Investigating the effects of rosmarinic acid on ovarian tissue, inflammatory markers, and sex hormones in polycystic ovary syndrome rats

Abstract

Polycystic ovary syndrome (PCOS) causes the impairment of female fertility and elevates the risk of metabolic disorders. The current study aimed to evaluate the effects of rosmarinic acid (Ros) on the ovarian histo-stereology, the level of reproductive hormones, and inflammation in a rat model of PCOS. Fifteen adult Sprague Dawley rats were randomly divided into three groups, including controls, PCOS, and PCOS+Ros (receiving 25 mg/kg of Ros for 39 days). After treatments, the ovarian histo-stereology, the level of reproductive hormones, and the level of inflammatory markers were analyzed. PCOS led to increased ovarian weight and volume, cortical and medullary expansion, reduced ovarian follicles, and enhanced follicular atresia. It also caused hormonal imbalances, elevating LH, FSH, and testosterone while decreasing estradiol, progesterone, and AMH. Additionally, PCOS increased pro-inflammatory markers (TNF-α and IL-6) and decreased anti-inflammatory markers (IL-4 and IL-10). However, Ros administration in PCOS animals improved ovarian structure, increased follicle numbers, reduced atresia, balanced reproductive hormones, and restored inflammatory markers (p value <0.05). The present findings may suggest Ros as a novel strategy for the management of PCOS, although further studies are necessary.

Keywords: histology; hormone; infertility; integrative medicine; phytotherapy; polycystic ovary syndrome; rosmarinic acid.

FIGURE 1

The weight and volume of ovarian tissues. The findings revealed that in PCOS animals the weight (a) and volume (b) of ovarian tissues significantly increased compared to controls. Nevertheless, rosmarinic acid represented ameliorative properties on the PCOS‐induced elevation in the weight and volume of ovarian tissue. There were five animals in each group as biological replicates. p value <0.05 was considered significant. One‐way ANOVA and Tukey's post hoc tests were performed. Con, healthy controls; PCOS, animals with polycystic ovary syndrome; PCOS+Ros, animals with polycystic ovary syndrome treated with rosmarinic acid.

FIGURE 2

Rosmarinic acid improved the volume of ovarian structures that pathologically changed in PCOS animals. The administration of rosmarinic acid to PCOS animals reduced the volume of cortex (a), medulla (b), and cystic follicles (d) while increasing corpus luteum volume (c). There were five animals in each group as biological replicates. p value <0.05 was considered significant. One‐way ANOVA and Tukey's post hoc tests were performed. Con, healthy controls; PCOS, animals with polycystic ovary syndrome; PCOS+Ros, animals with polycystic ovary syndrome treated with rosmarinic acid.

FIGURE 3

The stereological analysis of ovarian tissue. The number of primordial (a), unilaminar (b), multilaminar (c), antral (d), and graafian (e) follicles in PCOS animals significantly decreased, while PCOS increased the number of atretic (f) follicles. There were five animals in each group as biological replicates. p value <0.05 was considered significant. One‐way ANOVA and Tukey's post hoc tests were performed. Con, healthy controls; PCOS, animals with polycystic ovary syndrome; PCOS+Ros, animals with polycystic ovary syndrome treated with rosmarinic acid.

FIGURE 4

The histomorphology of ovarian tissue. Control group (a), PCOS group (b), PCOS+Ros group (c). a–c × 40 magnification is used in H&E staining. S.F, antral follicles; Cy, cystic follicles; A.F, atretic follicles; C.L, corpus luteum; Con, healthy controls; PCOS, animals with polycystic ovary syndrome; PCOS+Ros, animals with polycystic ovary syndrome treated with rosmarinic acid.

FIGURE 5

The restoration of PCOS‐induced alteration in the levels of reproductive hormones by rosmarinic acid. PCOS increased the levels of LH (a), FSH (b), estradiol (c), testosterone (e), and LH:FSH ratio (g), whereas reduced the levels of progesterone (d) and AMH (f). However, rosmarinic acid significantly restored the altered levels of reproductive hormones. There were five animals in each group as biological replicates, and two technical replicates were performed. A p value <0.05 was considered significant. One‐way ANOVA and Tukey's post hoc tests were performed. Con, healthy controls; PCOS, animals with polycystic ovary syndrome; PCOS+Ros, animals with polycystic ovary syndrome treated with rosmarinic acid.

FIGURE 6

Gene expression analysis of inflammatory markers. PCOS increased the expression of genes encoding TNF‐α (a) and IL‐6 (b), while the expression of IL‐4 (c) and IL‐10 (d) genes was significantly reduced in PCOS animals. The administration of rosmarinic acid restored the expression of inflammation‐related genes. There were five animals in each group as biological replicates, and two technical replicates were performed. A p value <0.05 was considered significant. One‐way ANOVA and Tukey's post hoc tests were performed. Con, healthy controls; PCOS, animals with polycystic ovary syndrome; PCOS+Ros, animals with polycystic ovary syndrome treated with rosmarinic acid.

FIGURE 7

Rosmarinic acid ameliorated PCOS caused changes in the levels of inflammatory markers. The levels of TNF‐α (a), IL‐6 (b), IL‐4 (c), and IL‐10 (d) were analyzed using the ELISA approach. There were five animals in each group as biological replicates, and two technical replicates were performed. A p value <0.05 was considered significant. One‐way ANOVA and Tukey's post hoc tests were performed. Con, healthy controls; PCOS, animals with polycystic ovary syndrome; PCOS+Ros, animals with polycystic ovary syndrome treated with rosmarinic acid.