Granulocytes and Vascularization Regulate Uterine Bleeding and Tissue Remodeling in a Mouse Menstruation Model

Astrid Menning,Isabella Gashaw,Karl-Heinrich Fritzemeier,Alexander Walter,Marion Rudolph,Lars Roese,Hyunjung Lim

doi:10.1371/journal.pone.0041800

Abstract

Menstruation-associated disorders negatively interfere with the quality of life of many women. However, mechanisms underlying pathogenesis of menstrual disorders remain poorly investigated up to date. Among others, this is based on a lack of appropriate pre-clinical animal models. We here employ a mouse menstruation model induced by priming mice with gonadal hormones and application of a physical stimulus into the uterus followed by progesterone removal. As in women, these events are accompanied by menstrual-like bleeding and tissue remodeling processes, i.e. disintegration of decidualized endometrium, as well as subsequent repair. We demonstrate that the onset of bleeding coincides with strong upregulation of inflammatory mediators and massive granulocyte influx into the uterus. Uterine granulocytes play a central role in regulating local tissue remodeling since depletion of these cells results in dysregulated expression of matrix modifying enzymes. As described here for the first time, uterine blood loss can be quantified by help of tampon-like cotton pads. Using this novel technique, we reveal that blood loss is strongly reduced upon inhibition of endometrial vascularization and thus, is a key regulator of menstrual bleeding. Taken together, we here identify angiogenesis and infiltrating granulocytes as critical determinants of uterine bleeding and tissue remodeling in a mouse menstruation model. Importantly, our study provides a technical and scientific basis allowing quantification of uterine blood loss in mice and thus, assessment of therapeutic intervention, proving great potential for future use in basic research and drug discovery.

Highlights

During a women’s reproductive phase the endometrium undergoes repetitive cycles of proliferation, differentiation, breakdown and repair preparing the uterus for implantation and growth of an embryo
Injection of oil into the uterus of ovariectomized mice sensitized by estrogen (E2) and P4 treatment induces decidualization of the endometrial stroma, which is reflected by a massive expansion of the uterus and an increase in uterine wet weight reaching a maximum between 0 - 24 h after P4 withdrawal and declining thereafter (Figure 2A–C)
Development of novel therapeutics is limited by the incomplete knowledge on mechanisms underlying menstrual bleeding and pathogenesis of associated disorders

Summary

Introduction

During a women’s reproductive phase the endometrium undergoes repetitive cycles of proliferation, differentiation, breakdown and repair preparing the uterus for implantation and growth of an embryo. As a result, coordinated inflammation, tissue injury and shedding of the functionalis take place being followed by complete, scarless repair and regeneration from the basal layer [1,2,3]. These repetitive cycles of tissue remodeling are highly controlled and disturbances are supposed to result in menstrual disorders such as prolonged and/or excessive blood loss (heavy menstrual bleeding/HMB, menorrhagia) [1,2]. HMB is defined as a blood loss exceeding 80 ml per cycle, which significantly interferes with a woman’s physical, social, emotional and sometimes even material quality of life [4]. HMB is associated with great socio-economic impact raising a high need to develop appropriate medication [5]

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Results

Conclusion