Browsing by Subject "Axolotl"

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    Axolotl Xenografts Improve Regeneration of Xenopus Hind Limbs
    (Office of the Vice Chancellor for Research, 2013-04-05) Chen, Xiaoping; Stocum, David L.
    Axolotls regenerate perfect copies of amputated limbs, whereas Xenopus froglet limbs regenerate only a spike of cartilage. We asked whether axolotl muscle and cartilage xenografted from normal or GFP-labeled limbs to amputated froglet limbs, with or without treatment with cyclosporin A (CSA) and/or retinoic acid (RA), would improve Xenopus limb regeneration via the release of regeneration-promoting factors into the host limb tissue. The grafted froglet limbs were allowed to regenerate for three months to two years. We detected initial symptoms of graft vs. host disease with or without CSA treatment that subsequently disappeared. The grafted limbs first formed a spike that subsequently grew wider at the tip and after three months began to separate into 2-5 digit-like structures that continued to grow. CSA and low-dose RA treatment decreased the time at which digit formation could be detected but were not necessary for digit formation. The digit pattern was not asymmetric, thus individual digits were not identifiable. Immature muscle was detected in the regenerated limbs by trichrome and MF-20 antibody staining, and nerve fibers were detected by Luxol Fast Blue staining. In one limb with a GFP graft, a few axolotl cells were detected around the base of the digits that may have stimulated digit separation. Although the mechanism of digit formation remains obscure, we conclude that factors released by degraded axolotl tissue or surviving axolotl cells can stimulate complex tissue regeneration and initiate the first step of digital anterior-posterior pattern formation in regenerating Xenopus hind limbs. These results have significance for the possibility of stimulating the regeneration of complex mammalian structures that have been injured by trauma or disease.
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    Leukocyte Tyrosine Kinase ( Ltk) Is the Mendelian Determinant of the Axolotl Melanoid Color Variant
    (MDPI, 2023-04-13) Kabangu, Mirindi; Cecil, Raissa; Strohl, Lloyd, II; Timoshevskaya, Nataliya; Smith, Jeramiah J.; Voss, Stephen R.; Medicine, School of Medicine
    The great diversity of color patterns observed among amphibians is largely explained by the differentiation of relatively few pigment cell types during development. Mexican axolotls present a variety of color phenotypes that span the continuum from leucistic to highly melanistic. The melanoid axolotl is a Mendelian variant characterized by large numbers of melanophores, proportionally fewer xanthophores, and no iridophores. Early studies of melanoid were influential in developing the single-origin hypothesis of pigment cell development, wherein it has been proposed that all three pigment cell types derive from a common progenitor cell, with pigment metabolites playing potential roles in directing the development of organelles that define different pigment cell types. Specifically, these studies identified xanthine dehydrogenase (XDH) activity as a mechanism for the permissive differentiation of melanophores at the expense of xanthophores and iridophores. We used bulked segregant RNA-Seq to screen the axolotl genome for melanoid candidate genes and identify the associated locus. Dissimilar frequencies of single-nucleotide polymorphisms were identified between pooled RNA samples of wild-type and melanoid siblings for a region on chromosome 14q. This region contains gephyrin (Gphn), an enzyme that catalyzes the synthesis of the molybdenum cofactor that is required for XDH activity, and leukocyte tyrosine kinase (Ltk), a cell surface signaling receptor that is required for iridophore differentiation in zebrafish. Wild-type Ltk crispants present similar pigment phenotypes to melanoid, strongly implicating Ltk as the melanoid locus. In concert with recent findings in zebrafish, our results support the idea of direct fate specification of pigment cells and, more generally, the single-origin hypothesis of pigment cell development.
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    The role of retinoids in the regeneration of the axolotl spinal cord
    (2015-07-17) Kirk, Maia P.; Chernoff, Ellen A. G.; Belecky-Adams, Teri; Baucum II, A. J.
    Retinoids play an important role in tissue patterning during development as well as in epithelial formation and health. In the mammalian central nervous system, the meninges are a source of retinoids for brain tissue. Retinoid production has been described in juvenile Axolotl ependymal cells. Retinoid effects may possess a significant role in the regeneration-permissive interaction of the meninges and ependyma of the Axolotl spinal cord after penetrating injury. During spinal cord regeneration in urodele amphibians, the pattern of retinoid production changes as the meninges interact with the injury-reactive ependymal cells reconstructing the injured spinal cord. In order to determine which components of the retinoid metabolism and intracellular signaling pathway act in Urodele spinal cord regeneration, we employed antibody/horseradish peroxidase staining of both intact and regenerating Axolotl spinal cord tissues obtained from adult animals as well as cell culture techniques to determine expression of three retinoid pathway components: Cellular Retinoic Acid Binding Protein II (CRABP 2), Cellular Retinol Binding Protein I (CRBP 1), and Retinaldehyde Dehydrogenase II (RALDH 2). Current results demonstrate the following in the intact cord: 1) CRBP 1 is expressed in the pia and dura mater meningeal layers, in gray matter neurons (including their axonal processes), and the ependymal cell radial processes that produce the glia limitans, 2) CRABP 2 is expressed in the arachnoid and/or dura mater meningeal layers surrounding the spinal cord, and 3) RALDH 2 is expressed in the meninges as well as cytoplasm of grey matter neurons and some ependymal/sub-ependymal cells. In the regenerating cord, CRBP 1 is expressed in ependymal cells that are undergoing epithelial-to-mesenchymal transition (EMT), as is CRABP 2. RALDH 2 staining is very strong in the reactive meninges; in addition, expression is also upregulated in the cytoplasmic and perinuclear regions of reactive grey matter neurons, including motor neurons and in the apical region of ependymal. Preliminary studies culturing reactive meninges and ependymal cells together suggested that the meninges could drive re-epithelialization of the reactive ependymal cells. Experiments to characterize this interaction show an unusual proliferation pattern: Proliferating Cell Nuclear Antigen (PCNA) labeling is present in intact and regenerating cord ependymal cells. However, in culture, the presence of meninges results in no proliferation proximal to the explant, but extensive proliferation in leading cell outgrowth; also, the cultured meninges is positive for RALDH2. In summary, the intact adult cord shows meningeal production of RA, which is upregulated following injury; in addition, during this time, RA production is upregulated in the adult ependymal cells as well. In culture, the reactive meninges appears to modulate the behavior of reactive ependymal cells.