P300 Acetyltransferase Mediates Stiffness-Induced Activation of Hepatic Stellate Cells Into Tumor-Promoting Myofibroblasts

Changwei Dou, Zhikui Liu, Kangsheng Tu, Hongbin Zhang, Chen Chen, Usman Yaqoob, Yuanguo Wang, Jialing Wen, Jan Van Deursen, Delphine Sicard, Daniel J Tschumperlin, Hongzhi Zou, Wei Chien Huang, Raul Urrutia, Vijay Shah, Ningling Kang

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

Background & Aims: Hepatic stellate cells (HSCs) contribute to desmoplasia and stiffness of liver metastases by differentiating into matrix-producing myofibroblasts. We investigated whether stiffness due to the presence of tumors increases activation of HSCs into myofibroblasts and their tumor-promoting effects, as well as the role of E1A binding protein p300, a histone acetyltransferase that regulates transcription, in these processes. Methods: HSCs were isolated from liver tissues of patients, mice in which the p300 gene was flanked by 2 loxP sites (p300F/F mice), and p300+/+ mice (controls). The HSCs were placed on polyacrylamide gels with precisely defined stiffness, and their activation (differentiation into myofibroblasts) was assessed by immunofluorescence and immunoblot analyses for alpha-smooth muscle actin. In HSCs from mice, the p300 gene was disrupted by cre recombinase. In human HSCs, levels of p300 were knocked down with small hairpin RNAs or a mutant form of p300 that is not phosphorylated by AKT (p300S1834A) was overexpressed. Human HSCs were also cultured with inhibitors of p300 (C646), PI3K signaling to AKT (LY294002), or RHOA (C3 transferase) and effects on stiffness-induced activation were measured. RNA sequencing and chromatin immunoprecipitation–quantitative polymerase chain reaction were used to identify HSC genes that changed expression levels in response to stiffness. We measured effects of HSC-conditioned media on proliferation of HT29 colon cancer cells and growth of tumors following subcutaneous injection of these cells into mice. MC38 colon cancer cells were injected into portal veins of p300F/Fcre and control mice, and liver metastases were measured. p300F/Fcre and control mice were given intraperitoneal injections of CCl4 to induce liver fibrosis. Liver tissues were collected and analyzed by immunofluorescence, immunoblot, and histology. Results: Substrate stiffness was sufficient to activate HSCs, leading to nuclear accumulation of p300. Disrupting p300 level or activity blocked stiffness-induced activation of HSCs. In HSCs, substrate stiffness activated AKT signaling via RHOA to induce phosphorylation of p300 at serine 1834; this caused p300 to translocate to the nucleus, where it up-regulated transcription of genes that increase activation of HSCs and metastasis, including CXCL12. MC38 cells, injected into portal veins, formed fewer metastases in livers of p300F/Fcre mice than control mice. Expression of p300 was increased in livers of mice following injection of CCl4; HSC activation and collagen deposition were reduced in livers of p300F/Fcre mice compared with control mice. Conclusions: In studies of mice, we found liver stiffness to activate HSC differentiation into myofibroblasts, which required nuclear accumulation of p300. p300 increases HSC expression of genes that promote metastasis.

Original languageEnglish (US)
Pages (from-to)2209-2221.e14
JournalGastroenterology
Volume154
Issue number8
DOIs
StatePublished - Jun 1 2018

Fingerprint

Hepatic Stellate Cells
Myofibroblasts
Neoplasms
Liver
Neoplasm Metastasis
p300-CBP-associated factor
Portal Vein
Colonic Neoplasms
Fluorescent Antibody Technique
RNA Sequence Analysis
Histone Acetyltransferases
Gene Expression
2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
Subcutaneous Injections
Conditioned Culture Medium
Transferases
Intraperitoneal Injections

Keywords

  • Chromatin Remodeling
  • Epigenetic Modification
  • Mechanotransduction
  • Tumor Progression

ASJC Scopus subject areas

  • Gastroenterology

Cite this

P300 Acetyltransferase Mediates Stiffness-Induced Activation of Hepatic Stellate Cells Into Tumor-Promoting Myofibroblasts. / Dou, Changwei; Liu, Zhikui; Tu, Kangsheng; Zhang, Hongbin; Chen, Chen; Yaqoob, Usman; Wang, Yuanguo; Wen, Jialing; Van Deursen, Jan; Sicard, Delphine; Tschumperlin, Daniel J; Zou, Hongzhi; Huang, Wei Chien; Urrutia, Raul; Shah, Vijay; Kang, Ningling.

In: Gastroenterology, Vol. 154, No. 8, 01.06.2018, p. 2209-2221.e14.

Research output: Contribution to journalArticle

Dou, C, Liu, Z, Tu, K, Zhang, H, Chen, C, Yaqoob, U, Wang, Y, Wen, J, Van Deursen, J, Sicard, D, Tschumperlin, DJ, Zou, H, Huang, WC, Urrutia, R, Shah, V & Kang, N 2018, 'P300 Acetyltransferase Mediates Stiffness-Induced Activation of Hepatic Stellate Cells Into Tumor-Promoting Myofibroblasts', Gastroenterology, vol. 154, no. 8, pp. 2209-2221.e14. https://doi.org/10.1053/j.gastro.2018.02.015
Dou, Changwei ; Liu, Zhikui ; Tu, Kangsheng ; Zhang, Hongbin ; Chen, Chen ; Yaqoob, Usman ; Wang, Yuanguo ; Wen, Jialing ; Van Deursen, Jan ; Sicard, Delphine ; Tschumperlin, Daniel J ; Zou, Hongzhi ; Huang, Wei Chien ; Urrutia, Raul ; Shah, Vijay ; Kang, Ningling. / P300 Acetyltransferase Mediates Stiffness-Induced Activation of Hepatic Stellate Cells Into Tumor-Promoting Myofibroblasts. In: Gastroenterology. 2018 ; Vol. 154, No. 8. pp. 2209-2221.e14.
@article{41b134ecda0c4aef8983205ac47809a4,
title = "P300 Acetyltransferase Mediates Stiffness-Induced Activation of Hepatic Stellate Cells Into Tumor-Promoting Myofibroblasts",
abstract = "Background & Aims: Hepatic stellate cells (HSCs) contribute to desmoplasia and stiffness of liver metastases by differentiating into matrix-producing myofibroblasts. We investigated whether stiffness due to the presence of tumors increases activation of HSCs into myofibroblasts and their tumor-promoting effects, as well as the role of E1A binding protein p300, a histone acetyltransferase that regulates transcription, in these processes. Methods: HSCs were isolated from liver tissues of patients, mice in which the p300 gene was flanked by 2 loxP sites (p300F/F mice), and p300+/+ mice (controls). The HSCs were placed on polyacrylamide gels with precisely defined stiffness, and their activation (differentiation into myofibroblasts) was assessed by immunofluorescence and immunoblot analyses for alpha-smooth muscle actin. In HSCs from mice, the p300 gene was disrupted by cre recombinase. In human HSCs, levels of p300 were knocked down with small hairpin RNAs or a mutant form of p300 that is not phosphorylated by AKT (p300S1834A) was overexpressed. Human HSCs were also cultured with inhibitors of p300 (C646), PI3K signaling to AKT (LY294002), or RHOA (C3 transferase) and effects on stiffness-induced activation were measured. RNA sequencing and chromatin immunoprecipitation–quantitative polymerase chain reaction were used to identify HSC genes that changed expression levels in response to stiffness. We measured effects of HSC-conditioned media on proliferation of HT29 colon cancer cells and growth of tumors following subcutaneous injection of these cells into mice. MC38 colon cancer cells were injected into portal veins of p300F/Fcre and control mice, and liver metastases were measured. p300F/Fcre and control mice were given intraperitoneal injections of CCl4 to induce liver fibrosis. Liver tissues were collected and analyzed by immunofluorescence, immunoblot, and histology. Results: Substrate stiffness was sufficient to activate HSCs, leading to nuclear accumulation of p300. Disrupting p300 level or activity blocked stiffness-induced activation of HSCs. In HSCs, substrate stiffness activated AKT signaling via RHOA to induce phosphorylation of p300 at serine 1834; this caused p300 to translocate to the nucleus, where it up-regulated transcription of genes that increase activation of HSCs and metastasis, including CXCL12. MC38 cells, injected into portal veins, formed fewer metastases in livers of p300F/Fcre mice than control mice. Expression of p300 was increased in livers of mice following injection of CCl4; HSC activation and collagen deposition were reduced in livers of p300F/Fcre mice compared with control mice. Conclusions: In studies of mice, we found liver stiffness to activate HSC differentiation into myofibroblasts, which required nuclear accumulation of p300. p300 increases HSC expression of genes that promote metastasis.",
keywords = "Chromatin Remodeling, Epigenetic Modification, Mechanotransduction, Tumor Progression",
author = "Changwei Dou and Zhikui Liu and Kangsheng Tu and Hongbin Zhang and Chen Chen and Usman Yaqoob and Yuanguo Wang and Jialing Wen and {Van Deursen}, Jan and Delphine Sicard and Tschumperlin, {Daniel J} and Hongzhi Zou and Huang, {Wei Chien} and Raul Urrutia and Vijay Shah and Ningling Kang",
year = "2018",
month = "6",
day = "1",
doi = "10.1053/j.gastro.2018.02.015",
language = "English (US)",
volume = "154",
pages = "2209--2221.e14",
journal = "Gastroenterology",
issn = "0016-5085",
publisher = "W.B. Saunders Ltd",
number = "8",

}

TY - JOUR

T1 - P300 Acetyltransferase Mediates Stiffness-Induced Activation of Hepatic Stellate Cells Into Tumor-Promoting Myofibroblasts

AU - Dou, Changwei

AU - Liu, Zhikui

AU - Tu, Kangsheng

AU - Zhang, Hongbin

AU - Chen, Chen

AU - Yaqoob, Usman

AU - Wang, Yuanguo

AU - Wen, Jialing

AU - Van Deursen, Jan

AU - Sicard, Delphine

AU - Tschumperlin, Daniel J

AU - Zou, Hongzhi

AU - Huang, Wei Chien

AU - Urrutia, Raul

AU - Shah, Vijay

AU - Kang, Ningling

PY - 2018/6/1

Y1 - 2018/6/1

N2 - Background & Aims: Hepatic stellate cells (HSCs) contribute to desmoplasia and stiffness of liver metastases by differentiating into matrix-producing myofibroblasts. We investigated whether stiffness due to the presence of tumors increases activation of HSCs into myofibroblasts and their tumor-promoting effects, as well as the role of E1A binding protein p300, a histone acetyltransferase that regulates transcription, in these processes. Methods: HSCs were isolated from liver tissues of patients, mice in which the p300 gene was flanked by 2 loxP sites (p300F/F mice), and p300+/+ mice (controls). The HSCs were placed on polyacrylamide gels with precisely defined stiffness, and their activation (differentiation into myofibroblasts) was assessed by immunofluorescence and immunoblot analyses for alpha-smooth muscle actin. In HSCs from mice, the p300 gene was disrupted by cre recombinase. In human HSCs, levels of p300 were knocked down with small hairpin RNAs or a mutant form of p300 that is not phosphorylated by AKT (p300S1834A) was overexpressed. Human HSCs were also cultured with inhibitors of p300 (C646), PI3K signaling to AKT (LY294002), or RHOA (C3 transferase) and effects on stiffness-induced activation were measured. RNA sequencing and chromatin immunoprecipitation–quantitative polymerase chain reaction were used to identify HSC genes that changed expression levels in response to stiffness. We measured effects of HSC-conditioned media on proliferation of HT29 colon cancer cells and growth of tumors following subcutaneous injection of these cells into mice. MC38 colon cancer cells were injected into portal veins of p300F/Fcre and control mice, and liver metastases were measured. p300F/Fcre and control mice were given intraperitoneal injections of CCl4 to induce liver fibrosis. Liver tissues were collected and analyzed by immunofluorescence, immunoblot, and histology. Results: Substrate stiffness was sufficient to activate HSCs, leading to nuclear accumulation of p300. Disrupting p300 level or activity blocked stiffness-induced activation of HSCs. In HSCs, substrate stiffness activated AKT signaling via RHOA to induce phosphorylation of p300 at serine 1834; this caused p300 to translocate to the nucleus, where it up-regulated transcription of genes that increase activation of HSCs and metastasis, including CXCL12. MC38 cells, injected into portal veins, formed fewer metastases in livers of p300F/Fcre mice than control mice. Expression of p300 was increased in livers of mice following injection of CCl4; HSC activation and collagen deposition were reduced in livers of p300F/Fcre mice compared with control mice. Conclusions: In studies of mice, we found liver stiffness to activate HSC differentiation into myofibroblasts, which required nuclear accumulation of p300. p300 increases HSC expression of genes that promote metastasis.

AB - Background & Aims: Hepatic stellate cells (HSCs) contribute to desmoplasia and stiffness of liver metastases by differentiating into matrix-producing myofibroblasts. We investigated whether stiffness due to the presence of tumors increases activation of HSCs into myofibroblasts and their tumor-promoting effects, as well as the role of E1A binding protein p300, a histone acetyltransferase that regulates transcription, in these processes. Methods: HSCs were isolated from liver tissues of patients, mice in which the p300 gene was flanked by 2 loxP sites (p300F/F mice), and p300+/+ mice (controls). The HSCs were placed on polyacrylamide gels with precisely defined stiffness, and their activation (differentiation into myofibroblasts) was assessed by immunofluorescence and immunoblot analyses for alpha-smooth muscle actin. In HSCs from mice, the p300 gene was disrupted by cre recombinase. In human HSCs, levels of p300 were knocked down with small hairpin RNAs or a mutant form of p300 that is not phosphorylated by AKT (p300S1834A) was overexpressed. Human HSCs were also cultured with inhibitors of p300 (C646), PI3K signaling to AKT (LY294002), or RHOA (C3 transferase) and effects on stiffness-induced activation were measured. RNA sequencing and chromatin immunoprecipitation–quantitative polymerase chain reaction were used to identify HSC genes that changed expression levels in response to stiffness. We measured effects of HSC-conditioned media on proliferation of HT29 colon cancer cells and growth of tumors following subcutaneous injection of these cells into mice. MC38 colon cancer cells were injected into portal veins of p300F/Fcre and control mice, and liver metastases were measured. p300F/Fcre and control mice were given intraperitoneal injections of CCl4 to induce liver fibrosis. Liver tissues were collected and analyzed by immunofluorescence, immunoblot, and histology. Results: Substrate stiffness was sufficient to activate HSCs, leading to nuclear accumulation of p300. Disrupting p300 level or activity blocked stiffness-induced activation of HSCs. In HSCs, substrate stiffness activated AKT signaling via RHOA to induce phosphorylation of p300 at serine 1834; this caused p300 to translocate to the nucleus, where it up-regulated transcription of genes that increase activation of HSCs and metastasis, including CXCL12. MC38 cells, injected into portal veins, formed fewer metastases in livers of p300F/Fcre mice than control mice. Expression of p300 was increased in livers of mice following injection of CCl4; HSC activation and collagen deposition were reduced in livers of p300F/Fcre mice compared with control mice. Conclusions: In studies of mice, we found liver stiffness to activate HSC differentiation into myofibroblasts, which required nuclear accumulation of p300. p300 increases HSC expression of genes that promote metastasis.

KW - Chromatin Remodeling

KW - Epigenetic Modification

KW - Mechanotransduction

KW - Tumor Progression

UR - http://www.scopus.com/inward/record.url?scp=85047768188&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85047768188&partnerID=8YFLogxK

U2 - 10.1053/j.gastro.2018.02.015

DO - 10.1053/j.gastro.2018.02.015

M3 - Article

C2 - 29454793

AN - SCOPUS:85047768188

VL - 154

SP - 2209-2221.e14

JO - Gastroenterology

JF - Gastroenterology

SN - 0016-5085

IS - 8

ER -