P53-dependent Signaling Research Articles

DNA damage caused by suicide gene therapy system under Tet-On regulation in breast cancer cells

To determine the effect and molecular mechanism of DNA damage caused by suicide gene therapy system HSV-TK/GCV under Tet-On regulation in human breast cancer cell line MCF-7 infected by recombinant adeno-associated virus (rAAV). We used comet assay to detect the effect of HSV-TK/GCV suicide gene regulation system on MCF-7 DNA damage, and analyzed the expression change of relative DNA damage response active genes and proteins with RT-PCR and Western blot. Compared with other control groups, the comet assay showed that MCF-7 cells with HSV-TK/GCV treatment had obvious comet tails, and the expression level of DNA damage response active genes and proteins changed obviously in the HSV-TK/GCV treatment group,such as ATM, p53 and p27,but CyclinE and CDK2 did not change. DNA damage on MCF-7 cells is resulted from HSV-TK/GCV in suicide gene therapy system through a p53-dependent signal pathway, causing cell cycle arrest and cell death.

Suppression of HPV E6 and E7 expression by BAF53 depletion in cervical cancer cells

Deregulation of the expression of human papillomavirus (HPV) oncogenes E6 and E7 plays a pivotal role in cervical carcinogenesis because the E6 and E7 proteins neutralize p53 and Rb tumor suppressor pathways, respectively. In approximately 90% of all cervical carcinomas, HPVs are found to be integrated into the host genome. Following integration, the core-enhancer element and P105 promoter that control expression of E6 and E7 adopt a chromatin structure that is different from that of episomal HPV, and this has been proposed to contribute to activation of E6 and E7 expression. However, the molecular basis underlying this chromatin structural change remains unknown. Previously, BAF53 has been shown to be essential for the integrity of higher-order chromatin structure and interchromosomal interactions. Here, we examined whether BAF53 is required for activated expression of E6 and E7 genes. We found that BAF53 knockdown led to suppression of expression of E6 and E7 genes from HPV integrants in cervical carcinoma cell lines HeLa and SiHa. Conversely, expression of transiently transfected HPV18-LCR-Luciferase was not suppressed by BAF53 knockdown. The level of the active histone marks H3K9Ac and H4K12Ac on the P105 promoter of integrated HPV 18 was decreased in BAF53 knockdown cells. BAF53 knockdown restored the p53-dependent signaling pathway in HeLa and SiHa cells. These results suggest that activated expression of the E6 and E7 genes of integrated HPV is dependent on BAF53-dependent higher-order chromatin structure or nuclear motor activity.

189 Hypomethylating agents induce apoptosis and cell cycle arrest with a concomitant activation of p53-dependent signalling pathways and Foxo3a activation

189 Hypomethylating agents induce apoptosis and cell cycle arrest with a concomitant activation of p53-dependent signalling pathways and Foxo3a activation

Cell growth inhibition and apoptosis by SDS-solubilized single-walled carbon nanotubes in normal rat kidney epithelial cells

Carbon nanotubes (CNTs), promising novel nanomaterials, have been applied to drug delivery and bio-imaging; however, their potential harmful effects on human health and environment have gained much attention recently. In the present study, we investigated cytotoxic effect of solubilized single-walled CNTs (SWCNTs), which were dispersed in water by sodium dodesyl sulfate (SDS), in normal rat kidney epithelial cells (NRK-52E). SDS-SWCNT (0.125-10 μg/mL)-treated NRK-52E cells showed decreased cell viability and enhanced cytotoxicity marker levels following 24-48 h incubation. In addition, SDS-SWCNT treatment evoked the cell growth inhibition: 8 μg/mL SDS-SWCNT induced the growth arrest at G(0)/G(1) phase and levels of cell cycle-related proteins such as CDK2, CDK6 and phosphorylated-retinoblastoma (pRB) were significantly reduced by CNT. Whereas, at higher concentration of SDS-SWCNT, the percentage of cell numbers in apoptotic sub-G(1) phase was substantially increased. Along with these changes, SDS-SWCNT treatment elevated protein levels for p53 and p21 with a concomitant increase in the single strand DNA breakage. Taken together, these results suggest that SDS-solubilized SWCNTs exert genotoxic effect in renal epithelial cells, and p53-dependent signaling can be associated with the growth arrest and apoptosis events upon CNT-induced DNA damage.

Apoptin sensitizes radiation-induced cell death via classic mitochondrial, caspase and p53-dependent signaling in HepG2 cells

Resistance or insensitivity to radiation therapy is one of the hallmarks of hepatocellular carcinoma. Sensitizing radioresistant cancer by combining radiation with other therapeutics to induce apoptosis has been widely investigated. Our previous study showed that chicken anaemia virus-derived apoptin protein induced the apoptosis of hepatic carcinoma HepG2 cells. In the present study, we demonstrated that apoptin sensitizes cells to radiation-induced apoptosis using a lentivirus-apoptin expression system in hepatic carcinoma HepG2 cells. Combination therapy with radiation and apoptin dramatically induced mitochondrial cytochrome c release and the cleavage of caspases -9, -3 and -7. Our findings are also the first to show that the combination of radiation and apoptin up-regulates p53 expression. Thus, apoptin treatment represents a potential method for enhancing the effectiveness of radiotherapy in poorly responding hepatocellular carcinoma.

Role of Adrenomedullin in Lyme Disease

Borrelia burgdorferi stimulates a strong inflammatory response during infection of a mammalian host. To understand the mechanisms of immune regulation employed by the host to control this inflammatory response, we focused our studies on adrenomedullin, a peptide produced in response to bacterial stimuli that exhibits antimicrobial activity and regulates inflammatory responses by modulating the expression of inflammatory cytokines. Specifically, we investigated the effect of B. burgdorferi on the expression of adrenomedullin as well as the ability of adrenomedullin to dampen host inflammatory responses to the spirochete. The concentration of adrenomedullin in the synovial fluid of untreated Lyme arthritis patients was elevated compared with that in control osteoarthritis patient samples. In addition, coculture with B. burgdorferi significantly increased the expression of adrenomedullin in RAW264.7 macrophages through MyD88-, phosphatidylinositol 3-kinase (PI3-K)-, and p38-dependent signaling cascades. Furthermore, the addition of exogenous adrenomedullin to B. burgdorferi-stimulated RAW264.7 macrophages resulted in a significant decrease in the induction of proinflammatory cytokines. Taken together, these results suggest that B. burgdorferi increases the production of adrenomedullin, which in turn negatively regulates the B. burgdorferi-stimulated inflammatory response.

Dcr3 inhibit p53-dependent apoptosis in γ-irradiated lung cancer cells

Purpose: To identify genes responsible for the radiosensitivity, we investigated the role of the differential gene expression profiles by comparing radioresistant H1299 with radiosensitive H460 lung cancer cell lines.Materials and methods: mRNA profiles of lung cancer cell lines were assessed using microarray, and subsequent validation was performed with qRT-PCR (Quantitative real time-polymerase chain reaction). The expression levels of differentially expressed genes were determined by Western blot and the radioresistance of lung cancer cell lines was measured by clonogenic assay.Results: From the differentially expressed apoptosis-related genes between H1299 and H460, we found Dcr3 (Decoy receptor 3, also known as TNFRSF6B; Tumour necrosis factor receptor super family member 6B) expression was significantly (P = 4.38 × 10−7) higher in H1299 cells than H460 cells. Moreover, the Dcr3 mRNA expression level in the radioresistant cell lines (H1299, A549, DLD1, MB231, MB157) was increased in comparison to the radiosensitive cell lines (ME180, Caski, U87MG, MCF7, H460). Overexpression of Dcr3 increased the survival rate of radiosensitive H460, MCF7, and U87MG cells, and knockdown of Dcr3 abolished the radioresistance of A549 cells. The survival rate of p53 (Tumour protein 53)-deficient H1299 after gamma-irradiation was not affected by the suppression of Dcr3 expression. However, when we introduced p53 into H1299 cells, siDcr3 (siRNA of Dcr3) suppressed the radioresistance of H1299 cells by inducing p53-dependent Fas (Fas receptor, also known as TNFRSF6; Tumour necrosis factor receptor super family member 6)-mediated apoptosis pathway.Conclusion: Characterisation of gene expression profiles in two lung cancer cell lines revealed that Dcr3 expression and p53-dependent apoptosis signalling pathway regulate cellular response to ionising radiation.

Differential Gene Expression Profiles of Radioresistant Non–Small-Cell Lung Cancer Cell Lines Established by Fractionated Irradiation: Tumor Protein p53-Inducible Protein 3 Confers Sensitivity to Ionizing Radiation

Despite the widespread use of radiotherapy as a local and regional modality for the treatment of cancer, some non-small-cell lung cancers commonly develop resistance to radiation. We thus sought to clarify the molecular mechanisms underlying resistance to radiation. We established the radioresistant cell line H460R from radiosensitive parental H460 cells. To identify the radioresistance-related genes, we performed microarray analysis and selected several candidate genes. Clonogenic and MTT assays showed that H460R was 10-fold more resistant to radiation than H460. Microarray analysis indicated that the expression levels of 1,463 genes were altered more than 1.5-fold in H460R compared with parental H460. To evaluate the putative functional role, we selected one interesting gene tumor protein p53-inducible protein 3 (TP53I3), because that this gene was significantly downregulated in radioresistant H460R cells and that it was predicted to link p53-dependent cell death signaling. Interestingly, messenger ribonucleic acid expression of TP53I3 differed in X-ray-irradiated H460 and H460R cells, and overexpression of TP53I3 significantly affected the cellular radiosensitivity of H460R cells. These results show that H460R may be useful in searching for candidate genes that are responsible for radioresistance and elucidating the molecular mechanism of radioresistance.

Benzo[ a ]pyrene diol epoxide stimulates an inflammatory response in normal human lung fibroblasts through a p53 and JNK mediated pathway

Cellular responses to carcinogens are typically studied in transformed cell lines, which do not reflect the physiological status of normal tissues. To address this question, we have characterized the transcriptional program and cellular responses of human lung WI-38 fibroblasts upon exposure to the ultimate carcinogen benzo[a]pyrene diol epoxide (BPDE). In contrast to observations in cell lines, we find that BPDE treatment induces a strong inflammatory response in these normal fibroblasts. Whole-genome microarrays show induction of numerous inflammatory factors, including genes that encode interleukins (ILs), growth factors and enzymes related to prostaglandin synthesis and signaling. Real-time reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay (ELISA) revealed a time- and dose-dependent-induced expression and production of cyclooxygenase 2, prostglandin E2 and IL1B, IL6 and IL8. In parallel, cell cycle progression and DNA repair processes were repressed, but DNA damage signaling was increased via p53-Ser15 phosphorylation and induced expression levels of GADD45A, CDKN1A, BTG2 and SESN1. Network analysis suggested that activator protein 1 transcription factors may link the cell cycle response and DNA damage signaling with the inflammatory stress-response in these cells. We confirmed this hypothesis by showing that p53-dependent signaling through c-jun N-terminal kinase (JNK) led to increased cJun-Ser63 phosphorylation and that inhibition of JNK-mediated cJun activation using p53- or JNK-specific inhibitors significantly reduced IL gene expression and subsequent production of IL8. This is the first demonstration that a strong inflammatory response is triggered in normal fibroblasts by BPDE and that this occurs through coordinated regulation with other cellular processes.

Human bone marrow stromal cells display variable anatomic site-dependent response and recovery from irradiation

Objectives Orofacial bone is commonly affected by osteoradionecrosis (ORN) during head and neck cancer radiotherapy possibly due to interactions of several factors including radiation damage to resident bone marrow stromal cells (BMSCs). Irradiation causes DNA damage, triggers p53-dependent signalling resulting in either cell-cycle arrest or apoptosis. In same individuals, disproportionately higher rapid growth of orofacial BMSCs relative to those of axial/appendicular bones suggests their response to radiation is skeletally site-specific. We hypothesised that survival and osteogenic recovery capacity of irradiated human BMSCs is site-dependent based on anatomic skeletal site of origin. Methods Early passage BMSCs from maxilla, mandible and iliac crest of four normal volunteers were exposed to 2.5 to 10 Gy gamma radiation to evaluate clonogenic survival, effects on cell cycle, DNA damage, p53-related response and in vivo osteogenic regenerative capacity. Results Orofacial bone marrow stromal cells (OF-MSCs) survived higher radiation doses and recovered quicker than iliac crest (IC-MSCs) based on clonogenic survival, proliferation and accumulation in G 0G 1 phase. Post-irradiation p53 level was relatively unchanged but expression of p21, a downstream effector was moderately increased in OF-MSCs. Re-establishment of in vivo bone regeneration was delayed more in irradiated IC-MSCs relative to OF-MSCs. Conclusions Effect of irradiation on human BMSCs was skeletal site-specific with OF-MSCs displaying higher radio-resistance and quicker recovery than IC-MSCs.

Suppression of B-cell lymphomagenesis by the BH3-only proteins Bmf and Bad

AbstractOncogenic c-Myc is known to balance excessive proliferation by apoptosis that can be triggered by p53-dependent and p53-independent signaling networks. Here, we provide evidence that the BH3-only proapoptotic Bcl-2 family members Bcl-2 modifying factor (Bmf) and Bcl-2 antagonist of cell death (Bad) are potent antagonists of c-Myc–driven B-cell lymphomagenesis. Tumor formation was preceded by the accumulation of preneoplastic pre-B and immature immunoglobulin M–positive (IgM+) B cells in hematopoietic organs of Eμ-myc/bmf−/− mice, whereas Eμ-myc/bad−/− mice showed an increase of pre-B cells limited to the spleen. Although the loss of Bad had no impact on the tumor immunophenotype, Bmf deficiency favored the development of IgM+ B cell over pre-B cell tumors. This phenomenon was caused by a strong protection of immature IgM+ B cells from oncogene-driven apoptosis caused by loss of bmf and c-Myc–induced repression of Bmf expression in premalignant pre-B cells. Steady-state levels of B-cell apoptosis also were reduced in the absence of Bad, in support of its role as a sentinel for trophic factor-deprivation. Loss of Bmf reduced the pressure to inactivate p53, whereas Bad deficiency did not, identifying Bmf as a novel component of the p53-independent tumor suppressor pathway triggered by c-Myc.

Edaravone, a known free radical scavenger, enhances X-ray-induced apoptosis at low concentrations

Edaravone has been reported to have a radioprotective effect at high concentrations. We now report that a lower dose of edaravone enhanced X-ray-induced apoptosis of some cell lines harboring p53 wild-type status, such as MOLT-4, Nalm-6, and HepG2. The knock-down of p53 using siRNA in MOLT-4 cells abolished the radiosensitizing effect of edaravone. Enhanced phosphorylations of p53 at Ser 15 and Ser 20 and up-regulation of PUMA, a p53 target protein, were observed after X-irradiation in the presence of edaravone. We conclude that the low dose of edaravone sensitized cells to X-irradiation by promoting the p53-dependent apoptotic signaling pathway.

Interaction of hepatitis C virus (HCV) with P53 family-induced apoptosis signaling pathways

Introduction: Chronic hepatitis C virus (HCV) infection constitutes a major risk factor for liver cirrhosis and hepatocellular carcinoma (HCC). Here we investigate the interaction of HCV with transcriptionally active (TA) and inactive (DeltaN) isoforms of the p53 family which play a central role in tumor development, treatment response and prognosis of HCC. Methods and results: Combined adenoviral transfer of wild-type p53 (wt p53) or TAp73 with HCV expression constructs led to a synergistic transactivation of the CD95 and the Bax gene (luciferase reporter assays) and to an increase in p53- and TAp73-mediated apoptosis in HepG2 cells (wt p53) and Hep3B cells (p53 -/-) (FACS analysis). For a biologically more relevant approach we used the HCV infectious tissue culture system replicating the Jc1 construct. HUH6 (wt p53) and HUH7-Lunet cells (mutant p53) were infected with the Jc1 construct following adenoviral transfer of wt p53, TAp73 or DeltaNp73. In addition, we performed silencing of the p53 family members by transfection of siRNA. HCV titers were measured using immunohistochemistry. Overexpression of TAp73 led to a decline in HCV infectivity whereas silencing of TAp73 enhanced HCV infectivity. Of note, upregulation of DeltaNp73 led to an increase of HCV titers. Conclusions: HCV, wt p53 and TAp73 cooperate in the activation of ‘classic’ p53-dependent apoptosis genes and p53 family-dependent apoptosis signaling pathways. Of clinical importance, TAp73 influences HCV infectivity. Thus, the interaction of HCV with anti-apoptotic (DeltaN) isoforms of the p53 family may be an underlying mechanism how the virus evades the cellular apoptotic defence and establishes a chronic infection. This may constitute a new mechanism by which HCV and oncogenic DeltaN isoforms of the p53 family could enhance the carcinogenic process in liver cells.

Nitrosative Stress as a Mediator of Apoptosis: Implications for Cancer Therapy

Nitric oxide (NO) is now recognised as one of the most important molecules influencing the development, progression and treatment of cancer. A key component of its action is as a negative and positive regulator of apoptosis. Broadly, constitutive levels of NO(nM), are capable of inhibiting numerous signalling pathways in both normal and cancer cells. These include soluble guanylate cyclase, leading to reduced Ca++ signalling, inhibition of caspases and scavenging of reactive oxygen species, all of which promote survival signalling. High concentrations (M-mM) on the other hand, generally promote apoptosis. Pathways involving cGMP, cytochrome c release, mitogen activated kinases, ceramide and poly(ADP)ribose polymerase have all been implicated. The role of p53 in NO-induced cell death has been widely studied. In many cell types p53-dependent signalling is involved, while in others, apoptosis occurs in the absence of functional p53. There is also evidence that the tumour microenvironment, where low oxygen and glucose levels prevail, enhances cell death signalling by NO and peroxynitrite, thus tumours may be more sensitive to high levels of NO than their normal tissue counterpart. The cytotoxicity of NO has been studied directly in many tumour models, both in vitro and in vivo. In all cases, high concentrations of NO, generated by donor drugs or by iNOS gene transfer caused extensive tumour cell death, which was enhanced by the ability of NO to diffuse readily from its source of generation to most cells within tumours. NO was also a very effective enhancer of conventional chemo- and radiotherapy. Thus, NO therapy has great potential to improve the treatment of cancer.

P53Gene Expression and 2-Methoxyestradiol Treatment Differentially Induce Nuclear Factor Kappa B Activation in Human Lung Cancer Cells with Different p53 Phenotypes

The p53 tumor suppressor gene is frequently mutated in multiple human cancers, leading to loss of wild-type p53 (wt-p53)-dependent functions and tumorigenesis. p53 gene therapy is used to induce apoptosis in human cancer cells and tumors. Activation of nuclear factor kappa B (NF-kappaB) causes resistance to both chemotherapy and apoptosis in tumor cells. We show that expression of wt-p53 from a recombinant adenovirus-p53 followed by treatment with 2-methoxyestradiol (2-ME), an endogenous, nontoxic, estrogenic metabolite, resulted in differential NF-kappaB activation and inhibitor kappaB alpha (IkappaB-alpha) degradation in three different human lung cancer cell lines with different p53 phenotypes. The H322J cells, with mutant (Arg248Gln) p53, showed NF-kappaB activation and IkappaB-alpha degradation after adeno-p53 expression + 2-ME treatment; however, these conditions separately did not activate NF-kappaB, rather caused accumulation of IkappaB-alpha. In contrast, either adeno-p53 expression or 2-ME treatment induced NF-kappaB activation in the p53-deleted H1299 cells, but H460 cells, containing wt-p53, did not show NF-kappaB activation under any of these conditions. This shows p53-dependent differential signaling to NF-kappaB by 2-ME. Since NF-kappaB activation inhibits apoptosis and causes resistance to chemotherapy, our study suggests the need to distinguish p53 phenotypes of tumors for p53 gene and 2-ME therapy.

β-Ionone-induced apoptosis in human osteosarcoma (U2os) cells occurs via a p53-dependent signaling pathway

beta-Ionone is a constituent of vegetables and fruits, and can induce apoptosis in some types of malignant cells. However, the mechanism of apoptosis in osteosarcoma (U2os) cells is currently unclear. In this study, we determined whether beta-ionone can induce apoptosis in U2os cells in vitro and which signal pathway(s) is involved. We found that beta-ionone inhibited cell proliferation in U2os cells in a concentration- and time-dependent manner and caused cell cycle arrest at the G1-S phase. TUNEL assay, DNA ladder and assessment of Caspase 3 activity showed that apoptosis was the determinant in the effects of beta-ionone. Furthermore, Expression of the p53 protein increased in a concentration-dependent and time-dependent manner according to immunocytochemistry and immunoblotting after beta-ionone treatment. In addition, beta-ionone upregulated Bax protein and downregulated Bcl2 protein which led to Bax translocation and cytochrome c release, subsequently activated Caspase 3, thus resulting in apoptosis. In summary, these data suggested that beta-ionone induced apoptosis in a concentration-dependent manner in U2os cells via a p53-dependent mitochondrial pathway.

Redox-Dependent Translocation of p53 to Mitochondria or Nucleus in Human Melanocytes after UVA- and UVB-Induced Apoptosis

The p53 protein is an important transcription factor and tumor suppressor that is induced in response to many forms of cellular stress. UVA irradiation of human melanocytes caused generation of reactive oxygen species, which altered the intracellular redox balance and was accompanied by translocation of p53 to mitochondria. In contrast, UVB did not affect the redox status and p53 was translocated to the nucleus. Although different intracellular location of p53, UVA/B induced apoptosis through the intrinsic pathway detected as translocation of Bax to mitochondria, release of cytochrome c, and activation of caspases. These events were all prevented by inhibition of p53 with pifithrin-alpha. Furthermore, inhibition of p53 prevented lysosomal membrane permeabilization, detected as translocation of cathepsins to the cytosol, after UVB exposure, whereas UVA-induced lysosomal release was unaffected by inhibition of p53. In control cells, p53 coimmunoprecipitated with the antiapoptotic proteins Bcl-2 and Bcl-x(L) and upon UVA exposure the interaction was replaced by binding to the proapoptotic proteins Bax, Noxa, and Puma. Our findings suggest that UVA-induced apoptosis is caused by extensive oxidative damage leading to p53-regulated mitochondrial release, whereas UVB induces DNA damage and apoptosis signaling upstream of lysosomal membrane permeabilization.

P38MAPK Controls Expression of Multiple Cell Cycle Inhibitors and Islet Proliferation with Advancing Age

Aging is a complex organismal process that is controlled by genetic, environmental, and behavioral factors. Accumulating evidence supports a role for different cell cycle inhibitors in mammalian aging. Little is known, however, about the upstream signals that induce their expression. Here, we explore the role of p38MAPK by generating a dominant-negative allele (p38(AF)) in which activating phosphorylation sites Thr180 and Tyr182 are mutated. Heterozygous p38(AF) mice show a marked attenuation of p38-dependent signaling and age-induced expression of multiple cell cycle inhibitors in different organs, including pancreatic islets. As a result, aged p38(AF/+) mice show enhanced proliferation and regeneration of islets when compared to wild-type littermates. We further find an age-related reduction in expression of the p38-specific phosphatase Wip1. Wip1-deficient mice demonstrate decreased islet proliferation, while Wip1 overexpression rescues aging-related decline in proliferation and regenerative capacity. We propose that modulation of p38MAPK activity may provide new avenues for treating certain age-related degenerative diseases.

Activation of several concurrent proapoptic pathways by sulforaphane in human colon cancer cells SW620

Despite the reported cytotoxicity and apoptosis-inducing properties of sulforaphane (SF) in colon cancer cells, the details concerning individual mechanisms and signaling cascades underlying SF-mediated apoptosis remain unclear. To understand different aspects of SF-induced proapoptic signaling in advanced colon carcinoma, we investigated its mechanisms in metastatic SW620 cell line. Our results indicate that in SW620 cells SF acts to induce multivariate cascades including DNA-damage response pathway whose proapoptotic signaling is nevertheless reduced owing to the mutant status of p53 and caspase-2-JNK pathway which seems to complement and enhance p53-dependent signaling, however only in wild-type p53. Furthermore, both pathways require the active role of mitochondria and do not depend on generation of ROS, making SF an attractive chemopreventive agent whose antitumor properties should be further investigated in colon cancer.

Trans-2-phenylcyclopropylamine induces nerve cells apoptosis in zebrafish mediated by depression of LSD1 activity

Trans-2-phenylcyclopropylamine (referred to as PCPA hereafter, also known as tranylcypromine and Parnate) is used clinically as an antidepressant. Here, we use a new model—zebrafish ( Danio rerio) to study the molecular mechanisms of its adverse reactions in vivo. Following a PCPA exposure (75 μM), embryos showed “sluggish” action (slow swim and slow escape action). Whole mount in situ hybridization showed that sox1a and huc expressions were downregulated in PCPA-treated embryos, which indicated a decrease in the number of nerve cells. TUNEL assay diplayed that the drop of nerve cells number due to excessive apoptosis. Moreover, lysine-specific demethylase 1 morpholino injection (LSD1 MO) also induced increased cellular apoptosis in embryos just as PCPA. RT-PCR at 24 hpf evaluated that the absence of LSD1 resulted in increased expression of two p53 target genes (p21 and bax2). These findings demonstrate for the first time that PCPA-induced apoptosis through inhibition of LSD1 demethylase activity and p53-dependent signaling pathway might be required for the maintenance of nerve cell apoptosis.