Ezrin is down-regulated in diabetic kidney glomeruli and regulates actin reorganization and glucose uptake via GLUT1 in cultured podocytes.

Diabetic nephropathy is a complication of diabetes and a major cause of end-stage renal disease. To characterize the early pathophysiological mechanisms leading to glomerular podocyte injury in diabetic nephropathy, we performed quantitative proteomic profiling of glomeruli isolated from rats with streptozotocin-induced diabetes and controls. Fluorescence-based two-dimensional difference gel electrophoresis, coupled with mass spectrometry, identified 29 differentially expressed spots, including actin-binding protein ezrin and its interaction partner, NHERF2, which were down-regulated in the streptozotocin group. Knockdown of ezrin by siRNA in cultured podocytes increased glucose uptake compared with control siRNA-transfected cells, apparently by increasing translocation of glucose transporter GLUT1 to the plasma membrane. Knockdown of ezrin also induced actin remodeling under basal conditions, but reduced insulin-stimulated actin reorganization. Ezrin-dependent actin remodeling involved cofilin-1 that is essential for the turnover and reorganization of actin filaments. Phosphorylated, inactive cofilin-1 was up-regulated in diabetic glomeruli, suggesting altered actin dynamics. Furthermore, IHC analysis revealed reduced expression of ezrin in the podocytes of patients with diabetes. Our findings suggest that ezrin may play a role in the development of the renal complication in diabetes by regulating transport of glucose and organization of the actin cytoskeleton in podocytes.

[Functional MRI investigation of brain activity triggered by taste stimulation]. / Ízstimulációval kiváltott agyi tevékenység funkcionális MR-vizsgálata elhízásban.

INTRODUCTION: Many factors contribute to the pathogenesis of morbid obesity, and the central nervous system - as one of those - also has an important role. Numerous studies focus on the central regulation of eating and metabolism, since associated problems like obesity, anorexia, diabetes or metabolic syndrome put an increasing burden on the health system of modern societies. Neither the pathophysiologic changes, nor the normal regulation of these systems are known adequately. Functional MR (fMRI) imaging, which has certainly gained popularity recently, aims to better understand these mechanisms. In this series we studied the brain fMRI activity changes of normal and obese persons, triggered by gustatory stimulation. METHODS: 10 obese and 10 normal weight healthy volunteers took part in the study, with comparable age and sex distribution. Gustatory stimulation was performed by 0.1 M sucrose (pleasant), 0.5 mM quinine HCl (unpleasant) and complex vanilla flavored (Nutridrink) solutions, which were administered through 0.5 mm PVC tubes, in 5-5 ml portions. For rinsing distilled water with neutral flavor was used. Imaging was performed in a 3T MRI, applying standard EPI sequences. Post processing of data was accomplished by FSL software package. RESULTS: Brain activation for gustatory stimuli was characteristically different between the two groups. There were high intensity activations in more cortical and subcortical regions of the obese volunteers compared to the normal ones. CONCLUSIONS: Our current fMRI investigations revealed different activations of numerous brain regions of normal and obese individuals, triggered by pleasant and unpleasant gustatory stimulation. Based on these results this method can help to recognize the role of the central nervous system in obesity, and may contribute to develop new therapies for weight loss.

Homeostatic alterations after IL-1beta microinjection into the nucleus accumbens of the rat.

The present study investigates the effects of direct administration of interleukin-1beta (IL-1beta) into the nucleus accumbens (NAcc) on homeostatic regulation. Short- and long-term food intakes (FI), water intakes (WI) and body temperature (BT) were measured before and after bilateral microinjection of IL-1beta (with or without paracetamol pretreatment) into the NAcc of Wistar rats, and the effects were compared with those found in vehicle treated control animals. In addition, blood glucose levels, along with a glucose tolerance test (GTT), and plasma concentrations of metabolic parameters, such as total cholesterol, triglycerides, HDL, LDL and uric acid were determined in cytokine treated and control rats. Short-term FI and WI were suppressed after intraaccumbens application of IL-1beta. A significant increase of BT was also observed after the cytokine microinjection. Pretreatment with paracetamol failed to influence the anorexigenic, adipsogenic, and pyrogenic effects of IL-1beta. A definite glucose intolerance of the cytokine treated animals and their pathologically elevated blood glucose levels became obvious in the acute GTT. Following IL-1beta microinjection, plasma levels of triglycerides, total cholesterol and LDL were found increased. Our present findings show that the NAcc is an important site of action of IL-1beta mediated processes in central homeostatic regulation.

Taste reactivity deficit in anorexia nervosa.

AIM: Anorexia nervosa is a complex psychiatric disorder posing a rapidly increasing burden on modern societies. Our purpose was to clarify perceptual-motivational aspects of gustatory disturbances in the disease. METHODS: A taste reactivity test, with the use of all five primary qualities in two concentrations, was performed in restrictive-type anorexic patients, and their hedonic evaluations were compared to those of age-matched healthy control subjects. RESULTS: The patients gave significantly lower pleasantness scores for pleasant taste stimuli compared with controls. The differences were the greatest for the lower concentration sucrose, umami and sodium chloride. Ratings given for the aversive taste stimuli were similar in both experimental groups. CONCLUSION: Our findings contribute to a better understanding of complex symptoms of anorexia nervosa, and may also help to develop more effective cognitive-behavioral therapies.

Ruptured Aortic Aneurysm and Dissection Related Death: an Autopsy Database Analysis.

Acute aortic catastrophes (AAC), mainly ruptured aneurysms and dissections, lead all other vascular conditions in morbidity and mortality, even if intervention occurs. The aim of our study was to give a descriptive overview of the demographic and pathological characteristics of AAC. Between 1994 and 2013, 80,469 autopsies were performed at Semmelweis University hospitals in Budapest. After collecting the autopsy reports we were able to create the AAC database upon which we conducted our analysis. We found 567 cases of AAC. The cause of death in 120 of them was classified as a non-ruptured aorta with malperfusion or distal embolization. Of the remaining 447 cases, in 305 the cause of death was a ruptured aortic aneurysm (rAA), and in 142 it was a ruptured aortic dissection (rAD). The distribution of rAA cases was 34.4% thoracal, 4.3% thoracoabdominal, and 61.3% abdominal. We found female dominance where the rAA was thoracal. In rAD cases, 84% were Stanford A and 16% Stanford B type. In both groups we found different pathological distributions. In the prehospital group, the number of thoracal ruptures was considerable. 88% of the patients with Stanford A dissection died in the prehospital or perioperative period. The most progressive AACs were ruptures of intrapericardial aneurysms and Stanford A dissections., however survival rate can be elevated by using rapid imaging examination and immediate surgical intervention. We want to highlight that our study contains such gender differences, which are worth to be taken into consideration.

Taste reactivity alterations after IL-1beta microinjection into the ventromedial hypothalamic nucleus of the rat.

The ventromedial hypothalamic nucleus (VMH) is a central site of action of interleukin-1beta (IL-1beta) induced feeding disturbances. This study was designed to elucidate taste-related perceptual and motivational processes potentially contributing to the anorexia and adipsia seen after bilateral IL-1beta microinjection into the VMH. A saccharin conditioned taste aversion (CTA) paradigm was tested after the central IL-1beta administration. To further investigate whether gustatory deficits are involved in development of the feeding alterations, IL-1beta induced changes of taste responsiveness were also studied in taste reactivity tests. Administration of the cytokine into the VMH did not cause the development of CTA. During taste reactivity tests, however, IL-1beta treated rats displayed significantly poorer ingestive reactions to pleasant taste stimuli than did animals of the control group. In addition, the aversive responses of IL-1beta injected rats to pleasant tastes were significantly more robust than those of control animals. The cytokine treated animals also showed stronger aversion than ingestion to hedonically positive tastes. The present findings indicate that (1) anorexigenic and adipsogenic consequences of IL-1beta microinjection into the VMH are not due to development of cytokine induced CTA; and (2) hedonic responsiveness to palatable tastes is processed by IL-1beta mediated neural mechanisms in the VMH.

History of simple febrile seizures is associated with hippocampal abnormalities in adults.

SUMMARY BACKGROUND: It is unclear whether the hippocampal abnormality in temporal lobe epilepsy (TLE) is a consequence or the cause of afebrile or febrile seizures (FSs). We investigated whether hippocampal abnormalities are present in healthy adults>15 years after a simple FS. METHODS: Eight healthy subjects (5 men) with a history of simple FS (FS+ group) and eight sex- and aged-matched control subjects (FS- group) were investigated by three MR methods: blinded visual inspection of the MRI pictures; automatic voxel-based volumetry; and T2 relaxation time measurements. RESULTS: The mean total volume of the two hippocampi was 5.36 +/- 1.33 cm(3)in the FS+ group and 6.63 +/- 1.46 cm(3)in the FS- group (p = 0.069). The T2 values in the anterior part of the left hippocampus (p = 0.036) and in the middle part of the right hippocampus (p = 0.025) were elevated in the FS+ subjects. The mean volume of the right hippocampus was 3.05 +/- 0.8 cm(3)in the FS+ men and 4.05 +/- 0.48 cm(3)in the FS- men (p = 0.043). The mean total volume of the two hippocampi was 5.38 +/- 1.4 cm(3)in the FS+ men and 7.48 +/- 1.14 cm(3)in the FS- men (p = 0.043). There were three FS+ men in whom hippocampal abnormalities including hippocampal sclerosis (HS) and dysgenesis were observed on visual inspection. CONCLUSIONS: A history of simple FS in childhood can be associated with hippocampal abnormalities in adults. These abnormalities are probably more pronounced in men. Simple FS may not be as a benign event as previously thought. Our findings suggest that hippocampal abnormalities associated with FS are not necessarily epileptogenic.

Right-left discrimination is related to the right hemisphere.

We aimed to determine the functional localisation of right-left discrimination (RLD) by functional MRI (fMRI). In this study, 16 male volunteers were examined. There were three task sessions: one active and two baseline tasks. During the baseline tasks participants were instructed to show numbers with their fingers. The first baseline task was performed with the right hand, the second one with the left hand. During the active (RLD) task participants were also instructed to show numbers. The difference between baseline and active tasks was that during the active task the hand with which the participant should perform the instruction was assigned randomly. Thus, participants were unaware which hand should be used before the instruction command. During RLD, activations occurred in the right-sided frontal, precuneus, postcentral, angular, lingual, and superior temporal gyri. Activations also appeared in the left-sided temporal gyri and precuneus. Of the activations, 76.7% appeared in the right hemisphere, 23.3% in the left hemisphere. Conclusively, we found that RLD is mainly related to the right hemisphere, and requires activation of the parieto-temporo-occipital junction and the visual system including cuneus, precuneus, and gyrus lingualis.

Glucose-monitoring neurons in the nucleus accumbens.

The nucleus accumbens, a key structure of the limbic circuitry, is involved in the regulation of motivated behaviors. The accumbens performs its roles via interconnections with brain areas where glucose-monitoring neurons have been localized. To search for such integrative chemosensory cells here, extracellular single neuron activity was recorded in the nucleus accumbens of Wistar rats by means of multibarreled glass microelectrodes during microelectrophoretic administration of D-glucose and other chemicals. Every fourth neuron tested changed in activity in response to glucose. Accumbens cells also displayed distinct gamma-amino-n-butyric acid type sensitivity. It is suggested that differential distribution of the chemosensory units, demonstrated between subdivisions of the nucleus accumbens, has particular significance with respect to functional dichotomy of the shell and core subregions.

Identifying non-toxic doses of manganese for manganese-enhanced magnetic resonance imaging to map brain areas activated by operant behavior in trained rats.

Manganese-enhanced magnetic resonance imaging (MEMRI) offers unique advantages such as studying brain activation in freely moving rats, but its usefulness has not been previously evaluated during operant behavior training. Manganese in a form of MnCl2, at a dose of 20mg/kg, was intraperitoneally infused. The administration was repeated and separated by 24h to reach the dose of 40mg/kg or 60mg/kg, respectively. Hepatotoxicity of the MnCl2 was evaluated by determining serum aspartate aminotransferase, alanine aminotransferase, total bilirubin, albumin and protein levels. Neurological examination was also carried out. The animals were tested in visual cue discriminated operant task. Imaging was performed using a 3T clinical MR scanner. T1 values were determined before and after MnCl2 administrations. Manganese-enhanced images of each animal were subtracted from their baseline images to calculate decrease in the T1 value (ΔT1) voxel by voxel. The subtracted T1 maps of trained animals performing visual cue discriminated operant task, and those of naive rats were compared. The dose of 60mg/kg MnCl2 showed hepatotoxic effect, but even these animals did not exhibit neurological symptoms. The dose of 20 and 40mg/kg MnCl2 increased the number of omissions and did not affect the accuracy of performing the visual cue discriminated operant task. Using the accumulated dose of 40mg/kg, voxels with a significant enhanced ΔT1 value were detected in the following brain areas of the visual cue discriminated operant behavior performed animals compared to those in the controls: the visual, somatosensory, motor and premotor cortices, the insula, cingulate, ectorhinal, entorhinal, perirhinal and piriform cortices, hippocampus, amygdala with amygdalohippocampal areas, dorsal striatum, nucleus accumbens core, substantia nigra, and retrorubral field. In conclusion, the MEMRI proved to be a reliable method to accomplish brain activity mapping in correlation with the operant behavior of freely moving rodents.

Oxidative/Nitrative Stress and Inflammation Drive Progression of Doxorubicin-Induced Renal Fibrosis in Rats as Revealed by Comparing a Normal and a Fibrosis-Resistant Rat Strain.

Chronic renal fibrosis is the final common pathway of end stage renal disease caused by glomerular or tubular pathologies. Genetic background has a strong influence on the progression of chronic renal fibrosis. We recently found that Rowett black hooded rats were resistant to renal fibrosis. We aimed to investigate the role of sustained inflammation and oxidative/nitrative stress in renal fibrosis progression using this new model. Our previous data suggested the involvement of podocytes, thus we investigated renal fibrosis initiated by doxorubicin-induced (5 mg/kg) podocyte damage. Doxorubicin induced progressive glomerular sclerosis followed by increasing proteinuria and reduced bodyweight gain in fibrosis-sensitive, Charles Dawley rats during an 8-week long observation period. In comparison, the fibrosis-resistant, Rowett black hooded rats had longer survival, milder proteinuria and reduced tubular damage as assessed by neutrophil gelatinase-associated lipocalin (NGAL) excretion, reduced loss of the slit diaphragm protein, nephrin, less glomerulosclerosis, tubulointerstitial fibrosis and matrix deposition assessed by periodic acid-Schiff, Picro-Sirius-red staining and fibronectin immunostaining. Less fibrosis was associated with reduced profibrotic transforming growth factor-beta, (TGF-ß1) connective tissue growth factor (CTGF), and collagen type I alpha 1 (COL-1a1) mRNA levels. Milder inflammation demonstrated by histology was confirmed by less monocyte chemotactic protein 1 (MCP-1) mRNA. As a consequence of less inflammation, less oxidative and nitrative stress was obvious by less neutrophil cytosolic factor 1 (p47phox) and NADPH oxidase-2 (p91phox) mRNA. Reduced oxidative enzyme expression was accompanied by less lipid peroxidation as demonstrated by 4-hydroxynonenal (HNE) and less protein nitrosylation demonstrated by nitrotyrosine (NT) immunohistochemistry and quantified by Western blot. Our results demonstrate that mediators of fibrosis, inflammation and oxidative/nitrative stress were suppressed in doxorubicin nephropathy in fibrosis-resistant Rowett black hooded rats underlying the importance of these pathomechanisms in the progression of renal fibrosis initiated by glomerular podocyte damage.

LPS-induced delayed preconditioning is mediated by Hsp90 and involves the heat shock response in mouse kidney.

INTRODUCTION: We and others demonstrated previously that preconditioning with endotoxin (LPS) protected from a subsequent lethal LPS challenge or from renal ischemia-reperfusion injury (IRI). LPS is effective in evoking the heat shock response, an ancient and essential cellular defense mechanism, which plays a role in resistance to, and recovery from diseases. Here, by using the pharmacological Hsp90 inhibitor novobiocin (NB), we investigated the role of Hsp90 and the heat shock response in LPS-induced delayed renal preconditioning. METHODS: Male C57BL/6 mice were treated with preconditioning (P: 2 mg/kg, i.p.) and subsequent lethal (L: 10 mg/kg, i.p.) doses of LPS alone or in combination with NB (100 mg/kg, i.p.). Controls received saline (C) or NB. RESULTS: Preconditioning LPS conferred protection from a subsequent lethal LPS treatment. Importantly, the protective effect of LPS preconditioning was completely abolished by a concomitant treatment with NB. LPS induced a marked heat shock protein increase as demonstrated by Western blots of Hsp70 and Hsp90. NB alone also stimulated Hsp70 and Hsp90 mRNA but not protein expression. However, Hsp70 and Hsp90 protein induction in LPS-treated mice was abolished by a concomitant NB treatment, demonstrating a NB-induced impairment of the heat shock response to LPS preconditioning. CONCLUSION: LPS-induced heat shock protein induction and tolerance to a subsequent lethal LPS treatment was prevented by the Hsp90 inhibitor, novobiocin. Our findings demonstrate a critical role of Hsp90 in LPS signaling, and a potential involvement of the heat shock response in LPS-induced preconditioning.

Activation of the miR-17 family and miR-21 during murine kidney ischemia-reperfusion injury.

BACKGROUND: Ischemia-reperfusion (I/R) is the main cause of acute kidney injury (AKI) in patients. We investigated renal microRNA (miRNA) expression profiles and the time course of changes in selected miRNA expressions after renal I/R to characterize the miRNA network activated during development and recovery from AKI. METHODS AND RESULTS: One day after lethal (30 minutes) and sublethal (20 minutes) renal ischemia, AKI was verified by renal histology (tubular necrosis, regeneration), blood urea nitrogen (BUN) level, renal mRNA expression, and plasma concentration of neutrophil gelatinase-associated lipocalin (NGAL) in C57BL/6J mice. On the first day after 30-minute, lethal I/R miR-21, miR-17-5p, and miR-106a were elevated out of the 21 miRNAs successfully profiled on the Luminex multiplex assay. After 20-minute, sublethal I/R, renal miR-17-5p and miR-106a expressions were elevated on the first and second days of reperfusion, while miR-21 expression increased later and lasted longer. Renal miR-17-5p and miR-21 expressions correlated with each other. Renal function returned to normal on the fourth day after sublethal I/R. CONCLUSIONS: Our results demonstrate that besides miR-21, miR-17-5p, and miR-106a are additionally activated during the maintenance and recovery phases of renal I/R injury. Furthermore, a correlation between renal miR-17-5p and miR-21 expressions warrants further investigation of how they may influence each other and the outcome of renal ischemia-reperfusion injury.

Glucose-monitoring neurons in the mediodorsal prefrontal cortex.

The mediodorsal prefrontal cortex (mdPFC), a key structure of the limbic neural circuitry, plays important roles in the central regulation of feeding. As an integrant part of the forebrain dopamine (DA) system, it performs complex roles via interconnections with various brain areas where glucose-monitoring (GM) neurons have been identified. The main goal of the present experiments was to examine whether similar GM neurons exist in the mediodorsal prefrontal cortex. To search for such chemosensory cells here, and to estimate their involvement in the DA circuitry, extracellular single neuron activity of the mediodorsal prefrontal cortex of anesthetized Wistar and Sprague-Dawley rats was recorded by means of tungsten wire multibarreled glass microelectrodes during microelectrophoretic administration of d-glucose and DA. One fourth of the neurons tested changed in firing rate in response to glucose, thus, proved to be elements of the forebrain GM neural network. DA responsive neurons in the mdPFC were found to represent similar proportion of all cells; the glucose-excited units were shown to display excitatory whereas the glucose-inhibited neurons were demonstrated to exert mainly inhibitory responses to dopamine. The glucose-monitoring neurons of the mdPFC and their distinct DA sensitivity are suggested to be of particular significance in adaptive processes of the central feeding control.

Gustatory perception alterations in obesity: an fMRI study.

The background of feeding associated and metabolic diseases is not sufficiently understood yet. Since gustatory alterations may be of particular significance in the above illnesses, in the present experiments, cerebral activation was detected by fMRI in twelve obese patients and twelve, age and gender matched healthy subjects. The gustatory stimulus solutions were delivered via intraorally positioned polyvinyl tubes. Each session consisted of three runs. Sucrose was used as a pleasant; quinine HCl as an aversive; and a high-calorie, vanilla flavored nourishment solution as a complex taste of high palatability. In each run, only one taste was used as a stimulus. During all runs, distilled water served as a neutral stimulus. Group analysis was made by using the FSL software package. The taste stimuli elicited characteristic and distinct activity changes of the two groups. In contrast to the controls, in the obese patients, stronger activation was detected in various cortical (anterior cingulate cortex, insular and opercular cortices, orbitofrontal cortex) and subcortical (amygdala, nucleus accumbens, putamen and pallidum) structures in case of all three stimuli. The present examinations elucidated differential activation of various brain structures to pleasant and unpleasant gustatory stimuli in obese patients compared to control subjects. These taste alterations are supposed to be of particular significance in obesity, and our findings may contribute to develop better strategies for prevention and effective therapies in the future.

Body weight and the reward system: the volume of the right amygdala may be associated with body mass index in young overweight men.

We aimed to investigate the relationship between BMI (body mass index) and the volumes of the structures of the reward system (hippocampus, amygdala, accumbens, caudatum, putamen, and orbitofrontal cortex). The right and left structures were examined separately. Their volumes were assessed using a 3-T MRI scanner and Freesurfer software. Ninety-two healthy subjects were involved (mean BMI: 22.3 ± 3.4 kg/m(2), mean age: 23.2 ± 2.7). We found that the volume of the right amygdala positively correlated with the BMI in men but not in women. Moreover, we could demonstrate this association only in the overweight male sub-population. We suggest that an association between body weight and the morphological variability of the reward system can be demonstrated by MRI. This may be further evidence for a different body-weight regulation in the two sexes. The potential relationship between the volume of the right amygdala and the BMI in heavier individuals requires further studies with larger samples.