Type 1 Diabetes and the Male Reproductive Axis

Glucose regulation is important in both spermatogenesis as well as mature sperm function. The impact of diabetes on sperm function is becoming increasingly clear, though much work is still needed to elucidate the mechanism by which diabetes impairs spermatogenesis. Diabetes causes disruptions in the hypothalamic pituitary gonadal (HPG) axis, the regulatory system that controls reproduction. The HPG axis functions by secreting GnRH pulses from the hypothalamus, which bind to receptors on the pituitary and cause secretion of follicle stimulating hormone and luteinizing hormone, which then bind to receptors in the testes and facilitate spermatogenesis. Type 1 diabetes can severely impact function of the HPG axis, leading to fertility defects. Using the Akita mouse model, a genetic model of type 1 diabetes, we show that in the absence of sufficient insulin, spermatogenesis is arrested and the mice are infertile. These mice have lower levels of luteinizing hormone, follicle stimulating hormone, and testosterone. Upon treatment with either insulin or leptin, spermatogenesis is restored, despite persistent levels of high blood glucose. We conclude that the lack of insulin and leptin during the pathogenesis of type 1 diabetes causes an arrest in spermatogenesis by abrogating function of the HPG axis. It is possible that insulin and leptin have redundant roles regulating reproductive function within the hypothalamus, and a mechanism for the precise molecular targets of both insulin and leptin that restore HPG axis function remains to be determined. We additionally found that insulin secretion occurs within the testes in addition to that secreted by the pancreas. Insulin protein was localized to the Sertoli, or nurse , cell of the testes, which is the cell responsible for providing hormones and nutrients for sperm cells as they develop from spermatogonia to spermatozoa. Additionally, we determined the presence of glucose transporter 4 (GLUT4) on the mature sperm cell. As this is an insulin responsive transporter, this provides the potential for insulin regulation of sperm cells. We show the critical nature of both GLUT4 and the basal glucose transporter, GLUT1, in glucose uptake and fertilization capacity of sperm cells. These findings demonstrate the importance of hexose utilization to sperm cell competence, and further identification and characterization of glucose transporters in sperm cells will clarify the role of hexose metabolism during sperm cell maturation and fertilization reactions

Establishment and characterization of bovine oviductal epithelial cells in culture and study of sperm binding capacity: "Feasibility studies towards the development of a semen fertility assessment assay for Norwegian red"

Mastergradsoppgave i næringsrettet bioteknologi, Avdeling for lærerutdanning og naturvitenskap, Høgskolen i Hedmark, 2011. Master of applied and commercial biotechnologyThe mammalian oviduct is the physiological site for key events in reproduction, such as capacitation of spermatozoa, fertilization and early embryonic developments. During passage through the oviduct, a fertilizing spermatozoon has to bind to and interact with epithelial cells at the caudal isthmus during the formation of functional sperm reservoir. Binding to these cells is thought to increase the fertile life span of sperm cells. In this study, bovine oviduct epithelial cells (BOECs) from NRF at estrus were cultured in monolayers and used to study sperm cells oviduct binding in vitro. The cultured cells were characterized by immunostaining and Real Time PCR was used to study the expression pattern of OVGP1 in cultured cells in the presence and absence of human chorionic gonadotrophin. Chlortetracycline staining was employed to study the capacitation status of sperm cells bound to monolayers of epithelial cells and Ca 2+ ionophore was used to induce sperm cell capacitation. Main findings demonstrate that (1) Primary BOECs cultures are a mixed population of cells; (2) Cultured BOECs loss the expression of OVGP1 over time during in vitro culture; (3) BOECs monolayers selectively bind uncapacitated sperm cells. These findings support the possible establishment of a sperm quality assessment assay through binding of sperm cells to BOECs monolayers

DNA-BASED SELF-ASSEMBLY AND NANOROBOTICS: THEORY AND EXPERIMENTS

We study the following fundamental questions in DNA-based self-assembly and nanorobotics: How to control errors in self-assembly? How to construct complex nanoscale objects in simpler ways? How to transport nanoscale objects in programmable manner? Fault tolerance in self-assembly: Fault tolerant self-assembly is important for nanofab-rication and nanocomputing applications. It is desirable to design compact error-resilient schemes that do not result in the increase in the original size of the assemblies. We present a comprehensive theory of compact error-resilient schemes for algorithmic self-assembly in two and three dimensions, and discuss the limitations and capabilities of redundancy based compact error correction schemes. New and powerful self-assembly model: We develop a reversible self-assembly model in which the glue strength between two juxtaposed tiles is a function of the time they have been in neighboring positions. Under our time-dependent glue model, we can rigorously study and demonstrate catalysis and self-replication in the tile assembly. We can assemble thin rectangles of size k × N using O

PNA-mediated Whiplash PCR

In Whiplash PCR( WPCR), autonomous molecular computation is achieved by the recursive, self-directed polymerase extension of a mixture of DNA hairpins. A barrier confronting efficient implementation, however, is a systematic tendency for encoded molecules towards backhybridization, a simple form of self-inhibition. In order to examine this effect, the length distribution of extended strands over the course of the reaction is examined by modeling the process of recursive extension as a Markov chain. The extension efficiency per polymerase encounter of WPCRi s then discussed within the framework of a statistical thermodynamic model. The efficiency predicted by this model is consistent with the premature halting of computation reported in a recent in vitro WPCRimp lementation. The predicted scaling behavior also indicates that completion times are long enough to render WPCR-based massive parallelism infeasible. A modified architecture, PNA-mediated WPCR (PWPCR) is then proposed in which the formation of backhybridized structures is inhibited by targeted PNA2/DNA triplex formation. The efficiency of PWPCRis discussed, using a modified form of the model developed for WPCR. Application of PWPCR is predicted to result in an increase in computational efficiency sufficient to allow the implementation of autonomous molecular computation on a massive scale