Papers by Kelvin Brockbank
Journal of Cardiac Surgery, Mar 1, 1987
For any tissue, there is a cell viability threshold below which the ability of the tissue to main... more For any tissue, there is a cell viability threshold below which the ability of the tissue to maintain itself and function will eventually be compromised. During cryopreservation and subsequent thawing of tissues there are many steps involved, each with attendant potential risks for reduction of viability. To determine the effectiveness of any cryopreservation procedure it is important to select appropriate assays. In this manuscript viability assays, in general, are reviewed from a biological viewpoint prior to review of methods employed for assessment of heart valve viability. Both in situ and in vitro assays of heart valve viability indicate that valve mechanical properties and the majority of fibroblasts, which are responsible for maintenance of the valve connective tissue, are retained after cryopreservation.
Transfusion Medicine and Hemotherapy, 2011
Using an electrical conductivity method, the effect of matrix composition on solute permeability ... more Using an electrical conductivity method, the effect of matrix composition on solute permeability has been studied in hydrogels and cartilaginous tissues [24, 25]. In this study, we adopted this method to study the impact of 4 °C storage on cartilage ECM solute permeability. Moreover, the transport of small solutes (e.g. ions, oxygen, and glucose)
BioProcessing Journal, Dec 21, 2007
Cells, May 16, 2022
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
The purpose of this study was to determine the effects of varying heart cold ischemia times on po... more The purpose of this study was to determine the effects of varying heart cold ischemia times on post-cryopreservation valve leaflet cellular functions. Cellular viability and function were assessed through measurement of protein synthesis, ribonucleic acid synthesis, and glucose phosphorylation. Cold ischemia times varied from 24 to 113 h before processing and cryopreservation. The results show that valve leaflet cellular functions could be detected for at least 42 h of post-procurement cold ischemia. This information can be used as a guide in further defining the acceptable limits for heart valve donation and processing.
Leukemia Research, 1986
Fibroblast colony-forming units (CFU-F) in bone marrow and spleen were investigated after infecti... more Fibroblast colony-forming units (CFU-F) in bone marrow and spleen were investigated after infection of BALB/c mice with the Rauscher leukemia virus complex (RLV) or with cellularly cloned helper virus (R-MuLV). Both viral preparations induced a transient suppression of CFU-F prior to the development of leukemia. A second CFU-F decrease was observed in the terminal phase of R-MuLV-induced chronic lymphoid or myeloid leukemias, but not in mice with a RLV-induced acute erythroleukemia, which are highly viremic. When a number of Rauscher virus transformed leukemic cell lines were injected intravenously an erythroid and a T-cell line suppressed CFU-F values, but a third non-virus producing B-cell line did not. In contrast to the in-vitro situation, in-vitro inhibition of CFU-F was only observed in conditoned medium of an erythroid cell line with undetectable reverse transcriptase activity, whereas conditioned medium of lymphoid and myeloid cell lines were not inhibitory irrespective of reverse transcriptase activity. Together these results indicate that murine leukemia viruses can suppress the numbers of detectable CFU-F in vivo in an early phase of the disease and that leukemic cells may inhibit CFU-F proliferation by a mechanism which is not related to viremic state of the animal or the production of virus by these cells.
Scientific Reports, Dec 5, 2017
Conventional frozen cryopreservation (CFC) is currently the gold standard for cardiovascular allo... more Conventional frozen cryopreservation (CFC) is currently the gold standard for cardiovascular allograft preservation. However, inflammation and structural deterioration limit transplant durability. Ice-free cryopreservation (IFC) already demonstrated matrix structure preservation combined with attenuated immune responses. In this study, we aim to explore the mechanisms of this diminished immunogenicity in vitro. First, we characterized factors released by human aortic tissue after CFC and IFC. Secondly, we analyzed co-cultures with human peripheral blood mononuclear cells, purified monocytes, T cells and monocyte-derived macrophages to examine functional immune effects triggered by the tissue or released cues. IFC tissue exhibited significantly lower metabolic activity and release of proinflammatory cytokines than CFC tissue, but surprisingly, more active transforming growth factor β. Due to reduced cytokine release by IFC tissue, less monocyte and T cell migration was detected in a chemotaxis system. Moreover, only cues from CFC tissue but not from IFC tissue amplified αCD3 triggered T cell proliferation. In a specifically designed macrophage-tissue assay, we could show that macrophages did not upregulate M1 polarization markers (CD80 or HLA-DR) on either tissue type. In conclusion, IFC selectively modulates tissue characteristics and thereby attenuates immune cell attraction and activation. Therefore, IFC treatment creates improved opportunities for cardiovascular graft preservation. Providing matrices for tissue replacements that fulfill all requirements for successful and appropriate treatment of cardiovascular diseases is still an important issue. The ability to replace tissues on demand would help to decrease the mortality rate in many clinical scenarios. Off-the-shelf available vascular grafts and heart valves for example would be needed for bypass surgery 1,2 , cardiac valvular pathologies including rheumatic fever, atherosclerosis in elderly patients or congenital malformation in children 3-6 , shunts for dialysis patients 7,8 and vessel reconstruction after organ transplantation 9,10. Therefore, improved strategies and techniques for tissue preservation have to be developed to tackle this urgent and so far unmet medical need 11. Biological vascular and heart valve matrices appear to be beneficial compared to synthetic or mechanical substitutes with respect to hemodynamic features, biocompatibility, anti-coagulation treatment, and their potential to grow and remodel especially in young patients 4,6,12. However, the availability of human biological matrices for an off-the-shelf allogeneic application is limited. Therefore, xenogeneic (porcine and bovine) tissue is often used for heart valve replacement. To avoid unwanted xeno-immune responses, the tissue is fixed with glutaraldehyde (GA). However, after fixing, the tissue is not viable and lacks remodeling capacity 13,14. Another treatment option to reduce immune responses to xenogeneic tissue following implantation is the introduction of a decellularization
The Annals of Thoracic Surgery, Jun 1, 2011
Background. Cryopreserved allogeneic heart valves are usually stored and transported below ؊135°C... more Background. Cryopreserved allogeneic heart valves are usually stored and transported below ؊135°C; however, such methods require expensive equipment for both storage and transportation. Methods. In this study, vitrified porcine aortic valves were stored on either side of the cryoprotectant formulation's glass transition temperature (؊119°C) at ؊80°C and ؊135°C, using a newly formulated vitrification solution (VS83) consisting of a combination of 4.65M dimethyl sulfoxide, 4.65M formamide, and 3.30M 1,2-propanediol. Three groups of valves were studied: (1) fresh; (2) VS83preserved, stored at ؊80°C; and (3) VS83-preserved, stored at ؊135°C. Results. Using the VS83 cryoprotectant concentration formulation, cracking was not observed during valve storage. No ice-related events were detectable during 5°C rewarming by differential scanning calorimetry. All cryopreserved tissue samples demonstrated significantly less viability than fresh samples (p < 0.01). No significant viability differences were observed between the VS83preserved groups stored at ؊80°C and ؊135°C. Material testing did not reveal any significant differences among the three test groups. Multiphoton imaging of VS83preserved heart valves stored at ؊80°C and ؊135°C demonstrated similar collagen and elastin structures. Conclusions. These results indicate that VS83-preserved heart valves can be stored and transported at temperatures in the vicinity of ؊80°C with retention of extracellular matrix integrity and material properties. The VS83 preservation of heart valves at ؊80°C without the need for liquid nitrogen should result in both decreased manufacturing costs and reduced employee safety hazards. Moreover, it is anticipated that low cell viability may result in less immunogenicity in vivo.
Journal of Cardiovascular Development and Disease, May 26, 2023
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
The FASEB Journal, Apr 1, 2012
Structural heart, Feb 13, 2020
Cryobiology, Dec 1, 2010
Cryopreserved vessel allografts have been used preferably for infectious diseases in the surgical... more Cryopreserved vessel allografts have been used preferably for infectious diseases in the surgical field. But even in non-infectious area, cryopreserved vessels can be applied. In our Tissue Bank, cryopreserved vein allografts have been used for portal system reconstruction in the living donor liver transplantations. The purpose of this study was to examine the feasibility of application of cryopreserved vein allografts in the living donor liver transplantation. For living donor liver transplantation there were three groups (strategies) of extraction of the liver graft from the donor body: (A) right liver graft, where donor middle hepatic vein (MHV) was resected and attached to the extracted liver graft to use for portal reconstruction in the recipient surgery, (B) right liver graft without middle hepatic vein where the middle hepatic vein was preserved in the donor body and cryopreserved vein allograft was used instead for the portal reconstruction in the recipient surgery, and (C) left liver graft where donor middle hepatic vein could not be resected for anatomical reasons and cryopreserved vein allografts were used in the recipient surgery for the portal vein reconstruction. A total of 101 living donor liver transplantations were examined. The procedures were divided into the above three groups, and the complication incidence was compared between the three groups. Also, in group B, vein allograft patency was examined by ultra sound echography up to 6 months. All recipients received the same immune suppressants, and no anticoagulants were received. Complication incidence of 3 groups (A, B, C) was 11 (73%), 23 (43%), and 15 (47%), respectively. Not all the complication cases needed treatment. In group A, 3 cases (27%) did not need any treatment, in group B 13 (57%), and in group C 8 (53%) did not need treatment. Concerning the vein allograft patency in group B, the patency rate in 2, 4, and 6 months was 96%, 80%, and 54%, respectively. In general, the living donor must not have any complications for any reason. In this study, group A (no cryopreserved vein allograft) treatment was needed for complications in 50%. On the other hand, in group B or C (cryopreserved vein allograft was used in the recipient surgery), complications needing treatment occurred less than 20%. Originally, cryopreserved vein allografts were used to improve the outcome of the recipients. Our data showed that the application of cryopreserved vein allografts improved the quality of donor surgery outcome. The application of cryopreserved vein allografts improved not only the outcome of recipients but also that of donors, which is fundamental for living donor organ transplantations. Conflict of interest: None declared. Source of funding: None declared.
The destructive effect of extracellular ice formation in organized multicellular tissues is known... more The destructive effect of extracellular ice formation in organized multicellular tissues is known to be a principal reason why conventional techniques of cryopreservation fail to provide effective protection during freezing. Avoidance of ice during cooling can physically be achieved by vitrification, which is the amorphous solidification of a supercooled liquid by adjusting the solute composition and cooling rate such that nucleation and growth of ice crystals is prevented. In principal a biological system can be stabilized in the glassy (vitreous) state without the inherent problems associated with crystallization and so-called solution effects injury that arise from the increased concentration of solutes due to removal of water as ice. This study was designed to evaluate a vitrification approach to storing a vascular tissue model (rabbit jugular vein) compared with a standard commercial method used clinically and employing slow cooling (l°C/min) with dimethylsulfoxide (DMSO) as the cryoprotective agent (CPA). A baseline vitrification medium (designated VS55) was used to replace at least 50% of the tissue water with a combination of CPAs. and vitrification was achieved using a relatively high cooling rate (&gt; 40°C/min). After rewarming and removal of the CPAs. contractility of the blood vessel segments was evaluated in vitro using standard physiological response tests to a panel of reagents. The maximum contractions achieved in vitrified vessels were all greater than 80% of fresh matched controls with similar drug sensitivities. In contrast, frozen/thawed vessels exhibited maximal contractions below 25% of fresh matched controls with concomitant decreases in drug sensitivity. This study clearly demonstrates a significant benefit of vitrification compared with conventional freezing and thawing for preserving smooth muscle contractile function of preserved blood vessels. Vitrification may prove to be an effective way to minimize the deleterious effects of freezing and thawing in cryopreserved tissues, or engineered tissue constructs.
Springer eBooks, 1989
Shortfalls in organ donation result in prolonged waiting periods for transplant recipients. Such ... more Shortfalls in organ donation result in prolonged waiting periods for transplant recipients. Such shortfalls could be significantly reduced through the efforts of health professionals who attend to, and are involved with, brain dead patients. The purpose of this chapter is to present some current and future developments in the field of tissue transplantation, with the aim of clarifying the importance and potential of transplantable tissues. Since large organs such as heart, liver, and kidneys are the subjects of other chapters in this volume, this chapter is restricted to small, less complex tissues, such as heart valves, veins, and islets of Langerhans. The importance of these tissues is clarified when it is realized that approximately 400,000 transplantation procedures have been performed in 1987 (Table I).
PubMed, 1980
We have discovered that human and rabbit bone marrow conditioned media (BMCM) promote the growth ... more We have discovered that human and rabbit bone marrow conditioned media (BMCM) promote the growth of bursts in cultures containing erythropoietin. We then demonstrated that the measurement of 59Fe incorporation into heme was a more quantitative assay for burst-promoting activity (BPA) than counting burst numbers. Furthermore, we have observed that the use of low (less than 15%) concentrations of fetal calf serum is a convenient way of reducing endogenous sources of BPA for studies of the BPA in the test samples. When 59Fe incorporation and cell numbers of individual rabbit erythropoietic bursts were analyzed simultaneously, BPA caused a shift in the cumulative frequency distributions without changes in their shapes. These results indicated that BPA augments hemoglobin synthesis by stimulating cell proliferation during the early phase of burst formation. In addition, human BPA increased the relative proportion of fetal hemoglobin in the hemoglobins synthesized by adult circulating BFU-e. BPA appears to augment cell proliferation of early precursors which are committed to produce F cells.
Annals of the New York Academy of Sciences, Jun 1, 2002
PubMed, Aug 1, 1987
Bone marrow fibroblasts have been shown to have a role in the support and regulation of hemopoies... more Bone marrow fibroblasts have been shown to have a role in the support and regulation of hemopoiesis, both in vivo and in vitro. In this study we examine the ability of skin-derived fibroblasts to interact with hemopoiesis in vitro. Murine skin and bone marrow-derived fibroblasts were similar with respect to their abilities to support granulopoiesis and release colony-stimulating activity. Detailed analysis of skin fibroblast cultures 1 week after seeding with stromal cell-depleted bone marrow demonstrated that both multipotential hemopoietic stem cells and erythroid stem cells were maintained, while granulocyte/macrophage colony-forming units far exceeded inoculum values. Immunostaining demonstrated the presence of foci of T200 positive hemopoietic cells on the surface of the fibroblasts with less frequent scattered M1/70 and F4/80 positive macrophages. The majority of cells (greater than 90%) released from the stromal layer were of the granulocytic series. These findings demonstrate that the hemopoietic regulatory properties previously attributed to bone marrow-derived fibroblasts are not unique to fibroblasts derived from hemopoietic tissues.
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Papers by Kelvin Brockbank