Piceatannol SNEDDS Attenuates Estradiol-Induced Endometrial Hyperplasia in Rats by Modulation of NF-κB and Nrf2/HO-1 Axes

Abstract

Endometrial hyperplasia (EH) is the most common risk factor for endometrial malignancy in females. The pathogenesis of EH has been directly linked to uterine inflammation, which can result in abnormal cell division and decreased apoptosis. Piceatannol (PIC), a natural polyphenolic stilbene, is known to exert anti-inflammatory, antioxidant and anti-proliferative activities. The aim of the present study was to examine the potential preventive role of PIC in estradiol benzoate (EB)-induced EH in rats. A self-nanoemulsifying drug delivery system (SNEDDS) was prepared to improve the solubility of the PIC. Therefore, thirty female Wistar rats were divided into five groups: (1) control, (2) PIC SNEDDS (10 mg/kg), (3) EB (0.6 mg/kg), (4) EB + PIC SNEDDS (5 mg/kg) and (5) EB + PIC SNEDDS (10 mg/kg). The administration of PIC SNEDDS prevented EB-induced increases in uterine weights and histopathological changes. Additionally, it displayed pro-apoptotic and antioxidant activity in the endometrium. Immunohistochemical staining of uterine sections co-treated with PIC SNEDDS showed significantly decreased expression of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and nuclear transcription factor-kappa B (NF-κB). This anti-inflammatory effect was further confirmed by a significant increase in Nrf2 and heme oxygenase-1 (HO-1) expression. These results indicate that SNEDDS nanoformulation of PIC possesses protective effects against experimentally induced EH.

Keywords: endometrial hyperplasia; estradiol benzoate; inflammation; oxidative stress; piceatannol.

Figure 1

Effect of piceatannol SNEDDS on uterine morphology in rats treated with estradiol benzoate. Gross images of rat uteri collected after four weeks of treatment with estradiol benzoate (0.6 mg/kg) in the absence or presence of piceatannol SNEDDS (5 or 10 mg/kg). EB is estradiol benzoate, PIC is piceatannol, SNEDDS is self-nanoemulsifying drug delivery system.

Figure 2

Effect of piceatannol SNEDDS on uterine histopathology in EB-induced EH in rats. Shown are uterus sections stained with Hematoxylin and eosin (H&E) from the following groups: (A) control group showing the normal histoarchitecture of the uterus; (B) piceatannol SNEDDS (PIC) 10 mg/kg group with no observable histological changes; (C) estradiol benzoate (EB) 0.6 mg/kg group showing increased endometrium thickness with hyperplasia and intraluminal papillary projections; (D) EB group co-treated with PIC (5 mg/kg) showing reduction in hyperplasia and projections; (E) EB group co-treated with PIC (10 mg/kg) showing a clear reduction in hyperplasia and projections. Arrows point to intraluminal papillary projections. Panel (F) represents a graphical presenation of endometrial thickness. Dara are expressed as mean ± S.D (n = 6). ns = not significant, ** p < 0.01, *** p < 0.001 and **** p < 0.0001 by one-way ANOVA with Tukey’s post hoc test.

Figure 3

The effect of PIC SNEDDS treatment on the mRNA expression of Bax (A) and Bcl-2 (B) and the Bax/Bcl-2 ratio (C) in the uterine tissue. Data are shown as Mean ± S.D (n = 6). ns = not significant, * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001 by one-way ANOVA with Tukey’s post hoc test.

Figure 4

Effect of PIC SNEDDS treatment on caspase-3 concentration in the uterine tissue. Data are shown as Mean ± S.D (n = 6). ns = not significant and **** p < 0.0001 by one-way ANOVA with Tukey’s post hoc test.

Figure 5

Effect of PIC SNEDDS on EB-induced alterations of IL-6, TNF-α, NFκB, HO-1 and Nrf2 expression in the uterine tissue of rats by immunohistochemical staining. Data presented in bar graphs are the mean of H-scores ± S.D (n = 6). ns = not significant, * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001.