CD4−CD8− double negative (DN) αβ T cells are legitimate components of the normal immune system. However, they are poorly understood and largely ignored by immunologists because of their historical association with the lymphoproliferation that occurs in mice (lpr and gld) and humans (ALPS patients) with impaired Fas-mediated apoptosis where they are considered abnormal T cells. We believe that the traditional view that DN T cells that cause lymphoproliferation (hereafter referred to as lpr DN T cells) are CD4 and CD8 T cells that lost their coreceptor, conceived more than two decades ago, is flawed and that conflating lpr DN T cells with DN T cells found in normal immune system (hereafter referred to as nDN T cells) is unnecessarily dampening interest of this potentially important cell type. To begin rectifying these misperceptions, we will revisit the traditional view of lpr DN T cells and show that it does not hold true in light of recent immunological advances. In lieu of it, we offer a new model proposing that Fas-mediated apoptosis actively removes normally existing DN T cells from the periphery and that impaired Fas-mediated apoptosis leads to accumulation of these cells rather than de novo generation of DN T cells from activated CD4 or CD8 T cells. By doing so, we hope to provoke a new discussion that may lead to a consensus about the origin of lpr DN T cells and regulation of their homeostasis by the Fas pathway and reignite wider interest in nDN T cells.
A comprehensive assessment of the association of patients’ renal replacement therapy (RRT) modality on their participation in life activities (physical function, travel, recreation, freedom, work) is needed.
Systematic review of peer-reviewed published studies.
Setting & Population
Adults undergoing RRT (hemodialysis, peritoneal dialysis, or transplantation).
Selection Criteria for Studies
We searched PubMed, Cochrane Library, and EMBASE from January 1980 through April 2012 for English-language articles that compared participation in life activities among patients receiving 1) hemodialysis compared with peritoneal dialysis, 2) hemodialysis compared with kidney transplantation, or 3) peritoneal dialysis compared with kidney transplantation.
Reported rates of physical function, travel, recreation, freedom, and work-related activities by RRT modality.
A total of 46 studies (6 prospective cohort, 38 cross-sectional, and 2 pre-post transplantation) provided relevant comparisons of life participation activities among patients treated with hemodialysis, peritoneal dialysis, and kidney transplantation. Studies were conducted from 1985 to 2011 among diverse patient populations in 16 distinct locations. A majority of studies reported greater life participation rates among patients with kidney transplants compared to patients receiving either hemodialysis or peritoneal dialysis. In contrast, a majority of studies reported no differences in outcomes between patients receiving hemodialysis and patients receiving peritoneal dialysis. These results were consistent throughout the study period, across diverse populations, and among the subset of studies that performed appropriate adjustments for potential confounding factors.
Many studies included in the review had significant design weaknesses.
Evidence suggests patients with kidney transplants may experience better rates of life participation compared to patients receiving dialysis, while patients receiving hemodialysis and patients receiving peritoneal dialysis may experience similar rates of life participation. Rigorously performed studies are needed to better inform patients about the association of RRT on these important patient reported outcomes.
dialysis; ESRD treatment; kidney transplantation; physical functioning; quality of life; social participation
Focal segmental glomerulosclerosis (FSGS) recurs after kidney transplantation in more than 30 % of cases and can lead to allograft loss. Serum soluble urokinase -type plasminogen activator receptor (suPAR) is implicated in the pathogenesis of native and recurrent FSGS.
We conducted a retrospective study of 25 adults with post-transplant FSGS. We investigated the relationship between suPAR levels and podocyte changes and the impact of therapy on podocyte structure. We assessed response to therapy by improvement in proteinuria, allograft function and resolution of histologic changes.
A median of 15 (interquartile range: 10–23) plasmapheresis sessions was administered; 13 of the subjects also received rituximab. Median pre-treatment suPAR levels were higher among those with severe (≥75%) versus those with mild (≤25%) podocyte foot process effacement (13,030 vs. 4,806 pg/mL; P=0.02). Overall, mean ± standard deviation of proteinuria improved from 5.1 ± 3.8 to 2.1 ± 2.8 mg/dL (P=0.003), mean podocyte effacement decreased from 57 ± 33% to 22 ± 22 % (P=0.0001), estimated glomerular filtration rates increased from median (interquartile range) of 32.9 (20.6 – 44.2) to 39.3 (28.8 – 63.4) (P<0.0001) and suPAR levels decreased from a median of 6,781 pg/ml to 4,129 pg/ml (P=0.02) with therapy.
Podocyte effacement is the first pathological manifestation of FSGS post-transplant. The degree of podocyte effacement correlates with suPAR levels at time of diagnosis. Response to therapy results in significant reduction of suPAR levels and complete or significant improvement of podocyte effacement.
kidney transplant; podocyte effacement; FSGS; suPAR; rituximab
Most previous studies of living kidney donors have been retrospective and have lacked suitable healthy controls. Needed are prospective controlled studies to better understand the effects of a mild reduction in kidney function from kidney donation in otherwise normal individuals.
Prospective, controlled, observational cohort study.
Setting & Participants
Consecutive patients approved for donation at 8 transplant centers in the US were asked to participate. For every donor enrolled, an equally healthy control with 2 kidneys who theoretically would have been suitable to donate a kidney was also enrolled.
At baseline pre-donation and at 6 months after donation, a medical history, vital signs, measured (iohexol) glomerular filtration rate and other measurements were collected. There were 201 donors and 198 controls that completed both baseline and 6 month visits and form the basis of this report.
Compared to controls, donors had 28% lower glomerular filtration rate at 6 months (94.6±15.1 [SD] v. 67.6±10.1 mL/min/1.73m2; P<0.001), associated with a 23% greater parathyroid hormone (42.8±15.6 v. 52.7±20.9 pg/mL; P<0.001), 5.4% lower serum phosphate (3.5±0.5 v. 3.3±0.5 mg/dL; P<0.001), 3.7% lower hemoglobin (13.6±1.4 v. 13.1±1.2 g/dL; P<0.001), 8.2% greater uric acid (4.9±1.2 v. 5.3±1.1 mg/dL; P<0.001), 24% greater homocysteine (1.20±0.34 v. 1.49±0.43 mg/L; P<0.001), and 1.5% lower high density lipoprotein cholesterol (54.9±16.4 v. 54.1±13.9 mg/dL; P=0.03) level. There were no differences in albumin-creatinine ratios (5.0 [IQR, 4.0-6.6] v. 5.0 [IQR, 3.3-5.4] mg/g; P=0.5), office blood pressure, or glucose homeostasis.
Short duration of follow-up and possible bias resulting from an inability to screen controls with kidney and vascular imaging performed in donors.
Kidney donors have some, but not all, abnormalities typically associated with mild chronic kidney disease 6 months after donation. Additional follow up is warranted.
We conducted focus group meetings of African American and non-African American patients with end-stage renal disease (six groups) and their family members (six groups), stratified by race/ethnicity and treatment. We elicited differences in participants’ experiences with shared decision making about initiating renal replacement therapy (RRT; that is, hemodialysis, peritoneal dialysis, or a kidney transplant). Patients were often very sick when initiating RRT, and had little, if any, time to make a decision about what type of RRT to initiate. They also lacked sufficient information about alternative treatment options prior to initiation. Family members played supportive roles and shared in decision making when possible. Reports were similar for African American and non-African American participants. Our findings suggest that a greater emphasis on the improved engagement of patients and their families in shared decision making about RRT initiation is needed for both ethnic/racial minorities and nonminorities.
African Americans; communication; medical; decision making; illness and disease; chronic; illness and disease; experiences; minorities; nephrology
Accumulated to-date microarray data on ischemia reperfusion injury (IRI) of kidney represent a powerful source for identifying new targets and mechanisms of kidney IRI. In this study, we conducted a meta-analysis of gene expression profiles of kidney IRI in human, pig, rat, and mouse models, using a new scoring method to correct for the bias of overrepresented species. The gene expression profiles were obtained from the public repositories for 24 different models. After filtering against inclusion criteria 21 experimental settings were selected for meta-analysis and were represented by 11 rat models, 6 mouse models, and 2 models each for pig and human, with a total of 150 samples. Meta-analysis was conducted using expression-based genome-wide association study (eGWAS). The eGWAS results were corrected for a rodent species bias using a new weighted scoring algorithm, which favors genes with unidirectional change in expression in all tested species.
Our meta-analysis corrected for a species bias, identified 46 upregulated and 1 downregulated genes, of which 26 (55%) were known to be associated with kidney IRI or kidney transplantation, including LCN2, CCL2, CXCL1, HMOX1, ICAM1, ANXA1, and TIMP1, which justified our approach. Pathway analysis of our candidates identified “Acute renal failure panel” as the most implicated pathway, which further validates our new method. Among new IRI candidates were 10 novel (<5 published reports related to kidney IRI) and 11 new candidates (0 reports related to kidney IRI) including the most prominent candidates ANXA2, CLDN4, and TYROBP. The cross-species expression pattern of these genes allowed us to generate three workable hypotheses of kidney IRI, one of which was confirmed by an additional study.
Our first in the field kidney IRI meta-analysis of 150 microarray samples, corrected for a species bias, identified 10 novel and 11 new candidate genes. Moreover, our new meta-analysis correction method improved gene candidate selection by identifying genes that are model and species independent, as a result, function of these genes can be directly extrapolated to the disease state in human and facilitate translation of potential diagnostic or therapeutic properties of these candidates to the bedside.
Kidney; Ischemia reperfusion injury; Bioinformatics; Meta-analysis
Little is known regarding the types of information African American and non-African American patients with chronic kidney disease (CKD) and their families need to inform renal replacement therapy (RRT) decisions.
In 20 structured group interviews, we elicited views of African American and non-African American patients with CKD and their families about factors that should be addressed in educational materials informing patients’ RRT selection decisions. We asked participants to select factors from a list and obtained their open-ended feedback.
Ten groups of patients (5 African American, 5 non-African American; total 68 individuals) and ten groups of family members (5 African American, 5 non-African American; total 62 individuals) participated. Patients and families had a range (none to extensive) of experiences with various RRTs. Patients identified morbidity or mortality, autonomy, treatment delivery, and symptoms as important factors to address. Family members identified similar factors but also cited the effects of RRT decisions on patients’ psychological well-being and finances. Views of African American and non-African American participants were largely similar.
Educational resources addressing the influence of RRT selection on patients’ morbidity and mortality, autonomy, treatment delivery, and symptoms could help patients and their families select RRT options closely aligned with their values. Including information about the influence of RRT selection on patients’ personal relationships and finances could enhance resources’ cultural relevance for African Americans.
Decision-making; Renal replacement therapy; Family members; African American
Acute kidney injury (AKI) often results from ischemia reperfusion, sepsis, or exposure to nephrotoxins and is associated with a high rate of mortality and morbidity. Advances in understanding the pathophysiology of AKI may lead to the development of specific therapies. Although there is evidence of an important role for immune cells in AKI, the specific relevant populations and the mechanisms of their actions are unclear. In this issue of the JCI, Li et al. demonstrate that adenosine manipulates DC responses to kidney injury, raising hope that immunotherapy could be a tangible approach to AKI.
To test the hypothesis that IL-6 contributes to the development of ventilator-associated lung injury (VALI), IL-6-deficient (IL6−/−) and wild-type control (WT) mice received intratracheal hydrochloric acid followed by randomization to MV (MV+IT HCl) or spontaneous ventilation (IT HCl). After 4 hr, injury was assessed by estimation of lung lavage protein concentration and total and differential cell counts, wet/dry lung weight ratio, pulmonary cell death, histologic inflammation score (LIS), and parenchymal myeloperoxidase (MPO) concentration. Vascular endothelial growth factor (VEGF) concentration was measured in lung lavage and homogenate, as IL-6 and stretch both regulate expression of this potent mediator of permeability. MV-induced increases in alveolar barrier dysfunction and lavage VEGF were attenuated in IL6−/− mice as compared with WT controls, whereas tissue VEGF concentration increased. The effects of IL-6 deletion on alveolar permeability and VEGF concentration were inflammation-independent, as parenchymal MPO concentration, LIS, and lavage total and differential cell counts did not differ between WT and IL6−/− mice following IT HCl+MV.
These data support a role for IL-6 in promoting VALI in this two-hit model. Strategies to interfere with IL-6 expression or signaling may represent important therapeutic targets to limit the injurious effects of MV in inflamed lungs.
mechanical ventilation; pulmonary barrier dysfunction; acid aspiration; vascular endothelial growth factor (VEGF)
Few educational resources have been developed to inform patients’ renal replacement therapy (RRT) selection decisions. Patients progressing toward end stage renal disease (ESRD) must decide among multiple treatment options with varying characteristics. Complex information about treatments must be adequately conveyed to patients with different educational backgrounds and informational needs. Decisions about treatment options also require family input, as families often participate in patients’ treatment and support patients’ decisions. We describe the development, design, and preliminary evaluation of an informational, evidence-based, and patient-and family-centered decision aid for patients with ESRD and varying levels of health literacy, health numeracy, and cognitive function.
We designed a decision aid comprising a complementary video and informational handbook. We based our development process on data previously obtained from qualitative focus groups and systematic literature reviews. We simultaneously developed the video and handbook in “stages.” For the video, stages included (1) directed interviews with culturally appropriate patients and families and preliminary script development, (2) video production, and (3) screening the video with patients and their families. For the handbook, stages comprised (1) preliminary content design, (2) a mixed-methods pilot study among diverse patients to assess comprehension of handbook material, and (3) screening the handbook with patients and their families.
The video and handbook both addressed potential benefits and trade-offs of treatment selections. The 50-minute video consisted of demographically diverse patients and their families describing their positive and negative experiences with selecting a treatment option. The video also incorporated health professionals’ testimonials regarding various considerations that might influence patients’ and families’ treatment selections. The handbook was comprised of written words, pictures of patients and health care providers, and diagrams describing the findings and quality of scientific studies comparing treatments. The handbook text was written at a 4th to 6th grade reading level. Pilot study results demonstrated that a majority of patients could understand information presented in the handbook. Patient and families screening the nearly completed video and handbook reviewed the materials favorably.
This rigorously designed decision aid may help patients and families make informed decisions about their treatment options for RRT that are well aligned with their values.
Kidney disease; Decision aid; Literacy; Numeracy; Cognitive function
Living related kidney transplantation (LRT) is underutilized, particularly among African Americans. The effectiveness of informational and financial interventions to enhance informed decision-making among African Americans with end stage renal disease (ESRD) and improve rates of LRT is unknown.
We report the protocol of the Providing Resources to Enhance African American Patients’ Readiness to Make Decisions about Kidney Disease (PREPARED) Study, a two-phase study utilizing qualitative and quantitative research methods to design and test the effectiveness of informational (focused on shared decision-making) and financial interventions to overcome barriers to pursuit of LRT among African American patients and their families. Study Phase I involved the evidence-based development of informational materials as well as a financial intervention to enhance African American patients’ and families’ proficiency in shared decision-making regarding LRT. In Study Phase 2, we are currently conducting a randomized controlled trial in which patients with new-onset ESRD receive 1) usual dialysis care by their nephrologists, 2) the informational intervention (educational video and handbook), or 3) the informational intervention in addition to the option of participating in a live kidney donor financial assistance program. The primary outcome of the randomized controlled trial will include patients’ self-reported rates of consideration of LRT (including family discussions of LRT, patient-physician discussions of LRT, and identification of a LRT donor).
Results from the PREPARED study will provide needed evidence on ways to enhance the decision to pursue LRT among African American patients with ESRD.
Shared decision-making; Live kidney transplantation; Live kidney donation; Chronic kidney disease; End stage renal disease
Acute kidney injury (AKI) leads to increased lung microvascular permeability, leukocyte infiltration and upregulation of soluble inflammatory proteins in rodents. Most work investigating connections between AKI and pulmonary dysfunction, however, has focused on characterizing whole lung tissue changes associated with AKI. Studies at the cellular level are essential to understanding the molecular basis of lung changes during AKI. Given that the pulmonary microvascular barrier is functionally abnormal during AKI, we hypothesized that AKI induces a specific pro-inflammatory and pro-apoptotic lung endothelial cell (EC) response. Four and 24 hours after kidney ischemia/reperfusion injury (IRI) or bilateral nephrectomy (BNx), murine pulmonary endothelial cells were isolated via tissue digestion followed by magnetic bead sorting. Purified lung ECs were analyzed for changes in mRNA expression using real time “Superarray” PCR analysis of genes related to endothelial cell function. In parallel experiments, confluent rat pulmonary microvascular ECs were treated with AKI or control serum to evaluate functional cellular alterations. Immunocytochemistry and FACS analysis of Annexin V/PI staining were employed to evaluate cytoskeletal changes and promotion of apoptosis. Isolated murine pulmonary endothelial cells exhibited significant changes in expression of gene products related to inflammation, vascular reactivity and programmed cell death. Further experiments using an in vitro rat pulmonary microvascular EC system revealed that AKI serum induced functional cellular changes related to apoptosis, including structural actin alterations and phosphatidylserine translocation. Analysis and segregation of both upregulated and downregulated genes into functional roles suggests that these transcriptional events likely participate in the transition to an activated pro-inflammatory and pro-apoptotic endothelial cell phenotype during AKI. Further mechanistic analysis of EC-specific events in the lung during AKI might reveal potential novel therapeutic targets for the deleterious kidney-lung crosstalk in the critically ill patient.
AKI; Ischemia-reperfusion; inflammation; apoptosis; Superarray; MODS
Radionuclide imaging of the kidneys with gamma cameras involves the use of labeled molecules seeking functionally critical molecular mechanisms in order to detect the pathophysiology of the diseased kidneys and achieve an early, sensitive and accurate diagnosis. The most recent imaging technology, PET, permits quantitative imaging of the kidney at a spatial resolution appropriate for the organ. H215O, 82RbCl, and [64Cu] ETS are the most important radiopharmaceuticals for measuring renal blood flow.
The renin angiotensin system is the most important regulator of renal blood flow; this role is being interrogated by detecting angiotensin receptor subtype AT1R using in vivo PET imaging. Membrane organic anion transporters are important for the function of the tubular epithelium; therefore, Tc-99m MAG3 as well as some novel radiopharmaceuticals such as copper-64 labeled mono oxo-tetraazamacrocyclic ligands have been utilized for molecular renal imaging. Additionally, other radioligands that interact with the organic cation transporters or peptide transporters have developed.
Focusing on early detection of kidney injury at the molecular level is an evolving field of great significance. Potential imaging targets are the kidney injury molecule- 1 (KIM-1) that is highly expressed in kidney injury and renal cancer but not in normal kidneys. While pelvic clearance, in addition to parenchymal transport, is an important measure in obstructive nephropathy, techniques that focus on upregulated molecules in response to tissue stress resulted from obstruction will be of great implication. Monocyte chemoattractant protein -1 (MCP-1) is a well-suited molecule in this case.
The greatest advances in molecular imaging of the kidneys have been recently achieved in detecting renal cancer. In addition to the ubiquitous [18F]FDG, other radioligands such as [11C]acetate and anti-[18F]FACBC have emerged. Radioimmuno-imaging with [124I]G250 could lead to radioimmunotherapy for renal cancer.
Considering the increasing age of general population, the incidence of kidney diseases such as atherosclerosis, diabetic nephropathy, and cancer is expected to increase. Successful management of these diseases offers an opportunity and a challenge for development of novel molecular imaging technologies.
There is an increase in the older incident end-stage renal disease population that is associated with an increasing prevalence of end-stage renal disease in the United States. This trend is paralleled by an increasing rate of kidney transplantation in the elderly. Although patient survival is lower in older versus younger kidney recipients, the elderly benefit from a reduction in mortality rate and improved quality of life with transplantation compared with dialysis. Immunologic, physiologic, and psychosocial factors influence transplant outcomes and should be recognized in the care of the elderly transplant patient. In this review, we discuss transplantation in the elderly patient, particularly the topics of access to transplantation, patient and graft survival, the impact of donor quality on transplant outcomes, immunology and immunosuppression of aging, and ethical considerations in the development of an equitable organ allocation scheme.
Elderly; kidney transplant; aging; patient survival; graft survival
Despite advances in renal replacement therapy, the mortality of acute kidney injury (AKI) has remained high, especially when associated with distant organ dysfunction such as acute lung injury (ALI). Mortality rates for combined AKI/ALI reach 80% in critically ill patients. While the clinical presentation of AKI-associated ALI is characterized by increased pulmonary edema, a defining feature of the syndrome, the AKI-induced lung effects extend beyond simple volume overload. Furthermore, ALI and associated mechanical ventilation frequently lead to a decline in renal hemodynamics, structure and function. New experimental data have emerged in recent years focusing on the interactive effects of kidney and lung dysfunction, and these studies have highlighted the pathophysiological importance of proinflammatory and proapoptotic pathways as well as the complex nature of interorgan crosstalk. This review will examine our current understanding of the deleterious kidney-lung crosstalk in the critically ill.
Acute kidney injury; Ischemia/reperfusion; Acute lung injury; Apoptosis
Autosomal dominant polycystic kidney disease (ADPKD) is characterized by age-dependent growth of kidney cysts with end-stage renal disease developing in about 50% of affected individuals. Living donors from ADPKD families are at risk for developing ADPKD and may be excluded from renal donation if the diagnosis cannot be conclusively ruled out. Radiographic imaging may be adequate to screen for kidney cysts in most at-risk donors but may fail to identify affected individuals younger than 40 or older individuals from families with mild disease. In this paper we report a strategy that incorporates genetic testing in the evaluation of live kidney donors at risk for ADPKD whose disease status cannot be established with certainty on the basis of imaging studies alone. We show that DNA diagnostics can be used to enhance safe donation for certain living donor candidates at risk for ADPKD.
living donation; polycystic kidney disease; genetic screening; molecular diagnostics; DNA testing
Kidney ischemia reperfusion injury is a major cause of morbidity in both allograft and native kidneys. Ischemia reperfusion-induced acute kidney injury is characterized by early, allo-antigen independent inflammation. Major components of the innate immune system are activated and participate in the pathogenesis of acute kidney injury, plus prime the allograft kidney for rejection. Soluble members of innate immunity implicated in acute kidney injury include the complement system, cytokines, and chemokines. Toll-like receptors (TLRs) are also important contributors. Effector cells that participate in ACUTE KIDNEY INJURY include the classic innate immune cells, neutrophils and macrophages. Recent data has unexpectedly identified lymphocytes as participants of early acute kidney injury responses. In this review, we will focus on immune mediators that participate in the pathogenesis of ischemic acute kidney injury.
Ischemia reperfusion injury; Innate immunity; Inflammation
It is well-established that significant ischemia-reperfusion injury during kidney transplantation results in increased incidence of long-term fibrosis and rejection. To test for a role of T cell infiltration and activation following ischemic injury, we induced both bilateral and unilateral renal ischemia in mice, followed by reperfusion, and then isolated mononuclear cells. Analysis of these cells by flow cytometry showed that 2 weeks after bilateral ischemia there was a significant increase of CD8 + T cells. Furthermore, both CD4 + and CD8 + T cells infiltrated the injured kidney 6 weeks after unilateral ischemia. These T cells had increased expression of CD69 + and CD44hiCD62L −, markers of activation and effector-memory, respectively. CD4 + NK1.1 + and CD19 + B cells were decreased in percentage both 6 and 11 weeks after bilateral or unilateral injury. There was a significant upregulation of IL-1β, IL-6, TNF-α, IFN-γ, MIP-2, and RANTES expression, measured by real-time PCR, 6 weeks after unilateral renal ischemia, further indicating T cell activation. Depletion of CD4 + and CD8 + T cells before ischemia caused less medullary damage and reduced kidney IFN-γ expression, whereas their depletion following ischemia increased kidney IL-1β; however, depletion of these cells had no effect on histological damage to the kidney. Our study demonstrates that moderate or severe kidney ischemia induces long-term T lymphocyte infiltration and cytokine/chemokine upregulation, leading to kidney structural changes.
cell depletion; long-term infiltration; renal ischemia; T lymphocytes
Ischemia reperfusion injury (IRI) is a common and important clinical problem in many different organ systems, including kidney, brain, heart, liver, lung, and intestine. IRI occurs during all deceased donor organ transplants. IRI is a highly complex cascade of events that includes interactions between vascular endothelium, interstitial compartments, circulating cells, and numerous biochemical entities. It is well established that the innate immune system, such as complement, neutrophils, cytokines, chemokines, and macrophages participate in IRI. Recent data demonstrates an important role for lymphocytes, particularly T cells but also B cells in IRI. Lymphocytes not only participate in augmenting injury responses after IRI, but could also be playing a protective role depending on the cell type and stage of injury. Furthermore, lymphocytes appear to be participating in the healing response from IRI. These new data open the possibility for lymphocyte targeted therapeutics to improve the short and long term outcomes from IRI.
Although neurological sequelae of acute kidney injury (AKI) are well established, the pathogenesis of acute uremic encephalopathy is poorly understood. We examined the short-term effect of severe ischemic AKI on mouse brain inflammatory and functional changes. C57BL/6 mice underwent a 60 min of bilateral renal ischemia or a sham operation and the brains were harvested and studied at 24 h after surgery. Compared to sham operated mice, the mice with AKI had significantly increased concentration of C reactive protein and chemokine G-CSF in the blood and increased the numbers of pyknotic neurons and microgliosis in the brain. AKI mice had increased levels of proinflammatory chemokine proteins KC and G-CSF in both cerebral cortex and hippocampus when compared to sham operated mice. There was increased glial fibrillary acidic protein (GFAP) expression in astrocytes in the cortex and corpus callosum after AKI. In contrast, no increased neuronal pyknosis or increased expression of GFAP was found in the brain from mice with ischemic acute liver injury and normal renal function. Mice with AKI also had increased brain Evans blue dye extravasation into the brain suggesting disruption of the blood-brain barrier.. To evaluate the functional effects of different severities of AKI, we studied locomotor activity using an open field test. Mice with either a 45 or 60 min renal ischemia or a bilateral nephrectomy had moderate to severe decline in locomotor activity 24 h after surgery when compared to sham operated mice. This is the first demonstration that severe ischemic AKI induces brain inflammation and ischemic time-dependent functional changes. Targeting these pathways could reduce morbidity and mortality in critically ill patients with severe AKI.
Increased organ ischemia time leads to delayed graft function (DGF), increased acute rejection (AR), enhanced chronic allograft nephropathy (CAN) and reduced long term allograft survival. The mechanisms by which IRI predisposes to AR and CAN are unknown. We hypothesized that gene expression profiling of IRI-affected kidney would identify how IRI predisposes to AR and CAN. Furthermore, we examined how current immunosuppressive drug molecular targets are altered by IRI.
C57BL/6J mice were exposed to 30 (n=3) or 60 (n=3) min of bilateral kidney ischemia or sham surgery (n=5). At 36 hr kidney tissue was collected and analyzed using Affymetrix 430MOEA (22626 genes) array and GC-RMA-SAM pipeline. Genes with the false discovery rate (q<1%) and ±50% fold change (FC) were considered affected by IRI. Genes coding for histocompatibility and antigen presenting factors, calcineurin and mTOR pathway-associated proteins were selected using Gene Ontology (GO) analysis. GO analysis identified 10 and 17 alloimmunity-related genes affected by IRI induced by 30 and 60 min of ischemia, respectively including Traf6 (FC=2.99) and H2-D1 (FC=2.58). We also detected significant IRI genomic responses in calcineurin and mTOR pathways represented by Fkbp5 (FC=4.18) and Fkbp1a (FC=2.0); Eif4ebp1 (FC=16.8) and Akt1 (FC=3.64), respectively.
These data demonstrated that IRI upregulates expression of several alloimmunity-associated genes, which can in turn enhance alloimune responses. Our discovery of IRI-induced upregulation of genes associated with calcineurin and mTOR pathways are consistent with clinical observations that FK506 and Rapamycin can alter course of DGF. Further validation and dissection of these pathways can lead to novel approaches by which improved management of early “non-immune” transplant events can decrease susceptibility to more classic “immune” changes and CAN.
Aberrant promoter hypermethylation, also known as epigenetics, is thought to be a promising biomarker approach to diagnose malignancies. Kidney repair after injury is a recapitulation of normal morphogenesis, with similarities to malignant transformation. We hypothesized that changes in urine epigenetics could be a biomarker approach during early kidney transplant injury and repair. We examined urine DNA for aberrant methylation of 2 gene promoters (DAPK CALCA) by quantitative methylation specific PCR from 13 deceased and 10 living donor kidney transplant recipients on postoperative day 2 and 65 healthy controls. Results were compared with clinical outcomes and to results of the kidney biopsy. Transplant recipients were significantly more likely to have aberrant hypermethylation of the CALCA gene promoter in urine than healthy controls (100% vs. 31%, p<0.0001). There was increased CALCA hypermethylation in the urine of deceased vs. living donor transplants (21.60 +/− 12.5 vs. 12.19 +/− 4.7 P=0.04). Furthermore, there was a trend towards increased aberrant hypermethylation of urine CALCA in patients with biopsy-proven acute tubular necrosis vs. acute rejection and slow or prompt graft function (mean: 20.40 +/− 6.9, 13.87 +/− 6.49, 17.17 +/− 13.4, P=0.67). However, there was no difference of CALCA hypermethylation in urine of patients with delayed graft function vs. those with slow or prompt graft function (16.9 +/− 6.2 vs. 18.5 +/− 13.7, respectively; P=0.5). There was no aberrant hypermethylation of DAPK in the urine of transplant patients. Urine epigenetics is a promising biomarker approach for acute ischemic injury in transplantation that merits future study.
biomarker; hypermethylation; acute kidney injury
Thrombospondin 1 (TSP-1) is a matricellular protein that inhibits angiogenesis and causes apoptosis in vivo and in vitro in several cancerous cells and tissues. Here we identify TSP-1 as the molecule with the highest induction level at 3 hours of IR injury in rat and mouse kidneys subjected to ischemia/reperfusion (IR) injury using the DNA microarray approach. Northern hybridizations demonstrated that TSP-1 expression was undetectable at baseline, induced at 3 and 12 hours, and returned to baseline levels at 48 hours of reperfusion. Immunocytochemical staining identified the injured proximal tubules as the predominant sites of expression of TSP-1 in IR injury and showed colocalization of TSP-1 with activated caspase-3. Addition of purified TSP-1 to normal kidney proximal tubule cells or cells subjected to ATP depletion in vitro induced injury as demonstrated by cytochrome c immunocytochemical staining and caspase-3 activity. The deleterious role of TSP-1 in ischemic kidney injury was demonstrated directly in TSP-1 null mice, which showed significant protection against IR injury–induced renal failure and tubular damage. We propose that TSP-1 is a novel regulator of ischemic damage in the kidney and may play an important role in the pathophysiology of ischemic kidney failure.