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Embryonic stem cells (ESCs) have the capacity to differentiate into every cell type in the body, and therefore can theoretically be used to generate cells and tissues to cure a variety of diseases. Our client in the Odorico Lab (Department of Surgery) has derived foregut-committed cell lines from ESCs (which correspond to progenitor cells of the gut region that develops primarily into pancreas) and would like to differentiate these ESCs into insulin-producing pancreatic beta-like cells. These cells could replace or supplement transplanted donor beta cells. The mechanisms required to differentiate ESCs into these pancreatic cells is currently unknown, and this device would aid in researching such mechanisms. Our client would like to test a large number of growth factors for their ability to affect conversion of these precursor cells to mature insulin-secreting cells. In addition, a recapitulation of the 3-dimentional embryonic environment to prompt cells to adopt a pancreatic cell fate, perhaps using a Matrigel substrate, is desirable. A small scale cell culture using microfluidics to set up growth factor gradients is one approach that could be successful.
From left to right: Kyle Herzog, Tim Pearce, Jonathan Baran, Dhaval Desai
Fully Formed Gradient using Texas Red labeled Dextran
MATLAB Image processing, shows the intensity of light over the length of the channel, for the fully formed gradient seen above. Shows a linear gradient was formed.
Time Lapse Series of Pictures showing Gradient Formation
Currently, we have tested our microfluidic device with a Texas Red dye to verify the proof of concept. Testing will then proceed with the use of actual growth factors and cell embedded in Matrigel.
| Week | Reporting Period Beginning | Activities |
|---|---|---|
| 1 | September 7 | Met with our client Dr. Browning and she described that she is looking for a way to grow stem cells in a environment with differing concentrations of growth factors. Our initial idea is that microfluidics is the best possible design idea, so we have contacted Dr. Beebe who is a specialist in this area. |
| 2 | September 14 | Our group met with Dr. Beebe and he told us about the two different systems to set up concentration gradients, the flow and no-flow system. Each of these systems have their own advantages and disadvantages, which our client needs to decide on before we can proceed. |
| 3 | September 21 | We met with Dr. Browning and discussed the advantages and disadvantages of the flow vs. no-flow system. It is apparent through discussion that the best option is the no flow system, since this will allow autocrine and paracrine signaling to occur in the experimental setup. |
| 4 | September 28 | We contacted Dr. Beebe to try to begin construction of an initial no-flow system. The system will be primarily based upon Dr. Beebe’s experimental system. The initial construction will allow us to view a working system and then allow us to develop our own system which we can implement for our client. |
| 5 | October 5 | We met with Erwin (BME Graduate student) and he went over the fabrication process with us and told us of things we need to consider when designing our device. He showed us the lab space and also assured us that Dr. Beebe would let us use his lab and supplies when we are ready to begin making devices. |
| 6 | October 12 | We continued to finalize design constraints including channel dimensions, sink and source volumes, and the time to setup up the concentration gradient. We also worked on completing our mid-semester presentation. |
| 7 | October 19 | We met as a group to finalize our design, and while working through the calculations we decided that a 3-d environment would be best to house the cells. This would simpify the setup and allow the cells to be cultured in a 3-d matrix. Erwin will be consulted ASAP to ensure that this option is viable and then a mask will be made. |
| 8 | October 26 | We met with Erwin talk about the use of a 3-d environment for the cells to live. We discussed our final channel dimmensions and an Adobe template was formed. The template was then sent to Erwin to be printed on a high resolution printer. |
| 9 | November 2 | We recieved the high-resolution printout, and next week we will create the mask, which is used as a template to pour PDMS. Also modeling software (Matlab, Comsol) are being used to model our system. |
| 10 | November 9 | Our team meet and we built our mold which can be used to create each set of channels. |
| 11 | November 16 | A MATlab model was finished to simulate the gradient formation in the channel. PDMS was poured over the channel and mounted to a glass slide. Experimentation began with Texas Red flourescense to determine diffusion coefficients. |
| 12 | November 23 | Thanksgiving |
| 13 | November 30 | Concentration gradients were formed through the use of Texas Red. This was verified through the use of MATLAB image processing, which verified the intensity of the light was proportional to the concentration. A COMSOL model was developed to further demonstrate the gradient formation. |
| 14 | December 7 | Poster and paper were completed. |
| 15 | December 14 | End of Semester. |
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Midsemester Presentation (Oct 18 2007, 5259 kb) |
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Final Poster (Dec 6 2007, 598 kb) |
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Final PDS (Dec 11 2007, 73 kb) |
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Final Paper (Dec 12 2007, 1156 kb) |
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Poster Presentation Powerpoint (Dec 12 2007, 885 kb) |
