Cells for Treating Organ Damage

The past decade has witnessed great scientific enthusiasm for regenerative medicine using stem cell therapy. Stem cells, prompted to differentiate into healthy, mature cells, can be used to replace diseased or dead cells and thus hypothetically restore structure and function. On this background, scientists have been injecting stem cells into diseased areas of a body to allow them to repopulate with healthy cells. With mixed results, this has been attempted extensively to repair myocardium in the hopes of creating functioning cardiomyocytes.1 Contrary to initial belief, it is surprising that among transplanted cells in many models, only a few actually engraft and ultimately function within host tissue2,3; rather, the benefit of stem cells seems to be largely mediated by paracrine secretion of growth factors indirectly promoting resident (stem) cell proliferation to reconstitute damaged tissue, ultimately leading to organ repair.4 These latter experimental notions have spurred investigators to explore new ways to improve cell therapy by increasing targeted delivery of growth factors to a local environment.5 This approach may allow tissue regeneration while overcoming a major concern on the use of stem cells for regenerative medicine, namely their capacity to interfere with host immune surveillance, promoting the development of cancers, teratomas, or other proliferative lesions. These conditions are a major potential limitation of embryonic stem cells (ESCs) in renal medicine.6 Alternatively, a new class of molecules, namely self-assembling peptides, have been designed to allow local delivery of proteins.7 Self-assembly is a process mediated by noncovalent interactions between molecules. Self-assembling peptides are typically eight to 16 amino acids long and composed of alternating hydrophilic and hydrophobic residues. On exposure to physiologic osmolarity and pH, the peptides spontaneously assemble into interwoven nanofibers (10 to 20 nm) that further organize to form hydrogels. Those nanofibers are biocompatible because they do not include toxic cross-linkers to initiate solution-gel transformation, and their degradation produces natural amino acids that can be metabolized.8 That the formation of hydrogels occurs under physiologic conditions renders them suitable for delivery of bioactive molecules and/or cells that can be co-injected into a given tissue. Self-assembling peptides have been successfully used for regeneration of cardiac tissue. Highly controlled delivery of the endothelium-derived pro-survival factor PDGF-BB into the damaged heart by self-assembling nanofibers preserved cardiac systolic function after myocardial infarction and decreased infarct size after ischemia/reperfusion injury.9 Proteins can also be tethered to self-assembling peptides using avidin–biotin complexes. Injection of biotinylated IGF-1 complexed with tetravalent streptavidin and then bound to biotinylated self-assembling peptides led to prolonged delivery of IGF-1 to the heart, favoring the integration of neonatal myocytes co-implanted with the tethered peptide.10 Similarly, administration of cardiac progenitor cells, possessing the IGF-1/IGF-1 receptor system, together with the prolonged release of IGF-1 by self-assembling peptide nanofibers enhances the recovery of myocardial structure and function after infarction.11 Evidence that hyaluronic acid hydrogel-encapsulated endothelial progenitor cells accelerate the recovery of collateral circulation in mice with hindlimb ischemia, more than intravenous infusions of endothelial progenitor cells, has also recently opened the perspective of regeneration to other organs.12 No data are available so far on the effect of preconditioned nanofibers with ESCs on the recovery of organs after acute injury. It is also unknown whether biodegradable nanofibers can also act as acellular delivery platforms for the secretome released from stem cells. In this issue of JASN, the article by Wang et al.13 offers unprecedented findings shedding light on these issues. Through an integrated in vitro and in vivo approach, they assessed the effect of self-assembling nanofibers preconditioned with secretome from mouse ESCs on the kidney after LPS-induced acute kidney injury (AKI). While performing in vitro experiments, they initially compared two separate strategies for delivering secretome from ESCs. Using encapsulated nanofibers obtained after embedding ESCs or using preconditioned nanofibers after exposure to ESC secretome. Both preparations prevented the hyperpermeability of renal microvascular endothelial cell monolayers induced by LPS and also by vascular endothelial growth factor, which could be taken to indicate that preconditioning of nanofibers allows delivery of bioactive molecules to targeted cells in vitro. These preconditioned nanofibers in these experiments exerted cytoprotective effects by preventing LPS-induced apoptosis of tubular cells and podocytes through modulation of Rho kinase activation. One enticing feature of the study by Wang et al.13 is the in vivo confirmation of in vitro results. Preconditioned nanofibers prevented proteinuria, reduced renal function impairment, and exhibited antifibrotic and anti-inflammatory properties in mice with LPS-induced AKI. This seminal observation, however, leaves a number of open questions. The first is the characterization of the protein profiling of ESCs secretome, which has not been reported so far. Efforts to identify components of stem cell secretome by proteomics have concentrated on solving mehodologic issues of protein analysis and development of bioinformatic tools.14 Despite recent advances, many proteins still remain unidentified, which hampers the study of their specific function and interaction with other proteins. Wang et al.13 found 36 differentially expressed proteins in the solution containing preconditioned nanofibers and elected to assess the protective effect on the kidney of follistatin, adiponectin, and secretory leukoprotease inhibitor, which were selected on the basis of preexisting literature. Simultaneous secretion of the three proteins was necessary to prevent LPS-induced AKI. However, a more integrated approach is needed to establish definitively the determinant of renoprotection afforded by the ESC secretome. Another important issue, which is not addressed in the article, is the duration of protection of ESC preconditioned nanofibers on renal damage. The authors observed the effect of ESC secretome very early after LPS exposure but did not evaluate how long it lasted. The assessment of the time course of the effect may have disclosed important clues on the timing of nanofiber injection. The potential of stem cells and especially ESCs to repair the heart, kidney, or liver as recently documented in animals and possibly in humans has dramatically challenged our thinking on how to address the hurdle of acute and chronic organ damage in clinical setting. Despite unprecedented excitement over cell therapy, the use of cells in clinical medicine will by no means be the final solution. Questions such as cell purity, the number of cells to inject, the safety of the cell transplant procedure, whether the cells have to be predifferentiated before transplantation, how long they are retained in the damaged tissue, and how important their spatial distribution after transplantation is have not been addressed and will require time and major regulatory effort to resolve. Progress in direct reprogramming of somatic cells into pluripotent stem cells has been made15,16; however, a number of technical issues also need to be addressed before the technology can be transferred to the clinic. The most relevant problem still rests on achieving a complete resetting of the epigenetic state from a differentiated into a pluripotent embryonic condition to eliminate transient epigenetic memory that could hamper proper commitment to a lineage.17 Moreover, genomic instability, commonly found in human pluripotent cell lines, requires careful monitoring of each cell clone.18 Preconditioning nanofibers with ESC secretomes can theoretically obviate all of these limitations. The acellular platform reported here to deliver effectively paracrine or endocrine factors critically needed for tissue reconstitution and repair is a major technical step forward. It is theoretically easier to standardize with better commercial potential than stem cells and is devoid of the toxicity associated with injection of cells in humans. Will this be the end of stem cell therapy? Within the limits of predicting the future, our answer to this question today is certainly yes. DISCLOSURES None. This work was supported by a grant from the European Community's Seventh Framework Programme (FP7), STAR-TREK project (HEALTH-F5-2008-223007).