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Updated: Jun 27, 2026
Isolation of Human Endometrial Stromal Cells for In Vitro Decidualization
Published on: September 1, 2018
[Influence of nuclear factor-kappaB decoy oligonucleotides on RANTES expression and monocyte chemotactic activity in
Xiu-li Wang1, Su-ping Han, Chun-li Fang
1Department of Obstetrics and Gynecology, First Affiliated Hospital of Nanjing Medical University, Nanjing 210036, China. xiuli_2266@163.com
Objective:
To study the inhibitory effect on the expression of regulated upon activation, normal T cell expressed and secreted (RANTES) and monocyte chemotactic activity of ectopic endometrial stromal cells by nuclear factor (NF)-kappaB decoy oligonucleotides (ODN).
Methods:
The stromal cells of ectopic endometrium were divided into 3 groups. Two groups were cultured with or without 10 microg/L of interleukin (IL)-1beta. Another group was transfected with NF-kappaB decoy ODN with the aid of a lipofectamine reagent. After 4 h of transfection, 10 microg/L of IL-1beta was added to induce the stromal cells to secrete RANTES. Concentration of RANTES in the supernatant at 4, 8, 12, 24 and 36 h was measured with the sandwich enzyme linked immunosorbent assay (ELISA). U937 monocyte chemotactic activity was assayed in Boyden chambers. The specific RANTES-neutralizing monoclonal antibodies at serial doses (0.5, 1, 2, 4 and 8 mg/L) were added into IL-1beta induced medium of 24 h to detect the monocyte chemotactic activity of RANTES in supernatant.
Results:
The concentration of RANTES secreted by stromal cells was respectively (58 +/- 10), (150 +/- 35), (360 +/- 46) and (586 +/- 42) ng/L after IL-1beta stimulation for 8, 12, 24 and 36 h, significantly higher than that of stromal cells cultured without IL-1beta. The concentrations of RANTES were respectively (86 +/- 16), (128 +/- 28) and (183 +/- 32) ng/L after IL-1beta stimulation for 12, 24 and 36 h in stromal cells transfected with NF-kappaB decoy ODN, evidently lower than that of stromal cells stimulated with IL-1beta alone. The monocyte chemotactic index of 12, 24, 36 h in conditioned medium of stromal cells transfected with NF-kappaB decoy ODN was respectively 10.3 +/- 0.9, 13.7 +/- 1.1, 18.6 +/- 1.2, which was evidently lower than that of stromal cells stimulated with IL-1beta alone. The anti-RANTES antibody at 0.5, 1, 2, 4 and 8 mg/L inhibited respectively 5%, 23%, 40%, 62% and 61% of the chemotactic activity in 12 h medium treated with IL-1beta.
Conclusions:
RANTES accounts for the majority of the monocyte chemotactic activity in IL-1beta induced medium of 24 h. NF-kappaB decoy ODN may influence the feed-forward inflammatory loop whereby IL-1beta from activated macrophages can lead to RANTES production by ectopic implants and further monocyte chemotaxis.
Insights
Nuclear factor-kappaB (NF-kappaB) decoy oligonucleotides (ODN) reduce regulated upon activation, normal T cell expressed and secreted (RANTES) production and monocyte chemotactic activity in ectopic endometrial stromal cells. This suggests NF-kappaB decoy ODN can disrupt inflammatory loops in endometriosis.
Area of Science:
- Reproductive biology
- Immunology
- Molecular biology
Context:
- Endometriosis is characterized by ectopic endometrial stromal cells.
- Interleukin-1beta (IL-1beta) stimulates RANTES production in these cells.
- RANTES plays a role in monocyte recruitment.
Purpose:
- To investigate the inhibitory effect of NF-kappaB decoy ODN on RANTES expression and monocyte chemotactic activity.
- To elucidate the role of NF-kappaB signaling in RANTES-mediated inflammation in endometriosis.
Summary:
- Ectopic endometrial stromal cells stimulated with IL-1beta showed increased RANTES secretion and monocyte chemotactic activity.
- Transfection with NF-kappaB decoy ODN significantly reduced RANTES levels and monocyte chemotaxis.
- RANTES was identified as a major mediator of IL-1beta-induced monocyte chemotactic activity.
Impact:
- NF-kappaB decoy ODN demonstrate potential as a therapeutic strategy for endometriosis.
- Understanding the NF-kappaB/RANTES axis offers insights into endometriosis pathogenesis.
- This study highlights a potential mechanism to modulate inflammatory processes in ectopic endometrial implants.
