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The molecular mechanisms governing vertebrate appendage regeneration remain poorly understood. Uncovering these mechanisms may lead to novel therapies aimed at alleviating human disfigurement and visible loss of function following injury. Here, we explore tadpole tail regeneration in Xenopus tropicalis, a diploid frog with a sequenced genome.
We fo...
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... contrast, the T 60h vs T 24h array were almost indistin- guishable using these analytical conditions. The largest number of significant gene expression changes and the largest fold change magnitudes occurred in the T 6h vs. T 0h comparison; there were 422 targets that possessed an over 5-fold change (up or down) accompanied by a q-value of under .05 in the T 6h vs T 0h comparison, while only 20 targets fit these stringency conditions in the T 60h vs. T 24h comparison (Table 1). ...
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Background
The senses of hearing and balance depend upon mechanoreception, a process that originates in the inner ear and shares features across species. Amphibians have been widely used for physiological studies of mechanotransduction by sensory hair cells. In contrast, much less is known of the genetic basis of auditory and vestibular function i...
Citations
... The process of tissue regeneration encompasses a multitude of intricate cellular events, wherein maintaining a balanced cellular metabolic profile becomes crucial to meet the energy demands of regenerative tissues (Love et al. 2014). During Xenopus tail regeneration, it has been observed that glucose enters the glycolytic pathway and subsequently branches into the pentose phosphate pathway (Love et al. 2011(Love et al. , 2014. Moreover, due to the distinct metabolic requirements of stem cells and their progeny, stem cell proliferation often favors non-oxidative glycolysis, while differentiated somatic cells rely on oxidative phosphorylation (Folmes et al. 2011(Folmes et al. , 2012Panopoulos et al. 2012;Ly et al. 2020). ...
Limb autotomy and regeneration represent distinctive responses of crustaceans to environmental stress. Glucose metabolism plays a pivotal role in energy generation for tissue development and regeneration across various species. However, the relationship between glucose metabolism and tissue regeneration in crustaceans remains elusive. Therefore, this study is aimed at analyzing the alterations of glucose metabolic profile during limb autotomy and regeneration in Eriocheir sinensis, while also evaluating the effects of carbohydrate supplementation on limb regeneration. The results demonstrated that limb autotomy triggered a metabolic profile adaption at the early stage of regeneration. Hemolymph glucose levels were elevated, and multiple glucose catabolic pathways were enhanced in the hepatopancreas. Additionally, glucose and ATP levels in the regenerative limb were upregulated, along with increased expression of glucose transporters. Furthermore, the gene expression and activity of enzymes involved in gluconeogenesis were repressed in the hepatopancreas. These findings indicate that limb regeneration triggers metabolic profile adaptations to meet the elevated energy requirements. Moreover, the study observed that supplementation with corn starch enhanced limb regeneration capacity by promoting wound healing and blastema growth. Interestingly, dietary carbohydrate addition influenced limb regeneration by stimulating gluconeogenesis rather than glycolysis in the regenerative limb. Thus, these results underscore the adaptation of glucose metabolism during limb autotomy and regeneration, highlighting its essential role in the limb regeneration process of E. sinensis.
... Second, it has remarkable regenerative abilities and completes regenerative process fast, within 2 weeks following amputation. Third, it has both regenerative and non-regenerative period during development [8,23,24], offering an opportunity to study how regeneration is regulated during development. Previous studies have shown that tail regeneration proceeds through three essential periods: the formation of specialized wound epidermis, blastema bud formation and subsequent patterning and outgrowth via cell proliferation [9,[25][26][27]. ...
... Previous studies have shown that tail regeneration proceeds through three essential periods: the formation of specialized wound epidermis, blastema bud formation and subsequent patterning and outgrowth via cell proliferation [9,[25][26][27]. A number of genes/pathways and cellular activities have been found to be involved during these different periods, including reactive oxygen species (ROS), apoptosis, leptin, matrix metalloproteinases (MMPs), Wnt/FGF pathways, mTOR, and cellular metabolism [24][25][26][28][29][30][31][32]. ...
... However, the fold induction by amputation was much higher in the TRDKO tadpoles for 4 of the 6 genes, mmp1 (20.5-fold vs 7.4-fold), mmp13l (65.8-fold vs 3.4-fold), mmp13 (10.0-fold vs 2.9-fold) and mmp25 (7.2-fold vs 2.3-fold), compared to that in wild type tadpoles (Fig. 6A). We next assessed the expression of 2 genes known to be among the most significantly regulated genes after tail amputation and likely involved in the regeneration of different tissues/organs, suggesting that they are the conserved markers that be more useful for our experiment: the upregulated gene leptin and downregulated gene cyp26a [24]. We found that leptin was induced to peak level at 6 h post-amputation and the expression level then dropped to lower levels at 24 and 72 h in both wild type and TRDKO tail (Fig. 6B). ...
Background
Animal regeneration is the natural process of replacing or restoring damaged or missing cells, tissues, organs, and even entire body to full function. Studies in mammals have revealed that many organs lose regenerative ability soon after birth when thyroid hormone (T3) level is high. This suggests that T3 play an important role in organ regeneration. Intriguingly, plasma T3 level peaks during amphibian metamorphosis, which is very similar to postembryonic development in humans. In addition, many organs, such as heart and tail, also lose their regenerative ability during metamorphosis. These make frogs as a good model to address how the organs gradually lose their regenerative ability during development and what roles T3 may play in this. Early tail regeneration studies have been done mainly in the tetraploid Xenopus laevis ( X. laevis ), which is difficult for gene knockout studies. Here we use the highly related but diploid anuran X. tropicalis to investigate the role of T3 signaling in tail regeneration with gene knockout approaches.
Results
We discovered that X. tropicalis tadpoles could regenerate their tail from premetamorphic stages up to the climax stage 59 then lose regenerative capacity as tail resorption begins, just like what observed for X. laevis . To test the hypothesis that T3-induced metamorphic program inhibits tail regeneration, we used TR double knockout (TRDKO) tadpoles lacking both TRα and TRβ, the only two receptor genes in vertebrates, for tail regeneration studies. Our results showed that TRs were not necessary for tail regeneration at all stages. However, unlike wild type tadpoles, TRDKO tadpoles retained regenerative capacity at the climax stages 60/61, likely in part by increasing apoptosis at the early regenerative period and enhancing subsequent cell proliferation. In addition, TRDKO animals had higher levels of amputation-induced expression of many genes implicated to be important for tail regeneration, compared to the non-regenerative wild type tadpoles at stage 61. Finally, the high level of apoptosis in the remaining uncut portion of the tail as wild type tadpoles undergo tail resorption after stage 61 appeared to also contribute to the loss of regenerative ability.
Conclusions
Our findings for the first time revealed an evolutionary conservation in the loss of tail regeneration capacity at metamorphic climax between X. laevis and X. tropicalis . Our studies with molecular and genetic approaches demonstrated that TR-mediated, T3-induced gene regulation program is responsible not only for tail resorption but also for the loss of tail regeneration capacity. Further studies by using the model should uncover how T3 modulates the regenerative outcome and offer potential new avenues for regenerative medicines toward human patients.
... Larval tail regeneration in zebrafish is characterized by increased glucose uptake; though hexosamine biosynthesis, rather than glycolysis, is required for regeneration by enabling blastema formation though transforming growth factor alpha (TGF-b) signaling (Sinclair et al., 2021). Transcriptionally, regenerating Xenopus tails increases expression of glucose transporters and the rate-limiting PPP enzyme g6p dehydrogenase (g6pd), suggesting increased PPP activity (Love et al., 2011). It has been proposed that increased PPP activity could provide biosynthetic intermediates and redox regulation needed for Xenopus tail regeneration (Love et al., 2011(Love et al., , 2014. ...
... Transcriptionally, regenerating Xenopus tails increases expression of glucose transporters and the rate-limiting PPP enzyme g6p dehydrogenase (g6pd), suggesting increased PPP activity (Love et al., 2011). It has been proposed that increased PPP activity could provide biosynthetic intermediates and redox regulation needed for Xenopus tail regeneration (Love et al., 2011(Love et al., , 2014. While these models center on glucose metabolism, it seems that different branches of glucose breakdown are prioritized in different wounding systems. ...
... Metabolic changes driving regeneration are attractive targets for improving wound-healing outcomes in non-regenerative contexts. Here, we identify a critical increase in PPP flux downstream of increased glucose uptake in the regenerating Xenopus tail, building on prior transcriptomic studies (Love et al., 2011(Love et al., , 2014. Further, we identify an increase in pro-proliferative metabolites during regeneration. ...
A fundamental step in regeneration is rapid growth to replace lost tissue. Cells must generate sufficient lipids, nucleotides, and proteins to fuel rapid cell division. To define metabolic pathways underlying regenerative growth, we undertake a multimodal investigation of metabolic reprogramming in Xenopus tropicalis appendage regeneration. Regenerating tissues have increased glucose uptake; however, inhibition of glycolysis does not decrease regeneration. Instead, glucose is funneled to the pentose phosphate pathway (PPP), which is essential for full tail regeneration. Liquid chromatography-mass spectrometry (LC-MS) metabolite profiling reveals increased nucleotide and nicotinamide intermediates required for cell division. Using single-cell RNA sequencing (scRNA-seq), we find that highly proliferative cells have increased transcription of PPP enzymes and not glycolytic enzymes. Further, PPP inhibition results in decreased cell division specifically in regenerating tissue. Our results inform a model wherein regenerating tissues direct glucose toward the PPP, yielding nucleotide precursors to drive regenerative cell proliferation.
... While OAA-dependent increases in glycolysis and OxPhos could be attributed to the differential effects of WNT signaling, it's also important to consider the critical signaling role of OxPhos-mediated ROS generation in limb regeneration. ROS production after tail amputation in Xenopus is necessary for regeneration, possibly through WNT signaling and proliferation (Love et al., 2013), and leptin and melanocortin 4 receptor, key molecules regulating energy balance, are critical in tadpole tail regeneration (Love et al., 2011) and digit regeneration (Kang et al., 2016;Zhang et al., 2018). While we chose OAA partially because it supplemented the elevated cell metabolism levels seen at D10, these increased metabolism levels may be an inherent and essential part of limb regeneration, with OAA potentially providing building blocks for cell growth and expansion, further increasing cell metabolism. ...
De novo limb regeneration after amputation is restricted in mammals to the distal digit tip. Central to this regenerative process is the blastema, a heterogeneous population of lineage-restricted, dedifferentiated cells that ultimately orchestrates regeneration of the amputated bone and surrounding soft tissue. To investigate skeletal regeneration, we made use of spatial transcriptomics to characterize the transcriptional profile specifically within the blastema. Using this technique, we generated a gene signature with high specificity for the blastema in both our spatial data, as well as other previously published single-cell RNA-sequencing transcriptomic studies. To elucidate potential mechanisms distinguishing regenerative from non-regenerative healing, we applied spatial transcriptomics to an aging model. Consistent with other forms of repair, our digit amputation mouse model showed a significant impairment in regeneration in aged mice. Contrasting young and aged mice, spatial analysis revealed a metabolic shift in aged blastema associated with an increased bioenergetic requirement. This enhanced metabolic turnover was associated with increased hypoxia and angiogenic signaling, leading to excessive vascularization and altered regenerated bone architecture in aged mice. Administration of the metabolite oxaloacetate decreased the oxygen consumption rate of the aged blastema and increased WNT signaling, leading to enhanced in vivo bone regeneration. Thus, targeting cell metabolism may be a promising strategy to mitigate aging-induced declines in tissue regeneration.
... This increasing pandemic of non-communicable/chronic diseases mandates urgent research into possible aetiopathogenic influences. Epidemiological studies show association to entity called metabolic syndrome and this suggests that chronic diseases may to an extent share common genetic and/or environmental predisposing factors [4]. ...
Background: Vegetable oils are commonly used food additives. Aim: To characterize and observe the toxic effect of sub chronic consumption of heated and unheated Palm Kernel oil (PKO) and Soya Oil (S0) after heating at 180 ℃ and at room temperature in albino Wistar rats. Methodology: The physicochemical properties were analyzed, sixty rats were divided into five groups of twelve rats and their baseline weight recorded. Group 1 (controls), received water and feed only, while 10mls of each oil was added (add libitum) to 100 grams of feed in each test group for six weeks and twelve weeks. Group 2 received feed, water and unheated PKO and Group 3 received feed, water and heated PKO. Group 4 were given feed, water and unheated SO while group 5 had feed, water and heated SO. The lipid profile and blood glucose were measured at six weeks exposure while the renal function and liver function were determined at twelve weeks exposure. Result: Heating the oils at 180 ℃ for 15 minutes caused a decrease in relative density, peroxide values, and iodine values in both the PKO and SBO and increased acid values and free fatty acids in both with no effect in the refractive index. Biochemical result showed that both heated and the unheated oil samples significantly increased the LDL levels, HPKO and UHSO caused increase in total cholesterol levels, UHPKO significantly decreased the TAG, HPKO significantly increased the ALT, HPKO significantly decreased the urea level and HSO increased the BG significantly. Conclusion: Heating alters the physicochemical properties of the oils thereby decreasing their qualities, while the oils also demonstrated evidence of hyperlipidemia.
... The oxidative response is also regulated in regenerative models; a transcriptomic study of the regenerative response after tail amputation in Xenopus tropicalis tadpoles, showed that "hydrogen peroxide metabolic process", "superoxide metabolic process", and "oxygen and reactive oxygen species metabolic process" were among the most upregulated processes during the first 24 h of regeneration [183]. Likewise, the early production of reactive oxygen species (ROS) is necessary for the activation of signaling pathways that regulate cellular proliferation, which is required for initiating the regenerative program in Xenopus tropicalis [182], Axolotl [5], and Zebrafish tail regeneration [91], as well as zebrafish fin regeneration [92]. ...
... RNAseq and proteomics experiments performed in our laboratory showed that mitochondrial proteins, as well as proteins and transcripts involved in metabolic processes, e.g., fatty acid and glucose metabolism, are differentially regulated in regenerative and non-regenerative stages after Xenopus laevis SCI [168,169]. In addition, a transcriptomic study of the regenerative response after tail amputation in Xenopus tropicalis tadpoles, showed that genes associated with organic acid metabolism and NADP/H metabolic process were among the most regulated [183]. These data lead to consider mitochondrial function and substrate utilization as key aspects that suffer adaptive responses after SCI and therefore, treatments targeting them have been proposed beneficial for improving regeneration [246]. ...
... Even though there are no studies showing how changes in the different aforementioned metabolic pathways affect NSPCs response after SCI, we previously demonstrated that SCI induces early changes in transcripts associated to metabolic processes, such as fatty acid and glucose metabolism, which are temporarily related to changes in transcripts associated to cell cycle [169] and the Sox2 + cells proliferative response to SCI [207] in regenerative X. laevis stages. The same has been observed by others in a X. tropicalis tail amputation model, where Affymetrix genome array [183] and RNAseq analyses [232], show changes in transcripts related to metabolic processes and cell proliferation during tail and spinal cord regeneration. All these evidences suggest a relationship between changes in metabolism, cell proliferation, and spinal cord regeneration, that should be addressed by future experiments. ...
Many people around the world suffer from some form of paralysis caused by spinal cord injury (SCI), which has an impact on quality and life expectancy. The spinal cord is part of the central nervous system (CNS), which in mammals is unable to regenerate, and to date, there is a lack of full functional recovery therapies for SCI. These injuries start with a rapid and mechanical insult, followed by a secondary phase leading progressively to greater damage. This secondary phase can be potentially modifiable through targeted therapies. The growing literature, derived from mammalian and regenerative model studies, supports a leading role for mitochondria in every cellular response after SCI: mitochondrial dysfunction is the common event of different triggers leading to cell death, cellular metabolism regulates the immune response, mitochondrial number and localization correlate with axon regenerative capacity, while mitochondrial abundance and substrate utilization regulate neural stem progenitor cells self-renewal and differentiation. Herein, we present a comprehensive review of the cellular responses during the secondary phase of SCI, the mitochondrial contribution to each of them, as well as evidence of mitochondrial involvement in spinal cord regeneration, suggesting that a more in-depth study of mitochondrial function and regulation is needed to identify potential targets for SCI therapeutic intervention.
... BMP, FGF, Wnt, and Notch signaling play an important role in tail regeneration (Beck et al. 2003). Furthermore, the Amaya lab has reported a key role for reactive oxygen species (ROSs) in the initiation of the regenerative response that seems to be evolutionarily conserved among many species (Love et al. 2011;Love et al. 2013;Phipps et al. 2020). ...
... The main aim of the study [25] was to calculate the variations in gene expression during the regeneration of tadpole tail. Particularly, the study required to develop a gene-expression dataset to act as a resource in finding genes as well as the processes that are involved in the tail regeneration. ...
The augmentation of regenerative capability is a powerful method for pursuing for the regulation of degeneration, traumatic injury and cancer. The tadpole, Clinotarsus curtipes and Xenopus laevis is a significant model system for addressing the fundamental regeneration mechanism that enables to understand the key aspects of regeneration medicine. The selected creatures Clinotarsus curtipes and Xenopus laevis could able to obtain both tissue regeneration and scar free healing during larval stage in spite of its predominant loss of such ability during the metamorphic process. Such transient capability associated with the evolutionary correlation with humans creates Clinotarsus curtipes and Xenopus a very good attractive model for uncovering the functional regeneration mechanisms. The study analysed the existing literatures on change in the levels of ROS that is required for the proper wnt-signaling in every regeneration system. Apart from that the paper provided the comprehensive review on the histopathological view, regeneration signals like TGFβ, FGF, BMP, Wnt etc for successful regeneration. Factors that affect the tail regeneration like O2 influx, epigenetics and HDAC activity have also been provided. Significant other such criteria like role of TRKA signaling, profiling and intracellular protein expression followed by its corresponding challenges adds value to the paper.The study presents an overview of Xenopus and Clinotarsus curtipesas a model organism for the research and highlighted the new insights.
... On the contrary, other species, including planarias, fish, amphibians and lampreys, have regenerative mechanisms that allow the anatomical and functional recovery of complex structures, such as organs, eyes, neural tissue, and appendages [10][11][12][13] . Amphibians and teleost fish are model organisms widely used to understand the molecular and cellular mechanisms involved in spinal cord regeneration 14,15 . ...
Xenopus laevis are able to regenerate the spinal cord during larvae stages through the activation of neural stem progenitor cells (NSPCs). Here we use high-resolution expression profiling to characterize the early transcriptome changes induced after spinal cord injury, aiming to identify the signals that trigger NSPC proliferation. The analysis delineates a pathway that starts with a rapid and transitory activation of immediate early genes, followed by migration processes and immune response genes, the pervasive increase of NSPC-specific ribosome biogenesis factors, and genes involved in stem cell proliferation. Western blot and immunofluorescence analysis showed that mTORC1 is rapidly and transiently activated after SCI, and its pharmacological inhibition impairs spinal cord regeneration and proliferation of NSPC through the downregulation of genes involved in the G1/S transition of cell cycle, with a strong effect on PCNA. We propose that the mTOR signaling pathway is a key player in the activation of NPSCs during the early steps of spinal cord regeneration.
... Spinal cord injuries (SCIs) are often irreversible and lead to the loss of motor and sensory function below the site of the damage (McDonald & Sadowsky, 2002). In contrast, amphibians such as Xenopus (X.) tadpoles have far greater regenerative abilities as they can regenerate a fully functional tail following amputation, including their spinal cord (Deuchar, 1975;Love et al, 2011;Kakebeen et al, 2020). The injured spinal cord is sealed within a day by the formation of the neural ampulla, and lineage tracing has revealed that the spinal cord regenerates from its original stump (Gargioli & Slack, 2004;Slack et al, 2008). ...
Xenopus tadpoles have the ability to regenerate their tails upon amputation. Although some of the molecular and cellular mechanisms that globally regulate tail regeneration have been characterised, tissue-specific response to injury remains poorly understood. Using a combination of bulk and single-cell RNA sequencing on isolated spinal cords before and after amputation, we identify a number of genes specifically expressed in the spinal cord during regeneration. We show that Foxm1, a transcription factor known to promote proliferation, is essential for spinal cord regeneration. Surprisingly, Foxm1 does not control the cell cycle length of neural progenitors but regulates their fate after division. In foxm1-/- tadpoles, we observe a reduction in the number of neurons in the regenerating spinal cord, suggesting that neuronal differentiation is necessary for the regenerative process. Altogether, our data uncover a spinal cord-specific response to injury and reveal a new role for neuronal differentiation during regeneration.
