Ribonuclease inhibitor and angiogenin system regulates cell type-specific global translation.

Translation of mRNAs is a fundamental process that occurs in all cell types of multicellular organisms. Conventionally, it has been considered a default step in gene expression, lacking specific regulation. However, recent studies have documented that certain mRNAs exhibit cell type-specific translation. Despite this, it remains unclear whether global translation is controlled in a cell type-specific manner. By using human cell lines and mouse models, we found that deletion of the ribosome-associated protein ribonuclease inhibitor 1 (RNH1) decreases global translation selectively in hematopoietic-origin cells but not in the non-hematopoietic-origin cells. RNH1-mediated cell type-specific translation is mechanistically linked to angiogenin-induced ribosomal biogenesis. Collectively, this study unravels the existence of cell type-specific global translation regulators and highlights the complex translation regulation in vertebrates.

Multiomics reveals multilevel control of renal and systemic metabolism by the renal tubular circadian clock.

Circadian rhythmicity in renal function suggests rhythmic adaptations in renal metabolism. To decipher the role of the circadian clock in renal metabolism, we studied diurnal changes in renal metabolic pathways using integrated transcriptomic, proteomic, and metabolomic analysis performed on control mice and mice with an inducible deletion of the circadian clock regulator Bmal1 in the renal tubule (cKOt). With this unique resource, we demonstrated that approximately 30% of RNAs, approximately 20% of proteins, and approximately 20% of metabolites are rhythmic in the kidneys of control mice. Several key metabolic pathways, including NAD+ biosynthesis, fatty acid transport, carnitine shuttle, and ß-oxidation, displayed impairments in kidneys of cKOt mice, resulting in perturbed mitochondrial activity. Carnitine reabsorption from primary urine was one of the most affected processes with an approximately 50% reduction in plasma carnitine levels and a parallel systemic decrease in tissue carnitine content. This suggests that the circadian clock in the renal tubule controls both kidney and systemic physiology.

Increased Inflammasome Activation Is Associated with Aging and Chronic Myelomonocytic Leukemia Disease Severity.

Aging causes chronic low-grade inflammation known as inflamm-aging. It is a risk factor for several chronic disorders, including chronic myelomonocytic leukemia (CMML), a hematological malignancy that is most prevalent in older people. Recent studies suggest a critical role for the NLRP3 (NOD-, LRR- and pyrin domain-containing protein 3) inflammasome in inflamm-aging. However, the mechanisms involved in NLRP3 activation in aging and its involvement in CMML progression are not fully understood. In this study, we report that aging increases IL-1ß production upon NLRP3 activation in human CD14+ monocytes. Interestingly, we found that the TLR1/2 agonist Pam3CSK4 directly activates the NLRP3 inflammasome in monocytes from older but not from younger healthy donors. Furthermore, we observed a dichotomous response to NLRP3 inflammasome activation in monocytes from a small cohort of CMML patients, and some patients produced high levels of IL-1ß and some patients produced low levels of IL-1ß compared with older healthy donors. Intriguingly, CMML patients with heightened NLRP3 activation showed increased treatment dependency and disease severity. Collectively, our results suggest that aging causes increased sensitivity to NLRP3 inflammasome activation at a cellular level, which may explain increased inflammation and immune dysregulation in older individuals. Furthermore, NLRP3 inflammasome activation was dysregulated in a small cohort of CMML patients and was positively correlated with disease severity.

Renal tubular peroxisomes are dispensable for normal kidney function.

Peroxisomes are specialized cellular organelles involved in a variety of metabolic processes. In humans, mutations leading to complete loss of peroxisomes cause multiorgan failure (Zellweger's spectrum disorders, ZSD), including renal impairment. However, the (patho)physiological role of peroxisomes in the kidney remains unknown. We addressed the role of peroxisomes in renal function in mice with conditional ablation of peroxisomal biogenesis in the renal tubule (cKO mice). Functional analyses did not reveal any overt kidney phenotype in cKO mice. However, infant male cKO mice had lower body and kidney weights, and adult male cKO mice exhibited substantial reductions in kidney weight and kidney weight/body weight ratio. Stereological analysis showed an increase in mitochondria density in proximal tubule cells of cKO mice. Integrated transcriptome and metabolome analyses revealed profound reprogramming of a number of metabolic pathways, including metabolism of glutathione and biosynthesis/biotransformation of several major classes of lipids. Although this analysis suggested compensated oxidative stress, challenge with high-fat feeding did not induce significant renal impairments in cKO mice. We demonstrate that renal tubular peroxisomes are dispensable for normal renal function. Our data also suggest that renal impairments in patients with ZSD are of extrarenal origin.

Dysfunction of the circadian clock in the kidney tubule leads to enhanced kidney gluconeogenesis and exacerbated hyperglycemia in diabetes.

The circadian clock is a ubiquitous molecular time-keeping mechanism which synchronizes cellular, tissue, and systemic biological functions with 24-hour environmental cycles. Local circadian clocks drive cell type- and tissue-specific rhythms and their dysregulation has been implicated in pathogenesis and/or progression of a broad spectrum of diseases. However, the pathophysiological role of intrinsic circadian clocks in the kidney of diabetics remains unknown. To address this question, we induced type I diabetes with streptozotocin in mice devoid of the circadian transcriptional regulator BMAL1 in podocytes (cKOp mice) or in the kidney tubule (cKOt mice). There was no association between dysfunction of the circadian clock and the development of diabetic nephropathy in cKOp and cKOt mice with diabetes. However, cKOt mice with diabetes exhibited exacerbated hyperglycemia, increased fractional excretion of glucose in the urine, enhanced polyuria, and a more pronounced kidney hypertrophy compared to streptozotocin-treated control mice. mRNA and protein expression analyses revealed substantial enhancement of the gluconeogenic pathway in kidneys of cKOt mice with diabetes as compared to diabetic control mice. Transcriptomic analysis along with functional analysis of cKOt mice with diabetes identified changes in multiple mechanisms directly or indirectly affecting the gluconeogenic pathway. Thus, we demonstrate that dysfunction of the intrinsic kidney tubule circadian clock can aggravate diabetic hyperglycemia via enhancement of gluconeogenesis in the kidney proximal tubule and further highlight the importance of circadian behavior in patients with diabetes.

Renal tubular arginase-2 participates in the formation of the corticomedullary urea gradient and attenuates kidney damage in ischemia-reperfusion injury in mice.

AIM: Arginase 2 (ARG2) is a mitochondrial enzyme that catalyses hydrolysis of l-arginine into urea and l-ornithine. In the kidney, ARG2 is localized to the S3 segment of the proximal tubule. It has been shown that expression and activity of this enzyme are upregulated in a variety of renal pathologies, including ischemia-reperfusion (IR) injury. However, the (patho)physiological role of ARG2 in the renal tubule remains largely unknown. METHODS: We addressed this question in mice with conditional knockout of Arg2 in renal tubular cells (Arg2lox/lox /Pax8-rtTA/LC1 or, cKO mice). RESULTS: We demonstrate that cKO mice exhibit impaired urea concentration and osmolality gradients along the corticomedullary axis. In a model of unilateral ischemia-reperfusion injury (UIRI) with an intact contralateral kidney, ischemia followed by 24 hours of reperfusion resulted in significantly more pronounced histological damage in ischemic kidneys from cKO mice compared to control and sham-operated mice. In parallel, UIRI-subjected cKO mice exhibited a broad range of renal functional abnormalities, including albuminuria and aminoaciduria. Fourteen days after UIRI, the cKO mice exhibited complex phenotype characterized by significantly lower body weight, increased plasma levels of early predictive markers of kidney disease progression (asymmetric dimethylarginine and symmetric dimethylarginine), impaired mitochondrial function in the ischemic kidney but no difference in kidney fibrosis as compared to control mice. CONCLUSION: Collectively, these results establish the role of ARG2 in the formation of corticomedullary urea and osmolality gradients and suggest that this enzyme attenuates kidney damage in ischemia-reperfusion injury.

Deletion of the serine protease CAP2/Tmprss4 leads to dysregulated renal water handling upon dietary potassium depletion.

The kidney needs to adapt daily to variable dietary K+ contents via various mechanisms including diuretic, acid-base and hormonal changes that are still not fully understood. In this study, we demonstrate that following a K+-deficient diet in wildtype mice, the serine protease CAP2/Tmprss4 is upregulated in connecting tubule and cortical collecting duct and also localizes to the medulla and transitional epithelium of the papilla and minor calyx. Male CAP2/Tmprss4 knockout mice display altered water handling and urine osmolality, enhanced vasopressin response leading to upregulated adenylate cyclase 6 expression and cAMP overproduction, and subsequently greater aquaporin 2 (AQP2) and Na+-K+-2Cl- cotransporter 2 (NKCC2) expression following K+-deficient diet. Urinary acidification coincides with significantly increased H+,K+-ATPase type 2 (HKA2) mRNA and protein expression, and decreased calcium and phosphate excretion. This is accompanied by increased glucocorticoid receptor (GR) protein levels and reduced 11ß-hydroxysteroid dehydrogenase 2 activity in knockout mice. Strikingly, genetic nephron-specific deletion of GR leads to the mirrored phenotype of CAP2/Tmprss4 knockouts, including increased water intake and urine output, urinary alkalinisation, downregulation of HKA2, AQP2 and NKCC2. Collectively, our data unveil a novel role of the serine protease CAP2/Tmprss4 and GR on renal water handling upon dietary K+ depletion.

The intrinsic circadian clock in podocytes controls glomerular filtration rate.

Glomerular filtration rate (GFR), or the rate of primary urine formation, is the key indicator of renal function. Studies have demonstrated that GFR exhibits significant circadian rhythmicity and, that these rhythms are disrupted in a number of pathologies. Here, we tested a hypothesis that the circadian rhythm of GFR is driven by intrinsic glomerular circadian clocks. We used mice lacking the circadian clock protein BMAL1 specifically in podocytes, highly specialized glomerular cells critically involved in the process of glomerular filtration (Bmal1lox/lox/Nphs2-rtTA/LC1 or, cKO mice). Circadian transcriptome profiling performed on isolated glomeruli from control and cKO mice revealed that the circadian clock controls expression of multiple genes encoding proteins essential for normal podocyte function. Direct assessment of glomerular filtration by inulin clearance demonstrated that circadian rhythmicity in GFR was lost in cKO mice that displayed an ultradian rhythm of GFR with 12-h periodicity. The disruption of circadian rhythmicity in GFR was paralleled by significant changes in circadian patterns of urinary creatinine, sodium, potassium and water excretion and by alteration in the diurnal pattern of plasma aldosterone levels. Collectively, these results indicate that the intrinsic circadian clock in podocytes participate in circadian rhythmicity of GFR.

Renal Fanconi Syndrome and Hypophosphatemic Rickets in the Absence of Xenotropic and Polytropic Retroviral Receptor in the Nephron.

Tight control of extracellular and intracellular inorganic phosphate (Pi) levels is critical to most biochemical and physiologic processes. Urinary Pi is freely filtered at the kidney glomerulus and is reabsorbed in the renal tubule by the action of the apical sodium-dependent phosphate transporters, NaPi-IIa/NaPi-IIc/Pit2. However, the molecular identity of the protein(s) participating in the basolateral Pi efflux remains unknown. Evidence has suggested that xenotropic and polytropic retroviral receptor 1 (XPR1) might be involved in this process. Here, we show that conditional inactivation of Xpr1 in the renal tubule in mice resulted in impaired renal Pi reabsorption. Analysis of Pi transport in primary cultures of proximal tubular cells or in freshly isolated renal tubules revealed that this Xpr1 deficiency significantly affected Pi efflux. Further, mice with conditional inactivation of Xpr1 in the renal tubule exhibited generalized proximal tubular dysfunction indicative of Fanconi syndrome, characterized by glycosuria, aminoaciduria, calciuria, and albuminuria. Dramatic alterations in the renal transcriptome, including a significant reduction in NaPi-IIa/NaPi-IIc expression, accompanied these functional changes. Additionally, Xpr1-deficient mice developed hypophosphatemic rickets secondary to renal dysfunction. These results identify XPR1 as a major regulator of Pi homeostasis and as a potential therapeutic target in bone and kidney disorders.

Nephron-Specific Deletion of Circadian Clock Gene Bmal1 Alters the Plasma and Renal Metabolome and Impairs Drug Disposition.

The circadian clock controls a wide variety of metabolic and homeostatic processes in a number of tissues, including the kidney. However, the role of the renal circadian clocks remains largely unknown. To address this question, we performed a combined functional, transcriptomic, and metabolomic analysis in mice with inducible conditional knockout (cKO) of BMAL1, which is critically involved in the circadian clock system, in renal tubular cells (Bmal1lox/lox/Pax8-rtTA/LC1 mice). Induction of cKO in adult mice did not produce obvious abnormalities in renal sodium, potassium, or water handling. Deep sequencing of the renal transcriptome revealed significant changes in the expression of genes related to metabolic pathways and organic anion transport in cKO mice compared with control littermates. Furthermore, kidneys from cKO mice exhibited a significant decrease in the NAD+-to-NADH ratio, which reflects the oxidative phosphorylation-to-glycolysis ratio and/or the status of mitochondrial function. Metabolome profiling showed significant changes in plasma levels of amino acids, biogenic amines, acylcarnitines, and lipids. In-depth analysis of two selected pathways revealed a significant increase in plasma urea level correlating with increased renal Arginase II activity, hyperargininemia, and increased kidney arginine content as well as a significant increase in plasma creatinine concentration and a reduced capacity of the kidney to secrete anionic drugs (furosemide) paralleled by an approximate 80% decrease in the expression level of organic anion transporter 3 (SLC22a8). Collectively, these results indicate that the renal circadian clocks control a variety of metabolic/homeostatic processes at the intrarenal and systemic levels and are involved in drug disposition.

Epithelial Sodium Channel-Mediated Sodium Transport Is Not Dependent on the Membrane-Bound Serine Protease CAP2/Tmprss4.

The membrane-bound serine protease CAP2/Tmprss4 has been previously identified in vitro as a positive regulator of the epithelial sodium channel (ENaC). To study its in vivo implication in ENaC-mediated sodium absorption, we generated a knockout mouse model for CAP2/Tmprss4. Mice deficient in CAP2/Tmprss4 were viable, fertile, and did not show any obvious histological abnormalities. Unexpectedly, when challenged with sodium-deficient diet, these mice did not develop any impairment in renal sodium handling as evidenced by normal plasma and urinary sodium and potassium electrolytes, as well as normal aldosterone levels. Despite minor alterations in ENaC mRNA expression, we found no evidence for altered proteolytic cleavage of ENaC subunits. In consequence, ENaC activity, as monitored by the amiloride-sensitive rectal potential difference (ΔPD), was not altered even under dietary sodium restriction. In summary, ENaC-mediated sodium balance is not affected by lack of CAP2/Tmprss4 expression and thus, does not seem to directly control ENaC expression and activity in vivo.

A fast automated screening method for the detection of blood transfusion in sports.

Currently, there is no simple direct screening method for the misuse of blood transfusions in sports. In this study, we investigated whether the measurement of iron in EDTA-plasma can serve as biomarker for such purpose. Our results revealed an increase of the plasma iron level up to 25-fold 6 h after blood re-infusion. The variable remained elevated 10-fold one day after the procedure. A specificity of 100% and a sensitivity of 93% were obtained with a proposed threshold at 45 µg/dL of plasma iron. Therefore, our test could be used as a simple, cost effective biomarker for the screening for blood transfusion misuse in sports.