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Finding ATF4/p75NTR/IL-8 signal pathway in endothelial-mesenchymal transition by safrole oxide.

Ge D, Jing Q, Zhao W, Yue H, Su L, Zhang S, Zhao J - PLoS ONE (2014)

Bottom Line: Safrole oxide (SFO), 50 µg/ml, could most effectively induce EndoMT within 12 h.Furthermore, knockdown of p75NTR inhibited the SFO-increased IL-8 expression and secretion, and knockdown of ATF4 inhibited SFO-increased p75NTR level significantly.The ATF4/p75NTR/IL-8 signal pathway may have an important role in EndoMT induced by SFO.

View Article: PubMed Central - PubMed

Affiliation: Shandong Provincial Key Laboratory of Animal Cells and Developmental Biology, School of Life Science, Shandong University, Jinan, China.

ABSTRACT
Targeting the endothelial-to-mesenchymal transition (EndoMT) may be a novel therapeutic strategy for cancer and various diseases induced by fibrosis. We aimed to identify a small chemical molecule as an inducer of EndoMT and find a new signal pathway by using the inducer. Safrole oxide (SFO), 50 µg/ml, could most effectively induce EndoMT within 12 h. To understand the underlying molecular mechanism, we performed microarray, quantitative real-time PCR and western blot analysis to find key factors involved in SFO-induced EndoMT and demonstrated the involvement of the factors by RNAi. The expression of activating transcription factor 4 (ATF4), p75 neurotrophin receptor (p75NTR), and interleukin 8 (IL-8) was greatly increased in SFO-induced EndoMT. Knockdown of ATF4 inhibited the SFO-induced EndoMT completely, and knockdown of p75NTR or IL-8 partially inhibited the EndoMT, which suggests that all three factors were involved in the process. Furthermore, knockdown of p75NTR inhibited the SFO-increased IL-8 expression and secretion, and knockdown of ATF4 inhibited SFO-increased p75NTR level significantly. The ATF4/p75NTR/IL-8 signal pathway may have an important role in EndoMT induced by SFO. Our findings support potential novel targets for the therapeutics of cancer and fibrosis disease.

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The ATF4/p75NTR/IL-8 signaling pathway was involved in the induction of EndoMT by SFO.(A and B) Cells were preincubated with ATF4 siRNA for 48 h, then 50 µg/ml SFO for 3 h for qPCR assay and 6 h for western blot assay. Effect of ATF4 siRNA on SFO-induced p75NTR expression in HUVECs was detected by qPCR assay (A) and western blot assay (B). Data below gels were used to quantify the density of the bands. (C to F) Cells were preincubated with p75NTR siRNA for 48 h, then 50 µg/ml of SFO for 3 h for qPCR assay and 6 h for western blot or ELISA assay. Effect of p75NTR siRNA on SFO-induced ATF4 expression in HUVECs was detected by qPCR assay (C) and western blot assay (D). Effect of p75NTR siRNA on SFO-induced IL-8 expression and release in HUVECs was detected by qPCR assay (E) and ELISA (F). G. A schematic of the signaling pathways of SFO in HUVEC transdifferentiation. (**p<0.01; & p>0.05, n = 3).
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pone-0099378-g006: The ATF4/p75NTR/IL-8 signaling pathway was involved in the induction of EndoMT by SFO.(A and B) Cells were preincubated with ATF4 siRNA for 48 h, then 50 µg/ml SFO for 3 h for qPCR assay and 6 h for western blot assay. Effect of ATF4 siRNA on SFO-induced p75NTR expression in HUVECs was detected by qPCR assay (A) and western blot assay (B). Data below gels were used to quantify the density of the bands. (C to F) Cells were preincubated with p75NTR siRNA for 48 h, then 50 µg/ml of SFO for 3 h for qPCR assay and 6 h for western blot or ELISA assay. Effect of p75NTR siRNA on SFO-induced ATF4 expression in HUVECs was detected by qPCR assay (C) and western blot assay (D). Effect of p75NTR siRNA on SFO-induced IL-8 expression and release in HUVECs was detected by qPCR assay (E) and ELISA (F). G. A schematic of the signaling pathways of SFO in HUVEC transdifferentiation. (**p<0.01; & p>0.05, n = 3).

Mentions: Transcription factors play an important role in the control of genes. Therefore, we detected whether ATF4 could regulate the expression of p75NTR. Suppressing the role of ATF4 inhibited SFO-increased p75NTR mRNA and protein levels (Fig. 6A, B), which suggested that ATF4 is upstream of p75NTR in SFO-induced EndoMT in HUVECs.


Finding ATF4/p75NTR/IL-8 signal pathway in endothelial-mesenchymal transition by safrole oxide.

Ge D, Jing Q, Zhao W, Yue H, Su L, Zhang S, Zhao J - PLoS ONE (2014)

The ATF4/p75NTR/IL-8 signaling pathway was involved in the induction of EndoMT by SFO.(A and B) Cells were preincubated with ATF4 siRNA for 48 h, then 50 µg/ml SFO for 3 h for qPCR assay and 6 h for western blot assay. Effect of ATF4 siRNA on SFO-induced p75NTR expression in HUVECs was detected by qPCR assay (A) and western blot assay (B). Data below gels were used to quantify the density of the bands. (C to F) Cells were preincubated with p75NTR siRNA for 48 h, then 50 µg/ml of SFO for 3 h for qPCR assay and 6 h for western blot or ELISA assay. Effect of p75NTR siRNA on SFO-induced ATF4 expression in HUVECs was detected by qPCR assay (C) and western blot assay (D). Effect of p75NTR siRNA on SFO-induced IL-8 expression and release in HUVECs was detected by qPCR assay (E) and ELISA (F). G. A schematic of the signaling pathways of SFO in HUVEC transdifferentiation. (**p<0.01; & p>0.05, n = 3).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4048316&req=5

pone-0099378-g006: The ATF4/p75NTR/IL-8 signaling pathway was involved in the induction of EndoMT by SFO.(A and B) Cells were preincubated with ATF4 siRNA for 48 h, then 50 µg/ml SFO for 3 h for qPCR assay and 6 h for western blot assay. Effect of ATF4 siRNA on SFO-induced p75NTR expression in HUVECs was detected by qPCR assay (A) and western blot assay (B). Data below gels were used to quantify the density of the bands. (C to F) Cells were preincubated with p75NTR siRNA for 48 h, then 50 µg/ml of SFO for 3 h for qPCR assay and 6 h for western blot or ELISA assay. Effect of p75NTR siRNA on SFO-induced ATF4 expression in HUVECs was detected by qPCR assay (C) and western blot assay (D). Effect of p75NTR siRNA on SFO-induced IL-8 expression and release in HUVECs was detected by qPCR assay (E) and ELISA (F). G. A schematic of the signaling pathways of SFO in HUVEC transdifferentiation. (**p<0.01; & p>0.05, n = 3).
Mentions: Transcription factors play an important role in the control of genes. Therefore, we detected whether ATF4 could regulate the expression of p75NTR. Suppressing the role of ATF4 inhibited SFO-increased p75NTR mRNA and protein levels (Fig. 6A, B), which suggested that ATF4 is upstream of p75NTR in SFO-induced EndoMT in HUVECs.

Bottom Line: Safrole oxide (SFO), 50 µg/ml, could most effectively induce EndoMT within 12 h.Furthermore, knockdown of p75NTR inhibited the SFO-increased IL-8 expression and secretion, and knockdown of ATF4 inhibited SFO-increased p75NTR level significantly.The ATF4/p75NTR/IL-8 signal pathway may have an important role in EndoMT induced by SFO.

View Article: PubMed Central - PubMed

Affiliation: Shandong Provincial Key Laboratory of Animal Cells and Developmental Biology, School of Life Science, Shandong University, Jinan, China.

ABSTRACT
Targeting the endothelial-to-mesenchymal transition (EndoMT) may be a novel therapeutic strategy for cancer and various diseases induced by fibrosis. We aimed to identify a small chemical molecule as an inducer of EndoMT and find a new signal pathway by using the inducer. Safrole oxide (SFO), 50 µg/ml, could most effectively induce EndoMT within 12 h. To understand the underlying molecular mechanism, we performed microarray, quantitative real-time PCR and western blot analysis to find key factors involved in SFO-induced EndoMT and demonstrated the involvement of the factors by RNAi. The expression of activating transcription factor 4 (ATF4), p75 neurotrophin receptor (p75NTR), and interleukin 8 (IL-8) was greatly increased in SFO-induced EndoMT. Knockdown of ATF4 inhibited the SFO-induced EndoMT completely, and knockdown of p75NTR or IL-8 partially inhibited the EndoMT, which suggests that all three factors were involved in the process. Furthermore, knockdown of p75NTR inhibited the SFO-increased IL-8 expression and secretion, and knockdown of ATF4 inhibited SFO-increased p75NTR level significantly. The ATF4/p75NTR/IL-8 signal pathway may have an important role in EndoMT induced by SFO. Our findings support potential novel targets for the therapeutics of cancer and fibrosis disease.

Show MeSH
Related in: MedlinePlus