Mesenchymal Stem/Stromal Cells in Organ Transplantation.

Organ transplantation is essential and crucial for saving and enhancing the lives of individuals suffering from end-stage organ failure. Major challenges in the medical field include the shortage of organ donors, high rates of organ rejection, and long wait times. To address the current limitations and shortcomings, cellular therapy approaches have been developed using mesenchymal stem/stromal cells (MSC). MSC have been isolated from various sources, have the ability to differentiate to important cell lineages, have anti-inflammatory and immunomodulatory properties, allow immunosuppressive drug minimization, and induce immune tolerance towards the transplanted organ. Additionally, rapid advances in the fields of tissue engineering and regenerative medicine have emerged that focus on either generating new organs and organ sources or maximizing the availability of existing organs. This review gives an overview of the various properties of MSC that have enabled its use as a cellular therapy for organ preservation and transplant. We also highlight emerging fields of tissue engineering and regenerative medicine along with their multiple sub-disciplines, underlining recent advances, widespread clinical applications, and potential impact on the future of tissue and organ transplantation.

A new flow cytometry assay identifies recipient IgG subtype antibodies binding donor cells: increasing donor availability for highly sensitised patients.

Objectives: There are four immunoglobulin (IgG) subtypes that have varying complement-activating ability: strong (IgG3 and IgG1) and weak (IgG2 and IgG4). The standard flow cytometric crossmatch (FCM) assay does not distinguish between the various subtypes of the IgG molecule. This study outlines the development and use of a novel cell-based IgG subtype-specific FCM assay that is able to detect the presence of and quantitate the IgG subtypes bound to donor cells. Methods: A six-colour lyophilised reagent was designed that specifically detects the four IgG subtypes, as well as distinguishes between T cells and B cells in the lymphocyte population. To test the efficacy of this reagent, a retrospective evaluation of a group of highly sensitised patients awaiting heart and kidney transplant was carried out, who, because of positive standard FCM results, had been deemed incompatible with numerous prior potential donors. Results: Observations in this study demonstrate that the positive standard FCM results were mainly because of the presence of noncomplement-activating IgG2 or IgG4 antibodies. The results were supported by the absence of C3d-binding donor-specific antibodies (DSA) and a negative complement-dependent cytotoxicity crossmatch (CDC). Conclusion: Preliminary data presented in this study demonstrate the reliability of the novel IgG subtype assay to detect the presence of pretransplant, complement-activating antibodies bound to donor cells. The knowledge gained from the IgG subtype assay and the C3d-binding specificities of DSAs provides improved identification of donor suitability in pretransplant patients, potentially increasing the number of transplants.

Differentiation of Human Deceased Donor, Adipose-Derived, Mesenchymal Stem Cells into Functional Beta Cells.

There is an emerging need for the rapid generation of functional beta cells that can be used in cell replacement therapy for the treatment of type 1 diabetes (T1D). Differentiation of stem cells into insulin-producing cells provides a promising strategy to restore pancreatic endocrine function. Stem cells can be isolated from various human tissues including adipose tissue (AT). Our study outlines a novel, non-enzymatic process to harvest mesenchymal stem cells (MSC) from research-consented, deceased donor AT. Following their expansion, MSC were characterised morphologically and phenotypically by flow cytometry to establish their use for downstream differentiation studies. MSC were induced to differentiate into insulin-producing beta cells using a step-wise differentiation medium. The differentiation was evaluated by analysing the morphology, dithizone staining, immunocytochemistry, and expression of pancreatic beta cell marker genes. We stimulated the beta cells with different concentrations of glucose and observed a dose-dependent increase in gene expression. In addition, an increase in insulin and c-Peptide secretion as a function of glucose challenge confirmed the functionality of the differentiated beta cells. The differentiation of adipose-derived MSC into beta cells has been well established. However, our data demonstrates, for the first time, that the ready availability and properties of MSC isolated from deceased donor adipose tissue render them well-suited as a source for increased production of functional beta cells. Consequently, these cells can be a promising therapeutic approach for cell replacement therapy to treat patients with T1D.

Structural and Functional Characterization of Deceased Donor Stem Cells: A Viable Alternative to Living Donor Stem Cells.

Stem cells can be isolated from various human tissues including bone marrow (BM) and adipose tissue (AT). Our study outlines a process to isolate adult stem cells from deceased donors. We have shown that cell counts obtained from deceased donor BM were within established living donor parameters. Evaluation of demographic information exhibited a higher percentage of hematopoietic stem cells (HSC) in males versus females, as well as a higher percentage of HSC in the age bracket of 25 years and under. For the first time, we show that deceased donor femur BM grew cell colonies. Our introduction of new technology for nonenzymatic AT processing significantly increased cell recovery over the traditional enzymatic processing method. Cell counts from the deceased donor AT exceeded living donor parameters. Furthermore, our data illustrated that AT from female donors yielded a much higher number of total nucleated cells (TNC) than males. Together, our data demonstrates that our approach to isolate stem cells from deceased donors could be a routine practice to provide a viable alternative to living donor stem cells. This will offer increased accessibility for patients awaiting stem cell therapies.