A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Xenopus Tadpoles

Increasing numbers of substances present in the environment are postulated to have endocrine-disrupting effects on vertebrate populations. However, data on disruption of thyroid signaling are fragmentary, particularly at the molecular level. Thyroid hormone (TH; triiodothyronine, T(3)) acts principally by modulating transcription from target genes; thus, thyroid signaling is particularly amenable to analysis with a transcriptional assay. Also, T(3) orchestrates amphibian metamorphosis, thereby providing an exceptional model for identifying thyroid-disrupting chemicals. We combined these two advantages to develop a method for following and quantifying the transcriptional action of T(3) in Xenopus laevis tadpoles. This technology provides a means of assessing thyroid activity at the molecular level in a physiologically relevant situation. Moreover, translucent tadpoles are amenable to “on-line” imaging with fluorescent reporter constructs that facilitate in vivo measurement of transcriptional activity. We adapted transgenesis with TH-responsive elements coupled to either luciferase or green fluorescent protein to follow T(3)-dependent transcription in vivo. To reduce time of exposure and to synchronize responses, we optimized a physiologic pre-treatment protocol that induced competence to respond to T(3) and thus to assess T(3) effects and T(3) disruption within 48 hr. This pretreatment protocol was based on a short (24 hr), weak (10(−12) M) pulse of T(3) that induced TH receptors, facilitating and synchronizing the transcriptional responses. This protocol was successfully applied to somatic and germinal transgenesis with both reporter systems. Finally, we show that the transcriptional assay allows detection of the thyroid-disrupting activity of environmentally relevant concentrations (10(−8) M) of acetochlor, a persistent herbicide

Stage-dependent cardiac regeneration in Xenopus is regulated by thyroid hormone availability

International audienceDespite therapeutic advances, heart failure is the major cause of morbidity and mortality worldwide, but why cardiac regenerative capacity is lost in adult humans remains an enigma. Cardiac regenerative capacity widely varies across vertebrates. Zebrafish and newt hearts regenerate throughout life. In mice, this ability is lost in the first postnatal week, a period physiologically similar to thyroid hormone (TH)-regulated metamorphosis in anuran amphibians. We thus assessed heart regeneration in Xenopus laevis before, during, and after TH-dependent metamorphosis. We found that tadpoles display efficient cardiac regeneration, but this capacity is abrogated during the metamorphic larval-to-adult switch. Therefore, we examined the consequence of TH excess and deprivation on the efficiently regenerating tadpole heart. We found that either acute TH treatment or blocking TH production before resection significantly but differentially altered gene expression and kinetics of extracellular matrix components deposition, and negatively impacted myocardial wall closure, both resulting in an impeded regenerative process. However, neither treatment significantly influenced DNA synthesis or mitosis in cardiac tissue after amputation. Overall, our data highlight an unexplored role of TH availability in modulating the cardiac regenerative outcome, and present X. laevis as an alternative model to decipher the developmental switches underlying stage-dependent constraint on cardiac regeneration

A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles-2

<p><b>Copyright information:</b></p><p>Taken from "A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles"</p><p>Environmental Health Perspectives 2005;113(11):1588-1593.</p><p>Published online 11 Jul 2005</p><p>PMCID:PMC1310923.</p><p>This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original DOI.</p

A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles-0

<p><b>Copyright information:</b></p><p>Taken from "A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles"</p><p>Environmental Health Perspectives 2005;113(11):1588-1593.</p><p>Published online 11 Jul 2005</p><p>PMCID:PMC1310923.</p><p>This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original DOI.</p

A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles-5

<p><b>Copyright information:</b></p><p>Taken from "A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles"</p><p>Environmental Health Perspectives 2005;113(11):1588-1593.</p><p>Published online 11 Jul 2005</p><p>PMCID:PMC1310923.</p><p>This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original DOI.</p

A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles-3

<p><b>Copyright information:</b></p><p>Taken from "A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles"</p><p>Environmental Health Perspectives 2005;113(11):1588-1593.</p><p>Published online 11 Jul 2005</p><p>PMCID:PMC1310923.</p><p>This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original DOI.</p

A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles-4

<p><b>Copyright information:</b></p><p>Taken from "A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles"</p><p>Environmental Health Perspectives 2005;113(11):1588-1593.</p><p>Published online 11 Jul 2005</p><p>PMCID:PMC1310923.</p><p>This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original DOI.</p

A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles-1

<p><b>Copyright information:</b></p><p>Taken from "A Rapid, Physiologic Protocol for Testing Transcriptional Effects of Thyroid-Disrupting Agents in Premetamorphic Tadpoles"</p><p>Environmental Health Perspectives 2005;113(11):1588-1593.</p><p>Published online 11 Jul 2005</p><p>PMCID:PMC1310923.</p><p>This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original DOI.</p

Colonization kinetics and implantation follow-up of the sewage microbiome in an urban wastewater treatment plant

The Seine-Morée wastewater treatment plant (SM_WWTP), with a capacity of 100,000 population-equivalents, was fed with raw domestic wastewater during all of its start-up phase. Its microbiome resulted from the spontaneous evolution of wastewater-borne microorganisms. This rare opportunity allowed us to analyze the sequential microbiota colonization and implantation follow up during the start-up phase of this WWTP by means of regular sampling carried out over 8 months until the establishment of a stable and functional ecosystem. During the study, biological nitrification-denitrification and dephosphatation occurred 68 days after the start-up of the WWTP, followed by flocs decantation 91 days later. High throughput sequencing of 18S and 16S rRNA genes was performed using Illumina's MiSeq and PGM Ion Torrent platforms respectively, generating 584,647 16S and 521,031 18S high-quality sequence rDNA reads. Analyses of 16S and 18S rDNA datasets show three colonization phases occurring concomitantly with nitrification, dephosphatation and floc development processes. Thus, we could define three microbiota profiles that sequentially colonized the SM_WWTP: the early colonizers, the late colonizers and the continuous spectrum population. Shannon and inverse Simpson diversity indices indicate that the highest microbiota diversity was reached at days 133 and 82 for prokaryotes and eukaryotes respectively; after that, the structure and complexity of the wastewater microbiome reached its functional stability. This study demonstrates that physicochemical parameters and microbial metabolic interactions are the main forces shaping microbial community structure, gradually building up and maintaining a functionally stable microbial ecosystem