Thyroid hormones act as mitogenic and pro survival factors in rat ovarian follicles

Abstract

Purpose

Thyroid disorders are clinically associated with impaired fertility in women, and these abnormalities can be improved by restoring the euthyroid state. The exact mechanisms of thyroid effect on female fertility are not well known; however, it is conceivable that thyroid hormones (THs) might act on ovarian physiology via receptors in granulosa cells. This work is aimed at evaluating the effects of THs on non-tumoral granulosa cells and follicles.

Methods

Freshly isolated rat ovarian follicles and granulosa cells were exposed to T3 or T4 (THs). Cell growth and viability were evaluated by cell counting and the MTT assay, respectively, follicle growth was evaluated by volume measurements. Apoptosis was evaluated by the TUNEL assay and active Caspase 3 staining. rGROV cells were exposed to T3, and apoptosis was induced by serum deprivation. Bcl2, Bcl-2-associated X protein (BAX), Akt and pAkt expression were evaluated by western blot.

Results

T3 induced a 40% increase in follicle volume (after 7 days). This increase was presumably due to the observed decrease (33%) in the apoptotic rate of the granulosa cell population. Both T3 and T4 caused a dose-dependent increase in rat granulosa cell number and viability. In addition, THs decreased the cell apoptotic rate in a dose-dependent manner. In both conditions, T3 appeared to be more efficient. In rGROV cells, 100 nM T3 induced cell growth and, in the absence of growth factors, reduced cell apoptosis by 40%, downregulating Caspase 3 and BAX. This effect was associated with an increase in pAkt levels. The involvement of the PI3 K pathway was confirmed by the ability of the PI3 K specific inhibitor (LY-294,002) to abolish T3 pro-survival action.

Conclusions

THs influence cell survival of ovarian granulosa cells. This effect likely contributes to the TH-induced follicle volume increase.

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Abbreviations

THs:

Thyroid hormones

THRs:

Thyroid Hormone Receptors

DIO:

Deiodinases

TI:

Theca/interstitial

GC:

Granulosa cells

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Funding

This study was supported by IBSA Institute Biochimique SA (Lugano, CH) (to M.C.) and by grants from Sapienza University of Rome, Ateneo Federato 2014–2015 (to R. C.).

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Correspondence to C. Verga Falzacappa.

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Animals were maintained in accordance with the Italian Department of Health Guide for Care and Use of Laboratory Animals. Experimental protocols were approved by the ‘La Sapienza’ University Committee for Animal Care and Use. All of the authors certify that: (1) this material has not been published in whole or in part elsewhere; (2) the manuscript is not currently being considered for publication in another journal; (3) they were personally and actively involved in substantive work leading to the manuscript and will hold themselves jointly and individually responsible for its content; (4) this article does not contain any studies with human participants performed by any of the authors.

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Canipari, R., Mangialardo, C., Di Paolo, V. et al. Thyroid hormones act as mitogenic and pro survival factors in rat ovarian follicles. J Endocrinol Invest 42, 271–282 (2019). https://doi.org/10.1007/s40618-018-0912-2

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Keywords

  • Thyroid hormones
  • Apoptosis
  • Ovary
  • Granulosa cells
  • Follicles