Applied Tissue Technologies (United States)

Partial heart transplantation (PHT) is a new type of transplant that delivers growing heart valve implants for children. However, the acceptable ischemia time for PHTs remains unexplored. Therefore, the ischemia time for PHTs is empirically limited to orthotopic heart transplant (OHT) ischemia time of 4-6 h because the valves contained in OHTs are known to grow. This limits the distance from where PHT grafts can be procured. Without longer procurement distances, children who need PHT must wait a long time for suitable donor hearts. We previously demonstrated that PHTs remain viable for an ischemia time of 48 h in a rat model. Here we expand on this work in a porcine model. Porcine pulmonary valve (PV) and aortic valve (AV) leaflets were preserved in DMEM culture medium, Belzer UW, Unisol, or HTK solution (n = 6/group) at 4°C. At preset intervals, the cellular viability was measured using the alamarBlue assay. The valves were also analyzed with flow cytometry and histology. While the metabolic activity of the valves slowly decreased over time, there was significant cellular viability for up to 21 days of cold preservation with Belzer UW solution. Flow cytometry and histology further corroborated the results, showing cellular preservation at 7 days of ischemia time. OHT preservation is limited to only 4-6 h because longer ischemia times are associated with primary graft dysfunction from reduced contractility of ventricular myocardial cells. In contrast, PHTs spare the native ventricles. Our results indicate that PHT grafts remain viable much longer than OHT grafts. Invivo data are needed to verify that PHT grafts can grow and function after this significantly increased ischemic time.

Transporting tissues and organs from the site of donation to the patient in need, while maintaining viability, is a limiting factor in transplantation medicine. One way in which the supply chain of organs for transplantation can be improved is to discover novel approaches and technologies that preserve the health of organs outside of the body. The dominant technologies that are currently in use in the supply chain for biological materials maintain tissue temperatures ranging from a controlled room temperature (+25 °C to +15 °C) to cryogenic (−120 °C to −196 °C) temperatures (reviewed in Criswell et al. Stem Cells Transl Med. 2022). However, there are many cells and tissues, as well as all major organs, that respond less robustly to preservation attempts, particularly when there is a need for transport over long distances that require more time. In this perspective article, we will highlight the current challenges and advances in biopreservation aimed at “freezing biological time,” and discuss the future directions and requirements needed in the field.

All common negative-pressure wound therapy systems include a material, usually foam or gauze, at the wound/device interface. In this preclinical study, the authors have compared the effects on different wound healing parameters in the three most common negative-pressure wound therapy systems (i.e., V.A.C.VIA, PREVENA, and PICO) with a new device without foam or gauze (i.e., Platform Wound Dressing). A strong effort was made to avoid bias. The study was conducted under good laboratory practice conditions, with the presence of an independent observer. In pigs, three types of wounds were studied: full-thickness excisions, open incisions, and sutured closed incisions. Several macroscopic and microscopic parameters were studied. The pigs were euthanized on day 9 and all wounds were processed for histology and excisions for immunohistochemistry. In general, the devices produced similar results, with only a few significant differences. In the excisions, the Platform Wound Dressing reduced wound area more than the V.A.C.VIA and the PICO. In the excisional wounds, reepithelialization was the same. In open incisions, PREVENA was better than the Platform Wound Dressing. Histologic examination showed that, in open incisions, there was less inflammation in the PREVENA-treated in comparison with the Platform Wound Dressing- and the PICO-treated wounds. Immunohistochemical analyses showed that the Platform Wound Dressing-treated excisions had significantly more blood vessels (von Willebrand factor) than the V.A.C.VIA-treated ones and that the PICO caused less T-cell activation (CD3) than the other two. The devices-with foam, with gauze, or without either and just an embossed membrane-performed equally in general.

We describe our efforts to date using Dendroides- and, to a lesser extent, Cujucus-derived recombinant antifreeze proteins for short- and long-term mammalian cell, tissue, and organ storage strategies. Some single Dendroides antifreeze proteins (DAFPs), DAFP-6 and DAFP-2 concentrations in combination with dimethylsulfoxide increased post-cryopreservation cell viability. However, no significant improvement in viability was observed with either DAFP-1 or DAFP-4. Furthermore, more complex DAFP formulations resulted in loss of viability enhancement observed with DAFP-6 and 2. Experiments were also performed to see if the combination of DAFPs-1, -2, and -4 plus thaumatin-like protein had a benefit for hypothermic storage at −7 versus +4 °C. Thaumatin-like proteins are polypeptides of about 200 residues originally described in plants that are synthesized in response to fungal infection. Wang and Duman (Biochemistry 45:1278–1284, 2006) had previously described a thaumatin-like protein from larvae of the beetle Dendroides canadensis that enhances the activity of AFPs.

Objective: There is an unmet need to improve immediate burn care, particularly when definitive treatment is delayed. Therefore, the purpose of this project was to formulate a hydrogel that contains very high concentrations of antibiotics and validate its use together with a platform wound device (PWD) for the immediate care of burns. Approach: The hydrogel properties were optimized by using a rheometer, differential scanning calorimetry, and liquid chromatography-mass spectrometry and were tested in an infected porcine burn model. Immediately, after burn creation, the burns were infected with different bacteria. Subsequently, the burns infected with Staphylococcus aureus, Pseudomonas aeruginosa, and Acinetobacter baumannii were covered with the PWD and treated with a single dose of hydrogel containing 1000 × minimum inhibitory concentration of vancomycin, gentamicin, and minocycline, respectively. On day 7 or 45, the animals were euthanized, and the burns were harvested for histology and quantitative bacteriology. Results: 0.625% was the best alginate concentration for the hydrogel in terms of viscosity, stability, and drug release. The porcine studies demonstrated that vancomycin-, gentamicin-, and minocycline-treated tissues contained significantly less bacteria and reduced depth of tissue necrosis in comparison to controls. Innovation: The PWD represents a platform technology that begins at the point of the first treatment by protecting the wound and allowing administration of topical therapeutics. The device can be adapted to enclose any size burn over any contour of the body. Conclusion: Antibiotics can be delivered safely in very high concentrations in a hydrogel using the PWD, and burn infections can be treated successfully with this method.

Negative pressure wound therapy (NPWT) promotes healing in acute or chronic wounds. Conventional NPWT devices consist of a filler (such as foam or gauze) that covers the wound and of a permeable membrane and tubing that connects the space under the membrane to a suction pump. The permeable membrane increases airflow and thus increases the required pump capacity that can cause patient discomfort or even ischemia in wounds with compromised vascularity. In addition, foam or gauze may fragment and become colonized with bacteria over time. To mitigate these, negative aspects, we have developed a new impermeable single layer component membrane dressing to deliver NPWT that does not need a foam or gauze to function. Therefore, the purpose of this study was to introduce this novel NPWT system (platform wound device, PWD) and evaluate its usability and effectiveness in the treatment of porcine full-thickness burns. A total of 48 burn wounds were created across four Yorkshire pigs on the dorsum. Wounds were created on day 0 and continuous NPWT with -50 mmHg and - 80 mmHg was initiated immediately. Subsequently, the burns were debrided on day 3 and animals were euthanized on day 7. The efficacy of the PWD on wound healing and reduction of bacterial burden was measured and compared to wounds that did not receive NPWT. The results showed that PWD promoted wound healing by outperforming the wounds that did not receive NPWT and that PWD was efficient at reducing bacteria from the burn eschar and from the wound bed. In conclusion, this study demonstrated that PWD promoted wound healing with a negative pressure as low as -50 mmHg, which likely benefits healing and avoids potential safety issues.

In regard to evaluating tissue banking methods used to preserve or otherwise treat (process) soft allograft tissue, current tests may not be sufficiently sensitive to detect potential damage inflicted before, during, and after processing. Using controlled parameters, we aim to examine the sensitivity of specific biomechanical, electrical, and biological tests in detecting mild damage to collagen. Fresh porcine pulmonary heart valves were treated with an enzyme, collagenase, and incubated using various times. Controls received no incubation. All valves were cryopreserved and stored at -135°C until being rewarmed for evaluation using biomechanical, permeability, and cell viability tests. Statistically significant time dependent changes in leaflet ultimate stress, (p=0.006), permeability (p=0.01), and viability (p≤0.02, four different days of culture) were found between heart valves subjected to 0-15min of collagenase treatment (ANOVA). However, no statistical significance was found between the tensile modulus of treated and untreated valves (p=0.07). Furthermore, the trends of decreasing and increasing ultimate stress and viability, respectively, were somewhat inconsistent across treatment times. These results suggest that permeability tests may offer a sensitive, quantitative assay to complement traditional biomechanical and viability tests in evaluating processing methods used for soft tissue allografts, or when making changes to current validated methods. Multiple test evaluation may also offer insight into the mechanism of potential tissue damage such as, as is the case here, reduced collagen content and increased tissue porosity.