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Role of CD15 and CD15s in the Cellular Mechanisms of Cancer Cell Metastasis from Lung to the Brain
Abstract and Figures
Non-small cell lung cancer is one of the most common primary tumours to metastasise to the brain in adults. The underlining molecular mechanisms of brain metastasis are still not fully understood. Interactions between brain endothelial cells and cancer cells play key roles in brain metastasis. CD15 and CD15s are cell-cell adhesion molecules which interact with E-selectin which is expressed on endothelial cells and known to be involved in the leukocyte homing process as well as being implicated in metastasis with many non-CNS neoplasms. The aim of this project was to investigate the role of CD15 and CD15s in cancer cell adhesion to brain endothelial cells and transendothelial migration of lung cancer cells during brain metastasis. Expression of CD15, CD15s and CD62E was characterised in human primary and brain metastatic lung cancer cells using immunocytochemistry, flow cytometry, Western blot and immunohistochemistry in human tissue sections. Effects of CD15 and CD15s expression on NSCLC metastatic to brain were assessed using genetic manipulation (overexpression and knockdown) followed by functional assays. Both CD15 and CD15s were overexpressed and knockdowned and cell-cell adhesion was then examined using qualitative and quantitative adhesion assays, under both static and flow physiological conditions. Trans-endothelial migration potential was also assessed using a voltometer, Electric Cell-Substrate Impedance sensing system and cell-monitoring system CellZscope™. Findings showed that CD15 and CD15s were prominently expressed on metastatic lung cancer cells (SEBTA-001, SEBTA-005 and NCI-H1299) and weakly expressed on both primary lung cancer cells (COR-L105 and A549) and brain endothelium (hCMEC/D3). The highest expression of CD62E was observed on brain endothelium stimulated with TNF-α (25pg/ml) (p<0.001). CD15, CD15s and CD62E expression was detected in human metastatic tissues. The absence of CD62E and immunoblocking and knockdown of CD15 and CD15s significantly reduced the adhesion of cancer cells under both static and shear stress conditions (p<0.0001).Overexpression of CD15 and CD15s significantly increased their adhesion on an endothelial monolayer (p<0.001). Metastatic cancer cells were able to transmigrate through a brain endothelial monolayer compared to primary and glioblastoma multiforme (GBM) cells. Knockdown of CD15 and CD15s decreased the trans-endothelial migration potential of cancer cells while even primary lung cancer cells and GBM cells transmigrated following overexpression of CD15 and CD15s. These results confirmed the correlation between CD15 and CD15s in adhesion as well as trans-endothelial migration of cancer cells during cerebral metastasis.
ISNI:0000 0004 6347 1408
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The belts of endothelial tight junctions, which impede diffusion between blood and brain, were reduced to fragmentary, small junctions in subcultured brain endothelium. When cocultured with the capillaries' nearest neighbor, the astrocytes, these endothelial tight junctions were enhanced in length, width, and complexity, as seen by en face views of the cell membranes with freeze-fracture electron microscopy. Gap junctions, common in brain endothelium in vitro but absent in mature brain capillaries in vivo, were markedly diminished in area from among the enhanced tight junctions of the cocultures. Thus, astrocytes in vitro play a role in the formation, extent, and configuration of the junctional complexes in brain endothelium, whose diffusion barrier may likewise be influenced by astrocytes in vivo.
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The American Joint Committee on Cancer's Cancer Staging Manual is used by physicians throughout the world to diagnose cancer and determine the extent to which cancer has progressed. All of the TNM staging information included in this Sixth Edition is uniform between the AJCC (American Joint Committee on Cancer) and the UICC (International Union Against Cancer).
In addition to the information found in the Handbook, the Manual provides standardized data forms for each anatomic site, which can be utilized as permanent patient records, enabling clinicians and cancer research scientists to maintain consistency in evaluating the efficacy of diagnosis and treatment. The CD-ROM packaged with each Manual contains printable copies of each of the book’s 45 Staging Forms.