Skip to main content
Advertisement

Main menu

  • Home
  • Articles
    • Accepted manuscripts
    • Issue in progress
    • Latest complete issue
    • Issue archive
    • Archive by article type
    • Subject collections
    • Interviews
    • Sign up for alerts
  • About us
    • About DMM
    • Editors and Board
    • Editor biographies
    • Travelling Fellowships
    • Grants and funding
    • Journal Meetings
    • Workshops
    • The Company of Biologists
    • Journal news
  • For authors
    • Submit a manuscript
    • Aims and scope
    • Presubmission enquiries
    • Article types
    • Manuscript preparation
    • Cover suggestions
    • Editorial process
    • Promoting your paper
    • Open Access
    • Outstanding paper prize
    • Biology Open transfer
  • Journal info
    • Journal policies
    • Rights and permissions
    • Media policies
    • Reviewer guide
    • Sign up for alerts
  • Contact
    • Contact DMM
    • Advertising
    • Feedback
  • COB
    • About The Company of Biologists
    • Development
    • Journal of Cell Science
    • Journal of Experimental Biology
    • Disease Models & Mechanisms
    • Biology Open

User menu

  • Log in

Search

  • Advanced search
Disease Models & Mechanisms
  • COB
    • About The Company of Biologists
    • Development
    • Journal of Cell Science
    • Journal of Experimental Biology
    • Disease Models & Mechanisms
    • Biology Open

supporting biologistsinspiring biology

Disease Models & Mechanisms

Advanced search

RSS   Twitter   Facebook   YouTube

  • Home
  • Articles
    • Accepted manuscripts
    • Issue in progress
    • Latest complete issue
    • Issue archive
    • Archive by article type
    • Subject collections
    • Interviews
    • Sign up for alerts
  • About us
    • About DMM
    • Editors and Board
    • Editor biographies
    • Travelling Fellowships
    • Grants and funding
    • Journal Meetings
    • Workshops
    • The Company of Biologists
    • Journal news
  • For authors
    • Submit a manuscript
    • Aims and scope
    • Presubmission enquiries
    • Article types
    • Manuscript preparation
    • Cover suggestions
    • Editorial process
    • Promoting your paper
    • Open Access
    • Outstanding paper prize
    • Biology Open transfer
  • Journal info
    • Journal policies
    • Rights and permissions
    • Media policies
    • Reviewer guide
    • Sign up for alerts
  • Contact
    • Contact DMM
    • Advertising
    • Feedback
RESEARCH ARTICLE
Establishment of a zebrafish hematological disease model induced by 1,4-benzoquinone
Ao Zhang, Mei Wu, Junliang Tan, Ning Yu, Mengchang Xu, Xutong Yu, Wei Liu, Yiyue Zhang
Disease Models & Mechanisms 2019 12: dmm037903 doi: 10.1242/dmm.037903 Published 28 March 2019
Ao Zhang
1Division of Cell, Developmental and Integrative Biology, School of Medicine, South China University of Technology, Guangzhou 510006, China
2Key Laboratory of Zebrafish Modeling and Drug Screening for Human Diseases of Guangdong Higher Education Institutes, Department of Developmental Biology, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, China
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Ao Zhang
Mei Wu
1Division of Cell, Developmental and Integrative Biology, School of Medicine, South China University of Technology, Guangzhou 510006, China
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Mei Wu
Junliang Tan
2Key Laboratory of Zebrafish Modeling and Drug Screening for Human Diseases of Guangdong Higher Education Institutes, Department of Developmental Biology, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, China
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Junliang Tan
Ning Yu
2Key Laboratory of Zebrafish Modeling and Drug Screening for Human Diseases of Guangdong Higher Education Institutes, Department of Developmental Biology, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, China
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Ning Yu
Mengchang Xu
2Key Laboratory of Zebrafish Modeling and Drug Screening for Human Diseases of Guangdong Higher Education Institutes, Department of Developmental Biology, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, China
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Mengchang Xu
Xutong Yu
2Key Laboratory of Zebrafish Modeling and Drug Screening for Human Diseases of Guangdong Higher Education Institutes, Department of Developmental Biology, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, China
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Xutong Yu
Wei Liu
1Division of Cell, Developmental and Integrative Biology, School of Medicine, South China University of Technology, Guangzhou 510006, China
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Wei Liu
Yiyue Zhang
1Division of Cell, Developmental and Integrative Biology, School of Medicine, South China University of Technology, Guangzhou 510006, China
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Yiyue Zhang
  • For correspondence: mczhangyy@scut.edu.cn
  • Article
  • Figures & tables
  • Info & metrics
  • PDF
Loading

ABSTRACT

Benzene exposure is associated with various hematological disorders, in particular leukemia. The reactive metabolite of benzene, 1,4-benzoquinone (BQ), generated in bone marrow, is suggested to be a key molecule in mediating benzene-induced hematotoxicity and carcinogenicity. However, its pathogenic role remains largely unknown due to a lack of suitable vertebrate whole-organism models. Here, we present an in vivo study to reveal the effect of BQ exposure on hematotoxicity in zebrafish. From embryonic stages to adulthood, BQ exposure suppressed erythroid and lymphoid hematopoiesis but led to abnormal accumulation of myeloid cells and precursors, which resembles benzene-induced cytopenia and myeloid dysplasia in humans. This myeloid expansion is caused by granulocyte, but not macrophage, lineage, emphasizing the significant role of lineage specificity in BQ-mediated hematopoietic toxicity. Analysis of the c-myb (also known as myb)-deficient mutant cmybhkz3 revealed that BQ induced neutrophilia in a c-myb-dependent manner, demonstrating that c-myb is a key intrinsic mediator of BQ hematotoxicity. Our study reveals that BQ causes lineage-specific hematotoxicity in zebrafish from embryonic stages to adulthood. Since c-myb is indispensable for BQ to induce neutrophilia, c-myb could serve as a potential drug target for reversing BQ hematotoxicity.

INTRODUCTION

For decades, epidemiological studies have found that occupational and environmental exposure to benzene can have harmful effects on the immunological, neurological, reproductive and, especially, hematological systems, causing cytopenia, aplastic anemia and leukemia (Wilbur et al., 2007). Benzene is classified as a Group 1 carcinogen in humans and animals (IARC, 1982). Benzene metabolism occurs principally in the liver and lungs, with secondary metabolism in bone marrow (BM) (McHale et al., 2012). 1,4-Benzoquinone (BQ) is generated by benzene in the BM, and it causes damage by forming protein and DNA adducts, and producing reactive oxygen species (ROS) (Snyder and Hedli, 1996; Wan and Winn, 2007), Subsequently, excess ROS induce oxidative stress and promote apoptosis (Circu and Aw, 2010). It has been suggested that BQ plays crucial roles in mediating benzene-induced hematological toxicity and carcinogenicity. In support of this, previous studies have demonstrated that the BQ-detoxifying enzyme NAD(P)H: quinone oxidoreductase 1, mutation of which is associated with human susceptibility to benzene hematological toxicity (Rothman et al., 1997), can protect mice against benzene-induced myelodysplasia (Long et al., 2002; Iskander and Jaiswal, 2005). Previous in vitro studies have shown that exposure of murine hematopoietic stem and progenitor cells (HSPCs) to BQ interferes with their physiological properties and changes their clonogenic potency by altering genes for apoptosis, DNA repair, cell cycle, self-renewal and differentiation (Chow et al., 2018; Faiola et al., 2004). Son et al. (2016) analyzed the impact that BQ exposure has on DNA-repair-defective mouse embryonic stem cells. They proposed that BQ suppresses type 1 topoisomerases to inhibit replication fork restart and progression, leading to chromosomal instability that has the potential to cause hematopoietic disorders. To date, there is no vertebrate model for hematological toxicity of BQ exposure, which limits the investigation at the whole-organism level.

Although BQ is known to be involved in hematological toxicity and cancer, how embryonic and adult hematopoiesis are affected and the molecular and cellular bases have not been fully elucidated. BQ metabolism and its molecular mechanisms have been studied for several decades, but animal models for evaluating the hematological effects of BQ from embryonic stages to adulthood are still lacking. The zebrafish is an effective vertebrate model for studying hematopoiesis in vivo and for investigating the pathogenesis of hematological disorders, owing to high fecundity, optical transparency and highly conserved hematopoiesis (Rasighaemi et al., 2015). In this study, we investigated the hematotoxicity of BQ in a zebrafish model, and found that the BQ-treated zebrafish exhibited cytopenia and myeloid dysplasia, which resembled benzene-induced hematotoxicity in mammals.

RESULTS

Embryotoxicity and teratogenicity of BQ in zebrafish

To investigate the effect of BQ on hematopoiesis, zebrafish embryos were treated with BQ. We first determined the embryotoxicity and teratogenicity of BQ by recording the survival and malformation rates every 24 h after BQ exposure. Absence of swim activity, heart beat and tail blood flow were used as criteria to differentiate a viable from a non-viable larval zebrafish. The Kaplan–Meier curve showed that, at 7 days post-fertilization (dpf), recorded survival rates were 94% and 77% in the control and 8 μM BQ groups, respectively. The survival rate was markedly decreased as the concentration of BQ increased to 10 μM (Fig. 1A). Malformations, such as yolk sac edema, spine malformation and pericardial edema, were occasionally observed in developing embryos (∼6%) (Fig. 1B,C). However, BQ exposure resulted in a dose-dependent increase in malformation rate, ∼13% and ∼31% in 8 μM BQ and 10 μM BQ groups, respectively, but the percentages of each malformation at all three doses were not significantly different (Fig. 1B). The above data demonstrated the fetal toxicity of BQ. Thus, the concentration of 8 μM BQ was chosen in subsequent experiments to evaluate BQ hematotoxicity in zebrafish.

Fig. 1.
  • Download figure
  • Open in new tab
  • Download powerpoint
Fig. 1.

Kaplan–Meier survival curve and analysis of zebrafish embryo phenotype following exposure to 1,4-benzoquinone (BQ). (A) Survival rates in zebrafish embryos following continuous exposure to BQ (n=278 per group) in the range 8–10 μM, from 2 dpf to 9 dpf (log-rank test, P<0.001). (B) Phenotypic traits (indicated by the arrows in C) observed following BQ exposure (Chi-squared test, mean±s.d., *P<0.05, **P<0.01). (C) Typical malformations were observed in zebrafish embryos exposed to BQ. Scale bars: 500 μm.

BQ exposure results in abnormal hematopoiesis in zebrafish embryos

Occupational and environmental exposure to benzene is associated with the incidence of hematological disorders and malignancies, such as aplastic anemia, myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) (Baan et al., 2009; Linet et al., 2015; McHale et al., 2012), as well as MDS progression to AML (Greenberg et al., 2012; Poynter et al., 2017; Schnatter et al., 2012). To explore the hematotoxicity of BQ, we monitored zebrafish embryonic hematopoiesis by performing whole-mount in situ hybridization (WISH) of lineage specific markers in 3 dpf and 5 dpf larval zebrafish. Expression of the erythroid marker, βe1-globin (also known as hbbe1.1) (Paffett-Lugassy et al., 2007), was decreased in BQ-exposed embryos (Fig. 2A,H). Likewise, expression of the lymphoid marker rag1 (Willett et al., 1997) was also reduced (Fig. 2B,I). These data indicate that BQ may lead to deficiencies of erythrocytes and lymphocytes, which is consistent with cytopenia and anemia in patients exposed to benzene (Aksoy, 1989). For myeloid lineage, expression of l-plastin (Herbomel et al., 2001), a marker for both neutrophils and macrophages, was significantly increased in the BQ-treated embryos (Fig. 2C,J). To unveil whether the myeloid expansion was caused by granulocyte or macrophage lineage cells, we checked the neutrophil and macrophage markers, respectively. Expression of the macrophage lineage marker mfap4 (Zakrzewska et al., 2010) was decreased in BQ-treated embryos (Fig. 2D,K). However, expression of the granulocytic marker mpx (Lieschke et al., 2001) (Fig. 2E,L) and Sudan Black B (SB) staining (Le Guyader et al., 2008) (Fig. 2F,M) were significantly elevated after BQ exposure, suggesting robust expansion of granulocytic lineage cells in BQ-exposed embryos. We also found that another granulocytic marker, lyz (Liu and Wen, 2002), was increased as well (Fig. 2G,N), which was further supported by flow cytometry analysis of Tg(lyz:GFP) embryos (Fig. 2O,P). These data demonstrate that BQ causes anemia and neutrophilia in zebrafish embryos, which resembles the hematotoxicity of benzene in mammals (Snyder and Kocsis, 1975).

Fig. 2.
  • Download figure
  • Open in new tab
  • Download powerpoint
Fig. 2.

Expression of lineage-specific markers in zebrafish was detected using WISH and SB staining. (A,B) Loss of βe1-globin (n=20) and rag1 (control group n=37, BQ group n=49) expression in 5 dpf embryos exposed to BQ. The red boxed regions show magnified views of the caudal hematopoietic tissue (CHT). Relative rag1+ thymocyte-signal areas were analyzed using ImageJ software. The circled regions show the thymus. (C) Increase in l-plastin expression upon BQ exposure at 3 dpf (control group n=25, BQ group n=30). (D) Expression of the macrophage marker mfap4 was reduced in the BQ exposure group (control group n=52, BQ group n=76). (E,G) The neutrophil markers mpx (E) (control group n=20, BQ group n=33) and lyz (G) (control group n=39, BQ group n=35) both showed increased expression in embryos at 3 dpf following BQ exposure. (F) Increased SB+ cells were observed in the BQ exposure group compared with the control group (n=21 per group). (H–N) Quantification of WISH and SB staining for A–E (Student's t-test, mean±s.d., **P<0.01, ***P<0.001). (O) Dot plot of flow cytometry analysis of lyz:GFP+ cells from control (left) and BQ-treated fish (right). Three independent experiments (in each group, 100 embryos are pooled together) were conducted. (P) Percentage of lyz+ cells in each group [0.30% in the control group and 0.40% in the BQ group; Chi-squared test (95% c.i.), ***P<0.001].

BQ exposure causes neutrophilia due to accelerated neutrophil proliferation

To establish whether the increase in neutrophils after BQ exposure is due to accelerated cell proliferation, we performed a bromodeoxyuridine (BrdU) incorporation assay, which is commonly used for detection of proliferating cells in living tissues (Lehner et al., 2011). Embryos of the neutrophil-specific transgenic line Tg(lyz:dsRed) were exposed to BQ and subjected to BrdU incorporation assay at 3 dpf. The proportion of BrdU and lyz:dsRed double-positive neutrophils among total lyz:dsRed+ neutrophils in the caudal hematopoietic tissue (CHT) region was increased in the BQ-treated compared with control group (Fig. 3A,B), indicating that neutrophil proliferation was accelerated by BQ.

Fig. 3.
  • Download figure
  • Open in new tab
  • Download powerpoint
Fig. 3.

Exposure to BQ induced neutrophil proliferation in zebrafish embryos. (A,B) BrdU incorporation assay. Double staining of BrdU/lyz (A) showed BrdU incorporation of CHT lyz+ cells in BQ-exposed embryos and controls at 3 dpf. Arrows indicate lyz/BrdU double-positive cells. Scale bars: 50 μm. Percentage of the CHT-localized lyz+ myeloid cells that incorporate BrdU (B) in lyz+ myeloid cells (Student's t-test, control group n=30, BQ group n=26, mean±s.d., **P<0.01).

BQ promotes neutrophilia through c-myb

The proto-oncogene c-myb (also known as myb) is a key regulator of hematopoietic cell proliferation and differentiation, and correct levels of c-myb are essential for regulating distinct differentiation steps during hematopoietic cell development, as well as in leukemogenesis (Sakamoto et al., 2006; Wolff, 1996). Our previous study in zebrafish showed that c-myb is essential for neutrophil differentiation (Jin et al., 2016), and c-myb activation causes MDS-like phenotypes due to enhanced myeloid cell proliferation (Liu et al., 2017). It has been shown that exposure to BQ results in increased c-myb transcriptional activity and phosphorylation of c-Myb in cell lines (Singh and Winn, 2008; Wan et al., 2005), although in vivo evidence is still required. We monitored whether c-myb expression was altered upon BQ exposure in zebrafish. By performing WISH and reverse transcription quantitative polymerase chain reaction (RT-qPCR), we found that, in BQ-treated embryos, c-myb+ signals were significantly elevated in the aorta-gonad-mesonephros (AGM) region (Fig. 4A,B), and c-myb expression was increased as well (Fig. 4C). To examine whether c-myb is a mediator for BQ-induced neutrophilia, we treated null c-mybhkz3/hkz3 mutants with BQ (Jin et al., 2016). Neutrophil-specific markers (lyz, mpx and SB staining) were elevated in siblings after BQ exposure (Fig. 5A,C,E), as described in wild-type embryos (Fig. 2D–F). However, neutrophils were not further induced by BQ when c-myb was absent, as demonstrated by the unaltered neutrophil markers with or without BQ treatment in c-mybhkz3/hkz3 mutants (Fig. 5B,D,F). These data suggest that c-myb is required for BQ-induced neutrophilia, and that it functions as a key mediator for BQ hematotoxicity in vivo.

Fig. 4.
  • Download figure
  • Open in new tab
  • Download powerpoint
Fig. 4.

Effect of BQ exposure on c-myb expression. (A–C) BQ exposure increased c-myb expression. Both WISH performed at 36 hpf (A,B) and RT-qPCR performed at 2 dpf (C) indicated increased expression of c-myb in BQ-exposed (right, A) and control (left, A) groups (Student's t-test, n=25 or 26, mean±s.d., *P<0.05, ***P<0.001). The red boxed regions show magnified views of the aorta-gonad-mesonephros (AGM).

Fig. 5.
  • Download figure
  • Open in new tab
  • Download powerpoint
Fig. 5.

BQ-induced neutrophil expansion was blocked in c-myb-deficient mutants. (A–I) Expression of the neutrophil markers lyz (A,B) (sibling control group n=65, BQ group n=38; mutant control group n=25, BQ group n=35) and mpx (C,D) (sibling control group n=83, BQ group n=47; mutant control group n=31, BQ group n=16) was analyzed using WISH, and SB staining (E,F) (sibling control group n=54, BQ group n=18; mutant control group n=25, BQ group n=22) was detected, in c-mybhkz3/+ intercrossed progenies in BQ-treated groups (right column) and untreated controls (left column). Signals increased following BQ exposure at 3 dpf in siblings (A,C,E) but were unaltered in c-mybhkz3/hkz3 mutants (B,D,F). The red boxed regions show magnified views of the CHT. (G–I) Quantification of WISH and SB staining for panels A–F (Student's t-test, mean±s.d., **P<0.01, ****P<0.0001; ns, non-significant).

BQ exposure causes myeloid dysplasia in adult fish

To explore whether adult hematopoiesis could be affected by BQ, we further investigated the cytological changes in myelogram of adult zebrafish following BQ exposure. Kidney marrow (KM) cells were collected from adult fish in the control and BQ exposure groups for cytological analysis and blood cell count. In BQ-exposed fish KM, the proportion of erythrocytes decreased, whereas the proportions of neutrophils and precursors were significantly increased (Fig. 6A,B). Exposure to benzene can lead to leukemia in humans, especially myeloid leukemia (Schnatter et al., 2012). The increase in hematopoietic precursors in the KM of BQ-treated fish resembled the hematopoietic disorders in benzene-exposed mice and humans, which suggested a tendency to progress to hematopoietic malignancy.

Fig. 6.
  • Download figure
  • Open in new tab
  • Download powerpoint
Fig. 6.

BQ hematotoxicity to adult fish. (A) May–Grunwald/Giemsa of KM cells in BQ-exposed (right) and control (left) fish. Red arrows, erythrocytes (oval-shaped nucleus and cytoplasm); black arrows, lymphocytes (large round-shaped nucleus surrounded by minimal cytoplasm); green stars, neutrophils (segmented nucleus and cytoplasm with distinct neutral granules); yellow stars, macrophages (large irregular-shaped cytoplasm with distinct vacuoles); blue arrows, precursors (large and dense nucleus with dark cytoplasm). Scale bars: 5 μm. (B) Blood cell counts of KM in each group were calculated manually based on their morphology (Student's t-test, n=9, mean±s.d., *P<0.05, **P<0.01).

DISCUSSION

BQ is a reactive metabolite of benzene that is produced in BM. It is believed to be mainly responsible for the myelotoxicity/myeloid neoplasms observed in the BM of people exposed to benzene (Hartwig, 2010). To establish the mechanism linking benzene exposure to hematotoxicity and carcinogenicity at the whole-organism level, we explored the effects of BQ on hematopoiesis and hematological changes in zebrafish. We showed that exposure to BQ increased the mortality and malformation rate in a dose-dependent manner, and the rate of malformations in multiple organs, suggesting BQ embryotoxicity in zebrafish. We demonstrated that BQ caused hematopoiesis perturbation from embryonic stages to adulthood, resembling human hematological diseases. At a molecular level, we uncovered c-myb as a key mediator of BQ-induced neutrophilia. Our study implies that c-myb serves as a molecular marker for evaluation of hematotoxicity upon BQ exposure, and that it could serve as a potential drug target for reversing BQ hematotoxicity.

We found that non-lethal doses of BQ impaired erythroid and lymphoid hematopoiesis and abnormally increased neutrophils. The myeloid dysplasia could be partially interpreted by the acceleration of myeloid cell proliferation upon BQ exposure. Likewise, in adult fish, BQ exposure also resulted in reduction in other blood cell lineages but an increase in neutrophils, while the proportion of precursors increased in KM as well, similar to the benzene-induced cytopenia and myeloid dysplasia in humans. Our data suggest that, from embryonic stages to adulthood, BQ induces abnormal expansion of myeloid cells, which may increase the risk of myeloid leukemia. However, more studies are required to confirm whether the duration of BQ treatment is long enough to induce hematological malignancies.

For the molecular and cellular mechanisms underlying BQ hematotoxicity, several molecular pathways have been proposed, such as type 1 topoisomerases (Son et al., 2016). It has been shown that BQ modulates the fate of HSPCs by altering the self-renewal- and differentiation-related genes, such as Bmi-1 and GATA3 (Chow et al., 2018). We found that c-myb is a key mediator in the effect of BQ on neutrophil expansion. How these molecules and pathways crosstalk upon BQ exposure requires further elucidation, which will help us to understand how benzene and BQ mediate leukemogenesis.

Clinically, c-myb is highly expressed in leukemic cells in patients with AML, chronic myeloid leukemia (CML) and acute lymphoblastic leukemia (Machov; Polakov; et al., 2011; Siegert et al., 1990). It is essential for the proliferation and maintenance of leukemic cells, and aberrant c-myb activity is associated with human myeloid leukemia (Westin et al., 1982). High expression of c-myb is believed to be associated with oncogenic activity and poor prognosis in human AML (Gopal et al., 1992). Our previous study revealed that zebrafish with c-mybhyper (c-myb with hyperactivity) display MDS-like phenotypes from embryonic stages, and can develop myeloid and lymphoid leukemia-like phenotypes in adulthood (Liu et al., 2017). In this study, BQ caused myeloid cell expansion in neutrophil, but not macrophage, lineage cells, which is similar to the hematopoietic phenotype in c-mybhyper zebrafish. Consistently, the upregulated c-myb level upon BQ treatment suggests that BQ leads to hematological disorders via c-myb. In the absence of c-myb, BQ had no effect on neutrophils, implying that BQ hematotoxicity could, at least partially, be reversed by targeting c-myb.

In summary, we generated an animal model for in vivo investigation of the hematotoxicity of BQ exposure. Our data indicate that BQ exposure promoted proliferation of myeloid cells but suppressed other blood lineage cells in a zebrafish model, which shares similar pathological features with myeloid dysplasia in humans. At a molecular level, c-myb is a key factor in the response to BQ-mediated hematotoxicity, and it could serve as a potential target for preventing or reversing BQ or benzene hematotoxicity.

MATERIALS AND METHODS

Zebrafish husbandry

All experiments were performed according to the guidelines laid down by the Institutional Animal Care and Use Committee of Southern Medical University and the South China University of Technology. The following strains were used and staged under standard conditions as described previously: wild-type ABSR, Tg(lyz:dsRed) (Hall et al., 2007) and c-mybhkz3 mutant (Zhang et al., 2011). Tg(lyz:dsRed)+/+; c-mybhkz3/+ were crossed with c-mybhkz3/+ and the progeny of c-mybhkz3/hkz3 mutants and siblings were separated based on dsRed (dsRed+ embryos represented siblings and dsRed− embryos were mutants). Embryos were obtained by natural breeding and maintained at 28±0.5°C for subsequent experiments.

BQ exposure

BQ (Sigma-Aldrich) was dissolved in 100% dimethyl sulfoxide (DMSO). The stock solution was 40 mmol/l, and the treatment solutions (8–10 μmol/l) were obtained by dilution with egg water. The final DMSO concentration in the control was equal to that in each exposure group. Dechorionated embryos were exposed to the treatment solutions of BQ at 24 h post-fertilization (hpf). Live embryos and malformed embryos were counted every 24 h, and dead embryos were removed from the culture.

WISH

WISH was performed with antisense digoxigenin-labeled RNA probes based on a standard protocol (Westerfield, 2007). The following probes were used: lyz, mpx, l-plastin, mfap4, rag1, βe1-globin and c-myb. For quantification, the relative rag1+ signal areas were analyzed using ImageJ (https://imagej.nih.gov/ij/) software, and signal counts for the other probes were calculated in the CHT region manually under the stereomicroscope (Olympus).

SB staining

Fixed embryos at 5 dpf were incubated in SB solution (Sigma-Aldrich) and washed, according to standard protocols. The stained neutrophils in the CHT region were counted under a stereomicroscope (Olympus).

RT-qPCR

Total RNA was extracted from the embryos at 2 dpf using TRIzol reagent (Invitrogen) and reverse transcribed with M-MLV Reverse Transcriptase (Promega) according to the manufacturer’s instructions. Complementary DNA from embryos was used for qPCR using a LightCycler+ 96 machine (Roche).

Cytological analysis

After adult zebrafish were soaked in 2 μmol/l BQ for 7 days, KM cells were re-suspended in 5% fetal bovine serum, followed by centrifugation at 18 g for 3 min. The cells were stained as described previously (Liu et al., 2017). Blood cells of KM were calculated manually based on their morphology (Carradice and Lieschke, 2008; Stachura and Traver, 2011).

BrdU labeling

Tg(lyz:dsRed) embryos at 3 dpf were incubated in 10 mM BrdU (Sigma-Aldrich) for 2 h. After BrdU treatment, the embryos were washed and fixed for immunohistochemistry as described previously (Lin et al., 2017). The embryos were stained with mouse anti-BrdU (1:50; 11170376001, Roche), followed by Alexa Fluor goat anti-mouse-488 (1:400; A-11001, Invitrogen) for fluorescent visualization.

Flow cytometry analysis

Tg(lyz:GFP) embryos were collected at 3 dpf, and 100 embryos in each group were pooled together and homogenized by needles and syringes. The homogenized samples were then treated with 0.25% Trypsin-EDTA (25200-072, Gibco) at 30°C for 30 min and the reaction was stopped by adding calcium chloride. Suspended cells were collected by centrifugation (400 g at 4°C), filtered by 35-μm nylon mesh (352235, FALCON) and finally subjected to flow cytometry analysis (FACS Aria IIIu, BD Biosciences). At least 10,000 events were collected for each sample. The live cells were gated using side scatter-A (SSC-A) (granularity) and forward scatter-A (FSC-A) (size) parameters. Discrimination of aggregates from singlets was performed using forward scatter-W (FSC-W) versus FSC-A, and the cell counts in the GFP+ gate were given as the percentages of the total singlet population.

Acknowledgements

We thank Xiaohui Chen for assisting with the maintenance of zebrafish lines, and Jin Xu and Zhibin Huang for providing helpful suggestions.

Footnotes

  • Competing interests

    The authors declare no competing or financial interests.

  • Author contributions

    Conceptualization: Y.Z.; Methodology: A.Z., M.W., W.L.; Validation: A.Z., M.W., J.T.; Formal analysis: A.Z.; Investigation: A.Z., M.W., J.T., N.Y., M.X., X.Y.; Writing - original draft: A.Z., Y.Z.; Writing - review & editing: A.Z., Y.Z.; Visualization: A.Z., M.W., J.T., N.Y., M.X., W.L.; Supervision: Y.Z.; Funding acquisition: W.L., Y.Z.

  • Funding

    This work was supported by the National Natural Science Foundation of China (31271574, 81770167) and the Natural Science Foundation of Guangdong Province (2016A030310378).

  • Received November 8, 2018.
  • Accepted March 11, 2019.
  • © 2019. Published by The Company of Biologists Ltd
http://creativecommons.org/licenses/by/4.0

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed.

References

  1. Agency for Toxic Substances and Disease Registry (ATSDR) (2007). Toxicological profile for benzene. Atlanta: US Department of Health and Human Services, Public Health Service.
  2. ↵
    1. Aksoy, M.
    (1989). Hematotoxicity and carcinogenicity of benzene. Environ. Health Perspect. 82, 193-197. doi:10.1289/ehp.8982193
    OpenUrlCrossRefPubMedWeb of Science
  3. ↵
    1. Baan, R.,
    2. Grosse, Y.,
    3. Straif, K.,
    4. Secretan, B.,
    5. El Ghissassi, F.,
    6. Bouvard, V.,
    7. Benbrahim-Tallaa, L.,
    8. Guha, N.,
    9. Freeman, C.,
    10. Galichet, L. et al.
    (2009). A review of human carcinogens–Part F: chemical agents and related occupations. Lancet Oncol. 10, 1143-1144. doi:10.1016/S1470-2045(09)70358-4
    OpenUrlCrossRefPubMedWeb of Science
  4. ↵
    1. Carradice, D. and
    2. Lieschke, G. J.
    (2008). Zebrafish in hematology: sushi or science? Blood 111, 3331-3342. doi:10.1182/blood-2007-10-052761
    OpenUrlAbstract/FREE Full Text
  5. ↵
    1. Chow, P. W.,
    2. Rajab, N. F.,
    3. Chua, K. H.,
    4. Chan, K. M. and
    5. Abd Hamid, Z.
    (2018). Differential responses of lineages-committed hematopoietic progenitors and altered expression of self-renewal and differentiation-related genes in 1,4-benzoquinone (1,4-BQ) exposure. Toxicol. In Vitro 46, 122-128. doi:10.1016/j.tiv.2017.10.001
    OpenUrlCrossRef
  6. ↵
    1. Circu, M. L. and
    2. Aw, T. Y.
    (2010). Reactive oxygen species, cellular redox systems, and apoptosis. Free Radic. Biol. Med. 48, 749-762. doi:10.1016/j.freeradbiomed.2009.12.022
    OpenUrlCrossRefPubMedWeb of Science
  7. ↵
    1. Faiola, B.,
    2. Fuller, E. S.,
    3. Wong, V. A.,
    4. Pluta, L.,
    5. Abernethy, D. J.,
    6. Rose, J. and
    7. Recio, L.
    (2004). Exposure of hematopoietic stem cells to benzene or 1,4-Benzoquinone induces Gender-Specific gene expression. Stem Cells 22, 750-758. doi:10.1634/stemcells.22-5-750
    OpenUrlCrossRefPubMedWeb of Science
  8. ↵
    1. Gopal, V.,
    2. Hulette, B.,
    3. Li, Y. Q.,
    4. Kuvelkar, R.,
    5. Raza, A.,
    6. Larson, R.,
    7. Goldberg, J.,
    8. Tricot, G.,
    9. Bennett, J. and
    10. Preisler, H.
    (1992). c-myc and c-myb expression in acute myelogenous leukemia. Leuk. Res. 16, 1003-1011. doi:10.1016/0145-2126(92)90080-Q
    OpenUrlCrossRefPubMed
  9. ↵
    1. Greenberg, P. L.,
    2. Tuechler, H.,
    3. Schanz, J.,
    4. Sanz, G.,
    5. Garcia-Manero, G.,
    6. Solé, F.,
    7. Bennett, J. M.,
    8. Bowen, D.,
    9. Fenaux, P.,
    10. Dreyfus, F. et al.
    (2012). Revised international prognostic scoring system for myelodysplastic syndromes. Blood 120, 2454-2465. doi:10.1182/blood-2012-03-420489
    OpenUrlAbstract/FREE Full Text
  10. ↵
    1. Hall, C.,
    2. Flores, M. V.,
    3. Storm, T.,
    4. Crosier, K. and
    5. Crosier, P.
    (2007). The zebrafish lysozyme C promoter drives myeloid-specific expression in transgenic fish. BMC Dev. Biol. 7, 42. doi:10.1186/1471-213X-7-42
    OpenUrlCrossRefPubMed
  11. ↵
    1. Hartwig, A.
    (2010). The role of DNA repair in benzene-induced carcinogenesis. Chem. Biol. Interact. 184, 269-272. doi:10.1016/j.cbi.2009.12.029
    OpenUrlCrossRefPubMed
  12. ↵
    1. Herbomel, P.,
    2. Thisse, B. and
    3. Thisse, C.
    (2001). Zebrafish early macrophages colonize cephalic mesenchyme and developing brain, retina, and epidermis through a M-CSF receptor-dependent invasive process. Dev. Biol. 238, 274-288. doi:10.1006/dbio.2001.0393
    OpenUrlCrossRefPubMedWeb of Science
  13. ↵
    IARC (1982). Some industrial chemicals and dyestuffs. IARC Monogr. Eval. Carcinog Risk Chem. Hum. 29, 1-398.
    OpenUrlPubMed
  14. ↵
    1. Iskander, K. and
    2. Jaiswal, A. K.
    (2005). Quinone oxidoreductases in protection against myelogenous hyperplasia and benzene toxicity. Chem-Biol. Interact. 153-154, 147-157. doi:10.1016/j.cbi.2005.03.019
    OpenUrlCrossRef
  15. ↵
    1. Jin, H.,
    2. Huang, Z.,
    3. Chi, Y.,
    4. Wu, M.,
    5. Zhou, R.,
    6. Zhao, L.,
    7. Xu, J.,
    8. Zhen, F.,
    9. Lan, Y.,
    10. Li, L. et al.
    (2016). c-Myb acts in parallel and cooperatively with Cebp1 to regulate neutrophil maturation in zebrafish. Blood 128, 415-426. doi:10.1182/blood-2015-12-686147
    OpenUrlAbstract/FREE Full Text
  16. ↵
    1. Le Guyader, D.,
    2. Redd, M. J.,
    3. Colucci-Guyon, E.,
    4. Murayama, E.,
    5. Kissa, K.,
    6. Briolat, V.,
    7. Mordelet, E.,
    8. Zapata, A.,
    9. Shinomiya, H. and
    10. Herbomel, P.
    (2008). Origins and unconventional behavior of neutrophils in developing zebrafish. Blood 111, 132-141. doi:10.1182/blood-2007-06-095398
    OpenUrlAbstract/FREE Full Text
  17. ↵
    1. Lehner, B.,
    2. Sandner, B.,
    3. Marschallinger, J.,
    4. Lehner, C.,
    5. Furtner, T.,
    6. Couillard-Despres, S.,
    7. Rivera, F. J.,
    8. Brockhoff, G.,
    9. Bauer, H.-C.,
    10. Weidner, N. et al.
    (2011). The dark side of BrdU in neural stem cell biology: detrimental effects on cell cycle, differentiation and survival. Cell Tissue Res. 345, 313-328. doi:10.1007/s00441-011-1213-7
    OpenUrlCrossRefPubMedWeb of Science
  18. ↵
    1. Lieschke, G.,
    2. Oates, A.,
    3. Crowhurst, M.,
    4. Ward, A. and
    5. Layton, J.
    (2001). Morphologic and functional characterization of granulocytes and macrophages in embryonic and adult zebrafish. Blood 98, 3087-3096. doi:10.1182/blood.V98.10.3087
    OpenUrlAbstract/FREE Full Text
  19. ↵
    1. Lin, Q.,
    2. Zhang, Y.,
    3. Zhou, R.,
    4. Zheng, Y.,
    5. Zhao, L.,
    6. Huang, M.,
    7. Zhang, X.,
    8. Leung, A. Y. H.,
    9. Zhang, W. and
    10. Zhang, Y.
    (2017). Establishment of a congenital amegakaryocytic thrombocytopenia model and a thrombocyte-specific reporter line in zebrafish. Leukemia 31, 1206-1216. doi:10.1038/leu.2016.320
    OpenUrlCrossRef
  20. ↵
    1. Linet, M. S.,
    2. Yin, S.-N.,
    3. Gilbert, E. S.,
    4. Dores, G. M.,
    5. Hayes, R. B.,
    6. Vermeulen, R.,
    7. Tian, H.-Y.,
    8. Lan, Q.,
    9. Portengen, L.,
    10. Ji, B.-T. et al.
    (2015). A retrospective cohort study of cause-specific mortality and incidence of hematopoietic malignancies in Chinese benzene-exposed workers. Int. J. Cancer 137, 2184-2197. doi:10.1002/ijc.29591
    OpenUrlCrossRef
  21. ↵
    1. Liu, F. and
    2. Wen, Z.
    (2002). Cloning and expression pattern of the lysozyme C gene in zebrafish. Mech. Dev. 113, 69-72. doi:10.1016/S0925-4773(01)00658-X
    OpenUrlCrossRefPubMed
  22. ↵
    1. Liu, W.,
    2. Wu, M.,
    3. Huang, Z.,
    4. Lian, J.,
    5. Chen, J.,
    6. Wang, T.,
    7. Leung, A. Y. H.,
    8. Liao, Y.,
    9. Zhang, Z.,
    10. Liu, Q. et al.
    (2017). c-myb hyperactivity leads to myeloid and lymphoid malignancies in zebrafish. Leukemia 31, 222-233. doi:10.1038/leu.2016.170
    OpenUrlCrossRef
  23. ↵
    1. Long, D. J., II.,
    2. Gaikwad, A.,
    3. Multani, A.,
    4. Pathak, S.,
    5. Montgomery, C. A.,
    6. Gonzalez, F. J. and
    7. Jaiswal, A. K.
    (2002). Disruption of the NAD(P)H: quinone oxidoreductase 1 (NQO1) gene in mice causes myelogenous hyperplasia. Cancer Res. 62, 3030-3036.
    OpenUrlAbstract/FREE Full Text
  24. ↵
    1. Machová Polaková, K.,
    2. Lopotová, T.,
    3. Klamová, H.,
    4. Burda, P.,
    5. Trněný, M.,
    6. Stopka, T. and
    7. Moravcová, J.
    (2011). Expression patterns of microRNAs associated with CML phases and their disease related targets. Mol. Cancer 10, 41. doi:10.1186/1476-4598-10-41
    OpenUrlCrossRefPubMed
  25. ↵
    1. McHale, C. M.,
    2. Zhang, L. and
    3. Smith, M. T.
    (2012). Current understanding of the mechanism of benzene-induced leukemia in humans: implications for risk assessment. Carcinogenesis 33, 240-252. doi:10.1093/carcin/bgr297
    OpenUrlCrossRefPubMedWeb of Science
  26. ↵
    1. Paffett-Lugassy, N.,
    2. Hsia, N.,
    3. Fraenkel, P. G.,
    4. Paw, B.,
    5. Leshinsky, I.,
    6. Barut, B.,
    7. Bahary, N.,
    8. Caro, J.,
    9. Handin, R. and
    10. Zon, L. I.
    (2007). Functional conservation of erythropoietin signaling in zebrafish. Blood 110, 2718-2726. doi:10.1182/blood-2006-04-016535
    OpenUrlAbstract/FREE Full Text
  27. ↵
    1. Poynter, J. N.,
    2. Richardson, M.,
    3. Roesler, M.,
    4. Blair, C. K.,
    5. Hirsch, B.,
    6. Nguyen, P.,
    7. Cioc, A.,
    8. Cerhan, J. R. and
    9. Warlick, E.
    (2017). Chemical exposures and risk of acute myeloid leukemia and myelodysplastic syndromes in a population-based study. Int. J. Cancer 140, 23-33. doi:10.1002/ijc.30420
    OpenUrlCrossRef
  28. ↵
    1. Rasighaemi, P.,
    2. Basheer, F.,
    3. Liongue, C. and
    4. Ward, A. C.
    (2015). Zebrafish as a model for leukemia and other hematopoietic disorders. J. Hematol. Oncol. 8, 29. doi:10.1186/s13045-015-0126-4
    OpenUrlCrossRef
  29. ↵
    1. Rothman, N.,
    2. Smith, M. T.,
    3. Hayes, R. B.,
    4. Traver, R. D.,
    5. Hoener, B.-A.,
    6. Campleman, S.,
    7. Li, G.-L.,
    8. Dosemeci, M.,
    9. Linet, M.,
    10. Zhang, L. et al.
    (1997). Benzene poisoning, a risk factor for hematological malignancy, is associated with the NQO1 609C–>T mutation and rapid fractional excretion of chlorzoxazone. Cancer Res. 57, 2839-2842.
    OpenUrlAbstract/FREE Full Text
  30. ↵
    1. Sakamoto, H.,
    2. Dai, G.,
    3. Tsujino, K.,
    4. Hashimoto, K.,
    5. Huang, X.,
    6. Fujimoto, T.,
    7. Mucenski, M.,
    8. Frampton, J. and
    9. Ogawa, M.
    (2006). Proper levels of c-Myb are discretely defined at distinct steps of hematopoietic cell development. Blood 108, 896-903. doi:10.1182/blood-2005-09-3846
    OpenUrlAbstract/FREE Full Text
  31. ↵
    1. Schnatter, A. R.,
    2. Glass, D. C.,
    3. Tang, G.,
    4. Irons, R. D. and
    5. Rushton, L.
    (2012). Myelodysplastic syndrome and benzene exposure among petroleum workers: an international pooled analysis. J. Natl. Cancer Inst. 104, 1724-1737. doi:10.1093/jnci/djs411
    OpenUrlCrossRefPubMedWeb of Science
  32. ↵
    1. Siegert, W.,
    2. Beutler, C.,
    3. Langmach, K.,
    4. Keitel, C. and
    5. Schmidt, C. A.
    (1990). Differential expression of the oncoproteins c-myc and c-myb in human lymphoproliferative disorders. Eur. J. Cancer 26, 733-737. doi:10.1016/0277-5379(90)90130-L
    OpenUrlCrossRefPubMed
  33. ↵
    1. Singh, R. and
    2. Winn, L. M.
    (2008). The effects of 1,4-benzoquinone on c-Myb and topoisomerase II in K-562 cells. Mutat. Res. 645, 33-38. doi:10.1016/j.mrfmmm.2008.08.007
    OpenUrlCrossRefPubMed
  34. ↵
    1. Snyder, R. and
    2. Hedli, C. C.
    (1996). An overview of benzene metabolism. Environ. Health Perspect. 104, 1165-1171.
    OpenUrlCrossRefPubMedWeb of Science
  35. ↵
    1. Snyder, R. and
    2. Kocsis, J. J.
    (1975). Current concepts of chronic benzene toxicity. CRC Crit. Rev. Toxicol. 3, 265-288. doi:10.3109/10408447509079860
    OpenUrlCrossRefPubMed
  36. ↵
    1. Son, M. Y.,
    2. Deng, C.-X.,
    3. Hoeijmarkers, J. H.,
    4. Rebel, V. I. and
    5. Hasty, P.
    (2016). A mechanism for 1, 4-Benzoquinone-induced genotoxicity. Oncotarget 7, 46433. doi:10.18632/oncotarget.10184
    OpenUrlCrossRef
  37. ↵
    1. Stachura, D. L. and
    2. Traver, D.
    (2011). Cellular dissection of zebrafish hematopoiesis. Methods Cell Biol. 101, 75-110. doi:10.1016/B978-0-12-387036-0.00004-9
    OpenUrlCrossRefPubMed
  38. ↵
    1. Wan, J. and
    2. Winn, L. M.
    (2007). Benzene's metabolites alter c-MYB activity via reactive oxygen species in HD3 cells. Toxicol. Appl. Pharmacol. 222, 180-189. doi:10.1016/j.taap.2007.04.016
    OpenUrlCrossRefPubMedWeb of Science
  39. ↵
    1. Wan, J.,
    2. Badham, H. J. and
    3. Winn, L.
    (2005). The role of c-MYB in benzene-initiated toxicity. Chem-Biol. Interact. 153, 171-178. doi:10.1016/j.cbi.2005.03.037
    OpenUrlCrossRef
  40. ↵
    1. Westerfield, M.
    (2007). The Zebrafish Book. A Guide for the Laboratory Use of Zebrafish (Danio rerio), 5th edn. Eugene: Univ. of Oregon Press.
  41. ↵
    1. Westin, E. H.,
    2. Gallo, R. C.,
    3. Arya, S. K.,
    4. Eva, A.,
    5. Souza, L. M.,
    6. Baluda, M. A.,
    7. Aaronson, S. A. and
    8. Wong-Staal, F.
    (1982). Differential expression of the amv gene in human hematopoietic cells. Proc. Natl. Acad. Sci. USA 79, 2194-2198. doi:10.1073/pnas.79.7.2194
    OpenUrlAbstract/FREE Full Text
  42. ↵
    1. Willett, C. E.,
    2. Cherry, J. J. and
    3. Steiner, L. A.
    (1997). Characterization and expression of the recombination activating genes (rag1 and rag2) of zebrafish. Immunogenetics 45, 394-404. doi:10.1007/s002510050221
    OpenUrlCrossRefPubMedWeb of Science
  43. ↵
    1. Wolff, L.
    (1996). Myb-induced transformation. Crit. Rev. Oncog. 7, 245-260. doi:10.1615/CritRevOncog.v7.i3-4.60
    OpenUrlCrossRefPubMed
  44. ↵
    1. Zakrzewska, A.,
    2. Cui, C.,
    3. Stockhammer, O. W.,
    4. Benard, E. L.,
    5. Spaink, H. P. and
    6. Meijer, A. H.
    (2010). Macrophage-specific gene functions in Spi1-directed innate immunity. Blood 116, e1-e11. doi:10.1182/blood-2010-01-262873
    OpenUrlAbstract/FREE Full Text
  45. ↵
    1. Zhang, Y.,
    2. Jin, H.,
    3. Li, L.,
    4. Qin, F.-X. and
    5. Wen, Z.
    (2011). cMyb regulates hematopoietic stem/progenitor cell mobilization during zebrafish hematopoiesis. Blood 118, 4093-4101. doi:10.1182/blood-2011-03-342501
    OpenUrlAbstract/FREE Full Text
Previous ArticleNext Article
Back to top
Previous ArticleNext Article

This Issue

RSSRSS

Keywords

  • 1,4-benzoquinone
  • Hematotoxicity
  • Neutrophilia
  • c-myb
  • Zebrafish

 Download PDF

Email

Thank you for your interest in spreading the word on Disease Models & Mechanisms.

NOTE: We only request your email address so that the person you are recommending the page to knows that you wanted them to see it, and that it is not junk mail. We do not capture any email address.

Enter multiple addresses on separate lines or separate them with commas.
Establishment of a zebrafish hematological disease model induced by 1,4-benzoquinone
(Your Name) has sent you a message from Disease Models & Mechanisms
(Your Name) thought you would like to see the Disease Models & Mechanisms web site.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
RESEARCH ARTICLE
Establishment of a zebrafish hematological disease model induced by 1,4-benzoquinone
Ao Zhang, Mei Wu, Junliang Tan, Ning Yu, Mengchang Xu, Xutong Yu, Wei Liu, Yiyue Zhang
Disease Models & Mechanisms 2019 12: dmm037903 doi: 10.1242/dmm.037903 Published 28 March 2019
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
Citation Tools
RESEARCH ARTICLE
Establishment of a zebrafish hematological disease model induced by 1,4-benzoquinone
Ao Zhang, Mei Wu, Junliang Tan, Ning Yu, Mengchang Xu, Xutong Yu, Wei Liu, Yiyue Zhang
Disease Models & Mechanisms 2019 12: dmm037903 doi: 10.1242/dmm.037903 Published 28 March 2019

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Alerts

Please log in to add an alert for this article.

Sign in to email alerts with your email address

Article navigation

  • Top
  • Article
    • ABSTRACT
    • INTRODUCTION
    • RESULTS
    • DISCUSSION
    • MATERIALS AND METHODS
    • Acknowledgements
    • Footnotes
    • References
  • Figures & tables
  • Info & metrics
  • PDF

Related articles

Cited by...

More in this TOC section

  • Hyperuricemia causes kidney damage by promoting autophagy and NLRP3-mediated inflammation in rats with urate oxidase deficiency
  • The infantile myofibromatosis NOTCH3 L1519P mutation leads to hyperactivated ligand-independent Notch signaling and increased PDGFRB expression
  • Altered cytoskeletal arrangement in induced pluripotent stem cells and motor neurons from patients with riboflavin transporter deficiency
Show more RESEARCH ARTICLE

Similar articles

Subject collections

  • Zebrafish as a Disease Model

Other journals from The Company of Biologists

Development

Journal of Cell Science

Journal of Experimental Biology

Biology Open

Advertisement

DMM and COVID-19

We are aware that the COVID-19 pandemic is having an unprecedented impact on researchers worldwide. The Editors of all The Company of Biologists’ journals have been considering ways in which we can alleviate concerns that members of our community may have around publishing activities during this time. Read about the actions we are taking at this time.

Please don’t hesitate to contact the Editorial Office if you have any questions or concerns.


The twin pillars of Disease Models & Mechanisms

In her first Editorial as Editor-in-Chief, Liz Patton sets out her vision and priorities for DMM focusing on four thematic challenges: mechanisms of disease, innovative technologies, disease progression through time and therapy.


Extended deadline - The RAS Pathway: Diseases, Therapeutics and Beyond

Our upcoming special issue is welcoming submissions until 3 May 2021. Guest-edited by Donita Brady (Perelman School of Medicine at the University of Pennsylvania, USA) and Arvin Dar (Icahn School of Medicine at Mount Sinai, USA), the issue will focus on the targeting the RAS pathway.

Find out more about the issue and how to submit your manuscript.


Perspective - Modelling the developmental origins of paediatric cancer to improve patient outcomes

James Amatruda authors our first Perspective, discussing some of the key challenges in paediatric cancer from his perspective as a physician-scientist.


A muscle growth-promoting treatment based on the attenuation of activin/myostatin signalling results in long-term testicular abnormalities

In this issue’s Editor’s choice, Ketan Patel and colleagues describe how even brief exposure to muscle-growth-promoting treatments exerts a long-term detrimental effect on the testes, and test promising therapeutics to mitigate this side-effect.

Articles

  • Accepted manuscripts
  • Issue in progress
  • Latest complete issue
  • Issue archive
  • Archive by article type
  • Subject collections
  • Interviews
  • Sign up for alerts

About us

  • About DMM
  • Editors and Board
  • Editor biographies
  • Travelling Fellowships
  • Grants and funding
  • Journal Meetings
  • Workshops
  • The Company of Biologists

For Authors

  • Submit a manuscript
  • Aims and scope
  • Presubmission enquiries
  • Article types
  • Manuscript preparation
  • Cover suggestions
  • Editorial process
  • Promoting your paper
  • Open Access
  • Biology Open transfer

Journal Info

  • Journal policies
  • Rights and permissions
  • Media policies
  • Reviewer guide
  • Sign up for alerts

Contact

  • Contact DMM
  • Advertising
  • Feedback

Twitter   YouTube   LinkedIn

© 2021   The Company of Biologists Ltd   Registered Charity 277992