Regeneration Bud Research Articles

Regeneration factors expressed on myeloid expression in macrophage-like cells is required for tail regeneration in Xenopus laevis tadpoles.

Xenopus laevis tadpoles can regenerate whole tails after amputation. We have previously reported that interleukin 11 (il11) is required for tail regeneration. In this study, we have screened for genes that support tail regeneration under Il11 signaling in a certain cell type and have identified the previously uncharacterized genes Xetrov90002578m.L and Xetrov90002579m.S [referred to hereafter as regeneration factors expressed on myeloid.L (rfem.L) and rfem.S]. Knockdown (KD) of rfem.L and rfem.S causes defects of tail regeneration, indicating that rfem.L and/or rfem.S are required for tail regeneration. Single-cell RNA sequencing (scRNA-seq) revealed that rfem.L and rfem.S are expressed in a subset of leukocytes with a macrophage-like gene expression profile. KD of colony-stimulating factor 1 (csf1), which is essential for macrophage differentiation and survival, reduced rfem.L and rfem.S expression levels and the number of rfem.L- and rfem.S-expressing cells in the regeneration bud. Furthermore, forced expression of rfem.L under control of the mpeg1 promoter, which drives rfem.L in macrophage-like cells, rescues rfem.L and rfem.S KD-induced tail regeneration defects. Our findings suggest that rfem.L or rfem.S expression in macrophage-like cells is required for tail regeneration.

Xvent-2 expression in regenerating Xenopus tails.

The tail of Xenopus tadpole is an excellent model for appendage regeneration studies. We analyzed the distribution pattern of the transcription factor Xvent-2 mRNA and protein in the beginning of the regeneration of Xenopus tadpole tail stumps after amputation. We revealed the emergence of Xvent-2 mRNA and protein in regeneration bud during the first day after amputation. The data obtained confirm that soon after amputation of the part of the Xenopus tadpole tail, there occurs the emergence of a structure, to some extend, resembling the early embryonic tail bud.

Role of TrkA signaling during tadpole tail regeneration and early embryonic development in Xenopus laevis.

Neurotrophic signaling regulates neural cell behaviors in development and physiology, although its role in regeneration has not been fully investigated. Here, we examined the role of neurotrophic signaling in Xenopus laevis tadpole tail regeneration. After the tadpole tails were amputated, the expression of neurotrophin ligand family genes, especially ngf and bdnf, was up-regulated as regeneration proceeded. Moreover, notochordal expression of the NGF receptor gene TrkA, but not that of other neurotrophin receptor genes TrkB and TrkC, became prominent in the regeneration bud, a structure arising from the tail stump after tail amputation. The regenerated tail length was significantly shortened by the pan-Trk inhibitor K252a or the TrkA inhibitor GW-441756, but not by the TrkB inhibitor ANA-12, suggesting that TrkA signaling is involved in elongation of regenerating tails. Furthermore, during Xenopus laevis embryonic development, TrkA expression was detected in the dorsal mesoderm at the gastrula stage and in the notochord at the neurula stage, and its knockdown led to gastrulation defects with subsequent shortening of the body axis length. These results suggest that Xenopus laevis TrkA signaling, which can act in the mesoderm/notochord, plays a key role in body axis elongation during embryogenesis as well as tail elongation during tadpole regeneration.

Regeneration of the acorn worm pygochord with the implication for its convergent evolution with the notochord.

The origin of the notochord is a central issue in chordate evolution. This study examined the development of the acorn worm pygochord, a putative homologue of the notochord. Because the pygochord differentiates only after metamorphosis, the developmental was followed process by inducing regeneration after artificial amputation in Ptychodera flava. It was found that although the regeneration of the posterior part of the body did not proceed via formation of an obvious regeneration bud, pygochord regeneration was observed within a few weeks, possibly via trans-differentiation of endoderm cells. The expression of the fibrillary collagen gene (Fcol) and elav in the pygochord during regeneration was detected. This indicates that pygochord cells are not part of gut epithelial cells, but that they differentiated as a distinct cell type. Our gene expression analyses do not provide supporting evidence for the homology between the pygochord and notochord, but rather favored the convergent evolution between them.

Early redox activities modulate Xenopus tail regeneration

Redox state sustained by reactive oxygen species (ROS) is crucial for regeneration; however, the interplay between oxygen (O2), ROS and hypoxia-inducible factors (HIF) remains elusive. Here we observe, using an optic-based probe (optrode), an elevated and steady O2 influx immediately upon amputation. The spatiotemporal O2 influx profile correlates with the regeneration of Xenopus laevis tadpole tails. Inhibition of ROS production but not ROS scavenging decreases O2 influx. Inhibition of HIF-1α impairs regeneration and stabilization of HIF-1α induces regeneration in the refractory period. In the regeneration bud, hypoxia correlates with O2 influx, ROS production, and HIF-1α stabilization that modulate regeneration. Further analyses reveal that heat shock protein 90 is a putative downstream target of HIF-1α while electric current reversal is a de facto downstream target of HIF-1α. Collectively, the results show a mechanism for regeneration via the orchestration of O2 influx, ROS production, and HIF-1α stabilization.

The Influence of the Xymedon Preparation on the Regeneration of Girardia tigrina Planarians

Synthesis of new medicines and analysis of their effects have become increasingly important lately. Hepatoprotectors is a general name of medicinal preparations for liver treatment and recovery. The Xymedon preparation (Hydroxyethyldimethyldihydropyrimidine), an immunostimulant, can be presumably used as a hepatoprotector. To date, the influence of this preparation has been insufficiently studied. The purpose of this paper is to investigate the effect of the pyrimidine derivative Xymedon on the regeneration of Girardia tigrina (Girard, 1850) planarians, one of the perspective models for analysis of the influence produced by various substances and preparations on proliferation rates of the regeneration bud (blastema). The study revealed that the Xymedon preparation is not toxic for G. tigrina planarians and does not cause their death even when its concentration is gradually increased in the solution. The selected working concentrations of the Xymedon preparation produced different effects on planarians during the experiment. The influence of small concentrations (0.0004 and 0.004 mg) was little different from that in the control group, i.e., the regeneration criterion was low. Higher concentrations (0.1 mg) stimulated regeneration processes, i.e., the regeneration criterion was high. The largest concentration of the Xymedon preparation (0.4 mg) led to a slight decrease of the regeneration criterion, being, however, characterized by the high regeneration coefficient.

Estimation of the toxicity of silver nanoparticles by using planarian flatworms.

The regeneration of planarian flatworms - specifically, changes to the area of the regeneration bud (blastema) after surgical dissection - was proposed for use as a robust tool for estimating the toxicity of silver nanoparticles. The use of Planaria species, due to their unique regenerative capacity, could result in a reduction in the use of more-traditional laboratory animals for toxicity testing. With our novel approach, silver nanoparticles were found to be moderately toxic to the planarian, Girardia tigrina.

Physiological basis of effective controlling Torpedo grass (Panicum repens L.)

Torpedo grass (Panicum repens L.) is a C4 plant species, present on different soil types. The rhizome system and tubers of grass well developed, although flowering but not yet recorded the presence of seeds and seedlings (Yêu cầu tác giả viết lại toàn bộ câu này, không thể sửa được. Lưu ý câu phải có động từ chính). Tuber has high drought tolerance. Regenerative ability of the grass decreases with water content of the tuber. Tuber inability to regenerate shoots when it has water less than 30% of water at first. Repeatedly cut grass will take the tuber reserve depletion, not sufficient to provide for the regeneration bud sprouts. Coordinate disposal of mowing and using systemic herbicides will lead to better results. 6-8 weeks after cutting, the grass grow well with multiple shoots, spraying glyphosate 480 SL or glyphosate trimethyl sulphonium (GTS) at doses of 6-8 l / ha, mixed with urea at concentrations from 1% to 1.5%. This way helps prevent the emergence of shoots from rhizomes and tubers.

Ecological characteristics and distribution of Asarum balansae Franch. in Ban Bung, Na Hang district, Tuyen Quang province

Asarum balansae Franch. is a perennial herbaceous plant, which began flowering from early October; flowers bloom in early December; ripe fruit and seed dispersal in July - August in the follwing year. Regeneration density of the plant was observed highest at the foot-hill of mountain (from 46,250 to 50,000 trees/ha), the plant originated from regeneration bud are dominant (from 76.6 to 97.5%), renewable particles have the relative low percentage (from 2.5 to 23.4%). At the mountain sides, the Asarum balansae is rarely observed, the regeneration desity is relatively low (from 0 to 417 trees/ha). The quality of regeneration is unequally observed in different areas: percentage of good, moderate and bad trees are 10 to 76.6%, 16.2 to 39% and 7.2 - 51%, res pectively. Asarum balansae distributes in the rain tropical forest in low land, altitude from 201 m to 600 m, concentrated in the valley along the trail at the foot-hill of mountain, they grow close to the soil surface, in the rich soil with humus; humidity, light porosity and high airflow; less coverage. The average distribution density is 42,083 trees/ha in forest structure comprising one timber floor at the foot-hill of mountain and two timber floor in mountain sides.

Long-Distance Signals Are Required for Morphogenesis of the Regenerating Xenopus Tadpole Tail, as Shown by Femtosecond-Laser Ablation

BackgroundWith the goal of learning to induce regeneration in human beings as a treatment for tissue loss, research is being conducted into the molecular and physiological details of the regeneration process. The tail of Xenopus laevis tadpoles has recently emerged as an important model for these studies; we explored the role of the spinal cord during tadpole tail regeneration.Methods and ResultsUsing ultrafast lasers to ablate cells, and Geometric Morphometrics to quantitatively analyze regenerate morphology, we explored the influence of different cell populations. For at least twenty-four hours after amputation (hpa), laser-induced damage to the dorsal midline affected the morphology of the regenerated tail; damage induced 48 hpa or later did not. Targeting different positions along the anterior-posterior (AP) axis caused different shape changes in the regenerate. Interestingly, damaging two positions affected regenerate morphology in a qualitatively different way than did damaging either position alone. Quantitative comparison of regenerate shapes provided strong evidence against a gradient and for the existence of position-specific morphogenetic information along the entire AP axis.ConclusionsWe infer that there is a conduit of morphology-influencing information that requires a continuous dorsal midline, particularly an undamaged spinal cord. Contrary to expectation, this information is not in a gradient and it is not localized to the regeneration bud. We present a model of morphogenetic information flow from tissue undamaged by amputation and conclude that studies of information coming from far outside the amputation plane and regeneration bud will be critical for understanding regeneration and for translating fundamental understanding into biomedical approaches.

Molecular cloning and expression of visinin-like protein 1 from Gekko japonicus spinal cord

Visinin-like protein 1 (VILIP-1), a myristoylated calcium sensor protein of the EF-hand Ca2+-binding protein superfamily, plays multiple physiological roles in the central nervous system and peripheral organs. In present study, the cDNA encoding VILIP-1 was identified from the brain and spinal cord cDNA library of Gekko japonicus. It contains a 573 bp open reading frame corresponding to a deduced protein of 191 amino acids. Gecko VILIP-1 shares more than 95.3% identity with vertebrate VILIP-1 proteins, and structurally consists of conserved four EF-hand Ca2+-binding motifs and one dsRNA-binding domain, suggesting that selective pressure must have been extremely high for the conservation of VILIP-1 during vertebrate evolution. Northern blot and RT-PCR showed that gecko VILIP-1 was ubiquitously expressed in all tissues examined. In situ hybridization revealed that the VILIP-1 transcript mainly appeared in the gray matter of the spinal cord, with less distribution in the white matter. Semiquantitative RT-PCR also showed that VILIP-1 expression in spinal cord after tail amputation remained stable at 1 day and 1 week, but decreased at 2 weeks, a time coinciding with regeneration bud formation. This suggests that VILIP-1 may function as a regeneration-associated factor in the form of a monomer or/and RNA-binding complex.

NaV-Mediated Sodium Currents Are Necessary For Vertebrate Appendage Regeneration

Mammals have a limited ability to regenerate tissues. In contrast, amphibians such as frogs can restore lost developmental structures, including the lens and tail. A detailed understanding of natural regeneration is important for developing therapies for organ repair. Recently, ion transport has been implicated as a regulator of regeneration. While voltage-gated sodium channels play a well-known and important role in propagating action potentials in excitable cells, we have identified a novel role in regeneration for the ion transport function mediated by the voltage-gated sodium channel, NaV1.2. After Xenopus tadpole tail amputation, a regeneration bud (containing progenitors required for regenerative growth) is formed within 1 day at the injury site, and a new tail is re-grown by 7 days. NaV1.2 is expressed early in the bud, and its function is required for regeneration. Inhibition of its activity causes regenerative failure by greatly reducing expression of downstream genes that drive tail outgrowth and patterning, leading to decreased proliferation and altered axonal patterning in the regeneration bud. Significantly, NaV1.2 is not expressed under non-regenerative conditions, suggesting that its activity is a determinant of regenerative ability. Most importantly, pharmacological induction of a brief, transient sodium current into the regeneration bud after tail amputation is sufficient to restore full regeneration of the tail during the refractory period (an endogenous developmental period when regeneration is blocked). Our study demonstrates that sodium transport is a critical mechanism for controlling regeneration, and suggests that short-term modulations of ion transport could represent an exciting new approach to tissue repair in mammals.

TGF-β signaling is required for multiple processes during Xenopus tail regeneration

Xenopus tadpoles can fully regenerate all major tissue types following tail amputation. TGF-β signaling plays essential roles in growth, repair, specification, and differentiation of tissues throughout development and adulthood. We examined the localization of key components of the TGF-β signaling pathway during regeneration and characterized the effects of loss of TGF-β signaling on multiple regenerative events. Phosphorylated Smad2 (p-Smad2) is initially restricted to the p63+ basal layer of the regenerative epithelium shortly after amputation, and is later found in multiple tissue types in the regeneration bud. TGF-β ligands are also upregulated throughout regeneration. Treatment of amputated tails with SB-431542, a specific and reversible inhibitor of TGF-β signaling, blocks tail regeneration at multiple points. Inhibition of TGF-β signaling immediately following tail amputation reversibly prevents formation of a wound epithelium over the future regeneration bud. Even brief inhibition immediately following amputation is sufficient, however, to irreversibly block the establishment of structures and cell types that characterize regenerating tissue and to prevent the proper activation of BMP and ERK signaling pathways. Inhibition of TGF-β signaling after regeneration has already commenced blocks cell proliferation in the regeneration bud. These data reveal several spatially and temporally distinct roles for TGF-β signaling during regeneration: (1) wound epithelium formation, (2) establishment of regeneration bud structures and signaling cascades, and (3) regulation of cell proliferation.

The Spark of Life: Electricity and Regeneration

A recent paper shows that a proton pump, the vacuolar-type adenosine triphosphatase (V-ATPase), is necessary for Xenopus tadpole tail regeneration. The V-ATPase is synthesized in distal cells of the regeneration bud as an early response to amputation, and its role in regeneration is specifically to pump H+ out of cells. Although the downstream effects of proton movement are not known, they may be mediated through changes in membrane potential. This work could reopen a consideration of the role of electric fields in regeneration, an issue that has caused much controversy in the past.

Making a New Tail

Adams et al . identified the V-ATPase H + pump in a pharmacological screen for ion transporters involved in Xenopus larval tail regeneration. When the pumping activity of the V-ATPase was inhibited with concanamycin, tail regeneration was inhibited, whereas primary tail development and wound healing were unaffected. Using a dominant-negative V-ATPase, the same inhibition of regeneration was observed, confirming V-ATPase as the target. Expression of a yeast P-type H + pump, which is insensitive to concanamycin, was localized to the plasma membrane and also prevented concanamycin inhibition of regeneration, confirming that it was transport of H + efflux at the plasma membrane that was important for regeneration. Six hours after amputation, the abundance of the mRNA for the c subunit of the V-ATPase had increased, and by 24 hours after amputation the H + pump was abundant in the plasma membrane of the cells of the regeneration bud. The cells of the regeneration bud were depolarized six hours after amputation; however, by 24 hours after amputation, the cells were repolarized, which is consistent with the hyperpolarizing activity of the H + pump. Furthermore, depolarization of the tails using a drug that converts the Na + ,K + ATPase into a Na + /K + channel also inhibited regeneration, supporting the concept that membrane voltage is a critical regulator of regeneration. Inhibition of the H + pump decreased the number of proliferating cells in the regeneration bud and inhibited the expression of the KCNK1 gene, which encodes a K + channel that is expressed at early stages of regeneration (12 hours after amputation). Axon patterning in the regeneration bud was also aberrant when the H + pump activity was inhibited. Thus, H + pump activity contributes to changes in membrane potential that are important for regulation of gene expression, cell proliferation, and proper patterning of the new tissues. D. S. Adams. A. Masi, M. Levin, H + pump-dependent changes in membrane voltage are an early mechanism necessary and sufficient to induce Xenopus tail regeneration. Development 134 , 1323-1335 (2007). [Abstract] [Full Text]

Apoptosis is required during early stages of tail regeneration in Xenopus laevis

The Xenopus tadpole is able to regenerate its tail, including skin, muscle, notochord, spinal cord and neurons and blood vessels. This process requires rapid tissue growth and morphogenesis. Here we show that a focus of apoptotic cells appears in the regeneration bud within 12 h of amputation. Surprisingly, when caspase-3 activity is specifically inhibited, regeneration is abolished. This is true of tails both before and after the refractory period. Programmed cell death is only required during the first 24 h after amputation, as later inhibition has no effect on regeneration. Inhibition of caspase-dependent apoptosis results in a failure to induce proliferation in the growth zone, a mispatterning of axons in the regenerate, and the appearance of ectopic otoliths in the neural tube, in the context of otherwise normal continued development of the larva. Larvae amputated during the refractory stage exhibit a much broader domain of caspase-3-positive cells, suggesting a window for the amount of apoptosis that is compatible with normal regeneration. These data reveal novel roles for apoptosis in development and indicate that a degree of apoptosis is an early and obligate component of normal tail regeneration, suggesting the possibility of the existence of endogenous inhibitory cells that must be destroyed by programmed cell death for regeneration to occur.

Temporal requirement for bone morphogenetic proteins in regeneration of the tail and limb of Xenopus tadpoles

Bone morphogenetic protein (BMP) signalling is necessary for both the development of the tail bud and for tail regeneration in Xenopus laevis tadpoles. Using a stable transgenic line in which expression of the soluble BMP inhibitor noggin is under the control of the temperature inducible hsp70 promoter, we have investigated the timing of the requirement for BMP signalling during tail regeneration. If noggin expression is induced followed by partial amputation of the tail, then wound closure and the formation of the neural ampulla occur normally but outgrowth of the regeneration bud is inhibited. Furthermore, we show that BMP signalling is also necessary for limb bud regeneration, which occurs in Xenopus tadpoles prior to differentiation. When noggin expression is induced, limb bud regeneration fails at an early stage and a stump is formed. The situation appears similar to the tail, with formation of the limb bud blastema occurring but renewed outgrowth inhibited. The transcriptional repressor Msx1, a direct target of BMP signalling with known roles in vertebrate appendage regeneration, fails to be re-expressed in both tail and limb in the presence of noggin. DNA labelling studies show that proliferation in the notochord and spinal cord of the tail, and of the blastema in the limb bud, is significantly inhibited by noggin induction, suggesting that in the context of these regenerating appendages BMP is mainly required, directly or indirectly, as a mitogenic factor.

Effects of amputation and subsequent regeneration of a leg on the duration of the intermoult period and the level of circulation ecdysteroids in Helleria brevicornis Ebner (ground isopod).

Amputation of one or two limbs from the ground isopod, Helleria brevicornis, carried out during stages C and D0 of the molting cycle, causes the cycle to be significantly longer. On the other hand, if amputation takes place during stages A-B, D1, or D2, there is no noticeable change in the duration of the intermolt period. Assays for ecdysteroids in the hemolymph demonstrate that amputation always brings on a drop in hormone concentration. About 18 days are required for the initial concentration to be restored after amputation during stage C. This coincides with the cell proliferation of the healing epidermis and development of the regeneration bud, which are indicative of the end of the “latency period” that follows the stump's healing. The existence of this period and the perturbations of hormone titer are probably at least partly responsible for the longer intermolt period. Ecdysis is also delayed after amputations during stage D0. In this case, the much longer duration of the stage and the drop in hormonal level, are likely preconditions for regeneration prior to the first postoperative ecdysis. By contrast, the very short-lived drop (24 hr) of hormonal level caused by amputations during stage D1 is not long enough to interfere with the preparation of the molt, which takes place about on schedule and only gives a healed stump.

Studies onSargassum II. Quantitative ultrastructural changes in differentiated stipe cells during wound regeneration and regrowth

Quantitative electron micrograph analytical techniques were used to study cytological changes associated with the process of dedifferentiation and redifferentiation in mature, specialized cells of the stipe in the brown algaSargassum filipendula following wounding. Early cytological changes associated with this process appear to be those involved with formation of a protective layer of material at the wound surface. An increase in the volume of cytoplasm occupied by mitochondria, dictyosomes, and type A and B vesicles was evident by three days postwound. Cytoplasmic volume occupied by chloroplasts decreased during the early stages of wound reaction while the development of internal membranes (thylakoids) in these organelles increased. Cells at the cut surface eventually differentiated into one of two types of tissues: 1. A wound surface epidermal layer similar in cytological aspect to the epidermal layers of the intact and regenerated plant. 2. A meristematic “regeneration bud” from which developed the blade-like regeneration growth. The morphology and anatomy of the tissues of the regeneration blade resembled those of the blade of the intact plant rather than those of the stipe from which it originated. Percent difference values were obtained by comparing cytological features of the tissues from the regeneration blade with those of the intact stipe and blade. The comparisons supported previously obtained data indicating that the regeneration blade should be considered as a typical blade organ rather than a morphological variant of the stipe. Results of this study raises questions about possible physiological differences which may exist between stipe and blade organs in this plant.

Studies on pteridines during tail regeneration in amphibia

1. 1. A comparative study of pteridine distribution and pattern, during Triturus cristatus tail regeneration was made, in order to investigate the place of tetrahydrobiopterin synthesis. 2. 2. The experiments show that tetrahydrobiopterin synthesis does not occur in the regenerating blastema, but in the rest of the body and then, this pteridine which is an important regulator of cell division, is transported into the regeneration bud.