Patel, Sanket N
(2009)
CHARACTERIZATION OF SMOOTH MUSCLE CELL PHENOTYPE AND FUNCTIONALITY FOR POTENTIAL TISSUE ENGINEERING APPLICATIONS.
Master's Thesis, University of Pittsburgh.
(Unpublished)
Abstract
Smooth muscle cell (SMC) embedded scaffolds have possible applications in treating diseased tissues that are rich in SMCs. Stress urinary incontinence (SUI) is an example of a disease that can be caused due to SMC dysfunction within the urinary sphincter. The goal of this thesis was to create a SMC-populated tissue engineered urethral wrap (TEUW) using autologous urethral SMCs (uSMCs), to be used as a cuff around the native urethra to integrate with the host tissue for providing mechanical and functional reinforcement to the diseased urethra. uSMCs were isolated from rat urethras. SMC phenotype was verified by immunofluorescence and western blotting. Isolation purity was assessed by staining uSMCs for skeletal muscle and urothelium markers since they are also present in the urethra. TEUWs were examined for SMC phenotype, apoptosis, mechanical and histological endpoints after culture.This thesis also evaluated the functionality of differentiated SMCs (dSMCs), which were derived via mechanical stimulation of bone marrow-derived mesenchymal stem cell (BMMSCs). The long-term objective is to use BMMSCs as an autologous source for SMCs in order to create TEUW-like tubular constructs for treating SMC related dysfunctions including, but not limited to SUI. uSMCs and dSMCs were assessed and compared for intracellular Ca2+ activity (fura-2) and contractile responses (live-cell) to various stimuli.Results of isolated uSMCs revealed expression of SMC markers and absence of skeletal and urothelium markers, suggesting isolation purity. uSMC-based TEUWs showed non-linear pressure-diameter profiles like soft tissues, greater compliance than the native urethra, and burst pressures similar to stem-cell based TEUWs. Both, uSMCs and dSMCs, exhibited intracellular Ca2+ activity, with and without extracellular Ca2+, vital for full SMC function. However, their failure to show morphological changes in the presence of agonists during contractility assessment indicated absence of mature SMCs. In summary, this study demonstrates proficient uSMC isolation, which represents an important step towards TEUW development, and that uSMCs and dSMCs are not fully functional at the differentiation stage tested. Future work should focus on increasing contractile protein expression by using matrix-like culture systems and/or biochemical stimulants. Following a systematic examination, SMC-populated TEUWs could be tested in an animal model.
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Details
| Item Type: |
University of Pittsburgh ETD
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| Status: |
Unpublished |
| Creators/Authors: |
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| ETD Committee: |
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| Date: |
25 September 2009 |
| Date Type: |
Completion |
| Defense Date: |
24 July 2009 |
| Approval Date: |
25 September 2009 |
| Submission Date: |
27 July 2009 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
Swanson School of Engineering > Bioengineering |
| Degree: |
MSBeng - Master of Science in Bioengineering |
| Thesis Type: |
Master's Thesis |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
smooth muscle; tissue engineering; urethra |
| Other ID: |
http://etd.library.pitt.edu/ETD/available/etd-07272009-083807/, etd-07272009-083807 |
| Date Deposited: |
10 Nov 2011 19:54 |
| Last Modified: |
15 Nov 2016 13:47 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/8658 |
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