Christof Westenfelder

VEGF is a mediator of the renoprotective effects of multipotent marrow stromal cells in acute kidney injury

Adult stem cell treatment of complex disorders is a promising therapeutic approach and multipotent marrow stromal cells (MSCs) have been shown to be effective in various animal models of diseases. Acute kidney injury (AKI) is a common and serious problem in hospitalized patients and bone marrow derived multipotent MSCs have been shown to be effective in different models of AKI. The mechanism of action of MSCs is complex but involves paracrine actions including growth factor secretion. Knockdown of vascular enthothelial growth factor (VEGF) by siRNA reduced effectiveness of MSCs in the treatment of ischemic AKI in a rat model. Animals treated with MSCs had increased renal microvessel density compared to VEGF knockdown MSC‐treated and vehicle‐treated animals. These results show that VEGF is an important mediator of the early and late phase of renoprotective action after AKI in the context of stem cell treatment.

Mesenchymal stem cells: a new therapeutic tool for AKI

Acute kidney injury (AKI) is a common clinical complication, associated with poor outcomes and the development of chronic kidney disease. Despite major advances in the understanding of its pathophysiology, available therapies for AKI are only supportive; therefore, adequate functional recovery from AKI must predominantly rely on the kidneys own reparative ability. An extensive body of preclinical data from our own and from other laboratories has shown that administration of adult multipotent marrow stromal cells (commonly referred to as mesenchymal stem cells [MSCs]), effectively ameliorates experimental AKI by exerting paracrine renoprotective effects and by stimulating tissue repair. Based on these findings, a clinical trial has been conducted to investigate the safety and efficacy of MSCs administered to open-heart surgery patients who are at high risk of postoperative AKI. In this Perspectives article, we discuss some of the early data from this trial and describe potential applications for stem cell therapies in other fields of nephrology.

Adult bone marrow–derived stem cells for organ regeneration and repair

Stem cells have been recognized as a potential tool for the development of innovative therapeutic strategies. There are in general two types of stem cells, embryonic and adult stem cells. While embryonic stem cell therapy has been riddled with problems of allogeneic rejection and ethical concerns, adult stem cells have long been used in the treatment of hematological malignancies. With the recognition of additional, potentially therapeutic characteristics, bone marrow–derived stem cells have become a tool in regenerative medicine. The bone marrow is an ideal source of stem cells because it is easily accessible and harbors two types of stem cells. Hematopoietic stem cells give rise to all blood cell types and have been shown to exhibit plasticity, while multipotent marrow stromal cells are the source of osteocytes, chondrocytes, and fat cells and have been shown to support and generate a large number of different cell types. This review describes the general characteristics of these stem cell populations and their current and potential future applications in regenerative medicine. Developmental Dynamics 236:3321–3331, 2007.

Bioluminescence imaging to monitor the in vivo distribution of administered mesenchymal stem cells in acute kidney injury.

Effective and targeted delivery of cells to injured organs is critical to the development of cell therapies. However, currently available in vivo cell tracking methods still lack sufficient sensitivity and specificity. We examined, therefore, whether a highly sensitive and specific bioluminescence method is suitable to noninvasively image the organ distribution of administered mesenchymal stem cells (MSCs) in vivo. MSCs were transfected with a luciferase/neomycin phosphotransferase construct (luc/neo-MSC). Bioluminescence of these cells was measured (charge-coupled device camera) after treatment with luciferin, showing a linear increase of photon emission with rising cell numbers. To track these cells in vivo, groups of mice were injected with 1 x 10(5) luc/neo-MSCs/animal and imaged with bioluminescence imaging at various time points. Injection of cells in the suprarenal aorta showed diffuse distribution of cells in normal animals, whereas distinct localization to the kidneys was observed in mice with ischemia- and reperfusion-induced acute kidney injury (AKI). Intrajugular infusion of MSCs demonstrated predominant accumulation of cells in both lungs. In animals with AKI, detectable cell numbers declined over time, as assessed by bioluminescence imaging and confirmed by PCR, a process that was associated with low apoptosis levels of intrarenally located MSCs. In conclusion, the described bioluminescence technology provides a sensitive and safe tool for the repeated in vivo tracking of infused luc/neo-MSCs in all major organs. This method will be of substantial utility in the preclinical testing and design of cell therapeutic strategies in kidney and other diseases.

In vivo magnetic resonance imaging of iron oxide-labeled, arterially-injected mesenchymal stem cells in kidneys of rats with acute ischemic kidney injury: detection and monitoring at 3T.

To evaluate MRI for a qualitative and quantitative in vivo tracking of intraaortal injected iron oxide–labeled mesenchymal stem cells (MSC) into rats with acute kidney injury (AKI).

Kidney Protection and Regeneration Following Acute Injury: Progress Through Stem Cell Therapy

Acute kidney injury (AKI) is a common clinical entity with high morbidity and mortality rates and ever increasing medical costs. A large number of patients who are hospitalized with morbidities such as diabetes, vascular disease, or chronic kidney disease are at high risk to develop AKI due to ischemic and nephrotoxic insults. The pathophysiology of ischemic and toxic forms of AKI is complex and includes tubular and vascular cell damage and inflammation. Given the seriousness of this essentially therapy-resistant complication, treatment beyond supportive measures and renal replacement therapy is urgently needed. Recent stem cell research has shown promising results, and cell therapy-based interventions are advancing into clinical trials. An example is our phase 1 clinical trial (NCT00733876) in which cardiac surgery patients at high risk of postoperative AKI were treated safely with allogeneic mesenchymal stem cells. Together with the introduction of biomarkers for an earlier and specific AKI diagnosis, currently tested stem cell-based therapies are expected to provide an entirely new class of diagnostic and therapeutic tools.

Stable isotopes of calcium as tracers: methodology

While radioisotopic tracers have been used extensively, stable isotopes may, in principle, give the same information and offer several advantages for some investigations. Since there is no radiation involved, experiments using such high risk groups as children and pregnant women become possible. In addition, some studies may require using several different tracers. In the case of calcium, there are five stable isotopes ( 42Ca, 43Ca, 44Ca, &Ca and 48Ca) which may be used as tracers. If stable isotopic tracers are used, experiments may be done independent of isotope production schedules and the samples may be stored indefinitely. In spite of these advantages, stable isotopes have rarely been used as tracers because there has not been a satisfactory method of detection. Although isotopic abundances of calcium have been determined by both neutron activation analysis [1,2] and thermal ionization mass spectrometry [3,4], neither method has been used extensively for clinical investigation. This is presumably due to requirements for extensive sample preparation, and the necessary instrumentation. It has recently been shown [5] that fast atom bombardment mass spectrometry can be used to measure tracer levels of stable isotopes of calcium in plasma and urine. This technique offers good precision, high sensitivity, rapid analysis, and requires little or no sample preparation. This paper describes the first application of fast atom bombardment mass spectrometry and the double isotope technique to measure the fractional true absorption of calcium.

Protective actions of administered mesenchymal stem cells in acute kidney injury: relevance to clinical trials

Current therapies for acute kidney injury remain primarily supportive and have failed to reduce morbidity, mortality (>50%), and associated costs. This prompted our studies in which rats with bilateral ischemia/reperfusion-induced acute kidney injury were treated with bone marrow-derived, culture-expanded allogeneic mesenchymal stem cells. Their administration into the suprarenal aorta after reflow significantly protected renal function and hastened repair, mediated by complex antiapoptotic, mitogenic, anti-inflammatory, and immune modulating actions that were not elicited by isogeneic fibroblasts. Infused mesenchymal stem cells, recruited to renal sites of injury, did not significantly differentiate into target cells but rather disappeared from kidneys and other organs within 72 h. Furthermore, at 3 months, compared with vehicle-treated controls, renal function was well preserved and interstitial fibrosis was absent. These preclinical data served as the scientific basis for a recently completed Phase I Clinical Trial (http://www.clinicaltrials.gov; # NCT00733876), in which patients at high risk for cardiac surgery-associated AKI were treated with allogeneic mesenchymal stem cells. Until now, MSC therapy in the study subjects has been safe, and none of the patients has developed postoperative AKI or subsequent loss of renal function, suggesting that this novel form of therapy may have promise in this group of high-risk patients, which will be further investigated in a Phase II Trial.

The role of multipotent marrow stromal cells (MSCs) in tissue regeneration.

An extensive body of preclinical and clinical data has shown that administration of adult multipotent marrow stromal cells (MSCs) effectively ameliorates experimental and clinical conditions of many different organ systems. Differentiation into organ parenchymal cells, however, is very rare, and the main mechanism for organ protection and regeneration from different types of injury is the exertion of paracrine effects and stimulation of tissue repair. A large number of clinical trials have been conducted and are ongoing to investigate the safety and efficacy of MSCs in different organs after various types of organ injury. This article intends to give a brief overview about current applications of MSCs and mechanisms involved in organ protection and regeneration.

Limited Immune-Modulating Activity of Porcine Mesenchymal Stromal Cells Abolishes Their Protective Efficacy in Acute Kidney Injury

We demonstrated previously that administration of mesenchymal stromal cells (MSCs) after renal ischemia/reperfusion injury (IRI) in rats protected renal function and hastened repair through complex paracrine mechanisms. Here we investigated kidney-protective actions of MSCs in a porcine IRI model that may have relevance to human acute kidney injury (AKI). Groups of female pigs with bilateral IRI were infused with autologous or male allogeneic MSCs. No acute or late complications were observed, but unexpectedly, MSC therapy also had no beneficial effects on kidney function and histology. In vitro, we demonstrated substantial functional and phenotypic overlaps between rodent, human, and porcine MSCs, all of which exhibited trilineage differentiation, characteristic antigen profiles, and secretion of renoprotective vascular endothelial growth factor (VEGF)-A and insulin-like growth factor-1 (IGF-1). However, in striking contrast to human MSCs, porcine MSCs failed to inhibit the mixed lymphocyte reaction (MLR) and induced robust production of proinflammatory interleukin-6 (IL-6). In summary, in contrast to rodent models, treatment of porcine IRI with MSCs was not kidney-protective. This, we conclude, is due to the fact that porcine MSCs exert inadequate immune-modulating effects, further demonstrating that successful therapy of IRI with MSCs critically depends on their anti-inflammatory actions. As a consequence, treatment of AKI with MSCs is not informative regarding the investigation of the underlying mechanisms in this large animal model. We expect, however, that the treatment of human IRI of the kidney with immune-modulating MSCs will be as effective as in rodent models.