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Mater Sci Eng C Mater Biol Appl. 2019 Nov;104:109903. doi: 10.1016/j.msec.2019.109903. Epub 2019 Jun 24.

Comparison of the impact of preservation methods on amniotic membrane properties for tissue engineering applications.

Author information

1
Univ. Bordeaux, INSERM, Laboratory BioTis, UMR 1026, F-33076 Bordeaux, France; CHU Bordeaux, Department of Oral Surgery, F-33076 Bordeaux, France. Electronic address: mathilde.fenelon@u-bordeaux.fr.
2
Univ. Bordeaux, INSERM, Laboratory BioTis, UMR 1026, F-33076 Bordeaux, France.
3
CHU Bordeaux, CIC 1401, 33000 Bordeaux, France; Inserm, CIC 1401, 33000 Bordeaux, France.
4
Univ. Bordeaux, INSERM, Laboratory BioTis, UMR 1026, F-33076 Bordeaux, France; CHU Bordeaux, CIC 1401, 33000 Bordeaux, France; Inserm, CIC 1401, 33000 Bordeaux, France.
5
University hospital, Gynecology-Obstetrics Service, F-33076 Bordeaux, France.
6
Univ. Bordeaux, INSERM, Laboratory BioTis, UMR 1026, F-33076 Bordeaux, France; CHU Bordeaux, Department of Oral Surgery, F-33076 Bordeaux, France.
7
Orthopedic, Traumatology & Plastic Surgery Department, University Hospital of Besançon, Besançon, France; Nanomedicine Lab, Imagery and Therapeutics (EA 4662), SFR FED 4234, University of Franche-Comté, Besançon, France.

Abstract

Human amniotic membrane (hAM) is considered as an attractive biological scaffold for tissue engineering. For this application, hAM has been mainly processed using cryopreservation, lyophilization and/or decellularization. However, no study has formally compared the influence of these treatments on hAM properties. The aim of this study was to develop a new decellularization-preservation process of hAM, and to compare it with other conventional treatments (fresh, cryopreserved and lyophilized). The hAM was decellularized (D-hAM) using an enzymatic method followed by a detergent decellularization method, and was then lyophilized and gamma-sterilized. Decellularization was assessed using DNA staining and quantification. D-hAM was compared to fresh (F-hAM), cryopreserved (C-hAM) and lyophilized/gamma-sterilized (L-hAM) hAM. Their cytotoxicity on human bone marrow mesenchymal stem cells (hBMSCs) and their biocompatibility in a rat subcutaneous model were also evaluated. The protocol was effective as judged by the absence of nuclei staining and the residual DNA lower than 50 ng/mg. Histological staining showed a disruption of the D-hAM architecture, and its thickness was 84% lower than fresh hAM (p < 0.001). Despite this, the labeling of type IV and type V collagen, elastin and laminin were preserved on D-hAM. Maximal force before rupture of D-hAM was 92% higher than C-hAM and L-hAM (p < 0.01), and D-hAM was 37% more stretchable than F-hAM (p < 0.05). None of the four hAM were cytotoxic, and D-hAM was the most suitable scaffold for hBMSCs proliferation. Finally, D-hAM was well integrated in vivo. In conclusion, this new hAM decellularization process appears promising for tissue engineering applications.

KEYWORDS:

Acellular scaffold; Amniotic membrane; Cryopreservation; Freeze-drying; Processed amnion; Rat; in vivo biocompatibility

PMID:
31500032
DOI:
10.1016/j.msec.2019.109903

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