外泌体在肿瘤细胞与TAMs之间相互作用中“桥梁”和调控作用的研究进展北大核心

外泌体是一类直径为30~100 nm的圆盘囊泡,其内包含许多组分,诸如复杂RNA和蛋白质等,主要参与细胞间的信号转导。肿瘤相关巨噬细胞(tumor-associated macrophages,TAMs)是肿瘤微环境中普遍存在的巨噬细胞,通过对肿瘤生长、免疫逃逸、侵袭和转移、耐药性等多方面的作用影响肿瘤进程。外泌体在肿瘤相关巨噬细胞的招募、极化及抗肿瘤免疫调控等方面发挥着重要的调节功能。同时,TAMs以外泌体为媒介作用于肿瘤细胞,从而构成了外泌体、TAMs与肿瘤细胞之间相互作用的调控通路。综上所述,本文旨在阐明肿瘤细胞与TAMs之间,以外泌体为“桥梁”相互影响的潜在机制,以及靶向肿瘤细胞和TAMs来源的外泌体在恶性肿瘤治疗中的展望。     

银椴苷对激活的原代小胶质细胞极化及神经炎症的影响＊

目的 银椴苷（Tiliroside，Tle）是一种天然产物，存在于结香花（Edgeworthia chrysantha Lindl.）、金英（Galphimia gracilis）、和Phlomoides spectabilis等生物体中。本文主要探讨Tle对脂多糖（lipopolysaccharide，LPS）激活的原代小胶质细胞的M1/M2极化的转化作用及其抑制神经炎症反应的影响。方法 MTT检测各浓度Tle对LPS激活的原代小胶质细胞活性的影响；Griess试剂检测LPS激活的原代小胶质细胞的一氧化氮（Nitric oxide，NO）产物生成量的影响；ELISA法检测Tle对LPS激活的原代小胶质细胞的肿瘤坏死因子-α（Tumor necrosis factor-α，TNF-α）、白细胞介素-6（Interleukin-6，IL-6）蛋白水平的影响；qRT-PCR检测Tle对LPS激活的原代小胶质细胞的精氨酸酶-1（Arginase-1，Arg-1）和胰岛素生长因子-1（Insulin growth factor-1，IGF-1）的mRNA表达水平。结果 Tle（0-80 μmol·L^-1）对LPS激活的原代小胶质细胞活性没有明显影响；Tle显著降低LPS激活的原代小胶质细胞亚硝酸盐的含量，并对炎症因子IL-6、TNF-α蛋白水平有显著抑制作用，同时显著提高抗炎因子Arg-1和IGF-1的mRNA水平。结论 Tle能够通过促进LPS激活的原代小胶质细胞M1型向M2型转化从而抑制神经炎症反应，因此Tle具有一定的神经保护作用。     

Excess salt intake promotes M1 microglia polarization via a p38/MAPK/AR-dependent pathway after cerebral ischemia in mice

A high salt diet (HSD) is among the most important risk factors for many diseases. One mechanism by which HSD aggravates cerebral ischemic injury is independent of blood pressure changes. The direct role of HSD in inflammation after cerebral ischemia is unclear. In this research, after twenty-one days of being fed a high salt diet, permanent focal ischemia was induced in mice via operation. At 12 h and 1, 3 and 5 days postischemia, the effects of HSD on the lesion volume, microglia polarization, aldose reductase (AR) expression, and inflammatory processes were analyzed. We report that in mice, surplus dietary salt promotes inflammation and increases the activation of classical lipopolysaccharide (LPS)-induced microglia/macrophages (M1). This effect depends on the expression of the AR protein in activated microglia after permanent middle cerebral artery ligation (pMCAL) in HSD mice. The administration of either the AR inhibitor Epalrestat or a p38-neutralizing antibody blocked the polarization of microglia and alleviated stroke injury.In conclusion, HSD promotes polarization in pro-inflammatory M1 microglia by upregulating the expression of the AR protein via p38/MAPK, thereby exacerbating the development of ischemia stroke.     

Kellerin alleviates cognitive impairment in mice after ischemic stroke by multiple mechanisms

Ischemic stroke is a global disease with high disability and mortality rates. Cognitive impairment is one of the major clinical features of ischemic stroke, and microglia‐mediated inflammation has been shown to be an important contributor to the pathogenesis of ischemic stroke. Kellerin, extracted from Ferula sinkiangensis, was previously shown to inhibit microglial activation and exert a strong anti‐neuroinflammatory effect. However, there is no report of the potential therapeutic effect of kellerin on ischemic stroke by targeting microglial cells. In this study, we wanted to examine the effects of kellerin on ischemic stroke in the bilateral common carotid artery occlusion (BCCAO) model and the lipopolysaccharide (LPS)‐activated microglia model. We found that kellerin alleviated cognitive impairment, decreased neuronal loss, suppressed microglial activation, and transformed microglia from the pro‐inflammatory M1 phenotype to the anti‐inflammatory M2 phenotype in BCCAO mice. Moreover, in in vitro studies, we found that kellerin regulated microglial polarization and inhibited the NLRP3 and MAPK signaling pathways after LPS treatment. These findings provide a new understanding of the function of kellerin in ischemic stroke, and suggest that kellerin could be a potential therapeutic agent for the treatment of ischemic stroke.     

Interactions between rat cortico-striatal slice cultures and neutrophil-like HL60 cells under thrombin challenge: Toward elucidation of pathological events in intracerebral hemorrhage

Neutrophils constitute the major population of infiltrating leukocytes after stroke including intracerebral hemorrhage (ICH), and these cells may exhibit pro-inflammatory and anti-inflammatory phenotypes depending on the external stimuli. Here we constructed an experimental system to evaluate how the properties of neutrophils were influenced by the injured brain tissues. HL60 cells differentiated into neutrophils were added to the culture medium of neonatal rat cortico-striatal slices maintained at liquid–air interface. Thrombin was applied to the cultures to mimic the pathogenic events associated with ICH. HL60 cells responded to thrombin by increasing mRNA expression of pro-inflammatory IL-1β and anti-inflammatory IL-10 with a different time course. Co-presence of cortico-striatal slice cultures significantly enhanced IL-1β mRNA expression, whereas attenuated IL-10 mRNA expression, in HL60 cells. Toll-like receptor 4 (TLR4) agonist lipopolysaccharide synergistically enhanced IL-1β mRNA expression with thrombin, and TLR4 inhibitor TAK-242 abolished thrombin-induced IL-1β mRNA expression in the presence of slice cultures. On the other hand, thrombin-induced cell death in cortico-striatal cultures was attenuated by the presence of HL60 cells. This experimental system may provide a unique platform to elucidate complex cell-to-tissue interactions during ICH pathogenesis.     

非编码RNA调控Kupffer细胞极化干预肝癌癌前病变恶变的研究进展

肝癌是临床常见消化道恶性肿瘤，其发生呈现多阶段性：肝炎-肝硬化-肝癌癌前病变-肝癌。其中肝癌癌前病变恶变成为肝癌的机制尚不完全明确，可能受Kupper细胞极化方向的影响。Kupffer细胞是肝脏中特殊的单核巨噬细胞，可在不同的微环境中分化出不同的表型即M1型和M2型，表现出不同的功能，在肝癌中发挥抑癌或促癌的作用。研究发现多种miRNA、LncRNA可通过靶向干预Kupffer极化相关蛋白进而调控其极化方向，从而影响肝癌癌前病变的发生、发展及恶变，降低肝癌的发生率。本文主要阐述上述研究的前沿进展，为肝癌的防治提供参考。     

活性多酚化合物LM49对RAW264.7巨噬细胞极化的影响

目的初步探讨LM49(2,4'三羟基-5,2'-二溴二苯甲酮)对脂多糖(LPS)联合干扰素γ(IFN-γ)诱导的小鼠单核巨噬细胞(RAW264.7)M1/M2极化的影响及其调控机制。方法四曱基偶氮唑蓝(MTT)法测定LM49对细胞活力的影响;流式细胞术、实时荧光定量聚合酶链反应(PCR)和Westem-blot法测定LM49(5,10,20μmol·L^-1)与LPS/INF-γ共同作用于RAW264.7细胞后,巨噬细胞亚型标志物的表达情况及对核因子(NF)-κB和JAK/STAT信号通路的影响。结果与LPS/INF-γ造模组相比,LM49显著抑制CD16/32^+细胞数及诱导型一氧化氮合酶(iNOS)、白细胞介素(IL)4和肿瘤坏死因子(TNF)-αmRNA的表达,升高CD206^+细胞数及Arg-1和IL-10 mRNA的表达,且降低巨噬细胞M1/M2的比值;Westem-blot法验证LM49可显著降低TLR4、Myd88、NF-κB和STAT1蛋白的表达量,同时抑制p-JAK2和p-STATl蛋白磷酸化水平。结论LM49通过抑制TLR4-Myd88-NF-κB和JAK2-STAT1信号通路,抑制巨噬细胞Ml型极化及促进巨噬细胞M2型极化,调节巨噬细胞M1/M2的平衡。     

Profilin 1 knockdown prevents ischemic brain damage by promoting M2 microglial polarization associated with the RhoA/ROCK pathway

Microglial polarization to the anti-inflammatory M2 phenotype is essential in resolving neuroinflammation, making it a promising therapeutic strategy for stroke intervention. The actin cytoskeleton is known to be important for the physiological functions of microglia, including migration and phagocytosis. Profilin 1 (PFN1), an actin-binding protein, is involved in the dynamic transformation and reorganization of actin. However, the role of PFN1 in microglial polarization and ischemia/reperfusion injury is unclear. The role of PFN1 on microglial polarization was examined in vitro in BV2 microglial cells subjected to oxygen-glucose deprivation/reoxygenation (OGDR) and in vivo in male mice after transient middle cerebral artery occlusion (MCAO). Knockdown of PFN1 inhibited M1 microglial polarization and promoted M2 microglia polarization 48 hr after OGDR stimulation in BV2 cells and 7 days after MCAO-induced injury in male mice. RhoA/ROCK pathway was involved in the regulation of PFN1 during microglial polarization. Knockdown of PFN1 also significantly attenuated brain infarcts and edema, improved cerebral blood flow and neurological deficits in MCAO-injured mice. Inhibition of PFN1 effectively protected the brain against ischemia/reperfusion injuries by promoting M2 microglial polarization in vitro and in vivo.     

A novel chrysin thiazole derivative polarizes macrophages to an M1 phenotype via targeting TLR4

Tumor-associated macrophages (TAMs) are an important cause of tumorigenesis and tumor development. M2 macrophages can promote tumor growth while M1 macrophages kill tumor cells, therefore, polarizing macrophages to achieve a functional M1 phenotype could effectively play its anti-tumor role. In the current study, we synthesized a novel chrysin derivative which is termed as ChR-TD. And we found ChR-TD might be a ligand of TLR4 that polarized the TAMs towards M1 phenotype and played its anti-tumor role. Further study indicated that ChR-TD reprogrammed the macrophages into an M1 phenotype via TLR4 activation. Moreover, ChR-TD activated TLR4/NF-κB signaling pathway and promoted the NF-κB/p65 translocated into the nuclear, leading to the activation of NF-κB and proinflammatory cytokines release. In addition, type I interferon signaling was also activated by ChR-TD, leading to the expressions of IFN-α and IFN-β and its targeted genes NOS2, MCP-1 and IP-10 were significantly increased in macrophages. Importantly, these effects were disturbed in TLR4^−/− macrophages, which are constructed by using CRISPR/Cas9 system. And the molecule docking simulation further indicated that ChR-TD could bind to TLR4 and might be a ligand of TLR4. Hence, these findings suggested that ChR-TD might be a ligand of TLR4 and can be used as a potential lead compound for tumors treatment.