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[Evaluation of recellularization level of bioprosthetic valve scaffold with AGNO3 staining technique in vitro]
Bin Feng1, Yinglong Liu, Ning Xie
1Department of Cardiac Surgery, Cardiovascular Institute and Fuwai Hospital, Chinese Academy of Medical Science and Beijing Union Medical College, Beijing, 100037, P. R. China. fsn9977@yahoo.com.cn
Objective:
To explore morphological recellularization level of bioprosthetic valve scaffold (BVS) and to provide researching means for fabricating tissue engineered heart valve in vitro.
Methods:
The homograft bioprosthetic aortic tube valve was selected as BVS, which was conserved by liquid nitrogen, and its endothelial cells (ECs) were removed by 0.1% sodium dodecylsulphate (SDS). As implantation cells, the endothelial cells (ECs) differentiating from human bone marrow mesenchymal stem cells (MSCs) in vitro were implanted with high-density seeding (>10(5) cells/cm2) on the BVS, which was covered by fibronectin (80 microg/ml) in advance. The complex structure was statically cultured in DMEM (high glucose) with 20% FBS and VEGF (10 ng/ml) for about 20 days in vitro and stained by 0. 5% AgNO3. The morphological structure was observed and photographed by stereomicroscope to detect the recellularization level.
Results:
The ECs of the bioprosthetic valve were not only removed completely, but also the collagen fiber and elastic fibers were reserved. The ECs differentiating from MSCs were successfully implanted on the HBS, whose recellularization levels on 7th, 14th and 20th day were 73%, 85%, and 92% respectively.
Conclusion:
AgNO3 staining technique is effective, convenient, and economic in evaluating the recellularization level of BVS. It is an effective method in morphological observation for fabricating tissue engineered heart valve in vitro.
