Vitrification of Intact Porcine Femoral Condyle Allografts Using an Optimized Approach

Wu, Kezhou; Laouar, Leila; Elliott, Janet A.W.; Jomha, Nadr M.

doi:10.1177/1947603520967077

Cited by 7 publications

(12 citation statements)

References 30 publications

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“…Our data revealed the inability of the traditional convection method for large OCA preservation and showed that improving the heating rate is needed to achieve large cartilage cryopreservation. Our findings are aligned with a recent literature report 47 . In this prior literature study, CPA solutions surrounding the condyle were depleted, enabling faster heat transfer due to direct contact between the cartilage and water bath 47 .…”

Section: Discussionsupporting

confidence: 93%

“…Our findings are aligned with a recent literature report 47 . In this prior literature study, CPA solutions surrounding the condyle were depleted, enabling faster heat transfer due to direct contact between the cartilage and water bath 47 . In contrast, in our nanowarming method, the surrounding CPAs remained and were rapidly warmed through nanowarming to achieve fast heat transfer and an elevated heating rate.…”

Section: Discussionsupporting

confidence: 93%

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Nanowarming and ice-free cryopreservation of large sized, intact porcine articular cartilage

et al. 2023

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Successful organ or tissue long-term preservation would revolutionize biomedicine. Cartilage cryopreservation enables prolonged shelf life of articular cartilage, posing the prospect to broaden the implementation of promising osteochondral allograft (OCA) transplantation for cartilage repair. However, cryopreserved large sized cartilage cannot be successfully warmed with the conventional convection warming approach due to its limited warming rate, blocking its clinical potential. Here, we develope a nanowarming and ice-free cryopreservation method for large sized, intact articular cartilage preservation. Our method achieves a heating rate of 76.8 °C min-1, over one order of magnitude higher than convection warming (4.8 °C min-1). Using systematic cell and tissue level tests, we demonstrate the superior performance of our method in preserving large cartilage. A depth-dependent preservation manner is also observed and recapitulated through magnetic resonance imaging and computational modeling. Finally, we show that the delivery of nanoparticles to the OCA bone side could be a feasible direction for further optimization of our method. This study pioneers the application of nanowarming and ice-free cryopreservation for large articular cartilage and provides valuable insights for future technique development, paving the way for clinical applications of cryopreserved cartilage.

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Section: Discussionsupporting

confidence: 93%

Section: Discussionsupporting

confidence: 93%

Nanowarming and ice-free cryopreservation of large sized, intact porcine articular cartilage

et al. 2023

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“…The ImageJ software was then used to count the cells in the images, and chondrocyte viability was calculated as the percentage of live cells relative to the total number of cells. In each cartilage slice, the chondrocyte viability was determined as the percentage of green cells relative to the total number of green and red cells 28,35 . The experimental procedures were repeated two times.…”

Section: Methodsmentioning

confidence: 99%

High‐Speed Centrifugation Efficiently Removes Immunogenic Elements in Osteochondral Allografts

Ma,

Lin,

Yang

et al. 2024

Orthopaedic Surgery

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ObjectivesThe current clinical pulse lavage technique for flushing fresh osteochondral allografts (OCAs) to remove immunogenic elements from the subchondral bone is ineffective. This study aimed to identify the optimal method for removing immunogenic elements from OCAs.MethodsWe examined five methods for the physical removal of immunogenic elements from OCAs from the femoral condyle of porcine knees. We distributed the OCAs randomly into the following seven groups: (1) control, (2) saline, (3) ultrasound, (4) vortex vibration (VV), (5) low‐pulse lavage (LPL), (6) high‐pulse lavage (HPL), and (7) high‐speed centrifugation (HSC). OCAs were evaluated using weight measurement, micro‐computed tomography (micro‐CT), macroscopic and histological evaluation, DNA quantification, and chondrocyte activity testing. Additionally, the subchondral bone was zoned to assess the bone marrow and nucleated cell contents. One‐way ANOVA and paired two‐tailed Student's t‐test are used for statistical analysis.ResultsHistological evaluation and DNA quantification showed no significant reduction in marrow elements compared to the control group after the OCAs were treated with saline, ultrasound, or VV treatments; however, there was a significant reduction in marrow elements after LPL, HPL, and HSC treatments. Furthermore, HSC more effectively reduced the marrow elements of OCAs in the middle and deep zones compared with LPL (p < 0.0001) and HPL (p < 0.0001). Macroscopic evaluation revealed a significant reduction in blood, lipid, and marrow elements in the subchondral bone after HSC. Micro‐CT, histological analyses, and chondrocyte viability results showed that HSC did not damage the subchondral bone and cartilage; however, LPL and HPL may damage the subchondral bone.ConclusionHSC may play an important role in decreasing immunogenicity and therefore potentially increasing the success of OCA transplantation.

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“… 7 , 9 , 19 , 25 Multiple attempts have been made to cryopreserve AC; however, it was not until recently that successful cryopreservation protocols were developed. 7 , 9 , 18 , 19 , 37 , 39 Optimized protocols for the long-term storage of osteochondral dowels 18 and more recently particulated AC cubes (~1-mm 3 cubes) have been developed. 37 Based on extensive previous research on the vitrification of AC, # mathematical modeling has been used to identify and screen potential loading parameters that included cryoprotectant concentration, exposure time, toxicity, and temperature.…”

mentioning

confidence: 99%

“… 7 , 9 , 18 , 19 , 37 , 39 Optimized protocols for the long-term storage of osteochondral dowels 18 and more recently particulated AC cubes (~1-mm 3 cubes) have been developed. 37 Based on extensive previous research on the vitrification of AC, # mathematical modeling has been used to identify and screen potential loading parameters that included cryoprotectant concentration, exposure time, toxicity, and temperature. 39 Protocols were tested on swine and human particulated cartilage to ensure that chondrocyte viability was maintained (>80% viable) after vitrification and rewarming.…”

mentioning

confidence: 99%

Vitrified Particulated Articular Cartilage for Joint Resurfacing: A Swine Model

Yong

Ead

et al. 2022

Am J Sports Med

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Background: The use of particulated articular cartilage for repairing cartilage defects has been well established, but its use is currently limited by the availability and short shelf life of donor cartilage. Vitrification is an ice-free cryopreservation technology at ultralow temperatures for tissue banking. An optimized vitrification protocol has been developed for particulated articular cartilage; however, the equivalency of the long-term clinical efficacy of vitrified particulated articular cartilage compared with fresh articular cartilage has not yet been determined. Hypothesis: The repair effect of vitrified particulated cartilage from pigs would be equivalent to or better than that of fresh particulated cartilage stored at 4°C for 21 days. Study Design: Controlled laboratory study. Methods: A total of 19 pigs were randomly divided into 3 experimental groups: fresh particulated cartilage group (n = 8), vitrified particulated cartilage group (n = 8), and negative control group (no particulated cartilage in the defect; n = 3). An additional pig was used as the initial cartilage donor for the first set of surgical procedures. Pigs were euthanized after 6 months to obtain femoral condyles, and the contralateral condyle was used as the positive (no defect) control. Samples were evaluated for gross morphology using the Outerbridge and Osteoarthritis Research Society International (OARSI) scoring systems, histology (safranin O, collagen type I/II, DAPI), and chondrocyte viability using live-dead membrane integrity staining. Results: There were no infections after surgery, and all 19 pigs were followed for the duration of the study. The OARSI grades for the fresh and vitrified particulated cartilage groups were 2.44 ± 1.35 and 2.00 ± 0.80, respectively, while the negative control group was graded significantly higher at 4.83 ± 0.29. Analysis of histological and fluorescent staining demonstrated that the fresh and vitrified particulated cartilage groups had equivalent regeneration within cartilage defects, with similar cell viability and densities and expression of proteoglycans and collagen type I/II. Conclusion: The implantation of fresh or vitrified particulated cartilage resulted in the equivalent repair of focal cartilage defects when evaluated at 6 months after surgery. Clinical Relevance: The vitrification of particulated cartilage is a viable option for long-term storage for cartilage tissue banking and could greatly increase the availability of donor tissue for transplantation.

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Vitrification of Intact Porcine Femoral Condyle Allografts Using an Optimized Approach

Cited by 7 publications

References 30 publications

Nanowarming and ice-free cryopreservation of large sized, intact porcine articular cartilage

Nanowarming and ice-free cryopreservation of large sized, intact porcine articular cartilage

High‐Speed Centrifugation Efficiently Removes Immunogenic Elements in Osteochondral Allografts

Vitrified Particulated Articular Cartilage for Joint Resurfacing: A Swine Model

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