Matrisomal components involved in regenerative wound healing in axolotl and Acomys: implications for biomaterial development.

Achieving regeneration in humans has been a long-standing goal of many researchers. Whereas amphibians like the axolotl (Ambystoma mexicanum) are capable of regenerating whole organs and even limbs, most mammals heal their wounds via fibrotic scarring. Recently, the African spiny mouse (Acomys sp.) has been shown to be injury resistant and capable of regenerating several tissue types. A major focal point of research with Acomys has been the identification of drivers of regeneration. In this search, the matrisome components related to the extracellular matrix (ECM) are often overlooked. In this review, we compare Acomys and axolotl skin wound healing and blastema-mediated regeneration by examining their wound healing responses and comparing the expression pattern of matrisome genes, including glycosaminoglycan (GAG) related genes. The goal of this review is to identify matrisome genes that are upregulated during regeneration and could be potential candidates for inclusion in pro-regenerative biomaterials. Research papers describing transcriptomic or proteomic coverage of either skin regeneration or blastema formation in Acomys and axolotl were selected. Matrisome and GAG related genes were extracted from each dataset and the resulting lists of genes were compared. In our analysis, we found several genes that were consistently upregulated, suggesting possible involvement in regenerative processes. Most of the components have been implicated in regulation of cell behavior, extracellular matrix remodeling and wound healing. Incorporation of such pro-regenerative factors into biomaterials may help to shift pro-fibrotic processes to regenerative responses in treated wounds.

mef2ca and mef2cb Double Mutant Zebrafish Show Altered Craniofacial Phenotype and Motor Behaviour.

The transcription factor MEF2C is crucial in neuronal, cardiac, bone and cartilage molecular processes, as well as for craniofacial development. MEF2C was associated with the human disease MRD20, whose patients show abnormal neuronal and craniofacial development. Zebrafish mef2ca;mef2cb double mutants were analysed for abnormalities in craniofacial and behaviour development through phenotypic analysis. Quantitative PCR was performed to investigate the expression levels of neuronal marker genes in mutant larvae. The motor behaviour was analysed by the swimming activity of 6 dpf larvae. We found that mef2ca;mef2cb double mutants display several abnormal phenotypes during early development, including those already described in zebrafish carrying mutations in each paralog, but also (i) a severe craniofacial phenotype (comprising both cartilaginous and dermal bone structures), (ii) developmental arrest due to the disruption of cardiac oedema and (iii) clear alterations in behaviour. We demonstrate that the defects observed in zebrafish mef2ca;mef2cb double mutants are similar to those previously described in MEF2C-null mice and MRD20 patients, confirming the usefulness of these mutant lines as a model for studies concerning MRD20 disease, the identification of new therapeutic targets and screening for possible rescue strategies.

Coronal brain atlas in stereotaxic coordinates of the African spiny mouse, Acomys cahirinus.

The African spiny mouse (Acomys cahirinus) is an emerging model of mammalian epimorphic regeneration that has aroused the interest of the scientific community in the last decade. To date, studies on brain repair have been hindered by the lack of knowledge on the neuroanatomy of this species. Here, we present a coronal brain atlas in stereotaxic coordinates, which allows for three-dimensional identification and localization of the brain structures of this species. The brain of 12-week-old spiny mice was mapped in stereotaxic coordinates using cresyl violet-stained brain sections obtained from coronal cryosectioning of the brain after transcardial perfusion with fixative. The atlas is presented in 42 plates representing sections spaced 240 µm apart. Stereotaxic coordinates were validated using both a model of Parkinsonian lesion of the striatum with 6-hydroxydopamine and labeling of the corticospinal tract in the spiny mouse spinal cord using AAV1/2-GFP intracortical injections. This work presents a new tool in A. cahirinus neurobiology and opens new avenues of research for the investigation of the regenerative ability of A. cahirinus in models of brain disorders.

Rewired glycosylation activity promotes scarless regeneration and functional recovery in spiny mice after complete spinal cord transection.

Regeneration of adult mammalian central nervous system (CNS) axons is abortive, resulting in inability to recover function after CNS lesion, including spinal cord injury (SCI). Here, we show that the spiny mouse (Acomys) is an exception to other mammals, being capable of spontaneous and fast restoration of function after severe SCI, re-establishing hind limb coordination. Remarkably, Acomys assembles a scarless pro-regenerative tissue at the injury site, providing a unique structural continuity of the initial spinal cord geometry. The Acomys SCI site shows robust axon regeneration of multiple tracts, synapse formation, and electrophysiological signal propagation. Transcriptomic analysis of the spinal cord following transcriptome reconstruction revealed that Acomys rewires glycosylation biosynthetic pathways, culminating in a specific pro-regenerative proteoglycan signature at SCI site. Our work uncovers that a glycosylation switch is critical for axon regeneration after SCI and identifies ß3gnt7, a crucial enzyme of keratan sulfate biosynthesis, as an enhancer of axon growth.

Expression pattern of cdkl5 during zebrafish early development: implications for use as model for atypical Rett syndrome.

Atypical Rett syndrome is a child neurodevelopmental disorder induced by mutations in CDKL5 gene and characterized by a progressive regression in development with loss of purposeful use of the hands, slowed brain and head growth, problems with walking, seizures, and intellectual disability. At the moment, there is no cure for this pathology and little information is available concerning animal models capable of mimicking its phenotypes, thus the development of additional animal models should be of interest to gain more knowledge about the disease. Zebrafish has been used successfully as model organism for many human genetic diseases; however, no information is available concerning the spatial and temporal expression of cdkl5 orthologous in this organism. In the present study, we identified the developmental expression patterns of cdkl5 in zebrafish by quantitative PCR and whole-mount in situ hybridization. cdkl5 is expressed maternally at low levels during the first 24 h of development. After that the expression of the gene increases significantly and it starts to be expressed mainly in the nervous system and in several brain structures, such as telencephalon, mesencephalon and diencephalon. The expression patterns of cdkl5 in zebrafish is in accordance with the tissues known to be affected in humans and associated to symptoms and deficits observed in Rett syndrome patients thus providing the first evidence that zebrafish could be an alternative model to study the molecular pathways of this disease as well as to test possible therapeutic approaches capable of rescuing the phenotype.

Xenopus Pkdcc1 and Pkdcc2 Are Two New Tyrosine Kinases Involved in the Regulation of JNK Dependent Wnt/PCP Signaling Pathway.

Protein Kinase Domain Containing, Cytoplasmic (PKDCC) is a protein kinase which has been implicated in longitudinal bone growth through regulation of chondrocytes formation. Nevertheless, the mechanism by which this occurs remains unknown. Here, we identified two new members of the PKDCC family, Pkdcc1 and Pkdcc2 from Xenopus laevis. Interestingly, our knockdown experiments revealed that these two proteins are both involved on blastopore and neural tube closure during gastrula and neurula stages, respectively. In vertebrates, tissue polarity and cell movement observed during gastrulation and neural tube closure are controlled by Wnt/Planar Cell Polarity (PCP) molecular pathway. Our results showed that Pkdcc1 and Pkdcc2 promote the recruitment of Dvl to the plasma membrane. But surprisingly, they revealed different roles in the induction of a luciferase reporter under the control of Atf2 promoter. While Pkdcc1 induces Atf2 expression, Pkdcc2 does not, and furthermore inhibits its normal induction by Wnt11 and Wnt5a. Altogether our data show, for the first time, that members of the PKDCC family are involved in the regulation of JNK dependent Wnt/PCP signaling pathway.

Expression pattern of zcchc24 during early Xenopus development.

We report the expression pattern of a novel Xenopus laevis gene, zcchc24, which encodes a protein containing two zinc finger domains from the zf-CCHC and zf-3CxxC superfamilies. This protein shares >84% amino acid identity with its vertebrate homologues. During X. laevis embryonic development, zcchc24 is expressed at gastrula stages in the dorsal mesoderm, including the cardiac precursors region. During neurula stages, zcchc24 is expressed as two stripes in the dorsal region, more precisely, in the somitogenic mesoderm until the cardiac mesoderm. At early tailbud stages, zcchc24 continues to be expressed in these regions, but starts to be expressed in the migrating neural crest. Later, this gene is expressed in the head, branchial arches, heart and somites. The zinc finger domains present in Zcchc24 protein and its dynamic gene expression pattern suggest that Zcchc24 might be involved in the regulation of heart, somites and of branchial arch formation/patterning, namely in the regulation of apoptosis.

Supported neodymium catalysts for isoprene and rac-ß-butyrolactone polymerization: modulation of reactivity by controlled grafting.

A series of hybrid materials, bearing neodymium silylamide initiating groups, have been shown to mediate isoprene polymerization when combined with alkyl aluminum activators [methylaluminoxane, AlEt(2)Cl, Al(iBu)(3)]. The surface species nature and relative distribution were correlated with isoprene polymerization activity and selectivity. This approach to stereocontrol modulation has been extended to racemic ß-butyrolactone isoselective ring opening polymerization.

Vsx2 in the zebrafish retina: restricted lineages through derepression.

BACKGROUND: The neurons in the vertebrate retina arise from multipotent retinal progenitor cells (RPCs). It is not clear, however, which progenitors are multipotent or why they are multipotent. RESULTS: In this study we show that the homeodomain transcription factor Vsx2 is initially expressed throughout the retinal epithelium, but later it is downregulated in all but a minor population of bipolar cells and all Müller glia. The Vsx2-negative daughters of Vsx2-positive RPCs divide and give rise to all other cell types in the retina. Vsx2 is a repressor whose targets include transcription factors such as Vsx1, which is expressed in the progenitors of distinct non-Vsx2 bipolars, and the basic helix-loop-helix transcription factor Ath5, which restricts the fate of progenitors to retinal ganglion cells, horizontal cells, amacrine cells and photoreceptors fates. Foxn4, expressed in the progenitors of amacrine and horizontal cells, is also negatively regulated by Vsx2. CONCLUSION: Our data thus suggest Vsx2-positive RPCs are fully multipotent retinal progenitors and that when Vsx2 is downregulated, Vsx2-negative progenitors escape Vsx2 repression and so are able to express factors that restrict lineage potential.

Generating asymmetries in the early vertebrate embryo: the role of the Cerberus-like family.

One fundamental aspect of vertebrate embryonic development is the formation of the body plan. For this process, asymmetries have to be generated during early stages of development along the three main body axes: Anterior-Posterior, Dorso-Ventral and Left-Right. We have been studying the role of a novel class of molecules, the Cerberus/Dan gene family. These are dedicated secreted antagonists of three major signaling pathways: Nodal, BMP and Wnt. Our studies contribute to the current view that the fine tuning of signaling is controlled by a set of inhibitory molecules rather than by activators. In this context, the Cerberus-like molecules emerge as key players in the regulation and generation of asymmetries in the early vertebrate embryo.

Developmental expression of Shisa-2 in Xenopus laevis.

Shisa is an antagonist of Wnt and FGF signaling, that functions cell autonomously in the endoplasmic reticulum (ER) to inhibit the post-translational maturation of Wnt and FGF receptors. In this paper we report the isolation of a second Xenopus shisa gene (Xshisa-2). Xenopus Shisa-2 shows 30.7% identity to Xshisa. RT-PCR analysis indicated that Xshisa-2 mRNA is present throughout early development and shows an increased expression during neurula and tailbud stages. At neurula stages Xenopus shisa-2 is initially expressed in the presomitic paraxial mesoderm and later in the developing somites. The expression profiles and pattern of Xshisa and Xshisa-2 differ significantly. During gastrulation only Xshisa mRNA is present in the Spemann-Mangold organizer and later on becomes restricted to the neuroectoderm and the prechordal plate.

Influences on neural lineage and mode of division in the zebrafish retina in vivo.

Cell determination in the retina has been under intense investigation since the discovery that retinal progenitors generate clones of apparently random composition (Price, J., D. Turner, and C. Cepko. 1987. Proc. Natl. Acad. Sci. USA. 84:156-160; Holt, C.E., T.W. Bertsch, H.M. Ellis, and W.A. Harris. 1988. Neuron. 1:15-26; Wetts, R., and S.E. Fraser. 1988. Science. 239:1142-1145). Examination of fixed tissue, however, sheds little light on lineage patterns or on the relationship between the orientation of division and cell fate. In this study, three-dimensional time-lapse analyses were used to trace lineages of retinal progenitors expressing green fluorescent protein under the control of the ath5 promoter. Surprisingly, these cells divide just once along the circumferential axis to produce two postmitotic daughters, one of which becomes a retinal ganglion cell (RGC). Interestingly, when these same progenitors are transplanted into a mutant environment lacking RGCs, they often divide along the central-peripheral axis and produce two RGCs. This study provides the first insight into reproducible lineage patterns of retinal progenitors in vivo and the first evidence that environmental signals influence the orientation of cell division and the lineage of neural progenitors.