protocol-for-microrna-transfer-into-adult-bone-marrow-derived

Protocol for MicroRNA Transfer into Adult Bone Marrow-derived Hematopoietic Stem Cells to Enable Cell Engineering Combined with Magnetic Targeting

While CD133+ hematopoietic stem cells (SCs) have been proven to provide high potential in the field of regenerative medicine, their low retention rates ...

Rostock University Medical Center

Rostock University

Aim. CD133(+) stem cells bear huge potential for regenerative medicine. However, low retention in the injured tissue and massive cell death reduce beneficial effects. In order to address these issues, we intended to develop a nonviral system for appropriate cell engineering. Materials and Methods. Modification of human CD133(+) stem cells with magnetic polyplexes carrying microRNA was studied in terms of efficiency, safety, and targeting potential. Results. High microRNA uptake rates (~80-90%) were achieved without affecting CD133(+) stem cell properties. Modified cells can be magnetically guided. Conclusion. We developed a safe and efficient protocol for CD133(+) stem cell modification. Our work may become a basis to improve stem cell therapeutical effects as well as their monitoring with magnetic resonance imaging.

Magnet-Bead Based MicroRNA Delivery System to Modify CD133(+) Stem Cells.

CD133(+) stem cells represent a promising subpopulation for innovative cell-based therapies in cardiovascular regeneration. Several clinical trials have shown remarkable beneficial effects following their intramyocardial transplantation. Yet, the purification of CD133(+) stem cells is typically performed in centralized clean room facilities using semi-automatic manufacturing processes based on magnetic cell sorting (MACS®). However, this requires time-consuming and cost-intensive logistics.

GMP-conformant on-site manufacturing of a CD133(+) stem cell product for cardiovascular regeneration.

Magnetic activated cell sorting (MACS(®)) is routinely used to isolate stem cell subpopulations intended for the treatment of cardiovascular diseases. In strong contrast, studies examining the amount, effect and intramyocardial distribution of iron nanoparticles used for magnetic cell labelling are missing, although iron excess can cause functional disorders in the heart.

Intramyocardial fate and effect of iron nanoparticles co-injected with MACS(®) purified stem cell products.

Adult cardiomyocytes (CMs) possess a highly restricted intrinsic regenerative potential - a major barrier to the effective treatment of a range of chronic degenerative cardiac disorders characterized by cellular loss and/or irreversible dysfunction and which underlies the majority of deaths in developed countries. Both stem cell programming and direct cell reprogramming hold promise as novel, potentially curative approaches to address this therapeutic challenge. The advent of induced pluripotent stem cells (iPSCs) has introduced a second pluripotent stem cell source besides embryonic stem cells (ESCs), enabling even autologous cardiomyocyte production. In addition, the recent achievement of directly reprogramming somatic cells into cardiomyocytes is likely to become of great importance. In either case, different clinical scenarios will require the generation of highly pure, specific cardiac cellular-subtypes. In this review, we discuss these themes as related to the cardiovascular stem cell and programming field, including a focus on the emergent topic of pacemaker cell generation for the development of biological pacemakers and in vitro drug testing.

(Re-)programming of subtype specific cardiomyocytes.

Many disorders are manifested by dysfunction of key cell types or their disturbed integration in complex organs. Thereby, adult organ systems often bear restricted self-renewal potential and are incapable of achieving functional regeneration. This underlies the need for novel strategies in the field of cell (re-)programming-based regenerative medicine as well as for drug development in vitro. The regenerative field has been hampered by restricted availability of adult stem cells and the potentially hazardous features of pluripotent embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). Moreover, ethical concerns and legal restrictions regarding the generation and use of ESCs still exist. The establishment of direct reprogramming protocols for various therapeutically valuable somatic cell types has overcome some of these limitations. Meanwhile, new perspectives for safe and efficient generation of different specified somatic cell types have emerged from numerous approaches relying on exogenous expression of lineage-specific transcription factors, coding and noncoding RNAs, and chemical compounds.It should be of highest priority to develop protocols for the production of mature and physiologically functional cells with properties ideally matching those of their endogenous counterparts. Their availability can bring together basic research, drug screening, safety testing, and ultimately clinical trials. Here, we highlight the remarkable successes in cellular (re-)programming, which have greatly advanced the field of regenerative medicine in recent years. In particular, we review recent progress on the generation of cardiomyocyte subtypes, with a focus on cardiac pacemaker cells. Graphical Abstract.

Frauke Hausburg

Reference and Translation Center for Cardiac Stem Cell Therapy (RTC),
Department of Cardiac Surgery,
Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), Department of Cardiac Surgery

Rostock University Medical Center

Protocol for MicroRNA Transfer into Adult Bone Marrow-derived Hematopoietic Stem Cells to Enable Cell Engineering Combined with Magnetic Targeting

Frauke Hausburg

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Reference and Translation Center for Cardiac Stem Cell Therapy (RTC),Department of Cardiac Surgery,Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), Department of Cardiac Surgery

Rostock University Medical Center

Protocol for MicroRNA Transfer into Adult Bone Marrow-derived Hematopoietic Stem Cells to Enable Cell Engineering Combined with Magnetic Targeting

Reference and Translation Center for Cardiac Stem Cell Therapy (RTC),
Department of Cardiac Surgery,
Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), Department of Cardiac Surgery