PubMed · 3102897
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J Hed. 1986. Methods for distinguishing ingested from adhering particles.. https://doi.org/10.1016/s0076-6879(86)32008-1
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Parallel-plate flow chambers have been used to model the vascular microcirculation and study the in vitro dynamic adhesive interactions of leukocytes and human umbilical vein endothelial cells (HUVECs). We describe here a high capacity system which can simultaneously monitor the adhesive interaction of neutrophils and HUVECs in ten flow chambers. Automated data collection was achieved with an image analyzer controlling the autostage and autofocus attachments of an inverted microscope. Images from the flow chambers were captured via phase-contrast microscopy using a video camera and laser videodisk recorder. The images were downloaded off-line into an image analyzer for automated counting of rolling and adherent cells. Neutrophils were detected by their "phase bright' characteristics. An automated optimization procedure allowed the computer to choose the best setting for the selective detection of neutrophils. In addition, a method which utilized image averaging was used to distinguish between rolling and adherent cells. A comparison of the results obtained from the manual and automated counting methods revealed linear relationships for the counting of both adherent (r = 0.98) and rolling cells (r = 0.96) with counting efficiencies of 59% and 46%, respectively. The utility of the system was demonstrated by its ability to measure the adhesive interaction between neutrophils and HUVEC in response to stimulus such as interleukin-1 alpha (IL-1 alpha), histamine, or formyl-1-methionyl-1-leucyl-1-phenylalanine (fMLP). In conclusion, we have developed an automated assay which combines the capacity of ten flow chambers with a computerized data analysis system; the result is an efficient and reproducible assay which minimizes operator associated errors and biological variability.
A "green beard" refers to a gene, or group of genes, that is able to recognize itself in other individuals and direct benefits to these individuals. Green-beard effects have been dismissed as implausible by authors who have implicitly assumed sophisticated mechanisms of perception and complex behavioral responses. However, many simple mechanisms for genes to "recognize" themselves exist at the maternal-fetal interface of viviparous organisms. Homophilic cell adhesion molecules, for example, are able to interact with copies of themselves on other cells. Thus, the necessary components of a green-beard effect -- feature, recognition, and response -- can be different aspects of the phenotype of a single gene. Other green-beard effects could involve coalitions of genes at closely linked loci. In fact, any form of epistasis between a locus expressed in a mother and a closely linked locus expressed in the fetus has the property of "self-recognition." Green-beard effects have many formal similarities to systems of meiotic drive and, like them, can be a source of intragenomic conflict.
We have investigated the effect of integrin antibodies to a well-characterized alpha 5 beta 1 (fibronectin receptor) and to a multi-specific alpha 3 beta 1 (laminin, collagen, and fibronectin receptor), on the expression of matrix metalloproteases and the invasion ability of two human glioblastoma cell lines, SNB19 and U251. Cell adhesion assays indicated that both cell lines adhere to fibronectin, type IV collagen and laminin. Adhesion of cells to fibronectin was inhibited by a RGD peptide. Cells treated with anti-alpha 3 beta 1 or anti-alpha 5 beta 1 antibodies expressed increased levels of MMP-2. An in vitro matrigel assay also showed that the alpha 3 beta 1 antibody-treated cells had greater invasive ability than the controls. Immunofluorescence data showed that glioma cells treated with either anti-alpha 3 beta 1 or anti-alpha 5 beta 1 antibodies expressed diminished alpha 3 beta-1 and alpha 5 beta 1 integrins relative to the controls. The data show that treatment of cells with alpha 3 beta 1 antibody diminishes the integrin expression on the cell surface and increases the MMP-2 activity and invasiveness.