The evolution of chemotaxis assays from static models to physiologically relevant platforms.

The role of chemotactic gradients in the immunological response is an area which elicits a lot of attention due to its impact on the outcome of the inflammatory process. Consequently there are numerous standard in vitro designs which attempt to mimic chemotactic gradients, albeit in static conditions, and with no control over the concentration of the chemokine gradient. In recent times the design of the standard chemotaxis assay has incorporated modern microfluidic platforms, computer controlled flow devices and cell tracking software. Assays under fluid flow which use biochips have provided data which highlight the importance of shear stress on cell attachment and migration towards a chemokine gradient. However, the in vivo environment is far more complex in comparison to conventional cell assay chambers. The designs of biochips are therefore in constant flux as advances in technology permit ever greater imitations of in vivo conditions. Researchers are focused on designing a generation of new biochips and enhancing the physiological relevance of the current assays. The challenge is to combine a shear flow with a 3D scaffold containing the endothelial layer and permitting a natural diffusion of chemokines through a tissue-like basal matrix. Here we review the latest range of chemotaxis assays and assess the innovative features of their designs which enable them to better imitate the in vivo environment. We also present some alternative designs that were initially employed in tissue engineering which could potentially be used in the establishment of novel chemotaxis assays.

A new microtubule-targeting compound PBOX-15 inhibits T-cell migration via post-translational modifications of tubulin.

The ordered, directional migration of T-lymphocytes is a key process during immune surveillance, immune response, and development. A novel series of pyrrolo-1,5-benzoxazepines have been shown to potently induce apoptosis in variety of human chemotherapy resistant cancer cell lines, indicating their potential in the treatment of both solid tumors and tumors derived from the hemopoietic system. Pyrrolobenzoxazepine 4-acetoxy-5-(1-naphtyl)naphtho[2,3-b]pyrrolo[1,2-d][1,4]-oxazepine (PBOX-15) has been shown to depolymerize tubulin in vitro and in the MCF7 breast cancer cell line. We hypothesized that this may suggest a role for this compound in modulating integrin-induced T-cell migration, which is largely dependent on the microtubule dynamics. Experiments were performed using human T lymphoma cell line Hut78 and peripheral blood T-lymphocytes isolated from healthy donors. We observed that human T-lymphocytes exposed to PBOX-15 have severely impaired ability to polarize and migrate in response to the triggering stimulus generated via cross-linking of integrin lymphocyte function associated antigen-1 receptor. Here, we show that PBOX-15 can dramatically impair microtubule network via destabilization of tubulin resulting in complete loss of the motile phenotype of T-cells. We demonstrate that PBOX-15 inhibitory mechanisms involve decreased tubulin polymerization and its post-translational modifications. Novel microtubule-targeting effects of PBOX-15 can possibly open new horizons in the treatment of overactive inflammatory conditions as well as cancer and cancer metastatic spreading.

Cytoskeletal reorganization and altered phagocytotic ability in primary cultures of rainbow trout hemopoietic tissue exposed to low-level ionizing radiation.

It has long been known that the hematopoietic tissue of mammals is one of the most radiosensitive tissues. In vitro studies on prawns have also shown that low doses of radiation have an extremely deleterious effect on cells cultured from this animal's blood-forming tissues. This raises questions about the relative effects of radiation in animals of different species. One of the most important aquatic animals, from both an economic and an ecological point of view, is the fish. With this in mind, primary cultures of the blood-forming tissues of rainbow trout were exposed to radiation followed by a morphological comparison between control and irradiated cultures. The cultured cells were characterized as macrophages after incubation with apoptotic human polymorphonuclear leukocytes and were classified as phagocytotic leukocytes. These cells were found in two morphological forms, stretched and rounded. It was shown that there was a commensurate increase in the number of stretched cells after irradiation. Radiation was also shown to cause a dose-dependent increase in the amounts of apoptosis in these cells over time. The phagocytotic efficacy of these cells was shown to inhibited by the exposure to low doses of radiation.