Wan-Jin Jeon1, Kyeong-Min Kim1, Eun-Jung Kim2, Won-Gu Jang3. 1. Department of Biotechnology, College of Engineering, Daegu University, Gyeongbuk 38453, Republic of Korea; Research Institute of Anti-Aging, Daegu University, Gyeongbuk 38453, Republic of Korea. 2. Brain Science and Engineering Institute, School of Medicine, Kyungpook National University, Daegu, Republic of Korea. Electronic address: eunjungkim@knu.ac.kr. 3. Department of Biotechnology, College of Engineering, Daegu University, Gyeongbuk 38453, Republic of Korea; Research Institute of Anti-Aging, Daegu University, Gyeongbuk 38453, Republic of Korea; Institute of Industrial and Technology, Daegu University, Gyeongbuk 38453, Republic of Korea. Electronic address: jangwg@daegu.ac.kr.
Abstract
AIMS: Costunolide is a sesquiterpene lactones used in many herbal medicines, with well-established anti-inflammatory and anti-oxidant functions modulating endoplasmic reticulum (ER) stress pathways, and which promotes the expression of anti-oxidant genes. The aim of this study is to investigate whether costunolide is involved in osteoblast differentiation and, determine the mechanisms of differentiation in mesenchymal stem cells. MAIN METHODS: The cytotoxicity of costunolide was identified using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The mRNA and protein expression levels of osteogenic genes were determined by RT-PCR and Western blot analysis. Alkaline phosphate (ALP) staining and Alizarin red S (ARS) staining were performed to evaluate ALP activity and matrix mineralization. Transcriptional activity was detected using a luciferase reporter assay. KEY FINDINGS: In this study, we determined that costunolide increased the expression of distal-less homeobox 5 (Dlx5), runt-related transcription factor 2 (Runx2), ALP, and osteocalcin (OC) in C3H10T 1/2 cells. Furthermore, costunolide increased ALP activity and matrix mineralization. Interestingly, costunolide increased ER stress by Bip, activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP). However, it did not exert effects on expression of activating transcription factor 6 (ATF6). ATF4 activation has a protective role in oxidative stress, and its transcription induces anti-oxidant genes in cells. Heme oxygenase-1 (HO-1) is a major anti-oxidant enzyme, and is regulated by ATF4. We showed that costunolide treatment increased HO-1 expression. Furthermore, the HO-1 inhibitor, Sn(IV) Protoporphyrin IX dichloride (SnPP) was blocked costunolide-induced Runx2 expression. SIGNIFICANCE: Our results revealed that costunolide-induced osteoblast differentiation is regulated by ATF4-dependent HO-1 expression.
AIMS: Costunolide is a sesquiterpene lactones used in many herbal medicines, with well-established anti-inflammatory and anti-oxidant functions modulating endoplasmic reticulum (ER) stress pathways, and which promotes the expression of anti-oxidant genes. The aim of this study is to investigate whether costunolide is involved in osteoblast differentiation and, determine the mechanisms of differentiation in mesenchymal stem cells. MAIN METHODS: The cytotoxicity of costunolide was identified using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The mRNA and protein expression levels of osteogenic genes were determined by RT-PCR and Western blot analysis. Alkaline phosphate (ALP) staining and Alizarin red S (ARS) staining were performed to evaluate ALP activity and matrix mineralization. Transcriptional activity was detected using a luciferase reporter assay. KEY FINDINGS: In this study, we determined that costunolide increased the expression of distal-less homeobox 5 (Dlx5), runt-related transcription factor 2 (Runx2), ALP, and osteocalcin (OC) in C3H10T 1/2 cells. Furthermore, costunolide increased ALP activity and matrix mineralization. Interestingly, costunolide increased ER stress by Bip, activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP). However, it did not exert effects on expression of activating transcription factor 6 (ATF6). ATF4 activation has a protective role in oxidative stress, and its transcription induces anti-oxidant genes in cells. Heme oxygenase-1 (HO-1) is a major anti-oxidant enzyme, and is regulated by ATF4. We showed that costunolide treatment increased HO-1 expression. Furthermore, the HO-1 inhibitor, Sn(IV) Protoporphyrin IX dichloride (SnPP) was blocked costunolide-induced Runx2 expression. SIGNIFICANCE: Our results revealed that costunolide-induced osteoblast differentiation is regulated by ATF4-dependent HO-1 expression.
Authors: Xueman Zhou; Wenxiu Yuan; Xin Xiong; Zhenzhen Zhang; Jiaqi Liu; Yingcheng Zheng; Jun Wang; Jin Liu Journal: Front Cell Dev Biol Date: 2021-11-30
Authors: Ana Paço; Teresa Brás; Jacqueline O Santos; Paula Sampaio; Andreia C Gomes; Maria F Duarte Journal: Molecules Date: 2022-02-08 Impact factor: 4.411