2026-09-20T00:11:38Zhttps://dspace.mit.edu/server/oai/requestoai:dspace.mit.edu:1721.1/180452022-01-13T07:53:48Zcom_1721.1_7582com_1721.1_7581col_1721.1_131023
Linda Griffith.
Schreiber, Brent M. (Brent Matthew), 1981-
Massachusetts Institute of Technology. Biological Engineering Division.
Massachusetts Institute of Technology. Department of Biological Engineering
2005-06-02T19:45:25Z
2005-06-02T19:45:25Z
2004
http://hdl.handle.net/1721.1/18045
57364170
Thesis (M. Eng.)--Massachusetts Institute of Technology, Biological Engineering Division, 2004.
Includes bibliographical references (leaves 84-93).
Non-parenchymal cells (NPC's) are integral to recreate the native hepatic microenvironment and necessary to maintain in vivo liver function. A variety of in vitro culture systems have been developed to address different aspects of liver physiology and architecture in order to recreate the microenvironment. These in vitro co-culture strategies have been limited by their inability to systematically characterize the addition of non-parenchymal cells. In this dissertation, I use gene expression levels quantified by real-time RT-PCR to determine tissue composition. The identified genes demonstrate significant cell-type specificity, magnitude, and stability of expression in vivo and throughout each step of cell isolation process. In the course of this development, we establish protocols to accurately isolate and count an enriched fraction of primary NPC's. Experiments on the perfusion and isolation process prove that there exists an inverse correlation between perfusion flow rate and NPC yield and viability. Further, we have characterized the tissue composition of each step in the cell isolation process and the resulting NPC population to confirm that a significant number of each NPC type is delivered to in vitro co-culture. System output analysis of spheroids co-cultured at physiological ratios and seeded into the milliF bioreactor shows the presence of stellate cells, but the absence of endothelial (EC) and kupffer cells (KC). The same analysis of 2D collagen gel sandwiches shows the presence of all NPC cell types. This indicates that our process is currently limited by the ability of EC's and KC's to incorporate into spheroid aggregates. Future work that validates the temporal expression stability of the identified genes in different in vitro culture systems
(cont.) and environments will enable determination of relative levels of NPC incorporation and will allow correlations to be made between operational features of in vitro systems, the resulting culture microenvironment, and observed tissue function.
by Brent M. Schreiber.
M.Eng.
120 leaves
8006001 bytes
8020925 bytes
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eng
Massachusetts Institute of Technology
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http://dspace.mit.edu/handle/1721.1/7582
Biological Engineering Division.
Use of gene expression to characterize heterogeneous liver cell populations
Characterization of heterogeneous hepatic cell population via use of gene expression
Thesis
application/pdf
Publication
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<Abstract>Non-parenchymal cells (NPC's) are integral to recreate the native hepatic microenvironment and necessary to maintain in vivo liver function. A variety of in vitro culture systems have been developed to address different aspects of liver physiology and architecture in order to recreate the microenvironment. These in vitro co-culture strategies have been limited by their inability to systematically characterize the addition of non-parenchymal cells. In this dissertation, I use gene expression levels quantified by real-time RT-PCR to determine tissue composition. The identified genes demonstrate significant cell-type specificity, magnitude, and stability of expression in vivo and throughout each step of cell isolation process. In the course of this development, we establish protocols to accurately isolate and count an enriched fraction of primary NPC's. Experiments on the perfusion and isolation process prove that there exists an inverse correlation between perfusion flow rate and NPC yield and viability. Further, we have characterized the tissue composition of each step in the cell isolation process and the resulting NPC population to confirm that a significant number of each NPC type is delivered to in vitro co-culture. System output analysis of spheroids co-cultured at physiological ratios and seeded into the milliF bioreactor shows the presence of stellate cells, but the absence of endothelial (EC) and kupffer cells (KC). The same analysis of 2D collagen gel sandwiches shows the presence of all NPC cell types. This indicates that our process is currently limited by the ability of EC's and KC's to incorporate into spheroid aggregates. Future work that validates the temporal expression stability of the identified genes in different in vitro culture systems</Abstract>
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