EGCG Targets Notch To Attenuate The Inflammatory Response In The Immediate Early Stage

The inflammatory response has an important role at different stages of diabetes mellitus(DM),tumorigenesis and cardiovascular diseases.Attenuation of the inflammatory response has been considered an important side effect in the therapy and prevention of cancer,DM,cardiovascular diseases,and Alzheimer's disease.The mechanisms of the inflammatory response have been studied extensively.The NF-?B molecule has been considered to be a key regulator/mediator of LPS-induced inflammation.The well-known mitogen-activated protein kinase(MAPK)pathway,which plays a key part in many other biologic processes,has also been confirmed to participate in the inflammatory response.In recent years,the Notch signaling pathway has been found to have an essential role in regulation of the inflammatory response.Basal activation of the Notch signal is required to amplify the LPS-deviced inflammation response.Inhibition activation of Notch can attenuate inflammatory responses effectively.(-)-Epigallocatechin gallate(EGCG)is found in Chinese tea.It has been proposed to have strong anti-inflammatory effects.The mechanisms of EGCG in the attenuation of inflammation have been explored.However,those studies have focused on the physiologic response of cells ?6 h after EGCG treatment.Such cells have undergone several rounds of signaling processes in ?6 h,and cytokine release and feedback may have already occurred in those cells.Thus,those results may indicate that EGCG treatment interferes with mediator proteins in the signal transduction pathway,though the original mechanism of EGCG treatment is not clear.To understand the detailed mechanisms of EGCG in inflammation,we examined the immediate early response of macrophages after EGCG addition and the underlying mechanisms.We studied the immediate early mechanism of EGCG on the inflammatory response induced by LPS in human macrophages derived from THP-1 cells.To evaluate the direct effects of EGCG on inflammation,THP-1 derived macrophages were treated with EGCG.To determine the effects of EGCG on inflammation in macrophages,we examined the inflammatory cytokines of THP-1 derived macrophages in culture supernatant using Elisa and human inflammation antibody array.These results showed that the EGCG inhibited proinflammatory cytokine release to the medium in THP-1-derived macrophages after during 3h of inflammation induced by LPS.To identify how EGCG attenuated the immediate inflammatory response on THP-1 derived macrophages.We found that EGCG did not alter the phosphorylation levels of NF-?B or the phosphorylation state of the key MAPK molecules p38,Erk and JNK after EGCG treatment using western blot.The Notch signaling pathway has a pivotal role in the inflammation response.EGCG eliminated mature Notch from the cell membrane independent of 67 LR.MG132 treatment inhibits EGCG-induced Notch degradation via the proteasome pathway.67 LR has been proposed to be a receptor for EGCG on cell membranes of mouse.To ascertain if 67 LR is a receptor for EGCG on cell membranes of human.EGCG attenuated the immediate inflammatory response via the same mechanism.Using 67 LR closed antibodies blocking the combination of EGCG and 67 LR.The effect did not disappear that EGCG can suppress production of inflammatory mediators induced by LPS in human derived macrophages.EGCG treatment can inhibit the expression of Notch protein,independent of 67 LR.These results showed that EGCG have a new target except 67 LR on cell membrane.To determine whether the EGCG attenuates the inflammatory response via the Notch pathway.Knockdown of Notch 1/2 expression by RNA interference impaired the downregulation of the inflammatory response by EGCG using human inflammation antibody array.These results uncovered that EGCG targets Notch to regulate the inflammatory response in the immediate early stage.However,a human leukemic cell line(THP-1)cell is derived from the blood of a boy with acute monocytic leukemia,and so it may demonstrate different mechanisms with regard to regulation of the inflammatory response.To evaluate the effects of EGCG on normal human cells,we examined the anti-inflammatory effect of EGCG on primary human macrophages differentiated from PBMCs collected from healthy human donors.To identify how EGCG attenuated the immediate inflammatory response on THP-1 derived macrophages through Notch signaling pathway and whether EGCG inhibited LPS-induced inflammation and turned off Notch signaling in PBMC derived macrophages.These results showed that EGCG could inhibit inflammation through the Notch signaling pathway in PBMC derived macrophages model.Taken together,our results uncovered that EGCG targets Notch to regulate the inflammatory response in the immediate early stage,a deeper understanding of the EGCG inflammation mechanism.Research results,could provide more scientific guidance for the people to drinking tea or tea products containing EGCG,theoretical evidence for the prevention and treatment of inflammation related diseases through scientific drinking tea,meanwhile,can also help people to tea analogues,as traditional Chinese medicine,to provide experience for the modern scientific research of traditional Chinese medicine.