趋化因子配体18重组成熟肽段的表达

目的构建PET SUMO-CCL18原核表达质粒,并表达、纯化获得重组趋化因子配体18成熟肽段。方法从人卵巢癌组织中克隆CCL18基因全长,继而扩增表达成熟蛋白质的核酸序列,连接人PETSUMO载体,重组阳性克隆转入感受态细胞BL21（DE3）中IPTG诱导表达,基质辅助激光解吸／电离质谱技术（MALDI-TOF）验证后以SUMO蛋白酶切除载体部分,纯化浓缩,MALDI-TOF鉴定表达结果。结果测序分析证实,克隆人PETSUMO载体的CCL18序列与Genbank中报道序列完全一致,纯化后蛋白质谱鉴定与天然CCL18成熟蛋白序列一致。结论成功构建了高表达CCL18蛋白的表达系统,得到重组CCL18成熟肽段,为进一步研究CCL18蛋白在卵巢癌中的作用提供实验基础。     

免疫细胞治疗联合化疗治疗晚期卵巢癌的疗效及对血清CCL-18 水平的影响

目的:观察免疫细胞治疗联合化疗治疗晚期卵巢癌的疗效,及该疗法对血清趋化因子配体18(CCL-18)的影响。方法:收集2013年1月-2014年6月我院收治的Ⅳ期卵巢癌患者180例,根据随机数字表法将患者分为对照组和治疗组,各90例,对照组采用单纯化疗,治疗组在化疗基础上联合免疫细胞治疗,观察两组患者的治疗疗效,用酶联免疫吸附实验(ELISA)检测两组治疗前后血清CCL-18水平。结果:治疗组总有效率(ORR)为65.56%,显著高于对照组的41.11%,差异有统计学意义(P=0.001);与治疗前比较,两组患者治疗后血清CCL-18均降低,且治疗组下降更明显(P=0.000)。结论:免疫细胞治疗联合化疗治疗晚期卵巢癌的疗效确切显著,能显著降低患者血清CCL-18的水平,值得临床推广使用。     

CC趋化因子配体2与肝疾病

近年来CC趋化因子配体2(chemokine(C-C motif)ligand 2,CCL2)在肝脏疾病发病机制中的作用越来越受到重视.大量研究表明,CCL2在各种肝损伤中表达上调.CCL2是炎症反应的主要调节因子,通过与其受体CCR2相互作用,使血液中的单核细胞穿过血管内皮向炎症部位迁移.白色脂肪组织分泌的CCL2能直接诱导肝细胞的脂肪聚集,与非酒精性肝病的发病机理密切相关.肝实质细胞分泌的CCL2能激活并募集肝星形细胞,参与肝纤维化甚至肝硬化的形成.CCL2能介导肝癌细胞的转移和浸润,刺激肿瘤血管生成,其与肿瘤的关系也成为研究的热点.本文将阐述CCL2与病毒性肝炎、酒精性肝炎、非酒精性脂肪性肝炎、肝纤维化、肝硬化和肝癌的研究进展.     

CXC趋化因子配体8促进结直肠癌微环境中M2型巨噬细胞趋化及浸润

CXC趋化因子配体8(CXC chemokine ligand 8,CXCL8)在结直肠癌等多种肿瘤中高表达,并促进肿瘤恶性进展。研究发现,结直肠癌微环境中有大量M2型巨噬细胞浸润,但CXCL8是否影响M2型巨噬细胞的浸润及其潜在机制尚未可知。本文旨在探讨CXCL8对结直肠癌中M2型巨噬细胞浸润及趋化作用的影响。本研究首先分析了TCGA数据库结直肠癌样本中CXCL8表达水平及免疫细胞浸润情况,并在临床组织中进行验证。随后Western 印迹及qRT-PCR检测5种结直肠癌细胞株CXCL8的表达情况。佛波酯(PMA)及IL-4诱导THP-1至M2型巨噬细胞后,与HCT116、SW480细胞及过表达CXCL8的HCT116(CXCL8/HCT116)、SW480(CXCL8/SW480)共培养,检测M2型巨噬细胞趋化情况。白细胞介素1β(IL-1β)处理HCT116、SW480细胞,检测CXCL8表达情况,与M2型巨噬细胞共培养,分析趋化结果。结果显示,患者癌组织CXCL8表达高于癌旁组织,CXCL8高表达癌组织中存在更多M2型巨噬细胞浸润;IL-1β作用于HCT116或SW480后,CXCL8的mRNA及蛋白质表达水平升高(P<0.05)。Transwell实验证实,CXCL8趋化M2型巨噬细胞(P<0.05)。综上所述,结直肠癌细胞中CXCL8可由IL-1β诱导产生,CXCL8表达增加能够促进M2型巨噬细胞的趋化,结直肠癌微环境中M2型巨噬细胞大量浸润可能与CXCL8表达升高有关。     

趋化因子CCL-18 在慢性鼻- 鼻窦炎中的表达及意义

目的:分析趋化因子CCL-18在不同组织病理特征慢性鼻-鼻窦炎和正常鼻黏膜的表达差异,探讨CCL-18在慢性鼻-鼻窦炎中的表达及意义。方法:采用苏木精-伊红染色(HE),Masson染色及过碘酸-雪夫(PAS)染色对慢性鼻-鼻窦炎组织进行病理分析。采用Western blot检测CCL-18蛋白水平在不同组织病理特征慢性鼻-鼻窦炎和正常鼻黏膜组织中的表达差异。结果:CCL-18蛋白水平在伴鼻息肉和不伴有鼻息肉慢性鼻窦炎均较正常鼻黏膜组织中显著上调(P<0.05)。CCL-18蛋白水平在嗜酸性粒细胞慢性鼻窦炎的表达水平明显高于非嗜酸性粒细胞慢性鼻窦炎(P<0.05)。腺体型,纤维炎症型及水肿型慢性鼻-鼻窦炎中CCL-18表达水平均高于正常鼻黏膜,以水肿型表达最为显著(P<0.05)。结论:CCL-18在嗜酸性粒细胞和水肿型慢性鼻窦炎中高度表达,提示CCL-18可能参与慢性鼻-鼻窦炎中嗜酸性粒细胞的浸润这一基本病理过程。     

ZCCHC10通过激活P53促进急性髓系白血病细胞中CCL18基因的转录

肿瘤抑制因子P53是一种转录因子,可激活一系列靶基因的转录,从而调控细胞周期停滞、DNA修复、免疫、炎症和细胞凋亡等多种生理或病理过程。前期研究表明, CCHC型锌指蛋白10 (zinc finger CCHC-type containing 10, ZCCHC10)通过激活P53在肺癌和急性髓系白血病中发挥抑癌作用。为进一步探讨ZCCHC10在急性髓系白血病中的作用机制,本文通过RNA测序(RNA sequencing, RNA-seq)技术对过表达ZCCHC10或空载体的ML2细胞进行了转录组分析,一共鉴定到1 284个差异基因[|log2(fold change)|≥1, q值<0.05],包括778个上调基因和506个下调基因。其中,趋化因子CCL18在过表达ZCCHC10的ML2细胞中上调18倍。生物信息学分析显示, CCL18基因启动子上含有两个P53反应元件。生物素标记DNA亲和实验和染色质免疫共沉淀实验证实, P53可结合到CCL18基因启动子上。荧光素酶活性分析表明, P53可以增强CCL18基因启动子的活性。这些研究表明ZCCHC10通过激活P53促进CCL1...     

CCL20及其受体CCR6在肝细胞肝癌中的作用

肝细胞肝癌(hepatocellular carcinoma,HCC)(简称肝癌)是常见的血管侵袭性高、耐药性强的恶性肿瘤之一.CC趋化因子配体20(CC chemokine ligand 20,CCL20)作为一种炎症介质能够诱导肿瘤细胞定向迁移,与其唯一受体趋化因子受体6(chemokine receptor 6,...     

CC类趋化因子配体5与冠状动脉粥样硬化的相关性研究

目的:探讨CC类趋化因子配体5(CC chemokine ligand-5,CCL5)的表达水平与冠状动脉粥样硬化(Atherosclerosis,AS)严重程度之间的相关性。方法:将颈动脉粥样硬化狭窄(70%)行颈动脉内膜切除术的5例手术标本作为AS组,尸检无动脉粥样硬化的正常颈动脉3例作为对照组。应用免疫组织化学、Western blot检测AS组和对照组中CCL5的表达。收集110例冠状动脉造影术患者,根据Gensini评分分为对照组(Gensini评分=0分,n=27),轻微病变组(0Gensini评分≤20,n=32)和严重病变组(Gensini评分20,n=51)。收集各组的临床基线资料(包括年龄、性别、血压、血脂、血糖等),ELISA方法检测各组血浆中CCL5表达水平,并分析各危险因素、CCL5表达水平与Gensini评分之间的相关性。结果:颈AS组AS组织中CCL5蛋白的表达明显高于正常颈动脉组(P0.05),冠心病患者血浆CCL5水平明显高于对照组,差异具有统计学意义(P0.05)。对照组、轻微病变组和严重病变组在性别、吸烟史、收缩压、舒张压、血糖及血浆CCL5的表达上存在显著的差别。相关性分析显示舒张压、血糖及血浆CCL5表达水平与Gensini评分间呈显著正相关(r=0.276、0.418、0.519,P0.05)。以Gensini评分为因变量,性别、年龄、吸烟史、收缩压、舒张压、LDL-C、HDL-C、血糖、血浆CCL5为自变量建立多元逐步线性回归模型,结果提示血浆CCL5为Gensini评分的独立预测因子(B值为8.775;P值为0.000)。结论:血浆CCL5水平与冠状动脉粥样硬化病变程度呈正相关,可能作为AS程度的独立预测因子。     

胆囊结石患者血清microRNA表达谱及其意义

目的:建立胆囊结石患者与健康人的血清microRNA(miRNA)表达谱,并分析其差异miRNA功能。方法:收集解放军总医院诊治的胆囊结石患者的血清(试验组)及健康人的血清(对照组),采用miRNA测序技术,建立胆囊结石患者的miRNA表达谱,并通过qPCR技术验证其差异miRNA,利用生物信息学技术预测其差异miRNA的生物功能。定量资料以x±s描述,两组数据间比较采用t检验。结果:试验组与对照组miRNA表达谱序列分别为14 899 245和11 783 121个,miRNA种类分别为686和633个,其差异表达miRNA为16个,均为下调。GO分析显示差异miRNA的靶基因在细胞过程上富集于细胞转化、生物调节和代谢过程等,在细胞组成上富集于细胞成分和细胞器成分等,在分子功能上富集于结合和活性催化等;KEGG功能分析显示差异miRNA的靶基因主要参与环腺苷酸、催产素、生物节律等代谢通路。利用qPCR方法验证miR-1228、miR-1249、miR-3614和miR-766在2组间差异趋势与测序结果基本一致,其中miR-1228、miR-3614和miR-766在2组间存在统计学差异,而miR-1249无统计学差异。结论:血清中的差异miRNA在胆囊结石的形成中可能发挥重要作用,其对胆囊结石疾病的防治具有重要意义。     

Surgical trauma-induced CCL18 promotes recruitment of regulatory T cells and colon cancer progression

Background: Surgical stress has been suggested to facilitate colon cancer growth and metastasis. However, the precise mechanisms by which surgical trauma promotes colon cancer progression remain poorly understood. Methods: To unravel the mechanisms underlying surgery-induced colon cancer progression, a syngenic transplantation tumor model was established with CT26 cells, and the effect of laparotomy on tumor progression was investigated. Especially, the expression of several chemokines was assessed, and their roles in recruiting CD4+ CD25+ regulatory T cells (Tregs) after surgery were analyzed. Results: Tregs population was significantly increased in the tumor tissue and peripheral blood of tumor-bearing mice after laparotomy. C-C motif chemokine ligand 18 (CCL18) expression was significantly upregulated after laparotomy in tumor tissue and the peritoneal cavity of tumor-bearing mice, and it was positively correlated with the recruitment of Tregs. Functionally, CCL18 knockdown significantly reduces tumor growth and angiogenesis compared with control. Through analysis of Tregs, we found an upregulated proportion of Tregs in tumor tissue, peritoneal cavity, and peripheral blood after laparotomy, but this enhancement was blocked after CCL18 knockdown. In patients with colon cancer, a higher Tregs proportion is positively correlated to more advanced clinical TNM stages and shorter survival. Furthermore, a positive correlation was found between the serum CCL18 level and the Treg proportion in clinical samples. Conclusion: Surgical trauma contributes to colon cancer progression by increasing CCL18 expression and hence promotes Treg recruitment, which leads to an immunosuppressive environment.     

RegIV和CCL7在胃癌中的表达及相关性分析

目的:研究再生基因4(RegIV)和趋化因子CCL7在胃癌中的表达及相关性。方法:选择2010年3月至2014年3月在我院接受手术治疗的胃癌患者72例作为研究对象。对比RegIV及CCL7在胃癌组织以及癌旁正常组织中的阳性表达情况,分析不同病理参数对RegIV及CCL7阳性表达的影响,RegIV与CCL7表达的相关性及影响RegIV及CCL7在胃癌中表达的危险因素。结果:胃癌组织的RegIV阳性率及CCL7阳性率均显著高于癌旁正常组织,差异均有统计学意义(均P0.05)。有浆膜浸润、有淋巴结转移以及TNM分期为Ⅲ~Ⅳ期者的RegIV与CCL7阳性表达率均显著高于无浆膜浸润、无淋巴结转移以及TNM分期为0~Ⅱ期者,差异均有统计学意义(均P0.05)。经Spearman法分析相关性可知,RegIV与CCL7表达呈正相关。根据Logistic回归分析法可知,有浆膜浸润、有淋巴结转移以及TNM分期为Ⅲ~Ⅳ期均为RegIV及CCL7在胃癌中表达的危险因素。结论:RegIV与CCL7均可在胃癌组织中呈现出较高的阳性表达,且二者与患者临床预后有密切关系,在临床治疗过程中值得关注与重视。     

胃癌患者血清CCL2、ANXA2含量与癌细胞浸润性生长的相关性研究

该文旨在探讨胃癌患者血清CC趋化因子配体2(CCL2)、膜联蛋白A2(ANXA2)含量与癌细胞浸润性生长的相关性,并分析其与胃癌临床病理特征的关系.选择187例胃癌患者为研究对象,并选择153例正常人群为对照组,使用酶联免疫吸附试验(ELISA)法检测所有研究对象血清CCL2、ANXA2含量,分析组间临床病理特征(癌细...     

Chemokine CCL2 modulation of neuronal excitability and synaptic transmission in rat hippocampal slices

In addition to its well-characterized effects in immune system, chemokine CC motif ligand 2 (CCL2, formerly known as monocyte chemoattractant protein-1) is believed to play an important role in brain physiological and pathological processes. It has been shown that CCL2 and its cognate receptor chemokine CC motif receptor 2 are constitutively expressed in several brain regions including the hippocampus, and the expression is up-regulated under pathological conditions. Whereas most investigations have so far focused on its involvement in CNS pathology, few studies have examined the effects of CCL2 on neuronal and synaptic physiology. In this study, we tested the effects of CCL2 on neuronal excitability and excitatory synaptic transmission in the CA1 region of rat hippocampal slices using whole-cell patch clamp techniques. Bath application of CCL2 depolarized membrane potential and increased spike firing in CA1 neuronal cells. Bath application of CCL2 also produced an increase of excitatory post-synaptic currents recorded in Schaffer-collateral fibers to CA1 synapses. Quantal analysis revealed that CCL2 increased the frequency of spontaneous excitatory post-synaptic current occurrence and mean quantal content. Taken together, our data indicate that CCL2 enhances neuronal excitability and synaptic transmission via pre-synaptic mechanisms. These results support the emerging concept that chemokines function as neuromodulators in the CNS.     

The Interaction of Heparin Tetrasaccharides with Chemokine CCL5 Is Modulated by Sulfation Pattern and pH

Interactions between chemokines such as CCL5 and glycosaminoglycans (GAGs) are essential for creating haptotactic gradients to guide the migration of leukocytes into inflammatory sites, and the GAGs that interact with CCL5 with the highest affinity are heparan sulfates/heparin. The interaction between CCL5 and its receptor on monocytes, CCR1, is mediated through residues Arg-17 and -47 in CCL5, which overlap with the GAG-binding ⁴⁴RKNR⁴⁷ “BBXB” motifs. Here we report that heparin and tetrasaccharide fragments of heparin are able to inhibit CCL5-CCR1 binding, with IC₅₀ values showing strong dependence on the pattern and extent of sulfation. Modeling of the CCL5-tetrasaccharide complexes suggested that interactions between specific sulfate and carboxylate groups of heparin and residues Arg-17 and -47 of the protein are essential for strong inhibition; tetrasaccharides lacking the specific sulfation pattern were found to preferentially bind CCL5 in positions less favorable for inhibition of the interaction with CCR1. Simulations of a 12-mer heparin fragment bound to CCL5 indicated that the oligosaccharide preferred to interact simultaneously with both ⁴⁴RKNR⁴⁷ motifs in the CCL5 homodimer and engaged residues Arg-47 and -17 from both chains. Direct engagement of these residues by the longer heparin oligosaccharide provides a rationalization for its effectiveness as an inhibitor of CCL5-CCR1 interaction. In this mode, histidine (His-23) may contribute to CCL5-GAG interactions when the pH drops just below neutral, as occurs during inflammation. Additionally, an examination of the contribution of pH to modulating CCL5-heparin interactions suggested a need for careful interpretation of experimental results when experiments are performed under non-physiological conditions.     

CCL18 promotes the metastasis of squamous cell carcinoma of the head and neck through MTDH‐NF‐κB signalling pathway

Metastasis is one of the primary causes for high mortality in patients with squamous cell carcinoma of the head and neck (SCCHN). Our previous study showed that chemokine (C‐C motif) ligand 18 (CCL18), derived from tumour‐associated macrophages (TAMs), regulates SCCHN metastasis by promoting epithelial‐mesenchymal transition (EMT) and preserving stemness. However, the underlying mechanism needs to be further investigation. Interestingly, metadherin (MTDH) expression was induced when SCCHN cells were stimulated with recombinant CCL18 protein in this study. Suppressing MTDH expression reversed CCL18‐induced migration, invasion and EMT in SCCHN cells. Furthermore, the NF‐κB signalling pathway was involved in the MTDH knock‐down cells with CCL18 stimulation. We performed ELISA to evaluate the CCL18 levels in the serums of 132 treatment‐naive SCCHN patients, 25 patients with precancerous lesion and 32 healthy donors. Our results demonstrated that serum CCL18 levels were significantly higher in SCCHN patients than patients with precancerous lesion and healthy individuals. CCL18 levels were found to be significantly correlated with tumour classification, clinical stage, lymph node metastasis and histological grade in SCCHN patients. Thus, our findings suggest that CCL18 may serve as a potential biomarker for diagnosis of SCCHN and promote SCCHN invasion, migration and EMT by MTDH‐NF‐κB signalling pathway.