The direct effect of SARS-CoV-2 virus vaccination on human ovarian granulosa cells explains menstrual irregularities

Abstract

Following administration of the SARS-CoV-2 vaccine, many women worldwide reported short-term menstrual irregularities. Although menstrual bleeding, "the fifth vital sign", is experienced by more than 300 million people on any given day worldwide, these changes were only partially studied. Irregular periods are important well beyond fertility and the discomfort they impose; they are associated with the risk of cardiovascular morbidity, chronic diseases, and premature mortality. Pre-clinical examination of the vaccine polymeric envelope indicates its accumulation in the ovaries. The somatic endocrine cells of the ovarian follicle - the granulosa cells (GCs)-participate in the strict hypothalamic-pituitary-ovarian (HPO) feedback loop that governs the menstrual cycle via endocrine and paracrine regulators, as AMH and Inhibins. We aimed to unravel the direct effect of the COVID-19 vaccine on GCs and link their post-vaccine activity to changes in menstrual patterns. Human primary GCs exposed in-vitro to the Pfizer COVID-19 vaccine BNT162b2, demonstrated no change in their viability but altered mRNA transcripts, specifically of the regulatory key factors: InhibinB was upregulated, whereas AMH was downregulated. We further examined pre- and post-vaccination blood samples from individual women and found a 2-3 folds change in the post-vaccination FSH/InhibinB protein level ratio, compared to their pre-vaccination values. This altered expression of InhibinB could significantly impact the HPO axis in vaccinated women and may ultimately influence the endometrium cyclicity, manifested clinically by the commonly reported changes in menstrual bleeding patterns.

Fig. 1. Human primary GCs (hpGCs) vitality after exposure to BNT162b2 COVID-19 vaccine.

hpGCs (cells were cultured in pools) were exposed to 2 vaccine concentrations: “injected dose” and “end-organ dose”. Non-treated cells served as control. hpGCs were harvested for vitality analysis using MTT assay at 24 or 48 h. Data are presented as Mean ± SEM of relative expression. *P < 0.05—significantly different from control value. Data were analyzed by Kruskal–Wallis followed by FDR correction for multiple comparisons. Each experiment was conducted three times in duplicates.

Fig. 2. mRNA Expression of selected genes in hpGCs Following 24-hour Exposure to BNT162b2 COVID-19 Vaccine.

a–f hpGCs (cells were cultured in pools) were exposed to 2 vaccine concentrations: “injected dose” and “end-organ dose” for 24 h and harvested for mRNA analysis via qPCR. Non-treated cells served as control. Data are presented as Mean ± SEM of relative expression. *PV < 0.05—significantly different from control value. Data were analyzed by Kruskal–Wallis followed by FDR correction for multiple comparisons. Experiments were conducted at least five times.

Fig. 3. mRNA Expression of selected genes in hpGCs Following 48-hour Exposure to BNT162b2 COVID-19 Vaccine.

a–f hpGCs (cells were cultured in pools) were exposed to 2 vaccine concentrations: “injected dose” and “end-organ dose”. Non-treated cells served as control. Forty-eight hours later, hpGCs were harvested for mRNA analysis. The corresponding mRNAs were subjected to qPCR analysis with specific primers for aromatase, FSHR, IL-8, AMH, InhibinA, and InhibinB and calibrated by HPRT1. Data are presented as Mean ± SEM of relative expression. *PV < 0.05—significantly different from control value. Data were analyzed by Kruskal–Wallis followed by FDR correction for multiple comparisons. Experiments were conducted at least five times.

Fig. 4. Menstrual changes post 3rd dose of the COVID-19 vaccination questionnaire summary.

Two hundred forty-eight women, attending Sourasky Medical Center were presented with an electronic questionnaire ~4 months after vaccination, following changes in their menstrual cycle after COVID-19 vaccine administration. Two hundred thirty-six completed the survey, 124 of them were included. OC oral contraceptives, IUD intrauterine device.

Fig. 5. FSH/InhibinB ratio changes in response to administration of the COVID-19 vaccine.

FSH/InhibinB ratio in 5 women vaccinated with the 3rd dose of the COVID-19 vaccine and who reported subsequent changes in menstrual bleeding patterns.