Abstract
Background and Purpose: Liver fibrosis is one of the leading
causes of morbidity and mortality worldwide of which no acceptable
therapy exists. Accumulating evidence supports
that glioma-associated oncogene
homologue 1(GLI1) is a potentially important therapeutic target for
liver fibrosis. This study investigates the antifibrotic activities and
potential mechanisms of Physalin B (PB), a natural Solanaceae compound.
Experimental Approach: Mice subjected to CCl4challenge and bile duct ligation were used to study the antifibrotic
effects of PB in vivo . Mouse primary hepatic stellate cells
(pHSCs) and human HSC line LX‐2 also served as an in vitro liver
fibrosis model. Liver fibrogenic genes, GLI1 downstream genes were
examined using western blot and real-time PCR analyses. GLI1 acetylation
and LAP2α-HDAC1 interaction were analyzed by coimmunoprecipitation.
Key Results: In animal models, PB administration attenuated
hepatic histopathological injury, collagen accumulation, and reduced the
expression of fibrogenic genes. PB dose‐dependently suppressed fibrotic
marker expression in LX‐2 cells and mouse pHSCs. Mechanistic studies
showed PB inhibited GLI activity in a non-canonical Hedgehog signaling.
PB blocked lamina-associated polypeptide 2 α (LAP2α)/ histone
deacetylase 1 (HDAC1) complex formation thereby inhibited HDAC1mediated
GLI1 deacetylation. PB downregulated the acetylation and expression of
GLI1, and subsequently inhibiting HSC activation.
Conclusions and Implications: PB exerted potent antifibrotic
effects in vitro and in vivo by disrupting the LAP2α/HDAC1
complex, increasing GLI1 acetylation and inactivating GLI1. This
indicates that PB may be a potential therapeutic candidate for the
treatment of liver fibrosis.
Introduction
Liver
fibrosis, a wound-healing response to chronic liver injury, is
characterized by excessive deposition of extracellular matrix (ECM) in
the liver. During chronic liver damage, HSCs are continuously activated
and converted into myofibroblasts, which are the major source of ECM and
the principal cell type in liver fibrogenesis(Friedman, 2008).
This
transient response must be tightly controlled, otherwise, it would
become persistent and lead to excessive matrix accumulation and
fibrosis(Lee, Wallace & Friedman, 2015). Thus, potential therapies for
these pathologies should ideally possess pro regenerative and
antifibrotic properties, for example, the ability of controlling
hepatocyte proliferation and inactivating activated HSC(Hernandez-Gea &
Friedman, 2011). Currently, there are no approved drugs that can
effectively reverse liver fibrosis, further highlighting the urgent
clinical need for novel antifibrotic therapies.
GLI
(glioma-associated oncogene
homologue) was identified as a gene that is amplified in
gliomas(McMillin et al.,
2014),
and it is one of the strongest transcriptional activators of Hh
signaling pathway, can also be regulated independent of the canonical Hh
signaling pathway. The TGFβ signaling pathway activates GLI1 even in the
presence of SMO antagonist (Dennler et al., 2007). In injured organs,
resident GLI1+ cells were committed to the
myofibroblast lineage, which directly regulates HSC fate (Machado &
Diehl, 2018). Genetic ablation of GLI1+ cells
ameliorated fibrosis and improved organ function, providing a proof of
principle for therapeutic targeting of GLI1+ cells in
fibrotic disease (Kramann et al., 2015). The aforementioned reports
emphasize the importance of the mechanisms that terminate GLI1 activity
and that may be impaired in disease. Hence, GLI1 is a potential
biomarker and therapeutic target aiming at controlling activation of
HSCs during liver fibrogenesis.
GLI1 acetylation, a modification that limits the activity of this
transcription factor (Miele et al., 2017). Acetylation represents a key
transcriptional event, finely tuned by histone acetyltransferases (HATs)
and histone deacetylases (HDACs). It has been observed that loss of
class I HDAC disrupts the transcriptional response to Hh activation,
HDAC1 is shown to deacetylate the transcription factor GLI1 (Canettieri,
Di Marcotullio, Coni, Greco & Gulino, 2010), which allows GLI1 to
associate with chromatin and initiate transcription (Canettieri et al.,
2010; Coni et al., 2013). Such deacetylation mechanisms are involved in
GLI1-dependent cell proliferation, migration, differentiation, and
survival.
Currently, few antifibrotic treatment strategies are available. Thus,
there is an urgent clinical need for the development of antifibrotic
candidates specifically targeting HSCs. Our study is engaged in
evaluating the antifibrotic activities of natural compounds. We have
utilized a high-throughput drug screening model based on the COL1A1
promoter to filtrate potential anti-hepatic fibrosis agents from natural
substances. Physalin B (PB), a natural component derived from one of the
best-known traditional Chinese medicinal plants, Physalis species,
Solanaceae, was speculated having potential therapeutic effect for liver
fibrosis by us. Previous studies have described the biological functions
of PB, including anti‐inflammatory (Hsu et al., 2012; Yang, Yi, Wang,
Xie, Sha & Dong, 2018), antimalarial(Magalhaes et al., 2006),
proapoptotic(Soares et al., 2006), antinociceptive (Vandenberghe et al.,
2008), and antitumor activities(Hsu et al., 2012; Ma, Han, Li, Hu &
Zhou, 2015), but the effects of PB on liver fibrosis have not been
reported thus far. We investigated the anti-fibrotic effects of PB on
human stellate cell line LX-2, primary HSCs, and experimental hepatic
fibrosis mice induced by CCl4 and bile duct ligation
(BDL), finally elucidated the potential mechanisms underlying its
function in vitro and in vivo .