Retrograde urethrogram - a novel approach to diagnosing a posterior urethral polyp in a neonate.

We present a case of antenatally detected fetal megacystis caused by an obstructing posterior urethral polyp. Antenatal and postnatal ultrasounds showed bladder wall thickening and bilateral hydroureteronephrosis, most marked antenatally. A working diagnosis of posterior urethral valves was therefore made. However, further postnatal assessment with a micturating cystourethrogram (MCUG) combined with a retrograde urethrogram identified a pedunculated urethral polyp as the cause. The addition of a retrograde urethrogram as an adjunct to the MCUG in the diagnosis of posterior urethral polyp has not previously been reported, and in this case provided diagnostic confidence of this rare condition, allowing for definitive surgical planning.

A rare ventriculoarterial connection: double outlet of both ventricles.

Ventriculoarterial connection is one of the important points of the segmental approach to congenital cardiac malformations. Double outlet of both ventricles is a rare form where both great arterial roots override the interventricular septum. In this article, we aimed to draw attention to this very rare form of ventriculoarterial connection by presenting an infant case diagnosed using echocardiography, CT angiography, and 3-dimensional modelling.

Achieving Robust Compressive Sensing Seismic Acquisition with a Two-Step Sampling Approach.

The compressive sensing (CS) framework offers a cost-effective alternative to dense alias-free sampling. Designing seismic layouts based on the CS technique imposes the use of specific sampling patterns in addition to the logistical and geophysical requirements. We propose a two-step design process for generating CS-based schemes suitable for seismic applications. During the first step, uniform random sampling is used to generate a random scheme, which is supported theoretically by the restricted isometry property. Following that, designated samples are added to the random scheme to control the maximum distance between adjacent sources (or receivers). The null space property theoretically justifies the additional samples of the second step. Our sampling method generates sampling patterns with a CS theoretical background, controlled distance between adjacent samples, and a flexible number of active and omitted samples. The robustness of two-step sampling schemes for reallocated samples is investigated and CS reconstruction tests are performed. In addition, using this approach, a CS-based 3D seismic survey is designed, and the distributions of traces in fold maps and rose diagrams are analyzed. It is shown that the two-step scheme is suitable for CS-based seismic surveys and field applications.

Morphological Subtypes of Intracranial Internal Carotid Artery Arteriosclerosis and the Risk of Stroke.

BACKGROUND: Accumulating evidence highlights the existence of distinct morphological subtypes of intracranial carotid arteriosclerosis. So far, little is known on the prevalence of these subtypes and subsequent stroke risk in the general population. We determined the prevalence of morphological subtypes of intracranial arteriosclerosis and assessed the risk of stroke associated with these subtypes. METHODS: Between 2003 and 2006, 2391 stroke-free participants (mean age 69.6, 51.7% women) from the population-based Rotterdam Study underwent noncontrast computed tomography to visualize calcification in the intracranial carotid arteries as a proxy for intracranial arteriosclerosis. Calcification morphology was evaluated according to a validated grading scale and categorized into intimal, internal elastic lamina (IEL), or mixed subtype. Follow-up for stroke was complete until January 1, 2016. We used multivariable Cox regression to assess associations of each subtype with incident stroke. RESULTS: The prevalence of calcification was 82% of which 39% had the intimal subtype, 48% IEL subtype, and 13% a mixed subtype. During a median follow-up of 10.4 years, 155 participants had a stroke. All 3 subtypes were associated with a higher risk of stroke (adjusted hazard ratio [95% CI] for intimal: 2.11 [1.07-4.13], IEL: 2.66 [1.39-5.11], and mixed subtype 2.57 [1.18-5.61]). The association of the IEL subtype with stroke was strongest among older participants. The association of the intimal subtype with stroke was noticeably stronger in women than in men. CONCLUSIONS: Calcification of the IEL was the most prevalent subtype of intracranial arteriosclerosis. All 3 subtypes were associated with an increased risk of stroke, with noticeable age and sex-specific differences.

Radio-frequency induced heating of intra-cranial EEG electrodes: The more the colder?

Many neurological disorders are analyzed and treated with implantable electrodes. Many patients with such electrodes have to undergo MRI examinations - often unrelated to their implant - at the risk of radio-frequency induced heating. The number of electrode contact sites of these implants keeps increasing due to improvements in manufacturing and computational algorithms. Electrode grids with multiple receive channels couple to the RF fields present in MRI, but, due to their proximity, a combination of leads has a coupling response which is not a superposition of the individual leads' response. To investigate the problem of RF-induced heating of coupled multi-lead implants, temperature mapping was performed on a set of intra-cranial electroencephalogram (icEEG) electrode grid prototypes with increasing number of contact sites (1-16). Additionally, electric field measurements were used to investigate the radio-frequency heating characteristics of the implants in different media combinations, simulating the device being partially immersed inside the patient. MR measurements show RF-induced heating up to 19.6 K for the single electrode, reducing monotonically with larger number of contact sites to a minimum of 0.9 K for the largest grid. The SAR calculated from temperature measurements agrees well with electric field mapping: The same trend is visible for different insertion lengths, however, the energy dissipated by the whole implant varies with the grid size and insertion length. Thus, in the tested circumstances, a larger electrode number either reduced or had a similar risk of RF induced heating, indicating, that the size of electrode grids is a design parameter, which can be used to change an implants RF response and in turn to reduce the risk of RF induced heating and improve the safety of patient with neuro-implants undergoing MRI examinations.

Mechanical Characterization of Thrombi Retrieved With Endovascular Thrombectomy in Patients With Acute Ischemic Stroke.

Background and Purpose: Mechanical properties of thromboemboli play an important role in the efficacy of endovascular thrombectomy (EVT) for acute ischemic stroke. However, very limited data on mechanical properties of human stroke thrombi are available. We aimed to mechanically characterize thrombi retrieved with EVT, and to assess the relationship between thrombus composition and thrombus stiffness. Methods: Forty-one thrombi from 19 patients with acute stroke who underwent EVT between July and October 2019 were mechanically analyzed, directly after EVT. We performed unconfined compression experiments and determined tangent modulus at 75% strain (Et75) as a measure for thrombus stiffness. Thrombi were histologically analyzed for fibrin/platelets, erythrocytes, leukocytes, and platelets, and we assessed the relationship between histological components and Et75 with univariable and multivariable linear mixed regression. Results: Median Et75 was 560 (interquartile range, 393­1161) kPa. In the multivariable analysis, fibrin/platelets were associated with increased Et75 (aß, 9 [95% CI, 5 to 13]) kPa, erythrocytes were associated with decreased Et75% (aß, −9 [95% CI, −5 to −13]) kPa. We found no association between leukocytes and Et75. High platelet values were strongly associated with increased Et75 (aß, 56 [95% CI, 38­73]). Conclusions: Fibrin/platelet content of thrombi retrieved with EVT for acute ischemic stroke is strongly associated with increased thrombus stiffness. For thrombi with high platelet values, there was a very strong relationship with thrombus stiffness. Our data provide a basis for future research on the development of next-generation EVT devices tailored to thrombus composition.

Iron sucrose ('RBT-3') activates the hepatic and renal HAMP1 gene, evoking renal hepcidin loading and resistance to cisplatin nephrotoxicity.

BACKGROUND: Iron sucrose (FeS) administration induces a state of renal preconditioning, protecting against selected forms of acute kidney injury (AKI). Recent evidence suggests that recombinant hepcidin also mitigates acute renal damage. Hence the goals of this study were to determine whether a new proprietary FeS formulation ('RBT-3') can acutely activate the hepcidin (HAMP1) gene in humans, raising plasma and renal hepcidin concentrations; assess whether the kidney participates in this posited RBT-3-hepcidin generation response; test whether RBT-3 can mitigate a clinically relevant AKI model (experimental cisplatin toxicity) and explore whether mechanisms in addition to hepcidin generation are operative in RBT-3's cytoprotective effects. METHODS: Healthy human volunteers (n = 9) and subjects with Stages 3-4 CKD (n = 9) received 120, 240 or 360 mg of RBT-3 (intravenously over 2 h). Plasma and urine samples were collected and assayed for hepcidin levels (0-72 h post-RBT-3 injection). In complementary mouse experiments, RBT-3 effects on hepatic versus renal hepcidin (HAMP1) messenger RNA (mRNA) and protein levels were compared. RBT-3's impact on the mouse Nrf2 pathway and on experimental cisplatin nephrotoxicity was assessed. Direct effects of exogenous hepcidin on in vivo and in vitro (HK-2 cells) cisplatin toxicity were also tested. RESULTS: RBT-3 induced rapid, dose-dependent and comparable plasma hepcidin increases in both healthy volunteers and chronic kidney disease subjects (∼15 times baseline within 24 h). Human kidney hepcidin exposure was confirmed by 4-fold urinary hepcidin increases. RBT-3 up-regulated mouse hepcidin mRNA, but much more so in kidney (>25 times) versus liver (∼2 times). RBT-3 also activated kidney Nrf2 [increased Nrf2 nuclear binding; increased Nrf2-responsive gene mRNAs: heme oxygenase-1, sulfiredoxin-1, glutamate-cysteine ligase catalytic subunit and NAD(P)H quinone dehydrogenase 1]. RBT-3 preconditioning (18 h time lapse) markedly attenuated experimental cisplatin nephrotoxicity (∼50% blood urea nitrogen/creatinine decrements), in part by reducing renal cisplatin uptake by 40%. Exogenous hepcidin (without RBT-3) treatment conferred protection against mild in vivo (but not in vitro) cisplatin toxicity. CONCLUSIONS: RBT-3 acutely and dramatically up-regulates cytoprotective hepcidin production, increasing renal hepcidin levels. However, additional cytoprotective mechanisms are activated by RBT-3 (e.g. Nrf2 activation; reduced cisplatin uptake). Thus RBT-3-induced preconditioning likely confers renal resistance to cisplatin via an interplay of multiple cytoprotective activities.

Predicting cell lineages using autoencoders and optimal transport.

Lineage tracing involves the identification of all ancestors and descendants of a given cell, and is an important tool for studying biological processes such as development and disease progression. However, in many settings, controlled time-course experiments are not feasible, for example when working with tissue samples from patients. Here we present ImageAEOT, a computational pipeline based on autoencoders and optimal transport for predicting the lineages of cells using time-labeled datasets from different stages of a cellular process. Given a single-cell image from one of the stages, ImageAEOT generates an artificial lineage of this cell based on the population characteristics of the other stages. These lineages can be used to connect subpopulations of cells through the different stages and identify image-based features and biomarkers underlying the biological process. To validate our method, we apply ImageAEOT to a benchmark task based on nuclear and chromatin images during the activation of fibroblasts by tumor cells in engineered 3D tissues. We further validate ImageAEOT on chromatin images of various breast cancer cell lines and human tissue samples, thereby linking alterations in chromatin condensation patterns to different stages of tumor progression. Our results demonstrate the promise of computational methods based on autoencoding and optimal transport principles for lineage tracing in settings where existing experimental strategies cannot be used.

Inositols: From Established Knowledge to Novel Approaches.

Myo-inositol (myo-Ins) and D-chiro-inositol (D-chiro-Ins) are natural compounds involved in many biological pathways. Since the discovery of their involvement in endocrine signal transduction, myo-Ins and D-chiro-Ins supplementation has contributed to clinical approaches in ameliorating many gynecological and endocrinological diseases. Currently both myo-Ins and D-chiro-Ins are well-tolerated, effective alternative candidates to the classical insulin sensitizers, and are useful treatments in preventing and treating metabolic and reproductive disorders such as polycystic ovary syndrome (PCOS), gestational diabetes mellitus (GDM), and male fertility disturbances, like sperm abnormalities. Moreover, besides metabolic activity, myo-Ins and D-chiro-Ins deeply influence steroidogenesis, regulating the pools of androgens and estrogens, likely in opposite ways. Given the complexity of inositol-related mechanisms of action, many of their beneficial effects are still under scrutiny. Therefore, continuing research aims to discover new emerging roles and mechanisms that can allow clinicians to tailor inositol therapy and to use it in other medical areas, hitherto unexplored. The present paper outlines the established evidence on inositols and updates on recent research, namely concerning D-chiro-Ins involvement into steroidogenesis. In particular, D-chiro-Ins mediates insulin-induced testosterone biosynthesis from ovarian thecal cells and directly affects synthesis of estrogens by modulating the expression of the aromatase enzyme. Ovaries, as well as other organs and tissues, are characterized by a specific ratio of myo-Ins to D-chiro-Ins, which ensures their healthy state and proper functionality. Altered inositol ratios may account for pathological conditions, causing an imbalance in sex hormones. Such situations usually occur in association with medical conditions, such as PCOS, or as a consequence of some pharmacological treatments. Based on the physiological role of inositols and the pathological implications of altered myo-Ins to D-chiro-Ins ratios, inositol therapy may be designed with two different aims: (1) restoring the inositol physiological ratio; (2) altering the ratio in a controlled way to achieve specific effects.

Magnetic resonance imaging for pathobiological assessment and interventional treatment of the coronary arteries.

X-ray-based fluoroscopy is the standard tool for diagnostics and intervention in coronary artery disease. In recent years, computed tomography has emerged as a non-invasive alternative to coronary angiography offering detection of coronary calcification and imaging of the vessel lumen by the use of iodinated contrast agents. Even though currently available invasive or non-invasive techniques can show the degree of vessel stenosis, they are unable to provide information about biofunctional plaque properties, e.g. plaque inflammation. Furthermore, the use of radiation and the necessity of iodinated contrast agents remain unfavourable prerequisites. Magnetic resonance imaging (MRI) is a radiation-free alternative to X-ray which offers anatomical and functional imaging contrasts fostering the idea of non-invasive biofunctional assessment of the coronary vessel wall. In combination with molecular contrast agents that target-specific epitopes of the vessel wall, MRI might reveal unique plaque properties rendering it, for example, 'vulnerable and prone to rupture'. Early detection of these lesions may allow for early or prophylactic treatment even before an adverse coronary event occurs. Besides diagnostic imaging, advances in real-time image acquisition and motion compensation now provide grounds for MRI-guided coronary interventions. In this article, we summarize our research on MRI-based molecular imaging in cardiovascular disease and feature our advances towards real-time MRI-based coronary interventions in a porcine model.

La fluoroscopia con rayos X es la herramienta estándar para el diagnóstico y la intervención de coronariopatías. En los últimos años, la tomografía computarizada se ha convertido en una alternativa atraumática a la coronariografía, ya que se puede detectar la calcificación coronaria y ver a través de imágenes las luces de los vasos sanguíneos mediante el uso de medios de contraste yodados. Si bien las técnicas traumáticas o atraumáticas disponibles actualmente pueden mostrar el grado de la estenosis vascular, no pueden proporcionar información sobre las propiedades biofuncionales de la placa de ateroma, por ejemplo, inflamación de la placa de ateroma. Por otra parte, el uso de radiación y la necesidad de agentes de contraste yodados siguen siendo requisitos desfavorables. La resonancia magnética (RM) es una alternativa sin radiación a los rayos X que proporciona contrates de imagen con información anatómica y funcional, lo cual refuerza la idea del diagnóstico biofuncional atraumático de las paredes de los vasos coronarios. En combinación con medios de contraste molecular que actúan sobre epítopos específicos de las paredes de los vasos, la RM puede poner de manifiesto propiedades particulares de la placa de ateroma mediante su representación, por ejemplo, «vulnerabilidad y predisposición a rotura¼. La detección precoz de este tipo de lesiones puede facilitar un tratamiento a tiempo o preventivo antes de que tenga lugar una complicación coronaria grave.Además del diagnóstico por imagen, los avances en la adquisición de imágenes en tiempo real y la compensación del movimiento sirven de base para las intervenciones coronarias guiadas por RM. En este artículo, ofrecemos un resumen de nuestra investigación sobre imagen molecular con resonancia magnética en enfermedades cardiovasculares y presentamos nuestros avances hacia las intervenciones coronarias con RM en tiempo real en un modelo porcino.

Pediatric functional hemispherectomy: operative techniques and complication avoidance.

Functional hemispherectomy/hemispherotomy is a disconnection procedure for severe medically refractory epilepsy where the seizure foci diffusely localize to one hemisphere. It is an improvement on anatomical hemispherectomy and was first performed by Rasmussen in 1974. Less invasive surgical approaches and refinements have been made to improve seizure freedom and minimize surgical morbidity and complications. Key anatomical structures that are disconnected include the 1) internal capsule and corona radiata, 2) mesial temporal structures, 3) insula, 4) corpus callosum, 5) parietooccipital connection, and 6) frontobasal connection. A stepwise approach is indicated to ensure adequate disconnection and prevent seizure persistence or recurrence. In young pediatric patients, careful patient selection and modern surgical techniques have resulted in > 80% seizure freedom and very good functional outcome. In this report, the authors summarize the history of hemispherectomy and its development and present a graphical guide for this anatomically challenging procedure. The use of the osteoplastic flap to improve outcome and the management of hydrocephalus are discussed.

Synergistic activation of mitochondrial metabolism and the glutathione redox couple protects HepG2 hepatocarcinoma cells from palmitoylcarnitine-induced stress.

Fatty acid stress can have divergent effects in various cancers. We explored how metabolic and redox flexibility in HepG2 hepatocarcinoma cells mediates protection from palmitoylcarnitine. HepG2 cells, along with HCT 116 and HT29 colorectal cancer cells were incubated with 100 µM palmitoylcarnitine for up to 48 h. Mitochondrial H2O2 emission, glutathione, and cell survival were assessed in HT29 and HepG2 cells. 100 µM palmitoylcarnitine promoted early growth in HepG2 cells by ~8% after 48 h versus decreased cell survival observed in HT29 and HCT 116 cells. Palmitoylcarnitine increased mitochondrial respiration at physiological and maximal concentrations of ADP, while lowering cellular lactate content in HepG2 cells, suggesting a switch to mitochondrial metabolism. HepG2 cell growth was associated with an early increase in H2O2 emission by 10 min, followed by a decrease in H2O2 at 24 h that corresponded with increased glutathione content, suggesting a redox-based compensatory mechanism. In contrast, abrogation of HT29 cell proliferation was related to decreased mitochondrial respiration (likely due to cell death) and decreased glutathione. Concurrent glutathione depletion with BSO prevented palmitoylcarnitine-induced growth in HepG2 cells, indicating that glutathione was critical for promoting growth following palmitoylcarnitine. Inhibiting UCP2 with genipin sensitized HepG2 cells to palmitoylcarnitine, suggesting that activation of UCP2 may be a 2nd redox-based mechanism conferring protection. These findings suggest that HepG2 cells possess inherent metabolic and redox flexibility relative to HT29 cells that confers protection from palmitoylcarnitine-induced stress via adaptive increases in mitochondrial respiratory control, glutathione buffering, and induction of UCP2.

It's the little things: On the complexity of planar electrode heating in MRI.

Neurological disorders are increasingly analysed and treated with implantable electrodes, and patients with such electrodes are studied with MRI despite the risk of radio-frequency (RF) induced heating during the MRI exam. Recent clinical research suggests that electrodes with smaller diameters of the electrical interface between implant and tissue are beneficial; however, the influence of this electrode contact diameter on RF-induced heating has not been investigated. In this work, electrode contact diameters between 0.3 and 4 mm of implantable electrodes appropriate for stimulation and electrocorticography were evaluated in a 1.5 T MRI system. In situ temperature measurements adapted from the ASTM standard test method were performed and complemented by simulations of the specific absorption rate (SAR) to assess local SAR values, temperature increase and the distribution of dissipated power. Measurements showed temperature changes between 0.8 K and 53 K for different electrode contact diameters, which is well above the legal limit of 1 K. Systematic errors in the temperature measurements are to be expected, as the temperature sensors may disturb the heating pattern near small electrodes. Compared to large electrodes, simulations suggest that small electrodes are subject to less dissipated power, but more localized power density. Thus, smaller electrodes might be classified as safe in current certification procedures but may be more likely to burn adjacent tissue. To assess these local heating phenomena, smaller temperature sensors or new non-invasive temperature sensing methods are needed.

Acute kidney injury induces dramatic p21 upregulation via a novel, glucocorticoid-activated, pathway.

The cyclin kinase inhibitor p21 is acutely upregulated during acute kidney injury (AKI) and exerts cytoprotective effects. A proposed mechanism is oxidant stress-induced activation of p53, the dominant p21 transcription factor. Glycerol-induced rhabdomyolysis induces profound renal oxidant stress. Hence, we studied this AKI model to determine whether p53 activation corresponds with p21 gene induction and/or whether alternative mechanism(s) might be involved. CD-1 mice were subjected to glycerol-induced AKI. After 4 or 18 h, plasma, urinary, and renal cortical p21 protein and mRNA levels were assessed. Renal p53 activation was gauged by measurement of both total and activated (Ser15-phosphorylated) p53 and p53 mRNA levels. Glycerol evoked acute, progressive increases in renal cortical p21 mRNA and protein levels. Corresponding plasma (~25-fold) and urinary (~75-fold) p21 elevations were also observed. Renal cortical ratio of total to phosphorylated (Ser15) p53 rose three- to fourfold. However, the p53 inhibitor pifithrin-α failed to block glycerol-induced p21 gene induction, suggesting that an alternative p21 activator might also be at play. To this end, it was established that glycerol-induced AKI 1) dramatically increased plasma (~5-fold) and urinary (~75-fold) cortisol levels, 2) the glucocorticoid receptor antagonist mifepristone blocked glycerol-induced p21 mRNA and protein accumulation, and 3) dexamethasone or cortisol injections markedly increased p21 protein and mRNA in both normal and glycerol-treated mice, although no discernible p53 protein or mRNA increases were observed. We conclude that AKI-induced "systemic stress" markedly increases plasma and urinary cortisol, which can then activate renal p21 gene expression, at least in part, via a glucocorticoid receptor-dependent signaling pathway. Discernible renal cortical p53 increases are not required for this dexamethasone-mediated p21 response.

Parenterial iron sucrose-induced renal preconditioning: differential ferritin heavy and light chain expression in plasma, urine, and internal organs.

Experimental data suggest that iron sucrose (FeS) injection, used either alone or in combination with other prooxidants, can induce "renal preconditioning," in part by upregulating cytoprotective ferritin levels. However, the rapidity, degree, composition (heavy vs. light chain), and renal ferritin changes after FeS administration in humans remain to be defined. To address these issues, healthy human volunteers (n = 9) and patients with stage 3-4 chronic kidney disease(n = 9) were injected once with FeS (120, 240, or 360 mg). Plasma ferritin was measured from 0 to 8 days postinjection as an overall index of ferritin generation. Urinary ferritin served as a "biomarker" of renal ferritin production. FeS induced rapid (≤2 h), dose-dependent, plasma ferritin increases in all study participants, peaking at approximately three to five times baseline within 24-48 h. Significant urinary ferritin increases (~3 times), without dose-dependent increases in albuminuria, neutrophil gelatinase-associated lipocalin, or N-acetyl-ß-d-glucosaminidase excretion, were observed. Western blot analysis with ferritin heavy chain (Fhc)- and light chain (Flc)-specific antibodies demonstrated that FeS raised plasma Flc but not Fhc levels. Conversely, FeS increased both Fhc and Flc in urine. To assess sites of FeS-induced ferritin generation, organs from FeS-treated mice were probed for Fhc, Flc, and their mRNAs. FeS predominantly raised hepatic Flc. Conversely, marked Fhc and Flc elevations developed in the kidney and spleen. No cardiopulmonary ferritin increases occurred. Ferritin mRNAs remained unchanged throughout, implying posttranscriptional ferritin production. We conclude that FeS induces rapid, dramatic, and differential Fhc and Flc upregulation in organs. Renal Fhc and Flc increases, in the absence of nephrotoxicity, suggest potential FeS utility as a clinical renal "preconditioning" agent.

Synchronous identification of a dysembryoplastic neuroepithelial tumor (DNET) and an oligodendroglioma in a patient: A case report.

Synchronous gliomas of different histopathology are quite rare in non-syndromic, non-irradiated patients. Although "mixed" gliomas are not infrequent, and malignant gliomas often contain areas of disparate differentiation (e.g., glioblastoma with ependymal differentiation), it is unusual to find gliomas of different lineage presenting concurrently. We present a case of synchronous gliomas, one dysembryoplastic neuroepithelial tumor (DNET) and the other oligodendroglioma.

Mechanisms Underlying Increased TIMP2 and IGFBP7 Urinary Excretion in Experimental AKI.

BACKGROUND: Recent clinical data support the utility/superiority of a new AKI biomarker ("NephroCheck"), the arithmetic product of urinary TIMP × IGFBP7 concentrations. However, the pathophysiologic basis for its utility remains ill defined. METHODS: To clarify this issue, CD-1 mice were subjected to either nephrotoxic (glycerol, maleate) or ischemic AKI. Urinary TIMP2/IGFBP7 concentrations were determined at 4 and 18 hours postinjury and compared with urinary albumin levels. Gene transcription was assessed by measuring renal cortical and/or medullary TIMP2/IGFBP7 mRNAs (4 and 18 hours after AKI induction). For comparison, the mRNAs of three renal "stress" biomarkers (NGAL, heme oxygenase 1, and p21) were assessed. Renal cortical TIMP2/IGFBP7 protein was gauged by ELISA. Proximal tubule-specific TIMP2/IGFBP7 was assessed by immunohistochemistry. RESULTS: Each AKI model induced prompt (4 hours) and marked urinary TIMP2/IGFBP7 increases without an increase in renal cortical concentrations. Furthermore, TIMP2/IGFBP7 mRNAs remained at normal levels. Endotoxemia also failed to increase TIMP2/IGFBP7 mRNAs. In contrast, each AKI model provoked massive NGAL, HO-1, and p21 mRNA increases, confirming that a renal "stress response" had occurred. Urinary albumin rose up to 100-fold and strongly correlated (r=0.87-0.91) with urinary TIMP2/IGFBP7 concentrations. Immunohistochemistry showed progressive TIMP2/IGFBP7 losses from injured proximal tubule cells. Competitive inhibition of endocytic protein reabsorption in normal mice tripled urinary TIMP2/IGFBP7 levels, confirming this pathway's role in determining urinary excretion. CONCLUSIONS: AKI-induced urinary TIMP2/IGFBP7 elevations are not due to stress-induced gene transcription. Rather, increased filtration, decreased tubule reabsorption, and proximal tubule cell TIMP2/IGFBP7 urinary leakage seem to be the most likely mechanisms.

Plasma and urinary p21: potential biomarkers of AKI and renal aging.

p21 is upregulated in renal tubules in response to acute kidney injury ( AKI). and localizes in the nucleus, where it induces cell cycle arrest (CCA). These events can mitigate early injury but can also facilitate the onset of the degenerative cell senescence/"aging" process. Hence, we asked the following: 1) can AKI-induced p21 upregulation be gauged by plasma and/or urinary p21 assay; 2) might p21 serve as an AKI/CCA biomarker; and 3) does p21 accumulate during normal renal aging, and might plasma p21 reflect this process? Mice were subjected to either ischemia-reperfusion (I/R) or nephotoxic (maleate) AKI. Renal cortical p21 expression (protein, mRNA) was assessed 2-18 h later and contrasted with plasma/urine p21 concentrations (ELISA). p21 mRNA/protein levels were also measured in aging mice (2, 12, 24 mo). AKI induced marked, progressive, increases in renal cortical p21 mRNA and protein levels. These changes were marked by acute (within 2-4 h) and profound increases (up to 200×) in both plasma and urine p21 concentrations. Renal I/R also activated p21 gene expression in extrarenal organs (heart, brain), consistent with so-called "organ cross talk". p21 efflux from damaged cells was confirmed with studies of hypoxia-injured, isolated proximal tubules. Aging was associated with progressive renal cortical p21 expression, which correlated ( r, 0.83) with rising plasma p21 concentrations. We concluded that 1) during AKI, renal p21 increases can be gauged by either plasma or urine p21 assay, serving as potentially useful AKI/CCA biomarkers; 2) AKI can activate p21 in extrarenal organs; and 3) plasma p21 levels may provide an index of the renal/systemic aging process.

Mechanisms and consequences of oxidant-induced renal preconditioning: an Nrf2-dependent, P21-independent, anti-senescence pathway.

Background: P21, a cyclin kinase inhibitor, is upregulated by renal 'ischemic preconditioning' (IPC), and induces a 'cytoresistant' state. However, P21-induced cell cycle inhibition can also contribute to cellular senescence, a potential adverse renal event. Hence, this study assessed whether: (i) IPC-induced P21 upregulation is associated with subsequent renal senescence; and (ii) preconditioning can be established 'independent' of P21 induction and avoid a post-ischemic senescent state? Methods: CD-1 mice were subjected to either IPC (5-15 min) or to a recently proposed 'oxidant-induced preconditioning' (OIP) strategy (tin protoporphyrin-induced heme oxygenase inhibition +/- parental iron administration). P21 induction [messenger RNA (mRNA)/protein], cell proliferation (KI-67, phosphohistone H3 nuclear staining), kidney senescence (P16ink4a; P19Arf mRNAs; senescence-associated beta-galactosidase levels) and resistance to ischemic acute kidney injury were assessed. Results: IPC induced dramatic (10-25×) and persistent P21 activation and 'downstream' tubular senescence. Conversely, OIP did not upregulate P21, it increased, rather than decreased, cell proliferation markers, and it avoided a senescence state. OIP markedly suppressed ischemia-induced P21 up-regulation, it inhibited the development of post-ischemic senescence and it conferred near-complete protection against ischemic acute renal failure (ARF). To assess OIP's impact on a non-P21-dependent cytoprotective pathway, its ability to activate Nrf2, the so-called 'master regulator' of endogenous cell defenses, was assessed. Within 4 h, OIP activated each of three canonical Nrf2-regulated genes (NQO1, SRXN1, GCLC; 3- to 5-fold mRNA increases). Conversely, this gene activation pathway was absent in Nrf2-/- mice, confirming Nrf2 specificity. Nrf2-/- mice also did not develop significant OIP-mediated protection against ischemic ARF. Conclusions: OIP (i) activates the cytoprotective Nrf2, but not the P21, pathway; (ii) suppresses post-ischemic P21 induction and renal senescence; and (iii) confers marked protection against ischemic ARF. In sum, these findings suggest that OIP may be a clinically feasible approach for safely activating the Nrf2 pathway, and thereby confer protection against clinical renal injury.