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The role of osteolineage cells in regulating hematopoietic stem cell (HSC) regeneration following myelosuppression is not well understood. Here we show that deletion of the pro-apoptotic genes Bak and Bax in osterix (Osx, also known as Sp7 transcription factor 7)-expressing cells in mice promotes HSC regeneration and hematopoietic radioprotection following total body irradiation. These mice showed increased bone marrow (BM) levels of the protein dickkopf-1 (Dkk1), which was produced in Osx-expressing BM cells. Treatment of irradiated HSCs with Dkk1 in vitro increased the recovery of both long-term repopulating HSCs and progenitor cells, and systemic administration of Dkk1 to irradiated mice increased hematopoietic recovery and improved survival. Conversely, inducible deletion of one allele of Dkk1 in Osx-expressing cells in adult mice inhibited the recovery of BM stem and progenitor cells and of complete blood counts following irradiation. Dkk1 promoted hematopoietic regeneration via both direct effects on HSCs, in which treatment with Dkk1 decreased the levels of mitochondrial reactive oxygen species and suppressed senescence, and indirect effects on BM endothelial cells, in which treatment with Dkk1 induced epidermal growth factor (EGF) secretion. Accordingly, blockade of the EGF receptor partially abrogated Dkk1-mediated hematopoietic recovery. These data identify Dkk1 as a regulator of hematopoietic regeneration and demonstrate paracrine cross-talk between BM osteolineage cells and endothelial cells in regulating hematopoietic reconstitution following injury., Author(s): Heather A Himburg [1]; Phuong L Doan [2]; Mamle Quarmyne [1, 3]; Xiao Yan [1, 3]; Joshua Sasine [1]; Liman Zhao [1]; Grace V Hancock [4]; Jenny Kan [1]; [...]

Age-related changes in the niche have long been postulated to impair the function of somatic stem cells. Here we demonstrate that the aged stem cell niche in skeletal muscle contains substantially reduced levels of fibronectin (FN), leading to detrimental consequences for the function and maintenance of muscle stem cells (MuSCs). Deletion of the gene encoding FN from young regenerating muscles replicates the aging phenotype and leads to a loss of MuSC numbers. By using an extracellular matrix (ECM) library screen and pathway profiling, we characterize FN as a preferred adhesion substrate for MuSCs and demonstrate that integrin-mediated signaling through focal adhesion kinase and the p38 mitogen-activated protein kinase pathway is strongly de-regulated in MuSCs from aged mice because of insufficient attachment to the niche. Reconstitution of FN levels in the aged niche remobilizes stem cells and restores youth-like muscle regeneration. Taken together, we identify the loss of stem cell adhesion to FN in the niche ECM as a previously unknown aging mechanism., Extrinsic signals that originate in the immediate cellular environment, commonly known as the stem cell niche, are critical for the regulation of MuSCs (1). Following injury, the stem cell niche [...]

Myocardial infarction results in compromised myocardial function and heart failure owing to insufficient cardiomyocyte self-renewal (1). Unlike many vertebrates, mammalian hearts have only a transient neonatal renewal capacity (2). Reactivating [...]

The small Intestine Is a dynamic and complex organ that Is characterized by constant epithelium turnover and crosstalk among various cell types and the microbiota. Lymphatic capillaries of the small intestine, called lacteals, play key roles in dietary fat absorption and the gut immune response; however, little is known about the molecular regulation of lacteal function. Here, we performed a high-resolution analysis of the small intestinal stroma and determined that lacteals reside in a permanent regenerative, proliferative state that is distinct from embryonic lymphangiogenesis or quiescent lymphatic vessels observed in other tissues. We further demonstrated that this continuous regeneration process is mediated by Notch signaling and that the expression of the Notch ligand delta-like 4 (DLL4) in lacteals requires activation of VEGFR3 and VEGFR2. Moreover, genetic inactivation of Dll4 in lymphatic endothelial cells led to lacteal regression and impaired dietary fat uptake. We propose that such a slow lymphatic regeneration mode is necessary to match a unique need of intestinal lymphatic vessels for both continuous maintenance, due to the constant exposure to dietary fat and mechanical strain, and efficient uptake of fat and immune cells. Our work reveals how lymphatic vessel responses are shaped by tissue specialization and uncover a role for continuous DLL4 signaling in the function of adult lymphatic vasculature., Introduction The small intestinal epithelium provides a barrier between the microorganism-rich intestinal lumen and the underlying stroma. Epithelial cells are constantly renewed from the stem cell niche at the crypt [...]

Postnatal tissue quiescence is thought to be a default state in the absence of a proliferative stimulus such as injury. Although previous studies have demonstrated that certain embryonic developmental programs are reactivated aberrantly in adult organs to drive repair and regeneration (1-3), it is not well understood how quiescence is maintained in organs such as the lung, which displays a remarkably low level of cellular turnover (4, 5). Here we demonstrate that quiescence in the adult lung is an actively maintained state and is regulated by hedgehog signalling. Epithelial-specific deletion of sonic hedgehog (Shh) during postnatal homeostasis in the murine lung results in a proliferative expansion of the adjacent lung mesenchyme. Hedgehog signalling is initially downregulated during the acute phase of epithelial injury as the mesenchyme proliferates in response, but returns to baseline during injury resolution as quiescence is restored. Activation of hedgehog during acute epithelial injury attenuates the proliferative expansion of the lung mesenchyme, whereas inactivation of hedgehog signalling prevents the restoration of quiescence during injury resolution. Finally, we show that hedgehog also regulates epithelial quiescence and regeneration in response to injury via a mesenchymal feedback mechanism. These results demonstrate that epithelial-mesenchymal interactions coordinated by hedgehog actively maintain postnatal tissue homeostasis, and deregulation of hedgehog during injury leads to aberrant repair and regeneration in the lung., The Hedgehog (Hh) pathway coordinates tissue-tissue interactions in multiple organs during embryonic development through paracrine activation of smoothened (Smo)-mediated downstream signalling events (6, 7). We have previously demonstrated that Shh [...]

The self-regenerating property of the adult myocardium is not a new discovery. Even though we could not confirm that the adult myocardium is a post-mitotic tissue, we should consider that its plasticity is extremely low. Studies are still in progress to decipher the mechanisms underlying the abovementioned potential fetal features of the adult heart. The modest results of several clinical trials based on the transplantation of millions of autologous stem cells into the dysfunctional heart have confirmed that the cross-talk of different signals, such as the microenvironment, promotes the regeneration of adult myocardium. Recent scientific evidence has revealed that cellular cross-talk does not depend on the action of a single cell phenotype. It is conceivable that the limited turnover of cardiomyocytes is ensured by the interplay of adult cardiac cells in response to environmental changes. The epigenetic state of a cell serves as a dynamic interface between the environment and phenotype. The epigenetic modulation of the adult cardiac cells by natural active compounds encourages further studies to improve myocardial plasticity. In this review, we will highlight the most relevant studies demonstrating the epigenetic modulation of myocardial regeneration without the use of stem cell transplantation. Key words: epigenetic, myocardial regeneration, microenvironment, heart, stem cells. La propriete d'auto-regeneration du myocarde adulte n'est pas une nouvelle decouverte. Bien que nous ne puissions confirmer que le myocarde adulte est un tissu post-mitotique, nous devrions considerer sa plasticite comme extremement faible. Des etudes sont toujours en cours afin de decortiquer les mecanismes qui sous-tendent ces caracteristiques possiblement foetales du coeur adulte. Les resultats modestes de plusieurs essais cliniques bases sur la transplantation de millions de cellules souches autologues dans le coeur dysfonctionnel ont confirme que la communication croisee de differents signaux comme le microenvironnement favorise la regeneration du myocarde adulte. Des donnees scientifiques recentes ont revele que la communication croisee cellulaire ne depend pas de l'action d'un phenotype cellulaire unique. On peut concevoir que le renouvellement limite des cardiomyocytes soit assure par l'interaction de cellules cardiaques adultes en reponse a des changements environnementaux. L'etat epigenetique d'une cellule sert d'interface dynamique entre l'environnement et le phenotype. La modulation epigenetique des cellules cardiaques adultes par des composes naturels actifs nous encourage a realiser davantage d'etudes afin d'ameliorer la plasticite myocardique. Dans cet article de synthese, nous mettrons en evidence les etudes les plus pertinentes qui demontrent une modulation epigenetique de la regeneration myocardique sans utilisation de la transplantation de cellules souches. [Traduit par la Redaction] Mots-cles : epigenetique, regeneration myocardique, microenvironnement, coeur, cellules souches., Introduction Up to 15 years ago, the adult myocardium was not widely considered a self-renewing tissue, even if the rare mitosis of adult cardiomyocytes was already detected in the human [...]

The pancreas is a complex organ comprised of three critical cell lineages: islet (endocrine), acinar, and ductal. This review will focus upon recent insights and advances in the biology of pancreatic ductal cells. In particular, emphasis will be placed upon the regulation of ductal cells by specific transcriptional factors during development as well as the underpinnings of acinarductal metaplasia as an important adaptive response during injury and regeneration. We also address the potential contributions of ductal cells to neoplastic transformation, specifically in pancreatic ductal adenocarcinoma., Introduction The pancreas is an organ that has endocrine and exocrine compartments (1). The endocrine compartment consists of α, β, δ, ζ, and pancreatic polypeptide cells organized into islets, whereas [...]

What gives an organism the ability to regrow tissues and to recover function where another organism fails is the central problem of regenerative biology. The challenge is to describe the mechanisms of regeneration at the molecular level, delivering detailed insights into the many components that are cross-regulated. In other words, a broad, yet deep dissection of the system-wide network of molecular interactions is needed. Functional genomics has been used to elucidate gene regulatory networks (GRNs) in developing tissues, which, like regeneration, are complex systems. Therefore, we reason that the GRN approach, aided by next generation technologies, can also be applied to study the molecular mechanisms underlying the complex functions of regeneration. We ask what characteristics a model system must have to support a GRN analysis. Our discussion focuses on regeneration in the central nervous system, where loss of function has particularly devastating consequences for an organism. We examine a cohort of cells conserved across all vertebrates, the reticulospinal (RS) neurons, which lend themselves well to experimental manipulations. In the lamprey, a jawless vertebrate, there are giant RS neurons whose large size and ability to regenerate make them particularly suited for a GRN analysis. Adding to their value, a distinct subset of lamprey RS neurons reproducibly fail to regenerate, presenting an opportunity for side-by-side comparison of gene networks that promote or inhibit regeneration. Thus, determining the GRN for regeneration in RS neurons will provide a mechanistic understanding of the fundamental cues that lead to success or failure to regenerate., Introduction What are the molecular factors that are necessary and sufficient to accomplish successful regeneration, as defined as regrowth after injury, leading to functional recovery? Many studies have beautifully illustrated [...]

The most extensive regenerative ability in adult vertebrates is found in the salamanders. Although it is often suggested that regeneration is an ancestral property for vertebrates, our studies on the cell-surface three-finger-protein Prod 1 provide clear evidence for the importance of local evolution of limb regeneration in salamanders. Prod 1 is implicated in both patterning and growth in the regeneration of limbs. It interacts with well-conserved proteins such as the epidermal growth-factor receptor and the anterior gradient protein that are widely expressed in phylogeny. A detailed analysis of the structure and sequence of Prod 1 in relation to other vertebrate three-finger proteins in mammals and zebra fish supports the view that it is a salamander-specific protein. This is the first example of a taxon-specific protein that is clearly implicated in the mechanisms of regeneration. We propose the hypothesis that regeneration depends on the activity of taxon-specific components in orchestrating a cellular machinery that is extensively conserved between regenerating and non-regenerating taxa. This hypothesis has significant implications for our outlook on regeneration in vertebrates, as well as for the strategies employed in extending regenerative ability in mammals. doi: 10.1093/icb/icq022

Angiogenesis occurs through a convergence of diverse signaling mechanisms with prominent pathways that include autocrine effects of endothelial nitric oxide (NO) synthase (eNOS)-derived NO and vascular endothelial growth factor (VEGF). However, the redundant and distinct roles of NO and VEGF in angiogenesis remain incompletely defined. Here, we use the partial hepatectomy model in mice genetically deficient in eNOS to ascertain the influence of eNOS-derived NO on the angiogenesis that accompanies liver regeneration. While sinusoidal endothelial cell (SEC) eNOS promotes angiogenesis in vitro, surprisingly the absence of eNOS did not influence the angiogenesis that occurs after partial hepatectomy in vivo. While this observation could not be attributed to induction of alternate NOS isoforms, it was associated with induction of VEGF signaling as evidenced by enhanced levels of VEGF ligand in regenerating livers from mice genetically deficient in eNOS. However, surprisingly, mice that were genetically heterozygous for deficiency in the VEGF receptor, fetal liver kinase-1, also maintained unimpaired capacity for liver regeneration. In summary, inhibition of VEGF-and NO-dependent angiogenesis does not impair liver regeneration, indicating signaling redundancies that allow liver regeneration to continue in the absence of this canonical vascular pathway. endothelial nitric oxide synthase; vascular endothelial growth factor; nitric oxide; fetal liver kinase-1; angiogenesis; partial hepatectomy doi: 10.1152/ajpregu.00836.2009.

Animals capable of regenerating multiple tissue types, organs, and appendages after injury are common yet sporadic and include some sponge, hydra, planarian, and salamander (i.e., newt and axolotl) species, but notably such regenerative capacity is rare in mammals. The adult MRL mouse strain is a rare exception to the rule that mammals do not regenerate appendage tissue. Certain commonalities, such as blastema formation and basement membrane breakdown at the wound site, suggest that MRL mice may share other features with classical regenerators. As reported here, MRL fibroblast-like cells have a distinct cell-cycle (G2/M accumulation) phenotype and a heightened basal and wound site DNA damage/repair response that is also common to classical regenerators and mammalian embryonic stem cells. Additionally, a neutral and alkaline comet assay displayed a persistent level of intrinsic DNA damage in cells derived from the MRL mouse. Similar to mouse ES cells, the p53-target p21 was not expressed in MRL ear fibroblasts. Because the p53/p21 axis plays a central role in the DNA damage response and cell cycle control, we directly tested the hypothesis that p21 down-regulation could functionally induce a regenerative response in an appendage of an otherwise non-regenerating mouse strain. Using the ear hole closure phenotype, a genetically mapped and reliable quantitative indicator of regeneration in the MRL mouse, we show that the unrelated [Cdkn1a.sup.tmi/Tyj]/J [p21.sup.-/-] mouse (unlike the B6129SF2/J WT control) closes ear holes similar to MRL mice, providing a firm link between cell cycle checkpoint control and tissue regeneration. G2/M checkpoint | MRL mouse | regeneration | DNA damage | DNA repair doi/10.1073/pnas.1000830107

In response to DNA damage, checkpoint proteins halt cell cycle progression and promote repair or apoptosis, thereby preventing mutation accumulation and suppressing tumor development. The DNA damage checkpoint protein Hus1 associates with Rad9 and Rad1 to form the 9-1-1 complex, which localizes to DNA lesions and promotes DNA damage signaling and repair. Because complete inactivation of mouse Hus1 results in embryonic lethality, we developed a system for regulated Hus1 inactivation in the mammary gland to examine roles for Hus1 in tissue homeostasis and tumor suppression. Hus1 inactivation in the mammary epithelium resulted in genome damage that induced apoptosis and led to depletion of Hus1-null cells from the mammary gland. Conditional Hus1 knockout females retained grossly normal mammary gland morphology, suggesting compensation by cells that failed to undergo Cre-mediated Hus1 deletion, p53-deficiency delayed the clearance of Hus1-null cells from conditional Hus1 knockout mice and caused the accumulation of damaged, dying cells in the mammary gland. Notably, compensatory responses were impaired following combined Hus1 and p53 loss, resulting in aberrant mammary gland morphology and lactation defects. Overall, these results establish a requirement for Hus1 in the survival and proliferation of mammary epithelium and identify a role for p53 in mammary gland tissue regeneration and homeostasis. breast cancer | DNA damage checkpoint doi/10.1073/pnas.0904965106

Urodele amphibians and teleost fish regenerate amputated body parts via a process called epimorphic regeneration. A hallmark of this phenomenon is the reactivation of silenCed developmental regulatory genes that previously functioned during embryonic patterning. We demonstrate that histone modifications silence promoters of numerous genes involved in zebrafish caudal fin regeneration. Silenced developmental regulatory genes contain bivalent [me.sup.3]K4/[me.sup.3]K27 H3 histone modifications created by the concerted action of Polycomb (PcG) and Trithorax histone methyltransferases. During regeneration, this silent, bivalent chromatin is converted to an active state by loss of repressive [me.sup.3]K27 H3 modifications, occurring at numerous genes that appear to function during regeneration. Loss-of-function studies demonstrate a requirement for a [me.sup.3]K27 H3 demethylase during fin regeneration. These results indicate that histone modifications at discreet genomic positions may serve as a crucial regulatory event in the initiation of fin regeneration. chromatin | polycomb doi/10.1073/pnas.0904132106

Neurons in the central nervous system (CNS) lose their ability to regenerate early in development, but the underlying mechanisms are unknown. By screening genes developmentally regulated in retinal ganglion cells (RGCs), we identified Kruppel-like factor-4 (KLF4) as a transcriptional repressor of axon growth in RGCs and other CNS neurons. RGCs lacking KLF4 showed increased axon growth both in vitro and after optic nerve injury in vivo. Related KLF fatuity members suppressed or enhanced axon growth to differing extents, and several growth-suppressive KLFs were up-regulated postnatally, whereas growth-enhancing KLFs were down-regulated. Thus, coordinated activities of different KLFs regulate the regenerative capacity of CNS neurons. 10.1126/science.1175737

Regeneration requires specification of the identity of new tissues to be made. Whether this process relies only on intrinsic regulative properties of regenerating tissues or whether wound signaling provides input into tissue repatterning is not known. The head-versus-tail regeneration polarity decision in planarians, which requires Wnt signaling, provides a paradigm to study the process of tissue identity specification during regeneration. The Smed-wntP-1 gene is required for regeneration polarity and is expressed at the posterior pole of intact animals. Surprisingly, wntP-1 was expressed at both anterior- and posterior-facing wounds rapidly after wounding, wntP-1 expression was induced by all types of wounds examined, regardless of whether wounding prompted tail regeneration. Regeneration polarity was found to require new expression of wntP-1. Inhibition of the wntP-2 gene enhanced the polarity phenotype due to wntP-1 inhibition, with new expression of wntP-2 in regeneration occurring subsequent to expression of wntP-1 and localized only to posterior-facing wounds. New expression of wntP-2 required wound-induced wntP-1. Finally, wntP-1 and wntP-2 expression changes occurred even in the absence of neoblast stem cells, which are required for regeneration, suggesting that the role of these genes in polarity is independent of and instructive for tail formation. These data indicate that wound-induced input is involved in resetting the normal polarized features of the body axis during regeneration. anteroposterior axis | [beta]-catenin | planaria

A number of genes have been implicated in regeneration, but the regulation of these genes, particularly pertaining to regeneration in higher vertebrates, remains an interesting and mostly open question. We have studied microRNA (miRNA) regulation of regeneration and found that an intact miRNA pathway is essential for caudal fin regeneration in zebrafish. We also showed that miR-203 directly targets the Wnt signaling transcription factor Lef1 during this process. Repression of Lef1 by miR-203 blocks regeneration, whereas loss of miR-203 results in excess Lef1 levels and fin overgrowth. Expression of Lef1 from mRNAs lacking 3' UTR recognition elements can rescue the effects of excess miR-203, demonstrating that these effects are due to specific regulation of lef1 by miR-203. Our data support a model in which regulation of Lef1 protein levels by miR-203 is a key limiting step during regeneration. lef1 | miR-203

One goal of regenerative medicine is to instructively convert adult cells into other cell types for tissue repair and regeneration. Although isolated examples of adult cell reprogramming are known, there [...]

Regeneration in echinoderms has been extensively characterized physiologically, but fewer studies have addressed the genetic mechanisms underlying the process. Telomeres, genetic sequences on the ends of chromosomes that shorten with cell division, are associated with cell senescence, and thus genetic age after cell growth. Relative telomere lengths in the starfish Luidia clathrata were measured in arms of juveniles, adults prior to injury, and adults after injury during initial regeneration. Additionally, relative telomerase expression was measured in arms of adults prior to injury, at wound closure, and during initial regeneration. Telomeres were longer in juveniles than uninjured adults, indicating genetic aging. Telomeres elongated in adults after injury. Telomerase was detected before and after injury. The combination of genetic aging and telomerase expression both before and after injury suggests possible post-transcriptional regulation of telomerase. Additionally, as previous studies did not detect elongated telomeres in regenerated arms, it is possible that telomere elongation occurs early in the regenerative process as a mechanism to permit cellular division for growth without aging the final regenerated arm., Author(s): Rebecca M. Varney [sup.1] , Christopher M. Pomory [sup.1] , Alexis M. Janosik [sup.1] Author Affiliations: (1) Department of Biology, University of West Florida, 0000 0001 2112 2427, grid.267436.2, [...]

We identified a mechanism whereby retina regeneration in the embryonic chick can be induced by the contribution of stem/ progenitor cells. We show that bone morphogenetic protein (BMP) signaling is sufficient and necessary to induce retina regeneration and that its action can be divided into two phases. By 3 days after postretinectomy (d PR), the BMP pathway directs proliferation and regeneration through the activation of Smad (canonical BMP pathway) and the up-regulation of FGF signaling by the MAPK pathway. By 7d PR, it induces apoptosis by activating p38 (a noncanonical BMP pathway) and down-regulating FGF signaling (by both MAPK and AKT pathways). Apoptosis at this later stage can be prevented, and BMP-induced regeneration can be further induced by inhibition of p38. These results unravel a mechanism for stem/ progenitor cell-mediated retina regeneration, where BMP activation establishes a cross-talk with the FGF pathway and selectively activates the canonical and noncanonical BMP pathways. Retina stem/progenitor cells exist in other species, including humans. Thus, our findings provide insights on how retinal stem cells can be activated for possible regenerative therapies. p38 | FGF

A study investigating the types of genes responsible for regeneration and asexual budding in Hydra is presented. Evidence was obtained by embryological research.

Newly identified genetic elements help to replace missing tails and other body parts. Newly identified genetic elements help to replace missing tails and other body parts., Author Affiliations: A killifish regrew its tail only six days after the appendage was clipped at the brightly dotted horizontal line. Credit: Dr Wei Wang Regenerating tail of a male [...]

Postnatal neovascularization is essential for wound healing, cancer progression, and many other physiological functions. However, its genetic mechanism is largely unknown. In this report, we study neovascularization in regenerating adult zebrafish fins using transgenic fish that express EGFP in blood vessel endothelial cells. We first describe the morphogenesis of regenerating vessels in wild-type animals and then the phenotypic analysis of a genetic mutation that disrupts blood vessel regeneration. In wild-type zebrafish caudal fins, amputated blood vessels heal their ends by 24 h postamputation (hpa) and then reconnect arteries and veins via anastomosis, to resume blood flow at wound sites by 48 hpa. The truncated vessels regenerate by first growing excess vessels to form unstructured plexuses, resembling the primary capillary plexuses formed during embryonic vasculogenesis. Interestingly, this mode of vessel growth switches by 8 days postamputation (dpa) to growth without a plexus intermediate. During blood vessel regeneration, vessel remodeling begins during early plexus formation and continues until the original vasculature pattern is reestablished at ~35 dpa. Temperature-sensitive mutants for reg6 have profound defects in blood vessel regeneration. At the restrictive temperature, reg6 regenerating blood vessels first fail to make reconnections between severed arteries and veins, and then form enlarged vascular sinuses rather than branched vascular plexuses. Reciprocal temperature-shift experiments show that reg6 function is required throughout plexus formation, but not during later growth. Our results suggest that the reg6 mutation causes defects in branch formation and/or angiogenic sprouting. Keywords: Zebrafish; Regeneration; Angiogenesis; Plexus; Branching; Anastomosis

Spinal muscular atrophy (SMA), a common autosomal recessive form of motoneuron disease in infants and young adults, is caused by mutations in the survival motoneuron 1 (SMN1) gene. The corresponding gene product is part of a multiprotein complex involved in the assembly of spliceosomal small nuclear ribonucleoprotein complexes. It is still not understood why reduced levels of the ubiquitously expressed SMN protein specifically cause motoneuron degeneration. Here, we show that motoneurons isolated from an SMA mouse model exhibit normal survival, but reduced axon growth. Overexpression of Smn or its binding partner, heterogeneous nuclear ribonucleoprotein (hnRNP) R, promotes neurite growth in differentiating PC12 cells. Reduced axon growth in Smn-deficient motoneurons correlates with reduced [beta]-actin protein and mRNA staining in distal axons and growth cones. We also show that hnRNP R associates with the 3' UTR of [beta]-actin mRNA. Together, these data suggest that a complex of Smn with its binding partner hnRNP R interacts with [beta]-actin mRNA and translocates to axons and growth cones of motoneurons.

The regenerative capacity of tissues to recover from injury or stress is dependent on stem cell competence, yet the underlying mechanisms that govern how stem cells detect stress and initiate appropriate responses are poorly understood. In this issue of the JCI, Cho and Yusuf et al. demonstrate that the purinergic receptor [P2Y.sub.14] may mediate the hematopoietic stem and progenitor cell regenerative response., Senescence and stem cell decline Cellular senescence, a state of permanent irreversible growth arrest, was initially described over half a century ago by Leonard Hayflick and Paul Moorhead, who observed [...]

The T-domain transcription factors Tbx4 and Tbx5 have been implicated, by virtue of their limb-type specific expression, in controlling the identity of vertebrate legs and arms, respectively. To study the roles of these genes in developing and regenerating limbs, we cloned Tbx4 and Tbx5 cDNAs from the newt, and generated antisera that recognize Tbx4 or Tbx5 proteins. We show here that, in two urodele amphibians, newts and axolotls, the regulation of Tbx4 and Tbx5 differs from higher vertebrates. At the mRNA and protein level, both Tbx4 and Tbx5 are expressed in developing hindlimbs as well as in developing forelimbs. The coexpression of these genes argues that additional factors are involved in the control of limb type-specific patterns. In addition, newt and axolotl Tbx4 and Tbx5 expression is regulated differently during embryogenesis and regenerative morphogenesis. During regeneration, Tbx5 is exclusively upregulated in the forelimbs, whereas Tbx4 is exclusively upregulated in the hindlimbs. This indicates that, on a molecular level, different regulatory mechanisms control the shaping of identical limb structures and that regeneration is not simply a reiteration of developmental gene programs. Key Words: limb development; limb regeneration; pattern formation; Tbx4; Tbx5.

Recent studies suggest that tissue-specific stem cells possess much wider potential for differentiation than previously thought and can, in some instances, even cross germ layer boundaries. However, information is lacking regarding the efficiency and the fidelity of their differentiation along heterologous lineages. To address these issues of transdifferentiation, we have analyzed the heterologous potential of stem cells within the same germ layer. We report the neural potential of cells isolated from the limbal epithelium of the adult cornea. Limbal epithelium, which, like the neuroepithelium, is ectodermally derived, participates in the regeneration of cornea throughout life. We have observed that limbal epithelial cells, when removed from their niche and cultured in the presence of mitogens, begin to express neural progenitor markers. Based on the self-renewal property, it is likely that the nestin-positive progenitors are derived from limbal stem cells rather than transit-amplitying (TA) cells that have limited proliferating potential. In differentiation conditions, a subset of these cells acquire neural morphology and express transcripts and proteins specific to neurons and glia, suggesting their differentiation along neural lineage. The acquisition of neural properties is regulated by BMP signaling. Neural differentiation of these cells is also observed upon heterotopic transplantation. Investigation of functional differentiation of cells by electrophysiological analysis reveals properties consistent with the presence of glia that are influenced by extracellulat cues. However, similar analyses coupled with [Ca.sup.2+] imaging suggest an incomplete differentiation of limbal epithelial-derived neural progenitors into neurons in the condition studied. Our study, therefore, draws attention toward the necessity for rigorous characterization of transditterentiation and offers a model for characterizing neural potential of heterologous stem cells/progenitors. Key Words: neural stem cells; progenitors; limbal epithelium; reprogramming; transdifferentiation; BMP signaling.

In previous studies, we have shown that dorsoventral (DV) interaction evokes not only blastema formation, but also morphogenetic events similar to those that occur in regeneration. However, it is still unclear what kinds of signal molecules are involved in the DV interaction. To investigate the signal systems involved in the DV interaction, we focused on a noggin-like gene (Djnlg) identified by the planarian EST project. Djnlg is the first noggin homologue, isolated from an invertebrate. In DjNLG, the positions of nine cysteine residues which may be essential for dimer formation were well conserved, but overall, the amino acid sequence of DjNLG did not show high similarity to the sequences of vertebrate Noggins. Expression of Djnlg was observed only in the proximal region of the branch structures in the brain of intact planarians, suggesting that Djnlg may have a role in pattern formation in the brain. Interestingly, transient strong expression of Djnlg was observed in the amputated region of regenerating planarians. Djnlg-expressing cells were detected beneath the muscle 9 h after amputation and were then detected in the ventral subepidermal region of the blastema. The induction of Djnlg expression by amputation was not affected by X-ray irradiation, even though the stem cells were completely eliminated, implying the existence of signal-producing cells which may provide a positional cue to the stem cells. In DV reversed grafting, expression of Djnlg was strongly induced in the DV boundary between the host and donor. These results suggest that ectopic DV interaction may induce expression of Djnlg in the positional cue-producing cells, and that it might be involved in stimulation of blastema formation as well as DV patterning of the body. Key Words: planarian; noggin; regeneration; dorsoventral patterning; brain; BMP signal.

Understanding the gene-expression patterns during liver regeneration may help to reveal how regenerative processes are initiated and controlled as well as shed new light onto processes that lead to liver disease. Using high-density oligonucleotide arrays, we have examined the gene-expression program in the livers of mice after partial hepatectomy. A time course was constructed for gene expression between 0 and 4 h after partial hepatectomy, corresponding to the priming phase of liver regeneration. The genomic program for liver regeneration involves transcription-factor generation, stress and inflammatory responses, cytoskeletal and extracellular matrix modification, and regulation of cell-cycle entry. The genome-wide changes that are observed provide a detailed and comprehensive map of the initial priming stage of liver regeneration.

Research has been conducted on the effect of genetic and environmental seasonal variations on larval color in swallowtail butterfly. The hypothesis that larval color plasticity depends on its ability to increase larval growth rate in the autumn via raising larval body temperature has been tested and the details are presented.

The novel peptide HEADY of a simple, radially symmetric metazoan, Hydra, has been found to specify apical fate. HEADY is a short amidated peptide and the HEADY gene transcripts accumulate at the apical organizing center. If HEADY function is disrupted by dsRNA-mediated interference, severe head formation defects result. This study has given insight into how first asymmetry is specified in lower metazoans.

Metazoan regeneration is one of the least understood fundamental problems of biology. The lack of progress in understanding this phenomenon at the molecular level has been due to the poor regenerative abilities of the genetic organisms used for developmental studies, as well as the difficulties encountered with molecular and genetic manipulations of the commonly studied vertebrate models (the urodele amphibians). Here, we demonstrate that introduction of double-stranded RNA selectively abrogates gene function in planarians, a classic model of regeneration. The ability to eliminate gene function in a regenerating organism such as the planarian overcomes previous experimental limitations and opens the study of animal regeneration to unprecedented levels of molecular detail.

The assembly of genome sequences for the flatworm Schmidtea mediterranea and the salamander Ambystoma mexicanum will provide insights into the remarkable regenerative characteristics of these two organisms. The assembly of genome sequences for the flatworm Schmidtea mediterranea and the salamander Ambystoma mexicanum will provide insights into the remarkable regenerative characteristics of these two organisms., Author(s): G. Parker Flowers, Craig M. Crews Author Affiliations: Regeneration writ large Although humans have a limited ability to regenerate after injury, other animals can perform extraordinary regenerative feats. Small [...]

Hox genes play a critical role in the development of the vertebrate axis and limbs, and previous studies have implicated them in the specification of positional identity, the control of growth, and the timing of differentiation. Axolotl limbs offer an opportunity to distinguish these alternatives because the sequence of skeletal differentiation is reversed along the anterior-posterior axis relative to that of other tetrapods. We report that during early limb development, expression patterns of HoxD genes in axolotls resemble those in amniotes and anuran amphibians. At later stages, the anterior boundary of Hoxd-11 expression is conserved with respect to morphological landmarks, but there is no anterior-distal expansion of the posterior domain of Hoxd-11 expression similar to that observed in mice and chicks. Since axolotls do not form an expanded paddle-like handplate prior to digit differentiation, we suggest that anterior expansion of expression in higher vertebrates is linked to the formation of the handplate, but is clearly not necessary for digit differentiation. We also show that the 5[prime] HoxD genes are reexpressed during limb regeneration. The change in the expression pattern of Hoxd-11 during the course of regeneration is consistent with the hypothesis that the distal tip of the regenerate is specified first, followed by intercalation of intermediate levels of the pattern. Both Hoxd-8 and Hoxd-10 are expressed in non-regenerating wounds, but Hoxd-11 is specific for regeneration. It is also expressed in the posterior half of nerve-induced supernumerary outgrowths. Key Words: Hoxd-8; Hoxd-10; Hoxd-11; limb development; limb regeneration; handplate; digit; digit sequence.

Cellular senescence is defined by the limited proliferative capacity of normal cultured cells. Immortal cells overcome this regulation and proliferate indefinitively. One step in the immortalization process may be reactivation of telomerase activity, a ribonucleoprotein complex, which, by de novo synthesized telomeric TTAGGG repeats, can prevent shortening of the telomeres. Here we show that immortal human skin keratinocytes, irrespective of whether they were immortalized by simian virus 40, human papillomavirus 16, or spontaneously, as well as cell lines established from human skin squamous cell carcinomas exhibit telomerase activity. Unexpectedly, four of nine samples of intact human skin also were telomerase positive. By dissecting the skin we could show that the dermis and cultured dermal fibroblasts were telomerase negative. The epidermis and cultured skin keratinocytes, however, reproducibly exhibited enzyme activity. By separating different cell layers of the epidermis this telomerase activity could be assigned to the proliferative basal cells. Thus, in addition to hematopoietic cells, the epidermis, another example of a permanently regenerating human tissue, provides a further exception of the hypothesis that all normal human somatic tissues are telomerase deficient. Instead, these data suggest that in addition to contributing to the permanent proliferation capacity of immortal and tumor-derived keratinocytes, telomerase activity may also play a similar role in the lifetime regenerative capacity of normal epidermis in vivo.

Changes in mRNA stability regulate the expression of genes involved in liver regeneration. Experiments using rats also suggest that changes in mRNA stability of genes modulated beyond the G1 phase of the cell cycle are associated with changes in steady-state transcript levels. It is concluded that allelic genomic methylation influences the regulation of transcript stability during liver regeneration.

A study was conducted to examine the processes involved in fin regeneration in amputated fins of adult zebrafish. The investigation focused on the stages of wound healing, establishment of the wound epithelium, recruitment of the blastema for mesenchymal cells underlying the wound epithelium and on differentiatio and outgrowth of the regenerate tissue.

Neurogeneticists exploring various techniques for the repair of myelin in people with central nervous system demyelinating disorders have begun focusing their research efforts on the usefulness of gene transfer. The trend can be attributed largely to the successful characterization of the molecular mechanisms governing demyelination, the identification and cloning of genes which cause leukodystrophies and the development of recombinant growth factors which affect the development of myelin-forming cells.