Linnan Zhu

Cellular Metabolism and Macrophage Functional Polarization

Macrophages are a functionally heterogeneous cell population that is mainly shaped by a variety of microenvironmental stimuli. Interferon γ (IFN-γ), interleukin-1β (IL-1β), and lipopolysaccharide (LPS) induce a classical activation of macrophages (M1), whereas IL-4 and IL-13 induce an alternative activation program in macrophages (M2). Reprogramming of intracellular metabolisms is required for the proper polarization and functions of activated macrophages. Similar to the Warburg effect observed in tumor cells, M1 macrophages increase glucose consumption and lactate release and decreased oxygen consumption rate. In comparison, M2 macrophages mainly employ oxidative glucose metabolism pathways. In addition, fatty acids, vitamins, and iron metabolisms are also related to macrophage polarization. However, detailed metabolic pathways involved in macrophages have remained elusive. Understanding the bidirectional interactions between cellular metabolism and macrophage functions in physiological and pathological situations and the regulatory pathways involved may offer novel therapies for macrophage-associated diseases.

Digital Earth: decadal experiences and some thoughts

Abstract The understanding that mankind should reasonably exploit and utilize earth resources and effectively protect the planet on which we live, is now widely accepted. However, effective actions can only be conducted if we better understand and visualize the earth. To meet this need, digital earth science and technology have been put forward and developed. This paper introduces the evolution and development process of digital earth, and presents an overview by reviewing and analyzing the 1999 and 2009 Beijing Declaration on Digital Earth, the scientific and commercial digital earth systems, global and regional digital earth research, and some existing platforms of digital earth science. It also presents some thoughts about digital earths future development.

Induction of M2-like macrophages in recipient NOD- scid mice by allogeneic donor CD4 + CD25 + regulatory T cells

CD4+CD25+ regulatory T cells (Tregs) play an important role in maintaining host immune tolerance via regulation of the phenotype and function of the innate and adaptive immune cells. Whether allogeneic CD4+CD25+ Tregs can regulate recipient mouse macrophages is unknown. The effect of allogeneic donor CD4+CD25+ Tregs on recipient mouse resident F4/80+macrophages was investigated using a mouse model in which allogeneic donor CD4+CD25+ Tregs were adoptively transferred into the peritoneal cavity of host NOD-scid mice. The phenotype and function of the recipient macrophages were then assayed. The peritoneal F4/80+ macrophages in the recipient mice that received the allogeneic CD4+CD25+ Tregs expressed significantly higher levels of CD23 and programmed cell death-ligand 1(PD-L1) and lower levels of CD80, CD86, CD40 and MHC II molecules compared to the mice that received either allogeneic CD4+CD25− T cells (Teffs) or no cells. The resident F4/80+ macrophages of the recipient mice injected with the allogeneic donor CD4+CD25+ Tregs displayed significantly increased phagocytosis of chicken red blood cells (cRBCs) and arginase activity together with increased IL-10 production, whereas these macrophages also showed decreased immunogenicity and nitric oxide (NO) production. Blocking arginase partially but significantly reversed the effects of CD4+CD25+ Tregs with regard to the induction of the M2 macrophages in vivo. Therefore, the allogeneic donor CD4+CD25+ Tregs can induce the M2 macrophages in recipient mice at least in part via an arginase pathway. We have provided in vivo evidence to support the unknown pathways by which allogeneic donor CD4+CD25+ Tregs regulate innate immunity in recipient mice by promoting the differentiation of M2 macrophages.

Lipopolysaccharide Increases the Incidence of Collagen‐Induced Arthritis in Mice Through Induction of Protease HTRA‐1 Expression

OBJECTIVE The protease HTRA-1 is closely associated with rheumatoid arthritis (RA). The molecular mechanisms that control HTRA-1 expression are currently unknown. This study was undertaken to determine the regulatory role of Toll-like receptors (TLRs) on HTRA-1 expression in mice with collagen-induced arthritis (CIA) and in synovial cells from RA patients. METHODS HTRA-1 messenger RNA and protein production in mouse fibroblasts, mouse macrophages, and freshly isolated RA patient synovial cells treated with TLR ligands were detected by real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Arthritis incidence and severity were determined using clinical scores and histopathologic analysis. Involvement of HTRA-1 in lipopolysaccharide (LPS)-increased arthritis incidence and severity in mice was determined using anti-HTRA-1 monoclonal antibody. The signal pathways involved in HTRA-1 expression were accessed by specific inhibitors, RNA interference, dual-luciferase reporter, and chromatin immunoprecipitation methods. RESULTS LPS and tenascin-C, but not the other TLR ligands tested, strongly induced HTRA-1 expression. LPS significantly increased HTRA-1 expression in the joint tissue as well as arthritis incidence and severity in mice with CIA. Blocking HTRA-1 by antibody significantly decreased LPS-promoted CIA severity. Inhibiting NF-κB significantly decreased LPS-induced HTRA-1 expression in mouse and human cells. Dual-luciferase reporter assay and ChIP analysis showed that p65 directly binds to HTRA-1 promoter (amino acid 347). CONCLUSION Our findings indicate that TLR-4 activation increases HTRA-1 expression through the NF-κB pathway in fibroblasts and macrophages. HTRA-1 expression is involved in the enhancing effects of LPS on CIA. This study offers new insights into the regulation of HTRA-1 expression via LPS/TLR-4 and the role of HTRA-1 in RA pathogenesis.

The Inhibitory Effect of IFN-γ on Protease HTRA1 Expression in Rheumatoid Arthritis

The high temperature requirement A1 (HTRA1) is a potent protease involved in many diseases, including rheumatoid arthritis (RA). However, the regulatory mechanisms that control HTRA1 expression need to be determined. In this study, we demonstrated that IFN-γ significantly inhibited the basal and LPS-induced HTRA1 expression in fibroblasts and macrophages, which are two major cells for HTRA1 production in RA. Importantly, the inhibitory effect of IFN-γ on HTRA1 expression was evidenced in collagen-induced arthritis (CIA) mouse models and in human RA synovial cells. In parallel with the enhanced CIA incidence and pathological changes in IFN-γ–deficient mice, HTRA1 expression in the joint tissues was also increased as determined by real-time PCR and Western blots. IFN-γ deficiency increased the incidence of CIA and the pathological severity in mice. Neutralization of HTRA1 by Ab significantly reversed the enhanced CIA frequency and severity in IFN-γ–deficient mice. Mechanistically, IFN-γ negatively controls HTRA1 expression through activation of p38 MAPK/STAT1 pathway. Dual luciferase reporter assay and chromatin immunoprecipitation analysis showed that STAT1 could directly bind to HTRA1 promoter after IFN-γ stimulation. This study offers new insights into the molecular regulation of HTRA1 expression and its role in RA pathogenesis, which may have significant impact on clinical therapy for RA and possibly other HTRA1-related diseases, including osteoarthritis, age-related macular degeneration, and cancer.

Microgravity inhibition of lipopolysaccharide-induced tumor necrosis factor-α expression in macrophage cells

AbstractObjective and design Microgravity environments in space can cause major abnormalities in human physiology, including decreased immunity. The underlying mechanisms of microgravity-induced inflammatory defects in macrophages are unclear.Material or subjectsRAW264.7 cells and primary mouse macrophages were used in the present study. Lipopolysaccharide (LPS)-induced cytokine expression in mouse macrophages was detected under either simulated microgravity or 1g control.MethodsFreshly isolated primary mouse macrophages and RAW264.7 cells were cultured in a standard simulated microgravity situation using a rotary cell culture system (RCCS-1) and 1g control conditions. The cytokine expression was determined by real-time PCR and ELISA assays. Western blots were used to investigate the related intracellular signals.ResultsLPS-induced tumor necrosis factor-α (TNF-α) expression, but not interleukin-1β expression, in mouse macrophages was significantly suppressed under simulated microgravity. The molecular mechanism studies showed that LPS-induced intracellular signal transduction including phosphorylation of IKK and JNK and nuclear translocation of NF-κB in macrophages was identical under normal gravity and simulated microgravity. Furthermore, TNF-α mRNA stability did not decrease under simulated microgravity. Finally, we found that heat shock factor-1 (HSF1), a known repressor of TNF-α promoter, was markedly activated under simulated microgravity.ConclusionsShort-term treatment with microgravity caused significantly decreased TNF-α production. Microgravity-activated HSF1 may contribute to the decreased TNF-α expression in macrophages directly caused by microgravity, while the LPS-induced NF-κB pathway is resistant to microgravity.

Cellular Metabolism on T-Cell Development and Function

Cell metabolism is closely related to the host immunity in many respects. We herein briefly summarized the recent progress on the roles of cellular metabolism in T-cell development, homeostasis, differentiation and functions. Relatively quiescent naïve T cells only require energy for survival and migration, and they mainly metabolize glucose to carbon dioxide through oxidative phosphorylation. However, activated T cells engage in robust cell proliferation, produce of a range of effector molecules and migrate through peripheral tissues, so they utilizes glycolysis to convert glucose to lactate (termed aerobic glycolysis) to meet the significantly increased metabolic demands. Importantly, the differentiation of T-cell subsets and memory T cells (Tm) was also significantly shaped by distinct cellular metabolic pathways including glucose, amino acids (AA), fatty acids (FA), and others. Understanding the regulatory metabolic networks on immunity may offer new insights into the immune-related disorders and open novel potential therapies to prevent and treat immune diseases.

TSC1 controls IL-1β expression in macrophages via mTORC1-dependent C/EBPβ pathway

The tuberous sclerosis complex 1 (TSC1) is a tumor suppressor that inhibits the mammalian target of rapamycin (mTOR), which serves as a key regulator of inflammatory responses after bacterial stimulation in monocytes, macrophages, and primary dendritic cells. Previous studies have shown that TSC1 knockout (KO) macrophages produced increased inflammatory responses including tumor necrosis factor-α (TNF-α) and IL-12 to pro-inflammatory stimuli, but whether and how TSC1 regulates pro-IL-1β expression remains unclear. Here using a mouse model in which myeloid lineage-specific deletion of TSC1 leads to constitutive mTORC1 activation, we found that TSC1 deficiency resulted in impaired expression of pro-IL-1β in macrophages following lipopolysaccharide stimulation. Such decreased pro-IL-1β expression in TSC1 KO macrophages was rescued by reducing mTORC1 activity with rapamycin or deletion of mTOR. Rictor deficiency has no detectable effect on pro-IL-1β synthesis, suggesting that TSC1 positively controls pro-IL-1β expression through mTORC1 pathway. Moreover, mechanism studies suggest that mTORC1-mediated downregulation of the CCAAT enhancer-binding protein (C/EBPβ) critically contributes to the defective pro-IL-1β expression. Overall, these findings highlight a critical role of TSC1 in regulating innate immunity by control of the mTOR1-C/EBPβ pathway.

Characterization and biological significance of IL-23-induced neutrophil polarization

Neutrophils are heterogeneous with distinct subsets, and can switch phenotypes to exert regulatory functions on immunity. We herein demonstrate that IL-23-treated neutrophils selectively produce IL-17A, IL-17F and IL-22, and display a distinct gene expression profile in contrast to resting and lipopolysaccharide-treated neutrophils. IL-17+ neutrophils are present in the colons of mice with dextran sulfate sodium-induced colitis. Adoptive transfer of IL-23-treated neutrophils significantly promotes pathogenesis in this model. IL-23 induces neutrophil polarization through STAT3-dependent RORγt and BATF pathways. Thus, IL-23-induced neutrophil polarization expresses a unique cytokine-producing profile, which may contribute to IL-23-mediated inflammatory diseases.

2-Deoxy-d-Glucose Treatment Decreases Anti-inflammatory M2 Macrophage Polarization in Mice with Tumor and Allergic Airway Inflammation

As important effector cells in inflammation, macrophages can be functionally polarized into either inflammatory M1 or alternatively activated anti-inflammatory M2 phenotype depending on surroundings. The key roles of glycolysis in M1 macrophage polarization have been well defined. However, the relationship between glycolysis and M2 polarized macrophages is still poorly understood. Here, we report that 2-deoxy-d-glucose (2-DG), an inhibitor of the glycolytic pathway, markedly inhibited the expressions of Arg, Ym-1, Fizz1, and CD206 molecules, the hall-markers for M2 macrophages, during macrophages were stimulated with interleukin 4. The impacted M2 macrophage polarization by 2-DG is not due to cell death but caused by the impaired cellular glycolysis. Molecular mechanism studies indicate that the effect of 2-DG on M2 polarized macrophages relies on AMPK-Hif-1α-dependent pathways. Importantly, 2-DG treatment significantly decreases anti-inflammatory M2 macrophage polarization and prevents disease progression in a series of mouse models with chitin administration, tumor, and allergic airway inflammation. Thus, the identification of the master role of glycolysis in M2 macrophage polarization offers potential molecular targets for M2 macrophages-mediated diseases. 2-DG therapy may have beneficial effects in patients with tumors or allergic airway inflammation by its negative regulation on M2 macrophage polarization.