An Evolutionarily Conserved Region in the Second lntron of the Human Nestin Gene Directs Gene Exmession to CNS Progenitor Cells and to Early Neural Ciest Cells

Central nervous system (CNS) progenitor cells transiently proliferate in the embryonic neural tube and give rise to neurons and glial cells. A characteristic feature of the CNS progenitor cells is expression of the intermediate filament nestin and it was previously shown that the rat nestin second intron functions as an enhancer, directing gene expression to CNS progenitor cells. In this report we characterize the nestin enhancer in further detail. Cloning and sequence analysis of the rat and human nestin second introns revealed local domains of high sequence similarity in the 3' portion of the introns. Transgenic mice were generated with the most conserved 714 bp in the 3' portion of the intron, or with the complete, 1852 bp, human second intron, coupled to the reporter gene lacZ. The two constructs gave a very similar nestin-like expression pattern, indicating that the important control elements reside in the 714 bp element. Expression was observed starting in embryonic day (E)7.5 neural plate, and at E10.5 CNS progenitor cells throughout the neural tube expressed lacZ. At E12.5, lacZ expression was more restricted and confined to proliferating regions in the neural tube. An interesting difference, compared to the rat nestin second intron, was that the human intron at E10.5 mediated lacZ expression also in early migrating neural crest cells, which is a site of endogenous nestin expression. In conclusion, these data show that a relatively short, evolutionarily conserved region is sufficient to control gene expression in CNS progenitor cells, but that the same region differs between rodents and primates in its capacity to control expression in neural crest cells.     

Bulbectomy Enhances Neurogenesis and Cell Turnover of Primary Olfactory Neurons But Does Not Abolish Carnosine Expression

Primary olfactory neurons located in the olfactory neuroepithelium project to the ipsilateral olfactory bulb and undergo a continuous process of neurogenesis and differentiation. We describe, in the adult rat, the kinetics of proliferation, differentiation and survival of primary olfactory neurons either in the presence or absence of their target, the olfactory bulb. The experimental design included unilateral bulbectomy, coupled with a single bromodeoxyuridine pulse 35 days after surgery. The rate of proliferation and survival of olfactory neurons was then examined by immunohistochemistry for bromodeoxyuridine, and the differentiation status by in situ hybridization for calmodulin messenger RNA in immature and mature olfactory neurons and immunohistochemistry for the dipeptide carnosine in mature olfactory neurons. We show that primary olfactory neurons can synthesize carnosine in the absence of the olfactory bulb. However, the number of carnosine-immunopositive neurons in the absence of their target is dramatically reduced to less than one-fourth, whereas the number of olfactory neurons expressing calmodulin messenger RNA is only slightly reduced. The numeric reduction of camosine-positive neurons in the target-deprived neuroepithelium is correlated with a dramatic reduction in the survival rate of olfactory neurons, since newly generated olfactory neurons are completely lost 35 days after the bromodeoxyuridine pulse. In contrast, in the normal olfactory neuroepithelium almost one-third of newly generated olfactory neurons survive 35 days after the bromodeoxyuridine pulse. On the whole, these data indicate that most of the primary olfactory neurons have a short lifespan but that once they have connected with the olfactory bulb they may persist longer, and suggest that throughout adulthood olfactory neurons are overproduced, differentiate independently from their target, and then undergo a process of target-induced neuronal selection.     

FGF2 promotes neuronal differentiation in explant cultures of adult and embryonic mouse olfactory epithelium

Neurogenesis in the adult olfactory epithelium is highly regulated in vivo. Little is known of the molecular signals which control this process, although contact with the olfactory bulb or with astrocytes has been implicated. Explants of mouse olfactory epithelium were grown in the presence or absence of several peptide growth factors. Basic fibroblast growth factor (FGF2) stimulated differentiation of sensory neurons in adult and embryonic olfactory epithelium. Other growth factors tested were ineffective. FGF2-stimulated neurons were born in vitro and expressed neurofilament, neural cell adhesion molecule, and β-tubulin. The cells also expressed olfactory marker protein, a marker for mature olfactory sensory neurons in vivo. These bipolar neurons did not express glial fibrillary acidic protein or low-affinity nerve growth factor receptor. These results indicate that neither astrocytes nor olfactory bulb are necessary for differentiation of olfactory sensory neurons in vitro. © 1996 Wiley-Liss, Inc.     

Neuronal progenitors identified by their inability to express class I histocompatibility antigens in response to interferon-γ

Interferon-γ (IFN-γ) can induce class I major histocompatibility complex (MHC) antigen (H-2) expression on virtually all neuroepithelial cells isolated from embryonic day 9 (E9) mice. However, a subpopulation of cells become refractory to H-2 induction (H-2l^?) by E10 and the percentage of H-2 noninducible cells increases during development. Cell sorting, by flow cytometry or magnetic bead immunoselection, has shown that H-2l^? cells give rise exclusively to neuronal cells, and by E12, the majority of the neuronal progenitors are found within this population. It has also been found that 98% of the H-2l^? also express the neuron-associated marker, A2B5. Cells of the glial cell lineage retain the ability to express class I antigens throughout development. From these studies, it is clear that the neuroepithelium contains cells committed to the neuronal cell lineage as early as E10 in the mouse. Copyright © 1994 Wiley-Liss, Inc.     

Neuroplasticity in the olfactory system: differential effects of central and peripheral lesions of the primary olfactory pathway on the expression of B-50/GAP43 and the olfactory marker protein

The regeneration of the olfactory neuroepithelium following olfactory bulbectomy or peripheral deafferentation was studied with mRNA probes and antibodies for B-50/GAP43 and for olfactory marker protein (OMP). Two stages in the regeneration of the olfactory epithelium could be discerned with these reagents. The first stage occurs following either peripheral deafferentation of the olfactory epithelium with Triton X-100 (TX-100) or after bulbectomy and is characterized by the formation of a large population of immature olfactory receptor neurons. These newly formed neurons express B-50/GAP43, a phosphoprotein related to neuronal growth and plasticity. During the second stage of the regeneration process the newly formed olfactory neurons mature, as evidenced by a decrease in their expression of B-50/GAP43 and an increase in the expression of OMP. This stage is only manifested if the developing neurons have access to the target olfactory bulb. Formation of a full complement of OMP-expressing neurons occurs only after peripheral lesion with TX-100. In contrast, following bulbectomy the reconstituted olfactory epithelium lacks its normal target and is compromised in its ability to recover from nerve damage, as evidenced by the presence of a large number of B-50/GAP43-expressing neurons up to 3 months after the lesion and its failure to establish a full complement of OMP-expressing neurons. These results demonstrate that the olfactory epithelium is capable of replacing its sensory neurons independently of the presence of its target, the olfactory bulb. However, the differential patterns of expression of B-50/GAP43 and OMP at long times after peripheral lesion with TX-100 or bulbectomy illustrate the profound effect the olfactory bulb has on neuronal maturation in reconstituted olfactory neuroepithelium.     

胚胎发育不良性神经上皮肿瘤的MRI表现

目的 探讨胚胎发育不良性神经上皮肿瘤(DNT)的MRI影像学表现及临床特点,以期提高对DNT的诊断水平.方法 收集经手术病理证实的12例DNT患者临床资料,回顾性分析其影像学表现及临床特点.结果 12例患者肿瘤均位于幕上大脑半球皮层内,颞叶8例,额叶及顶叶各2例,其中6例累及邻近脑白质.MRI平扫中,T1WI呈低或等低混杂信号,T2WI及FLAIR呈高信号,肿瘤表现为囊性或以囊性部分为主,部分内可见网状分隔.5例病灶表现为"三角征",5例表现为"脑回征",另2例表现为圆形病灶.肿瘤均无明显占位效应,1例瘤周轻度水肿,增强扫描仅1例有轻度不均匀强化.结论 DNT符合一般良性肿瘤的生物学特征,MRI诊断价值很大.大脑半球皮层内囊性病变,无占位效应,瘤周无水肿,出现"三角征"和"脑回征"等征象可提示DNT的诊断.

Abstract：

Objective To study the MRI appearance and clinical features of dysembryoplastic neuroepithelial tumor (DNT) to improve accurate diagnosis of DNT. Methods The clinical data and MRI appearance of 12 patients with DNT confirmed by surgery and pathology were analyzed retrospectively. Routine MRI was performed in all of the 12 patients, and also dynamic contrast-enhanced MR imaging in 10 of them. Results Eight lesions resided in the temporal lobes, 2in the frontal lobe, and the other 2 in the parietal lobe. All of the 12 DNTs located in supratentorial hemisphere cortex, and 6 of them encroached the adjacent white matter. The lesions appeared as hypointense or iso-hypointense signal on T1WI of MRI, and hyperintense on T2WI and FLAIR of MRI.Cystic structure was shown in all the lesions or as its main part, and mesh-separated structure was also noted in some of the tumors. Five lesions appeared as a triangle in shape, 5 in gyrus-like shape and the remaining 2 in round shape. All the tumors had no significant mass effect with 1 having mild edema around the tumor. Enhanced MR imaging showed only 1 lesion having slight and heterogeneous enhancement. Conclusion DNT is accorded with general tumor's biological characteristics. MRI has great value on DNT's diagnosis. If a cystic lesion locates in the hemisphere cortex with a triangle in shape or gyrus-like shape, having no significant mass effect and peripheral edema, it has a great possibility of being DNT.     

Histopathological analysis of the olfactory epithelium of zebrafish (Danio rerio) exposed to sublethal doses of urea

Chronic renal disease is known to alter olfactory function, but the specific changes induced in olfactory organs during this process remain unclear. Of the uraemic toxins generated during renal disease, high levels of urea are known to induce hyposmic conditions. In this study, the effects of environmental exposure to elevated concentrations of urea (7, 13.5 and 20 g L^?1) on the sensory mucosa of zebrafish in acute toxicity and chronic toxicity tests were described. It was observed that lamellae maintained structural integrity and epithelial thickness was slightly reduced, but only following exposure to the highest concentrations of urea. Pan‐neuronal labelling with anti‐Hu revealed a negative correlation with levels of urea, leading to investigation of whether distinct neuronal subtypes were equally sensitive. Using densitometric analysis of immunolabelled tissues, numbers of G_{α olf}‐, TRPC2‐ and TrkA‐expressing cells were compared, representing ciliated, microvillous and crypt neurons, respectively. The three neuronal subpopulations responded differently to increasing levels of urea. In particular, crypt cells were more severely affected than the other cell types, and G_{α olf}‐immunoreactivity was found to increase when fish were exposed to low doses of urea. It can be concluded that exposure to moderate levels of urea leads to sensory toxicity directly affecting olfactory organs, in accordance with the functional olfactometric measurements previously reported in the literature.     

Many roads lead to primary autosomal recessive microcephaly

Autosomal recessive primary microcephaly (MCPH), historically referred to as Microcephalia vera, is a genetically and clinically heterogeneous disease. Patients with MCPH typically exhibit congenital microcephaly as well as mental retardation, but usually no further neurological findings or malformations. Their microcephaly with grossly preserved macroscopic organization of the brain is a consequence of a reduced brain volume, which is evident particularly within the cerebral cortex and thus results to a large part from a reduction of grey matter. Some patients with MCPH further provide evidence of neuronal heterotopias, polymicrogyria or cortical dysplasia suggesting an associated neuronal migration defect. Genetic causes of MCPH subtypes 1–7 include mutations in genes encoding microcephalin, cyclin-dependent kinase 5 regulatory associated protein 2 (CDK5RAP2), abnormal spindle-like, microcephaly associated protein (ASPM), centromeric protein J (CENPJ), and SCL/TAL1-interrupting locus (STIL) as well as linkage to the two loci 19q13.1–13.2 and 15q15–q21. Here, we provide a timely overview of current knowledge on mechanisms leading to microcephaly in humans with MCPH and abnormalities in cell division/cell survival in corresponding animal models. Understanding the pathomechanisms leading to MCPH is of high importance not only for our understanding of physiologic brain development (particularly of cortex formation), but also for that of trends in mammalian evolution with a massive increase in size of the cerebral cortex in primates, of microcephalies of other etiologies including environmentally induced microcephalies, and of cancer formation.     

Ovarian germ cell tumour classification: views from the testis

The classification of ovarian germ cell tumours has remained unchanged for many years, while there have been considerable changes in the testicular classification. In recent years there has been concern about the overtreatment of clinical stage 1 testicular germ cell tumours with increasing use of surveillance for low-risk disease. We outline here the current classification of germ cell tumours of the ovary with particular regard to treatment and outcome and highlight some areas which may cause confusion, particularly pertaining to immature teratomas and mixed germ cell tumours. We suggest that some minor changes to the classification, evidenced by a recent retrospective series by some of the authors, may lead to less adjuvant chemotherapy for immature teratomas and may obviate the need for the grading of immature teratomas, by aligning with testicular experience in pure post-pubertal teratomas. Adoption of this will require retrospective and prospective re-evaluation, but may avoid long-term patient morbidity.     

Molecular cloning of three zebrafish lin7 genes and their expression patterns in the retina

The vertebrate retina develops from an undifferentiated sheet of neuroepithelial cells, whose differentiation requires the generation and maintenance of the correct cellular polarity. To examine the role of cell polarity in retinal development, we cloned three zebrafish lin7 genes (lin7a, lin7b, and lin7c), which each encodes a protein candidate that is required for generation/maintenance of neuroepithelial cell junctions. These three zebrafish Lin7 proteins share over 78% amino acid identity and contain both L27 and PDZ domains that are present in all Lin7 homologs. Immunoblots revealed that the Lin7b and Lin7c proteins were first expressed in the developing eye by 24hr postfertilization (hpf), while Lin7a was not detected in the eye until 72 hpf. At 33 hpf, the Lin7 proteins localized at, or slightly apical of, the actin-associated adherens junctions in the retinal neuroepithelium. This subcellular distribution required the expression of the Nok protein. In the absence of Nok, the Lin7 proteins failed to localize to either the ectopic adherens junctions or the cell membrane. At 4 days postfertilization, in situ hybridisation revealed that all three lin7 genes were expressed in both the ganglion cell layer and the bipolar cell region of the inner nuclear layer. The lin7a gene was also expressed in the amacrine and horizontal cell regions of the inner nuclear layer, while lin7c was also expressed in the outer nuclear layer. In the adult retina, where Lin7a is the predominant form expressed, the Lin7 proteins were localized to the outer and inner plexiform layers, the bipolar and horizontal cells of the inner nuclear layer, and the ganglion cells. These results suggest that the three zebrafish Lin7 proteins possess partially redundant, yet essential, roles in retinal development.