细胞膜磷脂与肿瘤侵袭、转移及靶向治疗的相关性研究进展

侵袭和转移是肿瘤进展的标志,是肿瘤临床治疗的难题及肿瘤患者死亡的主要原因。磷脂是生物膜的重要物质基础,参与肿瘤细胞增殖、运动、黏附、凋亡、信号转导、细胞周期调节等活动。本文对膜磷脂在肿瘤侵袭、转移、靶向治疗中的作用研究进展进行了综述。细胞膜磷脂成分及分布、磷脂代谢产物、磷脂酶及磷脂结合蛋白在肿瘤细胞膜流动性、黏附、凋亡、增殖、侵袭、转移、血管及淋巴管形成中具有重要作用,以细胞膜磷脂及其代谢产物为靶点治疗恶性肿瘤具有广阔前景。     

Membrane Fluidity and Adherence to Extracellular Matrix Components Are Related to Blast Cell Count in Acute Myeloid Leukemia

The level of blast cells in peripheral blood in acute myeloid leukemia (AML) varies in individual patients. The bone marrow egress of hematopoietic cells is an unclear phenomenon in which cell deformability and cytoadhesion to the extracellular matrix are involved. One component of deformability is the membrane fluidity. Using fluorescence polarization, we have studied the fluidity of blast cell membranes from 22 AML patients. This membrane was found to be highly fluid and a statistically significant correlation was found between the increase in membrane fluidity and the number of blast cells in the blood. Studying interaction between blast cells and several components of bone marrow stroma, we found adhesion to fibronectin and fibroblastic extracellular matrix. Adhesion to the extracellular matrix was inversally correlated to the level of blast cells in the blood. The observed increase in membrane fluidity and reduction of adhesion to bone marrow stroma may result in an increase of blast cells egress in AML.     

Shedding of Membrane Type 1 Matrix Metalloproteinase in a Human Breast Carcinoma Cell Line

Membrane type 1 matrix metalloproteinase (MT1-MMP) with a transmembrane domain is a new member of the MMP gene family and is expressed on the cell surfaces of many carcinoma cells to activate the zymogen of MMP-2 (gelatinase A). We have previously reported that MT1-MMP is released into culture media in a complex form with tissue inhibitor of metalloproteinases 2 (TIMP-2) from a human breast carcinoma cell line, MDA-MB-231, treated with concanavalin A (Con A). In the present study, we further studied the release mechanism of MT1-MMP. Immunoblot analysis indicated that the amounts of MT1-MMP in culture media increase with the time of exposure and the concentration of Con A, and those in cell lysates conversely decrease in a similar way. Time- and dose-dependent release of MT1-MMP into the media was confirmed by a sandwich enzyme immunoassay specific to MT1-MMP. The molecular weight of the immunoreactive MT1-MMP in the media was M_r 56,000, which was 4,000-M_r smaller than that in the cell lysates. Northern blot analysis demonstrated that the mRNA expression level of MT1-MMP is about 3-fold enhanced after a 24 h-exposure to Con A and this is maintained up to 72-h exposure. The release of MT1-MMP from the Con A-treated cells was inhibited by metalloproteinase inhibitors such as EDTA and o-phenanthroline, but not by MMP inhibitors including TIMP-1, TIMP-2 and BB94 or other proteinase inhibitors of serine, cysteine and aspartic proteinases. During the Con A treatment of the cells, cell viability decreased time- and dose-dependently and dead cells reacted positively in the TdT-mediated dUTP Nick-End Labeling (TUNEL) method. Con A-treated MDA cells showed apoptotic morphology when stained with Hoechst dye and hematoxylin and eosin. DNA ladder formation was detected by electrophoresis of the DNA from Con A-treated MDA cells. These results suggest that MT1-MMP release from Con A-treated cells is due to shedding mediated by metalloproteinase(s) other than MMPs, and is associated with apoptosis.     

Modulation of Membrane Permeability, Cell Proliferation and Cytotoxicity of Antitumor Agents by External ATP in Mouse Tumor Cells

External ATP causes a remarkable change in the passive permeability of the plasma membrane in several types of transformed cells. When mouse melanoma cells, Clone-M3, were exposed to ATP in Tris-buffered saline, a great increase in the passive permeability was induced within several minutes. Longer exposure of Clone-M3 cells to external ATP led to a decrease in cell viability. Similar results were obtained with Ehrlich ascites cells, but none of these ATP effects were noted in untransformed cells such as NIH 3T3 cells or BALB/c mouse embryonic fibroblasts. The in vitro cytotoxic effects of antitumor agents (5-fluorouracil, adriamycin, mitomicin C and nimustine hydrochloride) against Clone-M3 cells were additively potentiated by treatment with external ATP, which also synergistically enhanced the cytotoxicity of vincristine. However, the effects of these drugs on mouse embryonic fibroblasts were not modulated by ATP. These results suggest that ATP-treatment is a useful means of enhancing a selective toxicity for tumor cells.     

Low Incidence of Point Mutations in H-, K- and N-ras Oncogenes and p53 Tumor Suppressor Gene in Renal Cell Carcinoma and Peritoneal Mesothelioma of Wistar Rats Induced by Ferric Nitrilotriacetate

An iron chelate, ferric nitrilotriacetate (Fe-NTA), induces renal proximal tubular damage, a consequence of iron-catalyzed free radical reactions, that finally leads to a high incidence of renal cell carcinoma (RCC) in rodents. Previous studies have identified, within 24 h after administration of Fe-NTA, lipid peroxidation products, aldehyde-modified proteins and a variety of modified DNA bases such as 8-hydroxyguanine that may be mutagenic in vivo. In the present study, pathological features of the RCCs were studied, and, in an effort to correlate them with carcinogen-specific molecular events in Fe-NTA-induced carcinogenesis, the H-, K- and N- ras oncogenes and the p53 tumor suppressor gene were investigated for the presence of mutations. Fe-NTA-induced RCCs showed similarity to human RCCs in that they are often invasive, metastatic and fatal. None (0 of 12) of the tumors had mutation in codons 12, 13 and 61 of the H-, K- and N- ras genes by direct sequencing. Only one (1 of 12) tumor with high grade histology revealed a CGC-to-CTC (Arg to Leu) transversion in codon 246 of the p53 gene by the use of single strand conformation polymorphism (SSCP) analysis and direct sequencing. High expression of mutant p53 protein was confirmed by Western blotting and immunohistochemistry. Study of three peritoneal mesotheliomas induced by Fe-NTA revealed no mutation in ras and p53 genes. These results suggest that the ras and p53 genes are not the major targets of mutation in Fe-NTA-induced carcinogenesis of kidney and mesothelium. Instead, p53 mutation may work for potentiation of malignant character in Fe-NTA-induced renal carcinogenesis.     

Apoptotic Changes Precede Mitochondrial Dysfunction in Red Cell-type Pyruvate Kinase Mutant Mouse Erythroleukemia Cell Lines

Two erythroleukemia cell lines have been established from the splenic lesions of red blood celltype pyruvate kinase (R-PK) activity-deficient mice of CBA/N origin infected with a polycythemic strain of Friend leukemia virus complex (FVp). Ten to 30 % of the cells of these cell lines undergo apoptotic changes in routine passage, as shown by nuclear fragmentation, DNA laddering, DNA content (propidium iodide (PI) staining), and annexin V binding assay. In these cells, however, although adenosine 5'-triphosphate (ATP) levels were lower than in the control cells, the mitochondrial inner transmembrane potential (Δψ_m), detected by rhodamine 123 (R123) and diSC₃(5) staining, remained unchanged until the final stage of apoptosis. No evidence was obtained to relate this finding to R-PK mutation due to difficulty in cloning stable, conditionally inducible R-PK gene transfectants. However, low Δψ_m in the apoptotic cell population of the control T3-K-1 (K-1) and T3-Cl-2-0 (2-0) Friend erythroleukemia cells supports a possible relationship, as do results obtained in two Friend erythroleukemia cells recently isolated from normal CBA/N mice. These cell lines are expected to be useful for clarifying both the primary apoptotic changes independent of mitochondrial dysfunction and the PK-isozyme changes during erythrodifferentiation, for example, the decreased muscle type 2 (M2) PK level. Modification of growth signals in these cell lines may modulate differentiation and/or apoptosis and allow further elucidation of the signaling networks.     

肿瘤发生发展过程中红细胞免疫功能的变化

用小鼠可移植性组织细胞型淋巴瘤ＬⅡ接种于近交系６１５小鼠右后爪垫皮下，按宿主荷瘤时间以及肿瘤生长发展的不同阶段分期分批取血，检测荷瘤小鼠ＰＢＣ－Ｃ３ｂＲＲ和ＲＢＣ－ＩＣＲ。结果组织学观察，将肿瘤生长发展过程分为潜伏期，侵袭早，中，晚期，转移早，中，晚期等７个时期。观察ＲＢＣ－Ｃ３ｂＲＲ明显低于正常，并随肿瘤的发展呈下降趋势，而ＲＢＣ－ＩＣＲ高于正常，在肿瘤侵袭中期急骤增高，以后稳定在较高水平，提示     

荷瘤小鼠瘤苗治疗疗效与红细胞天然免疫活性相关性及其意义

[目的]探讨瘤苗对荷瘤小鼠红细胞天然免疫活性影响的临床意义.[方法]采用直向红细胞天然免疫黏附肿瘤细胞试验(DRIAC)和促红细胞天然免疫黏附肿瘤细胞试验(IRIAC)测定30例荷瘤小鼠.[结果]死瘤苗治疗组DRIAC水平(26.25±9.85)明显高于未治疗对照组水平(14.38±3.18,P<0.01).溶瘤苗治疗组IRIAC水平(12.14±3.18)明显低于未治疗对照组水平(17.25±3.8,P<0.01).[结论]瘤苗作用与红细胞天然免疫活性之间相关性分析表明红细胞天然免疫黏附瘤苗制品应有使用价值.     

Membrane proteomic analysis comparing squamous cell lung cancer tissue and tumour-adjacent normal tissue

Lung cancer is the leading cause of cancer-related deaths worldwide. Squamous cell carcinoma is one of the predominant histological subtypes of lung cancer. Detecting lung cancer at an early stage is essential for successful therapy and increasing survival. There are still no satisfactory biomarkers for the early detection of lung cancer. In this study, tumour tissue paired with tumour-adjacent normal bronchial epithelial tissue was obtained from patients with squamous cell lung carcinoma without metastasis. The proteins extracted from the cell membrane were separated by two-dimensional polyacrylamide gel electrophoresis (2-DE) and were analysed with the Image Master two-dimensional platinum software. Twenty-five significantly different protein spots were selected and identified with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS). A total of 19 proteins were successfully identified. Twelve proteins were up-regulated, and seven proteins were down-regulated in the cancerous tissue compared with the tumour-adjacent normal tissue. One up-regulated protein and one down-regulated protein in squamous cell lung carcinoma were verified by Western blot analysis and RT-PCR; the results were consistent with the 2-DE analysis. In conclusion, membrane proteomics identified a number of candidate biomarker proteins that were differentially expressed between squamous cell lung cancer tissue and adjacent normal tissue. These biomarker candidates have the potential to elucidate the underlying pathogenesis of squamous cell lung cancer.     

Membrane transporters and channels in chemoresistance and -sensitivity of tumor cells

Membrane transporters play important roles in mediating chemosensitivity and -resistance of tumor cells. ABC transporters, such as ABCB1/MDR1, ABCC1/MRP1 and ABCG2/BCRP, are frequently associated with decreased cellular accumulation of anticancer drugs and multidrug resistance of tumors. SLC transporters, such as folate, nucleoside, and amino acid transporters, commonly increase chemosensitivity by mediating the cellular uptake of hydrophilic drugs. Ion channels and pumps variably affect sensitivity to anticancer therapy by modulating viability of tumor cells. A pharmacogenomic approach, using correlations between drug potency and transporter gene expression in multiple cancer cell lines, has shown promise for identifying potential drug-transporter relationships and predicting anticancer drug response, in an effort to optimize chemotherapy for individual patients.     

七氟烷联合丙泊酚用于胸腔镜非小细胞肺癌切除术对术后患者炎性与氧化、抗氧化状态的影响

目的 研究与观察七氟烷联合丙泊酚用于胸腔镜非小细胞肺癌切除术对术后患者炎性与氧化、抗氧化状态的影响.方法 选取78例胸腔镜非小细胞肺癌切除术患者为研究对象,依据麻醉方式的不同分为对照组(丙泊酚组)39例和观察组(七氟烷联合丙泊酚组)39例.比较2组术前及术后3 d、7 d的血清炎性指标(白细胞介素及其他炎症因子)与氧化...     

Diagnosis Value of Membrane Glycolipids Biochemistry Index in Intracranial and Gastrointestinal Tumors

The diagnostic value of membrane glycolipid biochemistry index, the lipid-bound sialic acid (LSA) and totalsialic acid (TSA) in cerebrospinal fluid (CSF) was evaluated in 30 intracranial and 65 gastrointestinal tumors.The plasma LSA, TSA and red cell membrane sialic acid (R-SA) in were determined according to the method ofSevenmerhulm. Our results showed that the levels of LSA and TSA in CSF of intracranial tumor patients washigher than that of normal group(p<0.01). The concentration of TSA and LSA in patients with malignant gliomawas higher than that of benign meningioma patients(P<0.01). No significance was found between intracranialhalmatoma patients and normal control group for levels of membrane glycolipids (p>0.05). Results also found thatthe plasma LSA, TSA and R-SA of gastric carcinoma were significantly higher than those of control group (p<0.05);while no significant difference was found in the plasma LSA, TSA and R-SA levels between chronic gastritis,gastrohelcoma and normal control group (p>0.05). Plasma LSA, TSA and R-SA levels of gastric carcinoma patientwere significantly higher than those of chronic gastritis patients and gastrohelcoma patients(p<0.05). It was alsofound that plasma LSA, TSA and R-SA contents were significantly higher in large intestine carcinoma patientsthan in benign in stestine tumor patients (p<0.05) while no significant difference was found between intestinebenign tumor and normal control group (p>0.05). The levels of LSA, TSA and R-SA were obviously higher inthe patients with metastasis than in the ones without (p<0.05.) The membrane glycolipid biochemistry indexLSA and TSA in CSF are sensive markers for diagnosing intracranial tumors. For gastrointestinal malignanttumors the plasma LSA TSA and red blood cell membrane SA may be considered as auxiliary indicators fordiagnosis. They can be used for distinguishing benign from malignant tumors.     

Changes in DNA Damage Repair Gene Expression and Cell Cycle Gene Expression Do Not Explain Radioresistance in Tamoxifen-Resistant Breast Cancer

Tamoxifen-induced radioresistance, reported in vitro, might pose a problem for patients who receive neoadjuvant tamoxifen treatment and subsequently receive radiotherapy after surgery. Previous studies suggested that DNA damage repair or cell cycle genes are involved, and could therefore be targeted to preclude the occurrence of cross-resistance. We aimed to characterize the observed cross-resistance by investigating gene expression of DNA damage repair genes and cell cycle genes in estrogen receptor-positive MCF-7 breast cancer cells that were cultured to tamoxifen resistance. RNA sequencing was performed, and expression of genes characteristic for several DNA damage repair pathways was investigated, as well as expression of genes involved in different phases of the cell cycle. The association of differentially expressed genes with outcome after radiotherapy was assessed in silico in a large breast cancer cohort. None of the DNA damage repair pathways showed differential gene expression in tamoxifen-resistant cells compared to wild-type cells. Two DNA damage repair genes were more than two times upregulated (NEIL1 and EME2), and three DNA damage repair genes were more than two times downregulated (PCNA, BRIP1, and BARD1). However, these were not associated with outcome after radiotherapy in the TCGA breast cancer cohort. Genes involved in G₁, G₁/S, G₂, and G₂/M phases were lower expressed in tamoxifen-resistant cells compared to wild-type cells. Individual genes that were more than two times upregulated (MAPK13) or downregulated (E2F2, CKS2, GINS2, PCNA, MCM5, and EIF5A2) were not associated with response to radiotherapy in the patient cohort investigated. We assessed the expression of DNA damage repair genes and cell cycle genes in tamoxifen-resistant breast cancer cells. Though several genes in both pathways were differentially expressed, these could not explain the cross-resistance for irradiation in these cells, since no association to response to radiotherapy in the TCGA breast cancer cohort was found.     

Swords of Cell Death: Caspase Activation and Regulation

Apoptosis represents a crucial process in modulating organ development in the embryo, in organ homeostasis in the adult, and in fostering appropriate immunological function. Caspases represent two central class of molecules that are either involved with the stimulation of the apoptotic cascade (initiator caspases), or the various sequential biological pathways required for its execution (effector caspases). With an eye towards therapeutic opportunities, this review discusses in detail the lineage of initiator and effector caspases, how they are each activated, their substrates, their regulation, and maps out how they interact throughout the process from initiation of the first apoptotic signal to the final consequential breakdown of cellular integrity.     

Changes in Brain Energy and Membrane Metabolism in Glioblastoma following Chemoradiation

Brain parenchyma infiltration with glioblastoma (GB) cannot be entirely visualized by conventional magnetic resonance imaging (MRI). The aim of this study was to investigate changes in the energy and membrane metabolism measured with phosphorous MR spectroscopy (31P-MRS) in the presumably “normal-appearing” brain following chemoradiation therapy (CRT) in GB patients in comparison to healthy controls. Twenty (seven female, thirteen male) GB patients underwent a 31P-MRS scan prior to surgery (baseline) and after three months of standard CRT (follow-up examination. The regions of interest “contrast-enhancing (CE) tumor” (if present), “adjacent to the (former) tumor”, “ipsilateral distant” hemisphere, and “contralateral” hemisphere were compared, differentiating between patients with stable (SD) and progressive disease (PD). Metabolite ratios PCr/ATP, Pi/ATP, PCr/Pi, PME/PDE, PME/PCr, and PDE/ATP were investigated. In PD, energy and membrane metabolism in CE tumor areas have a tendency to “normalize” under therapy. In different “normal-appearing” brain areas of GB patients, the energy and membrane metabolism either “normalized” or were “disturbed”, in comparison to baseline or controls. Differences were also detected between patients with SD and PD. 31P-MRS might contribute as an additional imaging biomarker for outcome measurement, which remains to be investigated in a larger cohort.     

检测血清纤维连接蛋白（FN）在盆腔放射性损伤中的临床价值

目的：初步探讨血清纤维连接蛋白（FN）与盆腔放射性损伤的关系及其意义。方法：采用酶联免疫吸附法（ELISA）动态检测Ⅱ-Ⅲ期妇科肿瘤患者在放疗前、放疗中、放疗结束及放疗后随诊期间血清FN的含量变化。结果：血清FN水平在放疗开始后即出现明显的升高趋势,至1周时达第一高峰,此后在整个放疗期间呈间断性上升,至放疗结束时达第二高峰。发生放射性反应的患者在放疗结束后仍旧呈现持续性升高,而无放射性反应者表现为逐渐下降。结论：血清FN与放射治疗密切相关,可随放射治疗剂量的增加而升高,而与是否发生急性放射性损伤无关。在放射性损伤的发展过程中是一个迟发反应标记物。     

右美托咪定对顺铂诱导的非小细胞肺癌细胞凋亡及氧化应激的影响

目的 探讨右美托咪定(Dex)对顺铂诱导的人非小细胞肺癌(NSCLC)细胞凋亡的影响及其潜在机制.方法 将人非小细胞肺癌细胞系分为3组:对照组、顺铂组和Dex组.采用Western Blot法检测各组细胞中凋亡相关蛋白、细胞色素C、mTOR/ERK1/2信号分子及E-钙粘蛋白的表达,采用试剂盒法检测活性氧(ROS)的含...     

汞、镉对小鼠离体骨髓细胞和睾丸生殖细胞的DNA损伤作用

背景与目的: 研究氯化汞、氯化镉对小鼠离体骨髓细胞和睾丸生殖细胞的DNA损伤作用,比较两种细胞DNA损伤的敏感性差异。材料与方法: 分别以0、0.01、0.1、1 mmol/L剂量氯化汞和0、0.04、0.2、1、5 mmol/L剂量氯化镉处理离体小鼠骨髓细胞和睾丸生殖细胞1h,应用单细胞凝胶电泳(single cell gel electrophoresis, SCGE)技术检测氯化汞和氯化镉对细胞DNA的损伤率和细胞DNA迁移距离的影响。结果： 氯化汞、氯化镉处理组两种细胞DNA损伤率增高, DNA迁移距离增加。DNA损伤率和迁移距离存在明显的剂量效应关系。0.01 mmol/L剂量氯化汞、1 mmol/L氯化镉及以下剂量处理两种细胞引起睾丸生殖细胞DNA的损伤率高于骨髓细胞的DNA损伤率。结论： 氯化汞和氯化镉可引起小鼠骨髓细胞和睾丸生殖细胞DNA损伤,且睾丸生殖细胞DNA受氯化汞和氯化镉损伤更敏感。