Interactions between cancer cells and immune cells drive transitions to mesenchymal-like states in glioblastoma

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Ethanol upregulates the P2X7 purinergic receptor in human macrophages

Alcohol consumption is considered to be the third leading cause of death in the United States. In addition to its direct toxicity, ethanol has two contrasting effects on the immune system: the nucleotide oligomerization domain‐like receptor pyrin domain‐containing‐3 (NLRP3) inflammasome is inhibited by acute ethanol exposure but activated by chronic ethanol exposure. Purinergic receptors (especially the P2X7 receptor) are able to activate the NLRP3 inflammasome and are involved in many ethanol‐related diseases (such as gout, pulmonary fibrosis, alcoholic steatohepatitis, and certain cancers). We hypothesized that ethanol regulates purinergic receptors and thus modulates the NLRP3 inflammasome's activity. In experiments with monocyte‐derived macrophages, we found that interleukin (IL)‐1β secretion was inhibited after 7 h of exposure (but not 48 h of exposure) to ethanol. The disappearance of ethanol's inhibitory effect on IL‐1β secretion after 48 h was not mediated by the upregulated production of IL‐1β, IL‐1α, IL‐6 or the inflammasome components NLRP3, apoptosis‐associated speck‐like protein containing a caspase recruitment domain, and caspase 1. P2X7R expression was upregulated by ethanol, whereas expression of the P2X4 and P2X1 receptors was not. Taken as a whole, our results suggest that ethanol induces NLRP3 inflammasome activation by upregulating the P2X7 receptor. This observation might have revealed a new mechanism for inflammation in ethanol‐related diseases.     

艳山姜挥发油通过调控巨噬细胞CD36和ABCA1表达抑制泡沫细胞的形成

目的:观察艳山姜挥发油对氧化低密度脂蛋白(ox-LDL)诱导巨噬细胞转化为泡沫细胞的抑制作用,并探索其机制。方法:使用佛波酯(PMA,100μg·L^-1)诱导人白血病单核细胞(THP-1)24 h后形成巨噬细胞,实验分为4组,分别为空白组(无血清RPMI 1640),模型组(80 mg·L^-1ox-LDL),艳山姜挥发油低剂量组(80 mg·L^-1ox-LDL+4μg·L^-1艳山姜挥发油),艳山姜挥发油高剂量组(80 g·L^-1ox-LDL+20μg·L^-1艳山姜挥发油)。噻唑蓝(MTT)比色法检测艳山姜挥发油对巨噬细胞的活性的影响,蛋白免疫印迹法(Western blot)检测巨噬细胞中白细胞分化抗原36(CD36)和三磷酸腺苷结合盒转运体A1(ABCA1)的表达,酶联免疫吸附测定(ELISA)检测巨噬细胞内胆固醇酯含量,油红O染色法检测巨噬细胞中脂质小滴的含量。结果:艳山姜挥发油对巨噬细胞无毒性。与空白组比较,模型组的巨噬细胞内脂滴和胆固醇酯的含量显著增加(P<0.01),CD36蛋白表达显著上升(P<0.01),ABCA1蛋白表达无显著变化;与模型组比较,艳山姜挥发油显著抑制巨噬细胞中脂滴和胆固醇酯的含量(P<0.01),下调CD36的蛋白表达(P<0.01),上调ABCA1蛋白的表达(P<0.01),艳山姜挥发油可抑制巨噬细胞向泡沫细胞的转化。结论:艳山姜挥发油对ox-LDL诱导的巨噬细胞向泡沫细胞的形成具有抑制作用,该药理作用与艳山姜挥发油下调巨噬细胞CD36和上调ABCA1蛋白的表达有关。     

月华胶囊对耐多药结核分支杆菌感染自噬的影响

目的:以月华胶囊含药血清对耐多药结核分支杆菌感染小鼠单核巨噬细胞(RAW264.7)进行干预,探讨其对耐多药结核分支杆菌感染巨噬细胞自噬的作用及其机制。方法:低温冷冻干燥法制备月华胶囊,大鼠按3.02 g·kg^-1灌胃给药7 d,制备月华胶囊含药血清。体外培养RAW264.7,耐多药结核分子杆菌。以细胞增殖毒性检测试剂盒(CCK-8)检测含药血清对RAW264.7的增殖能力并选定其有效浓度。细胞分为模型组(10%胎牛血清);月华胶囊含药血清组(10%月华胶囊);自噬抑制剂3-甲基腺嘌呤(3-MA)+月华胶囊含药血清组(5 mg·L^-1的3-MA+10%月华胶囊含药血清);雷帕霉素(Rap)组(200 mg·L^-1的Rap+10%胎牛血清);正常组(10%胎牛血清)。除正常组外,各组细胞培养24 h后,按细胞-细菌1∶10感染4 h。透射电镜观察细胞自噬体的出现和形成;蛋白免疫印迹法(Western blot)检测细胞中I型微管相关蛋白轻链3(LC-3Ⅰ),Ⅱ型微管相关蛋白轻链3/I型微管相关蛋白1轻链3(LC-3Ⅱ/LC-3Ⅰ),Beclin-1蛋白的表达;间接免疫荧光染色法观察细胞中LC3B荧光颗粒、斑点及亮度;实时荧光定量聚合酶链式反应(Real-time PCR)检测细胞中LC3,Beclin-1 mRNA的表达。结果:与正常组比较,模型组细胞未见自噬体,细胞中LC-3Ⅱ,LC-3Ⅱ/LC-3Ⅰ,Beclin-1蛋白及LC3,Beclin-1 mRNA的相对表达量均无显著变化,细胞无荧光颗粒、斑点,无荧光亮度。与模型组比较,月华胶囊含药血清组及Rap组细胞,均可观察到自噬体的形成,细胞LC-3Ⅱ,LC-3Ⅱ/LC-3Ⅰ,Beclin-1蛋白及LC3,Beclin-1 mRNA的相对表达量值均明显升高(P<0.05),月华胶囊含药血清组细胞荧光颗粒和荧光斑点明显增多,Rap组细胞荧光颗粒和荧光斑点增多非常明显,荧光闪亮,可判定细胞荧光呈现为阳性;自噬抑制剂3-MA+月华胶囊含药血清组细胞未见自噬体,细胞中LC-3Ⅱ,LC-3Ⅱ/LC-3Ⅰ,Beclin-1蛋白及LC3,Beclin-1 mRNA的相对表达量值均无明显升高。结论:月华胶囊含药血清能使耐多药结核分支杆菌感染细胞产生自噬而发挥抗结核的免疫效应,其机制是通过调控自噬相关蛋白LC3,Beclin-1蛋白及其mRNA的表达水平,促使LC3-I趋向LC3-Ⅱ的转化加快而实现的。     

高糖饮食对小鼠真菌性角膜炎的影响

目的　探究高糖饮食对小鼠真菌性角膜炎的影响。方法　选取健康无眼疾的雄性C57BL/6J小鼠78只，随机分为高糖饮食组和模型对照组，每组36只，模型对照组给予正常饮用水，高糖饮食组给予含体积分数10%果糖溶液，每2 d测量两组小鼠体质量及血糖，10 d后建立真菌性角膜炎模型。造模后24 h、36 h、48 h、72 h、96 h、120 h、168 h裂隙灯显微镜下对角膜进行临床评分并拍照。处死小鼠后，取角膜组织进行HE染色和PAS染色；测定角膜内中性粒细胞和巨噬细胞浸润体积。利用酶联免疫吸附实验对小鼠角膜内的白细胞介素-1β含量进行测定。结果　造模后 0~14 d，两组小鼠体质量与血糖差异均无统计学意义（均为P>0.05）。造模后24 h、36 h、48 h、120 h、168 h，高糖饮食组小鼠角膜临床评分均明显高于模型对照组，差异均有统计学意义（均为P<0.05）。高糖饮食组小鼠角膜穿孔率79.5%，高于模型对照组的40.9%，差异有统计学意义（P=0.000）。造模后各时间点，高糖饮食组中性粒细胞浸润体积均高于模型对照组，差异均有统计学意义（均为P=0.000）。造模后72 h、96 h、120 h、168 h，高糖饮食组巨噬细胞浸润体积均高于模型对照组（均为P=0.000）。角膜组织病理学检查结果示，高糖饮食组炎症反应更重，角膜组织破坏更早且更为严重。造模后24 h、48 h高糖饮食组白细胞介素-1β含量均明显高于模型对照组（均为P<0.05）。结论　高糖饮食加重了真菌性角膜炎感染的严重程度，增强了中性粒细胞、巨噬细胞的趋化，促进了IL-1β的分泌。     

Quercetin protects against cisplatin‐induced acute kidney injury by inhibiting Mincle/Syk/NF‐κB signaling maintained macrophage inflammation

Acute kidney injury (AKI) with high incidence and mortality is the main cause of chronic kidney disease. Previous studies have indicated that quercetin, an abundant flavonoid in plants, exhibited renoprotective role in AKI. However, the underlying mechanism is largely unknown. In this study, we try to explore whether quercetin protects against AKI by inhibiting macrophage inflammation via regulation of Mincle/Syk/NF‐κB signaling. The results demonstrated that quercetin can significantly inhibit expression and secretion of IL‐1β, IL‐6, and TNF‐α in LPS‐induced bone marrow‐derived macrophages (BMDMs) and reduce activity of Mincle/Syk/NF‐κB signaling in vitro. We also found that quercetin can strongly reduce the concentration of serum creatinine, BUN, IL‐1β, IL‐6, and TNF‐α in cisplatin‐induced AKI model. Furthermore, quercetin down‐regulated protein levels of Mincle, phosphorylated Syk and NF‐κB in kidney macrophages of AKI, as well as inhibited M1, up‐regulated M2 macrophage activity. Notably, the down‐regulation of LPS‐induced inflammation by quercetin was reversed after adding TDB (an agonist of Mincle) in BMDMs, suggesting that quercetin suppresses macrophage inflammation may mainly through inhibiting Mincle and its downstream signaling. In summary, these findings clarified a new mechanism of quercetin improving AKI‐induced kidney inflammation and injury, which provides a new drug option for the clinical treatment of AKI.     

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。     

Niche-Specific Reprogramming of Epigenetic Landscapes Drives Myeloid Cell Diversity in Nonalcoholic Steatohepatitis

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