Bushen Huoxue Recipe inhibits endometrial epithelial-mesenchymal transition through the transforming growth factor-β/nuclear factor kappa-B pathway to improve polycystic ovary syndrome-mediated infertility

Figures & data

Figure 1. Network pharmacology analysis results in presentation. a: Disease target and drug-disease shared target acquisition. b: Bushen Huoxue Recipe (BSHX) " Chemical Composition-Common Targets-Disease Information Network Diagram. c: Protein-protein interaction (PPI) network diagram of key and core targets.

Table 1. Information of active ingredients and predicted targets in the Bushen Huoxue Recipe.

Chinese name	Scientific name	Active ingredient(s) (pcs)	Predicted targets (pcs)
Guizhi	Cinnamomum cassia (L.) J. Presl	7	58
Fuling	Poria cocos (Schw.) Wolf	15	23
Mudanpi	Paeonia suffruticosa Andr.	11	209
Baishao	Paeonia lactiflora Pall.	13	102
Taoren	Prunus persica(L.) Batsch	23	101
Tusizi	Cuscuta australis R.Br.	11	291
Sangjisheng	Taxillus chinensis(DC.)Danser	2	140
Xuduan	Dipsacus asper Wall. ex Henry	8	47

Table 2. Information on the ranking of active ingredients in the treatment of polycystic ovary syndrome with the Bushen Huoxue Recipe.

No.	Molecular ID	Chemical name	Degrees	Oral bioavailability(OB%)	Drug similarity(DL)	Source Chinese Medicine
1	MOL000098	quercetin	111	46.43	0.28	Cuscuta australis R.Br.、Taxillus chinensis (DC.) Danser、Paeonia suffruticosa Andr.
2	MOL000422	kaempferol	40	41.88	0.24	Cuscuta australis R.Br、Paeonia Suffruticosa Andr.、Paeonia lactiflora Pall.
3	MOL000354	isorhamnetin	25	49.60	0.31	Cuscuta australis R.Br.
4	MOL000358	beta-sitosterol	21	36.91	0.75	Cuscuta australis R.Br.、Prunus persica (L.) Batsch、Dipsacus asper Wall. ex Henry、Cinnamomum Cassia (L.) J. Presl、Paeonia lactiflora Pall.
5	MOL001558	sesamin	15	56.55	0.83	Cuscuta australis R.Br.
6	MOL000296	hederagenin	12	36.91	0.75	Prunus persica (L.) Batsch、Poria cocos (Schw.) Wolf
7	MOL003152	gentisin	11	64.06	0.21	Dipsacus asper Wall. ex Henry
8	MOL004576	taxifolin	9	57.84	0.27	Cinnamomum cassia (L.) J. Presl
9	MOL005944	matrine	9	63.77	0.25	Cuscuta australis R.Br.
10	MOL000492	(+)-catechin	8	54.83	0.24	Cinnamomum cassia (L.) J. Presl、MOUTAN CORTEX、Paeonia lactiflora Pall.
11	MOL000493	campesterol	5	37.58	0.71	Prunus persica (L.) Batsch
12	MOL000359	sitosterol	3	36.91	0.75	Dipsacus asper Wall. ex Henry、Taxillus chinensis(DC.) Danser、Cinnamomum cassia(L.) J. Presl、Paeonia Suffruticosa Andr.、Paeonia lactiflora Pall.
13	MOL000211	Mairin	1	55.38	0.78	Paeonia Suffruticosa Andr.、Paeonia lactiflora Pall.

Figure 2. GO and KEGG pathway analysis.

Figure 3. Molecular docking animation and binding energy demonstration. a: Docking animation of TNF-α (-8.7kcal/mol), NF-κB (-8.6kcal/mol), TAK1 (-8.6kcal/mol), TGFβ (-6.6kcal/mol) and quercetin molecules. b: Docking binding ability of quercetin, kaempferol, isorhamnetin, and beta-sitosterol with TNF-α, TGFβ, TAK1, and NF-κB molecules.

Figure 4. Basic conditions of rats in each group. a: Body mass growth index of rats and ovarian index of rats. Data are shown as mean ± SD (n = 8), and data between multiple groups were compared by one-way analysis of variance (ANOVA). b: Rat ovary sections (HE staining). c: Vaginal exfoliated cell map of rats in the control group. d: Estrus cycle curves diagram for each group of rats.

Figure 5. Uterine tissue morphology and pregnancy in rats of four groups. a: Rat endometrial tissue (HE staining). b: Pregnancy loci of rats.

Table 3. Diameter of the rat uterus, thickness of the endometrium, and number of endometrial glands in different groups.

Group	N	Diameter of uterus (mm)	Endometrial thickness(mm)	Number of uterine glands
Control	4	20.63 ± 7.97	7.00 ± 2.94	26.75 ± 9.22
Model	4	10.00 ± 2.97	3.12 ± 0.75	10.50 ± 5.56
BSHX	4	11.95 ± 1.72	3.63 ± 0.75	12.25 ± 5.32
Metformin	4	11.75 ± 2.50	3.50 ± 0.82	11.00 ± 3.37
F		4.475	4.98	6.236
P		0.025	0.018	0.009

BSHX: Bushen Huoxue Recipe.

Data are shown as mean ± SD, and data between multiple groups were compared by one-way ANOVA.

Table 4. Pregnancy rate and the number of embryos in different groups.

Group	N	Number of embryos	Pregnancy rate
Control	8	13.75 ± 1.04	1.00
Model	2	1.50 ± 0.71	0.25
BSHX	6	12.50 ± 2.43	0.75
Metformin	6	3.67 ± 1.03	0.75
F		74.328
P		0.000

BSHX: Bushen Huoxue Recipe.

Each group of eight female rats were mated with males in a 2:1 ratio.

Data are shown as mean ± SD, and data between multiple groups were compared by one-way ANOVA.

Figure 6. Serum TNF-α levels, and levels of target proteins and EMT-related proteins in rat endometrial tissue. a: Endometrial TGF-β, TAK, NF-κB, E-cadherin, and Vimentin WB results. b: Endometrial TGF-β, TAK, NF-κB, Vimentin Heatmap Display. c: Serum TNF-α levels in rats (*p < .05, **p < .01, ***p < .001). Levels of TGFβ, TAK, and NF-κB in rat endometrial tissue and levels of EMT-related proteins Vimentin and E-cadherin in the endometrium. Data are shown as mean ± SD (n = 5), and data between multiple groups were compared by one-way ANOVA. Compared with the control group: *p < .05, **p < .01, ***p < .001; compared with the model group: #p < .05, ##p < .01, ###p < .001; compared with the metformin group: ▲p < .05, ▲▲p < .01, ▲▲▲p < .001.

Data availability statement

We are open to providing our research data. The data that support the findings of this study are openly available in 10.6084/m9.figshare.24156384.