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。     

Recent insights into the characteristics and role of peritoneal macrophages from ascites of cirrhotic patients

Macrophages are a diverse myeloid cell population involved in innate and adaptive immune responses, embryonic development, wound repair, and regulation of tissue homeostasis. These cells link the innate and adaptive immunities and are crucial in the development and sustainment of various inflammatory diseases. Macrophages are tissue-resident cells in steady-state conditions; however, they are also recruited from blood monocytes after local pathogen invasion or tissue injury. Peritoneal macrophages vary based on their cell complexity, phenotype, and functional capabilities. These cells regulate inflammation and control bacterial infections in the ascites of decompensated cirrhotic patients. Our recent work reported several phenotypic and functional characteristics of these cells under both healthy and pathological conditions. A direct association between cell size, CD14/CD16 expression, intracellular level of GATA-6, and expression of CD206 and HLA-DR activation/maturation markers, indicate that the large peritoneal macrophage CD14^highCD16^high subset constitutes the mature phenotype of human resident peritoneal macrophages during homeostasis. Moreover, elevated expression of CD14/CD16 is related to the phagocytic capacity. The novel large CD14^highCD16^high peritoneal subpopulation is increased in the ascites of cirrhotic patients and is highly sensitive to lipopolysaccharide (LPS)-induced activation, thereby exhibiting features of inflammatory priming. Thus, phosphorylation of ERK1/2, PKB/Akt, and c-Jun is remarkably increased in response to LPS in vitro, whereas that of p38 MAPK is reduced compared with the monocyte-derived macrophages from the blood of healthy controls. Furthermore, in vitro activated monocyte-derived macrophages from ascites of cirrhotic patients secreted significantly higher levels of IL-6, IL-10, and TNF-α and lower amounts of IL-1β and IL-12 than the corresponding cells from healthy donor’s blood. Based on these results, other authors have recently reported that the surface expression level of CD206 can be used to identify mature, resident, inflammatory peritoneal macrophages in patients with cirrhosis. Soluble CD206 is released from activated large peritoneal macrophages, and increased concentrations in patients with cirrhosis and spontaneous bacterial peritonitis (SBP) indicate reduced odds of survival for 90 d. Hence, the level of soluble CD206 in ascites might be used to identify patients with SBP at risk of death. In conclusion, peritoneal macrophages present in ascites of cirrhotic patients display multiple phenotypic modifications characterized by reduced ratio of cells expressing several membrane markers, together with an increase in the ratios of complex and intermediate subpopulations and a decrease in the classic-like subset. These modifications may lead to the identification of novel pharmaceutical targets for prevention and treatment of hepatic damage.     

肿瘤相关巨噬细胞促进宫颈癌侵袭转移的机制研究

 目的 研究肿瘤相关巨噬细胞（tumor-associated macrophages, TAMs）对宫颈癌侵袭转移的影响及分子机制。方法 免疫组织化学法检测宫颈病变组织TAMs的浸润情况。对人单核巨噬细胞系THP1进行体外诱导分化，使其转化为M2型TAMs。取TAMs上清液刺激宫颈癌细胞SiHa和C33a。划痕法、Transwell法分别检测TAMs对宫颈癌细胞系迁移和侵袭能力的影响。Western blot检测刺激后宫颈癌细胞上皮间质转化进程及MMP-9的表达。结果 TAMs浸润数目与宫颈病变进展呈正相关。TAMs上清液刺激后，SiHa和C33a细胞出现间质样改变，而且迁移和侵袭能力显著增强（均P<0.5）。TAMs可下调E-Cadherin的表达，上调N-Cadherin、Vimentin及MMP-9的表达。结论 TAMs浸润与宫颈上皮恶性转化和进展密切相关，TAMs可能通过上调MMP-9的表达促进宫颈癌细胞侵袭转移。     

Toll样受体信号途径在RAW264.7细胞川崎病模型炎症反应中的作用及其机制研究

目的探讨Toll样受体(TLR)2信号途径在RAW264.7细胞川崎病模型炎症反应中的作用及其分子机制。方法构建TLR2-干扰小RNA(siRNA),合成2对特异性针对TLR2基因的siRNA序列,并用于转染RAW264.7细胞。实时荧光聚合酶链反应和蛋白质印迹法检测TLR2-siRNA对干酪乳杆菌细胞壁提取物(LCWE)诱导RAW264.7细胞p38促分裂原活化的蛋白激酶(MAPK)、基质多属蛋白酶(MMP)-9表达的影响,乳胶增强免疫比浊定量法检测培养基中高敏C反应蛋白(hsCRP)水平的变化;检测p38MAPK抑制剂SB 203580对LCWE诱导RAW264.7细胞MMP-9表达的影响。结果与对照组相比,LCWE刺激组p-p38MAPK、MMP-9的表达及hs-CRP水平都有明显升高,差异具有统计学意义(P<0.05),与LCWE刺激组相比,TLR2-siRNA+LCWE刺激组的p-p38MAPK、MMP-9表达和hs CRP水平降低,差异具有统计学意义(P<0.05)。与LCWE刺激组相比,SB203580+LCWE刺激组的MMP-9表达减轻,差异具有统计学意义(P<0.05)。结论 TLR2-siRNA可以有效减轻LCWE刺激造成的p-p38MAPK、MMP-9、hs-CRP表达升高。p38MAPK抑制剂SB203580对LCWE诱导RAW264.7细胞MMP-9表达有抑制作用。TLR2参与了RAW264.7细胞川崎病模型中的炎症反应,其机制可能与p38MAPK途径有关。     

Arginase and its mechanisms in Leishmania persistence

Leishmaniasis is a neglected infectious disease with clinical presentations ranging from asymptomatic or mild symptoms to chronic infection and eventual death. The mechanisms of disease susceptibility and pathology have been extensively studied, but there are no steadfast rules regarding leishmaniasis. A Th1 response is usually associated with infection control, while a predominant Th2 response is detrimental to the patient. In this scenario, the enzymes arginase and inducible nitric oxide synthase represent two possible pathways of immune response. While the former contributes to parasite replication, the latter is crucial for its control. In the present review, we collected study results that associate arginase expression in patients and in experimental models with disease susceptibility/chronicity and show some proposed mechanisms that explain the role of arginase in maintaining Leishmania infection, including polyamine and thiol synthesis, tissue-resident macrophage (TRM) proliferation and activation and T-cell suppression and exhaustion.     

以机油为分散剂高温热解人发合成新型碳点及其生物效应研究

目的以机油为分散剂高温热解人发后发现新型纳米碳点,通过动物实验评价了该碳点的生物效应。方法利用高温热解法对人发和机油进行炭化,对炭化产物进行萃取、过滤分离和透析得到一种具有水溶性的新型物质碳点,并命名为JYRF-CDs。利用透射电子显微镜(TEM)、高分辨透射电子显微镜(HR-TEM)、紫外-可见分光光谱、傅里叶变换红外光谱、荧光光谱、X射线光电子能谱分析(XPS)等多种方法对JYRF-CDs进行表征,利用小鼠单核巨噬细胞RAW264.7细胞进行CCK-8毒性实验来评价JYRF-CDs的安全性,并通过小鼠耳肿胀实验和小鼠醋酸扭体实验对JYRF-CDs的生物效应进行评价。结果 JYRF-CDs外形为类球形,粒径均匀分布在1.8～3.6 nm,晶格间距为0.219 7 nm。细胞毒性实验结果显示,JYRF-CDs具有低毒性,动物实验结果表明JYRF-CDs具有良好的抗炎和镇痛作用。结论首次以机油为分散剂高温热解人发后发现一种全新的碳点JYRF-CDs,以JYRF-CDs为突破口,更加明确阐释以机油为分散剂高温热解人发后炭化产物具有生物效应的物质基础,为纳米类成分的研究提供了一种新方法。     

Breast cancer cells promote CD169+ macrophage-associated immunosuppression through JAK2-mediated PD-L1 upregulation on macrophages

Macrophages are recognized as one of the major cell types in tumor microenvironment, and macrophage infiltration has been predominantly associated with poor prognosis among patients with breast cancer. Using the murine models of triple-negative breast cancer in CD169-DTR mice, we found that CD169⁺ macrophages support tumor growth and metastasis. CD169⁺ macrophage depletion resulted in increased accumulation of CD8⁺ T cells within tumor, and produced significant expansion of CD8⁺ T cells in circulation and spleen. In addition, we observed that CD169⁺ macrophage depletion alleviated tumor-induced splenomegaly in mice, but had no improvement in bone loss and repression of bone marrow erythropoiesis in tumor-bearing mice. Cancer cells and tumor associated macrophages exploit the upregulation of the immunosuppressive protein PD-L1 to subvert T cell-mediated immune surveillance. Within the tumor microenvironment, our understanding of the regulation of PD-L1 protein expression is limited. We showed that there was a 5-fold higher relative expression of PD-L1 on macrophages as compared with 4T1 tumor cells; coculture of macrophages with 4T1 cells augmented PD-L1 levels on macrophages, but did not upregulate the expression of PD-L1 on 4T1 cells. JAK2/STAT3 signaling pathway was activated in macrophages after coculture, and we further identified the JAK2 as a critical regulator of PD-L1 expression in macrophages during coculture with 4T1 cells. Collectively, our data reveal that breast cancer cells and CD169⁺ macrophages exhibit bidirectional interactions that play a critical role in tumor progression, and inhibition of JAK2 signaling pathway in CD169⁺ macrophages may be potential strategy to block tumor microenvironment-derived immune escape.     

Toxicity of ZnO nanoparticles (NPs) to THP-1 macrophages: interactions with saturated or unsaturated free fatty acids

In a biological microenvironment, free fatty acids (FFA) as ubiquitous biological molecules might interact with nanoparticles (NPs) and consequently change the toxicological responses. However, whether the chemical structures of FFA could influence their interactions with NPs remain unknown. This study investigated the interactions between ZnO NPs and saturated or unsaturated FFA (complexed to BSA), namely stearic acid (SA, C18:0), oleic acid (OA, C18:1), and α-linolenic acid (ALA, C18:3). It was shown that BSA, SA, OA, and ALA increased the atomic force microscope (AFM) heights as well the polydispersity index (PDI) of ZnO NPs. BSA modestly protected THP-1 macrophages from ZnO NP exposure, whereas OA and ALA led to relatively less cyto-protective effects of BSA. Moreover, only co-exposure to ZnO NPs and SA significantly promoted the release of interleukin-8. BSA, SA, OA, and ALA equally changed intracellular ROS and Zn ions associated with ZnO exposure, but co-exposure to ZnO NPs and OA/ALA particularly activated the expression of endoplasmic reticulum stress-apoptosis genes. In combination, these results showed that FFA could influence the colloidal aspects and toxicological signaling pathway of ZnO NPs, which is dependent on the number of unsaturated bonds of FFA.