Cell cycle-related genes p57kip2, Cdk5 and Spin in the pathogenesis of neural tube defects

In the field of developmental neurobiology, accurate and ordered regulation of the cell cycle and apoptosis are crucial factors contributing to the normal formation of the neural tube. Preliminary studies identified several genes involved in the development of neural tube defects. In this study, we established a model of developmental neural tube defects by administration of retinoic acid to pregnant rats. Gene chip hybridization analysis showed that genes related to the cell cycle and apoptosis, signal transduction, transcription and translation regulation, energy and metabolism, heat shock, and matrix and cytoskeletal proteins were all involved in the formation of developmental neural tube defects. Among these, cell cycle-related genes were predominant. Retinoic acid treat-ment caused differential expression of three cell cycle-related genes p57kip2, Cdk5 and Spin, the expression levels of which were downregulated by retinoic acid and upregulated during normal neural tube formation. The results of this study indicate that cell cycle-related genes play an im-portant role in the formation of neural tube defects. P57kip2, Cdk5 and Spin may be critical genes in the pathogenesis of neural tube defects.

Overexpression of the tissue inhibitor of metalloproteinase-3 during Xenopus embryogenesis affects head and axial tissue formation

Tissue inhibitors of metalloproteinases (TIMPs) modulate extracellular matrix remodeling during embryonic develop- ment and disease. TIMP-3 expression was examined during Xenopus laevis embryogenesis: TIMP-3 transcripts detected in the maternal pool of RNA increased at the mid-blastula transition, decreased dramatically during gastrulation and increased again during neurulation and axis elongation. Interestingly, the decrease during gastrulation was not seen in LiCl treated (dorsalized) embryos. Whole mount in situ hybridization of TIMP-3 using DIG-labeled RNA probes demonstrated that the transcripts were present in all dorsal tissues during embryogenesis, but were prominent only in head structures starting at stage 35. Overexpression of TIMP-3 through transgenesis and RNA injections led to devel- opmental abnormalities and death. Both overexpression strategies resulted in post-gastrulation perturbation including those to neural and head structures, as well as truncated axes. However, RNA injections resulted in more severe early defects such as failure of neural tube closure, and transgenesis caused truncated axes and head abnormalities. No transgenic embryo expressing TIMP-3 survived past stage 40.

Gene expression in retinoic acid-induced neural tube defects A cDNA microarray analysis

BACKGROUND： Neural tube defects can be induced by abnormal factors in vivo or in vitro during development. However, the molecular mechanisms of neural tube defect induction, and the related gene expression and regulation are still unknown. OBJECTIVE： To compare the differences in gene expression between normal embryos and those with neural tube defects. DESIGN, TIME AND SETTING： A neural development study was performed at the Department of Neurobiology, Third Military Medical University of Chinese PLA between January 2006 and October 2007. MATERIALS： Among 120 adult Kunming mice, 60 pregnant mice were randomly and evenly divided into a retinoic acid group （n = 30） and a normal control group （n =30）. The retinoic acid was produced by Sigma, USA, the gene microarray by the Amersham Pharmacia Company, Hong Kong, and the gene sequence was provided by the Incyte database, USA. METHODS： Retinoic acid was administered to prepare models of neural tube defects, and corn oil was similarly administered to the normal control group. Total RNA was extracted from embryonic tissue of the two groups using a Trizol kit, and a cDNA microarray containing 1 100 known genes was used to compare differences in gene expression between the normal control group and the retinoic acid group on embryonic （E） day 10.5 and 11.5. Several differentially expressed genes were randomly selected from the two groups for Northern blotting, to verify the results of the cDNA microarray. MAIN OUTCOME MEASURES： Morphological changes and differential gene expression between the normal control group and the retinoic acid group. RESULTS： Anatomical microscopy demonstrated that an intact closure of the brain was formed in the normal mouse embryos by days E10.5 and E11.5. The cerebral appearance was full and smooth, and the surface of the spine was intact. However, in the retinoic acid group on days E10.5 and E11.5, there were more dead embryos. Morphological malformations typically included non-closure at the top of the cranium and abnormal changes of the metencephalon and face. cDNA microarray analysis suggested that the changes in expression of seven different genes were similar on both days E10.5 and E11.5. These were downregulation of NekT, Igfbp5, Zw10, Csf3r, Psmc6 and Rb 1, and upregulation of Apoa-4. This study also indicated that Cdk5 expression was downregulated in the retinoic acid group on day E11.5. The results of the cDNA microarray analysis were partly confirmed by Northern blotting. CONCLUSION： Cdk5, Nek7, Igfbp5, Zw10, Csf3r, Psmc6, Rb1 and Apoa-4 may be key factors in retinoic acid-induced neural tube defects.

Cell extrusion in development and cancer,what MARCKS the difference for epithelial integrity?

Cell extrusion is an active mechanism to eliminate non-viable or supernumerary cells in healthy epithelia.It also plays a role in carcinogenesis,both in tumor growth(apical extrusion)and metastasis(basal extrusion).Embryonic tissues like the neuroepithelium,on the other hand,present rates of proliferation comparable to that of carcinomas,without the occurrence of cell extrusion.However,the downregulation or phosphorylation of actinmodulating proteins like MARCKS,causes extensive neuroepithelial apical cell extrusion.As changes in MARCKS proteins phosphorylation and expression have also been correlated to carcinogenesis,we propose here an integrated model for their functions in epithelial integrity.

Characterization of SoxB2 and SoxC genes in Amphi-oxus (Branchiostoma belcheri):Implications for their evolutionary conservation

Most Sox genes directly affect cell fate determination and differentiation. In this study,we isolated two Sox genes:SoxB2 and SoxC from amphioxus (Branchiostoma belcheri),the closest living invertebrate relative of the vertebrates. Alignments of SoxB2 and SoxC protein sequences and their vertebrate homologs show high conservation of their HMG domains. Phylogenic analysis shows that amphioxus SoxB2 and SoxC fall out of the vertebrate branches,suggesting that vertebrate homologs might arise from gene duplications during evolution. The two genes possess similar spatial and temporal expression patterns during embryogenesis and in adults. They are both maternally inherited. During neurulation,they are expressed in the neural ectoderm and archenterons. In adults,they are expressed not only in the nerve cord,but also in the gut,midgut diverticulum,gill and oocytes. These results suggest that amphioxus SoxB2 and SoxC might co-function and have conserved functions in the nervous system and gonads as their vertebrate homologs.

Cortactin mediated morphogenic cell movements during zebrafish (Danio rerio) gastrulation

Cell migration is essential to direct embryonic cells to specific sites at which their developmental fates are ultimately determined. However, the mechanism by which cell motility is regulated in embryonic development is largely unknown. Cortactin, a filamentous actin binding protein, is an activator of Arp2/3 complex in the nucleation of actin cytoskeleton at the cell leading edge and acts directly on the machinery of cell motility. To determine whether cortactin and Arp2/3 mediated actin assembly plays a role in the morphogenic cell movements during the early development of zebrafish, we initiated a study of cortactin expression in zebrafish embryos at gastrulating stages when massive cell migrations occur. Western blot analysis using a cortactin specific monoclonal antibody demonstrated that cortactin protein is abundantly present in em-bryos at the most early developmental stages. Immunostaining of whole-mounted embryo showed that cortactin immunoreactivity was associated with the embryonic shield, predominantly at the dorsal side of the embryos during gastrulation. In addition, cortactin was detected in the convergent cells of the epiblast and hypoblast, and later in the central nervous system. Im-munofluorescent staining with cortactin and Arp3 antibodies also revealed that cortactin and Arp2/3 complex colocalized at the periphery and many patches associated with the cell-to-cell junction in motile embryonic cells. Therefore, our data suggest that cortactin and Arp2/3 medi-ated actin polymerization is implicated in the cell movement during gastrulation and perhaps the development of the central neural system as well.