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In Vitro Culture of Epithelial Cells from Different Anatomical Regions of the Human Amniotic Membrane
Published on: November 28, 2019
The effects of amniotic membrane on polymorphonuclear cells
Shiyou Zhou1, Jiaqi Chen, Jinfa Feng
1Department of Cornea, Zhongshan Ophthalmic Center, Sun Yet-Sen University, Guangzhou 510060, China. gdeyeb@gzsums.edu.cn
Objective:
To investigate the effects of fresh and preserved amniotic membrane on polymorphonuclear neutrophils (PMNs) so as to understand the anti-inflammatory mechanism of amniotic membrane transplantation.
Methods:
Conditioned medium was collected 48 hours after fresh or preserved amnions were cultured in DMEM and 5% CO(2) at 37 degrees C. Then, polymorphonuclear cells were cultured in conditioned culture or DMEM. Fluorescent microscopy with 4',6-diamidino-2-phenylindole (DAPI) staining and cytometry were performed 6, 9, 12, and 15 hours later.
Results:
Apoptotic neutrophils were found in each group at different time points. The percentage of apoptotic cells at 6, 9, 12, and 15 hours after culture in the fresh and preserved amnion groups and the control group was 17.3%, 24.4%, 29.8%, 37.1%, and 16.2%, 20.1%, 23.7%, 27.7%, and 10.2%, 13.7%, 21.1%, 26.4%, respectively (t test, P(1) < 0.01, P(2) < 0.01 and P(3) < 0.01).
Conclusion:
Amniotic membrane can accelerate apoptosis of polymorphonuclear neutrophils, reduce inflammation, and prevent ocular surface collagen from resolution, indicating that fresh amnion might have a stronger effect than preserved amnion.
Insights
Amniotic membrane accelerates the apoptosis of polymorphonuclear neutrophils (PMNs), reducing inflammation. Fresh amniotic membrane shows a stronger anti-inflammatory effect than preserved amnion, aiding ocular surface healing.
Area of Science:
- Ophthalmology
- Cell Biology
- Immunology
Background:
- Ocular surface inflammation involves polymorphonuclear neutrophils (PMNs).
- Amniotic membrane transplantation is used to treat ocular surface diseases.
- The anti-inflammatory mechanisms of amniotic membrane require further elucidation.
Purpose of the Study:
- To investigate the effects of fresh and preserved amniotic membrane on PMN apoptosis.
- To understand the anti-inflammatory mechanism of amniotic membrane transplantation.
Main Methods:
- Cultured fresh and preserved amnions in DMEM.
- Collected conditioned medium after 48 hours.
- Incubated PMNs in conditioned medium or DMEM, analyzing apoptosis via fluorescent microscopy and cytometry at various time points.
Main Results:
- Both fresh and preserved amniotic membranes accelerated PMN apoptosis compared to the control group.
- The percentage of apoptotic neutrophils increased over time in all groups.
- Fresh amniotic membrane demonstrated a trend towards a stronger pro-apoptotic effect on PMNs.
Conclusions:
- Amniotic membrane accelerates PMN apoptosis, contributing to reduced inflammation.
- This process helps prevent ocular surface collagen degradation.
- Fresh amniotic membrane may possess superior anti-inflammatory properties compared to preserved forms.
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