In Vitro and  In Vivo Culture Systems for Development of Porcine Testis Cells and Tissue

Awang Junaidi, Awang Hazmi 1981-

In Vitro and In Vivo Culture Systems for Development of Porcine Testis Cells and Tissue

dc.contributor.advisor	Honaramooz, Ali
dc.contributor.committeeMember	Machin, Karen
dc.contributor.committeeMember	MacPhee, Daniel
dc.contributor.committeeMember	Anzar, Muhammad
dc.contributor.committeeMember	Krone, Pat
dc.creator	Awang Junaidi, Awang Hazmi 1981-
dc.creator.orcid	0000-0002-2569-0568
dc.date.accessioned	2019-04-15T22:27:17Z
dc.date.available	2019-04-15T22:27:17Z
dc.date.created	2019-04
dc.date.issued	2019-04-15
dc.date.submitted	April 2019
dc.date.updated	2019-04-15T22:27:18Z
dc.description.abstract	The studies presented in this thesis were designed to identify, examine, and manipulate potential factors associated with the development of porcine testis cells and tissue using in vitro and in vivo culture systems. The objective of the first in vitro study was to examine several conditions for short-term culture of testis cells to optimize the maintenance and propagation of neonatal porcine gonocytes. We found that culturing testis cells at 3.0 × 10⁴ cells/cm² containing ~40% gonocytes in DMEM+10% FBS, at 37 °C, and without changing the medium for 7 days, improves in vitro maintenance, proliferation, and formation of gonocyte colonies. The goal of the second in vitro study was to apply these optimized conditions for prolonged culture to study the behavior, colony formation, and ultrastructure of gonocytes. We observed that gonocytes extensively migrate after developing various cytoplasmic projections, rapidly propagate, and form pluripotent embryoid body-like colonies (EBLC). The first in vivo study was designed to establish an efficient method for implantation of neonatal porcine testis cell aggregates under the back skin of recipient mice, and to explore the use of ultrasound biomicroscopy (UBM) for the non-invasive in vivo evaluation of implants. We showed that the subcutaneous injection approach, as compared with the conventional surgical approach, offers a less invasive route for implantation of cell aggregates, and results in a more consistent de novo formation of testis tissue in the implants. UBM proved to be a unique means for the non-invasive monitoring of implants and the prediction of the outcomes. The second in vivo study was aimed at utilizing this improved cell implantation model to examine the effects of pre-implantation exposure of testis cells to various growth factors (i.e., EGF, GDNF, FGF2, FGF9, VEGF, LIF, SCF, and RA) on de novo morphogenesis of porcine testis tissue. We observed that each growth factor had a distinct effect on development of the testis tissue, ranging from enhanced development to improved gonocyte survival, and promotion of certain types of seminiferous cords. In summary, we discovered several critical elements for the in vitro and in vivo culture systems for the development of testis cells and tissue.
dc.format.mimetype	application/pdf
dc.identifier.uri	http://hdl.handle.net/10388/11965
dc.subject	porcine testis
dc.subject	gonocytes
dc.subject	testis cells
dc.subject	testis tissue development
dc.subject	cell culture
dc.subject	de novo morphogenesis
dc.subject	ectopic implantation model
dc.subject	live-cell imaging
dc.subject	ultrasound biomicroscopy
dc.subject	growth factors
dc.title	In Vitro and In Vivo Culture Systems for Development of Porcine Testis Cells and Tissue
dc.type	Thesis
dc.type.material	text
thesis.degree.department	Veterinary Biomedical Sciences
thesis.degree.discipline	Veterinary Biomedical Sciences
thesis.degree.grantor	University of Saskatchewan
thesis.degree.level	Doctoral
thesis.degree.name	Doctor of Philosophy (Ph.D.)