The Role Of SIRT5 In The Regulation Of Inflammatory Response And Glucose Homeostasis,and Lipidomics Study Of Macrophages

Immunometabolism is an emerging field of investigation at the interaction between the immunology pathway and metabolic pattern.Dissection of the molecular mechanism of the cross-talk between immunology and metabolism has become a priority.SIRT5 has a variety of diacylation functions,and regulates diverse key metabolic processes in cells.However,the role of SIRT5 in inflammation and metabolic regulation remain largely unknown.In the first part of this study,we shown that SIRT5 in maintaining glucose homeostasis for immune cell survival during bacterial challenge and for host survival in systemic infection.We systematically investigated the role of SIRT5 in the regulation of inflammatory reaction,cell metabolism and energy balance,providing important metabolic regulation strategy for the intervention of infection-related diseases or metabolic disorders.Traditionally,lipids are regarded as energy storage molecules or general membrane construction.Now,lipids are recognized as potent signaling molecules that regulate multiple cellular responses,such as energy transfer,signal transduction,cell growth and apoptosis.Disruption of lipid metabolic balance is an important mechanism for inducing cellular oxidative stress and inflammatory response.Lipid metabolism contributes to the pro-or anti-inflammatory functions of macrophages.However,there are few studies focused on lipid patterns in the integrity processes of monocyte-macrophage differentiation and macrophage polarization.With the development of mass spectrometry technique,the detection of lipids has become more comprehensively and rapidly.Mass spectrometry has become the most important technique for lipidomics research.In the second part of this study,we identified the changes of the lipid composition in the process of human and murine macrophages differentiation with the target mass spectrometry.This lipidomic analysis can help uncover new candidate markers of the inflammatory response and shed new light on monocyte-to-macrophage differentiation and macrophage polarization.This thesis was described as the following four chapters:The Chapter 1 is the introduction of this study.Firstly,the research progress of immunometabolism study was summarized from two parts.On the one hand,it has emerged that the immune systems could be mobilized by some non-immune pathologies,like metabolic disorders.On the other hand,the function of immune cells on many levels was controlled by internal metabolic patterns.Then,the role of SIRT5 in the regulation of metabolic pathway and inflammation,like fatty acid oxidation,ketone body synthesis,urea cycle and serine metabolism,ROS production,were introduced.Besides,the lipidomics methodology and present status in inflammation-related or metabolic diseases were summarily described.The Chapter 2 is about the investigation of SIRT5 in inflammation response and glucose homeostasis regulation.In this part,we first shown that the level of blood glucose was decreased,serum insulin and IL-1? was increased in Sirt5 KO mice challenged with LPS stimulation or metabolic stress.Moreover,LPS induced the activation of NLRP3 inflammasome and IL-1? secretion in Sirt5 KO BMDMs.The mechanism involved NF-?B,AMPK,and ROS production.Finally,we demonstrated that SIRT5 is important in maintaining glucose homeostasis for inflammation controlling during bacterial challenge.Our investigation provided an important basis for the intervention of macrophage-related immune diseases and infection diseases.In Chapter 3,by using a LC-MRM technology,the lipidomics study of mammal monocytes and macrophages was carried out.The present study investigated the dynamic lipid profiles in the complete processes of monocyte-macrophage differentiation and macrophage polarization.Firstly,we induced and identified different activated macrophages in vitro.Human monocyte THP-1 cells were induced into unactived macrophages(M0),M0 were then stimulated into the classically activated macrophage M1 or alternatively activated macrophage M2.And the lipids were extracted from different activated macrophages.Using the MRM scanning,we detected nearly 500 lipid molecules,including glycerophospholipids and sphingolipids: PC?PS?PE?PI?PG?SM and Cer.We observed a shift in the composition of the glycerophospholipids from short-chain monounsaturated lipids to long-chain polyunsaturated lipids in activated macrophages derived from THP-1 cells.A higher level of lyso GPs in mouse M2 macrophages than that in M1 macrophages.This lipidomic analysis can shed new light on monocyte differentiation and macrophage polarization,and may help uncover new candidate markers of the inflammatory response.The Chapter 4 is a review about the methodology of mass spectrometry,and the applications in life sciences and clinical medical research.Mass spectroscopy has become one of the most important analytical tools for biomedical research,depending on its high sensitivity,selectivity,and accuracy.The emergence of ionization technology of ESI and MALDI promoted the development and application of mass spectrometry in the life sciences.Nowadays,biological mass spectrometry is not only for the research of proteomics,metabolomics,and lipidomics,but also widely used for clinical diagnosis,disease screening and medical testing.ConclusionsIn the first part:1.SIRT5 has a potential role in regulating glucose homeostasis and Sirt5 deficient boosts IL-1? production in inflammatory response.2.Macrophage are the resources of IL-1? in Sirt5 KO mice,and the macrophages play an inportant role in regulating blood glucose and insulin in Sirt5 KO mice.3.LPS induced the activation of NLRP3 inflammasome and IL-1? secretion in Sirt5 KO BMDMs.The mechanism involved NF-?B,AMPK,and ROS production.4.Macrophage-derived IL-1? promotes insulin secretion and insulin can further stimulate IL-1? secretion by macrophages in Sirt5 KO mice.5.SIRT5 is important in maintaining glucose homeostasis for inflammation controlling during bacterial challenge.In the second part:1.We successfully induced different activated macrophages M1 and M2 in vitro.By using the LC-MRM method,368 lipid molecules were detected in human THP-1 cells,and 420 lipid molecules were detected in mouse RAW264.7 macrophages.2.We observed a shift in the composition of the glycerophospholipids from short-chain monounsaturated lipids to long-chain polyunsaturated lipids in activated macrophages derived from THP-1 cells.3.A higher level of lyso GPs in mouse M2 macrophages than that in M1 macrophages,especially the level of lyso PI.