TISSUE IMAGING IN CELL BIOLOGY

点击翻译按钮获取中文摘要

Continuing to develop imaging and culturing techniques for three- dimensional multicellular systems, we have grown live blocks of human and mouse lymphoid tissue in a low shear force rotating wall vessel bioreactor (RWV). Fragments of lymphoid tissue remain viable in the RWV for at least two weeks. Cells from the tissue migrate into the surrounding media. Migration of cells out of the tissue is reversible: labelled cells from the peripheral media re-populate the tissue and their fate can be followed by imaging with confocal microscopy. Migration of cells may be partially prevented by embedding the tissue fragments in collagen gels. Cell migration is completely stopped when the tissues are embedded in agarose gels. Agarose-embedded blocks of tissue remain alive: they consume oxygen and glucose, produce IgG into the media and retain the basic elements of their architecture, including B cell-rich germinal centers surrounded by T cells. Tall densities are similar that of tissue blocks cultured on the top of the gels (histocultures). Confocal imaging of agarose-embedded blocks of tissue reveal a network of follicular-dendritic cells in germinal centers. Agarose-embedded blocks of tissues retain more cells and have less apoptotic nuclei than non-embedded controls. Agarose-embedded blocks of tissues are productively infected with HIV-1. Long-term culturing of lymphoid tissue together with peripheral lymphocytes in a two-compartment system, in combination with the imaging technique, allows us to simulate the high level of complexity of the immune system and to address basic questions of lymphocyte migration, homing and kinetics. In the absence of animal models for various immune disorders, including HIV infection leading to AIDS, the developed model provides a unique system to study pathogenesis in human lymphoid tissue.