Identification of cell division cycle protein 20 in various forms of acute and chronic kidney injury in mice.

Tubular epithelial cell fate following exposure to various types of injurious stimuli can be decided at distinct cell cycle checkpoints. One such checkpoint occurs during mitosis, known as the spindle assembly checkpoint, and is tightly regulated through the actions of cell division cycle protein 20 (CDC20). Due to our paucity of knowledge about the role of CDC20 in the kidney, the present study was designed to investigate the expression levels and distribution of CDC20 within the kidney and how pharmacological inhibition of CDC20 function affects kidney recovery using various rodent models of kidney injury. CDC20 is normally detected in distal tubules, but upon injury by either cisplatin administration or ureter obstruction, CDC20 accumulation is considerably elevated. Blockade of CDC20 activity using a selective pharmacological inhibitor, Apcin, lowered serum creatinine, tubular damage, and DNA injury following acute kidney injury compared with vehicle-treated mice. In unilateral ureteral obstruction, Apcin reduced tissue kidney injury molecule-1 levels, sirius red staining, and tubulointerstitial α-smooth muscle actin staining in the tissue. The findings in the present study demonstrated that elevations in CDC20 levels in the kidney are associated with kidney injury and that inhibition of CDC20 can alleviate and reverse some of the pathological effects on the architecture and function of kidney.NEW & NOTEWORTHY To our knowledge, this is the first study to characterize the expression and localization of cell division cycle 20 protein (CDC20) in normal and acute, and chronically injured kidneys. Tubular epithelial cell damage was markedly reduced through the administration of a selective inhibitor of CDC20, Apcin. This study provides new evidence that CDC20 can be induced in damaged kidney cells and negatively impact the recovery of the kidney following acute kidney injury.

Incidence and outcomes of acute kidney injury including hepatorenal syndrome in hospitalized patients with cirrhosis in the US.

BACKGROUND & AIMS: Acute kidney injury (AKI) in cirrhosis is common and associated with high morbidity, but the incidence rates of different etiologies of AKI are not well described in the US. We compared incidence rates, practice patterns, and outcomes across etiologies of AKI in cirrhosis. METHODS: We performed a retrospective cohort study of 11 hospital networks, including consecutive adult patients admitted with AKI and cirrhosis in 2019. The etiology of AKI was adjudicated based on pre-specified clinical definitions (prerenal/hypovolemic AKI, hepatorenal syndrome [HRS-AKI], acute tubular necrosis [ATN], other). RESULTS: A total of 2,063 patients were included (median age 62 [IQR 54-69] years, 38.3% female, median MELD-Na score 26 [19-31]). The most common etiology was prerenal AKI (44.3%), followed by ATN (30.4%) and HRS-AKI (12.1%); 6.0% had other AKI, and 7.2% could not be classified. In our cohort, 8.1% of patients received a liver transplant and 36.5% died by 90 days. The lowest rate of death was observed in patients with prerenal AKI (22.2%; p <0.001), while death rates were higher but not significantly different from each other in those with HRS-AKI and ATN (49.0% vs. 52.7%; p = 0.42). Using prerenal AKI as a reference, the adjusted subdistribution hazard ratio (sHR) for 90-day mortality was higher for HRS-AKI (sHR 2.78; 95% CI 2.18-3.54; p <0.001) and ATN (sHR 2.83; 95% CI 2.36-3.41; p <0.001). In adjusted analysis, higher AKI stage and lack of complete response to treatment were associated with an increased risk of 90-day mortality (p <0.001 for all). CONCLUSION: AKI is a severe complication of cirrhosis. HRS-AKI is uncommon and is associated with similar outcomes to ATN. The etiology of AKI, AKI stage/severity, and non-response to treatment were associated with mortality. Further optimization of vasoconstrictors for HRS-AKI and supportive therapies for ATN are needed. IMPACT AND IMPLICATIONS: Acute kidney injury (AKI) in cirrhosis carries high morbidity, and management is determined by the etiology of injury. However, a large and well-adjudicated multicenter database from US centers that uses updated AKI definitions is lacking. Our findings demonstrate that acute tubular necrosis and hepatorenal syndrome have similar outcomes (∼50% mortality at 90 days), though hepatorenal syndrome is uncommon (12% of all AKI cases). These findings represent practice patterns at US transplant/tertiary centers and can be used as a baseline, presenting the situation prior to the adoption of terlipressin in the US.

Histologically resolved small RNA maps in primary focal segmental glomerulosclerosis indicate progressive changes within glomerular and tubulointerstitial regions.

Pathological heterogeneity is common in clinical tissue specimens and complicates the interpretation of molecular data obtained from the specimen. As a typical example, a kidney biopsy specimen often contains glomeruli and tubulointerstitial regions with different levels of histological injury, including some that are histologically normal. We reasoned that the molecular profiles of kidney tissue regions with specific histological injury scores could provide new insights into kidney injury progression. Therefore, we developed a strategy to perform small RNA deep sequencing analysis for individually scored glomerular and tubulointerstitial regions in formalin-fixed, paraffin-embedded kidney needle biopsies. This approach was applied to study focal segmental glomerulosclerosis (FSGS), the leading cause of nephrotic syndrome in adults. Large numbers of small RNAs, including microRNAs, 3'-tRFs, 5'-tRFs, and mitochondrial tRFs, were differentially expressed between histologically indistinguishable tissue regions from patients with FSGS and matched healthy controls. A majority of tRFs were upregulated in FSGS. Several small RNAs were differentially expressed between tissue regions with different histological scores in FSGS. Notably, with increasing levels of histological damage, miR-21-5p was upregulated progressively and miR-192-5p was downregulated progressively in glomerular and tubulointerstitial regions, respectively. This study marks the first genome scale molecular profiling conducted in histologically characterized glomerular and tubulointerstitial regions. Thus, substantial molecular changes in histologically normal kidney regions in FSGS might contribute to initiating tissue injury or represent compensatory mechanisms. In addition, several small RNAs might contribute to subsequent progression of glomerular and tubulointerstitial injury, and histologically mapping small RNA profiles may be applied to analyze tissue specimens in any disease.

Terlipressin and the Treatment of Hepatorenal Syndrome: How the CONFIRM Trial Moves the Story Forward.

Hepatorenal syndrome (HRS) is a form of acute kidney injury (AKI) occurring in patients with advanced cirrhosis and is associated with significant morbidity and mortality. The pathophysiology underlying HRS begins with increasing portal pressures leading to the release of vasodilatory substances that result in pooling blood in the splanchnic system and a corresponding reduction in effective circulating volume. Compensatory activation of the sympathetic nervous system and the renin-angiotensin-aldosterone system and release of arginine vasopressin serve to defend mean arterial pressure but at the cost of severe constriction of the renal vasculature, leading to a progressive, often fulminant form of AKI. There are no approved treatments for HRS in the United States, but multiple countries, including much of Europe, use terlipressin, a synthetic vasopressin analogue, as a first-line therapy. CONFIRM (A Multi-Center, Randomized, Placebo Controlled, Double-Blind Study to Confirm Efficacy and Safety of Terlipressin in Subjects With Hepatorenal Syndrome Type 1), the third randomized trial based in North America evaluating terlipressin, met its primary end point of showing greater rates of HRS reversal in the terlipressin arm. However, due to concerns about the apparent increased rates of respiratory adverse events and a lack of evidence for mortality benefit, terlipressin was not approved by the Food and Drug Administration (FDA). We explore the history of regulatory approval for terlipressin in the United States, examine the results from CONFIRM and the concerns they raised, and consider the future role of terlipressin in this critical clinical area of continued unmet need.

Endogenous miR-204 Protects the Kidney against Chronic Injury in Hypertension and Diabetes.

BACKGROUND: Aberrant microRNA (miRNA) expression affects biologic processes and downstream genes that are crucial to CKD initiation or progression. The miRNA miR-204-5p is highly expressed in the kidney but whether miR-204-5p plays any role in the development of chronic renal injury is unknown. METHODS: We used real-time PCR to determine levels of miR-204 in human kidney biopsies and animal models. We generated Mir204 knockout mice and used locked nucleic acid-modified anti-miR to knock down miR-204-5p in mice and rats. We used a number of physiologic, histologic, and molecular techniques to analyze the potential role of miR-204-5p in three models of renal injury. RESULTS: Kidneys of patients with hypertension, hypertensive nephrosclerosis, or diabetic nephropathy exhibited a significant decrease in miR-204-5p compared with controls. Dahl salt-sensitive rats displayed lower levels of renal miR-204-5p compared with partially protected congenic SS.13BN26 rats. Administering anti-miR-204-5p to SS.13BN26 rats exacerbated interlobular artery thickening and renal interstitial fibrosis. In a mouse model of hypertensive renal injury induced by uninephrectomy, angiotensin II, and a high-salt diet, Mir204 gene knockout significantly exacerbated albuminuria, renal interstitial fibrosis, and interlobular artery thickening, despite attenuation of hypertension. In diabetic db/db mice, administering anti-miR-204-5p exacerbated albuminuria and cortical fibrosis without influencing blood glucose levels. In all three models, inhibiting miR-204-5p or deleting Mir204 led to upregulation of protein tyrosine phosphatase SHP2, a target gene of miR-204-5p, and increased phosphorylation of signal transducer and activator of transcription 3, or STAT3, which is an injury-promoting effector of SHP2. CONCLUSIONS: These findings indicate that the highly expressed miR-204-5p plays a prominent role in safeguarding the kidneys against common causes of chronic renal injury.

MicroRNA-214-3p in the Kidney Contributes to the Development of Hypertension.

BACKGROUND: In spite of extensive study, the mechanisms for salt sensitivity of BP in humans and rodent models remain poorly understood. Several microRNAs (miRNAs) have been associated with hypertension, but few have been shown to contribute to its development. METHODS: We examined miRNA expression profiles in human kidney biopsy samples and rat models using small RNA deep sequencing. To inhibit an miRNA specifically in the kidney in conscious, freely moving rats, we placed indwelling catheters to allow both renal interstitial administration of a specific anti-miR and measurement of BP. A rat with heterozygous disruption of the gene encoding endothelial nitric oxide synthase (eNOS) was developed. We used bioinformatic analysis to evaluate the relationship between 283 BP-associated human single-nucleotide polymorphisms (SNPs) and 1870 human miRNA precursors, as well as other molecular and cellular methods. RESULTS: Compared with salt-insensitive SS.13BN26 rats, Dahl salt-sensitive (SS) rats showed an upregulation of miR-214-3p, encoded by a gene in the SS.13BN26 congenic region. Kidney-specific inhibition of miR-214-3p significantly attenuated salt-induced hypertension and albuminuria in SS rats. miR-214-3p directly targeted eNOS. The effect of miR-214-3p inhibition on hypertension and albuminuria was abrogated in SS rats with heterozygous loss of eNOS. Human kidney biopsy specimens from patients with hypertension or hypertensive nephrosclerosis showed upregulation of miR-214-3p; the gene encoding miR-214-3p was one of several differentially expressed miRNA genes located in proximity to human BP-associated SNPs. CONCLUSIONS: Renal miR-214-3p plays a functional and potentially genetic role in the development of hypertension, which might be mediated in part by targeting eNOS.

Artificial intelligence, physiological genomics, and precision medicine.

Big data are a major driver in the development of precision medicine. Efficient analysis methods are needed to transform big data into clinically-actionable knowledge. To accomplish this, many researchers are turning toward machine learning (ML), an approach of artificial intelligence (AI) that utilizes modern algorithms to give computers the ability to learn. Much of the effort to advance ML for precision medicine has been focused on the development and implementation of algorithms and the generation of ever larger quantities of genomic sequence data and electronic health records. However, relevance and accuracy of the data are as important as quantity of data in the advancement of ML for precision medicine. For common diseases, physiological genomic readouts in disease-applicable tissues may be an effective surrogate to measure the effect of genetic and environmental factors and their interactions that underlie disease development and progression. Disease-applicable tissue may be difficult to obtain, but there are important exceptions such as kidney needle biopsy specimens. As AI continues to advance, new analytical approaches, including those that go beyond data correlation, need to be developed and ethical issues of AI need to be addressed. Physiological genomic readouts in disease-relevant tissues, combined with advanced AI, can be a powerful approach for precision medicine for common diseases.

Tissue-Specific MicroRNA Expression Patterns in Four Types of Kidney Disease.

MicroRNAs contribute to the development of kidney disease. Previous analyses of microRNA expression in human kidneys, however, were limited by tissue heterogeneity or the inclusion of only one pathologic type. In this study, we used laser-capture microdissection to obtain glomeruli and proximal tubules from 98 human needle kidney biopsy specimens for microRNA expression analysis using deep sequencing. We analyzed specimens from patients with diabetic nephropathy (DN), FSGS, IgA nephropathy (IgAN), membranoproliferative GN (MPGN) (n=19-23 for each disease), and a control group (n=14). Compared with control glomeruli, DN, FSGS, IgAN, and MPGN glomeruli exhibited differential expression of 18, 12, two, and 17 known microRNAs, respectively. The expression of several microRNAs also differed between disease conditions. Specifically, compared with control or FSGS glomeruli, IgAN glomeruli exhibited downregulated expression of hsa-miR-3182. Furthermore, in combination, the expression levels of hsa-miR-146a-5p and hsa-miR-30a-5p distinguished DN from all other conditions except IgAN. Compared with control proximal tubules, DN, FSGS, IgAN, and MPGN proximal tubules had differential expression of 13, 14, eight, and eight microRNAs, respectively, but expression of microRNAs did not differ significantly between the disease conditions. The abundance of several microRNAs correlated with indexes of renal function. Finally, we validated the differential glomerular expression of select microRNAs in a second cohort of patients with DN (n=19) and FSGS (n=21). In conclusion, we identified tissue-specific microRNA expression patterns associated with several kidney pathologies. The identified microRNAs could be developed as biomarkers of kidney diseases and might be involved in disease mechanisms.

Deficiency in the Formation of 20-Hydroxyeicosatetraenoic Acid Enhances Renal Ischemia-Reperfusion Injury.

Ischemia-reperfusion (IR) injury is the most common cause of AKI. The susceptibility to develop AKI varies widely among patients. However, little is known about the genes involved. 20-Hydroxyeicosatetraenoic acid (20-HETE) has an important role in the regulation of renal tubular and vascular function and has been implicated in IR injury. In this study, we examined whether a deficiency in the renal formation of 20-HETE enhances the susceptibility of Dahl salt-sensitive (SS) rats to ischemic AKI. Transfer of chromosome 5 containing the CYP4A genes responsible for the formation of 20-HETE from the Brown Norway (BN) rat onto the SS genetic background increased renal 20-HETE levels after ischemia and reduced plasma creatinine levels (±SEM) 24 hours after IR from 3.7±0.1 to 2.0±0.2 mg/dl in an SS.5(BN)-consomic strain. Transfer of this chromosome also prevented the secondary decline in medullary blood flow and ischemia that develops 2 hours after IR in the susceptible SS strain. Blockade of the synthesis of 20-HETE with HET0016 reversed the renoprotective effects in SS.5(BN) rats. Similar results were observed in an SS.5(Lew)-congenic strain, in which a smaller region of chromosome 5 containing the CYP4A genes from a Lewis rat was introgressed onto the SS genetic background. These results indicate that 20-HETE has a protective role in renal IR injury by maintaining medullary blood flow and that a genetic deficiency in the formation of 20-HETE increases the susceptibility of SS rats to ischemic AKI.

Genetic susceptibility and loss of Nr4a1 enhances macrophage-mediated renal injury in CKD.

Nuclear hormone receptors of the NR4A subgroup have been implicated in cancer, atherosclerosis, and metabolic disease. However, little is known about the role of these receptors in kidney health or disease. Nr4a1-deficient rats (Nr4a1(-/-)) developed on a genetic background susceptible to kidney injury (fawn-hooded hypertensive rat [FHH]) were evaluated for BP, proteinuria, renal function, and metabolic parameters from 4 to 24 weeks-of-age. By week 24, Nr4a1(-/-) rats exhibited significantly higher proteinuria (approximately 4-fold) and decreased GFR compared with FHH controls. The severity of tubular atrophy, tubular casts, and interstitial fibrosis increased significantly in Nr4a1(-/-) rats and was accompanied by a large increase in immune cell infiltration, predominantly macrophages and to a lesser extent T cells and B cells. Global transcriptome and network analyses at weeks 8, 16, and 24 identified several proinflammatory genes and pathways differentially regulated between strains. Bone marrow crosstransplantation studies demonstrated that kidney injury in Nr4a1(-/-) rats was almost completely rescued by bone marrow transplanted from FHH controls. In vitro, macrophages isolated from Nr4a1(-/-) rats demonstrated increased immune activation compared with FHH-derived macrophages. In summary, the loss of Nr4a1 in immune cells appears to cause the increased kidney injury and reduced renal function observed in the Nr4a1(-/-) model.

G-protein ßγ subunit dimers modulate kidney repair after ischemia-reperfusion injury in rats.

Heterotrimeric G-proteins play a crucial role in the control of renal epithelial cell function during homeostasis and in response to injury. In this report, G-protein ßγ subunit (Gßγ) dimer activity was evaluated during the process of tubular repair after renal ischemia-reperfusion injury (IRI) in male Sprague Dawley rats. Rats were treated with a small molecule inhibitor of Gßγ activity, gallein (30 or 100 mg/kg), 1 hour after reperfusion and every 24 hours for 3 additional days. After IRI, renal dysfunction was prolonged after the high-dose gallein treatment in comparison with vehicle treatment during the 7-day recovery period. Renal tubular repair in the outer medulla 7 days after IRI was significantly (P < 0.001) attenuated after treatment with high-dose gallein (100 mg/kg) in comparison with low-dose gallein (30 mg/kg), or the vehicle and fluorescein control groups. Gallein treatment significantly reduced (P < 0.05) the number of proliferating cell nuclear antigen-positive tubular epithelial cells at 24 hours after the ischemia-reperfusion phase in vivo. In vitro application of gallein on normal rat kidney (NRK-52E) proximal tubule cells significantly reduced (P < 0.05) S-phase cell cycle entry compared with vehicle-treated cells as determined by 5'-bromo-2'-deoxyuridine incorporation. Taken together, these data suggest that Gßγ signaling contributes to the maintenance and repair of renal tubular epithelium and may be a novel therapeutic target for the development of drugs to treat acute kidney injury.

De novo expression of sodium-glucose cotransporter SGLT2 in Bowman's capsule coincides with replacement of parietal epithelial cell layer with proximal tubule-like epithelium.

In kidney nephron, parietal epithelial cells line the Bowman's capsule and function as a permeability barrier for the glomerular filtrate. Bowman's capsule cells with proximal tubule epithelial morphology have been found. However, the effects of tubular metaplasia in Bowman's capsule on kidney function remain poorly understood. Sodium-glucose cotransporter 2 (SGLT2) plays a major role in reabsorption of glucose in the kidney and is expressed on brush border membrane (BBM) of epithelial cells in the early segment of the proximal tubule. We hypothesized that SGLT2 is expressed in tubularized Bowman's capsule and used our novel antibody to test this hypothesis. Immunohistochemical analysis was performed with our SGLT2 antibody on C57BL/6 mouse kidney prone to have tubularized Bowman's capsules. Cell membrane was examined with periodic acid-Schiff (PAS) stain. The results showed that SGLT2 was localized on BBM of the proximal tubules in young and adult mice. Bowman's capsules were lined mostly with normal brush border-less parietal epithelial cells in young mice, while they were almost completely covered with proximal tubule-like cells in adult mice. Regardless of age, SGLT2 was expressed on BBM of the tubularized Bowman's capsule but did not co-localize with nephrin in the glomerulus. SGLT2-expressing tubular cells expanded from the urinary pole toward the vascular pole of the Bowman's capsule. This study identified the localization of SGLT2 in the Bowman's capsule. Bowman's capsules with tubular metaplasia may acquire roles in reabsorption of filtered glucose and sodium.

Phenytoin removal by continuous venovenous hemofiltration.

OBJECTIVE: To describe 2 cases of clinically significant phenytoin removal during continuous venovenous hemofiltration (CVVH) and review the relevant literature regarding phenytoin removal by renal replacement modalities. CASE SUMMARY: A 64-year-old female with chronic kidney disease and cirrhosis was admitted to the intensive care unit (ICU) with a traumatic subdural hematoma and seizures. The patient received a loading dose of intravenous phenytoin 1000 mg, followed by maintenance intravenous administration of phenytoin 100 mg and levetiracetam 250 mg every 12 hours. CVVH was initiated for acidosis. A 63-year-old male was admitted to the ICU after cardiac surgery complicated by hypotension. CVVH was initiated for fluid overload, and phenytoin was initiated 3 days later for seizures. A loading dose of intravenous phenytoin 2700 mg was administered, followed by maintenance dosing of intravenous phenytoin 150 mg every 8 hours. Concentrations of unbound phenytoin in serum and CVVH effluent samples were measured during concomitant treatment in each patient. In both patients, serum and effluent concentrations of unbound phenytoin fell steadily while they were on CVVH. Clearance of phenytoin by CVVH was calculated, as was the daily removal of phenytoin, as a percentage of total daily phenytoin dosage during each sampling period. Phenytoin clearance by CVVH ranged from 11 to 13 mL/min in these patients. DISCUSSION: The clearance of phenytoin with CVVH in these 2 patients was much higher than the renal clearance of phenytoin reported in healthy volunteers with normal renal function. Previous case reports have demonstrated that only small, clinically insignificant amounts of phenytoin are removed by hemodialysis, and the only published report of phenytoin removal by continuous renal replacement therapy used hemofiltration rates much lower than those used in the 2 cases described here. CONCLUSIONS: These cases demonstrate that a substantial amount-approximately 30%-of total daily phenytoin dose may be removed by CVVH, and patients may require higher than expected empiric doses. Phenytoin concentrations should be closely monitored in critically ill patients receiving CVVH.

Update on Assessment of Estimated Glomerular Filtration Rate in Patients With Cirrhosis.

Kidney disease is associated with adverse outcomes in patients with cirrhosis including increased post-liver transplantation (LT) mortality. Therefore, diagnosis and staging of kidney disease are critical to timely implementation of treatment and have important implications for transplant eligibility. Serum creatinine (sCr) is a key component of the Model for End-Stage Liver Disease score in LT candidates, and sCr-based estimated glomerular filtration rate (eGFR) values play an important role in determining medical urgency for LT. However, the use of sCr to assess kidney function may be limited in the cirrhotic milieu due to decreased creatinine production, interference of bilirubin with some laboratory assays for sCr, and expansion of the volume of distribution of creatinine. Therefore, conventional eGFR equations perform poorly in patients with cirrhosis and may overestimate kidney function leading to delayed diagnosis of acute kidney injury or lower priority for LT in patients with a truly low glomerular filtration rate. In this review, we will provide an update on the use of sCr for diagnosis and staging of kidney disease in patients with cirrhosis, discuss the limitations of sCr-based eGFR equations, and discuss novel eGFR equations that have been developed in patients with cirrhosis.

Increased susceptibility to kidney injury by transfer of genomic segment from SHR onto Dahl S genetic background.

The Dahl salt-sensitive (S) rat is a widely studied model of salt-sensitive hypertension and develops proteinuria, glomerulosclerosis, and renal interstitial fibrosis. An earlier genetic analysis using a population derived from the S and spontaneously hypertensive rat (SHR) identified eight genomic regions linked to renal injury in the S rat and one protective locus on chromosome 11. The "protective" locus in the S rat was replaced with the SHR genomic segment conferring "susceptibility" to kidney injury. The progression of kidney injury in the S.SHR(11) congenic strain was characterized in the present study. Groups of S and S.SHR(11) rats were followed for 12 wk on either a low-salt (0.3% NaCl) or high-salt (2% NaCl) diet. By week 12 (low-salt), S.SHR(11) demonstrated a significant decline in kidney function compared with the S. Blood pressure was significantly elevated in both strains on high salt. Despite similar blood pressure, the S.SHR(11) exhibited a more significant decline in kidney function compared with the S. The decline in S.SHR(11) kidney function was associated with more severe kidney injury including tubular loss, immune cell infiltration, and tubulointerstitial fibrosis compared with the S. Most prominently, the S.SHR(11) exhibited a high degree of medullary fibrosis and a significant increase in renal vascular medial hypertrophy. In summary, genetic modification of the S rat generated a model of accelerated renal disease that may provide a better system to study progression to renal failure as well as lead to the identification of genetic variants involved in kidney injury.

Role of medullary blood flow in the pathogenesis of renal ischemia-reperfusion injury.

PURPOSE OF REVIEW: Renal ischemia-reperfusion injury (IRI) is a common cause of acute kidney injury (AKI). Alterations in renal medullary blood flow (MBF) contribute to the pathogenesis of renal IRI. Here we review recent insights into the mechanisms of altered MBF in the pathogenesis of IRI. RECENT FINDINGS: Although cortical blood flow fully recovers following 30-45  min of bilateral IRI, recent studies have indicated that there is a prolonged secondary fall in MBF that is associated with a long-term decline in renal function. Recent findings indicate that angiopoietin-1, atrial natriuretic peptide, heme oxygenase-1, and the gasotransmitters CO and H(2)S, may limit the severity of IRI by preserving MBF. Additional studies have also suggested a role for cytochrome P450-derived 20-HETE in the postischemic fall in MBF. SUMMARY: Impaired MBF contributes to the pathogenesis of renal IRI. Measurement of renal MBF provides valuable insight into the underlying mechanisms of many renoprotective pathways. Identification of molecules that preserve renal MBF in IRI may lead to new therapies for AKI.

Loss of activator of G-protein signaling 3 impairs renal tubular regeneration following acute kidney injury in rodents.

The intracellular mechanisms underlying renal tubular epithelial cell proliferation and tubular repair following ischemia-reperfusion injury (IRI) remain poorly understood. In this report, we demonstrate that activator of G-protein signaling 3 (AGS3), an unconventional receptor-independent regulator of heterotrimeric G-protein function, influences renal tubular regeneration following IRI. In rat kidneys exposed to IRI, there was a temporal induction in renal AGS3 protein expression that peaked 72 h after reperfusion and corresponded to the repair and recovery phase following ischemic injury. Renal AGS3 expression was localized predominantly to the recovering outer medullary proximal tubular cells and was highly coexpressed with Ki-67, a marker of cell proliferation. Kidneys from mice deficient in the expression of AGS3 exhibited impaired renal tubular recovery 7 d following IRI compared to wild-type AGS3-expressing mice. Mechanistically, genetic knockdown of endogenous AGS3 mRNA and protein in renal tubular epithelial cells reduced cell proliferation in vitro. Similar reductions in renal tubular epithelial cell proliferation were observed following incubation with gallein, a selective inhibitor of Gßγ subunit activity, and lentiviral overexpression of the carboxyl-terminus of G-protein-coupled receptor kinase 2 (GRK2ct), a scavenger of Gßγ subunits. In summary, these data suggest that AGS3 acts through a novel receptor-independent mechanism to facilitate renal tubular epithelial cell proliferation and renal tubular regeneration.

Cannabinoid Type 2 Receptor Activation Reduces the Progression of Kidney Fibrosis Using a Mouse Model of Unilateral Ureteral Obstruction.

Background: Kidney fibrosis is a hallmark consequence of all forms of chronic kidney disease with few available treatment modalities. Material and Methods: In this study, we performed the unilateral ureteral obstruction (UUO) procedure to investigate the effects of a selective cannabinoid type 2 (CB2) agonist receptor, SMM-295, as a nephroprotective therapy. Results: SMM-295 was demonstrated to exhibit 50-fold selectivity over the cannabinoid type 1 (CB1) receptor with an EC50 ∼2 nM. Four other off-targets were identified in the safety panel, but only at the highest concentration (5 mM) tested in the assay demonstrating the relative selectivity and safety of our compound. Administration of SMM-295 (12 mg/kg IP daily) in UUO mice led to a significant decrease of 33% in tubular damage compared to the vehicle-treated UUO mice after 7 days. Consistent with these findings, there was a significant decrease in α-smooth muscle actin and fibronectin, which are markers of tubulointerstitial fibrosis, as determined by Western blot analysis. DNA damage as detected by a classic marker, γ-H2AX, was significantly reduced by 50% in the SMM-295 treatment group compared to vehicle treatment. Genetic knockout of CB2 or administration of a CB2 inverse agonist did not exhibit any beneficial effect on tubulointerstitial fibrosis or kidney tubule injury. Conclusions: In conclusion, our study provides new evidence that SMM-295 can therapeutically target the CB2 receptor with few, if any, physiological off-target sites to reduce kidney tissue damage and slow the fibrotic progression in a mouse model of kidney fibrosis.

20-HETE in acute kidney injury.

Ischemia/reperfusion injury (IRI) is a common cause of acute kidney injury (AKI) that is associated with a patient mortality of up to 50%. Currently there are not effective pharmacologic therapies for AKI. This Commentary highlights recent evidence indicating that 20-HETE plays an important role in IRI and that drugs that target this pathway have potential as therapeutic agents for AKI.