肿瘤及肿瘤基质细胞释放的外泌体对肿瘤耐药产生的影响

恶性肿瘤疾病对人类生命健康的影响程度日益升高,不仅早期筛查诊断难度较高,并且治疗预后效果往往较差。目前,国内外临床均将化疗作为患者的主要治疗手段,但治疗过程中不断出现的药物耐药问题对患者及医务人员均造成了严重困扰。国外研究已对肿瘤耐药与外泌体进行了相关研究,认为该物质对耐药传递形成了重要影响,其外泌体中可携带耐药相关分子不断在肿瘤细胞中进行传递与作用,并且能够外排相关介导药物,通过对其实施处理可为逆转耐药起到积极作用。

卵巢粘液性肿瘤细胞基质MMP-9的表达及其临床意义

目的 :研究卵巢粘液性肿瘤组织中细胞外基质金属蛋白酶 - 9(MMP - 9)的表达及其与临床病理因素的关系。方法 :通过免疫组织化学方法检测 4 3例卵巢粘液性囊腺瘤MMP - 9的表达情况。结果 :MMP - 9在粘液性囊腺瘤中无表达 ,在交界性粘液性囊腺瘤及粘液性囊腺癌中的阳性率分别为 2 2 2 %、 5 5 2 % ;MMP - 9的表达与粘液性囊腺癌的组织学分级、FIGO分期及术后复发和死亡有密切关系。 2例交界性粘液囊腺瘤MMP - 9阳性 ,同时这 2例出现复发。结论 :MMP - 9表达与卵巢粘液性囊腺瘤的恶性程度有关 ,可望作为交界性粘液性囊腺瘤及粘液性囊腺癌的诊断和分级的客观指标 ,并协助临床估计预后。

细胞外囊泡对肿瘤生物学特性的影响研究进展

肿瘤的生长、侵袭和转移依赖于肿瘤细胞与其所处微环境中基质细胞的“交叉对话”,这种“交叉对话”主要涉及这两种细胞的可溶因子的分泌、膜包囊颗粒的释放以及细胞与细胞间的直接接触。这些膜包囊颗粒被称为细胞外囊泡(EVSs),肿瘤细胞比正常细胞可分泌更多的EVSs,这些EVSs通过依赖于内体分选复合物蛋白(ESCRT)和不依赖于ESCRT的途径将蛋白质、DNA、RNA等“内容物”分选入体内。这些“内容物”促进肿瘤细胞与基质细胞之间的信息的转递,从而支持肿瘤血管生成,侵袭转移、耐药及使得肿瘤细胞获得恶性表征。

层状双氢氧化物调控肿瘤微环境的研究进展

肿瘤微环境是肿瘤发生和生长的内在环境,包括多种细胞类型(成纤维细胞、免疫细胞、免疫抑制细胞等)、细胞外成分(细胞因子、生长因子、激素、细胞外基质等)。它具有弱酸性、缺氧、免疫抑制等特点,与肿瘤的进展密不可分。作为一种新型二维纳米材料,层状双氢氧化物(layered doublehydroxides,LDH)被广泛应用于肿瘤治疗。本文结合近些年的研究报道,综述了LDH纳米材料针对肿瘤微环境的不同特点进行调控的研究现状,总结了LDH纳米材料存在的不足,并对LDH调控肿瘤微环境未来发展趋势进行了展望。

肿瘤微环境对胆管癌发生发展的影响及其机制研究进展

肿瘤微环境是指与肿瘤细胞生长、增殖及转移等发生发展过程相关的微环境,它不仅包括肿瘤本身的基质细胞,也与肿瘤细胞所处的细胞外基质有关。胆管癌细胞可以通过协同细胞外基质和基质细胞,分泌与肿瘤有关的调节介质来构建有利于自身的肿瘤微环境。微环境缺氧、外泌体等细胞外基质主要通过释放缺氧诱导因子和大量的免疫抑制因子,激活β-catenin和E-cadherin通路来促进胆管癌的增殖和侵袭性。促炎性细胞因子、胆管癌相关成纤维细胞、慢性炎症细胞、上皮细胞、间充质干细胞、上皮间充质干细胞和肿瘤干细胞等基质细胞则可以通过释放细胞因子,激活基质细胞相互间的多种代谢通路,调控胆管癌细胞的转录、翻译过程,在促进胆管癌细胞增殖、加速血管形成、促进肿瘤侵袭和防止细胞凋亡方面发挥着重要作用。

Transfection of the Human Sodium/Iodide Symporter(NIS) Gene with Liposomes and the Expression of the NIS Protein in Human Lung A549 Cancer Cells

OBJECTIVE To examine the possibility of human sodium iodide symporter （hNIS） protein expression in lung cancer cells. METHODS Human lung A549 cancer cells were thawed and cultured in vitro. The cells were divided into an experimental group transfected with a recombinant pcDNA3-hNIS plasmid and a control group transfected only with a pcDNA3 plasmid. The recombinant plasmid vector encoding the hNIS gene （pcDNA3-hNIS） was amplified, purified and identified. The hNIS gene was followed by DNA sequencing. A Western blot and an immunohistochemical assay were applied to detect the hNIS protein expression in the transfected human lung A549 cancer cells. RESULTS Restriction enzyme digestion and DNA sequencing results showed the size and direction of the inserted gene in the recombinant pcD- NA3-hNIS plasmid was correct. The Western blot method and immunohistochemical analysis showed a positive NIS protein expression in the experimental group. The NIS protein was detected mainly in the cell membranes showing a positive rate up to 70.6% with no expression of the NIS protein in the control group. There was a significant difference between two groups （P=0.000）. CONCLUSION The hNIS gene was transfected effectively into human lung A549 cancer cells mediated by Lipofectamine 2000, and was expressed with its protein in vitro.

Oncoprotein expression and inhibition of apoptosis during colorectal tumorigenesis

AIMS To study bcl-2 and P53 protein expression and inhibition of apoptosis during colorectal tumorigenesis. METHODS Expression of bcl -2 and p53 in 45 colorectal ade- nomas and 61 colorectal carcinomas was detected by immunohis- tochemical staining. RESULTS The bcl-2 and P53 protein expression was uniformly negative in normal mucosa,whereas bcl-2 and p53 positive rates were significantly higher in adenoma and carcinoma than in nor- reals(P<0.01 ).The area with strong bcl-2 expression was of- ten the area with severely dysplasia.In colorectal adenoma,ex- pression of p53 increased with the increasing size and dysplasia, in adenomas≥20 mm being higher than adenomas<10 mm(77, 8% vs 35.0%,P<0.05).p53 was relevant to differentiation and Duke's staging.A significant inverse correlation was found between bcl-2 and p53 in immunostaining in the adenomas,but not in the carcinomas.Furthermore,carcinomas with a high per- centage of bcl-2 positive cells were significantly more likely to have low rates of apoptosis. CONCLUSIONS These results suggest that bcl-2 gene appears to be an early event in colorectal tumorigenesis that can inhibit apoptosis,p53 expression plays an important role in the develop- ment and malignant change of colorectal adenoma,bcl-2 and p53 may be used as a good marker relating to cell apoptosis.

EFFECTS OF GENISTEIN ON INVASION AND MATRIX METALLOPROTEINASE ACTIVITIES OF HT1080 HUMAN FIBROSARCOMA CELLS

Effects of genistein on invasion and matrix metalloproteinase activities were investigated in HT1080 human sarcoma cells.Invasion of HT1080 cells through reconstituted basement membrane was inhibited when the cells were treated with 100 μ mol/L and 200 μ mol/L genistein.At the same concentrations,genistein not only suppressed latent forms of matrix metalloprotinese 2 and 9(MMP 2 and MMP 9) to convert into active forms,but also increase dramatically the tissue inhibitor of metalloproteinase(TIMP 1) mRNA contents and reverse the imbalance of MMPs and TIMPs.However,expressions of MMP 2 and MMP 9 were not significantly affected.Suppression of MMP activation and increase of TIMP 1 expression will decrease matrix degradation by MMPs,and consequently inhibit invasions of the cells.These results emphasized the existence of the imbalance between MMPs and TIMPs in tumor invasion and metastasis formation.The value of genistein as a drug for antiinvasion and anti metastasis chemotherapy was suggested.

Effect of gemcitabine on the expression of apoptosis-related genes in human pancreatic cancer cells

AIM： TO investigate the expression of genes involved in the gemcitabine-induced cytotoxicity in human pancreatic cancer cells. METHODS： A human pancreatic cancer cell line, PANC-1, was cultured. 1 × 10^4 PANG-1 cells were plated in 96-well microtiter plates. After being incubated for 24 h, gemcitabine was added to the medium at concentrations ranging 2.5 -1 000 mg/L. The AlamarBlue dye method was used for cell growth analysis. DNA fragmentation was quantitatively assayed using a DNA fragmentation enzyme-linked immunosorbent assay （ELISA） kit. PAP and TP53INP1 mRNA expression was determined using the reverse transcription-polymerase chain reaction with semi-quantitative analysis. The expression of GSK-3β and phospho-GSK-3β proteins was examined with Western blot analysis. RESULTS： The IC50 for the drug after a 48-h exposure to gemcitabine was 16 mg/L. The growth of PANC-1 cells was inhibited by gemcitabine in a concentration-dependent manner （P〈 0.0001） and the cell growth was also inhibited throughout the time course （P〈0.0001）. The DNA fragmentation rate in the gemcitabine-treated group at 48 h was 44.7 %, whereas it was 25.3 % in the untreated group. The PAP mRNA expression was decreased after being treal^l with gemcitabine, whereas the TP53INP1 mRNA was increased by the gemcitabine treatment. Western blot analysis showed that phospho- GSK-3βZ^ser9 was induced by the gemcitabine treatment. CONCLUSION： Gemcitabine suppresses PANC-1 cell proliferation and induces apoptosis. Apoptosis is considered to be associated with the inhibition of PAPand GSK-3β, and the activation of TP53INP1 and pospho- GSK-3β^ser9 .

Clinicopathological and Genetic Study of an Atypical Renal Hemangioblastoma

HEMANGIOBLASTOMA（HB）,a kind of benign tumor with uncertain histogenesis,is Characterized by the presence of stromal cells（SCs）and a rich vascular component.1 It occurs sporadi cally,

Mesenchymal stromal cells' role in tumor microenvironment:involvement of signaling pathways

Mesenchymal stromal cells(MSCs) are adult multipotent stem cells residing as pericytes in various tissues and organs where they can differentiate into specialized cells to replace dying cells and damaged tissues. These cells are commonly found at injury sites and in tumors that are known to behave like "wounds that do not heal." In this article, we discuss the mechanisms of MSCs in migrating, homing, and repairing injured tissues. We also review a number of reports showing that tumor microenvironment triggers plasticity mechanisms in MSCs to induce malignant neoplastic tissue formation, maintenance, and chemoresistance, as well as tumor growth. The antitumor properties and therapeutic potential of MSCs are also discussed.

Roles of Rap1 signaling in tumor cell migration and invasion

Ras-associated protein-1 （Rapl）, a small GTPase in the Ras-related protein family, is an important regulator of basic cellular functions （e.g., formation and control of cell adhesions and junctions）, cellular migration, and polarization. Through its interaction with other proteins, Rapl plays many roles during cell invasion and metastasis in different cancers. The basic function of Rapl is straightforward; it acts as a switch during cellular signaling transduction and regulated by its binding to either guanosine triphosphate （GTP） or guanosine diphosphate （GDP）. However, its remarkably diverse function is rendered by its interplay with a large number of distinct Rap guanine nucleotide exchange factors and Rap GTPase activating proteins. This review summarizes the mechanisms by which Rap 1 signaling can regulate cell invasion and metastasis, focusing on its roles in integrin and cadherin regulation, Rho GTPase control, and matrix metalloproteinase expression.

Role of the receptor for advanced glycation end products in hepatic fibrosis

AIM： To study the role of advanced glycation end products （AGE） and their specific receptor （RAGE） in the pathogenesis of liver fibrogenesis. METHODS： In vitro RAGE expression and extracellular matrix-related gene expression in both rat and human hepatic stellate cells （HSC） were measured after stimulation with the two RAGE ligands, advanced glycation end product-bovine serum albumin （AGE- BSA） and N＇-（carboxymethyl） lysine （CML）-BSA, or with tumor necrosis factor-α （TNF-α）. In vivo RAGE expression was examined in models of hepatic fibrosis induced by bile duct ligation or thioacetamide. The effects of AGE-BSA and CML-BSA on HSC proliferation, signal transduction and profibrogenic gene expression were studied in vitro. RESULTS： In hepatic fibrosis, RAGE expression was enhanced in activated HSC, and also in endothelial cells, inflammatory cells and activated bile duct epithelia. HSC expressed RAGE which was upregulated after stimulation with AGE-BSA, CML-BSA, and TNF-α.RAGE stimulation with AGE-BSA and CML-BSA did not alter HSC proliferation, apoptosis, fibrogenic signal transduction and fibrosis- or fibrolysis-related gene expression, except for marginal upregulation of procollagen α1（ I ） mRNA by AGE-BSA. CONCLUSION： Despite upregulation of RAGE in activated HSC, RAGE stimulation by AGE does not alter their fibrogenic activation. Therefore, RAGE does not contribute directly to hepatic fibrogenesis.

Esophagus and regenerative medicine

In addition to squamous cell carcinoma,the incidence of Barrett's esophagus with high-grade dysplasia and esophageal adenocarcinoma is rapidly increasing worldwide.Unfortunately,the current standard of care for esophageal pathology involves resection of the affected tissue,sometimes involving radical esophagectomy.Without exception,these procedures are associated with a high morbidity,compromised quality of life,and unacceptable mortality rates.Regenerative medicine approaches to functional tissue replacement include the use of biological and synthetic scaffolds to promote tissue remodeling and growth.In the case of esophageal repair,extracellular matrix(ECM) scaffolds have proven to be effective for the reconstruction of small patch defects,anastomosis reinforcement,and the prevention of stricture formation after endomucosal resection(EMR).More so,esophageal cancer patients treated with ECM scaffolds have shown complete restoration of a normal,functional,and disease-free epithelium after EMR.These studies provide evidence that a regenerative medicine approach may enable aggressive resection of neoplastic tissue without the need for radical esophagectomy and its associated complications.

Phosphorylation and biosynthesis of high molecular weight proteins of tumor nuclear matrix

Our previous studies showed a predominance of high molecular weight protein group in tumor nuclear matrices. Contrary to normal cells, proteins of this group are preferentially phosphorylated. Phosphoproteins of hepatoma nuclear matrix are selectively subjected to rapid proteolysis. By alkali treatment and a monoclonal antibody against phosphotyrosyl residue the presence of two high molecular weight bands of phosphotyrosyl-containing proteins was detected in nuclear matrices of tumor but not of normal liver cells. High molecular weight protein group of tumor nuclear matrices revealed also a rapid turnover and preferential incorporation of labeled amino acids selectively inhibited by chloramphenicol.

Experimental study on siRNA expressing vector-based RNA interference targeting c-Myc in human hepatocellular carcinoma cell line BEL-7402

Objective: To explore the inhibition of the expression of c-Myc in human hepatocellular carcinoma via vector-based RNA interference (RNAi). Methods: Two RNA interference DNA templates targeting c-Myc oncogene were designed via online software and synthesized. By ligation, the fragments were inserted into pSilencer 1.0-U6 to construct the recombinant siRNA expressing plasmids. The identified recombinants were introduced into BEL-7402 cells with Lipofactamine. The inhibition of c-Myc expression, together with the expression of CDK4, hTERT and Gadd45β in c-Myc down-regulated BEL-7402 cells, were analyzed by semi-quantitative RT-PCR. Results: Two recombinant plasmids pSic-myc-1 and pSic-myc-2, which direct the yields of siRNAs targeting c-Myc in cells, were constructed. Among which, pSic-myc-2 was shown to trigger a RNAi-mediated inhibition of expression of c-Myc in BEL-7402 by up to 90%. In c-Myc knockdown BEL-7402 cells, the expression of CDK4 and hTERT were down-regulated with a ratio of 85% and 57%, respectively, while the expression of Gadd45β was up-regulated by up to 110%. Conclusion: The expression of c-Myc in BEL-7402 could be suppressed by vector-based RNA interference successfully. The knockdown of c-Myc in turn resulted in the changes of expression of genes related to cell proliferation and apoptosis. Thus, our study provided a preliminary data in searching of a c-Myc-targeted RNAi therapy of human hepatocellular carcinoma.

The Influence of Glucose on Numerical Simulation of a Vascular Solid Tumor Growth

Glucose is the mainly nutrient substances in tumor growth,which played an important role in tumor cells' growth,proliferation and immigration.Numerical simulation will help a good understanding for the influence of glucose which affected on a vascular solid tumor growth.We present a hybrid on-Lattice Model to simulate the influence of glucose on a-vascular tumor growth.The hybrid model we developed focuses on five key variables implicated in the invasion process:tumor cells,extracellular matrix,matrix-degradative enzymes,oxygen and glucose.And about the discrete model,we consider cell evolution dynamics on cell level.Results indicate that the number of proliferation and quiescent cells is decreasing by decreasing the initial glucose concentration,consequently increase necrotic area relatively.Thus there is inhabitation effect on tumor growth by decreasing initial glucose concentration.

Oncologic Trogocytosis Protects Tumour Stromal Cells from γδ Cell Cytotoxicity

Tumours progressively develop chemoresistance and immunoescape abilities thanks to support from their stromal microenvironment. In ovarian carcinomas, for instance, tumour-associated mesenchymal stem cells （TAMC） can transfer multi-drug-resistant proteins to develop metastases. However, since the microenvironment of such carcinomas is frequently infiltrated by both TAMC and γδ T lymphocytes, the consequences of interactions between these cell types were unclear. Here, we report that whilst γδ T lymphocytes were not activated when co-incubated in vitro with TAMC, their cell membranes were trogocytosed by the TAMC. Since TAMC constitutively express a low level of HLA class I, which is increased by trogocytosis of γδ cell-derived HLA class I, the interaction increased the expression of HLA class-I molecules on TAMC. In addition, γδ T lymphocytes are HLA-unrestricted cytolytic cells and their activity is regulated by inhibitory receptors （KIR） for self-HLA class I. Hence, although the lytic activity of γδ T lymphocytes for unrelated target cells was unaffected by trogocytosis, it spared the TAMC. Therefore, interactions between TAMC and cytolytic γδ T cells avoided the killing of these stromal cells due to an active transfer of their protective HLA class-I molecules. These results suggest that trogocytosis contributes to the maintenance of cancer-associated stromal cells.

Effects of carbon dioxide pneumoperitoneum on metastasis of ovarian carcinoma cell line SKOV_3

The long suspicion of the potential harm of carbon dioxide （CO2） pneumoperitoneum exists in laparoscopic cancer surgery. For better understanding of this problem, we targeted this study at the effects of CO2 pneumoperitoneum on the invasive ability of ovarian carcinoma cell line and the possible mechanism within it. To study the effects of CO2 pneumoperitoneum on carcinoma cell, SKOV3 cells were divided into 2 groups, respectively exposed to pneumoperitoneal CO2-insuffiation and normal conditions. To study the possible mechanism, SKOV3 cells were divided into 3 groups, one was exposed to CO2 pneumoperitoneum, one to N2 and the other to normal conditions served as control. The in vitro adhesive and invasive ability of the cells was analyzed through 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyl tetrazolium bromide （MTT） assay and Boyden filters metastasis model; the expressions of vascular endothelial growth factor C （VEGF-C） and matrix metalloproteinase 9 （MMP9） were determined by reverse transcription polymerase chain reaction（RT-PCR）, and Western blot. We found that the adhesive ratio of SKOV3 cells exposed to CO2 was significantly higher than that of the control group; cells exposed to CO2 invaded the matrigel with a greater number （P〈0.01）; the expression of VEGF-C exposed to both CO2 and N2 was significantly increased compared with control group （P〈0.05）; the MMP9 expression level of CO2 group was higher than that of N2 group, P〈0.05. We concluded that carbon dioxide pneumoperitoneum may improve the adhesive and invasive ability of ovarian carcinoma cell line in vitro and CO2 can also be an independent factor to stimulate the expression of MMP9.

Chromosome 14q may harbor multiple tumor suppressor genes in primary glioblastoma multiforme

OBJECTIVE: To evaluate whether deletion of chromosome 14q is involved in the carcinogenesis of primary glioblastoma multiforme and to identify possibly common deletion regions. METHJODS: Fourteen fluorescent dye-labeled polymorphic markers were used and polymerase chain reaction-based microsatellite analysis was employed to investigate loss of heterozygosity (LOH) on chromosome 14q in 20 primary glioblastoma multiforme (GBM). RESULTS: Ten of twenty (50%) GBM displayed LOH at one or more of the markers on chromosome 14q. Five tumors showed either LOH or non-informative on all markers tested. The most frequent LOH was observed at locus D14S65 (57.1%) on 14q32.1, and in the chromosomal region spanning from D14S63 (47.1%) to D14S74 (46.7%) on 14q23-31. None of the informative loci exhibited microsatellite instability. CONCLUSIONS: Allelic deletion on chromosome 14q plays an important role in the pathogenesis of GBM. Chromosomal regions at locus D14S65 on 14q32.1 and spanning from D14S63 to D14S74 on 14q23-31 may harbor multiple tumor suppressor genes associated with GBM.