Human eosinophil cationic protein enhances stress fiber formation in Balb/c 3T3 fibroblasts and differentiation of rat neonatal cardiomyocytes

Fukuda, Takayuki; Iwata, Miki; Kitazoe, Midori; Maeda, Takashi; Salomon, David; Hirohata, Satoshi; Tanizawa, Katsuyuki; Kuroda, Shinya; Seno, Masaharu

doi:10.1080/08977190902987149

Cited by 6 publications

(2 citation statements)

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“…The GFP signals in HUVEC were observed by a fluorescence microscope (model BZ‐9000; Keyence) as previously reported (Sou et al, 2008; Fukuda et al, 2009; Yonezawa et al, 2010). For in vitro analysis, GFP‐positive cells per each hypoxia‐stimulated cell ( n =1000) were counted using software Dynamic cell count BZ‐HIC (Keyence).…”

Section: Methodsmentioning

confidence: 99%

AHR, a novel acute hypoxia-response sequence, drives reporter gene expression under hypoxiain vitroandin vivo

et al. 2010

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ADAMTS1 (a disintegrin and metalloproteinase with thrombospondin motifs 1) is an early immediate gene. We have previously reported that ADAMTS1 was strongly induced by hypoxia. In this study, we investigated whether ADAMTS1 promoter-driven reporter signal is detectable by acute hypoxia. We constructed the GFP (green fluorescent protein) expression vector [AHR (acute hypoxia-response sequence)-GFP] under the control of ADAMTS1 promoter and compared it with the constitutive GFP-expressing vector under the control of CMV (cytomegalovirus promoter-GFP). We transduced AHR-GFP and examined whether GFP signals can be detected under the acute hypoxia. When the human umbilical vein [HUVEC (human umbilical vein endothelial cells)] was transduced under normoxia, there were few GFP signals, while CMV-GFP showed considerable GFP signals. When HUVEC was stimulated with hypoxia, GFP signals from AHR-GFP gene were induced under hypoxic conditions. Notably, the GFP signals peaked at 3 h under hypoxia. In ischaemic hind limb model, transduced AHR-GFP showed hypoxic induction of GFP signals. In summary, we have demonstrated that the AHR system induced the reporter gene expression by acute hypoxia, and its induction is transient. This is the first report showing the unique acute hypoxia-activated gene expression system.

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Section: Methodsmentioning

confidence: 99%

AHR, a novel acute hypoxia-response sequence, drives reporter gene expression under hypoxiain vitroandin vivo

et al. 2010

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“…ECP was reported to show neurotoxicity 2 and apoptotic effects on several eukaryotic cell lines, 3 while its did not induce cell death but inhibited the growth of cancer-derived cells. 4 Furthermore, human ECP enhanced differentiation of rat neonatal cardiomyocytes and stress fiber formation in Balb/c 3T3 mouse fibroblasts, 5 while the effects of human ECP on human fibroblasts are unknown. Despite their usefulness in determination of eosinophil physiology in vivo, murine models differ from humans in several major respects.…”

Section: Introductionmentioning

confidence: 99%

Cytokine expression in human dermal fibroblasts stimulated with eosinophil cationic protein measured by protein array

Sato

Soga²,

Yamaguchi

et al. 2013

Asian Pac J Allergy Immunol

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Summa ryBackground: Eosinophil cationic protein (ECP) was reported previously to be involved in allergic inflammation with cytotoxic activity. On the other hand, recent studies showed that ECP did not induce cell death but inhibited the growth of cancer-derived cells. Our previous study indicated that human ECP enhanced differentiation of rat neonatal cardiomyocytes and stress fiber formation in Balb/c 3T3 mouse fibroblasts, while the effects of human ECP on human fibroblasts are unknown.Objective: The present study was performed to determine the effects of human ECP on cytokine expression in human fibroblasts by protein array. Methods:The effects of recombinant human ECP (rhECP) on normal human dermal fibroblasts (NHDF) were examined by assaying cell growth. Furthermore, cytokine expression of NHDF stimulated by ECP, which could influence cell growth, was evaluated by protein array.Results: ECP was not cytotoxic but enhanced the growth of NHDF. The peak rhECP concentration that enhanced the cell counts by 1.56-fold was 100 ng/mL, which was significantly different from cultures without ECP stimulation (ANOVA/ Scheffe's test, P < 0.05). Array analyses indicated that ciliary neurotrophic factor (CNTF), neutrophilactivating peptide (NAP)-2, and neurotrophin (NT)-3 were significantly upregulated in NHDF stimulated with 100 ng/mL ECP compared to those without stimulation. Conclusion

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Eosinophil cationic protein enhances stabilization of β-catenin during cardiomyocyte differentiation in P19CL6 embryonal carcinoma cells

et al. 2012

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Prior to gastrulation, the Wnt signaling pathway through stabilized β-catenin enhances the differentiation of mouse ES cell into cardiomyocytes. We have recently shown that cardiomyocyte differentiation is enhanced by eosinophil cationic protein (ECP) through accelerated expression of marker genes of early cardiac differentiation. Furthermore, ECP enhanced the expression of Wnt3a in P19CL6 cells which were stimulated to differentiate into cardiomyocytes by DMSO. Following these findings, we evaluated in this study the potential of ECP to activate the Wnt/β-catenin signaling pathway during cardiomyocyte differentiation. Analysis by real time qPCR revealed that ECP increased the expression of Frizzled genes such as Frizzled-1, -2, -4 and -10 in P19CL6 cells in the presence of DMSO. The increased expression of those Wnt receptors was found to inhibit the phosphorylation of β-catenin resulting in the stabilization and translocation of β-catenin into the nucleus of P19CL6 cells during the early stages of cardiomyocyte differentiation. When assessed for β-catenin/TCF transcriptional activity with a TCF-luciferase (TOP/FOP) assay, ECP enhanced luciferase activity in P19CL6 cells during 48 h after transfection with TOP/FOP flash reporter in a stoichiometric manner. Collectively, this suggests that ECP can activate a canonical Wnt/β-catenin signaling pathway by enhancing the stabilization of β-catenin during cardiomyocyte differentiation.

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Human eosinophil cationic protein enhances stress fiber formation in Balb/c 3T3 fibroblasts and differentiation of rat neonatal cardiomyocytes

Cited by 6 publications

References 40 publications

AHR, a novel acute hypoxia-response sequence, drives reporter gene expression under hypoxiain vitroandin vivo

AHR, a novel acute hypoxia-response sequence, drives reporter gene expression under hypoxiain vitroandin vivo

Cytokine expression in human dermal fibroblasts stimulated with eosinophil cationic protein measured by protein array

Eosinophil cationic protein enhances stabilization of β-catenin during cardiomyocyte differentiation in P19CL6 embryonal carcinoma cells

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