1-phenyl 2-thiourea (PTU) activates autophagy in zebrafish embryos

Chen, Xiang Ke; Kwan, Joseph; Chang, Raymond Chuen‐Chung; Hang, Alvin Chun

doi:10.1080/15548627.2020.1755119

Cited by 37 publications

(25 citation statements)

References 40 publications

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“…For instance, zebrafish has emerged as an important model organism for human cancer and models of hematological malignancies including myeloproliferative neoplasm (MPN), acute myeloid leukemia (AML), chronic myeloid leukemia (CML) and acute lymphoblastic leukemia (ALL) with highly conserved oncogenic pathways and pharmacologic responses were reported. [59][60][61][62][63] With the well-developed methodologies in autophagy study 32,39,64 and genetic engineering, 65 zebrafish could be developed into a unique modelling platform to define the role of autophagy in hematological malignancies, which will provide important information for the development of autophagy-related therapeutic strategies against these heterogeneous diseases.…”

Section: Discussionmentioning

confidence: 99%

“…29, 35 Conventional pigmentation inhibition with 1-phenyl 2-thiourea (PTU) was refrained to avoid unwanted autophagy induction. 32 Fixed embryos were bleached with 1% KOH and 3% H2O2 before WISH. Live imaging was performed using Lightsheet fluorescent microscope.…”

Section: Methodsmentioning

confidence: 99%

“…The small vertebrate has been widely used to study hematopoiesis and model human hematopoietic malignancies. [29][30][31] While it remains difficult to experimentally observed autophagy in other whole-animal model organisms, zebrafish has been demonstrated as a unique vertebrate model to study autophagy in vivo at cellular level, 32 highlighted the potential of using zebrafish model to study the complex role of autophagy in hematopoiesis. Here we used zebrafish ulk1b and ulk2 mutant models as the paradigm of autophagy dysfunction and depicted the previously unknown role of ulk-dependent autophagy in regulating definitive myelopoiesis.…”

Section: Introductionmentioning

confidence: 99%

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Genetic and chemical inhibition of autophagy in zebrafish induced myeloproliferation

Hasan

Chen

Zhang

et al. 2021

Preprint

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Autophagy is an evolutionary conserved and dynamic lysosomal degradation process for cellular homeostasis and remodelling, which is essential for the development and maintenance of different hematopoietic fates. However, the roles of autophagy in definitive hematopoiesis remain elusive. Here, we exploited zebrafish (Danio rerio) to investigate the effect of knocking-out unc-51 like autophagy activating kinase 1b and 2 (ulk1b and ulk2), homologous of human ULK1 and ULK2, respectively, on definitive hematopoiesis. Upon ulk1b or ulk2 mutation, autophagosome formation was blocked in zebrafish embryos. More importantly, pan-leukocytes (lcp1), common myeloid progenitors (spi1b), neutrophils (mpx), and macrophages (mpeg1.1) significantly elevated, while the hematopoietic stem and progenitor cell (HSPC) (myb), erythroid progenitors (gata1), and embryonic hemoglobin (hbae1.1) significantly reduced in the caudal hematopoietic tissue (CHT) of ulk1b or ulk2 mutant zebrafish embryos. On the other hand, chemically modulated autophagy induction by calpeptin, a downstream autophagy inducer for ulk complex, was insufficient to ameliorate dysregulated hematopoiesis in both ulk1b and ulk2 mutants. Conversely, autophagy inhibitor 3-Methyladenine functioned parallelly with the ulk mutants to maintain defective hematopoiesis. These observations raised a link between autophagy and definitive hematopoiesis and potentiates the fact that autophagy deficiency incorporates with myeloproliferation and anemia, which warrants the significance of autophagy in regulating definitive hematopoiesis.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Genetic and chemical inhibition of autophagy in zebrafish induced myeloproliferation

Hasan

Chen

Zhang

et al. 2021

Preprint

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“…Acridine orange at 10 µg/ml (Sigma-Aldrich) and ApopTag® Fluorescein In Situ Apoptosis Detection Kit (Millipore) were used to detect apoptotic cells in live and xed zebra sh embryos, respectively, following the protocols described previously [32]. In addition, immunostaining together with phospho-Histone H3 (PH3) primary antibody (Cell Signaling Technology) was applied to detect the cells undergoing mitosis according to the previously described protocol [33].…”

Section: Acridine Orange Tunel and Ph3 Stainingmentioning

confidence: 99%

IL-1β-mediated macrophage infiltration into the brain after peripheral tissue injury

Chen

Kwan

Wong

et al. 2020

Preprint

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Background: Infiltration of macrophages into the central nervous system (CNS) is involved in many neurological disorders, such as Alzheimer’s disease (AD), amyotrophic lateral sclerosis (ALS) and autism. Despite extensive studies into neuroinflammation associated macrophage infiltration into the CNS, its underlying mechanisms and pathological roles remain unclear, especially when triggered by peripheral inflammation. Methods: To further elucidate the role and mechanism of peripheral inflammation in neurological disorders, we exploited interleukin 1 beta (il1b) mutant transgenic zebrafish (Danio rerio) with fluorescent protein expression restricted to macrophages to track the macrophage migration under peripheral inflammation following tail amputation.Results: We found that macrophage infiltration into the brain of zebrafish embryo following peripheral tissue injury can be alleviated via genetically targeting il1b. In addition, through circulation-independent migration, macrophages infiltrate brains with evidence of increased apoptosis. We further identified the expression of camk2g1 in the brains of zebrafish with hyperactive behavior following peripheral tissue injury. This il1b-regulated protein is associated with neuropsychiatry disorders. Conclusion: These findings demonstrated that peripheral tissue injury induces il1b-mediated macrophage infiltration into the brain and a hyperactive behavior.

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“…Although zebrafish are relatively transparent during early embryogenesis, onset of pigmentation begins at approximately 24 hours post-fertilization (hpf), progressively obscuring and limiting imaging of different tissues and structures within the developing animal 2 . The formation of the highly opaque melanin pigment in melanocytes can be suppressed by treatment with 1-phenyl 2-thiourea (PTU) beginning prior to 24 hpf [3][4][5] , but PTU is toxic and prolonged treatment leads to developmental defects 6,7 . Genetic mutations that interfere with the development of pigment-producing cells provide an alternative to PTU treatment.…”

Section: Introductionmentioning

confidence: 99%

Rapid generation of pigment free, immobile zebrafish embryos and larvae in any genetic background using CRISPR-Cas9 dgRNPs

Davis

Castranova

Weinstein

2021

Preprint

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The ability to carry out high-resolution, high-magnification optical imaging of living animals is one of the most attractive features of the zebrafish as a model organism. However, formation of obscuring pigmentation as development proceeds and difficulties in maintaining sustained immobilization of healthy, living animals remain challenges that limit the application of live imaging. Chemical treatments can be used to suppress pigment formation and movement, but these treatments can lead to developmental defects. Genetic mutants can also be used to eliminate pigment formation and immobilize animals but maintaining these mutants in lines carrying other combinations of transgenes and mutants is difficult and laborious. Here, we show that CRISPR duplex guide ribonucleoproteins (dgRNPs) targeting the slc45a2 (albino) and chrna1 (nic1) genes can be used to efficiently suppress pigment formation in and immobilize F0 injected animals. CRISPR dgRNPs can be used to generate pigment-free, immobile zebrafish embryos and larvae in any transgenic and/or mutant-carrying background, greatly facilitating high-resolution imaging and analysis of the many transgenic and mutant lines available in the zebrafish.

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1-phenyl 2-thiourea (PTU) activates autophagy in zebrafish embryos

Cited by 37 publications

References 40 publications

Genetic and chemical inhibition of autophagy in zebrafish induced myeloproliferation

Genetic and chemical inhibition of autophagy in zebrafish induced myeloproliferation

IL-1β-mediated macrophage infiltration into the brain after peripheral tissue injury

Rapid generation of pigment free, immobile zebrafish embryos and larvae in any genetic background using CRISPR-Cas9 dgRNPs

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