Huayu Sanjie Enema Liquid Relieves Pain in Endometriosis Model Rats by Inhibiting Inflammation, Peripheral Sensitization, and Pelvic Adhesion.

The objective of this study is to observe the effect of relieving pain of Huayu Sanjie enema liquid (HYSJ-EL) on endometriosis model rats and to explore its mechanism of action. Of 24 female Sprague Dawley rats, six were randomly selected as the sham operation group (normal control group). The remaining rats were used to establish rat models of endometriosis through autologous endometrial transplantation combined with estrogen injection. Successfully modeled rats were randomly divided into the model, indomethacin (Western medicine group), and HYSJ-EL (Chinese herbs group) treatment groups. The thermal pain threshold of rats was measured, and hematoxylin and eosin staining was used to observe pathological changes after sampling. Serum levels of prostaglandin E2 (PGE2), interleukin-6 (IL-6), macrophage inflammatory protein-2 (MIP-2), plasminogen activator inhibitor-1 (PAI-1), and transforming growth factor-ß (TGF-ß) were measured using an enzyme-linked immunosorbent assay (ELISA). Furthermore, the protein and mRNA expression levels of transient receptor potential vanilloid-1 (TRPV1) and tumor necrosis factor-α (TNF-α) in the endometrium and endometriotic lesions were measured using Western blotting and quantitative real-time PCR assays, respectively. Compared to the model group, the heat pain threshold of rats in the HYSJ-EL group was significantly increased (P < 0.01), and the serum levels of PGE2, IL-6, MIP-2, PAI-1, and TGF-ß were significantly decreased (P < 0.01), as well as the expression of TRPV1 and TNF-α protein and mRNA in the tissue of the ectopic lesion was significantly decreased (P < 0.05). These results indicate that the Huayu Sanjie enema liquid exerts analgesic effects on endometriosis by inhibiting inflammation, peripheral nerve sensitization, and pelvic adhesion.

Proteomic Analysis of Human Follicular Fluid Reveals the Pharmacological Mechanisms of the Chinese Patent Drug Kunling Pill for Improving Diminished Ovarian Reserve.

Objective: To explore the pharmacological mechanism of a Chinese patent drug (Kunling Pill (KLP)) on improving diminished ovarian reserve based on proteomic analysis. Methods: A total of 18 patients divided into three groups (the normal ovary reserve (NOR), diminished ovary reserve (DOR), and KLP groups) undergoing assisted reproductive technology by standard ovarian stimulation protocols were recruited to collect follicular fluid. Data-independent acquisition mass spectrometry was used to identify differentially expressed proteins by nano-LC-MS/MS. Bioinformatic analysis was conducted to predict the functions and pathways of the identified proteins. Clinical, hormonal, and biochemical parameters were also analyzed in the three groups. Results: A total of 144 differentially expressed proteins were screened out, including 56 proteins that were downregulated and 88 proteins that were upregulated in the DOR group compared with the NOR group, while 27 proteins were shared in the KLP-treated group. Among them, 10 proteins were upregulated and 17 proteins were downregulated in the KLP-treated group compared with the DOR group. The most enriched biological processes accounted for 28 GO terms, including cellular process, biological regulation, metabolic process, and regulation of biological process. Significant pathways were associated with fatty acid elongation, fatty acid degradation, fatty acid metabolism, nicotinate and nicotinamide metabolism, and valine, leucine, and isoleucine degradation. Conclusion: Our study provides the proteome profiles of human follicular fluid from DOR patients treated by KLP. Functional analyses of proteome datasets revealed that core proteins (SAA1, MIF, and PRDX5) and related pathways (fatty acid metabolism, nicotinate and nicotinamide metabolism, and tyrosine and purine metabolism) are possible pharmacological mechanisms through which KLP improves DOR. Therefore, these findings may help better understand the complex mechanisms through which DOR is treated by the Chinese patent drug KLP.

Using network pharmacology and molecular docking to explore the underlying anti-inflammatory mechanism of Wuyao-Danshen to treat endometriosis.

Background: This study sought to explore the anti-inflammatory mechanism of Wuyao (radix linderae)-Danshen (salviae miltiorrhiza) in endometriosis (EMS) based on network pharmacology and molecular docking. Methods: The active constituents of Wuyao-Danshen were collected and identified using the Traditional Chinese Medicine Systems Pharmacology Database, and used to predict and identify the protein targets. The EMS targets and anti-inflammatory targets were obtained from Genecards, Online Mendelian Inheritance in Man, and Drugbank. The Search Tool for the Retrieval of Interacting Genes/Proteins database was used to analyze the protein interactions (PPIs) and core targets, and a target PPI network was constructed by importing the software of Cytoscape. The Metascape database was used to conduct enrichment analyses of the Gene Ontology (GO) functions and the Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathways for the key anti-inflammatory targets of EMS. Finally, Autodock Vina software was used to verify the results of the active ingredients and key anti-inflammatory targets. Results: There were 8 active components in Wuyao, 65 in Danshen, and 591 corresponding protein targets in Danshen, and 375 in Wuyao, including luteolin, quercetin, vancomyl alcohol, and salvianol. One thousand and six hundred eighty-nine disease targets, 1,216 anti-inflammatory targets, and 144 key anti-inflammatory targets were identified, including the (signal transduction and transcriptional activator 3) STAT3, phosphatidyl inositol-3 kinase regulates subunit 1 (PIK3R1) and mitogen-activated protein kinase 1 (MAPK1) protein kinase B. Three hundred and fifty-three biological processes (BPs), 21 cellular components, and 25 molecular functions (MFs) were enriched with GO functions, and 108 KEGG pathways were enriched and analyzed, including the MAPK and PI3K-Akt signaling pathways. Molecular docking confirmed that luteolin, coumarin, and quercetin could bind to the key target proteins (i.e., STAT3, PIK3R1, and MAPK1). Conclusions: Based on network pharmacology and molecular docking, Wuyao-Danshen was found to act on EMS through anti-inflammatory targets and related signaling pathways. Our findings provide a basis for further research.

Association between omega-3 fatty acid supplementation and lower risk of preterm delivery: a systematic review and meta-analysis.

OBJECTIVE: To explore whether omega-3 fatty acid supplementation is associated with lower risk of preterm delivery. METHODS: Searching the RCTs which were compared preterm birth between women with omega-3 fatty acid supplementation and without before December 2019 on Medline, EMBASE and Cochrane's Library, then performing a meta-analysis. RESULTS: 26 trials were identified, included 20124 women. There was almost no association between omega-3 fatty acid supplementation and lower risk of preterm delivery (risk ratio 0.92, 95% confidence interval 0.85 to 1.01, I2 = 9%), gestational duration (0.30, -0.05 to 0.64, I2 = 48%). In subgroup analyses, preterm delivery lower rate occurred in groups with mixed DHA and EPA supplementation not only DHA supplementation groups (P for interaction = 0.02); The dose of equivalent greater than 1 g made a higher reduction in preterm birth significantly. CONCLUSIONS: Omega-3 fatty acid supplementation was not associated with reduced risk of preterm delivery compared with placebo or no treatment during pregnancy. Relationship between Omega-3 fatty acid supplementation and other pregnant outcomes need more evidence and clinical studies.

Effect of Bushen Huoxue recipe on women with thin endometrial ovulation disorder and a rat model of thin endometrium resulted from kidney deficiency-related blood stasis.

To examine the therapeutic effect of Bushen Huoxue recipe (BHR) on women with thin endometrial ovulation disorder and on a rat model of kidney deficiency-related blood stasis. A total of 60 women with thin endometrial ovulation disorder was enrolled. The primary outcome of the study was the pregnancy rate three menstrual cycles after treatment. The study also examined the changes in the type and thickness of uterine artery, uterine artery pulsatility index (PI) and endometrial resistance index (RI). To establish kidney deficiency-related blood stasis in Sprague Dawley (SD) rats, an intragastric administration of hydroxyurea and a tail vein injection of Dextran were given, following with a flashing of the uterine cavity with 95% anhydrous ethanol. A combined regimen of BHR and estradiol valerate significantly increased the rate of pregnancy in women with thin endometrial ovulation disorder. The treatment was accompanied by a significant increase in endometrial thickness and decreases in uterine artery PI and endometrial RI. In rats, kidney deficiency-related blood stasis caused severe loss in endometrial architecture, thickness, and numbers of gland and blood vessel compared to the healthy SD rats. Treatment with BHR could ameliorate the endometrial damages associated with kidney deficiency-related blood stasis.

Expression of IL17A in endometrial carcinoma and effects of IL17A on biological behaviour in Ishikawa cells.

OBJECTIVE: A growing body of evidence suggests chronic inflammation triggers the process of endometrial carcinogenesis. Interleukin (IL) 17A is an important proinflammatory factor involved in the tumour angiogenesis processes of many solid tumours. This study aimed to characterize the function of IL17A in endometrioid-type endometrial carcinoma. METHODS: Levels of IL17A in human endometrial tissues were analysed by immunohistochemistry. In vitro proliferation and migration were analysed in Ishikawa cells treated with IL17A, using cell counting kit-8, wound healing and transwell assays. Western blots were used to analyse levels of oestrogen receptor (ER)α and ERß proteins in Ishikawa cells treated with IL17A. RESULTS: IL17A levels were significantly higher in endometrial carcinoma tissues than in endometrial hyperplasic tissues. Significantly increased proliferation and migration was observed in Ishikawa cells treated with IL17A versus controls. Investigation of the molecular mechanism revealed that IL17A treatment upregulated the ERα/ERß protein ratio in Ishikawa cells. CONCLUSIONS: IL17A may be an important proinflammatory factor involved in promoting endometrial carcinogenesis.

Flavonoids from Scutellaria barbata inhibit activation of tumor-associated macrophages by blocking the Toll-like receptor 4/myeloid differentiation factor 88/nuclear factor-κB signaling pathway.

OBJECTIVE: To determine the efficacy of Scutellaria barbata flavonoids and polysaccharides on Ishikawa endometrial carcinoma cells co-cultured with U937 macrophages. METHODS: The presence of CD163 and CD206 was determined by flow cytometry. Thiazolyl Blue Tetrazolium Bromide assays were used to assess the proliferation effect of tumor-associated macrophages (TAMs) on Ishikawa cells. The secretion of interleukin (IL)-10 in the co-culture conditioned media was examined using an enzyme-linked immunosorbent assay. The protein expression levels of Toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88) and nuclear factor (NF)-κB p65 were detected by Western blot. The mRNA expression levels of TLR4 and MyD88 were analyzed by real-time polymerase chain reaction (PCR). The expression levels of IL-12, IL-1ß and tumor necrosis factor-α (TNF-α) were evaluated with real-time PCR. RESULTS: Compared with the U937 control group, the expression levels of CD163 and CD206 in the TAM group were higher (P < 0.05). TAMs co-cultured with Ishikawa cells for 24 or 48 h showed higher proliferation rates (P < 0.05). The expression levels of IL-12 decreased than compared with those in the U937 untreated group (P < 0.05) and those of the Scutellaria barbata flavonoids group (P < 0.05). The expression levels of CD206, CD163, IL-10, IL-1ß and TNF-α, NF-κB p65 and TLR4/MyD88 in the TAMs control group were greater than those in the U937 untreated group (P < 0.05) and those of the Scutellaria barbata flavonoids group (P < 0.05). CONCLUSION: Scutellaria barbata flavonoids may inhibit TAM activation by blocking the TLR4/MyD88/NF-κB signaling pathway.

Homing and restorative effects of bone marrow-derived mesenchymal stem cells on cisplatin injured ovaries in rats.

Premature ovarian failure (POF) is a long-term adverse effect of chemotherapy treatment. However, current available treatment regimens are not optimal. Emerging evidence suggests that bone marrow-derived mesenchymal stem cells (BMSCs) could restore the structure and function of injured tissues, but the homing and restorative effects of BMSCs on chemotherapy injured ovaries are still not clear. In this study, we found that granulosa cell (GC) apoptosis induced by cisplatin was reduced when BMSCs were migrated to granulosa cells (GCs) in vitro. Chemotherapy-induced POF was induced by intraperitoneal injection of cisplatin in rats. BMSCs labeled with enhanced green fluorescent protein (EGFP) were injected into the rats via the tail vein to investigate the homing and distribution of BMSCs in vivo. The number of BMSCs in the ovarian hilum and medulla was greater than in the cortex, but no BMSCs were found in the follicles and corpus lutea. In addition, the BMSCs treatment group's antral follicle count and estradiol levels increased after 30 days, compared with the POF group. Hence, our study demonstrates that intravenously delivered BMSCs can home to the ovaries, and restore its structure and function in POF model rats.

Immediate analgesia effect of single point acupuncture in primary dysmenorrhea: a randomized controlled trial.

BACKGROUND: Acupuncture is often used for primary dysmenorrhea. OBJECTIVE: To assess the efficacy of a single point of acupuncture in the management of primary dysmenorrhea compared with sham acupuncture and no acupuncture. METHODOLOGY: Patients with primary dysmenorrhea were randomly assigned to acupoint group (n=50), unrelated acupoint group (n=50), nonacupoint group (n=46), or no acupuncture group (n=48). Acupuncture and sham acupuncture were administered once-daily for 3 days with electro-acupuncture at Sanyinjiao (SP6) that was specifically designed to treat primary dysmenorrhea, or an unrelated acupoint (Xuanzhong, GB39), or nonacupoint location. The primary outcome was pain intensity as measured by a 100-mm visual analog scale (VAS) at baseline; 5, 10, 30, and 60 minutes following the start of the first intervention. Cox retrospective symptom scale (RSS), verbal rating scale (VRS), pain total time, and proportion of participants using analgesics were also recorded during three menstrual cycles. RESULTS: The primary comparison of VAS scores demonstrated that patients receiving acupuncture (-15.56 mm, 95% CI -22.16 to -8.95, P<0.001), unrelated acupoint (-18.14 mm, 95% CI -24.81 to -11.47, P<0.001), and nonacupoint (-10.96 mm, 95% CI -17.62 to -4.30, P=0.001) treatment presented significant improvements compared with no acupuncture group. There were no significant differences among the four groups with respect to secondary outcomes. CONCLUSION: Acupuncture was better than no acupuncture for relieving the pain of dysmenorrhea following a single point of acupuncture, but no differences were detected between acupoint acupuncture and unrelated acupoint acupuncture, acupoint acupuncture and nonacupoint acupuncture.

[Effect of ferulic acid on proliferation and mechanism in human breast cancer cells].

OBJECTIVE: To investigate phytoestrogenic effects of ferulic acid in ER-positive T47D and ER-negative MDA-MB231 cells in culture. METHOD: T47D and MDA-MB231 human breast cancer cells were treated with ferulic acid and examined cell proliferation by means of MTT assay. Cell cycle distribution, ERalpha and ERbeta expression were treated by flow cytometer. The pS2 mRNA expressions were detected by real-time fluorescence quantitative PCR. RESULT: The proliferations were enhanced significantly by treatment with ferulic acid on T47D cells and the proliferation effects were inhibited by adding Faslodex (1 x 10(-8) mol x L(-1)). However, there was no significant difference on the proliferation in MDA-MB-231 cells compared with solvent control group by both treatment with ferulic acid and co-treatment with Faslodex (1 x 10(-8) mol x L(-1)). Ferulic acid stimulated the amount of T47D cells in phase S and proliferation index increased significantly. The effects were inhibited by treatment with Faslodex (1 x 10(-8) mol x L(-1)), and the amount of cells in phase S and proliferation index decreased, the amount of cells in G0/G1 phase increased, cell cycle of T47D was arrested in G0/G1 phase. Ferulic acid up-regulated pS2 mRNA expressions and increased the level of ERalpha protein expression in T47D cells. Ferulic acid did not show remarkable effect to the level of ERbeta protein expression in T47D cells. CONCLUSION: Ferulic acid possessed phytoestrogenic effect by up-regulating pS2 gene expression and the receptor subtype of ERalpha.

[Study on phytoestrogenic-like effects of four kinds of Chinese medicine including Radix Rehmanniae Preparata, Radix Paeoniae Alba, Radix Angelicae Sinensis, Rhizoma Chuanxiong].

OBJECTIVE: To explore the phytoestrogenic-like effects of four kinds of Chinese medicine including Radix rehmanniae preparata, Radix Paeoniae Alba, Radix Angelicae Sinensis, Rhizoma Chuanxiong. METHOD: Sixty immature female SD rats weighting (70 +/- 5) g were randomly divided into six groups: normal control group, positive control group and 4 Chinese medicine groups. The rats in different groups were treated for 4 days. On the fifth day, animals were sacrificed and uteri were separated solely and weighed. The blood was collected, and serum was separated. The effect of the pharmacological serum on proliferation assay with human breast cancer cell line (MCF7) by MTT method. Cell-cycle analyses were carried out with propidium iodide staining by flow cytometer. The expressions of subtypes-estrogen receptor-alpha (ERalpha) were detected by flow cytometry. RESULT: Radix Rehmanniae Preparata, Radix Paeoniae Alba and Radix Angelicae Sinensis could increase the immature rat's uterus wet weight and the ratio of uterus to body weight (P < 0.05). The pharmacological serum of the four kinds of Chinese medicine stimulated proliferation of MCF7 cell respectively compared with normal control group (P < 0.01). The cell cycle was impulsed from G1 to S, DNA synthesizing was inhanced, and PI was also increased. The pharmacological serum of Radix Angelicae Sinensis and Rhizoma Chuanxiong could increase the expressions of (ERalpha) (P < 0.05). CONCLUSION: Radix Rehmanniae Preparata, Radix Paeoniae Alba and Radix Angelicae sinensis have phytoestrogenic effects. But the data werent consistent with in vitro and in vivo assay of Rhizoma Chuanxiong.

[Regulation of genistein on the levels of ERalpha, ERbeta mRNA in uterine endometrial cancer cells].

OBJECTIVE: To explore the mechanism of phytoestrogen (genistein) on uterine endometrial cancer cells and provide the theory foundation for using phytoestrogen in clinical practice. METHODS: Uterine endometrial cancer cell lines (ishikawa) were cultured in vitro at different genistein concentration, and the expressions of the two estrogen subtypes of endometrial cancer cells were observed by real-time PCR. And two treatment groups, estradiol and progestin, were used as positive controls. RESULTS: (1) Low concentration of genistein could enhance the expression level of ERalpha mRNA of endometrial cancer cells, which was obviously weaker than that of the estradiol group. On the other hand, low concentration of genistein could decrease the expression level of ERbeta mRNA of endometrial cancer cells, however estradiol had hardly any effect on this. (2) High concentration of genistein could decline the expression levels of ERalpha and ERbeta mRNA of endometrial cancer cells, whose effect was weaker than that of progestin. CONCLUSION: The expression levels of ERalpha mRNA and ERbeta mRNA of endometrial cancer cells vary with genistein concentrations. The experimental results suggest that genistein may act as estrogen receptor regulator.

[Effects of extracts of root of kudzu vine on mammary gland and uterus development in rats].

OBJECTIVE: To study the effects of extracts of root of kudzu vine on mammary gland and uterus development in rats. METHOD: 40 Wistar rats weighting 65-85 g were randomly divided into 4 groups: control group, estrogen group, extracts of root of kudzu vine group of high dose, extracts of root of kudzu vine group of low dose. (10 rats in each group). After having been treated for 7 days, the rats were killed; mammary glands and uterus were removed and weighed. Serum was isolated and kept at 4 degrees C for determination of hormones. RESULT: 1. Administration of the root of kudzu vine significantly increased the weigh of mammary gland and uterus in rats. 2. Administration of the root of kudzu vine increased serum FSH, LH, E2 and decreased PRL. CONCLUSION: Extracts of root of kudzu vine could enhance the weight of mammary gland and uterus growth in rats, which may provide experimental evidence for the development of new drug used for promoting mammary gland and uterus.