Date of Award

Winter 1996

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Program/Concentration

Cellular Endocrinology and Reproductive Biology Track

Committee Director

William G. Kearns

Committee Director

Anibal A. Acosta

Committee Member

Stanton F. Hoegerman

Committee Member

Joleen L. Zackowski

Committee Member

Christopher J. Osgood

Committee Member

Gerald J. Pepe

Abstract

Artificial fertilization protocols have been developed which bypass natural barriers for reproduction. One aspect of infertility which has received little attention is the potential importance of aneuploidy in sperm. To more clearly understand the cytogenetic make-up of sperm from OAT patients, multi-probe, multi-color FISH was performed to determine aneuploidy.

The introduction of intracytoplasmic sperm injection (ICSI) has revolutionized protocols used in in vitro fertilization centers. The pregnancies resulting from ICSI suggest an increased frequency of sex chromosome aneuploidy in livebirths. Preliminary data by others' suggest that sperm tail swelling patterns following hypo-osmotic swelling (HOST) can be predictive of fertilizing ability. To clarify the relationship between sperm tail swelling patterns, aneuploidy, and fertilizing ability, we developed a technique combining HOST and FISH.

Using multi-probe, multi color FISH and developed formulae, we determined total aneuploidy in sperm from proven fertile donors and OAT patients. In controls, the total aneuploidy ranged between 4.1 and 7.7%. For OAT patients, the total aneuploidy was between 43 and 69%. Diploidy was found in 0.04% of sperm in controls and ranged between 0.4 and 9.6% in OAT patients.

To determine if sperm selection by swim-up can separate haploid from aneuploid sperm, we used three-probe, three-color FISH on sperm in the swim-up and pellet fractions. Between 35 and 52% of sperm from OAT patients in the swim-up fraction and 36 to 49% from the pellet were aneuploid. No significant difference was observed. However, the percent of diploid sperm predominantly resided within the pellet for all OAT and proven fertile donors.

We performed HOST and FISH on sperm from OAT patients undergoing ICSI. Data suggests that sperm tail swelling patterns A, B, D, E, F and G may be aneuploid in OAT patients. No aneuploidy was found in sperm with a C type tail swelling pattern in either OAT or normal controls. Diploid sperm in controls and OAT patients were only found in types A and G tail swelling patterns. Preliminary data suggests that there may be a correlation between HOST and aneuploidy for OAT patients.

Of the 30 OAT patients studied, 26 series of ICSI were performed. Sixty-eight percent of oocytes fertilized (165 embryos/242 oocytes). Following fresh embryo transfer, 1 preclinical abortion, 1 first trimester loss, and 2 term deliveries (2/26, 7.7%) occurred.

Our data show significant increases in the frequencies of diploidy, autosomal disomy, autosomal nullisomy, sex chromosome number, and total cytogenetic abnormalities in sperm from OAT patients versus controls. The data suggest that meiotic errors occur at highly elevated frequencies in the germ cells of severely affected OAT patients.

Comments

Dissertation submitted to the Faculty of Eastern Virginia Medical School and Old Dominion University in Partial Fulfillment of the Requirement for the Degree of Doctor of Philosophy in Biomedical Sciences.

DOI

10.25777/v96r-dm76

ISBN

9780591262216

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