Species-specific oxygen sensing governs the initiation of vertebrate limb regeneration

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Dec. 20, 2024

Why mammals cannot regenerate limbs, unlike amphibians, presents a longstanding puzzle in biology. We show that exposing ex vivo amputated embryonic mouse limbs to subatmospheric oxygen environment, or stabilizing oxygen-sensitive HIF1A enables not only rapid wound healing, but alters cellular mechanics, and reshapes the histone landscape prime regenerative fates. Conversely, Xenopus tadpole display low oxygen-sensing capacity, robust landscape, glycolytic programs even under high oxygen. This reduced stark contrast mammals, associates with decreased HIF1A-regulating gene expressions. Our findings thus uncover species-specific sensing as unifying mechanism for limb regeneration initiation across vertebrates, reveal how aquatic habitats may enhance capabilities, identify targetable barriers unlock latent adult mammals.

Language: Английский

---

Multi-species atlas resolves an axolotl limb development and regeneration paradox

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2023, Volume and Issue: unknown

Published: March 1, 2023

Abstract Humans and other tetrapods are considered to require apical-ectodermal-ridge, AER, cells for limb development, AER-like suggested be re-formed initiate regeneration. Paradoxically, the presence of AER in axolotl, primary regeneration model organism, remains controversial. Here, by leveraging a single-cell transcriptomics-based multi-species atlas, composed human, mouse, chicken, frog cells, we first established that axolotls contain with characteristics. Surprisingly, further analyses spatial transcriptomics revealed axolotl limbs do not fully re-form during Moreover, mesoderm displays part machinery, revealing novel program (re)growth. These results clarify debate about extent which developmental is recapitulated

Language: Английский

---

Hybridization Chain Reaction for mRNA Localization in Single Cells from Mouse and Human Cryosections

Current Protocols, Journal Year: 2022, Volume and Issue: 2(5)

Published: May 1, 2022

Abstract In situ hybridization has been a robust method for detection of mRNA expression in whole‐mount samples or tissue sections more than 50 years. Recent technical advances have incorporated oligo‐based probes that attain greater penetration and signal amplification steps with restricted localization visualization specific mRNAs within single cells. One such is third‐generation chain reaction (V3HCR). Here, we report an optimized protocol V3HCR gene using sectioned frozen tissues from mouse human on microscope slides. Our methods modifications cryosectioning, fixation, processing over three‐day are detailed along recommendations aliquoting storing single‐stranded DNA hairpin amplifiers. addition, describe blocking background lipofuscin, highly autofluorescent material widespread neurons often complicates imaging efforts. After testing multiple strategies reduction determined application lipofuscin quencher dye compatible V3HCR, contrast to other like cupric sulfate quenching Sudan Black B cause loss. This adaptation enables neuronal populations otherwise problematic due autofluorescence. © 2022 Wiley Periodicals LLC. Basic Protocol : Mouse fresh‐frozen slides Support Aliquoting HCR hairpins

Language: Английский

---

Gene expression analysis of the Xenopus laevis early limb bud proximodistal axis

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2022, Volume and Issue: unknown

Published: May 4, 2022

Abstract Background Limb buds develop as bilateral outgrowths of the lateral plate mesoderm and are patterned along three axes. Current models proximal to distal patterning early amniote limb suggest that two signals, a organising signal from apical epithelial ridge (AER, Fgfs) an opposing (retinoic acid) act on pattern this axis. Results Transcriptional analysis stage 51 Xenopus laevis hindlimb sectioned proximal-distal axis, showed region is distinct rest limb. Expression capn8 . 3 , novel calpain, was located in cells immediately flanking AER. The Wnt antagonist Dkk1 AER-specific limbs. Two transcription factors, sall1 zic5 were expressed mesenchyme. Zic 5 has no described association with development. We also describe expression genes, gata5 tnn not previously associated Differentially genes Fgf, retinoic acid (RA) signalling well differential cell adhesion proliferation. Conclusions identify new candidate for proximodistal patterning. Our RA-regulated supports role transient RA gradients bud proximal-to-distal anamniote model organism.

Language: Английский

---

Fgf10mutant newts can regenerate normal limbs despite severe developmental hindlimb defects

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2023, Volume and Issue: unknown

Published: May 24, 2023

Abstract In the amniote limb, FGF10 is essential for limb bud initiation and outgrowth. However, whether this function broadly conserved in tetrapods and/or involved adult regeneration remains unknown. To tackle question, we established an Fgf10 null mutant line newt Pleurodeles waltl which have amazing regenerative ability. While forelimbs develop normally, hindlimbs exhibit severe digit reduction, fail to ossify zeugopod, downregulate FGF target genes like Sall1, Runx1 Hoxa11/d11 . Despite these developmental defects, mutants were able regenerate near-normal hindlimbs. Together, our results suggest important role hindlimb formation zeugopod ossification during development, but little or no regeneration, suggesting that different mechanisms operate versus development.

Language: Английский

---