Cryopreserved amniotic membrane as transplant allograft: viability and post-transplant outcome

Abstract

Amniotic membrane (AM) transplantation is increasingly used in ophthalmological and dermatological surgeries to promote re-epithelialization and wound healing. Biologically active cells in the epithelial and stromal layers deliver growth factors and cytokines with anti-inflammatory, anti-bacterial, anti-immunogenic and anti-fibrotic properties. In this work, confocal microscopy was used to show that our cryopreservation protocol for AM yielded viable cells in both the stromal and epithelial layers with favorable post-transplant outcome. AM was obtained from Caesarean-section placenta, processed into allograft pieces of different sizes (3 cm × 3 cm, 5 cm × 5 cm, and 10 cm × 10 cm) and cryopreserved in 10 % dimethyl sulfoxide using non-linear controlled rate freezing. Post-thaw cell viability in the entire piece of AM and in the stromal and epithelial cell layers was assessed using a dual fluorescent nuclear dye and compared to hypothermically stored AM, while surveys from surgical end-users provided information on post-transplant patient outcomes. There was no significant statistical difference in the cell viability in the entire piece, epithelial and stromal layers regardless of the size of allograft piece (p = 0.092, 0.188 and 0.581, respectively), and in the entire piece and stromal layer of hypothermically stored versus cryopreserved AM (p = 0.054 and 0.646, respectively). Surgical end-user feedback (n = 49) indicated that 16.3 % of AM allografts were excellent and 61.2 % were satisfactory. These results support the expanded clinical use of different sizes of cryopreserved AM allografts and address the issue of orientation of the AM during transplant for the treatment of dermatological defects and ocular surface disorders.

Properties of viable lyopreserved amnion are equivalent to viable cryopreserved amnion with the convenience of ambient storage

“…VCAM retrospective and prospective clinical studies reported positive clinical outcomes in patients with acute and chronic wounds of various etiologies and in a variety of surgical procedures without adverse events attributed to VCAM application [ 22 – 25 ]. Data from these VCAM clinical studies are consistent with other reports describing the clinic use of DMSO-cryopreserved viable AM prepared using different cryopreservation protocols [ 48 , 49 ]. In a randomized controlled clinical trial, 75% of pressure ulcers were closed in the cryopreserved AM group versus 0% in the control group [ 48 ].…”

“…In a randomized controlled clinical trial, 75% of pressure ulcers were closed in the cryopreserved AM group versus 0% in the control group [ 48 ]. A different retrospective study reported positive outcomes with DMSO-cryopreserved viable AM for a broad variety of ocular diseases and dermatological defects without adverse events [ 49 ].…”

Cryopreservation of Human Mesenchymal Stem Cells in an Allogeneic Bioscaffold based on Platelet Rich Plasma and Synovial Fluid

“…Penetrant CPAs, such as DMSO, displace the internal water from the cell, minimizing the intracellular ice crystal formation while, non-penetrant CPAs, such as sucrose, act from the outside of the cells, promoting their dehydration 23 . DMSO shows several disadvantages in the preservation process such as toxicity or loss of multipotency 41 , but its replacement is difficult since no other CPA has shown the same results maintaining embedded cell viabilities after cryopreservation 42 . However, the combination of DMSO and sucrose enhance also the post-thawing viability of human MSCs 43 , 44 .…”

Lyopreserved amniotic membrane is cellularly and clinically similar to cryopreserved construct for treating foot ulcers

“…Our independent results were similar to other published findings that compared cryopreserved and lyopreserved amniotic tissue from this construct. 18 The primary outcome of the prospective cohort study was to evaluate the proportion of ulcers that achieved complete closure in 12 weeks in patients treated with LAM. The proportion of patients who achieved complete closure was 48%.…”