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This dissertation, The Study of Chinese Herbal Medicinal Compound on Implantation: in Vitro Spheroid-endometrium Co-culture by Hoi-yan, Cheung, 張凱恩, was obtained from The University of Hong Kong (Pokfulam, Hong Kong) and is being sold pursuant to Creative Commons: Attribution 3.0 Hong Kong License. The content of this dissertation has not been altered in any way. We have altered the formatting in order to facilitate the ease of printing and reading of the dissertation. All rights not granted by the above license are retained by the author. Abstract: Traditional Chinese medicine (TCM) plays an important role in the Chinese healthcare system for over five thousand years. It includes the use of herbal medicine, acupuncture, Tui Na (推拿), and diet therapy. TCM helps to maintain a balance of Yin-Yang (阴阳), Five Phases (五行), Meridians (经络) and Qi (气) inside the body. In practise, pregnant women take tocolytic drugs to tonify the blood and qi to provide a continuous supply of nutrients for baby. Traditional Chinese herbal medicines usually prescribed as a complex formula to produce synergistic or agonistic effect to maintain a well balance of the above components in human bodies. Moreover, TCM usually cannot produce immediate effect on patients, therefore, the efficacy of individual component remains largely unknown. This study aims to investigate whether Chinese tocolytic drug components could modulate fertility by affecting the in vitro spheroid (blastocyte surrogate) attachment process by using trophoblastic (JEG-3) and endometrial epithelial (Ishikawa) cells to mimic the embryo-endometrial implantation process. Nine Chinese herbal medicinal compounds (Atractylenolide I(白术内酯), Atractylenolide II(白术内酯II), Atractylenolide III(白术内酯III), Paeoniflorin(芍药苷), Albiflorin(芍药内酯苷), Nuzhenide(女贞子甙), Ecliptasaponin A(旱莲甙A), Wedelolactone(蟛蜞菊内酯) and Columbianadin(二氢欧山芹醇当归酸酯)) which are commonly found in traditional Chinese tocolytic drug formula were selected to study (1) the toxicity of the drugs on trophoblastic (JEG-3) and endometrial epithelial (Ishikawa) cells growth, (2) the effect of three tocolytic drugs (Atractylenolide I, Atractylenolide II and Atractylenolide III) on spheroid attachment, and (3) their effect of the expression of Wingless (Wnt) signaling molecules (Active-β-Catenin, Axin-2, β-catenin, E-cadherin, GSK-3β, and Mucin-1). It was found that the nine compounds, Atractylenolide I, Atractylenolide II, Atractylenolide III, Paeoniflorin, Albiflorin, Nuzhenide, Ecliptasaponin A, Wedelolactone and Columbianadin did not affect cell viability at 25μM, 25μM, 5μM, 0.2μM, 125μM, 125μM, 125μM, 5μM and 25μM, respectively, by cell proliferation assay. However, at these concentrations, the spheroid attachment was not significantly increased by Atractylenolide I, Atractylenolide II and Atractylenolide III. Interestingly, the protein expression of GSK-3β and Active-β-catenin were up-regulated by the three compounds in both cells and JEG-3 cells respectively. The expressions of Axin-2 and E-cadherin were up-regulated by Atractylenolide III in Ishikawa cells and Atractylenolide II in JEG-3 cells. Atractylenolide I and Atractylenolide III increase the Ishikawa cells expression of Active-β-catenin and β-catenin respectively and together suppress the JEG-3 cells Mucin-1 and β-catenin expression. In conclusion, the nine tocolytic compounds have different effect on cell proliferation. Atractylenolide I, Atractylenolide II and Atractylenolide III did not enhance the attachment rate of JEG-3 spheroid onto Ishikawa monolayer. However, they affected Wnt-signaling molecules expression, suggesting that they may modulate endometrial receptivity. Further experiments are needed to study their combined effect on co-culture and expression of Wnt-signaling molecules. DOI: 10.5353/th_b515

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