Letrozole and the Traditional Chinese Medicine, Shaofu Zhuyu Decoction, Reduce Endometriotic Disease Progression in Rats: A Potential Role for Gut Microbiota

Abstract

We previously showed that the Chinese herbal medicine, Shaofu Zhuyu decoction (SFZYD), shrank the size of endometriotic lesions in rats with endometriosis. We therefore conducted the present study to investigate the effects of letrozole and SFZYD on gut microbiota in endometriotic rats. Rats were divided into four groups: a blank group, model group, letrozole group, and SFZY group. Ectopic lesion size and COX-2 expression in the endometrium and endometriotic lesions were compared, and the community of gut microbiota was detected using 16S rRNA gene sequencing. Both letrozole and SFZYD reduced the size of ectopic lesions as well as lowered the expression of COX-2, thus reducing the inflammatory response. Compared with the blank group, the α-diversity of gut microbiota in endometriotic rats decreased, the Firmicutes/Bacteroidetes ratio increased, and the abundance of Ruminococcaceae was reduced. The α-diversity of gut microbiota in the letrozole group was similar to that in the model group, but the Firmicutes/Bacteroidetes ratio was diminished. The α-diversity in the SFZY group was similar to that in the blank group, the Firmicutes/Bacteroidetes ratio was attenuated, and the abundance of Ruminococcaceae was elevated compared with the model group. These results indicated that the therapeutic mechanisms of both letrozole and SFZYD were related to the restoration of gut microbiota.

Figure 1

Body weight gain of rats in each group (two-way ANOVA with post hoc Student's t-tests, ^∗P < 0.05).

Figure 2

Representative images of immunohistochemical staining of COX-2 in endometriotic lesions and normal endometrium. (a) Sections stained for COX-2 in endometriotic lesions (magnification, ×400). (b) Mean optical density (MOD) values of the COX-2 expression (n = 6); one-way ANOVA with Tukey's test, ^∗∗P < 0.01 compared with the model group. (c) Sections stained for COX-2 in the endometrium (magnification, ×400). (d) Mean optical density (MOD) values of the COX-2 expression (n = 6); one-way ANOVA with Tukey's test, ^ΔΔP < 0.01 compared with the blank group and ^∗∗P < 0.01 compared with the model group.

Figure 3

Gut microbiota composition is analyzed by α-diversity (n = 6). (a) A species accumulation boxplot is used to evaluate species richness and whether the sample size is sufficient. (b, c) ACE and Simpson analyses are used to evaluate community richness and diversity within each group (Kruskal–Wallis test, a nonparametric test is used; ^∗P < 0.05 and ^∗∗P < 0.01).

Figure 4

Analysis of species composition of gut microbiota among groups using OUT analysis (n = 6). (a) Heatmap representation of relative abundances of the phyla in fecal samples from each group. Microbiota distribution at the phylum level (b), class level (c), family level (d), and genus level (e).

Figure 5

Comparison of gut microbiota among groups using PCoA and LEfSe analyses. (a, b) PCoA analysis based on unweighted and weighted UniFrac algorithms among groups. (c, d) Histogram of LDA scores and cladogram for differentially abundant genera between blank and model groups. (e, f) Histogram of LDA scores and cladogram for differentially abundant genera among blank, model, and cc vcx letrozole groups. (g, h) Histogram of LDA scores and cladogram for differentially abundant genera among blank, model, and SFZY groups.