Enhancement of organogenesis with small molecule drugs

βββ -- using a bio-degradable scaffold. seed myocytes onto it. before scaffold breaks down, seed with angioblasts. use statins instead of VEGF to stimulate endothelial production. evaluate with fluoro microscopy.

Project Details and Methods

1. Grow mesenchyme cells in a Wharton's Jelly-like solution (significantly containing hyaluronan). Hyaluronan is a significant component of the extracellular matrix and has been shown to contribute to tumor growth. In this step, we need to grow a layer of mesenchyme cells roughly five cells-thick. Hopefully the Jelly will promote this growth pattern.

(Several day's growth...)

2. Transfect the cells with Invitrogen™ Qtracker® 505 dye.

3. Apply statins to layers of cells to induce differentiation.

(Several day's growth and differentiation...)

4. Transfect the cells with Invitrogen™ Qtracker® 655 dye.

5. Quickly freeze sections of the cell layers to enable preparation of thin cross-sectional slices of tissue for observation. Then view under fluorescent microscope.

Resources

• Researchers were able to grow rat ventricular cells in rotating bioreactors that quickly formed aggregates that rhythmically contracted in unison.
  • R.E. Akins, R.A. Boyce, M.L. Madonna, N.A. Schroedl, S.R. Gonda, T.A. McLaughlin, C.R. Hartzell. Tissue Engineering. April 1, 1999, 5(2): 103-118. doi:10.1089/ten.1999.5.103. [Article]

• Asahara T, et al (1997). "Isolation of putative progenitor endothelial cells for angiogenesis." Science 275: 964-7. PMID 9020076. [Article]

• Asahara T, et al (1999). "Bone marrow origin of endothelial progenitor cells responsible for postnatal vasculogenesis in physiological and pathological neovascularization." Circulation Research 85 (6): 221-8. PMID 10436164. [Article]

• Hristov, et al (2003). "Endothelial progenitor cells: isolation and characterization." Trends in Cardiovascular Medicine 13 (5): 201-6. PMID 12837583. [Article]

• Shaw J, et al (2004). "Hematopoietic stem cells and endothelial cell precursors express Tie-2, CD31 and CD45.". Blood Cells, Molecules, and Diseases 32 (1): 168-75. PMID 14757432. [Article]

• Werner N, et al (2005). "Circulating Endothelial Progenitor Cells and Cardiovascular Outcomes.". New England Journal of Medicine 353: 999-1007. PMID 16148285. [Article]

• A specific study of HMG-CoA reductase inhibitors (statins). The researchers found that the statins controlled hematopoietic progenitor cell differentiation.
  • Stefanie Dimmeler, Alexandra Aicher, Mariuca Vasa, Christiane Mildner-Rihm, Klaudia Adler, Michaela Tiemann, Hartmut Rütten, Stephan Fichtlscherer, Hans Martin and Andreas M. Zeiher. HMG-CoA reductase inhibitors (statins) increase endothelial progenitor cells via the PI 3-kinase/Akt pathway. J. Clin. Invest., 2001; 108(3): 391 - 397. [Article]

• The Pax3 protein induces dense multi-layer aggregation of mammalian mesenchymal cells and facilitates the mesenchymal to endothelial transition.
  • Wiggan O'Neil, Mark Fadel, and Paul Hamel. Pax3 induces cell aggregation and regulates phenotypic mesenchymal-epithelial interconversion. Journal of Cell Science 115, 517-529 (2002) [Article]

• Pax3 protein production in E. coli.
  • Stéphane C. Boutet, Marie-Hélène Disatnik, Lauren S. Chan, Kevin Iori and Thomas A. Rando. Regulation of Pax3 by Proteasomal Degradation of Monoubiquitinated Protein in Skeletal Muscle Progenitors. Cell 130, 342-362 (2007) [Article]

People

• Eddie Liu
• Zach Bjørnson

Steps (not for final page)

• http://science.nasa.gov/headlines/y2002/14feb_heart.htm
• http://www.nature.com/nbt/journal/v23/n10/full/nbt1005-1237.html
• http://www.nibib.nih.gov/HealthEdu/eAdvances/31May06
• http://www3.interscience.wiley.com/cgi-bin/fulltext/112395581/PDFSTART

• looking at cells in layers
• scaffolds
• statins and other signals for vasculogensis