Page 28 - HKSEMR2020 Programme book

Poster Presentation (Basic Science) Abstracts

Investigating the role of notch signaling in endometrial

mesenchymal stem-like cells

1,2
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1
Sisi Zhang , RWS Chan , EHY Ng , WSB Yeung 2
1.Department of Obstetrics and Gynecology, University of Hong Kong, Hong Kong, SAR, China.; 2.Shenzhen Key
Laboratory of Fertility Regulation, The University of Hong Kong Shenzhen Hospital, Shenzhen, China

Introduction / Background / Objectives: Western blotting confirmed the activation or inhibition of Notch-
related proteins in eMSCs.
Human endometrium undergoes cycles of proliferation,
differentiation and shedding during the female reproductive years.
Endometrial mesenchymal stem-like cells (eMSCs) contribute to Results / Outcomes:
this regenerative process. Notch signaling pathway is known to
play a vital role in cell fate decisions in many somatic stem cells. The expression of Notch target gene HES-1 was significantly
However, its role in endometrial stem cells remains unclear. We higher in eMSCs compared with unfractionated and progenitor
first examined the gene and protein expression related to Notch cells (n = 12, P < 0.01). The gene expression levels of NOTCH1,
signaling in three subpopulations of endometrial stromal cells: NOTCH2, NOTCH3, HEY-1, HEY-2 and HEY-L were similar (n =
unfractionated stromal cells, progenitor cells (CD140b+CD146- 10). Immunofluorescence revealed that stem cells expressed
cells) and eMSCs (CD140b+CD146+ cells). Second, the importance more Notch 1 than other stromal subpopulations (n = 5, P <
of Notch signalling by gain or loss of function approaches in eMSCs 0.01). Notch signalling relies on the communication between two
was evaluated. adjacent cells. EMSCs cultured at higher density expressed more
Notch intracellular domain (NICD) than cells at low density (n =
5, P < 0.05). Inhibition of Notch with DAPT significantly reduced
Methods: the proportion of eMSCs at high seeding density compared to
untreated cells (n = 6, P < 0.01). The relative percentage of the
Full-thickness endometrial tissues were obtained from women
undergoing hysterectomy. After mechanical and enzymatic cells co-expressing CD140b and CD146 obviously increased
dissociation, endometrial stromal cells were purified from epithelial when Notch was activated by Jagged-1 (n = 6, P < 0.01). Western
cells using EpCAM magnetic beads, and leukocytes were removed blotting confirmed that Jagged-1 up-regulated the expression of
using CD45 magnetic beads. The eMSCs were then obtained Notch-related proteins NICD, HES-1 and HEY-2 in eMSCs, while
using CD140b and CD146 magnetic beads. The gene and protein DAPT showed the opposite effect (n = 5, P < 0.05).
expression of Notch signalling in three stromal subpopulations
were evaluated by quantitative PCR and immunofluorescence, Conclusion:
respectively. For functional assays, eMSCs were seeded at different
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densities (1000, 2000, 4000 cells/cm ) and cultured for 7 days. The eMSCs can activate endogenous Notch signalling and may
Flow cytometry was used to assess the phenotypic expression have a role in cell-fate specification of stem cells.
of eMSCs after Notch activation (Jagged-1) or inhibition (DAPT).

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