Birthdating demonstrated that different molecularly defined types were produced sequentially, in overlapping waves. Cells adopted laminar fates characteristic of their molecular phenotypes, regardless of cell birthday. Molecular markers also revealed the locations of different projection cell types in the malformed cortex of reeler mice. These studies suggest that molecular profiles can be used advantageously for classifying cortical projection cells, for analyzing their neurogenesis and fate specification, and for evaluating cortical malformations.