The long term objectives of this research are to develop methods for measuring the major physiological aspects of cellular motility, namely chemotaxin reception, cell orientation and cell movement for individual cells and to apply these methods as clinically useful tests. During the first years of this grant we developed an image analysis assay for measuring cellular orientation and motility. This renewal application is designed to extend this capability to include measurement of chemotaxin receptor sites and to test our current assay system's ability to differentiate bacterial sepsis from nonseptic inflammation. The first studies use tetramethyl rhodamine isothiocyanate (TRITC) labeled formyl methionyl leucyl phenylalanine lysine (FMLPL) as the chemoattractant and computer programs written in FORTRAN and C to quantitate cellular fluorescence at three 10 second time periods during a six minute exposure to TRITC-FMLPL. Cellular orientation, chemokinesis and chemotaxis are measured by phase contrast microscopy throughout the six minute period except for the three ten second intervals. The results should allow correlation of receptor function, orientation and motility thereby improving understanding of these physiological interrelationships. The second set of studies determines if the already developed methods for precisely measuring cellular chemokinesis and chemotaxis can detect the septic state in patients with severe burns and trauma and patients with bacterial infections and if abnormalities in PMM motility are related to infection in patients with ARC and AIDS. Longitudinal studies will test the hypothesis that in vitro measurements-of chemotaxis using formyl methionyl leucyl phenylalanine (FMLP) for the chemoattractant are impaired to a significantly greater degree during septic periods because FMLP like products released by proliferating bacteria occupy cellular receptor sites. Similar longitudinal studies are proposed for patients with ARC and AIDS to determine if abnormalities in PMN motility contribute to the enhanced susceptibility to infection of these patients. A third set of experiments will improve the computer assisted assay of cell motion by increasing the number of cells tracked in the assay. Cellular motility will be followed with low power optics (100X) using newly written computer programs. This system which w.11 simultaneously track.100 or more cells will improve the assay's sensitivity and practicality.
| Dickover, R E; Donovan, R M; Goldstein, E et al. (1992) Decreases in unintegrated HIV DNA are associated with antiretroviral therapy in AIDS patients. J Acquir Immune Defic Syndr 5:31-6 |
| Donovan, R M; Dickover, R E; Goldstein, E et al. (1991) HIV-1 proviral copy number in blood mononuclear cells from AIDS patients on zidovudine therapy. J Acquir Immune Defic Syndr 4:766-9 |
| Hart, L; Donovan, R M; Goldstein, E et al. (1990) Detection of human immunodeficiency virus in infected CEM cells using fluorescent DNA probes and a laser-based computerized image cytofluorometry system. Anal Quant Cytol Histol 12:127-34 |
| Kim, Y; Goldstein, E; Lippert, W et al. (1989) Polymorphonuclear leucocyte motility in patients with severe burns. Burns 15:93-7 |
| Donovan, R M; Goldstein, E; Kim, Y et al. (1987) A computer-assisted image-analysis system for analyzing polymorphonuclear leukocyte chemotaxis in patients with diabetes mellitus. J Infect Dis 155:737-41 |
