Tadpole Tail Research Articles

An improved method for whole-mount in situ hybridization in regenerating tails of Xenopus laevis tadpoles

Whole-mount in situ hybridization (WISH) is a widely used method that supports the concept of “seeing is believing” by enabling the visualization of gene expression patterns in whole-mount multicellular samples or sections. This technique is essential in the study of epimorphic regeneration in cold-blooded vertebrates, where complex three-dimensional organs such as tails, limbs, and eyes are completely restored after loss. The tadpoles of the frog X. laevis serve as a convenient model for studying regeneration, as they can regenerate their tails within a week after amputation. Modern high-throughput sequencing methods have identified various cell populations involved in the regeneration process and determined the repertoire of genes activated during this time. Specifically, a population of reparative myeloid cells expressing mmp9 as a marker gene has been shown to be crucial for the initial stages of tail regeneration in X. laevis tadpoles. The validation of these data and further examination using WISH offers the advantage of providing detailed information on the spatial and temporal dynamics of target gene expression levels. However, detecting mRNA by WISH can be challenging when mRNA levels are very low, transcripts are localized in hard-to-access areas, or tissue samples are prone to background staining, as is the case with X. laevis regenerating tail samples. Here, we describe additional treatments for regenerating tail samples that minimize background staining and enhance the visualization of cells containing target RNA through in situ hybridization. Using an optimized WISH protocol on X. laevis tadpole tail regenerates, we obtained novel data on the mmp9 expression pattern during the first day post-amputation at the regeneration-competent stage 40 and the regeneration-incompetent stage 47 (refractory period). The significant differences in the expression patterns indicate that mmp9 activity is positively correlated with regeneration competence.

SoxC and MmpReg promote blastema formation in whole-body regeneration of fragmenting potworms Enchytraeus japonensis

Regeneration in many animals involves the formation of a blastema, which differentiates and organizes into the appropriate missing body parts. Although the mechanisms underlying blastema formation are often fundamental to regeneration biology, information on the cellular and molecular basis of blastema formation remains limited. Here, we focus on a fragmenting potworm (Enchytraeus japonensis), which can regenerate its whole body from small fragments. We find soxC and mmpReg as upregulated genes in the blastema. RNAi of soxC and mmpReg reduce the number of blastema cells, indicating that soxC and mmpReg promote blastema formation. Expression analyses show that soxC-expressing cells appear to gradually accumulate in blastema and constitute a large part of the blastema. Additionally, similar expression dynamics of SoxC orthologue genes in frog (Xenopus laevis) are found in the regeneration blastema of tadpole tail. Our findings provide insights into the cellular and molecular mechanisms underlying blastema formation across species.

Appendage regeneration requires IMPDH2 and creates a sensitized environment for enzyme filament formation.

Regeneration of lost tissue requires biosynthesis of metabolites needed for cell proliferation and growth. Among these are the critical purine nucleotides ATP and GTP. The abundance and balance of these purines is regulated by inosine monophosphate dehydrogenase 2 (IMPDH2), which catalyzes the committing step of GTP synthesis. IMPDH2 assembles into filaments that resist allosteric inhibition under conditions of high GTP demand. Here we asked whether IMPDH2 is required in the highly proliferative context of regeneration, and whether its assembly into filaments takes place in regenerating tissue. We find that inhibition of IMPDH2 leads to impaired tail regeneration and reduced cell proliferation in the tadpole Xenopus tropicalis. We find that both endogenous and fluorescent fusions of IMPDH2 robustly assemble into filaments throughout the tadpole tail, and that the regenerating tail creates a sensitized condition for filament formation. These findings clarify the role of purine biosynthesis in regeneration and reveal that IMPDH2 enzyme filament formation is a biologically relevant mechanism of regulation in vertebrate regeneration.

Adsorption characteristics of Janus tadpole polymers

The shape of Janus particles is directly connected to their adsorption behavior. Janus tadpole polymers offer a unique topological architecture that includes competition between entropic, enthalpic, and topological terms in the adsorption free energy; accordingly, non-trivial adsorption behavior patterns are expected. We study the surface adsorption of Janus tadpole polymers by means of Monte Carlo simulations, finding that, depending on which part of the tadpole polymers is preferentially adsorbing on the surface, very different types of behavior for both the adsorbed polymeric phase and of the brush arise. The adsorbed phase and the brush mutually influence each other, leading to a variety of phenomena such as nematic ordering of the adsorbed stiff tadpole tails and intriguing changes in the territoriality of adsorbed ring polymers on the surface. We analyze in detail our findings, revealing the mechanisms behind the organization and ordering, and opening up new possibilities to tune and control the structure of such systems.

Injury-induced cooperation of InhibinβA and JunB is essential for cell proliferation in Xenopus tadpole tail regeneration

In animal species that have the capability of regenerating tissues and limbs, cell proliferation is enhanced after wound healing and is essential for the reconstruction of injured tissue. Although the ability to induce cell proliferation is a common feature of such species, the molecular mechanisms that regulate the transition from wound healing to regenerative cell proliferation remain unclear. Here, we show that upon injury, InhibinβA and JunB cooperatively function for this transition during Xenopus tadpole tail regeneration. We found that the expression of inhibin subunit beta A (inhba) and junB proto-oncogene (junb) is induced by injury-activated TGF-β/Smad and MEK/ERK signaling in regenerating tails. Similarly to junb knockout (KO) tadpoles, inhba KO tadpoles show a delay in tail regeneration, and inhba/junb double KO (DKO) tadpoles exhibit severe impairment of tail regeneration compared with either inhba KO or junb KO tadpoles. Importantly, this impairment is associated with a significant reduction of cell proliferation in regenerating tissue. Moreover, JunB regulates tail regeneration via FGF signaling, while InhibinβA likely acts through different mechanisms. These results demonstrate that the cooperation of injury-induced InhibinβA and JunB is critical for regenerative cell proliferation, which is necessary for re-outgrowth of regenerating Xenopus tadpole tails.

Tadpole fingerprinting: Using tail venation patterns to photo‐identify tadpole individuals of a threatened frog

AbstractTraditional methods for identifying individual amphibians in capture–mark–recapture (CMR) studies have been primarily confined to post‐metamorphic stages, using artificial markers that come with a variety of limitations. An alternative that may open CMR studies to earlier life stages involves the use of a species' natural external markers in photo‐based identification. In this study, we investigated whether it was possible to distinguish tadpoles of the threatened green and golden bell frog (Litoria aurea) at the individual level based on tail venation patterns. We collected photographs of the tails of captive‐raised tadpoles using a smartphone over a 4‐week period. This photo‐library was used to create an electronic survey where participants were asked to detect matches for query tadpoles from small image pools. We found that most participants agreed on a match for each query, with perfect consensus achieved for most queries (83%). We detected a 14% decline in perfect consensus when participants were asked to match images of tadpoles separated by longer time intervals, suggesting that it is more difficult to visually identify recapture events of L. aurea tadpoles over extended periods due to changes to tail appearance. However, consensus was obtained by participants for all queries, with all matches verified as being correct by the primary researcher. The strength of agreement among participants with no prior experience in matching tadpole tails suggests that there is sufficient inter‐individual variation in this feature for individuals to be manually identified. We thus propose that photo‐identification is likely to be a valid, non‐invasive technique that can be used for short‐term studies on tadpole populations that display tail venation. This offers an alternative to artificial markers that may not allow for individual identification, while also opening up tadpole monitoring programmes to citizen scientists who can be recruited online to process image data from home.

Characterization of a novel corticosterone response gene in Xenopus tropicalis tadpole tails.

Corticosteroids are critical for development and for mediating stress responses across diverse vertebrate taxa. Study of frog metamorphosis has made significant breakthroughs in our understanding of corticosteroid signaling during development in non-mammalian vertebrate species. However, lack of adequate corticosterone (CORT) response genes in tadpoles make identification and quantification of CORT responses challenging. Here, we characterized a CORT-response gene frzb (frizzled related protein) previously identified in Xenopus tropicalis tadpole tail skin by an RNA-seq study. We validated the RNA-seq results that CORT and not thyroid hormone induces frzb in the tails using quantitative PCR. Further, maximum frzb expression was achieved by 100-250 nM CORT within 12-24 hours. frzb is not significantly induced in the liver and brain in response to 100 nM CORT. We also found no change in frzb expression across natural metamorphosis when endogenous CORT levels peak. Surprisingly, frzb is only induced by CORT in X. tropicalis tails and not in Xenopus laevis tails. The exact downstream function of increased frzb expression in tails in response to CORT is not known, but the specificity of hormone response and its high mRNA expression levels in the tail render frzb a useful marker of exogenous CORT-response independent of thyroid hormone for exogenous hormone treatments and in-vivo endocrine disruption studies.

Comparative Analysis of Transcriptome Profiles Reveals Distinct and Organ-Dependent Genomic and Nongenomic Actions of Thyroid Hormone in Xenopus tropicalis Tadpoles.

Background: Thyroid hormone (triiodothyronine [T3]) is essential for development and organ metabolism in all vertebrates. T3 has both genomic and nongenomic effects on target cells. While much has been learnt on its genomic effects via T3 receptors (TRs) in vertebrate development, mostly through TR-knockout and TR-knockin studies, little is known about the effects of T3 on gene expression in animals in the absence of TR. We have been studying Xenopus metamorphosis as a model for mammalian postembryonic development, a period around birth when plasma T3 level peaks and many organs/tissues mature into their adult forms. We have recently generated TR double knockout (TRDKO) Xenopus tropicalis animals. This offers an opportunity to compare the effects of T3 on global gene expression in tadpole tissues in the presence or absence of TR. Methods: We analyzed the effects of T3 on gene expression in tadpole tail and intestine by using RNA-seq analysis on wild-type and TRDKO tadpoles with or without T3 treatment. Results: We observed that removing TRs reduced the number of genes regulated by T3 in both organs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses revealed that T3 affected distinct biological processes and pathways in wild-type and TRDKO tadpoles. Many GO terms and KEGG pathways that were enriched among genes regulated in wild-type tissues are likely involved in mediating the effects of T3 on metamorphosis, for example, those related to development, stem cells, apoptosis, and cell cycle/cell proliferation. However, such GO terms and pathways were not enriched among T3-regulated genes in TRDKO tadpoles. Instead, in TRDKO tadpoles, GO terms and pathways related to "metabolism" and "immune response" were highly enriched among T3-regulated genes. We further observed strong divergence in the TR-independent nongenomic effects of T3 in the intestine and tail. Conclusions: Our data suggest that T3 has distinct and organ-dependent effects on gene expression in developing tadpoles. The TR-mediated effects are consistent with the metamorphic changes, in agreement with the fact that TR is necessary and sufficient to mediate the effects of T3 on metamorphosis. T3 appears to have a major effect on metabolism and immune response via TR-independent nongenomic processes.

Strongylopus grayii tadpole blastema extract exerts cytotoxic effects on embryonal rhabdomyosarcoma cells

Amphibians have regenerative capacity and are resistant to developing cancer. This suggests that the developing blastema, located at the tissue regeneration site, may secrete anti-cancer factors. Here, we investigate the anti-cancer potential of tadpole tail blastema extracts (TAD) from the stream frog, Strongylopus grayii, in embryonal rhabdomyosarcoma (ERMS) cells. ERMS originates in skeletal muscle tissue and is a common pediatric soft tissue sarcoma. We show using MTT assays that TAD inhibited ERMS cell viability in a concentration-dependent manner, and phase contrast/fluorescent microscopy revealed that it induced morphological markers of senescence and apoptosis. Western blotting showed that this was associated with DNA damage (γH2AX) and activation of the p38/MAPK stress signaling pathway as well as molecular markers of senescence (p16INK4a), apoptosis (cleaved PARP), and inhibition of cell cycle promoters (cyclin A, CDK2, and cyclin B1). Furthermore, proteomics followed by gene ontology analyses showed that TAD treatment inhibited known tumor promoters and proteins required for cancer cell survival. Lastly, using the LINCS drug perturbation library, we show that there is an overlap between the proteomics signature induced by TAD and common anti-cancer drugs. Taken together, this study provides novel evidence that TAD exhibits cytotoxicity in ERMS cells.

Upregulation of proto-oncogene ski by thyroid hormone in the intestine and tail during Xenopus metamorphosis

Thyroid hormone (T3) is important for adult organ function and vertebrate development, particularly during the postembryonic period when many organs develop/mature into their adult forms. Amphibian metamorphosis is totally dependent on T3 and can be easily manipulated, thus offering a unique opportunity for studying how T3 controls postembryonic development in vertebrates. Numerous early studies have demonstrated that T3 affects frog metamorphosis through T3 receptor (TR)-mediated regulation of T3 response genes, where TR forms a heterodimer with RXR (9-cis retinoic acid receptor) and binds to T3 response elements (TREs) in T3 response genes to regulate their expression. We have previously identified many candidate direct T3 response genes in Xenopus tropicalis tadpole intestine. Among them is the proto-oncogene Ski, which encodes a nuclear protein with complex function in regulating cell fate. We show here that Ski is upregulated in the intestine and tail of premetamorphic tadpoles upon T3 treatment and its expression peaks at stage 62, the climax of metamorphosis. We have further discovered a putative TRE in the first exon that can bind to TR/RXR in vitro and mediate T3 regulation of the promoter in vivo. These data demonstrate that Ski is activated by T3 through TR binding to a TRE in the first exon during Xenopus tropicalis metamorphosis, implicating a role of Ski in regulating cell fate during metamorphosis.

Effective enrichment of stem cells in regenerating Xenopus laevis tadpole tails using the side population method.

Xenopus laevis tadpoles have a strong regenerative ability and can regenerate their whole tails after tail amputation. Lineage-restricted tissue stem cells are thought to provide sources for the regenerating tissues by producing undifferentiated progenitor cells in response to tail amputation. However, elucidating the behavioral dynamics of tissue stem cells during tail regeneration is difficult because of their rarity, and there are few established methods of isolating these cells in amphibians. Here, to detect and analyze rare tissue stem cells, we attempted to enrich tissue stem cells from tail regeneration buds. High Hoechst dye efflux capacity is thought to be a common characteristic of several types of mammalian tissue stem cells; these stem cells, designated as the "side population (SP)," may be enriched by flow cytometry (SP method). To evaluate the effectiveness of stem cell enrichment using the SP method in regenerating X. laevis tadpole tails, we performed single-cell RNA sequencing (scRNA-seq) of SP cells from regeneration buds and analyzed the frequency of satellite cells, which are muscle stem/progenitor cells expressing pax7. The pax7-expressing cells were enriched in the SP compared with whole normal tails and regeneration buds. Furthermore, hes1-expressing cells, which are assumed to be neural stem/progenitor cells, were also enriched in the SP. Our findings suggest that the SP method is efficient for successfully enriching tissue stem cells in regenerating X. laevis tadpole tails, indicating that the combination of the SP method and scRNA-seq is useful for studying tissue stem cells that contribute to tail regeneration.

Double peak quasi-periodic pulsations in a circular-ribbon flare

We study quasi-periodic pulsations (QPPs) during the impulsive phase of the C8.3 flare SOL2002-08-06T01:43. The shape of an extended 5.7 GHz source is similar to a tadpole with the head located above the region of a negative magnetic polarity, surrounded by positive polarity patches and with a remote tail source. The flare configuration includes bright extreme ultraviolet (EUV) ropes with footpoints near the boundary of the negative magnetic field region and it can be identified as a circular ribbon flare. We use simultaneous observations carried out by the Siberian Solar Radio Telescope at 5.7 GHz, the Nobeyama Radio Heliograph (NoRH) at 17 and 34 GHz, the Reuven Ramaty High Energy Solar Spectroscopic Imager (RHESSI)/HXR, and the Transition Region and Coronal Explorer (TRACE) imaging in the extreme ultraviolet. The flare HXR emission is produced by a compact source located at the south periphery of the Negative Magnetic field Region (NMR). The QPPs are observed during a one-minute interval after the start of the impulsive phase, when this HXR source appeared. The remote source is detected on the variation maps of the of the brightness temperature at 17 GHz and is located at the end of tadpole tail about 60 arcsec eastward. More than a dozen cotemporal HXR and microwave pulses with timescales from 1.5 s up to about 8 s were observed in the flare kernel. At 5.7 GHz, the pulses are more prominent near the remote source where they are highly polarized and generated by the electron beams propagating from the flare kernel. The main tone of the QPP periodicity corresponds to the oscillations with a period of 8 s and is accompanied by the variations in the hardness of nonthermal electrons, that is, in the efficiency of the acceleration mechanism. The second intensity harmonic (about a 3-s period) appears due to a double peak structure of the QPP event. Such pulse shapes suggest oscillations of the current sheet during the loop coalescence as a modulation mechanism of the flare energy release.

Inducible and tissue-specific cell labeling in Cre-ERT2 transgenic Xenopus lines.

Investigating cell lineage requires genetic tools that label cells in a temporal and tissue‐specific manner. The bacteriophage‐derived Cre‐ER T2 /loxP system has been developed as a genetic tool for lineage tracing in many organisms. We recently reported a stable transgenic Xenopus line with a Cre‐ER T2 /loxP system driven by the mouse Prrx1 (mPrrx1) enhancer to trace limb fibroblasts during the regeneration process (Prrx1:CreER line). Here we describe the detailed technological development and characterization of such line. Transgenic lines carrying a CAG promoter‐driven Cre‐ER T2 /loxP system showed conditional labeling of muscle, epidermal, and interstitial cells in both the tadpole tail and the froglet leg upon 4‐hydroxytamoxifen (4OHT) treatment. We further improved the labeling efficiency in the Prrx1:CreER lines from 12.0% to 32.9% using the optimized 4OHT treatment regime. Careful histological examination showed that Prrx1:CreER lines also sparsely labeled cells in the brain, spinal cord, head dermis, and fibroblasts in the tail. This work provides the first demonstration of conditional, tissue‐specific cell labeling with the Cre‐ER T2 /loxP system in stable transgenic Xenopus lines.

Influence of tail injury on the development of Neotropical elegant treefrog tadpoles

Anuran larvae in aquatic environments are important prey items for several vertebrate and invertebrate species. Besides avoiding predation, there are some strategies that may reduce the physical damage in those tadpoles that survive the predation attempt. For example, the injured tadpole tail can regrow after a predator bite, but few studies have examined the consequences of such injury. We examined the consequences of three levels of injury to the tail and how this influenced development and feeding behavior of tadpoles of the Neotropical elegant treefrog, Dendropsophus elegans. We collected spawns and kept them in the laboratory until tadpoles reached Gosner’s stages 28 to 35. Then, they were separated in four experimental groups: individuals with tail trimmed in 30, 50 or 70 % of its length, and a control group, with no tail removing. We counted the days until metamorphosis, calculated the Scaled Mass Index (SMI) through weight and length of newly-metamorphosed, and evaluated the feeding frequency to evaluate the influence of tail amputation on them. We found that the time until metamorphosis was positively related to the extent of the amputation, but SMI and feeding behavior were not influenced. As the time to metamorphose is related to the survivorship chances of individuals: i.e., if the aquatic environment is with high density of predators, it would be advantageous to rapidly metamorphose out of the water. However, tail injury delays the metamorphose process, which could influence the survival of the individual.

Uncovering early thyroid hormone signalling events through temperature-mediated activation of molecular memory in the cultured bullfrog tadpole tail fin

Thyroid hormone (TH) is a critical signalling molecule for all vertebrate organisms, playing a crucial role in postembryonic development. The best-studied mechanism of TH response is through modulating gene expression, however TH’s involvement in coordinating the early steps in the TH signal transduction pathway is still poorly understood. The American bullfrog, Rana [Lithobates] catesbeiana, is a useful model to study these early responses as tadpole post-embryonic development in the form of metamorphosis of the tadpole into a frog can be experimentally induced by TH exposure. The rate of TH-induced metamorphosis can be modulated by temperature where sufficiently cold temperatures (5 °C) completely halt precocious metamorphosis. Interestingly, when premetamorphic tadpoles exposed to exogenous THs at 5 °C are shifted to permissive temperatures (24 °C), their metamorphic rate exceeds that of TH-exposed tadpoles at the permissive temperature. This suggests that a molecular memory of TH exposure is retained at 5 °C even after THs are cleared at this low temperature. However, the molecular memory machinery is poorly understood. Herein we use RNA-seq analysis to identify potential components of the molecular memory in cultured tail fin that allows for the recapitulation of the molecular memory phenomenon. Eighty-one gene transcripts were TH-responsive at 5 °C compared to matched controls indicating that the molecular memory is more complex than previously thought. Many of these transcripts encode transcription factors including thyroid hormone-induced B/Zip, thibz, and a novel krüppel-like factor family member, klfX. Actinomycin D and cycloheximide treatment had no effect on their TH induction suggesting that a change in transcription or translation is not required. Rather a change in RNA stability may be a possible mechanism contributing to the molecular memory. The ability to manipulate temperature and TH response in cultured organs provide an exciting opportunity to further elucidate the early TH signalling mechanisms during postembryonic development.

Methodology for intravital mitotic chromosome preparation from regenerated tissue derived from the tail tips of tadpoles

We propose a modified and updated protocol to obtain mitotic chromosomes from the regenerated tissue of Pelophylax tadpole tail tips. Chromosomal preparations from regenerated tissue results in high-quality and clean slides suitable for further staining and study. Tadpoles remain alive, undergo minimum suffering, and can be grown to adulthood for further investigation. The method could be used for other groups of Anura and modified for other species with the ability to regenerate their tissues.

Manipulating the microbiome alters regenerative outcomes in Xenopus laevis tadpoles via lipopolysaccharide signalling.

Xenopus laevis tadpoles can regenerate functional tails, containing the spinal cord, notochord, muscle, fin, blood vessels and nerves, except for a brief refractory period at around 1 week of age. At this stage, amputation of the tadpole's tail may either result in scarless wound healing or the activation of a regeneration programme, which replaces the lost tissues. We recently demonstrated a link between bacterial lipopolysaccharides and successful tail regeneration in refractory stage tadpoles and proposed that this could result from lipopolysaccharides binding to Toll-like receptor 4 (TLR4). Here, we have used 16S rRNA sequencing to show that the tadpole skin microbiome is highly variable between sibships and that the community can be altered by raising embryos in the antibiotic gentamicin. Six Gram-negative genera, including Delftia and Chryseobacterium, were over-represented in tadpoles that underwent tail regeneration. Lipopolysaccharides purified from a commensal Chryseobacterium spp. XDS4, an exogenous Delftia spp. or Escherichia coli, could significantly increase the number of antibiotic-raised tadpoles that attempted regeneration. Conversely, the quality of regeneration was impaired in native-raised tadpoles exposed to the antagonistic lipopolysaccharide of Rhodobacter sphaeroides. Editing TLR4 using CRISPR/Cas9 also reduced regeneration quality, but not quantity, at the level of the cohort. However, we found that the editing level of individual tadpoles was a poor predictor of regenerative outcome. In conclusion, our results suggest that variable regeneration in refractory stage tadpoles depends at least in part on the skin microbiome and lipopolysaccharide signalling, but that signalling via TLR4 cannot account for all of this effect.

Xenopus laevis il11ra.L is an experimentally proven interleukin-11 receptor component that is required for tadpole tail regeneration

Xenopus laevis tadpoles possess high regenerative ability and can regenerate functional tails after amputation. An early event in regeneration is the induction of undifferentiated cells that form the regenerated tail. We previously reported that interleukin-11 (il11) is upregulated immediately after tail amputation to induce undifferentiated cells of different cell lineages, indicating a key role of il11 in initiating tail regeneration. As Il11 is a secretory factor, Il11 receptor-expressing cells are thought to mediate its function. X. laevis has a gene annotated as interleukin 11 receptor subunit alpha on chromosome 1L (il11ra.L), a putative subunit of the Il11 receptor complex, but its function has not been investigated. Here, we show that nuclear localization of phosphorylated Stat3 induced by Il11 is abolished in il11ra.L knocked-out culture cells, strongly suggesting that il11ra.L encodes an Il11 receptor component. Moreover, knockdown of il11ra.L impaired tadpole tail regeneration, suggesting its indispensable role in tail regeneration. We also provide a model showing that Il11 functions via il11ra.L-expressing cells in a non-cell autonomous manner. These results highlight the importance of il11ra.L-expressing cells in tail regeneration.

FGF2 and FGF10 expression patterns in the epidermis and mesenchyme during homeotic transformation of tail into hindlimbs in frog tadpoles

Limbs are trunk quintessential in tetrapods. Their development relies on the Retinoic acid (RA) gradient in association with the Fibroblast Growth Factors (FGFs). The role of various FGFs have been probed extensively and confirmed during the induction of ectopic limbs in vertebrates. On such factual backdrops, we studied the expression patterns of FGF2 and FGF10 in the epidermis and mesenchyme by immunohistochemical localization in the regenerating tails of tadpoles of the Indian tree frog, Polypedates maculatus. These tadpoles are known to exhibit a kind of homeotic transformation of tail to limbs during regeneration, whose exact mechanism is still to be established by scientific investigations. Here in this study, we provide the first evidence of the putative involvement of FGF2 and FGF10 during such ectopic appendage development.

Triiodothyronine, diluted according homeopathic techniques, modifies the programmed cell death of tadpole tail's explants

High Dilution is a solution beyond the Avogadro limits that, in the dependence of the applied succussion elicits a suppressive or a stimulant effect on a living cell, with a consequent generation of an oscillatory dose-effect curve. According to Bonamin et al. [1], “Perhaps, the most enigmatic feature regarding the properties of high dilutions is the non-linearity of their effects. In several studies employing in vivo and ex vivo models, especially involving iso-endopathy, an oscillatory potency-effect curve has appeared. The first observations were initially considered as artifacts, but the repetition of this pattern in different studies involving completely different experimental models, in times and places equally different, points out to the existence of a property intrinsic to dynamized systems.” ...