Regeneration In Animals Research Articles

A novel Axin2 knock-in mouse model for visualization and lineage tracing of WNT/CTNNB1 responsive cells.

SummaryWnt signal transduction controls tissue morphogenesis, maintenance and regeneration in all multicellular animals. In mammals, the WNT/CTNNB1 (Wnt/β‐catenin) pathway controls cell proliferation and cell fate decisions before and after birth. It plays a critical role at multiple stages of embryonic development, but also governs stem cell maintenance and homeostasis in adult tissues. However, it remains challenging to monitor endogenous WNT/CTNNB1 signaling dynamics in vivo. Here, we report the generation and characterization of a new knock‐in mouse strain that doubles as a fluorescent reporter and lineage tracing driver for WNT/CTNNB1 responsive cells. We introduced a multi‐cistronic targeting cassette at the 3′ end of the universal WNT/CTNNB1 target gene Axin2. The resulting knock‐in allele expresses a bright fluorescent reporter (3xNLS‐SGFP2) and a doxycycline‐inducible driver for lineage tracing (rtTA3). We show that the Axin2 P2A‐rtTA3‐T2A‐3xNLS‐SGFP2 strain labels WNT/CTNNB1 responsive cells at multiple anatomical sites during different stages of embryonic and postnatal development. It faithfully reports the subtle and dynamic changes in physiological WNT/CTNNB1 signaling activity that occur in vivo. We expect this mouse strain to be a useful resource for biologists who want to track and trace the location and developmental fate of WNT/CTNNB1 responsive stem cells in different contexts.

Endothelial Senescence in Regenerative Lung Growth after Pneumonectomy

Impairment of lung regeneration and repair in aging animals contributes to the pathogenesis of aging‐associated diseases, including chronic obstructive pulmonary disease (COPD). Angiogenesis – the formation of new blood capillaries ‐ plays a key role in organ regeneration. We have reported that inhibition of angiogenesis attenuates lung growth after unilateral pneumonectomy (PNX) and that post‐PNX regenerative lung growth is inhibited in aged mice. While accumulation of senescent cells in aged adults is linked to the age‐related decline in regenerative ability, senescent cells also play important roles in injury repair and tissue remodeling. Here we investigate the role of endothelial cell (EC) senescence in regenerative lung growth after PNX in young vs. aged mice. The levels of the senescence marker, p16INK4A and a major angiogenic factor, angiopoietin2 (Ang2) transiently increase in ECs isolated from 2‐months (2M) old post‐PNX mouse lungs compared to those from sham‐operated control mice and decrease at the later stage. The baseline levels of p16INK4A are high in 24M old mouse lung ECs but post‐PNX transient induction of p16INK4A and Ang2 is inhibited in 24M old mouse lung ECs. Depletion of senescent cells from 2M old p16‐3MR mice by treating with ganciclovir suppresses post‐PNX lung growth. When ECs isolated from 2M old post‐PNX mouse lungs are mixed in the gel and implanted on the mouse lungs, vascular network formation is stimulated in the gel, while these effects are inhibited in the gel mixed with 24M old post‐PNX mouse lung ECs. These results suggest that post‐PNX transient induction of EC senescence is required for vascular morphogenesis after PNX. Deregulation of this mechanism may contribute to age‐dependent inhibition of regenerative lung growth. Understanding this mechanism will lead to the development of efficient strategy for aging‐associated lung diseases.Support or Funding InformationNIH R21AG054830, NIH R21AG062893, NIH R01HL142578, AHA 18TPA34170129

Dissecting the Molecular Signature of Spinal Cord Regeneration in the Axolotl Model.

Thousands of people are affected by central nervous system (CNS) dysfunctions each year, with stroke and spinal cord injury (SCI) being the most frequent causes. Although there is some evidence of partial CNS self-repair (via migration of neural stem cells to the injury zone and adult neurogenesis), due to restricted regeneration capacity in mammals, acute or chronic spinal cord injuries cannot be repaired completely. Therefore, to expand the availability of treatment options for SCI, research on highly regenerative animals has become essential. Among vertebrates, axolotl, a salamander species, has been emerging as a powerful model to explore the molecular mechanisms of regeneration due to its exceptional regenerative capacity. In this study, gene expression modulation for regenerative-capable neotenic axolotl during spinal cord regeneration has been investigated. Next-generation sequencing was applied for the collected regeneration samples at zero and seven days post-amputation (dpa). The data obtained from the analyzed samples revealed 363 genes differentially expressed, mostly downregulated, between zero dpa and seven dpa. The extracellular matrix, cell-cell adhesion, and immune system-related processes and pathways were enriched by gene ontology and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Based on these data, we conclude that the downregulation of immune system-related biological processes is crucial for spinal cord regeneration.

The role of fasting on spine regeneration and bacteremia in the purple sea urchin Strongylocentrotus purpuratus.

Fasting has been shown to increase longevity and alter immune function in a variety of animals, but little is understood about how reduced caloric intake may impact regeneration and infections in animals that must regularly repair and regenerate tissue in marine environments that contain high levels of bacteria. We examined the possibility that fasting could enhance spine regeneration and reduce bacteremia in the purple sea urchin Strongylocentrotus purpuratus. A small number of spines were removed from urchins and rates of spine regrowth and levels of culturable bacteria from the coelomic fluid were measured for 21 days in fed and fasted urchins. Fasted urchins had higher rates of spine regrowth and lower levels of colony-forming units (CFU) per milliliter of coeolomic fluid. The predominant bacteria in the coelomic fluid was isolated and identified by DNA sequence-based methods as Vibrio cyclitrophicus. After 21 days, fasted and fed urchins were injected with V. cyclitrophicus. Two hours after injection, fed urchins had about 25% more culturable bacteria remaining in their coelomic fluid compared to fasted urchins. We found no evidence that fasting altered coelomic fluid cell number or righting response, indicators of physiologic and behavioral stress in urchins. Our results demonstrate that V. cyclitrophicus is present in purple urchin coelomic fluid, that fasting can increase spine regeneration and that fasted urchins have much lower levels of culturable bacteria in their coelomic fluid than fed urchins. Overall, our data suggests that fasting may ultimately reduce bacteremia and infection in injured or damaged urchins.

Markers of liver regeneration\u2014the role of growth factors and cytokines: a systematic review

BackgroundPost-hepatectomy liver failure contributes significantly to postoperative mortality after liver resection. The prediction of the individual risk for liver failure is challenging. This review aimed to provide an overview of cytokine and growth factor triggered signaling pathways involved in liver regeneration after resection.MethodsMEDLINE and Cochrane databases were searched without language restrictions for articles from the time of inception of the databases till March 2019. All studies with comparative data on the effect of cytokines and growth factors on liver regeneration in animals and humans were included.ResultsOverall 3.353 articles comprising 40 studies involving 1.498 patients and 101 animal studies were identified and met the inclusion criteria. All included trials on humans were retrospective cohort/observational studies. There was substantial heterogeneity across all included studies with respect to the analyzed cytokines and growth factors and the described endpoints.ConclusionHigh-level evidence on serial measurements of growth factors and cytokines in blood samples used to predict liver regeneration after resection is still lacking. To address the heterogeneity of patients and potential markers, high throughput serial analyses may offer a method to predict an individual’s regenerative potential in the future.

The Eupentacta fraudatrix transcriptome provides insights into regulation of cell transdifferentiation

The holothurian Eupentacta fraudatrix is a unique organism for studying regeneration mechanisms. Moreover, E. fraudatrix can quickly restore parts of its body and entire organ systems, yet at the moment, there is no data on the participation of stem cells in the process. To the contrary, it has been repeatedly confirmed that this process is only due to the transformation of terminally differentiated cells. In this study, we examine changes in gene expression during gut regeneration of the holothurian E. fraudatrix. Transcriptomes of intestinal anlage of the three stages of regeneration, as well as the normal gut, were sequenced with an Illumina sequencer (San Diego, CA, USA). We identified 14,617 sea urchin protein homologs, of which 308 were transcription factors. After analysing the dynamics of gene expression during regeneration and the map of biological processes in which they participate, we identified 11 factors: Ef-EGR1, Ef-ELF, Ef-GATA3, Ef-ID2, Ef-KLF1/2/4, Ef-MSC, Ef-PCGF2, Ef-PRDM9, Ef-SNAI2, Ef-TBX20, and Ef-TCF24. With the exception of TCF24, they are all involved in the regeneration, development, epithelial-mesenchymal transition, and immune response in other animals. We suggest that these transcription factors may also be involved in the transdifferentiation of coelomic epithelial cells into enterocytes in holothurians.

GLI STUDI DI ALDO PERRONCITO SULLA RIGENERAZIONE DEL NERVO PERIFERICO

Animal regeneration was a major subject of investigation in the 18th century Europe. Lazzaro Spallanzani was one of the most active experimenters, he was able to demonstrate the regenerative abilities of many invertebrates and even some vertebrates. At the beginning of the 20th century in the General Pathology laboratory of the University of Pavia directed by Camillo Golgi, a young researcher, Aldo Perroncito, succeeded for the first time in understanding and describing the process of regeneration of peripheral nerves after experimental cut. His research had a great impact also for the subsequent surgical applications. Perroncito’s studies paved the way for the first attempts at operations on patients wounded during the World War I for the functional restoration of the injured nerves conducted by the University of Pavia graduated medical doctors Giovanni Verga and Guido Sala.

Pathogenetic substantiation of the combined transplantation use of multipotent mesenchymal stromal cells and hepatic stellate cells to restore the liver morphofunctional state after acute toxic hepatitis in the old body

The purpose of this study was to study the cotransplantation influence of multipotent mesenchymal stromal (MMSC) and hepatic stellate (HSC) cells on liver regeneration of old laboratory animals in conditions of its toxic damage. Acute toxic hepatitis was caused by single intraperitoneal CC14 injection at a dose of 50 μg/kg. The introduction of MMSC and HSC was carried out at doses of 4 million cl/kg and 9 million cl/kg respectively 1 hour after toxic hepatitis modelling. The morphofunctional liver state of old laboratory mice was evaluated on the 1st, 3rd, 7th day after combined injection of MMSC and HSC in laboratory animals with toxic hepatitis. As a result of the study, it was obtained that MMSC and HSC cotransplantation leads to cellular and intracellular liver regeneration activation in old mice with acute toxic hepatitis. Also, the introduction of these cell types leads to decreased liver mutagenesis, inhibition of programmed cellular hepatocytes death. Thus, the conducted studies indicate the ability of combined MMSC and HSC transplantation to restore the morphofunctional liver state of the old organism under the conditions of its toxic damage.

Transdifferentiation and mesenchymal-to-epithelial transition during regeneration in Demospongiae (Porifera).

Origin and early evolution of regeneration mechanisms remain among the most pressing questions in animal regeneration biology. Porifera have exceptional regenerative capacities and, as early Metazoan lineage, are a promising model for studying evolutionary aspects of regeneration. Here, we focus on reparative regeneration of the body wall in the Mediterranean demosponge Aplysina cavernicola. The epithelialization of the wound surface is completed within 2 days, and the wound is completely healed within 2 weeks. The regeneration is accompanied with the formation of a mass of undifferentiated cells (blastema), which consists of archaeocytes, dedifferentiated choanocytes, anucleated amoebocytes, and differentiated spherulous cells. The main mechanisms of A. cavernicola regeneration are cell dedifferentiation with active migration and subsequent redifferentiation or transdifferentiation of polypotent cells through the mesenchymal-to-epithelial transformation. The main cell sources of the regeneration are archaeocytes and choanocytes. At early stages of the regeneration, the blastema almost devoid of cell proliferation, but after 24 hr postoperation (hpo) and up to 72 hpo numerous DNA-synthesizing cells appear there. In contrast to intact tissues, where vast majority of DNA-synthesizing cells are choanocytes, all 5-ethynyl-2'-deoxyuridine-labeled cells in the blastema are mesohyl cells. Intact tissues, distant from the wound, retains intact level of cell proliferation during whole regeneration process. For the first time, the apoptosis was studied during the regeneration of sponges. Two waves of apoptosis were detected during A. cavernicola regeneration: The first wave at 6-12 hpo and the second wave at 48-72 hpo.

A single session of brief electrical stimulation enhances axon regeneration through nerve autografts

Nerve graft reconstruction of gap defects may result in poor clinical outcomes, particularly with long regeneration distances. Electrical stimulation (ES) of nerves may improve outcomes in such patients. A single session of ES at 20 Hz for 1 h significantly enhances axon regeneration in animals and human subjects after nerve crush or nerve transection and repair. The objectives of this study were to evaluate if ES enhances axon regeneration through nerve grafts and if there is added benefit of a second, delayed session of ES (serial ES) on axon regeneration as compared to a single session only of ES. In female rats, a gap defect was created in the hindlimb common peroneal (CP) nerve and immediately reconstructed with a 10 mm nerve autograft (Experiment 1) or a 20 mm nerve autograft (Experiment 2). In Experiment 1, rats were randomized to 1 h of CP nerve ES or sham stimulation. In Experiment 2, rats were randomized to control (sham ES + sham ES), single ES (ES + sham ES), or serial ES (ES + ES), which consisted of an initial 1 h session of either ES or sham stimulation of the CP nerve, followed by a second 1 h session of ES or sham stimulation of the CP nerve 4 weeks later. In both experiments, after a 6 week period of nerve regeneration, CP neurons that had regenerated axons distal to the autograft were retrograde labelled for enumeration, and the CP nerve distal to the autograft was harvested for histomorphometry. In Experiment 1, rats that received CP nerve ES had statistically significantly more motor (p < .05) and sensory (p < .05) neurons that regenerated axons distal to the 10 mm nerve autograft, with more myelinated axons on histomorphometry (p < .001). Similarly, in Experiment 2, significantly more motor (p < .01) and sensory (p < .05) neurons regenerated axons distal to the 20 mm nerve autograft after a single session or two sessions of CP nerve ES. There was no significant difference in the number of regenerated motor or sensory neurons between rats with 20 mm CP nerve autografts receiving either one or two sessions of CP nerve ES (p > .05). In conclusion, a single session of ES enhances axon regeneration following nerve autografting with no added effect of a second, delayed session of ES. These findings support previous studies in animals and humans of the robust effect of a single session of ES in promoting nerve regeneration following injury and repair.

Regeneration in the ctenophore Mnemiopsis leidyi occurs in the absence of a blastema, requires cell division, and is temporally separable from wound healing

BackgroundThe ability to regenerate is a widely distributed but highly variable trait among metazoans. A variety of modes of regeneration has been described for different organisms; however, many questions regarding the origin and evolution of these strategies remain unanswered. Most species of ctenophore (or “comb jellies”), a clade of marine animals that branch off at the base of the animal tree of life, possess an outstanding capacity to regenerate. However, the cellular and molecular mechanisms underlying this ability are unknown. We have used the ctenophore Mnemiopsis leidyi as a system to study wound healing and adult regeneration and provide some first-time insights of the cellular mechanisms involved in the regeneration of one of the most ancient extant group of multicellular animals.ResultsWe show that cell proliferation is activated at the wound site and is indispensable for whole-body regeneration. Wound healing occurs normally in the absence of cell proliferation forming a scar-less wound epithelium. No blastema-like structure is generated at the cut site, and pulse-chase experiments and surgical intervention show that cells originating in the main regions of cell proliferation (the tentacle bulbs) do not seem to contribute to the formation of new structures after surgical challenge, suggesting a local source of cells during regeneration. While exposure to cell-proliferation blocking treatment inhibits regeneration, the ability to regenerate is recovered when the treatment ends (days after the original cut), suggesting that ctenophore regenerative capabilities are constantly ready to be triggered and they are somehow separable of the wound healing process.ConclusionsCtenophore regeneration takes place through a process of cell proliferation-dependent non-blastemal-like regeneration and is temporally separable of the wound healing process. We propose that undifferentiated cells assume the correct location of missing structures and differentiate in place. The remarkable ability to replace missing tissue, the many favorable experimental features (e.g., optical clarity, high fecundity, rapid regenerative performance, stereotyped cell lineage, sequenced genome), and the early branching phylogenetic position in the animal tree, all point to the emergence of ctenophores as a new model system to study the evolution of animal regeneration.

Regulation of Genomic Output and (Pluri)potency in Regeneration.

Regeneration is a remarkable phenomenon that has been the subject of awe and bafflement for hundreds of years. Although regeneration competence is found in highly divergent organisms throughout the animal kingdom, recent advances in tools used for molecular and genomic characterization have uncovered common genes, molecular mechanisms, and genomic features in regenerating animals. In this review we focus on what is known about how genome regulation modulates cellular potency during regeneration. We discuss this regulation in the context of complex tissue regeneration in animals, from Hydra to humans, with reference to ex vivo-cultured cell models of pluripotency when appropriate. We emphasize the importance of a detailed molecular understanding of both the mechanisms that regulate genomic output and the functional assays that assess the biological relevance of such molecular characterizations.

7th Euro Evo Devo meeting: Report on the “Evolution of regeneration in Metazoa” symposium

Regeneration, the ability to restore lost parts of the body, is a widespread phenomenon in animals. While this ability is somehow limited in classical developmental model organisms, a variety of animals are able to regenerate complex structures such as limbs or important parts of their body, upon injury. Despite the recent emergence of regenerative studies using a large variety of metazoans, we still lack a general view of the evolution of animal regeneration. In the context of the 7th EvoDevo meeting that took place in June 2018 in Galway, Ireland, the "Evolution of regeneration in Metazoa" symposium gathered scientists studying the regenerative potential of evolutionarily distant animal species.

Model systems for regeneration: salamanders.

Salamanders have been hailed as champions of regeneration, exhibiting a remarkable ability to regrow tissues, organs and even whole body parts, e.g. their limbs. As such, salamanders have provided key insights into the mechanisms by which cells, tissues and organs sense and regenerate missing or damaged parts. In this Primer, we cover the evolutionary context in which salamanders emerged. We outline the varieties of mechanisms deployed during salamander regeneration, and discuss how these mechanisms are currently being explored and how they have advanced our understanding of animal regeneration. We also present arguments about why it is important to study closely related species in regeneration research.

Poly(ε-Caprolactone) Nanofiber Wrap Improves Nerve Regeneration and Functional Outcomes after Delayed Nerve Repair.

The purpose of this study was to assess the efficacy of biodegradable, electrospun poly(ε-caprolactone) nanofiber nerve conduits in improving nerve regeneration. The authors used a rat forelimb chronic denervation model to assess the effects of poly(ε-caprolactone) conduits on improving nerve regeneration and upper extremity function. Three groups of rats were examined: (1) negative-control animals (n = 5), which underwent 8 weeks of median nerve chronic denervation injury followed by repair with no conduit; (2) experimental animals (n = 5), which underwent 8 weeks of median nerve chronic denervation followed by repair and poly(ε-caprolactone) nerve conduit wrapping of the nerve coaptation site; and (3) positive-control animals (n = 5), which were naive controls. All animals underwent compound muscle action potential and functional testing. At 14 weeks after repair, the median nerve and flexor muscles were harvested for histologic analysis. Histomorphometric analysis of regenerating median nerves demonstrated augmented axonal regeneration in experimental versus negative control animals (total axon count, 1769 ± 672 versus 1072 ± 123.80; p = 0.0468). With regard to functional recovery, experimental and negative-control animals (1.67 ± 0.04 versus 0.97 ± 0.39; p = 0.036) had regained 34.9 percent and 25.4 percent, respectively, of baseline hand grip strength at 14 weeks after repair. Lastly, less collagen deposition at the nerve coaptation site of experimental animals was found when compared to control animals (p < 0.05). Biodegradable, poly(ε-caprolactone) nanofiber nerve conduits can improve nerve regeneration and subsequent physiologic extremity function in the setting of delayed nerve repair by decreasing the scar burden at nerve coaptation sites.

Whole-Tooth Regeneration by Allogeneic Cell Reassociation in Pig Jawbone.

The methods developed in this study to manipulate pig tooth germ cells in vitro and in vivo provide a reference for studying whole-tooth regeneration and tooth development in large animals. Of importance, compared with conventional ectopic tooth regeneration, conducted in the omentum, subcutaneous tissues, or kidney capsule (among other locations) with low with immune reactivity in rodent models, this study achieved orthotopic regeneration and development of whole teeth in a large mammal, representing a large stride toward the realization of tooth regenerative therapy for humans with missing teeth.

The Val66Met BDNF Polymorphism and Peripheral Nerve Injury: Enhanced Regeneration in Mouse Met-Carriers Is Not Further Improved With Activity-Dependent Treatment

Activity-dependent treatments to enhance peripheral nerve regeneration after injury have shown great promise, and clinical trials implementing them have begun. Success of these treatments requires activity-dependent release of brain-derived neurotrophic factor (BDNF). A single nucleotide polymorphism (SNP) in the bdnf gene known as Val66Met, which is found in nearly one third of the human population, results in defective activity-dependent BDNF secretion and could impact the effectiveness of these therapies. Here, we used a mouse model of this SNP to test the efficacy of treadmill exercise in enhancing axon regeneration in animals both heterozygous (V/M) and homozygous (M/M) for the SNP. Axon regeneration was studied 4 weeks after complete transection and repair of the sciatic nerve in both male and female animals, using both electrophysiological and histological outcome measures. Regeneration was enhanced significantly without treatment in V/M mice, compared with wild type (V/V) controls. Unlike V/V mice, treatment of both V/M and M/M mice with treadmill exercise did not result in enhanced regeneration. These results were recapitulated in vitro using dissociated neurons containing the light-sensitive cation channel, channelrhodopsin. Three days after plating, neurites of neurons from V/M and M/M mice were longer than those of V/V neurons. In neurons from V/V mice, but not those from V/M or M/M animals, longer neurites were found after optogenetic stimulation. Taken together, Met-carriers possess an intrinsically greater capacity to regenerate axons in peripheral nerves, but this cannot be enhanced further by activity-dependent treatments.

Nuclear receptor NR4A is required for patterning at the ends of the planarian anterior-posterior axis.

Positional information is fundamental to animal regeneration and tissue turnover. In planarians, muscle cells express signaling molecules to promote positional identity. At the ends of the anterior-posterior (AP) axis, positional identity is determined by anterior and posterior poles, which are putative organizers. We identified a gene, nr4A, that is required for anterior- and posterior-pole localization to axis extremes. nr4A encodes a nuclear receptor expressed predominantly in planarian muscle, including strongly at AP-axis ends and the poles. nr4A RNAi causes patterning gene expression domains to retract from head and tail tips, and ectopic anterior and posterior anatomy (e.g., eyes) to iteratively appear more internally. Our study reveals a novel patterning phenotype, in which pattern-organizing cells (poles) shift from their normal locations (axis extremes), triggering abnormal tissue pattern that fails to reach equilibrium. We propose that nr4A promotes pattern at planarian AP axis ends through restriction of patterning gene expression domains.

Hair Cell Development and Regeneration in Zebrafish Lacking the Cannabinoid Receptor2

Hair cells (HCs) are mechanosensory receptors found in sensory epithelia in the vertebrate inner ear that detect sound and head movements. HCs are fragile and prone to damage from various causes like disease, aging, and noise pollution. Unlike mammals, lower vertebrates including birds, amphibians and fish can regenerate HCs. To understand the underlying molecular mechanisms of HCs regeneration, we study the lateral line (LL) in a powerful genetic and regenerative animal model: the zebrafish. The LL is a skin‐deep fish sensory organ that is evolutionarily linked to the vertebrate sensory epithelia of the inner ear. It is composed of stereotypical distributed sensory patches called neuromasts (NMs) which contain HCs that are highly similar in function, cellular structure and gene expression to HCs of the inner ear. LL‐HCs are deflected and activate by surrounding water movements. Using CRISPR‐Cas9 genome editing technology, our laboratory has previously generated a null allele of the cnr2 gene (cnr2upr1) which is encoding the cannabinoid receptor 2 (CB2). We found that cnr2 mRNA is highly expressed in LL‐HCs. So, we hypothesized that CB2 has a role in the development and the regeneration of LL‐HCs. We counted LL‐HCs at different developmental stages in wild type and homozygote mutant larvae (cnr2upr1/upr1) to see if their number is affected in the absence of a functional CB2 protein. Next, to trigger vigorous HCs regeneration, we synchronously ablated LL‐HCs by copper treatment of 5‐day post‐fertilization (dpf) wild type and homozygote animals. We counted regenerating LL‐HCs for 3‐day post‐treatment. We will present our preliminary results that will ultimately allow us to conclude on the involvement of CB2 in development and in regeneration of the LL and on a potential link between endocannabinoid signaling and sensory epithelia development and regeneration.Support or Funding InformationCRESTThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Transcriptomic responses of regenerating earthworms (Eisenia foetida) to retinoic acid reveals the role of pluripotency genes

Exogenous retinoic acid (RA) delays and disturbs the regeneration of Eisenia foetida and inhibits the expression of pluripotent gene Sox2. However, studies of E. foetida conducted at the molecular level have been unable to elucidate its regeneration and mechanisms of RA effects on its regeneration. We merged existing transcriptomic data for E. foetida to generate a high-confidence set of transcriptomes. The de novo assembly of transcriptomes was performed by using the Trinity method, and functional annotations were analysed. We performed RNA-seq on four samples of regenerating tail fragments, three across a time-course (0, 3 and 7 days post amputation) and the fourth sample exposed to RA (7 days post amputation). E. foetida regeneration genes underwent significant upregulation and downregulation over the examined time periods, which may have been caused by a shared regulatory programme controlled by multiple gene families. The inhibition of RA against earthworm regeneration is likely related to the expression of these genes. Using annotation data and clustering, we also identified specific transcripts of 6 gene superfamilies enriched among genes exhibiting differential expression during regeneration periods and exhibiting the same expression patterns as those of the Sox2 gene. The regeneration transcriptome of tail fragment regeneration serves as a strong resource for investigating global expression changes that occur during regeneration and the toxicity of RA. This study offers insight for better understanding the regeneration of lower animals and molecular mechanisms of RA toxicity in invertebrates.