Paracrine cell signaling, not differentiation, appears to be how stem cells work in initial phase of organ protection, repair
BETHESDA, Md. (August 16, 2005) – Acute renal failure, or ARF, is as serious as it sounds. An estimated 40% of critical care hospital admissions experience ARF. Estimates of their death rate range from 50% to 80%, complicated by the fact that patients with ARF often simultaneously suffer failure of other major organ systems.
The most serious form of ARF is caused by ischemia, or loss of blood supply to the kidneys caused by shock, blood infection or major cardiovascular surgery, particularly in such high- risk patients as those with diabetes, underlying renal disease, and the elderly. In the kidneys, such "insults" lead to destruction of kidney tubular and vascular cells, initiating a significant inflammatory response. Recovery of kidney function that is adequate for patient survival depends primarily on the protection and regeneration of destroyed and injured cells.
Yet virtually no progress has been made toward development of any highly effective ARF therapy for decades. "Basically, catastrophic loss of kidney function has remained treatment-resistant despite dialysis and intensive care. Treating patients with ARF thus presents a major clinical dilemma, particularly when severe ARF occurs with multiple organ failure," Christof Westenfelder from the University of Utah explained. "Our laboratory has therefore been pursuing development of novel therapeutic interventions that are urgently needed to treat this common, devastating and costly human disease," Westenfelder said.
Challenging the accepted theory of stem cell operation
In a new study, Westenfelder's team reported that injecting stem cells similar to the type used in bone marrow transplants is "highly renoprotective, showing almost immediate improvement in both kidney function and degree of tissue injury, followed by accelerated regeneration and return of function," he said. Furthermore, "these beneficial effects are predominantly mediated, as our data suggest, by paracrine rather than transdifferentiation-dependent mechanisms," the paper reported. Paracrine indicates action instigated by nearby cells.
These new results "challenge the most popular hypothesis of how stem cells work in kidney protection and repair, which holds that administered stem cells enter an injured organ where they differentiate into those cells that have been destroyed, and thus replace them both anatomically and functionally," Westenfelder said.
MSC paracrine effects prompt a positive cascade, halt inflammatory response
Rather, the Utah team found that "administered stem cells don't stay in the kidney that has ARF long enough to differentiate into kidney cells, but rather appear to alter the course of ARF by a number of identifiable and some still unexplored paracrine mechanisms. The former include the induction of organ-protective and repair-supporting genes in surviving renal cells, robust suppression of proinflammatory cytokines in the ARF kidney and upregulation of anti-inflammatory genes, as well as the delivery and release at the site of injury of organ-protective and other beneficial gene products by the stem cells per se. Collectively, these and as yet unidentified mechanisms represent a highly potent intervention in ARF," Westenfelder stated.
The study, "Administered mesenchymal stem cells protect against ischemic acute renal failure through differentiation-independent mechanisms," appears in the American Journal of Physiology-Renal Physiology, published by the American Physiological Society. Research was conducted by Florian Tögel, Zhuma Hu, Kathleen Weiss, and Christof Westenfelder of the University of Utah and the Veterans Affairs Medical Center; Jorge Isaac, University of Utah; and Claudia Lange, Bone Marrow Transplantation Center, Hamburg.
Stem cell time in the kidney: about two hours
At first, Westenfelder conceded, the researchers experienced "substantial frustration because the cells were detectable in the kidney for only two hours and then 'disappeared.' But after entry into the kidneys with ARF, we found that they fundamentally changed a number of important gene expression profiles. In fact, the intrarenal location of administered stem cells is such that it facilitates the delivery of renoprotective growth factors and cytokines to the sites where kidney cells are primarily destroyed from both the blood and urinary aspects of the cells that must be protected and repaired!" Westenfelder reported.
With their rapid mode of action within only two hours in the kidney, the researchers deduced that the mechanism mediating the protective effects of MSC (mesenchymal stem cells) "must be primarily paracrine, as implied by their demonstrated expression of several growth factors such as HGF (hepatocyte growth factor), VEGF and IGF-1, all known to improve renal function in ARF, mediated by their antiapoptotic, mitogenic and other cytokine actions," the paper said.
"Specifically, these as yet incompletely defined paracrine actions of MSC result in the renal downregulation of proinflammatory cytokines IL-1-beta, TNF-alpha, and IFN-gamma, as well as iNOS (inducible nitric oxide synthase), and upregulation of anti-inflammatory and organ-protective interleukin-10, as well as bFGF (basic fibroblast growth factor), TGF-alpha (transforming growth factor alpha) and antiapoptotic Bc1-2," the paper added.
Beneficial paracrine actions are elicited early and late after onset of ARF
The study said it "provides the first clear evidence that therapy with MSC affords significant renoprotection in rats with ischemic/reperfusion (I/R) ARF. Animals infused with MSC either immediately or 24 hours after reperfusion had significantly better renal function, lower renal injury and cell-death scores, and higher cell division indices than vehicle-treated control animals," the paper stated. Indeed, administration after 24 hours of more severe ARF had an even greater beneficial effect.
Based on utilization of several genomic and nongenomic cell-tagging techniques, the researchers were able to follow the stem cells as they circulated through the renal microcirculation of two different strains of rats used in the experiments. "Although we did not detect transdifferentiation events during the 72-hour period of observation, it is possible that cell transdifferentiation and integration may be important at later states of organ repair," the study noted.
The Utah team used MSC for several reasons, notably the fact that they can be harvested easily from bone marrow, isolated, grown in culture and genetically engineered. To test that MSC subsequently used in these studies retained their phenotype after being cultured, the researchers routinely tested whether their characteristic potential to differentiate into fat and bone cells was preserved.
The researchers conceded that it "is surprising that the very transient presence of MSC in the injured kidney, as we document, is sufficient to greatly ameliorate the course" of I/R ARF and also note that the rat model of ARF used in the study is "a suitable if somewhat imperfect model of the most common and the most treatment-resistant type of human ischemic ARF."
Next steps
- Current studies are investigating the collective and individual renoprotective capacity of cytokines temporarily released by MSC.
- Two clinical efforts are planned. First will be a compassionate study in Europe next year in patients with complicated bone marrow transplants, a setting where such stem cells already are approved for use. If successful, a compassionate-use study in patients with severe ARF and multiorgan failure will be designed, eventually leading to clinical trials.
- Though no adverse effects were seen in these experiments, long-term studies are needed to prove that no adverse effects occur after in vivo administration of adult MSC.
- Also to be determined is whether renoprotective and gene-modulating effects of MSC "are primary actions that are humorally elicited by these cells or whether they result from the improvement of tissue injury by as yet unknown factors released by them."
- Future studies "will also have to define the possible contribution to organ protection made by the immunomodulatory effects of MSC, as well as potential late phase contributions to organ repair via differentiation of stem cells into cells that have been destroyed in ARF," the study noted.
Source and funding
The study, entitled "Administered mesenchymal stem cells protect against ischemic acute renal failure through differentiation-independent mechanisms," appears in the American Journal of Physiology-Renal Physiology, published by the American Physiological Society. The research was conducted by Florian Tögel, Zhuma Hu, Kathleen Weiss, and Christof Westenfelder of the Division of Nephrology, Dept. of Medicine, University of Utah, Salt Lake City, and the Veterans Affairs Medical Center; Tögel and Westenfelder are also at the Dept. of Physiology; Jorge Isaac of the Dept. of Pathology; and Claudia Lange of the Bone Marrow Transplantation Center, Hamburg, Germany.
Research was supported in part by the U.S. Dept. of Veterans Affairs, American Heart Association, Dialysis Research Foundation, National Kidney Foundation, Western Institute for Biomedical Research and NHLBI (NIH).
Editor's note: The media may obtain an electronic version of Tögel et al. and interview members of the research team by contacting Mayer Resnick at the American Physiological Society, 301.634.7209, cell 301.332.4402 or mresnick@the-aps.org.
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