At the gliovascular interface, reciprocal inductive influences between brain microvascular endothelial cells (BMVEC) and astrocytes occur. Most of the knowledge in this area of research is derived from in vitro eo-culture models in which astrocytes are cultured on a stiff, two-dimensional (2D) surface. Three-dimensional (3D) culture models closely mimic the in lJZVO cellular architecture and they bridge the gap between 2D culture models and animal models. Hence, an in vitro 3D eo-culture model was developed and characterised, to study the interactions between BMVEC and astrocytes. In this model, human astrocytes (HA) were seeded inside a collagen type--I gel while human immortalised cerebral microvascular endothelial cells (hCMEC/D3) were cultured on the gel surface. Both cell types were of human origin to improve the translatability of findings to humans in vivo. Additional important features of the model are the culture of endothelial cells on a soft matrix, and the simulation of the geometric relationship that exists in vivo i.e., the interaction of astrocytes with BMVEC from their ab luminal side. To determine the effect of the 3D environment on the HA, the proliferation rate and expression of four molecules namely, glial fibrillary acidic protein (GF AP), aquaporin-4 (AQP4), endothelin-l (Et-l) and endothelin receptor type-B (EDNRB), were compared between 3D and 2D cultured HA. The decreased expression of AQP4 and EDNRB and the much-decreased proliferation rate of 3D HA suggested their reduced reactivity and a similarity to their in lJiVO counterparts. However, 3D HA did not differ from the 2D HA in their ability to release soluble factors that induce barrier properties on BMVEC, as observed by similar levels of three expression markers of barrier phenotype on hCMEC/D3 cells namely, zonula occludens-l, claudin-5, and P-glycoprotein and similar paracellular permeability coefficients to fluorescent-dextrans (70 kDa).
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:582802 |
| Date | January 2012 |
| Creators | Peddagangannagari, Sreekanth Reddy |
| Publisher | Open University |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://oro.open.ac.uk/54831/ |
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