Purchase this article with an account.
L. Zhu, Sr.; Immunosuppressive Property of Dried Human Amniotic Membrane. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4301.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
To report the immunosuppressive property of dry human amniotic membrane (dHAM).
All animal procedures were conducted in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research.Spleens were harvested from fresh -sacrificed, 15 week old, Balb/c mice. Splenocytes were collected after lysis of red blood cells using RBC lysis buffer (eBiosciences, San Diego, CA). Fifteen wells of a single 96- well plate were coated with functional-grade anti-mouse CD3e antibodies (eBiosciences, San Diego, CA) 1 day before splenocyte application and another 5 wells were used as control without antibodies coating. The splenocytes (5 x 105 cells per well) were plated in each well (n=20) and a small piece (2 mm x 2 mm) of dHAM (Ambiodry 1 or 2, OKTO Ophtho, Inc, Costa Mesa, CA) was added to 5 study wells each (n=10). After 4 days of incubation, the MTT (3-(4,5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide) assay (Cell Growth Determination Kit, MTT based, Sigma, St. Louis, MO) was performed according to the manufacturer’s instructions. The absorbance was measured at 590nm using a standard plate reader (Synergy HT, Biotek, Winooski, VT).The histologic examination was performed.
On histologic examination, Ambiodry 1 lacked an epithelial layer and consisted of an even thickness of stroma in contrast to Ambiodry 2, which had a well preserved monolayer of amniotic membrane epithelium. Both types of dHAMs significantly decreased mouse splenocyte proliferation in response to anti-mouse CD3e antibodies, and Ambiodry 2 did so to a significantly greater extent. dHAMs significantly suppressed mouse splenocyte proliferation compared to control. The suppression by dHAM with intact amniotic epithelium (Ambiodry 2) was significantly stronger than dHAM without epithelium(Ambiodry 1).
Human amniotic membrane has been reported to produce various immunosuppressive cytokines such as IL-4, IL-10, TGF . Moreover, supernatant from amniotic epithelial culture has been reported to suppress inflammation.This study showed that the immunosuppressive property of dHAM was demonstrated using an allo-reactive splenocyte proliferation assay. The devitalized dry preparation of human amniotic membrane maintained its immunosuppressive property and that this characteristic was potentiated by the preservation of the amniotic epithelium.Thus both amniotic membrane epithelium and stroma appear to be important for achieving optimal immunosuppression by amniotic membrane.
This PDF is available to Subscribers Only