Using the Cecal Ligation and Puncture Model of Sepsis to Induce Rats to Multiple Organ Dysfunction.

Sepsis is a dysregulated hyperinflammatory disease caused by infection. Sepsis leads to multiple organ dysfunction syndrome (MODS), which is associated with high rates of mortality. The cecal ligation and puncture (CLP) model has been widely used in animals and has become the gold-standard method of replicating features of sepsis in humans. Despite several studies and modified CLP protocols, there are still open questions regarding the multifactorial determinants of its reproducibility and medical significance. In our protocol, which is also aimed at mimicking the sepsis observed in clinical practice, male Wistar rats are submitted to CLP with adequate fluid resuscitation (0.15 M NaCl, 25 ml/kg BW i.p.) immediately after surgery. At 6 h after CLP, additional fluid therapy (0.15 M NaCl, 25 ml/kg BW s.c.) and antibiotic therapy with imipenem-cilastatin (single dose of 14 mg/kg BW s.c.) are administered. The timing of the fluid and antibiotic therapy correspond to the initial care given when patients are admitted to the intensive care unit. This model of sepsis provides a useful platform for simulating human sepsis and could lay the groundwork for the development of new treatments.

Tamoxifen and bone morphogenic protein-7 modulate fibrosis and inflammation in the peritoneal fibrosis model developed in uremic rats.

BACKGROUND: Peritoneal fibrosis (PF) represents a long-term complication of peritoneal dialysis (PD), affecting peritoneal membrane (PM) integrity and function. Understanding the mechanisms underlying PF development in an uremic environment aiming alternative therapeutic strategies for treating this process is of great interest. The aim of this study was to analyze the effects of tamoxifen (TAM) and recombinant BMP7 (rBMP7) in an experimental model of PF developed in uremic rats. METHODS: To mimic the clinical situation of patients on long-term PD, a combo model, characterized by the combination of PF and CKD with severe uremia, was developed in Wistar rats. PF was induced by intraperitoneal (IP) injections of chlorhexidine gluconate (CG), and CKD was induced by an adenine-rich diet. Uremia was confirmed by severe hypertension, increased blood urea nitrogen (BUN> 120 mg/dL) and serum creatinine levels (> 2 mg/dL). Uremic rats with PF were treated with TAM (10 mg/Kg by gavage) or BMP7 (30 µg/Kg, IP). Animals were followed up for 30 days. RESULTS: CG administration in uremic rats induced a striking increase in PM thickness, neoangiogenesis, demonstrated by increased capillary density, and failure of ultrafiltration capacity. These morphological and functional changes were blocked by TAM or rBMP7 treatment. In parallel, TAM and rBMP7 significantly ameliorated the PM fibrotic response by reducing α-SMA, extracellular matrix proteins and TGF-ß expression. TAM or rBMP7 administration significantly inhibited peritoneal Smad3 expression in uremic rats with PF, prevented Smad3 phosphorylation, and induced a remarkable up-regulation of Smad7, an intracellular inhibitor of TGFß/Smad signaling, contributing to a negative modulation of profibrotic genes. Both treatments were also effective in reducing local inflammation, possibly by upregulating IκB-α expression in the PM of uremic rats with PF. In vitro experiments using primary peritoneal fibroblasts activated by TGF-ß confirmed the capacity of TAM or rBMP7 in blocking inflammatory mediators, such as IL-1ß expression. CONCLUSIONS: In conclusion, these findings indicate important roles of TGF-ß/Smad signaling in PF aggravated by uremia, providing data regarding potential therapeutic approaches with TAM or rBMP7 to block this process.

Progenitor/Stem Cell Delivery by Suprarenal Aorta Route in Acute Kidney Injury.

Progenitor/stem cell-based kidney regenerative strategies are a key step towards the development of novel therapeutic regimens for kidney disease treatment. However, the route of cell delivery, e.g., intravenous, intra-arterial, or intra-parenchymal, may affect the efficiency for kidney repair in different models of acute and chronic injury. Here, we describe a protocol of intra-aorta progenitor/stem cell injection in rats following either acute ischemia-reperfusion injury or acute proteinuria induced by puromycin aminonucleoside (PAN) - the experimental prototype of human minimal change disease and early stages of focal and segmental glomerulosclerosis. Vascular clips were applied across both renal pedicles for 35 min, or a single dose of PAN was injected via intra-peritoneal route, respectively. Subsequently, 2 x 106 stem cells [green fluorescent protein (GFP)-labeled c-Kit+ progenitor/stem cells or GFP-mesenchymal stem cells] or saline were injected into the suprarenal aorta, above the renal arteries, after application of a vascular clip to the abdominal aorta below the renal arteries. This approach contributed to engraftment rates of ∼10% at day 8 post ischemia-reperfusion injury, when c-Kit+ progenitor/stem cells were injected, which accelerated kidney recovery. Similar rates of engraftment were found after PAN-induced podocyte damage at day 21. With practice and gentle surgical technique, 100% of the rats could be injected successfully, and, in the week following injection, ∼ 85% of the injected rats will recover completely. Given the similarities in mammals, much of the data obtained from intra-arterial delivery of progenitor/stem cells in rodents can be tested in translational research and clinical trials with endovascular catheters in humans.

Kidney-derived c-kit+ progenitor/stem cells contribute to podocyte recovery in a model of acute proteinuria.

Kidney-derived c-kit+ cells exhibit progenitor/stem cell properties and can regenerate epithelial tubular cells following ischemia-reperfusion injury in rats. We therefore investigated whether c-kit+ progenitor/stem cells contribute to podocyte repair in a rat model of acute proteinuria induced by puromycin aminonucleoside (PAN), the experimental prototype of human minimal change disease and early stages of focal and segmental glomerulosclerosis. We found that c-kit+ progenitor/stem cells accelerated kidney recovery by improving foot process effacement (foot process width was lower in c-kit group vs saline treated animals, P = 0.03). In particular, these cells engrafted in small quantity into tubules, vessels, and glomeruli, where they occasionally differentiated into podocyte-like cells. This effect was related to an up regulation of α-Actinin-4 and mTORC2-Rictor pathway. Activation of autophagy by c-kit+ progenitor/stem cells also contributed to kidney regeneration and intracellular homeostasis (autophagosomes and autophagolysosomes number and LC3A/B-I and LC3A/B-II expression were higher in the c-kit group vs saline treated animals, P = 0.0031 and P = 0.0009, respectively). Taken together, our findings suggest that kidney-derived c-kit+ progenitor/stem cells exert reparative effects on glomerular disease processes through paracrine effects, to a lesser extent differentiation into podocyte-like cells and contribution to maintenance of podocyte cytoskeleton after injury. These findings have clinical implications for cell therapy of glomerular pathobiology.

Development of urinary organs in domestic cat during the embryonic and fetal periods.

The embryonic origin of the urogenital system came from the intermediate mesoderm. Kidney development involves three successive renal systems with a fast chronological overlap: the pronephro, the mesonephro, and the metanephro. Due to the lack of specific knowledge about this system in cats the present work aimed to describe their urinary organs development, focusing on the structures seen in pronephro, mesonephro, and metanephro during the embryonic and fetal stages of development. The techniques used in this study were: light microscopy, immunohistochemistry, scanning electron microscopy, and transmission electron microscopy. For that, embryos and fetuses from 12 pregnant mixed-breed domestic cats in different gestational stages were used to describe the proposed organs. The pronephro is present at early stages of embryonary development in embryos from 15 to 19 days with the presence of pronephro's corpuscles, ducts and tubules. The mesonephro is found, in general, between days 17 and 37, and contains mesonephric ducts, mesonephric tubules, and glomeruli. The metanephro is seen since 21 days of pregnancy with the presence of glomeruli, proximal and distal contorted tubules and at day 37, the cortex-medullary region is already differentiated. The evaluation of these structures enhances the knowledge about embryology of the urinary system in cats, aiding a better anatomical understanding of the system in the specie allowing the correlation with other species.

Acute Kidney Injury as a Condition of Renal Senescence.

Acute kidney injury (AKI), characterized by a sharp drop in glomerular filtration, continues to be a significant health burden because it is associated with high initial mortality, morbidity, and substantial health-care costs. There is a strong connection between AKI and mechanisms of senescence activation. After ischemic or nephrotoxic insults, a wide range of pathophysiological events occur. Renal tubular cell injury is characterized by cell membrane damage, cytoskeleton disruption, and DNA degradation, leading to tubular cell death by necrosis and apoptosis. The senescence mechanism involves interstitial fibrosis, tubular atrophy, and capillary rarefaction, all of which impede the morphological and functional recovery of the kidneys, suggesting a strong link between AKI and the progression of chronic kidney disease. During abnormal kidney repair, tubular epithelial cells can assume a senescence-like phenotype. Cellular senescence can occur as a result of cell cycle arrest due to increased expression of cyclin kinase inhibitors (mainly p21), downregulation of Klotho expression, and telomere shortening. In AKI, cellular senescence is aggravated by other factors including oxidative stress and autophagy. Given this scenario, the main question is whether AKI can be repaired and how to avoid the senescence process. Stem cells might constitute a new therapeutic approach. Mesenchymal stem cells (MSCs) can ameliorate kidney injury through angiogenesis, immunomodulation, and fibrosis pathway blockade, as well as through antiapoptotic and promitotic processes. Young umbilical cord-derived MSCs are better at increasing Klotho levels, and thus protecting tissues from senescence, than are adipose-derived MSCs. Umbilical cord-derived MSCs improve glomerular filtration and tubular function to a greater degree than do those obtained from adult tissue. Although senescence-related proteins and microRNA are upregulated in AKI, they can be downregulated by treatment with umbilical cord-derived MSCs. In summary, stem cells derived from young tissues, such as umbilical cord-derived MSCs, could slow the post-AKI senescence process.

Kidney-Derived c-Kit+ Cells Possess Regenerative Potential.

Kidney-derived c-Kit+ cells exhibit progenitor/stem cell properties in vitro (self-renewal capacity, clonogenicity, and multipotentiality). These cells can regenerate epithelial tubular cells following ischemia-reperfusion injury and accelerate foot processes effacement reversal in a model of acute proteinuria in rats. Several mechanisms are involved in kidney regeneration by kidney-derived c-Kit+ cells, including cell engraftment and differentiation into renal-like structures, such as tubules, vessels, and podocytes. Moreover, paracrine mechanisms could also account for kidney regeneration, either by stimulating proliferation of surviving cells or modulating autophagy and podocyte cytoskeleton rearrangement through mTOR-Raptor and -Rictor signaling, which ultimately lead to morphological and functional improvement. To gain insights into the functional properties of c-Kit+ cells during kidney development, homeostasis, and disease, studies on lineage tracing using transgenic mice will unveil their fate. The results obtained from these studies will set the basis for establishing further investigation on the therapeutic potential of c-Kit+ cells for treatment of kidney disease in preclinical and clinical studies. Stem Cells Translational Medicine 2018;7:317-324.

Randomized Comparison of Allogeneic Versus Autologous Mesenchymal Stem Cells for Nonischemic Dilated Cardiomyopathy: POSEIDON-DCM Trial.

BACKGROUND: Although human mesenchymal stem cells (hMSCs) have been tested in ischemic cardiomyopathy, few studies exist in chronic nonischemic dilated cardiomyopathy (NIDCM). OBJECTIVES: The authors conducted a randomized comparison of safety and efficacy of autologous (auto) versus allogeneic (allo) bone marrow-derived hMSCs in NIDCM. METHODS: Thirty-seven patients were randomized to either allo- or auto-hMSCs in a 1:1 ratio. Patients were recruited between December 2011 and July 2015 at the University of Miami Hospital. Patients received hMSCs (100 million) by transendocardial stem cell injection in 10 left ventricular sites. Treated patients were evaluated at baseline, 30 days, and 3-, 6-, and 12-months for safety (serious adverse events [SAE]), and efficacy endpoints: ejection fraction, Minnesota Living with Heart Failure Questionnaire, 6-min walk test, major adverse cardiac events, and immune biomarkers. RESULTS: There were no 30-day treatment-emergent SAEs. Twelve-month SAE incidence was 28.2% with allo-hMSCs versus 63.5% with auto-hMSCs (p = 0.1004 for the comparison). One allo-hMSC patient developed an elevated (>80) donor-specific calculated panel reactive antibody level. The ejection fraction increased in allo-hMSC patients by 8.0 percentage points (p = 0.004) compared with 5.4 with auto-hMSCs (p = 0.116; allo vs. auto p = 0.4887). The 6-min walk test increased with allo-hMSCs by 37.0 m (p = 0.04), but not auto-hMSCs at 7.3 m (p = 0.71; auto vs. allo p = 0.0168). MLHFQ score decreased in allo-hMSC (p = 0.0022) and auto-hMSC patients (p = 0.463; auto vs. allo p = 0.172). The major adverse cardiac event rate was lower, too, in the allo group (p = 0.0186 vs. auto). Tumor necrosis factor-α decreased (p = 0.0001 for each), to a greater extent with allo-hMSCs versus auto-hMSCs at 6 months (p = 0.05). CONCLUSIONS: These findings demonstrated safety and clinically meaningful efficacy of allo-hMSC versus auto-hMSC in NIDCM patients. Pivotal trials of allo-hMSCs are warranted based on these results. (Percutaneous Stem Cell Injection Delivery Effects on Neomyogenesis in Dilated Cardiomyopathy [PoseidonDCM]; NCT01392625).

Mesenchymal Stem Cells as Therapeutic Candidates for Halting the Progression of Diabetic Nephropathy.

Mesenchymal stem cells (MSCs) possess pleiotropic properties that include immunomodulation, inhibition of apoptosis, fibrosis and oxidative stress, secretion of trophic factors, and enhancement of angiogenesis. These properties provide a broad spectrum for their potential in a wide range of injuries and diseases, including diabetic nephropathy (DN). MSCs are characterized by adherence to plastic, expression of the surface molecules CD73, CD90, and CD105 in the absence of CD34, CD45, HLA-DR, and CD14 or CD11b and CD79a or CD19 surface molecules, and multidifferentiation capacity in vitro. MSCs can be derived from many tissue sources, consistent with their broad, possibly ubiquitous distribution. This article reviews the existing literature and knowledge of MSC therapy in DN, as well as the most appropriate rodent models to verify the therapeutic potential of MSCs in DN setting. Some preclinical relevant studies are highlighted and new perspectives of combined therapies for decreasing DN progression are discussed. Hence, improved comprehension and interpretation of experimental data will accelerate the progress towards clinical trials that should assess the feasibility and safety of this therapeutic approach in humans. Therefore, MSC-based therapies may bring substantial benefit for patients suffering from DN.

Synergistic Effects of Combined Cell Therapy for Chronic Ischemic Cardiomyopathy.

BACKGROUND: Both bone marrow-derived mesenchymal stem cells (MSCs) and c-kit(+) cardiac stem cells (CSCs) improve left ventricular remodeling in porcine models and clinical trials. Using xenogeneic (human) cells in immunosuppressed animals with acute ischemic heart disease, we previously showed that these 2 cell types act synergistically. OBJECTIVES: To more accurately model clinical applications for heart failure, this study tested whether the combination of autologous MSCs and CSCs produce greater improvement in cardiac performance than MSCs alone in a nonimmunosuppressed porcine model of chronic ischemic cardiomyopathy. METHODS: Three months after ischemia/reperfusion injury, Göttingen swine received transendocardial injections with MSCs alone (n = 6) or in combination with cardiac-derived CSCs (n = 8), or placebo (vehicle; n = 6). Cardiac functional and anatomic parameters were assessed using cardiac magnetic resonance at baseline and before and after therapy. RESULTS: Both groups of cell-treated animals exhibited significantly reduced scar size (MSCs -44.1 ± 6.8%; CSC/MSC -37.2 ± 5.4%; placebo -12.9 ± 4.2%; p < 0.0001), increased viable tissue, and improved wall motion relative to placebo 3 months post-injection. Ejection fraction (EF) improved (MSCs 2.9 ± 1.6 EF units; CSC/MSC 6.9 ± 2.8 EF units; placebo 2.5 ± 1.6 EF units; p = 0.0009), as did stroke volume, cardiac output, and diastolic strain only in the combination-treated animals, which also exhibited increased cardiomyocyte mitotic activity. CONCLUSIONS: These findings illustrate that interactions between MSCs and CSCs enhance cardiac performance more than MSCs alone, establish the safety of autologous cell combination strategies, and support the development of second-generation cell therapeutic products.

S-nitrosoglutathione reductase-dependent PPARγ denitrosylation participates in MSC-derived adipogenesis and osteogenesis.

Bone marrow-derived mesenchymal stem cells (MSCs) are a common precursor of both adipocytes and osteoblasts. While it is appreciated that PPARγ regulates the balance between adipogenesis and osteogenesis, the roles of additional regulators of this process remain controversial. Here, we show that MSCs isolated from mice lacking S-nitrosoglutathione reductase, a denitrosylase that regulates protein S-nitrosylation, exhibited decreased adipogenesis and increased osteoblastogenesis compared with WT MSCs. Consistent with this cellular phenotype, S-nitrosoglutathione reductase-deficient mice were smaller, with reduced fat mass and increased bone formation that was accompanied by elevated bone resorption. WT and S-nitrosoglutathione reductase-deficient MSCs exhibited equivalent PPARγ expression; however, S-nitrosylation of PPARγ was elevated in S-nitrosoglutathione reductase-deficient MSCs, diminishing binding to its downstream target fatty acid-binding protein 4 (FABP4). We further identified Cys 139 of PPARγ as an S-nitrosylation site and demonstrated that S-nitrosylation of PPARγ inhibits its transcriptional activity, suggesting a feedback regulation of PPARγ transcriptional activity by NO-mediated S-nitrosylation. Together, these results reveal that S-nitrosoglutathione reductase-dependent modification of PPARγ alters the balance between adipocyte and osteoblast differentiation and provides checkpoint regulation of the lineage bifurcation of these 2 lineages. Moreover, these findings provide pathophysiological and therapeutic insights regarding MSC participation in adipogenesis and osteogenesis.

miR-30e targets IGF2-regulated osteogenesis in bone marrow-derived mesenchymal stem cells, aortic smooth muscle cells, and ApoE-/- mice.

AIMS: Activation of an osteogenic transcriptional program contributes to the initiation of aortic calcification in atherosclerosis. The role of microRNAs in regulating aortic calcification is understudied. We tested the hypothesis that miR-30e regulates an osteogenic program in bone marrow-derived mesenchymal stem cells (MSCs), aortic smooth muscle cells (SMCs), and ApoE(-/-) mice. METHODS AND RESULTS: In aortas of wild-type mice, we found that miR-30e is highly expressed in medial SMCs. In aortas of old ApoE(-/-) mice, we found that miR-30e transcripts are down-regulated in an inverse relation to the osteogenic markers Runx2, Opn, and Igf2. In vitro, miR-30e over-expression reduced the proliferation of MSCs and SMCs while increasing adipogenic differentiation of MSCs and smooth muscle differentiation of SMCs. In MSCs and SMCs over-expressing miR-30e, microarrays and qPCR showed repression of an osteogenic gene panel including Igf2. Inhibiting miR-30e in MSCs increased Igf2 transcripts. In SMCs, IGF2 recombinant protein rescued miR-30e-repressed osteogenic differentiation. Luciferase and mutagenesis assays showed binding of miR-30e to a novel and essential site at the 3'UTR of Igf2. In ApoE(-/-) mice, injections of antimiR-30e oligos increased Igf2 expression in the aortas and livers and significantly enhanced OPN protein expression and calcium deposition in aortic valves. CONCLUSION: miR-30e represses the osteogenic program in MSCs and SMCs by targeting IGF2 and drives their differentiation into adipogenic or smooth muscle lineage, respectively. Our data suggest that down-regulation of miR-30e in aortas with age and atherosclerosis triggers vascular calcification. The miR-30e pathway plays an important regulatory role in vascular diseases.

Rationale and design of the Percutaneous Stem Cell Injection Delivery Effects on Neomyogenesis in Dilated Cardiomyopathy (the POSEIDON-DCM study): a phase I/II, randomized pilot study of the comparative safety and efficacy of transendocardial injection of autologous mesenchymal stem cell vs. allogeneic mesenchymal stem cells in patients with non-ischemic dilated cardiomyopathy.

While accumulating clinical trials have focused on the impact of cell therapy in patients with acute myocardial infarction (MI) and ischemic cardiomyopathy, there are fewer efforts to examine cell-based therapy in patients with non-ischemic cardiomyopathy (NICM). We hypothesized that cell therapy could have a similar impact in NICM. The POSEIDON-DCM trial is a phase I/II trial designed to address autologous vs. allogeneic bone marrow-derived mesenchymal stem cells (MSCs) in patients with NICM. In this study, cells will be administered transendocardially with the NOGA injection-catheter system to patients (n = 36) randomly allocated to two treatment groups: group 1 (n = 18 auto-human mesenchymal stem cells (hMSC)) and group 2 (n = 18 allo-hMSCs). The primary and secondary objectives are, respectively, to demonstrate the safety and efficacy of allo-hMSCS vs. auto-hMSCs in patients with NICM. This study will establish safety of transendocardial injection of stem cells (TESI), compare phenotypic outcomes, and offer promising advances in the field of cell-based therapy in patients with NICM.

C-kit(+) cells isolated from developing kidneys are a novel population of stem cells with regenerative potential.

The presence of tissue specific precursor cells is an emerging concept in organ formation and tissue homeostasis. Several progenitors are described in the kidneys. However, their identity as a true stem cell remains elusive. Here, we identify a neonatal kidney-derived c-kit(+) cell population that fulfills all of the criteria as a stem cell. These cells were found in the thick ascending limb of Henle's loop and exhibited clonogenicity, self-renewal, and multipotentiality with differentiation capacity into mesoderm and ectoderm progeny. Additionally, c-kit(+) cells formed spheres in nonadherent conditions when plated at clonal density and expressed markers of stem cells, progenitors, and differentiated cells. Ex vivo expanded c-kit(+) cells integrated into several compartments of the kidney, including tubules, vessels, and glomeruli, and contributed to functional and morphological improvement of the kidney following acute ischemia-reperfusion injury in rats. Together, these findings document a novel neonatal rat kidney c-kit(+) stem cell population that can be isolated, expanded, cloned, differentiated, and used for kidney repair following acute kidney injury. These cells have important biological and therapeutic implications.

S-nitrosoglutathione reductase (GSNOR) enhances vasculogenesis by mesenchymal stem cells.

Although nitric oxide (NO) signaling promotes differentiation and maturation of endothelial progenitor cells, its role in the differentiation of mesenchymal stem cells (MSCs) into endothelial cells remains controversial. We tested the role of NO signaling in MSCs derived from WT mice and mice homozygous for a deletion of S-nitrosoglutathione reductase (GSNOR(-/-)), a denitrosylase that regulates S-nitrosylation. GSNOR(-/-) MSCs exhibited markedly diminished capacity for vasculogenesis in an in vitro Matrigel tube-forming assay and in vivo relative to WT MSCs. This decrease was associated with down-regulation of the PDGF receptorα (PDGFRα) in GSNOR(-/-) MSCs, a receptor essential for VEGF-A action in MSCs. Pharmacologic inhibition of NO synthase with L-N(G)-nitroarginine methyl ester (L-NAME) and stimulation of growth hormone-releasing hormone receptor (GHRHR) with GHRH agonists augmented VEGF-A production and normalized tube formation in GSNOR(-/-) MSCs, whereas NO donors or PDGFR antagonist reduced tube formation ∼50% by murine and human MSCs. The antagonist also blocked the rescue of tube formation in GSNOR(-/-) MSCs by L-NAME or the GHRH agonists JI-38, MR-409, and MR-356. Therefore, GSNOR(-/-) MSCs have a deficient capacity for endothelial differentiation due to downregulation of PDGFRα related to NO/GSNOR imbalance. These findings unravel important aspects of modulation of MSCs by VEGF-A activation of the PDGFR and illustrate a paradoxical inhibitory role of S-nitrosylation signaling in MSC vasculogenesis. Accordingly, disease states characterized by NO deficiency may trigger MSC-mediated vasculogenesis. These findings have important implications for therapeutic application of GHRH agonists to ischemic disorders.

Charcot neuroarthropathy after simultaneous pancreas-kidney transplant.

BACKGROUND: Immunosuppressive regimen is associated with several metabolic adverse effects. Bone loss and fractures are frequent after transplantation and involve multifactorial mechanisms. METHODS: A retrospective analysis of 130 patients submitted to simultaneous pancreas-kidney transplantation (SPKT) and an identification of risk factors involved in de novo Charcot neuroarthropathy by multivariate analysis were used; P<0.05 was considered significant. RESULTS: Charcot neuroarthropathy was diagnosed in 4.6% of SPKT recipients during the first year. Cumulative glucocorticoid doses (daily dose plus methylprednisolone pulse) during the first 6 months both adjusted to body weight (>78 mg/kg) and not adjusted to body weight were associated with Charcot neuroarthropathy (P=0.001 and P<0.0001, respectively). Age, gender, race, time on dialysis, time of diabetes history, and posttransplantation hyperparathyroidism were not related to Charcot neuroarthropathy after SPKT. CONCLUSIONS: Glucocorticoids are the main risk factors for de novo Charcot neuroarthropathy after SPKT. Protocols including glucocorticoid avoidance or minimization should be considered.