Sirt6 Alleviated Liver Fibrosis by Deacetylating Conserved Lysine 54 on Smad2 in Hepatic Stellate Cells

Abstract

Backgrounds and Aims Activation of hepatic stellate cells (HSCs) is a central driver of fibrosis. This study aimed to elucidate the role of the deacetylase sirtuin 6 (Sirt6) in HSC activation and liver fibrosis. Approach and Results Gain‐of‐function and loss‐of‐function models were used to study the function of Sirt6 in HSC activation. Mass spectrometry was used to determine the specific acetylation site. The lecithin retinol acyltransferase–driven cyclization recombination recombinase construct (CreERT2) mouse line was created to generate HSC‐specific conditional Sirt6‐knockout mice (Sirt6△HSC). We found that Sirt6 is most abundantly expressed in HSCs as compared with other liver cell types. The expression of Sirt6 was decreased in activated HSCs and fibrotic livers of mice and humans. Sirt6 knockdown and Sirt6 overexpression increased and decreased fibrogenic gene expression, respectively, in HSCs. Mechanistically, Sirt6 inhibited the phosphorylation and nuclear localization of mothers against decapentaplegic homolog (Smad) 2. Further study demonstrated that Sirt6 could directly interact with Smad2, deacetylate Smad2, and decrease the transcription of transforming growth factor β/Smad2 signaling. Mass spectrometry revealed that Sirt6 deacetylated conserved lysine 54 on Smad2. Mutation of lysine 54 to Arginine in Smad2 abolished the regulatory effect of Sirt6. In vivo, specific ablation of Sirt6 in HSCs exacerbated hepatocyte injury and cholestasis‐induced liver fibrosis in mice. With targeted delivery of the Sirt6 agonist MDL‐800, its concentration was 9.28‐fold higher in HSCs as compared with other liver cells and alleviated hepatic fibrosis. Conclusions Sirt6 plays a key role in HSC activation and liver fibrosis by deacetylating the profibrogenic transcription factor Smad2. Sirt6 may be a potential therapeutic target for liver fibrosis.

Inhibition of 5-Lipoxygenase in Hepatic Stellate Cells Alleviates Liver Fibrosis

“…cRGD is pentapeptide that binds with high affinity to integrin αV and β3 which are highly expressed in a-HSC ( Li et al, 2019 ). It was also confirmed that cRGD-guided Lips specifically target activated HSC in vitro and vivo ( Li et al, 2019 ; Zhang et al, 2020 ). In agreement with a previous report, we found that RGD-Lip-delivered zileuton was highly enriched in HSC but not hepatocytes, Kupffer cells, endothelial cells or biliary cells ( Supplementary Figure S9B ).…”

“…We used an HSC-specific drug delivery system by modifying sterically stable liposome (Lip) with cRGDyK, a pentapeptide that binds to integrin αvβ3 on the surface of a-HSC ( Li et al, 2019 ). cRGDyK-guided Lip showed high selectivity toward activated but not quiescent HSC, and preferentially accumulated in the fibrotic liver ( Li et al, 2019 ; Zhang et al, 2020 ). We loaded with zileuton, an inhibitor of 5-LO, into this delivery system (RGD-Lip/zileuton).…”

“…Liposome (Lip) were prepared by the thin-film hydration method as described ( Li et al, 2019 ; Zhang et al, 2020 ). Zileuton-loaded RGD-Lips (RGD-Lip/zileuton) were prepared by adding zileuton to the lipid organic solution before the solvent evaporation.…”

Emerging roles of SIRT6 in human diseases and its modulators

“…Moreover, SIRT6‐mediated activation of SOD2/Prdx6 pathway is pivotal for antidepressant response, which requires directing binding of SIRT6 to superoxide dismutase 2 (SOD2) and peroxiredoxin 6 (Prdx6), subsequently deacetylating them at Lys68/122 and Lys63/209, respectively 51 . Recently, SIRT6 has also been observed to suppress β–Smad family members (SMADs) 2 and 3 through deacetylation at Lys54 and Lys333/378, respectively, to protect against liver fibrosis 52,53 . Last but not least, SIRT6 was demonstrated to deacetylate p53 on K382, 54 which may significantly affect various physiological functions such as cancer prevention and stress resistance 54–57 …”

“…Moreover, SIRT6 deacetylation may also exert liver protection by transcriptionally activating XBP1s, which confer resistance to ER stress‐induced hepatic steatosis 50 . In addition, SIRT6 activation may alleviate liver fibrosis by suppressing SMAD 2/3 signaling to affect hepatic stellate cells activation or transcriptionally inhibiting activity of orphan nuclear receptor estrogen‐related receptor γ (ERR‐γ) to impact bile acid production, representing a new therapeutic potential for treating nonalcoholic steatohepatitis (NASH) and cholestatic liver injury, respectively 52,53,261 . In terms of attenuating NASH progression, SIRT6 can also curb inflammation and oxidative stress through enhancing the nuclear import of nuclear factor Nrf2 and transcriptionally activating phase II/antioxidant genes 256 .…”