ABSTRACT: Renal recovery following injury relies on cellular regeneration. In the mouse kidney following injury, injured epithelial cells undergoes de-differentiate, proliferate and re-differentiate into functional cells, following a a tightly controlled genetic programme where specific sets of genes are up-regulated. We used microarrays to detail the global programme of gene expression underlying cellular regeneration following injury with or without a HDAC inhibitor, m4PTB, treatment and identified distinct classes of up-regulated genes during this process. Male BALB/c mice underwent 26 minute unilateral ischemia-reperfusion (IR) with contralateral nephrectomy treated with vehicle or 100 mg/kg m4PTB 24 h after inducing renal injury, and kidneys harvested for RNA extraction 12 h later. We sought to obtain kidneys within a similar degree of injury in order to carefully evaluate the effect of m4PTB on expression profiles. To that end, we selected kidneys according to three different criteria of injury: (1) level of serum creatinine at Day1, (2) the level of blood urea nitrogen (BUN) (3) Kim1 expression by qPCR at Day1.5 using haevested kidneys
Project description:Renal recovery following injury relies on cellular regeneration. In the mouse kidney following injury, injured epithelial cells undergoes de-differentiate, proliferate and re-differentiate into functional cells, following a a tightly controlled genetic programme where specific sets of genes are up-regulated. We used microarrays to detail the global programme of gene expression underlying cellular regeneration following injury with or without a HDAC inhibitor, m4PTB, treatment and identified distinct classes of up-regulated genes during this process.
Project description:8 week-old male C57BL6J mice were given Gram-negative endotoxin (LPS O111:B4, 10 mg/kg) intraperitoneally at time 0. 18 hrs thereafter, they were administered 10 ml/kg 0.9% saline. Mice were sacrificed at 0, 18, or 42 hrs after LPS challenge. Kidneys were immediately collected into TRIzol for RNA preparation. Renal function was measured on blood collected at the time of tissue harvest At t=0hr, mice had normal baseline renal function. At t=18hr, mice exhibited early renal injury, At t=42hr, mice had either recovered normal renal function or had persistent renal injury. We collected kidneys from 3 mice per time point. For the 42 hr time point, we collected kidneys from 3 mice with recovered renal function and kidneys from 3 mice with persistent renal injury. Mouse kidneys selected at successive stages of renal injury and recovery following systemic LPS challenge and volume resuscitation following LPS challenge.
Project description:Previous studies have suggested that CD133+ cells isolated from human kidney biopsies have the potential to ameliorate injury following intravenous (IV) administration in rodent models of kidney disease by integrating into damaged renal tissue and generating specialised renal cells. However, whether renal engraftment of CD133+ cells is prerequisite for ameliorating injury has not yet been unequivocally resolved. Here, we have established a cisplatin-induced nephropathy model in immunodeficient rats to assess the efficacy of CD133+ human kidney cells in restoring renal health, and to determine the fate of these cells after systemic administration. Specifically, following IV administration, we evaluated the impact of the CD133+ cells on renal function by undertaking longitudinal measurements of the glomerular filtration rate using a novel transcutaneous device. Furthermore, using histological assays, we assessed whether the human kidney cells could promote renal regeneration, and if this was related to their ability to integrate into the damaged kidneys. Our results show that both CD133+ and CD133- cells improve renal function and promote renal regeneration to a similar degree. However, this was not associated with engraftment of the cells into the kidneys. Instead, after IV administration, both cell types were exclusively located in the lungs, and had disappeared by 24 hours. Our data therefore indicate that renal repair is not mediated by CD133+ cells homing to the kidneys and generating specialised renal cells. Instead, renal repair is likely to be mediated by paracrine or endocrine factors.
Project description:Introduction: Renal ischemia-reperfusion (IR) causes acute kidney injury (AKI) with high mortality and morbidity. The objective of this study was to ameliorate kidney IR injury and identify novel biomarkers for kidney injury and repair. Methods: Left renal ischemia was induced in rats by clamping renal artery for 45 minutes, followed by reperfusion and right nephrectomy. Thirty minutes prior to ischemia, rats (n=8/group) received Valproic Acid (150 mg/kg; VPA), Dexamethasone (3 mg/kg; Dex) or Vehicle (Saline) intraperitoneally. Animals were sacrificed at 3h, 24h or 120h post- IR and blood, urine and kidney were collected. Results: Serum creatinine (mg/dL) at 24 h IR in VPA (2.7±1.8) and Dex (2.3±1.2) was reduced (P<0.05) compared to Vehicle (3.8±0.5). At 3h post-IR, urine albumin (mg/ml) was higher in Vehicle (1.47±0.10), VPA (0.84±0.62) and Dex (1.04±0.73) compared to uninjured/untreated control (0.14±0.26) group. At 24h post-IR urine Lipocalin-2 (µg/ml) was significantly higher (P<0.05) in VPA, Dex and Vehicle groups (9.61-11.36) compared to uninjured/untreated control (0.67±o.29); also, Kidney Injury Molecule-1 (KIM-1; ng/ml) was significantly higher in VPA, Dex and Vehicle groups (13.7-18.7) compared uninjured/untreated control (1.7±1.9). KIM-1 levels were significantly (P<0.05) higher in all groups compared to uninjured/untreated control levels. Histopathology at 3h post IR demonstrated (P<0.05) reduction in ischemic injury in the renal cortex in VPA (Grade 1.6± 1.5) compared to Vehicle (Grade 2.9±1.1) group. Inflammatory cytokines IL1β and IL6 were down-regulated in VPA and Dex groups. BCL2 was higher in VPA group. DNA microarray analysis demonstrated reduced stress response and injury, and improved recovery related gene expression in the kidneys of VPA treated animals. Conclusions: VPA administration reduced kidney IR injury and improved regeneration. KIM-1 and Lipocalin-2 appear to be promising early urine biomarkers of acute ischemic kidney injury. We had three experimental groups. Group A, VPA treatment; Group B, Dexamethasone treatment; and Group C, No treatment (vehicle saline control). Treatments were administered prior to the induction of left renal ischemia. Animals underwent 45 minutes of left renal ischemia, followed by reperfusion, and right nephrectomy as described above. Following reperfusion, animals were sacrificed at 3, 24 or 120 hours (n=8/group). In the 3 hour group, rats were maintained under anesthesia after surgery until sacrifice. In the 24 and 120 hour groups, the rats were recovered and returned to the cages for normal housing. Analgesic buprenorphine (0.05mg/kg) was administered every 12 h for three days post-operatively. After animal sacrifice, urine, blood, and kidney were collected for kidney functional biomarker assays, histology and / or molecular analyses. Urine was obtained via cystocentesis and blood was obtained via the left renal vein. Tissue and urine samples collected from normal (naïve) animals (n=5) were used for baseline measurements. A subset of 46 animals (n = 4-5 per group) were selected for microarray analysis.
Project description:Congenital obstructive nephropathy is a common cause of chronic kidney disease and a leading indication for renal transplant in children. The cellular and molecular responses of the kidney to congenital obstruction are incompletely characterized. In this study, we evaluated global transcription in kidneys with graded hydronephrosis in the megabladder (mgb-/-) mouse to better understand the pathophysiology of congenital obstructive nephropathy. Three primary pathways associated with kidney remodeling/repair were induced in mgb-/- kidneys independent of the degree of hydronephrosis. These pathways included retinoid signaling, steroid hormone metabolism, and renal response to injury. Urothelial proliferation and the expression of genes with roles in the integrity and maintenance of the renal urothelium were selectively increased in mgb-/- kidneys. Ngal/Lcn2, a marker of acute kidney injury, was elevated in 36% of kidneys with higher grades of hydronephrosis. Evaluation of Ngalhigh versus Ngallow kidneys identified the expression of several novel candidate markers of renal injury. This study indicates that the development of progressive hydronephrosis in mgb-/- mice results in renal adaptation that includes significant changes in the morphology and potential functionality of the renal urothelium. These observations will permit the development of novel biomarkers and therapeutic approaches to progressive renal injury in the context of congenital obstruction. Gene expression was measured in control, mild, moderate and severely hydronephrotic megabladder mouse kidneys. A total of 6 control kidneys were compared to 18 mutant kidneys from age-matched male animals.
Project description:8 week-old male C57BL6J mice were given Gram-negative endotoxin (LPS O111:B4, 10 mg/kg) intraperitoneally at time 0. 18 hrs thereafter, they were administered 10 ml/kg 0.9% saline. Mice were sacrificed at 0, 18, or 42 hrs after LPS challenge. Kidneys were immediately collected into TRIzol for RNA preparation. Renal function was measured on blood collected at the time of tissue harvest At t=0hr, mice had normal baseline renal function. At t=18hr, mice exhibited early renal injury, At t=42hr, mice had either recovered normal renal function or had persistent renal injury. We collected kidneys from 3 mice per time point. For the 42 hr time point, we collected kidneys from 3 mice with recovered renal function and kidneys from 3 mice with persistent renal injury.
Project description:Despite the recent advances in our understanding of the role of lipids, metabolites and related enzymes in mediating kidney injury, there is limited integrated multi-omics data identifying potential metabolic pathways driving human kidney damage (KD). The limited availability of kidney biopsies from living donors with kidney disease has remained a major constraint. Here, we validated the use of deceased transplant donor kidneys as a good model to study kidney disease in humans and characterized these kidneys using imaging and multi-omics approaches. We demonstrated that changes in kidney injury and inflammatory markers following KD were consistent with the changes in pre-donation renal function in donors. Neighborhood and correlation analyses of imaging mass cytometry data showed that a subset of renal cells (e.g., fibroblasts) are associated with the expression profile of renal immune cells, potentially linking these cells to kidney inflammation. Integrated transcriptomic and metabolomic analysis of human kidneys showed that renal arachidonic acid metabolism and seven other metabolic pathways were upregulated following KD. To validate the therapeutic potential of targeting the arachidonic acid pathway, we demonstrated increased levels of cytosolic phospholipase A2 (cPLA2) protein and related lipid mediators (e.g., prostaglandin E2) in the injured kidneys. The inhibition of cPLA2 reduced injury and inflammation in human renal proximal tubular epithelial cells (RPTEC) in vitro. This study identifies cell types and metabolic pathways that may be critical for controlling inflammation associated with KD in humans.
Project description:We sequenced the transcriptome of gentamicin induced renal regeneration in adult zebrafish. Specifically, zebrafish kidney tissues in the first, third, fifth and seventh days of kidney injury and the control group were selected. Each sample contains three kidneys. Three samples were taken in each period, and then the total RNA of the kidney was extracted for second-generation sequencing.
Project description:<p>Acute kidney injury (AKI) is a known risk factor for the development of chronic kidney disease (CKD), with no satisfactory strategy to prevent the progression of AKI to CKD. Damage to the renal vascular system and subsequent hypoxia are common contributors to both AKI and CKD. Hypoxia inducible factor (HIF) is reported to protect the kidney from acute ischemic damage and a novel HIF stabilizer, FG4592 (Roxadustat), has become available in the clinic as an anti-anemia drug. However, the role of FG4592 in the AKI-to-CKD transition remains elusive. In the present study, we investigated the role of FG4592 in the AKI-to-CKD transition induced by unilateral kidney ischemia-reperfusion (UIR). The results showed that FG4592, given to mice 3 days after UIR, markedly alleviated kidney fibrosis and enhanced renal vascular regeneration, possibly via activating the HIF-1α/vascular endothelial growth factor A (VEGFA)/VEGF receptor 1 (VEGFR1) signaling pathway and driving the expression of the endogenous antioxidant superoxide dismutase 2 (SOD2). In accordance with the improved renal vascular regeneration and redox balance, the metabolic disorders of the UIR mice kidneys were also attenuated by treatment with FG4592. However, the inflammatory response in the UIR kidneys was not affected significantly by FG-4592. Importantly, in the kidneys of CKD patients, we also observed enhanced HIF-1α expression which was positively correlated with the renal levels of VEGFA and SOD2. Together, these findings demonstrated the therapeutic effect of the anti-anemia drug FG-4592 in preventing the AKI-to-CKD transition related to ischemia and the redox imbalance.</p><p><br></p><p>Linked study:</p><p><strong>UPLC-MS assay</strong> of mice kidney tissue sacrificed at<strong> day 10 </strong>after UIR is reported in <a href='https://www.ebi.ac.uk/metabolights/MTBLS3056' rel='noopener noreferrer' target='_blank'>MTBLS3056</a></p>
Project description:<p>Acute kidney injury (AKI) is a known risk factor for the development of chronic kidney disease (CKD), with no satisfactory strategy to prevent the progression of AKI to CKD. Damage to the renal vascular system and subsequent hypoxia are common contributors to both AKI and CKD. Hypoxia inducible factor (HIF) is reported to protect the kidney from acute ischemic damage and a novel HIF stabilizer, FG4592 (Roxadustat), has become available in the clinic as an anti-anemia drug. However, the role of FG4592 in the AKI-to-CKD transition remains elusive. In the present study, we investigated the role of FG4592 in the AKI-to-CKD transition induced by unilateral kidney ischemia-reperfusion (UIR). The results showed that FG4592, given to mice 3 days after UIR, markedly alleviated kidney fibrosis and enhanced renal vascular regeneration, possibly via activating the HIF-1α/vascular endothelial growth factor A (VEGFA)/VEGF receptor 1 (VEGFR1) signaling pathway and driving the expression of the endogenous antioxidant superoxide dismutase 2 (SOD2). In accordance with the improved renal vascular regeneration and redox balance, the metabolic disorders of the UIR mice kidneys were also attenuated by treatment with FG4592. However, the inflammatory response in the UIR kidneys was not affected significantly by FG-4592. Importantly, in the kidneys of CKD patients, we also observed enhanced HIF-1α expression which was positively correlated with the renal levels of VEGFA and SOD2. Together, these findings demonstrated the therapeutic effect of the anti-anemia drug FG-4592 in preventing the AKI-to-CKD transition related to ischemia and the redox imbalance.</p><p><br></p><p>Linked study:</p><p><strong>UPLC-MS assay</strong> of mice kidney tissues sacrificed at <strong>day 21 </strong>after UIR is reported in <a href='https://www.ebi.ac.uk/metabolights/MTBLS3003' rel='noopener noreferrer' target='_blank'>MTBLS3003</a></p>