S-Space College of Natural Sciences (자연과학대학) Dept. of Biological Sciences (생명과학부) Theses (Ph.D. / Sc.D._생명과학부)
Molecular Characterization of Anti-Inflammatory and Anti-Osteoporotic Activities of Dehydrodiconiferyl Alcohol
Dehydrodiconiferyl Alcohol의 항염증 및 항골다공증 활성에 대한 분자 기전 연구
- 자연과학대학 생명과학부
- Issue Date
- 서울대학교 대학원
- IMQ-induced psoriasis-like skin inflammation model; ovariectomy-induced mouse model; estrogen receptor; RANKL; BMP-2; Dehydrodiconiferyl alcohol; inflammation; phytoestrogen; osteoclast; osteoblast; IL-17
- 학위논문 (박사)-- 서울대학교 대학원 : 자연과학대학 생명과학부, 2018. 2. 김선영.
- Dehydrodiconiferyl alcohol (DHCA) is a lignan compound isolated from Cucurbita moschata. Synthetic DHCA has previously been shown to contain anti-adipogenic, anti-oxidative stress and anti-inflammatory activities. In an effort to understand the underlying mechanisms of such multiple bioactivities of this lignan molecule, my thesis research was focused on the effects of DHCA on IL-17-mediated inflammation and also on its potential estrogenic activities, both in vitro and in vivo.
The effects of DHCA on IL-17-mediated inflammation were investigated using HaCaT keratinocyte cell line. DHCA significantly inhibited the IL-17-mediated cell proliferation and suppressed the expression of various inflammatory mediators, such as TNF-α, IL-6, IL-1β and various chemokines by inhibiting p38 MAPK signaling pathway. Consistent with these in vitro data, in the imiquimod-induced psoriasis-like skin inflammation mouse model, DHCA ameliorated psoriatic symptoms, histological phenotypes and expression of various inflammatory mediators. Data from immunohistochemical analysis and ex vivo culture experiments suggested that DHCA reduced the infiltration of IL-17 producing inflammatory cells by suppressing various chemokines such as CXCL1, CXCL8 and CCL20.
Being a lignan molecule, DHCA is a member of the phytoestrogen family. Therefore, it was investigated whether DHCA contains anti-osteoporotic activities similar to estrogen. The effects of DHCA were studied on RANKL-induced osteoclastogenesis using RAW264.7 pre-osteoclast cell line. DHCA effectively inhibited the RANKL-induced differentiation and function of osteoclast in a dose-dependent manner. DHCA also suppressed the expression of various osteoclastogenic genes, including NFATc1, TRAP, c-Fos, DC-STAMP, MMP-9, and Cathepsin K, through the inhibition of NF-ĸB and p38 MAPK signaling pathways. These anti-osteoclastogenic effects of DHCA were suppressed when cells were transfected with siRNAs for AMPKα1 or ERα, whereas ERβ siRNA did not have any effect.
The effects of DHCA on BMP-2-induced osteoblastogenesis were also studied using MC3T3-E1 pre-osteoblast cell line. DHCA promoted BMP-2-induced differentiation of osteoblast in a dose-dependent manner. This lignan molecule further up-regulated the BMP-2 mediated activation of Smad1/5/9 and AMPK signaling pathways, the expression of RUNX2 and subsequently that of ALP, osteocalcin and OPG. Gene knockdown analysis, involving specific siRNAs for ERα or ERβ, indicated that DHCA might interact with either ERα or ERβ to promote the BMP-2-induced osteoblast differentiation.
Above data indicated that DHCA might produce anti-osteoporotic activities through its agonistic effect on estrogen receptor. When an ovariectomized mouse model was used, DHCA indeed improved a variety of bone morphometric parameters as determined by 3D-structure analysis. DHCA also reduced the blood level of NTx and CTx, biochemical markers for bone degradation which also regulate the expression of osteoclastogenic and osteoblastogenic genes in the bone marrow.
Together with our previous findings, data from my thesis work demonstrated that DHCA had a wide range of bioactivities including anti-adipogenic, anti-oxidative stress, anti-inflammatory, anti-osteoclastogenic and osteoblastogenic activities. Such multiple bioactivities of DHCA could be best explained if this molecule acts like estrogen. Data from molecular docking simulation suggested that DHCA could bind to both ERα and ERβ with a similar binding pose to estrogen. Furthermore, the 2D ligand-receptor interaction diagram showed that intermolecular forces and MM-GBSA binding energy were analogous to the case of estradiol. Indeed, results from estrogen receptor competition assay indicated that DHCA could efficiently bind to ERα and ERβ.
In conclusion, high therapeutic effects of DHCA observed in psoriasis and osteoporosis mouse models could be explained with DHCA acting as an estrogen receptor agonist, and thus producing anti-inflammatory, anti-osteoclastogenic and osteoblastogenic activities. Taken together, with previous findings, DHCA may be developed as a safe and effective therapeutic agent for the treatment of various diseases where inflammation and/or estrogen play prominent role(s).