Graduation Year

2016

Document Type

Dissertation

Degree

Ph.D.

Degree Name

Doctor of Philosophy (Ph.D.)

Degree Granting Department

Engineering

Major Professor

Robert D. Frisina, Ph.D.

Committee Member

Ann C. Eddins, Ph.D.

Committee Member

Nathan D. Gallant, Ph.D.

Committee Member

Anna Pyayt, Ph.D.

Committee Member

Joseph P. Walton, Ph.D.

Keywords

Estrogen, Progesterone, Auditory Brainstem Responses (ABRs), Temporal Processing, IGF-1 Pathway, FOXO3

Abstract

After the findings were reported for the Women’s Health Initiative (WHI) study in the past decade, there has been a significant decline in the overall use of hormone replacement therapy (HRT) among women. However, there are still millions of middle-aged, menopausal women in the U.S. who are currently undergoing hormone therapy. Their reasons for continuing treatment include relief of severe menopausal symptoms, aid in the management of osteoporosis and reduction in the risk of colon cancer (Ness et al., 2005). The purpose of the following investigation was to evaluate the impact of HRT on the central and peripheral auditory systems both during and after treatment. Over the course of the study, hormone treatments were administered to female aging CBA/CaJ mice to observe what effects estrogen (E) and progestin (P) have on the peripheral and central auditory systems. Female CBA/CaJ middle age mice were ovariectomized and placed into 4 HRT groups (E, P, E+P and Placebo [Pb]). Hormone treatment lasted 6 months followed by a recovery/washout period of 1 month. During this time, electrophysiology tests such as auditory brainstem responses (ABR) and ABR gap in noise (GIN) were used to measure neural activity for the auditory nerve and brainstem. Distortion product otoacoustic emission (DPOAE) testing was also implemented to assess the functional status of the outer hair cells (OHC) and their ability to amplify sound in the cochlea. After 6 months of treatment, animals treated with E exhibited the least amount of changes in ABR thresholds and ABR GIN amplitudes than any other subject groups. Interestingly, P animals exhibited an abrupt increase in ABR thresholds only 3 months after treatment; however, for ABR GIN amplitude levels a progressive reduction observed throughout the study. E+P and Pb animals showed signs of accelerated age-related hearing loss (ARHL) with significantly elevated ABR thresholds and dwindling ABR GIN amplitude levels. No significant signs of recovery were observed for any of the hormone groups. Therefore, in the present murine investigation, the effects of HRT were long lasting.

To further expand on the results obtained for the electrophysiology tests, molecular biology experiments were performed to evaluate the expression of IGF-1R and FoxO3 in the cochlea during hormone therapy, from both in vitro and in vivo perspectives. Both genes play significant roles in the PI3K/AKT pathway and were specifically chosen because of their role in anti-apoptotic responses and cell survival. It was hypothesized that E attenuates the effects of ARHL via the PI3K/AKT pathway by up-regulating IGF-1R and FoxO3 to counteract the effects of oxidative stress in the aging mammalian cochlea. qPCR experiments were performed with stria vascularis (SV) lateral wall cells extracted from the cochlea of each animal in the hormone groups post-treatment (in vivo) and in SVK-1 cells treated with HRT over various lengths of time (in vitro) to evaluate the expression levels of IGF-1R and FoxO3. In-vivo experiments showed that the E-treated animals had significantly higher IG-1R levels compared to the other subject groups after treatment was discontinued. Similarly, IGF-1R levels steadily increased for E-treated SVK-1 cells over the course of hormone therapy, compared to P and E+P cells. FoxO3 expression, on the other hand, declined for all of the hormone-treated cells groups, relative to control SVK-1 cells (in vitro), and no statistical differences were detected for FoxO3 levels among the post-treatment animals (in vivo). These findings indicate that there is cross talk between E and IGF-1R involving the PI3K/AKT pathway, which contributes to the delayed onset of ARHL observed during HRT with E. Meanwhile, FoxO3 may not play a role in neuro-protective properties in the cochlea during HRT, as initially hypothesized.

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