Human bone marrow stromal cells have mitogenic activity on SK-Hep-1 cells.

Siu, Yeung Tung.Thesis (M.Phil.)--Chinese University of Hong Kong, 2001.Includes bibliographical references (leaves 65-75).Abstracts in English and Chinese.Title Page --- p.iAbstract in English --- p.iiAbstract in Chinese --- p.iiiAcknowledgement --- p.ivTable of Contents --- p.v-viiiList of Figures --- p.ixList of Tables --- p.xAbbreviations --- p.xi-xiiChapter Chapter 1 --- IntroductionChapter 1.1 --- Growth factors involved in hepatocytes proliferation --- p.1-6Chapter 1.1.1 --- Hepatocyte growth factor (HGF) --- p.1Chapter 1.1.2 --- Tumor necrosis factor-a (TNF-α) and interleukin-6 (IL-6) --- p.2Chapter 1.1.3 --- Epidermal growth factor (EGF) and transforming growth factor-α (TGF-α) --- p.3Chapter 1.1.4 --- Other comitogens --- p.4Chapter 1.1.5 --- Transforming growth factor-β (TGF-β) --- p.5Chapter 1.2 --- Bone marrow stromal cells and hepatocytes proliferation --- p.7-12Chapter 1.2.1 --- Role of bone marrow stromal cells in bone marrow --- p.7Chapter 1.2.2 --- Bone marrow as a source of hepatic oval cells --- p.8Chapter 1.2.3 --- Growth factors secreted by bone marrow stromal cells involved in hepatocytes proliferation --- p.9Chapter 1.2.4 --- Endocrine in hepatocytes proliferation --- p.12Chapter 1.3 --- Objective of this study --- p.13-15Chapter Chapter 2 --- Materials and MethodsChapter 2.1 --- Cell cultures --- p.16Chapter 2.2 --- Selection of human hepatic cell line for the detection of mitogenic activity --- p.17-18Chapter 2.2.1 --- "Enrichment of human hepatic cell lines, Hep 3B, Hep G2, C3A, SK-Hep-1 and Chang cells at G0-G1 phases by serum deprivation" --- p.17Chapter 2.2.2 --- "Incubation of serum deprived Hep 3B, Hep G2, C3A, SK- Hep-1 and Chang cells with mitogenic stimuli" --- p.17Chapter 2.2.3 --- Cell cycle analysis by flow cytometry using propidium iodide staining --- p.17Chapter 2.3 --- "Detection of mitogenic activity of human bone marrow stromal cells on the selected cell line, SK-Hep-1 cells" --- p.18-20Chapter 2.3.1 --- Partially growth arrested human SK-Hep-1 cells --- p.18Chapter 2.3.2 --- Human bone marrow stromal cells --- p.19Chapter 2.3.2.1 --- Bone marrow stromal cellular extract --- p.19Chapter 2.3.2.2 --- Total protein assay --- p.19Chapter 2.3.3 --- Incubation of SK-Hep-1 cells with bone marrow stromal cellular extracts --- p.20Chapter 2.4 --- Characterization of hepatocyte mitogenic activity of bone marrow stromal cellular extract --- p.21-22Chapter 2.4.1 --- Dialysis --- p.21Chapter 2.4.2 --- Temperature treatment --- p.21Chapter 2.4.3 --- Proteolysis --- p.22Chapter 2.5 --- Performing a preliminary test on the difference between bone marrow stromal cellular extract and other growth factors --- p.22-26Chapter 2.5.1 --- Incubation of SK-Hep-1 cells with bone marrow stromal cellular extract or other growth factors --- p.22Chapter 2.5.2 --- Metabolic labeling of SK-Hep-1 cells with [32P]orthophosphate --- p.23Chapter 2.5.3 --- Incubation of labeled SK-Hep-1 cells with bone marrow stromal cellular extract or other growth factors --- p.23Chapter 2.5.4 --- SK-Hep-1 cells lysate extraction --- p.23Chapter 2.5.5 --- Two-dimensional electrophoresis --- p.24Chapter 2.5.5.1 --- First dimension isoelectric focusing --- p.24Chapter 2.5.5.2 --- Second dimension sodium dodecyl sulfate-polyacrylamide gel electrophoresis --- p.25Chapter 2.5.6 --- Amplification of radiolabeled signal by EN3HANCE --- p.25Chapter 2.5.7 --- Visualization of autoradiography --- p.26Chapter 2.5.8 --- Visualization by silver staining --- p.26Chapter Chapter 3 --- ResultsChapter 3.1 --- Selection of human hepatic cell line for the detection of mitogenic activity --- p.27-30Chapter 3.1.1 --- "Enrichment of human hepatic cell lines, Hep 3B, Hep G2, C3A, SK-Hep-1 and Chang cells at G0-G1 phases by serum deprivation" --- p.27Chapter 3.1.2 --- DNA synthesis of hepatic cell lines in response to 10 % FBS after serum deprivation --- p.29Chapter 3.2 --- "Detection of mitogenic activity of human bone marrow stromal cells on the selected cell line, SK-Hep-1 cells" --- p.31-39Chapter 3.2.1 --- Cell cycle distribution of partially growth arrested SK-Hep-1 cells in response to mitogens --- p.31Chapter 3.2.2 --- Time course on DNA synthesis of partially growth arrested SK-Hep-1 cells in response to FBS and bone marrow stromal cellular extract --- p.36Chapter 3.2.3 --- Dose response on DNA synthesis of partially growth arrested SK-Hep-1 cells in response to bone marrow stromal cellular extracts --- p.38Chapter 3.3 --- Characterization of hepatocyte mitogenic activity of bone marrow stromal cellular extract --- p.40-44Chapter 3.4 --- Performing a preliminary test on the difference between bone marrow stromal cellular extract and other growth factors --- p.45-49Chapter 3.4.1 --- Mitogenic response of SK-Hep-1 cells in response to bone marrow stromal cellular extract and other growth factors --- p.45Chapter 3.4.2 --- Early intracellular signaling of SK-Hep-1 cells in response to bone marrow stromal cellular extract and other growth factors --- p.47Chapter Chapter 4 --- DiscussionChapter 4.1 --- Selection of human hepatic cell line for the detection of mitogenic activity --- p.50Chapter 4.2 --- "Mitogenic activity of human bone marrow stromal cells on the selected cell line, SK-Hep-1 cells" --- p.51Chapter 4.3 --- Characterization of hepatocyte mitogenic activity of bone marrow stromal cellular extract --- p.52Chapter 4.4 --- Performing a preliminary test on the difference between bone marrow stromal cellular extract and other growth factors --- p.53Chapter 4.5 --- Possible directions for future investigation --- p.55Chapter 4.6 --- Conclusions --- p.56Chapter Chapter 5 --- AppendicesChapter 5.1 --- Reagents and solutiuons --- p.57-64Chapter 5.1.1 --- Selection of human hepatic cell line for the detection of mitogenic activity --- p.57Chapter 5.1.2 --- "Detection of mitogenic activity of human bone marrow stromal cells on the selected cell line, SK-Hep-1 cells" --- p.59Chapter 5.1.3 --- Characterization of hepatocyte mitogenic activity of bone marrow stromal cellular extract --- p.60Chapter 5.1.4 --- Performing a preliminary test on the difference between bone marrow stromal cellular extract and other growth factors --- p.61Chapter Chapter 6 --- References --- p.65-7