Investigations on formation and specification of neural precursor cells in the central nervous system of the Drosophila melanogaster embryo

Investigations on formation and specification of neural precursor cells in the central nervous system of the Drosophila melanogaster embryoSpecification of a unique cell fate during development of a multicellular organism often is a function of its position. The Drosophila central nervous system (CNS) provides an ideal system to dissect signalling events during development that lead to cell specific patterns. Different cell types in the CNS are formed from a relatively few precursor cells, the neuroblasts (NBs), which delaminate from the neurogenic region of the ectoderm. The delamination occurs in five waves, S1-S5, finally leading to a subepidermal layer consisting of about 30 NBs, each with a unique identity, arranged in a stereotyped spatial pattern in each hemisegment. This information depends on several factors such as the concentrations of various morphogens, cell-cell interactions and long range signals present at the position and time of its birth. The early NBs, delaminating during S1 and S2, form an orthogonal array of four rows (2/3,4,5,6/7) and three columns (medial, intermediate, and lateral) . However, the three column and four row-arrangement pattern is only transitory during early stages of neurogenesis which is obscured by late emerging (S3-S5) neuroblasts (Doe and Goodman, 1985; Goodman and Doe, 1993). Therefore the aim of my study has been to identify novel genes which play a role in the formation or specification of late delaminating NBs.In this study the gene anterior open or yan was picked up in a genetic screen to identity novel and yet unidentified genes in the process of late neuroblast formation and specification. I have shown that the gene yan is responsible for maintaining the cells of the neuroectoderm in an undifferentiated state by interfering with the Notch signalling mechanism. Secondly, I have studied the function and interactions of segment polarity genes within a certain neuroectodermal region, namely the engrailed (en) expressing domain, with regard to the fate specification of a set of late neuroblasts, namely NB 6-4 and NB 7-3. I have dissected the regulatory interaction of the segment polarity genes wingless (wg), hedgehog (hh) and engrailed (en) as they maintain each other’s expression to show that En is a prerequisite for neurogenesis and show that the interplay of the segmentation genes naked (nkd) and gooseberry (gsb), both of which are targets of wingless (wg) activity, leads to differential commitment of NB 7-3 and NB 6-4 cell fate. I have shown that in the absence of either nkd or gsb one NB fate is replaced by the other. However, the temporal sequence of delamination is maintained, suggesting that formation and specification of these two NBs are under independent control.

Hox genes and the regulation of programmed cell death in the embryonic central nervous system of Drosophila melanogaster

This study deals with the function and regulation of programmed cell death, or apoptosis, in the development of the embryonic central nervous system of Drosophila melanogaster. The first part provides a description of apoptosis-deficient embryos, which showed that preventing apoptosis does not cause gross morphological defects in the CNS, as it appears well organized despite the presence of too many cells. An analysis of the incidence and pattern of apoptosis over the course of development discloses a partly very orderly pattern suggesting tight spatio-temporal control, but also reveals random apoptotic cells, which suggests a certain amount of plasticity in the embryo. This analysis also allowed precise identification of some of the dying neural cells in the embryo, and establishment of single cell models for studying regulation of segment-specific apoptosis in the embryonic CNS. In the second part of the work, further investigations into mechanisms controlling segment-specific apoptosis revealed the involvement of two Hox genes, Antennapedia (Antp) and Ultrabithorax (Ubx), in this process. Hox genes control the formation of segment-specific structures in their domains of expression, but also regulate organ and tissue morphogenesis. The study presented here shows that Antp and Ubx play antagonistic roles in motoneuron survival in the embryo. Ubx expression in the CNS is strongly upregulated at a late point in development, when most cells have begun to differentiate. This upregulation shortly precedes Ubx-dependent, segment-specific apoptosis of two differentiated motoneurons. It could further be demonstrated that Antp is required for proper development of the NB7-3 lineage and for survival of the NB7-3 motoneuron in the anterior thoracic segments. In segments where Antp and Ubx expression overlaps, Ubx counteracts the anti-apoptotic function of Antp, resulting in cell death. Thus, these two Hox genes play opposing roles in the survival of differentiated neurons in the late developing nervous system. They thereby contribute to establishment of correct connections between outward-projecting neurons and their targets, which is crucial for the assembly of functional neural circuits, as these have to fulfill region-specific locomotion and sensory requirements along the antero-posterior body axis.

Automatic risk evaluation in elderly patients based on Autonomic Nervous System assessment

Dysfunction of Autonomic Nervous System (ANS) is a typical feature of chronic heart failure and other cardiovascular disease. As a simple non-invasive technology, heart rate variability (HRV) analysis provides reliable information on autonomic modulation of heart rate. The aim of this thesis was to research and develop automatic methods based on ANS assessment for evaluation of risk in cardiac patients. Several features selection and machine learning algorithms have been combined to achieve the goals. Automatic assessment of disease severity in Congestive Heart Failure (CHF) patients: a completely automatic method, based on long-term HRV was proposed in order to automatically assess the severity of CHF, achieving a sensitivity rate of 93% and a specificity rate of 64% in discriminating severe versus mild patients. Automatic identification of hypertensive patients at high risk of vascular events: a completely automatic system was proposed in order to identify hypertensive patients at higher risk to develop vascular events in the 12 months following the electrocardiographic recordings, achieving a sensitivity rate of 71% and a specificity rate of 86% in identifying high-risk subjects among hypertensive patients. Automatic identification of hypertensive patients with history of fall: it was explored whether an automatic identification of fallers among hypertensive patients based on HRV was feasible. The results obtained in this thesis could have implications both in clinical practice and in clinical research. The system has been designed and developed in order to be clinically feasible. Moreover, since 5-minute ECG recording is inexpensive, easy to assess, and non-invasive, future research will focus on the clinical applicability of the system as a screening tool in non-specialized ambulatories, in order to identify high-risk patients to be shortlisted for more complex investigations.

Inherited cavernous malformations of the central nervous system: clinical and genetic features in 19 Swiss families

Cavernous malformations (CCMs) are benign, well-circumscribed, and mulberry-like vascular malformations that may be found in the central nervous system in up to 0.5% of the population. Cavernous malformations can be sporadic or inherited. The common symptoms are epilepsy, hemorrhages, focal neurological deficits, and headaches. However, CCMs are often asymptomatic. The familiar form is associated with three gene loci, namely 7q21-q22 (CCM1), 7p13-p15 (CCM2), and 3q25.2-q27 (CCM3) and is inherited as an autosomal dominant trait with incomplete penetrance. The CCM genes are identified as Krit 1 (CCM1), MGC4607 (CCM2), and PDCD10 (CCM3). Here, we present the clinical and genetic features of CCMs in 19 Swiss families. Furthermore, surgical aspects in such families are also discussed.

Rapid assessment of internodal myelin integrity in central nervous system tissue

Monitoring pathology/regeneration in experimental models of de-/remyelination requires an accurate measure not only of functional changes but also of the amount of myelin. We tested whether X-ray diffraction (XRD), which measures periodicity in unfixed myelin, can assess the structural integrity of myelin in fixed tissue. From laboratories involved in spinal cord injury research and in studying the aging primate brain, we solicited "blind" samples and used an electronic detector to record rapidly the diffraction patterns (30 min each pattern) from them. We assessed myelin integrity by measuring its periodicity and relative amount. Fixation of tissue itself introduced +/-10% variation in periodicity and +/-40% variation in relative amount of myelin. For samples having the most native-like periods, the relative amounts of myelin detected allowed distinctions to be made between normal and demyelinating segments, between motor and sensory tracts within the spinal cord, and between aged and young primate CNS. Different periodicities also allowed distinctions to be made between samples from spinal cord and nerve roots and between well-fixed and poorly fixed samples. Our findings suggest that, in addition to evaluating the effectiveness of different fixatives, XRD could also be used as a robust and rapid technique for quantitating the relative amount of myelin among spinal cords and other CNS tissue samples from experimental models of de- and remyelination.

Meningial perineurioma: a benign peripheral nerve sheath tumor in a previously unrecognized central nervous system location, mimicking meningioma

Perineurioma is an uncommon, mostly benign, spindle-cell tumor of peripheral nerve sheath origin with a predilection for the soft tissues. Although increasing awareness points to the sites of involvement by perineurioma possibly being as ubiquitous as those frequented by schwannian tumors, only one intracerebral example has been described to date. We report on a surgically resected perineurioma of the falx cerebri in an 86-year-old woman. Preoperative imaging showed an enhancing extraaxial mass of 6 cm × 5.7 cm × 3.7 cm. Histologically, the tumor consisted of a proliferation of spindle cells interwoven by a lattice of basal lamina. Alongside a prevailing soft tissue perineurioma pattern, sclerosing and reticular areas were seen as well. Tumor cells coexpressed EMA and GLUT-1, and a minority immunoreacted for smooth muscle actin. Pericellular basal lamina was decorated with collagen type IV. No staining for S100 protein was detected. Mitotic activity was virtually absent, and the MIB1 labeling index averaged 2%. Ultrastructural examination revealed abundant pinocytotic vesicles within and conspicuous tight junctions between slender cytoplasmic processes which, in turn, were encased by discontinuous basal lamina. FISH analysis confirmed loss of at least part of one chromosome 22q. This observation calls attention to perineurioma as a novel item in the repertoire of low-grade meningial spindle cell neoplasms, in the differential diagnostic context of which it is apt to being misconstrued as either meningioma, solitary fibrous tumor, or neurofibroma. Confusion with the latter bears the risk of overgrading innocuous features of perineurioma as criteria for malignancy.

Atypical presentation of Behçet's disease with central nervous system involvement successfully treated with infliximab

Central nervous system involvement is a rare and serious complication of Behçet's disease (BD). Herein, we describe a patient with an atypical central lesion, who experienced progressive hypesthesia of the right arm and sensory loss of the trigeminal nerve together with intense headache. A repeated biopsy was necessary to conclusively establish the diagnosis of BD. Therapy with infusions of infliximab led to a remarkable full remission. TNFα-blocking therapy was successfully replaced by azathioprine. The present well-illustrated case demonstrates the difficulty of establishing the diagnosis of BD with central nervous system involvement, the dramatic benefit of short given TNF-α-blocking agent, and the long-term remission with azathioprin.

Ultrabithorax Confers Spatial Identity in a Context-Specific Manner in the Drosophila Postembryonic Ventral Nervous System

In holometabolous insects such as Drosophila melanogaster, neuroblasts produce an initial population of diverse neurons during embryogenesis and a much larger set of adult-specific neurons during larval life. In the ventral CNS, many of these secondary neuronal lineages differ significantly from one body segment to another, suggesting a role for anteroposterior patterning genes. Here we systematically characterize the expression pattern and function of the Hox gene Ultrabithorax (Ubx) in all 25 postembryonic lineages. We find that Ubx is expressed in a segment-, lineage-, and hemilineage-specific manner in the thoracic and anterior abdominal segments. When Ubx is removed from neuroblasts via mitotic recombination, neurons in these segments exhibit the morphologies and survival patterns of their anterior thoracic counterparts. Conversely, when Ubx is ectopically expressed in anterior thoracic segments, neurons exhibit complementary posterior transformation phenotypes. Our findings demonstrate that Ubx plays a critical role in conferring segment-appropriate morphology and survival on individual neurons in the adult-specific ventral CNS. Moreover, while always conferring spatial identity in some sense, Ubx has been co-opted during evolution for distinct and even opposite functions in different neuronal hemilineages.

Tight junctions in brain barriers during central nervous system inflammation

Homeostasis within the central nervous system (CNS) is a prerequisite to elicit proper neuronal function. The CNS is tightly sealed from the changeable milieu of the blood stream by the blood-brain barrier (BBB) and the blood-cerebrospinal fluid (CSF) barrier (BCSFB). Whereas the BBB is established by specialized endothelial cells of CNS microvessels, the BCSFB is formed by the epithelial cells of the choroid plexus. Both constitute physical barriers by a complex network of tight junctions (TJs) between adjacent cells. During many CNS inflammatory disorders, such as multiple sclerosis, human immunodeficiency virus infection, or Alzheimer's disease, production of pro-inflammatory cytokines, matrix metalloproteases, and reactive oxygen species are responsible for alterations of CNS barriers. Barrier dysfunction can contribute to neurological disorders in a passive way by vascular leakage of blood-borne molecules into the CNS and in an active way by guiding the migration of inflammatory cells into the CNS. Both ways may directly be linked to alterations in molecular composition, function, and dynamics of the TJ proteins. This review summarizes current knowledge on the cellular and molecular aspects of the functional and dysfunctional TJ complexes at the BBB and the BCSFB, with a particular emphasis on CNS inflammation and the role of reactive oxygen species.

Training-related modulations of the autonomic nervous system in endurance athletes: is female gender cardioprotective?

The risk of sudden death is increased in athletes with a male predominance. Regular physical activity increases vagal tone, and may protect against exercise-induced ventricular arrhythmias. We investigated training-related modulations of the autonomic nervous system in female and male endurance athletes. Runners of a 10-mile race were invited. Of 873 applicants, 68 female and 70 male athletes were randomly selected and stratified according to their average weekly training hours in a low (≤4 h) and high (>4 h) volume training group. Analysis of heart rate variability was performed over 24 h. Spectral components (high frequency [HF] and low frequency [LF] power in normalized units) were analyzed for hourly 5 min segments and averaged for day- and nighttime. One hundred and fourteen athletes (50 % female, mean age 42 ± 7 years) were included. No significant gender difference was observed for training volume and 10-mile race time. Over the 24-h period, female athletes exhibited a higher HF and lower LF power for each hourly time-point. Female gender and endurance training hours were independent predictors of a higher HF and lower LF power. In female athletes, higher training hours were associated with a higher HF and lower LF power during nighttime. In male athletes, the same was true during daytime. In conclusion, female and male athletes showed a different circadian pattern of the training-related increase in markers of vagal tone. For a comparable amount of training volume, female athletes maintained their higher markers of vagal tone, possibly indicating a superior protection against exercise-induced ventricular arrhythmias.

The overexpression of homeotic complex gene Ultrabithorax in the post-embryonic neuronal lineages of the ventral nervous system in Drosophila melanogaster

This study uses a molecular technique called MARCM (Mosaic Analysis with a Repressible Cell Marker) to label neuronal lineages that overexpress the Hox gene Ultrabithorax (Ubx) in an unlabeled, wild type background. The results indicate that the overexpression of Ubx is sufficient to transform more anterior neuronal lineages to themorphology of their more posterior counterparts. The data presented here begin to elucidate the role that the Hox genes have in shaping segment-specific neural connections in the post-embryonic ventral nervous system.

Dosing algorithm to target a predefined AUC in patients with primary central nervous system lymphoma receiving high dose methotrexate

There is no consensus regarding optimal dosing of high dose methotrexate (HDMTX) in patients with primary CNS lymphoma. Our aim was to develop a convenient dosing algorithm to target AUC(MTX) in the range between 1000 and 1100 µmol l(-1) h.

The Role of Antennapedia in Anteroposterior Patterning of the Drosophila Melanogaster Ventral Nervous System

A major unresolved question in developmental neurobiology is how the nervous system is adapted to the specific needs of the organism at different life stages. In the holometabolous insect Drosophila melanogaster, the larval ventral nervous system (VNS) is comprised of similar repeating segments, as opposed to the adult VNS, which varies greatly from segment to segment both in number and types of neurons. The adult-specific neurons of each segment are generated by 25 distinct types of neuronal progenitor cells called neuroblasts (NBs) that appear in a stereotyped array (Truman et al., 2004). Each NB divides repeatedly to produce a distinct set of daughter cells termed a lineage, which is bilaterally symmetric but present to varying degrees in each segment. These daughter cells can be distinguished by their position within the nervous system as well as by their axonal projections. Each of the 25 NBs produces neurons; if both daughter cells are present in a lineage then both sibling populations survived, whereas if only one projection is seen cell death occurred, leaving a hemilineage (half lineage). In some lineages, the same sibling type survives in all segments in which the lineage appears, but in others, the surviving sibling type varies across segments, resulting in a different morphology for the same lineage in different segments. How are these differences in survival and morphology controlled? The Hox genes provide positional information for developing structures along the anterior-posterior (AP) axis of animals. They encode transcription factors, thereby controlling the activity of genes down stream. In the postembryonic VNS, each NB lineage features its own characteristic expression pattern of Hox genes Antp and Ubx, which can vary from segment-to-segment, and can thereby cause variation in the number of neural cells and axonal projections that survive. This study defines the wild-type expression pattern of Antp and elucidates the role of Antp in gain of function studies. These studies are possible due to the MARCM (Mosaic Analysis with a Repressible Cell Marker) method, which allows the genetically manipulated cells to be specifically labeled in an otherwise normal, unlabeled organism. The results indicate that Antp is expressed in a segment-, lineage-, and hemilineage-specific manner. Antp is sufficient for both anterior and posterior transformations of particular lineages, including promotion of cell death and/or survival as well as axon guidance.

The anatomical and cellular basis of immune surveillance in the central nervous system

The central nervous system (CNS) comprises the brain, spinal cord, optic nerves and retina, and contains post-mitotic, delicate cells. As the rigid coverings of the CNS render swelling dangerous and destructive, inflammatory reactions must be carefully controlled in CNS tissues. Nevertheless, effector immune responses that protect the host during CNS infection still occur in the CNS. Here, we describe the anatomical and cellular basis of immune surveillance in the CNS, and explain how this shapes the unique immunology of these tissues. The Review focuses principally on insights gained from the study of autoimmune responses in the CNS and to a lesser extent on models of infectious disease. Furthermore, we propose a new model to explain how antigen-specific T cell responses occur in the CNS.