P006 Engineered propionate-producing bacteria attenuates murine colitis by modulating the immune function of resident macrophages via histone deacetylase
Feng, Z.(1)*;Wang, X.(1);Kang, G.(2);Zhao, J.(1);Ye, Y.(1);Liu, L.(1);Huang, H.(2);Cao, X.(1);
(1)Tianjin Medical University General Hospital, Department of Gastroenterology and Hepatology, Tianjin, China;(2)Tianjin University, Department of Biochemical Engineering- School of Chemical Engineering and Technology, Tianjin, China;
Background
Most inflammatory bowel disease (IBD) patients are unable to maintain long-term remission by current treatment. Propionate has been proven to be able to exert anti-inflammatory effects in intestinal inflammation, but direct supplement of propionate has actual disadvantages including low bioavailability and poor sustained effect, et. al.
Methods
In this study, the genes related to acetate synthesis and metabolism were knocked out through the double plasmid CRISPR/cas9 gene editing system to increase the yield of propionate to ultimately form the engineered propionate-producing bacteria (EPB). The dextran sulfate sodium（DSS）-induced colitis model was used to explore the protective effect of EPB in vivo. Flow cytometry was used to evaluate the ratio of resident macrophage. Peritoneal macrophage and RAW264.7 cell lines were used to explore the effect and mechanism of EPB on the immune function of macrophages in vitro. Real-time polymerase chain reaction (RT-PCR) was performed to detect the expression of several cytokines in colon tissue and cells. Enzyme-linked immunosorbent assay (ELISA) was used to detect the level of cytokines in the cell supernatant. Western bolt was used to detect the levels of histone deacetylase protein and transcription factor GATA6.
Results
EPB exhibited enhanced propionic acid production. In vivo results showed that EPB group had less barrier damage and less inflammatory infiltration than control groups indicating it could prominently ameliorate DSS-induced colitis. EPB promoted the production of anti-inflammatory cytokine IL-10 and reduced the production of IL-2、IL-5 and IL-6 in DSS-induced mice. In the further exploration of the effects of EPB, we found that EPB increased the proportion of resident macrophages (F4/80hiCD11bhi) in the peritoneal cavity, And the transcription factor GATA6, which is specific express in peritoneal macrophages, was detected in the intestinal tissue. Moreover, we investigated the effects of EPB on peritoneal macrophages in vitro and found EPB not only affected the cytokine production ability of resident macrophages but also reduced cell apoptosis. Further analysis showed the regulatory effect of EPB on macrophages is associated with G protein-coupled receptors 41/43 and histone deacetylase 2 (HDAC2), which is closely related to propionate. GPR41/43 inhibitor weakened the effects of EPB on macrophages, while HDAC inhibitor trichostatin A (TSA) exerted a similar effect to EPB.
Conclusion
The engineered propionate-producing bacteria (EPB) could attenuate DSS-induced colitis in the mouse by modulating the immune ability of resident macrophage and the production of classic inflammation-related cytokines, which indicates an effective and safe treatment strategy for ulcerative colitis.