TNFSF15 facilitates the differentiation of CD11b^(+) myeloid cells into vascular pericytes in tumors

Objective:Immature vasculature lacking pericyte coverage substantially contributes to tumor growth,drug resistance,and cancer cell dissemination.We previously demonstrated that tumor necrosis factor superfamily 15(TNFSF15)is a cytokine with important roles in modulating hematopoiesis and vascular homeostasis.The main purpose of this study was to explore whether TNFSF15 might promote freshly isolated myeloid cells to differentiate into CD11b^(+) cells and further into pericytes.Methods:A model of Lewis lung cancer was established in mice with red fluorescent bone marrow.After TNFSF15 treatment,CD11b^(+) myeloid cells and vascular pericytes in the tumors,and the co-localization of pericytes and vascular endothelial cells,were assessed.Additionally,CD11b^(+) cells were isolated from wild-type mice and treated with TNFSF15 to determine the effects on the differentiation of these cells.Results:We observed elevated percentages of bone marrow-derived CD11b^(+)myeloid cells and vascular pericytes in TNFSF15-treated tumors,and the latter cells co-localized with vascular endothelial cells.TNFSF15 protected against CD11b^(+)cell apoptosis and facilitated the differentiation of these cells into pericytes by down-regulating Wnt3a-VEGFR1 and up-regulating CD49e-FN signaling pathways.Conclusions:TNFSF15 facilitates the production of CD11b^(+) cells in the bone marrow and promotes the differentiation of these cells into pericytes,which may stabilize the tumor neovasculature.

Hepatopoietin Cn suppresses apoptosis of human hepatocellular carcinoma cells by up-regulating myeloid cell leukemia-1

AIM:To investigate the role of hepatopoietin Cn(HPPCn) in apoptosis of hepatocellular carcinoma(HCC)cells and its mechanism. METHODS:Two human HCC cell lines,SMMC7721 and HepG2,were used in this study.Immunostaining, Western blotting and enzyme linked immunosorbent assay were conducted to identify the expression of HPPCn and the existence of an autocrine loop of HPPCn/ HPPCn receptor in SMMC7721 and HepG2.Apoptotic cells were detected using fluorescein isothiocyanate (FITC)-conjugated Annexin V and propidium iodide.RESULTS:The HPPCn was highly expressed in human HCC cells and secreted into culture medium(CM). FITC-labeled recombinant human protein(rhHPPCn) could specifically bind to its receptor on HepaG2 cells. Treatment with 400 ng/mL rhHPPCn dramatically increased the viability of HCC-derived cells from 48.1% and 36.9%to 85.6%and 88.4%,respectively(P< 0.05).HPPCn silenced by small-interfering RNA reduced the expression and secretion of HPPCn and increased the apoptosis induced by trichostatin A.Additionally, HPPCn could up-regulate the expression of myeloid cell leukemia-1(Mcl-1)in HCC cells via mitogen-activated protein kinase(MAPK)and sphingosine kinase-1. CONCLUSION:HPPCn is a novel hepatic growth factor that can be secreted to CM and suppresses apoptosis of HCC cells by up-regulating Mcl-1 expression.

Heterogeneity of tumor-infiltrating myeloid cells in era of single-cell genomics

Tumor microenvironment(TME)is highly heterogeneous and composed of complex cellular components,including multiple kinds of immune cells.Among all immune cells in TME,tumor-infiltrating myeloid cells(TIMs)account for a large proportion and play roles as key regulators in a variety of functions,ranging from immune-mediated tumor killing to tumor immune evasion.Understanding the heterogeneity of TIMs will provide valuable insights for new therapeutic targeting of myeloid cells.Single-cell genomic technologies deciphering cell composition and gene expression at single-cell resolution have largely improved our understanding of the cellular heterogeneity of TIMs and highlighted several novel cell subtypes contributing to the variation of patient survival and treatment response.However,these cell subtypes were defined based on limited data without a concordant nomenclature,which makes it difficult to understand whether they exist in different studies.Thus,in this review,we comprehensively summarized the common agreements and current different opinions on the heterogeneity of TIMs gained from single-cell studies;evaluated the feasibility of current myeloid cell targets at single-cell level and proposed a uniform nomenclature for TIM subsets.

Expression of triggering receptor-1 in myeloid cells of mice with acute lung injury

BACKGROUND： Myeloid cell （TREM-1） is an important mediator of the signal transduction pathway in inflammatory response. In this study, a mouse model of acute lung injury （ALl） by intraperitoneal injection of lipopolysaccharide （LPS） was established to observe the expression pattern of TREM-1 in lung tissue and the role of TREM-1 in pulmonary inflammatory response to ALl.METHODS： Thirty BALB/C mice were randomly divided into a normal control group （n=6） and an ALl group （n=24）. The model of ALl was made by intraperitonal injection of LPS in dose of 10 mg/ kg. Specimens from peripheral blood and lung tissue were collected 6, 12, 24 and 48 hours after LPS injection. RT-PCR was used to detect TREM-1 mRNA, and ELISA was employed for detection of TREM-1 protein and TNF-a protein, and HE staining was performed for the pathological Smith lung scoring under a light microscope.RESULTS： The expressions of TREM-1 mRNAin lung tissue and blood of the ALl group 6, 12, 24, and 48 hours after injection of LPS were higher than those in the control group. The levels of TREM- 1 protein and the levels of TNF-a protein in lung tissue of the ALl group 6, 12, 24, and 48 hours after LPS injection were higher than those of the control group; the level of TREM-1 protein peaked 12 hours after LPS injection, but it was not significantly correlated with the expression of TREM-1 mRNA （P=0.14）; the TNF-a concentration was positively correlated with TREM-1 levels in lung tissue and with Smith pathological score （r=0.795, P=0.001 ：r=0.499, P=0.034）, but not with the expression of TREM-1 mRNA （P=0.176）.CONCLUSION： The expression of TREM-1 mRNA in lung tissue of mice with ALl is elevated, and the expression of TREM-1 mRNA is related to the level of TNF-a and the severity of inflammatory response to ALl. The expressions of the TREM-1 gene are not consistent with the levels of TREM-1 protein, suggesting a new functional protein involved in immune regulation.

Relationships between genetic polymorphisms of triggering receptor expressed on myeloid cells-1 and septic shock in a Chinese Han population

BACKGROUND: Triggering receptor expressed on myeloid cells-1(TREM-1) is a cell surface receptor expressed on neutrophils and monocytes. TREM-1 acts to amplify infl ammation and serves as a critical mediator of infl ammatory response in the context of sepsis. To date, the predisposition of TREM-1 gene polymorphisms to septic shock has not been reported. This study was designed to investigate whether TREM-1 genomic variations are associated with the development of septic shock.METHODS: We genotyped two TREM-1 single nucleotide polymorphisms(SNPs, rs2234237 and rs2234246) and evaluated the relationships between these SNPs and septic shock on susceptibility and prognosis.RESULTS: TREM-1 rs2234246 A allele in the promoter region was signifi cantly associated with the susceptibility of septic shock in recessive model(AA, OR=3.10, 95%CI 1.15 to 8.32, P=0.02), and in codominant model(AG, OR=0.72, 95%CI 0.43–1.19, P=0.02; AA, OR=2.71, 95%CI 1.00–7.42; P=0.03). However, in three inherited models(dominant model, recessive model, and codominant model), none of the assayed loci was signif icantly associated with the prognosis of septic shock. The nonsurvivor group demonstrated higher plasma IL-6 levels(99.7±34.7 pg/mL vs. 61.2±26.5 pg/mL, P<0.01) than the survivor group. Plasma concentrations of IL-6 among the three genotypes of rs2234246 were AA 99.4±48.9 pg/m L, AG 85.4±43 pg/m L, and GG 65.3±30.7 pg/m L(P<0.01). The plasma concentrations of IL-6 in patients with AA genotypes were signifi cantly higher than those in patients with GG genotypes(P<0.01).CONCLUSION: TREM-1 genetic polymorphisms rs2234246 may be significantly correlated only with susceptibility to septic shock in the Chinese Han population.

CHARACTERIZING FLUORESCENCE LIFETIME OF NAD(P)H IN HUMAN LEUKEMIC MYELOID CELLS AND MONONUCLEAR CELLS

The aim of this er vito study was to explore the potential of using the fluorescence lifetime of intraellular reduced nicotinamide adenine dinucleotide(phosphate)(NAD(P)H)as a label-free indicator to characterize the di ferencs between human leukemic myeloid cells and normal mononuclear cells(MNC).The steady-state and time-resolved autofuorescence of two human leukemic myeloid cell lines(K562,HL60)and MNC were measured by a spectrofuorimeter.According to excitation-enmission matrix(EEM)analysis,the optimal emission of NAD(P)H in these cells suspensions occurred at 445 nm.Furthermore,the fuorescence lifetimes of NAD(P)H in leukemic myeloid cells and MNC were determined by fitting the time-resolved autofuorescence data.The mean fuorescence lifetimes of NAD(P)H in K562,HL60,and MNC cells were 557±1.19,4.45±0.71,and 7.31±0.60 ns,respectively.There was a significant diference in the mean lifetime of NAD(P)H between leukemic myeloid cells and MNC(p<0.05).The difference was essentally caused by the change in relative concentration of free and protein-bound NAD(P)H.This study suggests that the mean fuorescence lifetime of NAD(P)H might be a potential label-free indicator for differentiating leukemic myeloid cells from MNC.

Engagement of sialylated glycans with Siglec receptors on suppressive myeloid cells inhibits anticancer immunity via CCL2

The overexpression of sialic acids on glycans,called hypersialylation,is a common alteration found in cancer cells.Sialylated glycans can enhance immune evasion by interacting with sialic acid-binding immunoglobulin-like lectin(Siglec)receptors on tumorinfiltrating immune cells.Here,we investigated the effect of sialylated glycans and their interaction with Siglec receptors on myeloid-derived suppressor cells(MDSCs).We found that MDSCs derived from the blood of lung cancer patients and tumor-bearing mice strongly express inhibitory Siglec receptors and are highly sialylated.In murine cancer models of emergency myelopoiesis,Siglec-E knockout in myeloid cells resulted in prolonged survival and increased tumor infiltration of activated T cells.Targeting suppressive myeloid cells by blocking Siglec receptors or desialylation strongly reduced their suppressive potential.We further identified CCL2 as a mediator involved in T-cell suppression upon interaction between sialoglycans and Siglec receptors on MDSCs.Our results demonstrated that sialylated glycans inhibit anticancer immunity by modulating CCL2 expression.

ACAT1 deficiency in myeloid cells promotes glioblastoma progression by enhancing the accumulation of myeloid-derived suppressor cells

Glioblastoma(GBM)is a highly aggressive and lethal brain tumor with an immunosuppressive tumor microenvironment(TME).In this environment,myeloid cells,such as myeloid-derived suppressor cells(MDSCs),play a pivotal role in suppressing antitumor immunity.Lipometabolism is closely related to the function of myeloid cells.Here,our study reports that acetyl-CoA acetyltransferase 1(ACAT1),the key enzyme of fatty acid oxidation(FAO)and ketogenesis,is significantly downregulated in the MDSCs infiltrated in GBM patients.To investigate the effects of ACAT1 on myeloid cells,we generated mice with myeloid-specific(LyzM-cre)depletion of ACAT1.The results show that these mice exhibited a remarkable accumulation of MDSCs and increased tumor progression both ectopically and orthotopically.The mechanism behind this effect is elevated secretion of C-X-C motif ligand 1(CXCLI)of macrophages(Mo).Overall,our findings demonstrate that ACAT1 could serve as a promising drug target for GBM by regulating the function of MDSCs in the TME.

The triggering receptor expressed on myeloid cells 2-apolipoprotein E signaling pathway in diseases

Triggering receptor expressed on myeloid cells 2(TREM2)is a membrane receptor on myeloid cells and plays an important role in the body’s immune defense.Recently,TREM2 has received extensive attention from researchers,and its activity has been found in Alzheimer’s disease,neuroinflammation,and traumatic brain injury.The appearance of TREM2 is usually accompanied by changes in apolipoprotein E(ApoE),and there has been a lot of research into their structure,as well as the interaction mode and signal pathways involved in them.As two molecules with broad and important roles in the human body,understanding their correlation may provide therapeutic targets for certain diseases.In this article,we reviewed several diseases in which TREM2 and ApoE are synergistically involved in the development.We further discussed the positive or negative effects of the TREM2-ApoE pathway on nervous system immunity and inflammation.

Meningeal lymphatic vessel crosstalk with central nervous system immune cells in aging and neurodegenerative diseases

Meningeal lymphatic vessels form a relationship between the nervous system and periphery, which is relevant in both health and disease. Meningeal lymphatic vessels not only play a key role in the drainage of brain metabolites but also contribute to antigen delivery and immune cell activation. The advent of novel genomic technologies has enabled rapid progress in the characterization of myeloid and lymphoid cells and their interactions with meningeal lymphatic vessels within the central nervous system. In this review, we provide an overview of the multifaceted roles of meningeal lymphatic vessels within the context of the central nervous system immune network, highlighting recent discoveries on the immunological niche provided by meningeal lymphatic vessels. Furthermore, we delve into the mechanisms of crosstalk between meningeal lymphatic vessels and immune cells in the central nervous system under both homeostatic conditions and neurodegenerative diseases, discussing how these interactions shape the pathological outcomes. Regulation of meningeal lymphatic vessel function and structure can influence lymphatic drainage, cerebrospinal fluid-borne immune modulators, and immune cell populations in aging and neurodegenerative disorders, thereby playing a key role in shaping meningeal and brain parenchyma immunity.

Comprehensive Understanding of Immune Cells in The Pathogenesis of Non-alcoholic Fatty Liver Disease

Non-alcoholic fatty liver disease(NAFLD)is the most common chronic liver disease,defined by several phases,ranging from benign fat accumulation to non-alcoholic steatohepatitis(NASH),which can lead to liver cancer and cirrhosis.Although NAFLD is a disease of disordered metabolism,it also involves several immune cell-mediated inflammatory processes,either promoting and/or suppressing hepatocyte inflammation through the secretion of pro-inflammatory and/or anti-inflammatory factors to influence the NAFLD process.However,the underlying disease mechanism and the role of immune cells in NAFLD are still under investigation,leaving many open-ended questions.In this review,we presented the recent concepts about the interplay of immune cells in the onset and pathogenesis of NAFLD.We also highlighted the specific non-immune cells exhibiting immunological properties of therapeutic significance in NAFLD.We hope that this review will help guide the development of future NAFLD therapeutics.

Soluble triggering receptor expressed on myeloid cell-1 （sTREM- 1）： a potential biomarker for the diagnosis of infectious diseases

Sensitive and useful biomarkers for the diagnosis and prognosis of infectious diseases have been widely developed. An example of these biomarkers is triggering receptor expressed on myeloid cell-1 （TREM-1）, which is a cell surface receptor expressed on monocytes/macrophages and neutrophils. TREM-1 amplifies inflammation by activating the TREM-1/DAP12 pathway. This pathway is triggered by the interaction of TREM-1 with ligands or stimulation by bacterial lipopolysaccharide. Consequently, pro-inflammatory cytokines and chemokines are secreted. Soluble TREM-1 （sTREM-1） is a special form of TREM-1 that can be directly tested in human body fluids and well-known biomarker for infectious diseases, sTREM-1 level can be potentially used for the early diagnosis and prognosis prediction of some infectious diseases, including infectious pleural effusion, lung infections, sepsis, bacterial meningitis, viral infections （e.g., Crimean Congo hemorrhagic fever and dengue fever）, fungal infections （e.g., Aspergillus infection）, and burn-related infections, sTREM-1 is a more sensitive and specific biomarker than traditional indices, such as C-reactive protein and procalcitonin levels, for these infectious diseases. Therefore, sTREM-1 is a feasible biomarker for the targeted therapy and rapid and early diagnosis of infectious diseases.

Conditional Knockout of Src Homology 2 Domain-containing Protein Tyrosine Phosphatase-2 in Myeloid Cells Attenuates Renal Fibrosis after Unilateral Ureter Obstruction

Background：Src homology 2 domain-containing protein tyrosine phosphatase-2 （SHP-2） is a kind of intracellular protein tyrosine phosphatase.Studies have revealed its roles in various disease,however,whether SHP-2 involves in renal fibrosis remains unclear.The aim of this study was to explore the roles of myeloid cells SHP-2 in renal interstitial fibrosis.Methods：Myeloid cells SHP-2 gene was conditionally knocked-out （CKO） in mice using loxP-Cre system,and renal interstitial fibrosis was induced by unilateral ureter obstruction （UUO）.The total collagen deposition in the renal interstitium was assessed using picrosirius red stain.F4/80 immunostaing was used to evaluate macrophage infiltration in renal tubular interstitium.Quantitative real-time polymerase chain reaction and enzyme linked immunosorbent assay were used to analyze the production of cytokines in the kidney.Transferase-mediated dUTP nick-end labeling stain was used to assess the apoptotic renal tubular epithelial cells.Results：Src homology 2 domain-containing protein tyrosine phosphatase-2 gene CKO in myeloid cells significantly reduced collagen deposition in the renal interstitium after UUO.Macrophage infiltration was evidently decreased in renal tubular interstitium of SHP-2 CKO mice.Meanwhile,the production of pro-inflammatory cytokines was significantly suppressed in SHP-2 CKO mice.However,no significant difference was observed in the number of apoptotic renal tubular epithelial cells between wild-type and SHP-2 CKO mice.Conclusions：Our observations suggested that SHP-2 in myeloid cells plays a pivotal role in the pathogenesis of renal fibrosis,and that silencing of SHP-2 gene in myeloid cells may protect renal from inflammatory damage and prevent renal fibrosis after renal injury.

Role of Triggering Receptor Expressed on Myeloid Cell-1 Expression in Mammalian Target of Rapamycin Modulation of CD8＋ T-cell Differentiation during the Immune Response to Invasive Pulmonary Aspergillosis

Background： Triggering receptor expressed on myeloid cell- 1 （TREM- 1） may play a vital role in mammalian target ofrapamycin （mTOR） modulation ofCD8＋ T-cell differentiation through the transcription factors T-box expressed in T-cells and eomesodermin during the immune response to invasive pulmonary aspergillosis （IPA）. This study aimed to investigate whether the roTOR signaling pathway modulates the proliferation and differentiation of CD8＋ T-cells during the immune response to I PA and the role TREM-1 plays in this process. Methods： Cyclophosphamide （CTX） was injected intraperitoneally, and Asl？e;gillus.[mnigams spore suspension was inoculated intranasally to establish the immunosuppressed IPA mouse model. After inoculation, rapamycin （2 mg-kg ·d -1） or interleukin （IL）-12 （5 μg/kg every other day） was given for 7 days. The number of CD8＋ effector memory T-cells （Tern）, expression of interferon （IFN）-y, roTOR, and ribosomal protein $6 kinase （S6K）, and the levels of IL-6, IL- 10, galactomannan （GM）, and soluble TREM- 1 （sTREM-I） were measured. Results： Viable A. fumigatus was cultured from the lung tissue of the inoculated mice. Histological examination indicated greater inflammation, hemorrhage, and lung tissue injury in both IPA and CTX ＋ IPA mice groups. The expression of mTOR and S6K was significantly increased in the CTX ＋ IPA ＋ I L- 12 group compared with the control, I PA （P = 0.01 ; P - 0.001 ）, and CTX ＋ 1PA （P = 0.034; P = 0.032） groups, but significantly decreased in the CTX ＋ IPA ＋ RAPA group （P 〈 0.001 ）. Compared with the CTX ＋ IPA group, the proportion of Tern, expression of IFN-y, and the level ofsTREM-I were significantly higher after IL-12 treatment （P = 0.024, P = 0.032, and P = 0.017, respectively）, and the opposite results were observed when the roTOR pathway was blocked by rapamycin （P 〈 0.001）. Compared with the CTX ＋ I PA and CTX ＋ I PA ＋ RAPA groups, IL-12 treatment increased IL-6 and downregulated IL- 10 as well as G M, which strengthened the immune response to the IPA infection. Conclusions： mTOR modulates CD8＋ T-cell differentiation during the immune response to IPA. TREM-1 may play a vital role in signal transduction between mTOR and the downstream immune response.

LILRB4, an immune checkpoint on myeloid cells

Leukocyte immunoglobulin-like receptor B4(LILRB4)is an inhibitory receptor in the LILR family mainly expressed on normal and malignant human cells of myeloid origin.By binding to ligands,LILRB4 is activated and subsequently recruits adaptors to cytoplasmic immunoreceptor tyrosine inhibitory motifs to initiate different signaling cascades,thus playing an important role in physiological and pathological conditions,including autoimmune diseases,microbial infections,and cancers.In normal myeloid cells,LILRB4 regulates intrinsic cell activation and differentiation.In disease-associated or malignant myeloid cells,LILRB4 is significantly correlated with disease severity or patient survival and suppresses T cells,thereby participating in the pathogenesis of various diseases.In summary,LILRB4 functions as an immune checkpoint on myeloid cells and may be a promising therapeutic target for various human immune diseases,especially for cancer immunotherapy.

Soluble Triggering Receptor Expressed on Myeloid Cells-1 and Inflammatory Markers in Colorectal Cancer Surgery： A Prospective Cohort Study

Background： Major abdominal surgery, including colorectal cancer （CRC） surgery, leads to systemic inflammatory response syndrome that can be detected and monitored with inflammatory markers testing. The aims of the study were to evaluate the usefulness of soluble triggering receptor expressed on myeloid cells-l （sTREM-1 ）, interleukin-6 （IL-6）, procalcitonin （PCT）, and C-reactive protein （CRP） in following the inflammatory response in CRC surgery and postoperative period, as well as to determine if duration of the surgery and the time that the colon has been opened during the surgery （open colon time [OCT]） refect a larger surgical stress through inflammatory markers rise. Methods： The study included 20 patients who underwent CRC surgery and 19 healthy volunteers from June 2011 to September 2012. We determined inflammatory markers 1 day before surgery （T0）, 24 h （T1）, 48 h （T2）, and 7 days after the surgery （T3）. All statistical analyses were calculated using MedCalc Statistical Software version 14.8.1 （MedCalc Software bvba, Ostend, Belgium）. Results： Concentrations ofCRP, PCT, and I L-6 in all measurement times were statistically different and sTREM- 1 did not yield statistical significance. A weak positive correlation was/bund between l L-6 in T 1 and T2 with the duration of the surgery （T 1 ： r= 0.4060, P 〈 0.0001 ; T2：r =0.3430, P〈0.0001）andOCT（T1：r= 0.3640, P〈0.0001,T2：r=0.3430, P〈0.0001）.AweakpositivecorrelationbetweenCRP in T2 and OCT （r = 0.4210, P 〈 0.0001 ） was also found. The interconnectivity of tested parameters showed a weak positive correlation between CRP and IL-6 in T1 （r= 0.3680; P 〈 0.0001 ）, moderate positive correlation in T2 （r = 0.6770; P 〈 0.0001）, and a strong positive correlation in T3 （r = 0.8651; P 〈 0.0001）. Conclusions： CRP, IL-6, and PCT were shown to be reliable for postoperative monitoring. Simultaneous determination of CRP and IL-6 might not be useful as they follow similar kinetics, sTREM- 1 might not be useful in CRC postoperative monitoring.

Expressions of farnesoid X receptor and myeloid cell leukemia sequence 1 protein are associated with poor prognosis in patients with gallbladder cancer

Background Farnesoid X receptor （FXR） regulates tumorigenesis, but its clinical significance in gallbladder cancer （GBC） remains unclear. This study investigated its clinical and prognostic significance in GBC patients, as well as its association with the anti-apoptotic protein, myeloid cell leukemia sequence 1 （MCL1） protein. Methods FXR and MCL1 expression in 42 primary GBC and 15 normal gallbladder tissues were analyzed by immunohistochemistry. The patients and samples were collected from Ren Ji Hospital from January 2005 to December 2010. Their association with clinicopathologic factors and prognosis, as well as the correlation between FXR and MCL1 protein expression were analyzed by statistical analyses. Results Compared with normal gallbladder tissues, FXR expression was decreased and MCL1 expression was increased in GBC, during progression of tumor node metastasis （TNM） stage. The Kaplan-Meier survival analysis showed that FXR low-expression and MCL1 over-expression were significantly associated with overall poor survival. Furthermore, multivariate analysis showed that FXR and MCL1 are both prognostic factors for GBC patients. FXR low-expression was significantly correlated with MCL1 over-expression. Conclusion FXR might be a new molecular marker to predict the prognosis of patients with GBC and a novel therapeutic target. Chin Med J 2014;127 （14）： 2637-2642

Establishment of induced pluripotent stem cells from aged mice using bone marrow-derived myeloid cells

If induced pluripotent stem(iPS)cells are to be used to treat damaged tissues or repair organs in elderly patients,it will be necessary to establish iPS cells from their tissues.To determine the feasibility of using this technology with elderly patients,we asked if it was indeed possible to establish iPS cells from the bone marrow(BM)of aged mice.BM cells from aged C57BL/6 mice carrying the green fluorescence protein(GFP)gene were cultured with granulocyte macrophage-colony stimulating factor(GM-CSF)for 4 days.Four factors(Oct3/4,Sox2,Klf4 and c-Myc)were introduced into the BM-derived myeloid(BM-M)cells.The efficiency of generating iPS cells from aged BM cultured in GM-CSF was low.However,we succeeded in obtaining BM-M-iPS cells from aged C57BL/6 mice,which carried GFP.Our BM-M-iPS cells expressed SSEA-1 and Pou5f1 and were positive for alkaline phosphatase staining.The iPS cells did make teratoma with three germ layers following injection into syngeneic C57BL/6 mice,and can be differentiated to three germ layers in vitro.By co-culturing with OP9,the BM-M-iPS cells can be differentiated to the myeloid lineage.The differentiated BM-M-iPS cells proliferated well in the presence of GM-CSF,and lost expression of Nanog and Pou5f1,at least in part,due to methylation of their promoters.On the contrary,Tnf and Il1b gene expression was upregulated and their promoters were hypomethylated.

Pik3c3 deficiency in myeloid cells imparts partial resistance to experimental autoimmune encephalomyelitis associated with reduced IL-1βproduction

The PIK3C3/VPS34 subunit of the class III phosphatidylinositol 3-kinase(PtdIns3K)complex plays a role in both canonical and noncanonical autophagy,key processes that control immune-cell responsiveness to a variety of stimuli.Our previous studies found that PIK3C3 is a critical regulator that controls the development,homeostasis,and function of dendritic and T cells.In this study,we investigated the role of PIK3C3 in myeloid cell biology using myeloid cell-specific Pik3c3-deficient mice.We found that Pik3c3-deficient macrophages express increased surface levels of major histocompatibility complex(MHC)class I and class II molecules.In addition,myeloid cell-specific Pik3c3 ablation in mice caused a partial impairment in the homeostatic maintenance of macrophages expressing the apoptotic cell uptake receptor TIM-4.Pik3c3 deficiency caused phenotypic changes in myeloid cells that were dependent on the early machinery(initiation/nucleation)of the classical autophagy pathway.Consequently,myeloid cell-specific Pik3c3-deficient animals showed significantly reduced severity of experimental autoimmune encephalomyelitis(EAE),a primarily CD4^(^(+))T-cell-mediated mouse model of multiple sclerosis(MS).This disease protection was associated with reduced accumulation of myelin-specific CD4^(^(+))T cells in the central nervous system and decreased myeloid cell IL-1βproduction.Further,administration of SAR405,a selective PIK3C3 inhibitor,delayed disease progression.Collectively,our studies establish PIK3C3 as an important regulator of macrophage functions and myeloid cell-mediated regulation of EAE.Our findings also have important implications for the development of small-molecule inhibitors of PIK3C3 as therapeutic modulators of MS and other autoimmune diseases.

Degradation-resistant implanted biomaterials establish an immunosuppressive microenvironment that induces T cell exhaustion by recruiting myeloid cells

Implanted biomaterials have transformed healthcare and the treatment of injury and disease,but their infuence on the local immune landscape remains unclear.Here we discovered that degradation-resistant titanium-based implants establish an immunosuppressive microenvironment by recruiting myeloid cells,including monocytes,macrophages,neutrophils,and myeloid-lineage dendritic cells.Unlike normal tissues,the tissues nearby implants exhibit an chronic inflamed and immunosuppressive status characterised by myeloid-rich,T cell-exhaustion gene signature by single-cell RNA sequencing.Vitamin C treatment provides an effective strategy to rescue the immunosuppressive microenvironment,which can be used as a regular supplement to reduce the risk of malignant cell survival around the implants.