- Title
- Investigation of equine fertility mechanisms through proteomic analysis of stallion spermatozoa, early equine embryos and pregnant mare uterine fluid
- Creator
- Swegen, Aleona
- Relation
- University of Newcastle Research Higher Degree Thesis
- Resource Type
- thesis
- Date
- 2017
- Description
- Research Doctorate - Doctor of Philosophy (PhD)
- Description
- Equine breeding industries suffer from many inefficiencies arising from deficits in our understanding of reproductive physiology in this species. This includes cases of idiopathic infertility and suboptimal resistance of spermatozoa to chilled storage and cryopreservation in stallions, and high rates of early embryo loss in mares. At the same time, unchecked breeding in populations of feral horses, an introduced species in the Australian landscape, contributes to damage of sensitive ecosystems and poses an ethical conundrum in light of controversy surrounding lethal population control methods. Existing fertility control agents are lacking in their efficacy and specificity. Improving our understanding of the molecular mechanisms underlying reproductive function in horses is paramount if we are to achieve better outcomes in breeding industries and develop novel contraceptive strategies for feral horses. The advent of highly sensitive mass spectrometry technologies provides a distinct opportunity to characterize the proteomic profiles of the major phenotypic elements of equine reproduction – the spermatozoon and the early embryo. This thesis encompasses a series of studies whose aims were 1) to utilize cutting edge mass spectrometry technology to characterize the proteomes of stallion spermatozoa and of early equine embryos together with their secretome and immediate uterine environment, 2) to integrate proteomic data with bioinformatics and in vitro experimentation and thus elicit an improved understanding of equine reproductive function and 3) to deliver applied outcomes in fertility manipulation (including contraceptive targets and strategies for improving fertility). Mass spectrometry driven proteomic analysis of equine spermatozoa revealed 1030 proteins. Together with gene ontology analyses these provided new information about the metabolism, antioxidant defenses and receptors of stallion spermatozoa. Mitochondrial proteins and those involved in catabolic processes constituted dominant categories. Several enzymes specific to beta-oxidation of fatty acids were identified, and further in vitro experiments demonstrated that beta-oxidation contributes to metabolism and motility in stallion spermatozoa. Identification of a series of receptors, kinases and metabolic enzymes in the sperm proteome led to an investigation of the roles of peroxisome proliferator activating receptor gamma (PPARG) and the metabolic switch, AMP-activated protein kinase (AMPK), in regulation of stallion sperm metabolism and in mediating the remarkable effects of an anti-diabetic pharmaceutical, rosiglitazone, on sperm motility, reactive oxygen species production, mitochondrial function and ATP production. The characterization of functional AMPK in stallion spermatozoa provided a novel avenue for the manipulation of sperm metabolism and improvement of sperm storage methods. Accordingly, rosiglitazone substantially alleviated the time-dependent deterioration of stallion spermatozoa during ambient temperature storage by diverting metabolism away from oxidative phosphorylation and towards glycolysis, with novel implications for preservation of stallion spermatozoa. In the final study, we present the first successful proteomic analysis of several components of the equine pregnancy: proteins secreted by early embryos, blastocoel fluid composition, the embryonic capsule, and uterine luminal fluid proteins. Integrating these findings with existing literature yields novel insights into how these proteins fit into the complex puzzle of pregnancy establishment and maintenance. The studies presented herewith demonstrate the successful utilization of cutting edge mass spectrometric technology for comprehensive analysis of the proteins that define the male gamete in the stallion and early pregnancy in the mare, with implications for both equine fertility control and fertility enhancement.
- Subject
- antigens; contraceptive; reactive oxygen species; AMPK; ATP; rosiglitazone; thiazolidinedione; PPAR; equine reproduction; equine embryo; early pregnancy loss; histograph; embryo; thesis by publication; oocyte; reproduction; spermatozoa; stallion; fertility; mitochondria; metabolism
- Identifier
- http://hdl.handle.net/1959.13/1336113
- Identifier
- uon:27549
- Rights
- Copyright 2017 Aleona Swegen
- Language
- eng
- Full Text
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| Thumbnail | File | Description | Size | Format | |||
|---|---|---|---|---|---|---|---|
| View Details Download | ATTACHMENT01 | Thesis | 30 MB | Adobe Acrobat PDF | View Details Download | ||
| View Details Download | ATTACHMENT02 | Abstract | 73 KB | Adobe Acrobat PDF | View Details Download |